INNATE IMMUNITY AND INFLAMMATION Shizuo Akira. Laboratory of Host Defense, WPI Immunology Frontier Research Center, Osaka University, Osaka, Japan
Gene expression is controlled at multiple points, including signal transduction, transcription and mRNA stability. So far, transcriptional regulation has been extensively studied. However, recent studies have revealed that control of gene expression at the mRNA level is as important as transcriptional control in the immune response.
The innate immune system is an evolutionally conserved host defense mechanism against pathogens. Innate immune responses are initiated by pattern recognition receptors (PRRs), which recognize specific structures of microorganisms. Among them, Toll-like receptors (TLRs) are capable of sensing organisms ranging from bacteria to fungi, protozoa and viruses, and play a major role in innate immunity. Individual TLRs recognize different microbial components, activate different signaling pathways via selective usage of adaptor molecules, and give rise to different patterns in gene expression.
We are now focusing on the role of genes induced in response to TLR stimulation, particularly the genes that are rapidly induced in a MyD88-dependent manner within 30 min after LPS stimulation. Among them, we have recently identified a novel gene named Zc3h12a which has a CCCH-type zinc finger domain. The knockout mice developed spontaneous autoimmune diseases accompanied by splenomegaly and lymphadenopathy. Subsequent studies showed that Zc3h12a is a nuclease involved in destabilization of IL-6 and IL-12mRNA via the stem loop structure present in the 3′UTR of these genes. We renamed it Regulatory RNase-1 (Regnase-1) based on the function. I would like to discuss the role of Regnase-1 in the immune response.
MOLECULAR FOUNDATIONS OF BIOELECTRONIC MEDICINE Kevin J. Tracey. The Feinstein Institute for Medical Research, Manhasset, NY, USA
The fundamental building blocks of the nervous system are reflex circuits that confer stability to organ system function. Reflex circuits maintain physiological homeostasis during fighting, fleeing, resting, digesting, and other prerequisites for species survival. Reflex activity is initiated when the environment changes. This activates action potentials in the sensory (afferent) arc that travel toward the central nervous system. The resultant output from this input is relayed through other neural signals that return via motor (efferent) neurons to the innervated system. This provides exquisite neural modulation of organ system function. The discovery of the inflammatory reflex as a prototypical reflex that modulates immunity established that reflex mechanisms regulate the immune system. This discovery arose during studies of an experimental cytokine-inhibiting molecule (CNI-1493). When administered into the brain, CNI-1493 unexpectedly inhibited the production of TNF by macrophages in the spleen and other organs. Cutting the vagus nerve, a major nerve to the body's organs, reversed this effect, indicating that the efferent signaling in the vagus nerve inhibited macrophage activation in the spleen and other organs of the reticulo-endothelial system. The neurophysiological and molecular mechanistic basis for this inflammatory reflex has been delineated in detail. Signals traveling in the vagus nerve modulate the activity of the splenic nerve, which secretes norepinephrine in spleen. Norepinephrine signals via beta-2AR expressed on specific T cells that synthesize and release acetylcholine. This is the terminal neurotransmitter in the reflex that binds to alpha-7 nicotinic acetylcholine receptors expressed on splenic and other macrophages to effectively inhibit cytokine release. IL-1, TNF, LPS, and other mediators stimulate sensory signals in the afferent vagus nerve. This can initiate the sensory arc of the inflammatory reflex; the motor arc, which can be activated using nerve-stimulating electrodes, counter-regulates inflammation. This rapidly expanding new field has been extended into several important clinical trials, where implantable nerve stimulators have been shown to stimulate the inflammatory reflex, suppress cytokine production, and alleviate disease in patients with rheumatoid arthritis and inflammatory bowel disease. The field of developing devices that target reflex molecular mechanisms using electrons is termed bioelectronic medicine. Progress stems from the convergence of molecular mechanisms with neural circuits to guide the development of therapeutic devices. Future patients will use electrons, instead of drugs, because of basic discoveries in understanding fundamental reflex principles in immunity and other organ systems.
THE STATE OF SHOCK SOCIETY, US Ping Wang. Shock Society, USA and Center for Immunology & Inflammation, The Feinstein Institute for Medical Research, Manhasset, NY, USA
The Shock Society was originated in 1978 to facilitate the integration of basic and clinical disciplines in the study of the pathophysiology of trauma and shock and to promote awareness of these indications both nationally and internationally. The society's mission is to improve the care of victims of trauma, shock and sepsis through promoting clinically relevant research, providing a multidisciplinary forum to integrate the basic and clinical aspects and to educate the next generation investigators in the field of trauma, shock and sepsis. The shock has aimed to accomplish these goals by several means. The journal SHOCK publishes original basic and clinical studies on shock and trauma. The Annual Conference on Shock provides a multidisciplinary forum to integrate the basic research and clinical arena, and promote awareness of shock and trauma to the members as well as students in the field. Every year, the society invites a world renowned researcher as the Keynote Speaker to share the latest research findings and expertise. In addition, the Shock Society offers a number of awards. The Research Faculty Award for Early Career Faculty Investigators is to support the career development of young investigators in the area of trauma, shock and sepsis. New Investigator Award and Travel Awards are given to qualifying students and postdoctoral fellows in research training to present their latest findings at the annual conference on Shock. This year in conjunction with the Department of Defense, two additional awards, the Shock-DoD Postdoctoral Fellowships will be given to qualifying candidates to conduct research in DoD labs. The society's newsletter is distributed twice a year and includes a message from the President, highlights of the upcoming Annual conference and an update of the society's finances to keep the membership abreast of the academic and business activities of the society. The founding meeting of the Shock Society held on April 5, 1977 in Chicago had 30 participants. Now the society is comprised of over 400 members, a highly structured executive council and numerous committees to facilitate and oversee various activities of the society. In the future, the society will concentrate more on the next generation of shock investigators as well as public awareness of the field of trauma, shock and sepsis. We intend to recruit young bright minds and continue to provide financial and mentoring support to young shock investigators at both basic and clinical settings. We expect to make the latest ground breaking studies presented at the annual meeting available to the public. With the avenues already in place and our ambitious plans, we envision that the Shock Society can achieve its mission to improve the care and treatment of victims of trauma, shock and sepsis.
ACHIEVEMENT AND FUTURE PERSPECTIVES OF THE BRAZILIAN SHOCK SOCIETY AND LATIN-AMERICAN SEPSIS INSTITUTE Reinaldo Salomao. Brazilian Shock Society (BSS) Latin American Sepsis Institute; Infectious Diseases/Hospital São Paulo, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, Brazil
Brazilian Shock Society (BSS) was founded in 1993 with Maurício Rocha e Silva as the president and Luíz Francisco Poli de Figueiredo as secretary. The first meetings in Brazil were focused in trauma and burns, with emphasis in the role of hypertonic resuscitation.
The emerging problem of sepsis in world and lack of organized efforts to face its challenges in developing countries led to the foundation of the Latin-American Sepsis Institute (LASI - www.ilas.org.br) by Eliezer Silva and colleagues in July 2004. Since then activities of BSS and LASI converged as one force.
Our major goals are:
- improve the quality of care for septic patients, with consequent reduction of the prevalence, rates of morbidity and mortality;
- generate knowledge in the area of sepsis;
- disseminate this information to lay and scientific communities.
For accomplishment of these goals, LASI has three major areas of actuation: quality improvement initiatives, knowledge generation and diffusion of knowledge.
QUALITY IMPROVEMENT PROGRAMS
In 2005, LASI started the quality improvement program based on the Surviving Sepsis Campaign bundles. Since then more than 150 hospitals adhered to the initiative and LASI database has now almost 40.000 patients. For the global LASI database, the reduction in sepsis mortality in our country from 2005 to 2015 was from 54% to 40%.
KNOWLEDGE GENERATION IN SEPSIS
We have carried out studies to investigate the burden of sepsis in Brazil. The first study done was a multicenter prospective evaluation of the economic impact of sepsis (COSTS study) and the results demonstrated that sepsis was associated with significant healthcare costs (Sogayar et al, Pharmacoeconomics. 2008;26(5):425–34). In 2014, LASI carried out the SPREAD study, a point-prevalence study with outcome assessment, to estimate prevalence and mortality of sepsis in Brazil. Using a random sampling of 13% of Brazilian ICU in all geoeconomic regions, this study collected data from 229 ICUs and demonstrated that sepsis prevalence in our country was 30% and mortality was 55% (Machado et al ICM Experimental 2015, 3 (Suppl 1):A642), showing again the importance of sepsis as a health care issue in Brazil.
KNOWLEDGE DISSEMINATION IN SEPSIS
Since 2004 we promote every year the Sepsis Forum (www.forumsepse.com.br), devoted to healthcare personnel, a two-day event that back on 2004 had only 100 delegates and now has about 600 delegates from all over the country. The event merges clinical research with pre-clinical science, being an important opportunity to share expertise both for bench scientists and bedside professionals.
As for increasing knowledge of sepsis among lay public, since its foundation LASI has continuously provided press releases related to sepsis. This activity increased substantially since 2012 when World Sepsis Day was initiated. In 2014, LASI performed a poll on sepsis awareness that demonstrated a very restrict knowledge on the syndrome by the lay public, since only 7% of the people in Brazil have ever heard the word sepsis. This result highlights the importance of disseminating media campaigns on sepsis in Brazil.
THE UPDATE OF SEVERE SEPSIS IN CHINA Yong-Ming Yao. Chinese Shock Society (CSS) and Trauma Research Center, First Hospital Affiliated to the Chinese PLA General Hospital, Beijing, China
China is the most populous country in the world, but epidemiology of severe sepsis within this country is still not well understood. A study was conducted as a multicenter, nationwide, epidemiologic survey of severe sepsis in surgical ICU patients. The results suggested that severe sepsis is a common, expensive, and frequently fatal syndrome in critically ill surgical patients in China. Other than the microbiological patterns, the incidence, mortality, and major characteristics of severe sepsis in Chinese surgical ICU are close to those documented in developed countries.
Immunological function seems to have become dissonant, leading to progressive rampant inflammatory responses on one hand, and immunosuppression on the other hand. In extensively burned patients, it was found that in the patients who developed severe sepsis, the value of plasma C5a was over 300 fold of that of normal controls on the first day after acute insults. Meanwhile, chemiluminescence of neutrophils was astoundingly low in patients with extensive burns. Since chemiluminescence of neutrophils was positively correlated with bactericidal index of neutrophils, it implied that the bactericidal function of neutrophils was greatly impaired. Human leukocyte antigen locus DR (HLA-DR) is essential in the process of antigen presentation. We quantitatively determined HLA-DR expression in patients with burn injury. Comparing the results between patients developing sepsis and those without sepsis, the lowering of HLA-DR was more remarkable in the former group. The results also arouse our attention that in patients with major burns, one link of the immunologic function represented by HLA-DR level persisted to be suppressed even when all open wounds have been soundly closed.
In a multi-center randomized control clinical trial, we gave Ulinastatin, a human urinary trypsin inhibitor, aiming at suppressing the production of inflammatory mediators and inhibiting a series of proteases and hydrolase, combined by the administration of alpha thymosin to raise the level of HLA-DR and inhibit caspase 3. In a group of 342 patients with severe sepsis, the effect of the treatment was salutary, as shown by lowering of 28-day mortality from 38.3% to 25.1% (P < 0.01), and 90-day mortality from 52.1% to 37.2% (P < 0.01). As for other indices of therapeutic evaluation, such as length of ICU stay, length of intubation and length of antibiotics usage, there was no significant difference between two groups. These results demonstrated that the combined administration of Ulinastatin and thymosin alpha1 for the treatment of severe sepsis appears to be successful.
EUROPEAN SHOCK SOCIETY: PAST, PRESENT AND FUTURE Jean-Marc Cavaillon. European Shock Society (ESS) and Cytokines & Inflammation Unit, Institut Pasteur, Paris, France
The European Shock Society was founded in Malmo (Sweden) on April 1983, after David Lewis (Linkoping, Sweden) had called his European colleagues to gather for the European Shock Society Constitutional Meeting. The inauguration meeting had two major aims: first, to address the contemporary shock research projects run in Europe, and second, to establish the European Shock Society (ESS) with a profile similar to that of the already 5-year old US Shock Society. The constitution document and the bylaws were approved at that meeting as well. The founding members were: Rod Little (Manchester, UK), Lambert Thijs (Amsterdam, The Netherlands), Ulf Haglund (Malmo, Sweden), Konrad Messmer (Heidelberg, Germany), Sandor Nagy (Szeged, Hungary), Jean-Louis Vincent (Brussels, Belgium), Gian Paolo Novelli (Florence, Italy), and lan Ledingham (Glasgow, Scotland) who became the very first president of the ESS. Since then, the ESS meetings have been regularly organized in different European cities on a biennial basis. The most recent ones were held in Lisbon (2009, Chris Thiemermann, president), Taormina (2011, Salvatore Cuzzocrea, president), Vienna (2013, Soheyl Bahrami, president), and Cologne (2015, Edmund Neugebauer, president). For the very first time the ESS congress will be held in France (Institut Pasteur, Paris) in September 2017. As for many international scientific societies, an important task of the ESS is to remain sufficiently attractive and vibrant to maintain the connection with the past members and to attract participation of the new and/or youngest ones. Furthermore, the ESS constitutes a rather small society with very limited operational funds. These issues remain a permanent challenge that is only possible to effectively cope with thanks to the good will and devotion of highly motivated present and past council members, and a robust executive committee. Luckily, the ESS has the chance to have the most efficient General Secretary, Inge Bauer (Dusseldorf, Germany), and the most helpful treasurer, Marcin Osuchowski (Vienna, Austria). Our councilors (see the list on our WEB site: http://www.europeanshocksociety.org) are of great help to maintain an attractive WEB site and to allow the publication of the ESS Newsletter. Indeed, since the ESS meets every two years, one of the key tools maintaining the link among the members and keeping our society up-and-running is the ESS Newsletter that has recently received a robust upgrade. For the first time, during the next ESS congress, a common session will be organized with the Chinese Shock Society, and another one with the American Shock Society. Following the idea of our president-elect, Markus Huber-Lang (Ulm, Germany), a first summer school addressing several up-to-date sepsis topics will be held at beginning of the congress, notably on the world sepsis day (Sept.13th).
PRESENT STATUS AND FUTURE PERSPECTIVE OF JAPAN SHOCK SOCIETY Shigeto Oda. Japanese Shock Society (JSS) and Department of Emergency and Critical Care Medicine, Chiba University Graduate School of Medicine, Chiba, Japan
The Japan Shock Society was established in 1986. The annual congress is held once a year, and this year's congress is the 31st. The number of members is almost 400, and almost unchanged in recent 5 years. The society members comprised physicians of emergency and critical care medicine (48%), anesthesiologist (17%), surgeons (17%), internal medicine (9%), and basic researcher (6%). It is characterized that clinical doctors is the majority and the number of basic researchers is very small. The annual congress is held in every June, and the number of the participant is about 150. Around forty papers including 30 original papers in average are presented in one day session of the annual congress. In the original papers, 55% is basic research and 45% is clinical research, which is also the feature of our society despite the large number of members is a physician, showing our society is interdisciplinary. The theme of special session, such as symposia or panel discussions in recent 5 years are “tissue dysoxia”, “sepsis and organ dysfunction”, “microcirculation”, “basic research in sepsis pathophysiology”, and “coagulopathy in shock”. The problem of our society is firstly, decreasing of young members and participants to the congress. Recent trend of loss of interest in research in young doctors is the serious common problem not only for our society but for many interdisciplinary societies. Therefore, joint meeting with other society interested in similar area is the future option of the annual congress. The second problem is that translational research between basic and clinical application is lacking despite excellent basic research has been carried out. Therefore, participation of more basic researchers and involvement of companies are the key for future development of shock research and improvement of clinical outcome.
SOCIETY FOR SHOCK STUDIES (RUSSIA) - CURRENT STATE AND FUTURE Viktor V. Moroz. Society for Shock Studies (Russia); V.A. Negovsky Research Institute of General Reanimatology, Moscow, Russia
Shock is an issue that joins forces of many people from biologists and physiologists to professionals working in emergency medicine.
Shock is a collective term for the critical states which acutely arise after a sudden impact of exo- or endogenic factors on balanced organism and is characterized by hemodynamic hemorheological and metabolic disorders. Experts in reanimatology and anesthesiology, surgeons, trauma surgeons and doctors of other specialities deal with the shock and its consequences on a daily basis. Shock is a process that accompanies most of the critical states. The Russian Society for shock studies History was founded in 2012. The primary goal of the Society is to improve the quality of care in shock states. The objectives of the Society are: promotion of basic and clinical researches in the field of shock; bringing together researchers and physicians of different specialties for the spread of new knowledge about the shock; encouraging professional education and mentoring of the society members, researchers and young specialists; implementation of the latest scientific achievements in the field of shock studies; ensuring the continuity of knowledge and improving the quality of education the next generations of researchers; representing the interests of the Russian school of reanimatology in the European and international medical organizations and associations.
CIRCULATING HISTONES AND COAGULATION DISORDER Toshiaki Iba. Department of Emergency and Disaster Medicine, Juntendo University, Tokyo, Japan
The role of damage-associated molecular patterns (DAMPs) in the progression of inflammation has been widely accepted. Histones are the major DAMPs increased in the circulating blood along with the various types of cell-death during severe sepsis. The former studies revealed that histones have extremely strong cytotoxicity to the vascular endothelial cells (ECs) and activate coagulation system. Together with the aforementioned background, it was also reported that the damage of histone is significantly attenuated in the serum. Several plasma proteins such as albumin, activated protein C (APC) and pentraxin 3 (PTX3) are considered to have potential to inhibit the actions of histones. Hence, we intended to examine the protective effects of these physiological proteins against histone in the experimental model using cultured ECs. As a result, all of albumin, APC and PTX3 have demonstrated significant protective effects in a dose dependent manner. In addition, since albumin and PTX3 inhibited histone-induced damage at physiological levels found in serum, these proteins are considered to be the major histone inhibitors in the clinical situation.
Regarding the pharmaceutical agents, heparins are expected to neutralize the histone toxicity, and we examined the effects of unfractionated (UFH) and low-molecular weight heparin (LMWH) both in vivo and in vitro settings. The results are as follows; both UFH and LMWH significantly suppressed the histone-induced decrease of the WBC and platelet counts in the animal models, both UFH and LMWH attenuated histone-induced hepatic and renal dysfunction, high-dose UFH and LMWH reduced the mortality significantly. The in vitro study revealed that both vascular EC death and leukocyte cell death were significantly attenuated by UFH and LMWH. From these results, we speculated that histone-neutralizing effect of heparins might contribute to the beneficial effects of heparins.
In summary, since some plasma proteins and heparins have shown the protective effects, we think the administration of these agents may be a choice of treatments for the patients suffering from severe sepsis.
MOLECULAR MANAGEMENT IN POLYTRAUMA Markus Huber-Lang. Department of Orthopaedic Trauma Surgery, University Hospital of Ulm, Ulm, Germany
Abstract not available yet.
THE ROLE OF FIBRINOGEN IN ACUTE TRAUMA-INDUCED COAGULOPATHY: NOVEL INSIGHTS AND THERAPEUTIC OPTIONS Herbert Schöchl. AUVA Trauma Centre Salzburg, Salzburg; Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Vienna, Austria
Low fibrinogen concentrations are associated with an increased risk of bleeding and massive transfusion.
Fibrinogen seems to be the most vulnerable coagulation factor, reaching a critically low threshold earlier than any other coagulation protein. Fibrinogen level l<1 g/l has been reported as the first laboratory indicator of coagulopathy in trauma.
Measurement of fibrinogen concentration is highly recommended in major trauma patients when they are admitted to the ER. In emergency situations, a rapid, simple and reliable method for quantifying clottable fibrinogen is needed. Specific ROTEM and TEG assays (FIBTEM and Functional Fibrinogen (FF)) are designed to assess the functional capacity of fibrinogen. This is achieved by assessing clotting in the presence of a platelet inhibitor: Cytochalasin D for the FIBTEM assay and abciximab for the FF assay. It allows specific evaluation of the fibrin component of the clot.
Therapy of hypofibrinogenemia
The threshold at which the plasma concentration of fibrinogen may be considered as critically low in trauma is not well established. The recent updates of the European Trauma Guidelines, recommend that plasma fibrinogen concentration in bleeding trauma patients should be not lower than 1.5–2 g/l. Until now no randomized controlled studies are available to support these suggested values. Experimental and initial clinical studies have reported promising outcomes in response to replacement of fibrinogen as the initial step in managing trauma-induced coagulopathy.
There are three treatment options for fibrinogen replacement:
- FFP: Plasma is usually donated by healthy volunteers, so it contains only low concentrations of fibrinogen, and there is considerable variation between donors. Mean fibrinogen concentrations between 2 and 2.9 g/l have been reported in FFP and solvent- detergent (SD) plasma.
- Cryoprecipitate: The fibrinogen content of cryoprecipitate is not well standardized and may vary widely (120–796 mg per single unit). Cryoprecipitate is generally used as a pooled product comprising single units from 6–10 donors, and this increases recipients’ donor exposure. Moreover, cryoprecipitate is not virus inactivated and must be cross matched.
- Fibrinogen concentrate (FC): FC contains a well-defined concentration of fibrinogen. FC undergoes numerous purification and viral inactivation steps during manufacture. The initial dose is 25 – 50 mg/kg BW.
THE UPDATED GUIDELINE FOR THE CLINICAL MANAGEMENT OF ACUTE TRAUMA HEMORRHAGE AND COAGULOPATHY: THE EUROPEAN PERSPECTIVE Marc Maegele. Department of Traumatology, Orthopedic Surgery and Sporttraumatology, Cologne-Merheim Medical Center (CMMC)/University Witten-Herdecke (Germany), Cologne, Germany
Trauma remains the leading cause of death with bleeding the primary cause of preventable death during the first 24 hours after trauma. One out of four severe trauma patients to arrive in the Emergency Department (ED) is already in the state of shock and acute traumatic coagulopathy (ATC). The principle drivers of ATC have been characterized by tissue hypoperfusion, inflammation, and the acute activation of neurohumoral systems. In contrast, there is an iatrogenic coagulopathy (IC) which occurs secondary to uncritical volume therapy leading to acidosis, hypothermia and hemodilution. It has been shown that early recognition prompted by appropriate and aggressive management can correct coagulopathy, control bleeding, reduce blood product use and improve outcomes in severely injured patients. In accordance with the 2016 European guideline for the management of bleeding and coagulopathy following major trauma there is broad consensus that monitoring and measures to support coagulation should be initiated as early as possible. Numerous retrospective studies have suggested improved outcomes when using a blood product ratio-based approach in patients with massive hemorrhage and to date the 1:1:1 transfusion strategy of blood products has been widely adopted. However, substantial methodological limitations to these studies exist which preclude any definitive conclusion on the potential benefit of this strategy with regard to efficacy and safety. Several studies have raised concerns regarding the potential increase in morbidity in particular when patients were overtriaged to 1:1:1 in cases where massive transfusion was unlikely. Alternatively, viscoelastic methods have been advocated to assist in characterizing the coagulopathy, guiding hemostatic therapy and their recommendation has been lifted to grade 1C within the updated 2016 European guideline. Early variables of clot firmness assessed by viscoelastic methods have been demonstrated to be good predictors for the need of massive transfusion and outcomes. Despite the rapidly increasing number of publications on the use of viscoelastic methods for the early detection of hemostatic disorders after trauma, controversy still remains on the standardisation of this technology. Undisputable advantages of this technology remain its rapid availability in the ED or at the bedside thus improving the availability of real-time point-of-care data to guide therapy, as well as its ability to visualize the dynamics and the quality of clot formation.
COAGULATION ABNORMALITY DURING SEPSIS AND ACUTE PANCREATITIS IN AGED ANIMALS Hiroshi Saito1, Daiki Okamura2,3, Charles T. Esmon4, B. Mark Evers5, Starr E. Starr1. 1Department of Surgery, University of Kentucky, Lexington, KY, USA, 2Department of Surgery, East Chiba Medical Center, Chiba, Japan, 3Department of General Medical Science, Chiba University Graduate School of Medicine, Chiba, Japan, 4Oklahoma Medical Research Foundation, University of Oklahoma Health Science Center, Oklahoma City, OK, USA, 5Markey Cancer Center, University of Kentucky, Lexington, KY, USA
Severe disseminated intravascular coagulation (DIC) is an independent predictor of clinical outcome in patients with critical illnesses such as sepsis and acute pancreatitis (AP). DIC is also considered to be a major factor in the pathogenesis of microvascular dysfunction and resulting organ failure. Because a majority of deaths by sepsis and AP occur among the elderly population, our laboratory has been studying the effects of aging on pathophysiology of these critical illnesses.
Under both experimental abdominal sepsis and AP, aged C57BL/6 mice (23–25 months old) as compared to young adult (4–6 months) and middle-aged mice (12–14 months), exhibit significantly increased mortality rates accompanied with enhanced fibrin deposition in multiple organs, most notably lung and kidney. The age-dependent increase in mortality and thrombosis is evident in polymicrobial abdominal sepsis, sterile endotoxemia, and caerulein-induced AP models.
During both sepsis and AP, young mice show signs of mild coagulation, such as increased plasma levels of d-dimer and plasminogen activator inhibitor-1 (PAI-1); however, even under severe lethal sepsis, young mice hardly develop DIC as confirmed by the absence of fibrin deposition.
Augmented production of PAI-1 is a characteristic of aged mice during sepsis and AP. PAI-1 is expressed most strongly in visceral adipose tissues among major organs, and the age-associated increase in PAI-1 expression in the adipose tissue is independent from obesity. High levels of PAI-1 may impair fibrin degradation resulting in enhanced intravascular fibrin deposition. However, plasma d-dimer levels in the aged were equivalent or higher compared to those of young mice during sepsis and AP, suggesting that aged animals with high levels of induced PAI-1 may still maintain fibrinolysis capability to some extent.
During an inflammatory response, thrombin and thrombomodulin convert protein C (PC) to activated PC (aPC) which is a potent anti-coagulant and anti-inflammatory factor. However, in contrast to young mice, aged mice cannot produce aPC efficiently. The blunted low level of aPC in the aged is likely to contribute to enhanced DIC during sepsis. Age-associated down-regulation of thrombomodulin may be one of the mechanisms that partly explain the suppression of aPC production in the aged.
EXPERIMENTALLY APPROACHING THE TRAUMA-INDUCED COAGULOPATHY Wenjun Martini. Hemostasis – Damage Control Resuscitation, US Army Institute of Surgical Research, Fort Sam Houston, TX, USA
Trauma injury is one of the leading causes of death, with uncontrolled hemorrhage as one of the main preventable causes for the mortality. Hypothermia, acidosis, and resuscitative hemodilution have been considered as significant contributors to hemostatic manifestations following trauma. Over the past decade, clinical data showed that coagulopathy may be present at hospital admission in some severely injured trauma patients, extending our recognition of trauma induced coagulopathy to shortly after injury. The high mortality from trauma induced coagulopathy has initiated tremendous interest and effort in trauma community to improve our understanding of its complex pathophysiology, using experimental approaches and clinical trials. This presentation will review contributions of animal experiments to reveal the underlying mechanisms and test potential effective treatments to restore hemostasis. The dynamic and multifactorial nature of trauma induced coagulopathy will be discussed with the effects of injury, shock, hypothermia, acidosis, fluid resuscitation, and fibrinogen concentrate supplementation on changes of endogenous fibrinogen metabolism, coagulation and hemodynamics. Although some mechanisms have been characterized, continuous effort is warranted to improve and translate our knowledge for better clinical outcomes in trauma patients.
PERSISTENT INFLAMMATION, IMMUNOSUPPRESSION AND CATABOLISM SYNDROME: A NEW FORM OF CHRONIC CRITICAL ILLNESS Lyle L. Moldawer. Department of Surgery, University of Florida College of Medicine, Gainesville, FL, USA
Over the past four decades the epidemiology of multiple organ failure (MOF) has evolved as a result of advances in care. A series of paradigms have been described to explain the pathophysiology of the new emerging predominant phenotypes. With the most recent improvements in the delivery of critical care, patients are less-frequently expiring early in their clinical course and in-hospital MOF-related mortality is on the decline. Unfortunately, this has resulted in a dramatic increase in the number of chronically critically ill patients (CCI) who linger in the intensive care unit (ICU), have high resource utilization, are discharged to non-home locations, experience sepsis recidivism requiring readmission, have persistent cognitive and functional impairments, and poor long-term survival. Within this population, we have proposed that a substantial subset of these patients suffer from a new phenotype termed Persistent Inflammation, Immunosuppression, and Catabolism Syndrome (PICS) which underlies these poor outcomes. While the mechanism(s) of PICS are under investigations, there is evidence that myelodysplasia with expansion of myeloid derived suppressor cells, innate and adaptive immune suppression and protein catabolism with malnutrition are major contributors. Optimal care of these patients will require a novel multimodality intervention using pharmacotherapy, physiotherapy and nutritional support.
CONCOMITANT INFLAMMATION AND IMMUNOSUPPRESSION Jean-Marc Cavaillon. Cytokines & Inflammation Unit, Institut Pasteur, Paris, France
Immediately after the occurrence of an insult, either sterile or infectious, an inflammatory response is initiated. During trauma, hemorrhagic shock, ischemia/reperfusion, severe surgery, pancreatitis however, a systemic inflammatory response syndrome (SIRS) is elicited. Similarly during sepsis, although SIRS does not belong to the new definition of sepsis (sepsis 3.0) anymore, systemic inflammation can be observed, which corresponds to an innate immune response. Soon after the beginning of SIRS, numerous mechanisms will be initiated to prevent an overzealous and deleterious inflammatory response. Accordingly, in most clinical settings, one can consider that both processes are almost concomitant. The price to pay is named compensatory anti-inflammatory response syndrome (CARS), which is often considered to reflect a global immunosuppression. But the word is far too excessive, since there is no global defect: some activities are maintained or even enhanced (e.g. phagocytosis, anti-microbial activity), and the most appropriate terms would be reprogramming. Leukocyte reprogramming is the consequence of inhibitory signals delivered at the cell surface (anti-inflammatory cytokines, activation of alpha-7 nicotinic receptor and resolvin receptor, and PD1/PDL1 cross-talk). It also involves numerous intracellular players (miRNA, negative signaling molecules), and events that are occurring within the nucleus (binding of the glucocorticoid-receptor complexes, of the aryl hydrocarbon receptor (AhR) linked with its nuclear translocator (ARNT), and epigenetic regulation). Importantly, the immune status of circulating leukocytes is completely different from that observed within tissues where the inflammatory response may predominate over the counter-regulatory mechanisms. For example, hemophagocytosis has been regularly observed in bone marrow of SIRS and sepsis patients. Despite the fact that we claimed 15 years ago that both SIRS and CARS are concomitant (J. Endotoxin Res. 2001, 7, 85), numerous reviews in most prestigious journals have continued to provide figures where CARS follows SIRS. Of note, when the concept of persistent inflammation, immunosuppression, and catabolism syndrome (PICS) was first proposed (Gentile et al. J Trauma Acute Care Surg 2012, 72, 1491), the authors fully integrated that at the very beginning, SIRS and CARS are concomitant. Indeed, this has been repeatedly illustrated in patients with sepsis, trauma, surgery, or resuscitated after cardiac arrest in studies of the ex vivo capacity of the cells to produce cytokines, in analyses of the circulating cytokines, of the HLA-DR expression and of the gene transcriptomes.
BENCH TO BEDSIDE RESEARCH ON CRITICAL ILLNESS MYOPATHY (CIM) AND VENTILATOR INDUCED DIAPHRAGM MUSCLE DYSFUNCTION (VIDD): MECHANISMS AND INTERVENTIONS Lars G. Larsson. Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden
Significant improvements in modern critical care related to technological advances, improved understanding of the pathogenesis of disease process, new therapies and the removal of inefficient/harmful interventions have led to improved survival from critical illness. However, the improved survival is associated with an increased number of patients with complications related to modern critical care. Severe muscle wasting and impaired muscle function are frequently observed in immobilized and mechanically ventilated intensive care unit (ICU) patients. Approximately 30% of mechanically ventilated and immobilized ICU patients for durations of 5 days and longer develop generalized muscle paralysis of all limb and trunk muscles, a condition known as CIM. Mechanical ventilation is a lifesaving treatment in critically ill ICU patients; however, the being on a ventilator creates dependence, and the weaning process occupies as much as 40% of the total time of mechanical ventilation. Furthermore, 20–30% of patients require prolonged intensive care due to VIDD, resulting in poorer outcomes, and greatly increased costs for health care providers. CIM and VIDD in ICU patients may be related to the primary disease, but there is heterogeneity of underlying disease and pharmacological treatment among patients exhibiting similar outcomes. Thus, it is highly likely that a common component of ICU treatment per se is directly involved in the progressive impairment of muscle function and muscle wasting during long-term ICU treatment. The specific mechanisms underlying the muscle wasting and impaired muscle function associated with the ICU intervention are poorly understood in the clinical setting. There is, accordingly, compelling need for experimental animal models closely mimicking the ICU condition, including long-term exposure to mechanical ventilation and immobilization. In this project, the muscle dysfunction, which by far exceeds the loss in muscle mass in limb and respiratory muscles in patients with CIM and VIDD have been investigated in detail at the cellular and molecular levels in rodent and porcine experimental ICU models, allowing detailed studies in immobilized and mechanically ventilated animals for long durations. Results demonstrate that the motor protein myosin is highly involved in the pathogenesis of both CIM and VIDD, but mechanisms are different. In CIM there is a preferential loss of myosin due to transcriptional down-regulation and enhanced degradation while post-translational modifications of myosin play a significant role for the diaphragm muscle dysfunction in VIDD. Specific intervention strategies targeting the mechanisms underlying CIM and VIDD will be presented and the translation of these interventions to the clinic.
INFLUENCE OF AGE ON PICS Philip A. Efron. University of Florida, Gainesville, FL, USA
The phenotype of the intensive care unit (ICU) patient has changed over the last several decades. Now, patients with Chronic Critical Illness and/or the Persistent Inflammation, Immunosuppression and Catabolism Syndrome (PICS) are becoming the main cohort after the critically ill are admitted to the hospital. These patients have increased morbidity and in-hospital and post-discharge one to two year mortality, as well as having significantly increased cost to their treatment. Interestingly, the elderly demonstrate the same characteristics prior to hospitalization, including chronic low grade inflammation, immunosuppression and catabolism. This presentation will explore how the elderly are at elevated risk for requiring prolonged ICU care, for PICS and for poor outcomes. It will review how the elderly have a fundamentally different biological response to severe inflammation than their younger counterparts. This is associated with subsequent worse morbidity and mortality in aged populations. This session will also summarize some of the science regarding the elderly response to sepsis, trauma and burns. In turn, this will hopefully allow practitioners to alter their future approach at the bedside to these patient populations, as well as the researcher's approach to the study of infection, injury and shock in the elderly so that these patients can improve their long term mortality and quality of life.
LONG TERM COGNITIVE AND FUNCTIONAL DECLINES FOLLOWING SEPSIS Nicholas Heming. Raymond Poincaré Hospital (APHP), University of Versailles, Garches, France
A growing number of patients who survive sepsis in the short term and are discharged home, thanks to improved quality of care of the critically ill. However, roughly one out of two of these survivors may present with long-term neurological sequelae, particularly altered cognitive function and neuromuscular weakness. The main underlying mechanisms for sepsis induces cognitive dysfunction, may include both vascular damage and inflammation of the brain. Brain's dysfunction in sepsis is likely mediated by metabolism disorders (e.g. dysglycemia, dysnatremia, hypoxia) and overwhelming inflammation which result from disruption of blood brain barrier, oxidative stress and severe microglial activation, particularly within the limbic system. Several interventions that may restore the blood brain barrier, reduce glial activation and oxidative stress, have shown favorable effects in preventing cognitive dysfunction in various experimental models of sepsis. Except adequate control of metabolic disorders and prevention of vascular damage, there is so far no routine specific treatment for cognitive dysfunction after sepsis.
INFLAMMATION ASSOCIATED EPIGENETICS AND METABOLISM: NOVEL TREATMENT TARGETS Charles E. McCall. Department of Molecular Medicine, Wake Forest University Medical Center, Winston Salem, NC, USA
Premise: Severe systemic inflammatory responses often overcome evolution's intent to defend, mend, and restore homeostasis. This cell and organism extreme stress state with lethal organ dysfunction and infection, called sepsis, remains a major cause of mortality. No molecular-based treatments are available. Mounting data support that lethal organ failure occurs when the nutrient and energy demands from sepsis cannot be met. During this time, inflammation-directed bioenergetics rapidly progresses through a transient anabolic glucose dependent activation state to a sustained phenotype of catabolic inertia. Immunometabolic paralysis accompanies catabolic inertia, which is most life threatening when mitochondrial bioenergy reserves are marginal, such as aging and chronic inflammatory and metabolic diseases, where mortality rates often exceed 50%.
Novel Treatment Targets: We have overcome catabolic inertia, reversing immunometabolic paralysis and markedly improving survival in septic mice in two ways: 1) Inhibiting nuclear NAD+-dependent nuclear sirtuin 1 (SIRT1) epigenetic control over homeostasis; 2) Inhibiting mitochondrial pyruvate dehydrogenase kinase 1 (PDHK1) posttranslational control over pyruvate dehydrogenase (PDH)-dependent fueling the Krebs Cycle and oxidative phosphorylation. Both treatments similarly promote immune, metabolic, and mitochondrial energy homeostasis and require only a single drug dose delivered after anabolic activation switches to catabolism.
Conclusion: An adjustable and highly integrated immune, metabolic, and bioenergy homeostasis network regulates the sepsis stress response and may provide novel checkpoint-based treatments. Biomarkers are urgently needed to identify the right way and time to treat this global killer.
Support: NIH grants RO1 s AI079144, AI065791, GM102497, GM099807
SEPSIS NEW DEFINITIONS: WHAT CHANGES? Clifford S. Deutschman. Center for Pediatric Research, The Feinstein Institute for Medical Research, Manhasset, NY, USA
Abstract not available yet.
CURRENT SITUATION OF DIAGNOSIS AND TREATMENT OF SEPSIS IN JAPAN Hiroyasu Ishikura. Department of Emergency and Critical Care Medicine, Faculty of Medicine, Fukuoka University, Fukuoka, Japan
Sepsis is often a complication that occurs in the clinical course of medical and surgical patients treated for other diseases, and is a global health problem that carries a high risk of death. A recent global assessment of the mortality rate of patients with sepsis treated in an intensive care unit found that over 30% of these patients died without leaving hospital.
Additionally, inflammation and coagulation play pivotal roles in the pathogenesis of sepsis. So the majority of ill patients with sepsis present with coagulation abnormalities and sepsis is most common disease associated with disseminated intravascular coagulation (DIC). Approximately 20–40% of all sepsis patients are complicated with DIC. And the mortality rate of sepsis patients complicated with DIC is clearly higher than that of patients without DIC. Although the mortality rate of septic shock has tended to gradually decrease during the past decade because of significant technological advances in supportive therapies, it remains high, and effective specific diagnosis and treatments are still very limited.
To improve clinical management and outcome of critically ill patients, the Surviving Sepsis Campaign (CCS): International Guidelines for Management of Severe Sepsis and Septic Shock were published approximately a decade ago and were most recently revised in 2012. On the other hand, in Japan, an original guideline for the management of sepsis was published in 2013. This Japanese Guideline was developed on the basis of Japanese evidence-based medicine and focuses on unique treatments that have not been mentioned in the international SSC guidelines (SSCG), as well as treatments that are viewed differently in Japan and in Western countries.
In this time we want to introduce about Japanese, may be Japanese original, diagnosis and treatment of sepsis.
In regards to the diagnosis of sepsis various biomarkers (e.g. CRP, IL-6, PCT) have been studied until now. Most recent years Presepsin (PSEP) is collecting a lot of attention as diagnosis marker of sepsis in Japan.
For the treatment, direct hemoperfusion with polymyxin B-immobilized fiber (PMX-DHP), a technique to remove circulating endotoxin, and/or intravenous immunoglobulin (IVlg) have been proposed as an adjuvant therapy for sepsis again in Japan. Moreover, sepsis induced DIC treatment is provided actively using recombinant human soluble thrombomodulin.
THE IMPACT OF SEPSIS IN USA Philip A. Efron. University of Florida, Gainesville, FL, USA
Abstract not available yet.
SEPSIS IN DEVELOPING COUNTRIES Reinaldo Salomao, Luciano C. Azevedo, Flavia R. Machado. Infectious Diseases/Internal Medicine, Escola Paulista de Medicina, Universidade Federal de São Paulo, São Paulo, Brazil
Incidence of sepsis is associated to social and biological factors, and outcomes, deeply influenced by timely diagnosis and therapy, are affected by access to health care facilities, including emergency rooms and intensive care units. Thus, the global estimative of 20–30 million cases of sepsis per year, certainly represent a heterogeneous burden for high income and low to medium income countries. Notably, estimates of worldwide incidence of sepsis usually lack epidemiological data from developing countries.
Two epidemiological studies evaluated incidence and outcomes of sepsis in Brazilian adults ICUs in the last decade, one including 5 and the other 75 units. The incidence density rates for sepsis, severe sepsis and septic shock in the first study were 61.4, 35.6 and 30.0 per 1000 patient-days, respectively. Both studies showed mortality rates around 50% for severe sepsis and septic shock.
To obtain nationwide epidemiologic data, the Latin American Sepsis Institute (LASI - www.ilas.org.br) conducted a one day national survey (SPREAD - Sepsis PREvalence Assessment Database) with strata created based on geo-economic region, stratified by type of institution and ICU size. The study included 229 ICUs and revealed that around 30% of ICU beds were occupied by septic patients, with a mortality rate of 55.7%. The LASI database, which includes over 30,000 septic patients, also reinforce the Brazilian high mortality rates for sepsis of 42.2%, higher in public than in private hospitals. In a retrospective analysis of Brazilian multiple-cause-of-death data the number of sepsis associated deaths increased both in absolute numbers and proportions from 95,972 (9.77% of total deaths) in 2002 to 186,712 deaths (16.46%) in 2010.
Sepsis is also related to increased costs, an additional burden for healthcare providers. One study estimates the costs of sepsis during ICU stay in U$ 9632 dollars per patient in Brazil, the median daily costs higher for non-survivors than for survivors (U$ 1,094.00 and U$ 826, respectively).
Quality improvement programs are of paramount importance in such adverse scenario and have been a priority for LASI that trained over 150 institutions since 2005. For the global LASI database, sepsis mortality in our country reduced from 54% to 40% between 2005 and 2015. Positive results were also obtained in a private network of hospitals in Sao Paulo, where implementation of protocols was associated with a reduction of sepsis mortality from 55% to 26%. Besides reducing mortality this intervention was also cost-effective.
In conclusion, sepsis pursues an increased burden in low to medium income countries. Generation of own clinical and epidemiological data, as well as efforts in education and health care are of pivotal importance to design and implement strategies to improve sepsis outcomes.
A NOVEL THERAPEUTIC TARGET FOR SEPSIS Ping Wang, Weng-Lang Yang. Center for Immunology & Inflammation, The Feinstein Institute for Medical Research, Manhasset, NY, USA
From recent the third international consensus definitions, sepsis should be defined as life-threatening organ dysfunction caused by a dysregulated host response to infection (JAMA, 2016). In the United States, approximately 750,000 cases of sepsis occur each year, of which at least 225,000 are fatal. Despite numerous studies aimed at advancing the understanding of the pathophysiology of sepsis and developing therapeutic candidates for clinical trials, currently no effective pharmacotherapy exists for the treatment of sepsis. The failure of clinical trials can be attributed to the heterogeneous medical conditions of enrolled patients as well as the lack of adequate animal models for evaluating drug efficacy. So far, sepsis has been referred to as the “graveyard” for pharmaceutical companies. However, there is still an urgent, unmet medical need for developing therapies to treat septic patients. Identification of novel therapeutic targets will bring the hope and excitement for sepsis research field and is needed for drug discovery. Recently, our laboratory has discovered that cold-inducible RNA-binding protein (CIRP) can act as damage-associated molecular pattern (DAMP) molecule in animal models of hemorrhage and sepsis. The serum levels of CIRP are elevated in the patients admitted to the surgical intensive care unit with shock. Extracellular CIRP stimulates the release of proinflammatory cytokine tumor necrosis factor-alpha from macrophages and induces inflammatory responses and causes tissue injury when injected to healthy animals. When CIRP activity is blocked by neutralizing antisera to CIRP, it attenuates inflammatory responses and improves the survival of sepsis and shock animals. Thus, CIRP may be targeted therapeutically to reduce morbidity and mortality in patients with sepsis and shock.
TARGETING HMGB1 IN THE TREATMENT OF SEPSIS Haichao Wang. Laboratory of Emergency Medicine, The Feinstein Institute for Medical Research, Manhasset, NY, USA
Sepsis refers to a systemic inflammatory response syndrome resulting from microbial infections, and its pathogenesis is partly attributable to dysregulated inflammatory responses orchestrated by various types of innate immune cells (e.g., macrophages and monocytes) which sequentially release early (e.g., TNF, IL-1, and IFN-gamma) and late (e.g., HMGB1) pro-inflammatory mediators. As a ubiquitous nuclear protein, HMGB1 is constitutively expressed in innate immune cells, and can be actively secreted in response to exogenous pathogen-associated molecular pattern molecules (PAMPs, e.g., ds-RNA, CpG-DNA and endotoxin) or endogenous cytokines [e.g., interferon (IFN)-gamma or IFN-beta]. In addition to active secretion, HMGB1 can also be passively released from damaged cells following ischemia/reperfusion, trauma, or toxemia, thereby serving as damage-associated molecular pattern molecule (DAMP). At a late stage of microbial infection, the PAMP-elicited inflammatory response may be accompanied by an unintended cell injury and DAMP release that further amplifies the cytokine storm to precipitate organ dysfunction. Here we discuss the evidence that support extracellular HMGB1 as a late mediator of inflammatory diseases, and discuss the potential of several HMGB1-targeting therapies in animal models of lethal sepsis. Although microbial infection-induced sepsis is indistinguishable from sterile injury-elicited systemic inflammatory response syndrome, it may be more advantageous to develop strategies that specifically attenuate DAMP-mediated inflammatory responses without compromising the PAMP-mediated innate immunity.
EFFECT OF HIGH MOBILITY GROUP BOX-1 PROTEIN (HMGB1) ON HOST IMMUNE RESPONSE AND ITS REGULATORY MECHANISM AFTER MAJOR BURNS Yong-Ming Yao. Trauma Research Center, First Hospital Affiliated to the Chinese PLA General Hospital, Beijing, China
With both animal experiments and clinical investigations, current studies were conducted to explore the potential role and its regulatory mechanism of high mobility group box-1 protein (HMGB1) in host immune dysfunction after major trauma/burns. It was found that HMGB1 formation could dendritic cells (DCs). Excessive release of HMGB1 induced CD4+CD25+ Tregs to maturation, and enhanced the marked differentiation of IL-10-producing CD11clowCD45RBhigh DCs, thereby mediating immunosuppression of T lymphocytes. It was noticed that HMGB1 could stimulate the activities of CD4+CD25+ Tregs as well as DCs via binding receptor for advanced glycation end products (RAGE) on the surface of cells, and trigger shift of helper T cell (Th)1 to Th2 with down-regulation of T lymphocyte immune response following acute insults. Furthermore, gene silence of X-box binding protein 1 (XBP-1) in splenic DCs decreased the levels of CD80, CD86 as well as major histocompatibility complex (MHC)-II expression and cytokine secretion after HMGB1 treatment, when compared with untransfected or nontargeting-transfected DCs. XBP-1 silenced DCs showed immunosuppressive or tolerogenic DCs and failed to stimulate notable proliferation response of T cells, even after treatment with HMGB1, suggesting an important role for endoplasmic reticulum stress (ERS) in HMGB1-induced maturation and activation of DCs. While treatment with HMGB1 inhibitors, the proliferative activation of splenic T lymphocytes increased from the 1st day to the 7th day after thermal injury, meanwhile, the expression levels of CTLA-4, Foxp3 on CD4+CD25+ Tregs and IL-10 production were markedly decreased after major burns. In addition, mortality rates of animals were significantly reduced compared to those without treatment with HMGB1 inhibitors. Clinical data showed that plasma HMGB1 levels were markedly elevated on postburn day 1 in severely burned patients, of them HMGB1 levels were obviously higher in sepsis group than in non-sepsis group, and HMGB1 levels were negatively correlated with T cell immune function, including lymphproliferation response, IL-2 release, and the ratio of CD4+/CD8+ T lymphocytes in patients suffering from extensive burns. These data proved that HMGB1 is not only identified as a novel late inflammatory mediator but also closely associated with immune depression in the setting of major burns.
FROM PAMPS AND ALARMINS TO TOLEROGENS IN SHOCK/SEPSIS: WHAT WE ARE LEARNING FROM INNATE REGULATORY LYMPHOCYTES Alfred Ayala, Tristen T. Chun, Eleanor A. Fallon, Whitney A. Young, Chun-Shiang Chung, Daithi S. Heffernan. Department of Surgery, Rhode Island Hospital; Division of Surgical Research, Brown University, Providence, RI, USA
Sepsis, a leading cause of death worldwide, is thought to involve expression of an overzealous inflammatory response (increased systemic/local pro-inflammatory cytokine/chemokine/mediator levels, etc.) and the concomitant development of an ineffective functional innate as well as adaptive immune response (decreased cytokine release capacity/decreased microbial phagocytosis/reduced antigen presenting capacity, reduced lymphocyte number [via altered cell death mechanisms], increased check-point protein/co-inhibitory molecule expression, etc.). Both the extent to which these aspects of the innate immune response diverge from a homeostatic response (either an inappropriately low response or the inability to rein in such responses as needed), as well as the sustained duration of this state of dys-homeostasis; appear to contribute to the morbid status of the septic patient/experimental animal. While much is understood about how sensing of PAMPs, DAMPs and/or Alarmins by a diverse family of pattern recognition receptors on various cellular components of the innate immune response, such as myeloid cells, phagocytes, etc., drives pro-inflammation; we have only recently begun to appreciate the role a novel family of innate regulatory lymphocytes (a sub-population of cells that are also responsive primarily to select novel PAMPs/DAMPs) appear to be able to play in shaping the extent and/or duration of the septic innate immune response. Here we will discuss some of our recent findings examining the contributions of a few of these innate regulatory lymphoid cell sub-sets, e.g., gamma/delta-T-cells, invariant NKT-cells (iNKT), as well as the type-2 innate lymphoid cells, in the response to experimental sepsis and the evidence that they may be affecting similar responses in critically ill humans. Finally, using the iNKT-cell as an example, we will consider how a tolerogen such as Programmed Cell Death Receptor-1 ligand (PD-L1), which is upregulated on these cells, may be playing a role outside its more classically defined adaptive immune response T-cell/check point protein biology, to regulate the innate septic immune response.
Supported by NIH R35 GM-118097 [AA] & NIH K08 GM-110495 [DSH] and an Armand D. Versaci Research Scholar in Surgical Sciences Fellowship award [TTC, EAF, WAY]
MOUSE SEPSIS MODELS IN THE POST-PNAS REALITY: STILL VIABLE OR UTTERLY HELPLESS? Marcin F. Osuchowski. Department of Intensive Care, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Vienna, Austria
The mouse has been on the forefront of pre-clinical sepsis research. The vast majority of animal sepsis experiments are performed in mice and the most wide-ranging selection of sepsis models is available in this species. Mice have been employed to study the multifaceted pathogenesis of sepsis syndromes and related inflammatory conditions as well as to verify efficacy of multiple experimental therapeutics against those illnesses. Yet, the popularity of mice in pre-clinical sepsis field is matched by the controversy regarding their translational utility. The two recent articles (2013 Seok et al.; 2015 Takao and Miyakawa) published in the Proceedings of the National Academy of Sciences of the United States of America (PNAS) expose the complexity of the mouse-to-human data transpolation. Namely, cross-comparison (using the same dataset) of the mouse and human gene expression responses in circulating leukocytes in burns, trauma, sepsis, endotoxemia led to two contradictory conclusions regarding the translational value of the mouse data. Given the rich body of available scientific evidence, it is plausible to state that the translational utility of the existing mouse models can be both excellent as well as poor. The final success or failure of mouse sepsis experimentation depends on many objective and subjective factors. It is clear that the mouse-human species gap is vast and similarities in investigated pathophysiological traits cannot be reflexively assumed given that some of them simply do not exist and others overlap only partially. On the other hand, high-quality sepsis modeling incorporating superior study design has demonstrated many solid similarities in mouse-versus-human immuno-inflammatory responses. A responsible and balanced approach towards modeling and subsequent data interpretation is the key to the effective use of mouse-based experimentation in the field of sepsis and related critical care conditions. Additionally, it is advisable to inject more clarity and direction into the complex and often confusing field of pre-clinical mouse modeling. This could be achieved, for example, by generating consensus guidelines that would support scientists in their study design and modeling decision-making. An implementation of such hypothetical “Minimum Modeling information in Pre-clinical Sepsis Studies” (MMiiPSS) guidelines, either just for the mouse or beyond, has a strong potential for making mouse sepsis studies more reliable and transpolatable.
IMPACT OF IMMUNOSENESCENCE ON SEPSIS: T CELL EXHAUSTION AND SECONDARY INFECTION AFTER SEPSIS IN THE ELDERLY Shigeaki Inoue, Nobuo Watanabe, Bunsei Yamamoto, Sadaki Inokuchi. Tokai University School of Medicine, Kanagawa, Japan
Background: The expanding aging population in the world will increase the incidence and mortality of sepsis. The aim of the presentation is to review the pathophysiological differences in sepsis and its clinical impact on the elderly. The impact of immunosenescence on acquired immunity is associated with relative immunosuppression that may favor the spreading of inflammation.
Methods: Fifty-five patients with severe sepsis and 30 healthy donors were prospectively enrolled, and 90-day survival was compared between elderly (≥65 years) and adult (18–64 years) septic patients. Within 24 h after diagnosis of severe sepsis, peripheral blood mononuclear cells were stimulated ex vivo to measure expression of the activation maker CD25 in T cells, IL-2 levels in the supernatant, and proliferation. In the mouse study, young (6–8 weeks) and aged (20–22 months) C57/B6 mice were subjected to cecal ligation and puncture (CLP), and survival was compared after 7 days. Expression of the negative co-stimulatory molecules, CD25, and IL-2 in CD4+ T cells was measured.
Results: The survival rate in elderly sepsis patients and aged septic mice was significantly lower than that in adult patients and young septic mice (60% vs. 93% in septic patients, 0% vs. 63% in septic mice, P < 0.05). Expression of negative co-stimulatory molecules in CD4+ T cells in the spleen, lymph nodes, and peripheral blood was significantly higher in aged mice than in young mice (P < 0.01). Ex vivo stimulation decreased CD25 expression, IL-2 production, and proliferation to a greater extent in CD4+ T cells from elderly patients and aged septic mice than in those from adult patients and young septic mice. Elderly patients demonstrated increased detection of gram-negative bacteria at days 14–16 and 28–32 after sepsis (P < 0.05).
Conclusions: T cell exhaustion with increased secondary infection may be associated with decreased survival in elderly patients and mice after sepsis. The specific immunological changes underlie the increased mortality in such patients and prompts research on therapeutic strategies with particular benefits to elderly septic patients.
UNDERSTANDING THE HETEROGENEITY OF THE HOST RESPONSE TO INFECTION MAY DIRECT THERAPY Daniel Remick. Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA, USA
The host response to a septic insult is best described as nonlinear and complex. The recent sepsis-3 lay definition that sepsis is a life-threatening condition that arises when the body's response to an infection injures its own tissues and organs provide a context for further understanding. Organ and tissue injury may occur, but there is a spectrum of host responses. Prior attempts to improve sepsis survival through global immunosuppression with glucocorticoids or inhibiting specific inflammatory mediators have generally not been successful. The concept that one-size-fits-all is the appropriate approach to the therapeutic treatment of sepsis has probably been guided by prior publications demonstrating that in animal models of acute, lethal sepsis there is an early, robust, and potentially injurious inflammatory response. In real life, the septic response is not always injurious. During the 21st century the survival of septic patients is approximately 75%. This indicates that 3 out of 4 septic patients mount an appropriate response to clear the offending pathogen without causing undue injury to the host. Virtually all prior sepsis trials used the nonspecific enrollment criteria of the systemic inflammatory response syndrome. Precision medicine is currently being widely viewed as a major advance for the treatment of cancer. Within intensive care units there is a precision medicine approach where physiologic monitoring directs fluid resuscitation and organ support such as dialysis or mechanical ventilation. Understanding the individual patient's host response may be used to direct immunotherapy. Septic patients may be divided into 3 broad groups, with current limited understanding about an individual patient's need for therapy. Those patients with an exuberant, organ injury response may benefit from modest immunosuppression. Those patients with an appropriate response which clears the pathogens while inducing limited organ injury may not require any immunotherapy. The final group would be those patients who fail to mount a sufficiently robust inflammatory response and subsequently succumbed to overwhelming infection. The murine model of cecal ligation and puncture may be used to identify the individual host response to a severe bacterial infection.
ARTESUNATE PROTECTS AGAINST THE ORGAN INJURY AND DYSFUNCTION INDUCED BY SEVERE HEMORRHAGE AND RESUSCITATION Chris Thiemermann1, Regina Sordi1, Kieran Nandra1, Fausto Chiazza2, Florence Johnson1, Claudia Cabrera1, Hew Torrence1, Noriaki Yamada1, Mike Barnes1, Karim Brohi1, Massimo Collino2. 1William Harvey Research Institute, Queen Mary University of London, London, UK, 2Department of Drug Science and Technology, University of Turin, Turin, Italy
Rational: HS is still a common cause of death in severely injured patients and is characterized by impairment of organ perfusion, systemic inflammatory response, and multiple organ failure. There is no specific therapy that reduces organ injury/dysfunction. Artesunate exhibits pharmacological actions beyond its antimalarial activity, such as anticancer, antiviral, and anti-inflammatory effects.
Objectives: To evaluate the effects of artesunate on organ injury and dysfunction associated with hemorrhagic shock (HS) in the rat.
Methods: Male Wistar rats were submitted to HS. Mean arterial pressure was reduced to 30 mm Hg for 90 inutes, followed by resuscitation. Rats were randomly treated with artesunate (2.4 or 4.8 mg/kg i.v.) or vehicle upon resuscitation. Four hours later, parameters of organ injury and dysfunction were assessed.
Results: Artesunate attenuated the multiple organ injury and dysfunction caused by HS. Pathway analysis of RNA sequencing provided good evidence to support an effect of artesunate on the Akt-survival pathway, leading to downregulation of interleukin-1 receptor-associated kinase 1. Using Western blot analysis, we confirmed that treatment of HS rats with artesunate enhanced the phosphorylation (activation) of Protein kinase B (Akt) and endothelial nitric oxide synthase and the phosphorylation (inhibition) of glycogen synthase kinase-3β (GSK-3β). Moreover, artesunate attenuated the HS-induced activation of nuclear factor kappa B and reduced the expression of proinflammatory proteins (inducible nitric oxide synthase, tumor necrosis factor-α, and interleukin 6).
Conclusions: Artesunate attenuated the organ injury/dysfunction associated with HS by a mechanism that involves the activation of the Akt-endothelial nitric oxide synthase survival pathway, and the inhibition of glycogen synthase kinase-3β and nuclear factor kappa B. A phase II clinical trial evaluating the effects of good manufacturing practice-artesunate in patients with trauma and severe hemorrhage is planned.
THE EFFECTIVE THERAPY USING ENDOTOXIN TOLERANCE AGAINST LETHAL SEPTICEMIA Manabu Kinoshita. Department of Immunology and Microbiology, National Defense Medical College, Saitama, Japan
Objectives: We investigated the effective therapy using in vivo endotoxin (LPS) tolerance against lethal septicemia, focusing on the alteration of Kupffer cell subsets.
Methods: LPS tolerance was induced in mice by intraperitoneal injections with 5 μg/kg of LPS for three consecutive days, then the mice were infected with Escherichia coli intravenously.
Results: All LPS-primed mice survived lethal bacterial infection (100% survivals vs. 0%). Drastic enhancement of bactericidal activity of Kupffer cells strongly contributed to bacterial clearance. Although the LPS-primed mice produced substantial amounts of TNF inside the liver, TNF efflux into the systemic circulation via the hepatic vein was markedly suppressed. These mice showed a dramatic increase in the number of CD11b+ recruited Kupffer cells in the liver. While potent TNF-producing capacity and TLR4 expression were suppressed in those CD11b+ Kupffer cells, they showed strong bactericidal activity with up-regulated expression of Fcγ receptor I. LPS priming did not affect the CD68+ resident Kupffer cell population, and CD68+ Kupffer cell-depleted mice still showed LPS tolerance with strong resistance to bacteremia.
Conclusions: LPS priming recruited CD11b+ Kupffer cells with enhanced bactericidal activity and low TNF production to the liver, which played a central role in the resistance to lethal bacteremia. In vivo LPS tolerance is a quite beneficial status that could be used for the treatment of bacterial septicemia.
POST-CARDIAC ARREST SYNDROME: THE NEW SCIENCE Robert W. Neumar. Department of Emergency Medicine, University of Michigan Medical School, Ann Arbor, MI, USA; International Liaison Committee on Resuscitation
The first ILCOR Scientific Statement on Post-Cardiac Arrest Syndrome was published in 2008, defining both the pathophysiology and treatment strategies based the available science. The key components of post-cardiac arrest syndrome were defined as 1) post-cardiac arrest brain injury, 2) post-cardiac arrest myocardial dysfunction, 3) systemic ischemia/reperfusion response, and 4) persistent precipitating pathology. The statement also highlighted important scientific knowledge gaps. Since that time, significant laboratory and clinical research has been published that has expanded our understanding of the pathophysiology of post-cardiac arrest syndrome and refined the therapeutic approach. Despite these advances, important knowledge gaps persist. This presentation will provide an update on the significant laboratory and clinical research that has been published since the original ILCOR Post-Cardiac Arrest Syndrome Scientific Statement, and highlight the most important knowledge gaps that need to be addressed through new scientific investigation.
MOLECULAR MECHANISMS OF ISCHEMIC BRAIN DAMAGE Hiroyuki Uchino, Shusuke Sekine, Naomi Hara, Miyuki Chijiiwa, Yosuke Fujita, Tomoki Nagakura, Takayuki Kobayashi, Eisuke Mutoh, Toshio Okada, Hidekimi Fukui, Yukihiko Ogihara. Department of Anesthesiology, Tokyo Medical University, Tokyo, Japan
Cardiac arrest induces the cessation of cerebral blood flow, which can result in brain damage. The primary intervention to salvage the brain under such a pathological condition is to restore the cerebral blood flow to the ischemic region. Ischemia is defined as a reduction in blood flow to a level that is sufficient to alter normal cellular function. Brain tissue is highly sensitive to ischemia, such that even brief ischemic periods in neurons can initiate a complex sequence of events that may ultimately culminate in cell death. However, paradoxically, restoration of blood flow can cause additional damage and exacerbate the neurocognitive deficits in patients who suffered a brain ischemic event, which is a phenomenon referred to as “reperfusion injury”. Transient brain ischemia following cardiac arrest results from the complex interplay of multiple pathways including excitotoxicity, acidotoxicity, ionic imbalance, peri-infarct depolarization, oxidative and nitrative stress, inflammation, and apoptosis. The pathophysiology of post-cardiac arrest brain injury involves a complex cascade of molecular events, most of which remain unknown. Many lines of evidence have shown that mitochondria suffer severe damage in response to ischemic injury. Mitochondrial dysfunction based on the mitochondrial permeability transition after reperfusion, particularly involving the calcineurin/immunophilin signal transduction pathway, appears to play a pivotal role in the induction of neuronal cell death. The aim of this article is to discuss the underlying pathophysiology of brain damage, which is a devastating pathological condition, and highlight the central signal transduction pathway involved in brain damage, which reveals potential targets for therapeutic intervention.
POST-CARDIAC ARREST BRAIN INJURY Clifton W. Callaway. Department of Emergency Medicine, University of Pittsburgh, Pittsburgh, PA, USA
After resuscitation from cardiac arrest, brain injury is the most common barrier to good quality of survival. It is useful to understand the clinical features of post-cardiac arrest brain injury and how to adjust medical support of the patient to favor brain recovery. The recovering brain may have electrical disturbances (seizures or bursting patterns), perfusion disturbances (loss of autoregulation and focal hypoperfusion), and physical disturbances (edema and cerebral hypertension). Adequate electrical and hemodynamic monitoring allows clinicians to mitigate secondary injury from these disturbances. Epileptiform discharges are sometimes responsive to anticonvulsant drugs. Adequate or elevated arterial blood pressure can improve brain tissue oxygenation. Careful fluid management and osmotherapy can prevent herniation from brain edema. Brain recovery may be assessed using clinical exam, EEG, neurophysiological testing, blood levels of cerebral proteins, and imaging. Brainstem and subcortical recovery occurs independently of cortical recovery. Recovery of the cortex is essential for cognitive function and supporting a favorable post-arrest quality of life. Assessment of cognitive impairments in patients who do awaken is critical for identifying opportunities for rehabilitation therapies.
CARDIOLOGIST'S PERSPECTIVE OF POST-CARDIAC ARREST SYNDROME Shoji Kawakami, Yoshio Tahara, Teruo Noguchi, Satoshi Yasuda. Department of Cardiovascular Medicine, National Cerebral and Cardiovascular Center, Osaka, Japan
Cardiac arrest is critical presentation of cardiovascular disease and results in subsequent cerebral and cardiovascular dysfunction. Even if patients who initially achieve return of spontaneous circulation, those accompanie prolonged whole-body ischemia and reperfusion injury. The advances in cardiopulmonary resuscitation and post cardiac arrest care including target temperature management and emergent coronary intervention could contribute to the improvement of clinical outcomes in patients with post-cardiac arrest syndrome.
In our institute of the national center in Japan that provides advanced and specialized cardiovascular care, extracorporeal cardiopulmonary resuscitation (ECPR) with/without target temperature management has been introduced by dedicated team. Usually, the veno-arterial bypass for ECPR is percutaneously performed and bypass flow is changed according to patients’ hemodynamics and cardiac performance, measurement by using echocardiography and pulmonary artery catheter. It should be noted that aortic valve ejection time and velocity time integral are useful parameters for repeatedly estimating cardiac performance and timing of weaning from mechanical support.
ECPR was effective to improve the survival rate in patients with in- or out-of-hospital cardiac arrest. In Japan, ECPR has become a common procedure for severe cardiogenic shock or cardiac arrest. However, the proper candidates for ECPR and protocol of withdrawal from ECPR were not well-established. In this session, we present general outline and would discuss clinical point of post-cardiac arrest care with veno-arterial cardiopulmonary bypass.
HOW TO PROVIDE THE CARDIOPROTECTION FOR THE ISCHEMIC HEART - A LESSON FROM THE J-WIND TRIAL - Masafumi Kitakaze. Department of Clinical Medicine and Development, National Cerebral and Cardiovascular Center, Osaka, Japan
Since both mortality and morbidity following acute ischemic heart diseases are high in the developed counties including Japan, the investigation of ischemic heart disease is becoming important worldwide to provide the effective methods for the prevention and treatment. There are three different strategies; the prevention of acute myocardial infarction, the treatment of heart failure following myocardial infarction and the attenuation of infarct size during acute myocardial infarction.
First of all, the plaque rapture causes coronary artery diseases, and endothelial dysfunction is primarily considered as one of its underlying mechanisms. Secondly, the prevention of cardiac hypertrophy and remodeling following acute myocardial infarction is important. Thirdly, we should reduce the infarct size once plaque of coronary artery is ruptured. However, there is no drug for the reduction of infarct size after the successful percutaneous coronary intervention.
Following our extensive research on the cellular mechanisms of ischemia and reperfusion injury, we culminated in the idea that both adenosine and NO are potential cardioprotective endogenous substances. Since atrial natriuretic peptide (ANP) shares the same signaling pathway with NO, and KATP channel opening is located at the downstream of adenosine, we evaluated the effects of either ANP or the KATP channel opener (nicorandil) on infarct size and CV outcome. In 2 independent prospective, blinded, placebo-controlled, randomized studies conducted at 65 hospitals, AMI subjects undergoing reperfusion therapy received ANP or nicorandil or the matching placebo. Overall, 603 subjects received ANP or placebo, and 613 subjects received nicorandil or placebo. ANP, but not nicorandil, reduced sigma creatine kinase compared with placebo by 14.7%, increased left ventricular ejection fraction (EF) gauged by left ventriculography at 6–12 months by 5.1% and reduced reperfusion injury by 26%. ANP decreased and nicorandil marginally decreased the rate of either re-hospitalization due to heart failure or cardiovascular death. ANP may be used for the patients with AMI to reduce both infarct size and reperfusion injury, and improve outcomes, while nicorandil provided additional cardiovascular protection.
Taken together, the current study opens new era of adjunctive therapy in patients with acute myocardial infarction to reduce ischemic heart failure via these strategies. We will further show and discuss the several advanced hypotheses derived from the fruitful results in basic research of pathophysiology of acute ischemic injury.
ECMO IN SEPTIC SHOCK: WHEN TO USE IT AND NOT TO USE IT? Tao-Min Huang. Department of Internal Medicine, National Taiwan University Hospital and College of Medicine, Taichung, Taiwan
Extracorporeal membrane oxygenation (ECMO) provides temporary supports for heart and lung function, and is potential lifesaving in selected patient group, including severe acute respiratory failure or cardiac failure. Sepsis and septic shock is the leading cause of morbidity and mortality in modern critical care. Patients with septic shock refractory to medical treatment have multiple organ failure, including compromised heart and lung function. However, the role of ECMO in patients with severe sepsis or septic shock is under great debates. Sepsis involved altered immune system, coagulopathy and resultant cardiopulmonary compromise. The coagulopathy and mechanical complications make ECMO not a good choice for patients with septic shock. However, the utilization of ECMO in septic shock patients is increasing. The discussion will focus on the current evidence of ECMO use in severe sepsis with multiple organ failure.
THE PATHOPHYSIOLOGY OF AKI AND THE RATIONAL FOR BLOOD PURIFICATION Hernando Gomez1, Yujie Ma1, Kui Jin1, Brian S. Zuckerbraun2, Michael R. Pinsky3, John A. Kellum1. 1Critical Care Medicine, Center for Critical Care Nephrology and The CRISMA Center, University of Pittsburgh, Pittsburgh, PA, USA, 2Department of Surgery, University of Pittsburgh, Pittsburgh, PA, USA, 3Department of Critical Care Medicine, The CRISMA Center, University of Pittsburgh, Pittsburgh, PA, USA
Acute kidney injury (AKI) is a common complication in critically ill patients, and is associated with increased morbidity and mortality. Sepsis is the most common cause of AKI in the critically ill. Considerable evidence has shown that AKI can occur in the absence of overt clinical signs of shock, and in the setting of increased renal blood flow. This has challenged the traditional paradigm that renal dysfunction was solely on the basis of hypoperfusion and ischemia. Animal and human data have further shown that sepsis-induced AKI is characterized not by acute tubular necrosis, but by a paucity of apoptosis and necrosis in the context of a very bland histology, by inflammation, by microvascular dysfunction, and cellular bioenergetics adaptive responses. These novel findings suggest that other potential mechanisms centered in these three domains may help explain the pathophysiology of sepsis-induced AKI. Although it remains unknown what the contribution of any of these three domains to sepsis induced AKI is, the metabolic re-programming of the tubular epithelial cell that frames the adaptive response has received much attention, as it not only relates to protection of the organ and the host during the early stages of inflammation, but also conditions the repair response during convalescence. We have shown that activation of a master regulator of energy balance, AMP activated protein kinase, can protect from AKI in a rodent model of cecal ligation and puncture (CLP) induced severe sepsis. Our preliminary data further suggests that AMPK activation in this setting can limit energy depletion (in the form of adenosine triphosphate or ATP) and may improve 7-day survival. This supports the findings from other groups who have demonstrated that activation of SirT1, a downstream AMPK target, early in the course of CLP-induced sepsis can improve survival. Thus, emerging evidence supports a more direct role of metabolic adaptations as defense cellular strategies that may impact relevant clinical outcomes like the development of organ dysfunction and survival. Furthermore, many of the novel pathophysiological mechanisms that will be discussed may yield viable biomarker candidates and more importantly, targets for extracorporeal therapy in the future.
ENDOTOXIN ADSORPTION THERAPY FOR SEPTIC SHOCK Kent Doi. Department of Emergency and Critical Care Medicine, The University of Tokyo, Tokyo, Japan
Although endotoxin has been regarded as a key factor in the sepsis cascade and a potential therapeutic target, the survival benefit of endotoxin adsorption therapy for septic shock has not been sufficiently established. Several multi-center randomized clinical trials that enrolled post-abdominal surgery sepsis did not show any significant improvement by endotoxin adsorption therapy using polymyxin B hemoperfusion (Shock 2005, JAMA 2009, ICM 2015). A previous retrospective analysis of a Japanese nationwide inpatient database also suggested that endotoxin adsorption provides no survival benefit for septic shock patients with lower gastrointestinal perforation (CCM 2014). However, another analysis that targeted septic shock patients complicated with severe acute kidney injury requiring continuous renal replacement therapy revealed that polymyxin B hemoperfusion provided survival benefit for this severely ill population (Blood Purif 2016). This observation encourages the appropriate identification of responders benefitting from the endotoxin adsorption therapy.
IMMUNOMODULATING BLOOD PURIFICATION SYSTEM IN SEPSIS AND SEPTIC SHOCK Yoshitaka Hara1, Tomoyuki Nakamura1, Yu Kato1, Toshikazu Sakai1, Yasuyo Shimomura1, Hidefumi Komura1, Chizuru Yamashita1, Junpei Shibata1, Kazuhiro Moriyama2, Osamu Nishida1. 1Department of Anesthesiology and Critical Care Medicine, Fujita Health University School of Medicine, Aichi, Japan, 2Laboratory for Immune Response and Regulatory Medicine, Fujita Health University School of Medicine, Aichi, Japan
The novel blood purification system, which we have devised and named immunomodulating blood purification system (IMBPS), aims to regulate not only the excessive mediators, but rather the entire immune reaction. This system relocates the excessive systemic inflammation occurs in vivo to ex vivo IMBPS, preventing the development of organ dysfunction. It is a “reversal strategy from a different angle”. We focused our attention on Adacolumn (JIMRO) for selective leukocytapheresis for ulcerative colitis and AN69ST hemofilter (Baxter), a mediator adsorbing-hemofilter. In IMBPS, the first column in the circuit (Adacolumn) selectively captures activated leukocytes. The mediators produced in the capturing process by the activated leukocytes are adsorbed by a downstream 2nd column (AN69ST hemofilter). The results from ex vivo basic research showed that within a single pass through IMBPS, cytokine activity was increased after passing Adacolumn and decreased after passing AN69ST (Ther Apher Dial 2015; 19: 308–315). With IMBPS, granulocytes and monocytes decreased, whereas lymphocytes did not. Phagocytosis and the adhesive capacity of granulocytes were also reduced. After we confirmed both the effectiveness and safety of IMBPS in living porcines, we moved on to clinical application of IMBPS. SHEDD-fA system was applied as the 2nd column, instead of unapproved AN69ST hemofilter.
Case 1: A 25-year-old male with ulcerative colitis (UC) and infectious gastroenteritis admitted to intensive care unit. We demonstrated IMBPS for sepsis during 4 days, and for the acute exacerbation UC during 3 days.
Case 2: A 66-year-old male with chronic Epstein Barr virus infection has invasive pulmonary aspergillosis, progress acute respiratory distress syndrome, and need to Veno-venous extracorporeal membrane oxygenation. His leukocyte was excessively high and IMBPS protected the artificial lung from activated leukocyte attacks, resulting long lifetime of the oxygenator. Circulating histone 3, a key effector in sepsis, was absorbed to Adacolumn, indicating the adsorption of activated leukocytes.
In the two cases reported here IMBPS was safe and effective. Phagocytosis and the adhesive capacity of granulocytes were also reduced. We here report the potential of IMBPS, with its results from ex vivo research and in vivo data from living porcine and clinical use.
CONTINUOUS HEMODIAFILTRATION WITH A CYTOKINE-ADSORBING HEMOFILTER IN PATIENTS WITH SEPTIC SHOCK Hidetoshi Shiga. Emergency and Intensive Care Center, Teikyo University Chiba Medical Center, Chiba, Japan
Introduction: It is widely accepted that hypercytokinemia caused by the inflammatory response to infection and abnormal tissue oxygen metabolism play pivotal roles in the pathophysiology of sepsis. While a number of blood purification therapies for the treatment of sepsis are available, our group has reported that continuous hemodiafiltration (CHDF) using a polymethylmethacrylate (PMMA) hemofilter, which has cytokine adsorbing capacity can continuously and effectively remove cytokines from the blood.
The AN69ST membrane has excellent cytokine adsorption capacities. The aim of the present study was to evaluate the clinical efficacy of cytokine removal by CHDF using an AN69ST hemofilter (AN69ST-CHDF) in patients with septic shock, even for the septic patients without renal dysfunction.
Patients and Methods: All patients admitted to the ICU were screened for eligibility. The inclusion criteria were the presence of sepsis, according to the definitions of the American College of Chest Physicians/Society of Critical Care Medicine Consensus Conference Committee, and shock diagnosed by an initial blood lactate level of 36 mg/dL or higher. The exclusion criteria were pregnancy, immunosuppressive therapy, and uncontrollable bleeding. No patients were excluded due to severity.
A prospective, multicenter, single-arm study was conducted.
CHDF was initiated immediately after a patient fulfilled the entry criteria, regardless of their kidney function. CHDF was performed using the Prismaflex with the ST filter set. The operating conditions were as follows: QB, 100–150 mL/min; dialysate flow rate, 500 mL/hr; QF, 600 mL/hr. Sublood-BS was used as both the dialysate and replacement fluid. Nafamostat mesilate was administered as an anticoagulant at a rate of 30 mg/hr. AN69ST-CHDF was intended to perform at least for 72 hr, and the hemofilter was scheduled to be replaced every 24 hr, even when there were no clots in the hemofilter or extracorporeal circuit.
Results: Thirty-four patients were enrolled. On ICU admission, the mean blood IL-6 level was 44,800 77,700 pg/mL, and the mean blood lactate level was 69.0 49.4 mg/dL. Both the mean blood IL-6 and lactate levels had significantly decreased to normal ranges after 72 hr of AN69ST-CHDF. Though the mean APACHE II score was 32.7 9.8, 28-day survival was 73.5%
Conclusion: AN69ST-CHDF eliminated the causative cytokines of septic shock from the bloodstream, mainly by adsorption.
Following the initiation of AN69ST-CHDF, both hypercytokinemia and dysoxia were improved. As a result, the AN69ST-CHDF might lead to good outcomes for patients with septic shock. Although further investigations are apparently needed, the present findings suggest that it might be an effective strategy for the treatment of septic shock.
MEMBRANE ADSORPTION AS A KEY MODALITY IN BLOOD PURIFICATION FOR SEPSIS IN THE FUTURE Patrick M. Honore, Rita Jacobs, Herbert D. Spapen. ICU Department, Universitair Ziekenhuis Brussel, Vrije Universiteit Brussel, Brussels, Belgium
After several negative studies upon High Volume Hemofiltration and High Cut-Off Hemofiltration targeted at Sepsis and septic shock, highly adsorptive membranes remains a major hope for the future regarding blood purification in Sepsis. Indeed, those membranes have the capability of adsorbing very effectively small size mediators such as high mobility group box protein 1 (HMGB-1). Indeed, those mediators although small in size could not be removed by convection alone. Probably many other small mediators like HMGB-1 are also playing a pivotal role in sepsis without being removed by convection for decades. This represents the most promising way for targeting septic shock including acute kidney injury in the future. Adsorption with or without the help of convection can be seen nowadays as the major player for fixing septic shock through a blood purification targeted approach.
Keywords: Acute kidney injury, Septic Shock, Sepsis, Blood purification, membrane adsorption, HMGB-1.
ALCOHOL-NEUROIMMUNE INTERACTIONS IN TRAUMATIC BRAIN INJURY Patricia E. Molina. Department of Physiology and Alcohol & Drug Abuse Center of Excellence, Louisiana State University Health Sciences Center, New Orleans, LA, USA
Traumatic brain injury (TBI) is a debilitating condition in military and civilian populations. The post-TBI period is characterized by an early and sharp rise in neuroinflammation followed by lasting neurological and behavioral deficits that include sleep disturbances, neuroendocrine dysfunction, increased incidence of anxiety and depression, and impulse control deficits. Using a rat model of mild to moderate “single-hit” TBI produced by fluid percussion, we have shown an early neuroinflammatory phase characterized by microgliosis, astrocytosis, and increased gene expression of inflammatory cytokines, that is resolved 10–14 days post-TBI. Post-TBI chronic intermittent alcohol exposure (CIE) via vapor inhalation profoundly impairs recovery of neurobehavioral function, exacerbates gliosis, and prevents resolution of neuroinflammation. The potential synergistic detrimental effects on alcohol and post-TBI neuroinflammation are further supported by our data showing that TBI produces transient escalation of operant alcohol self-administration, sufficient to exacerbate post-TBI neuroinflammation. Encouraging from a therapeutic perspective is the finding that a single post-TBI injection of JZL184 a monoacylglycerol lipase (MAGL) inhibitor that prevents endocannabinoid degradation, attenuates neuroinflammation, improves neurobehavioral function, and prevents increased PR responding for alcohol.
TRAUMATIC BRAIN INJURY INDUCES APOPTOSIS AND EXHAUSTION OF T CELLS WITH INCREASED MORTALITY AFTER SEPSIS Shigeaki Inoue, Usuke Uehara, Maiko Tada, Yusuke Suzuki, Nobuo Watanabe, Sadaki Inokuchi. Tokai University School of Medicine, Kanagawa, Japan
Background: Pneumonia and secondary infections after traumatic brain injury (TBI) are serious problems in ICU. Although brain infraction is known to be a cause of immunosuppression with decreased number and function of lymphocytes, the mechanism of immunosuppression after TBI is still unclear. The purpose of the study is to elucidate the immunological changes occurring in mice after TBI.
Methods: Six to eight weeks-old C57/B6 female mice were subjected to Sham or TBI using weigh-drop method (weight: 15.7 g, height: 47 cm, E = 0.07J).Mice were killed at 48 h after TBI or sham surgery, and the spleen, and lymph nodes were harvested. Apoptosis was measured by flow cytometry by using caspase 3. To determine the impact of TBI on the susceptibility to bacterial infection, survival study of two-hit model, that is cecal ligatioin puncture model (CLP) at 48 h after TBI/Sham, was conducted.
Results: Compared to sham mice, TBI mice showed 1) decreased population of splenocytes, 2) increased Caspase 3-positive CD8+ T and B cells from spleen and lymph nodes, 3) increased proportion of programmed death-1, cytotoxic T-lymphocyte-associated protein-4, and Foxp-3 in CD4+ T cells in lymph nodes at 48 hrs after TBI. The 7-days survival in mice with TBI following CLP was significantly decreased compared to that of the Sham group following CLP (17% vs 83%).
Conclusion: TBI induces apoptosis and exhaustion of T cells in mice. These immunological changes after TBI may be related to immunosuppression, following to increased secondary infection and sepsis in the delayed phase in ICU.
THE ROLE OF INNATE IMMUNITY IN THE TRAUMATIC BRAIN INJURY-INDUCED IMMUNE SUPPRESSION SYNDROME Steven J. Schwulst. Department of Surgery, Northwestern University, Chicago, IL, USA
Traumatic brain injury (TBI) is a growing and under-recognized public health concern. The Centers for Disease Control and Prevention estimates nearly 2.5 million people sustain a traumatic brain injury each year in the United States resulting in over 300,000 hospitalizations and 50,000 deaths. The impact of TBI is highlighted not just by its high mortality, but also by the significant long-term complications suffered by survivors. It is now recognized that even mild TBI, or concussions, may lead to significant long term morbidity and the insideous onset of neurodegenerative disease. The immune response to TBI represents a complex interplay between peripheral immunity and the resident immune system of the injured brain. The result is a microenvironment of simultaneous inflammation and repair. These competing backgrounds can produce a microenvironment geared towards either repair and regeneration or a local milieu that propells and propagates the index injury. To date, few studies have addressed the intersection of peripheral immunity with the resident innate immune system of the injured brain. Accordingly, there is a critical unmet need to determine whether advancements in our understanding of these neuro-immune interactions will lead to new therapies to combat this highly lethal injury process.
ESTROGENS FOR THE TREATMENT OF TRAUMATIC BRAIN INJURY Irshad H. Chaudry1, Harrison Kim2, Kurt Zinn2, Thomas van Groen3, Betul Cam1, Ahmar Ayub1, William Hubbard1. 1Department of Surgery, University of Alabama at Birmingham, Birmingham, AL, USA, 2Department of Radiology, University of Alabama at Birmingham, Birmingham, AL, USA, 3Department of Cell, Developmental & Integrative Biology, University of Alabama at Birmingham, Birmingham, AL, USA
Traumatic brain injury (TBI) is a worldwide problem leading to mortality and long-term disability. This injury often causes cerebral edema, leading to increased intracranial pressure and decreased cerebral perfusion pressure. These result in cerebral hypoxia and eventually cerebral ischemia and neuronal degeneration. Although the serious consequences of TBI were not fully appreciated for many years, more recent awareness of this problem has dramatically changed public perceptions for the consequences of TBI. It is also evident that estrogen can ameliorate neurological injury. Thus, if estrogen is to be used for the treatment of TBI, it must be administered as soon as possible after injury. Our results have shown that that the best means to deliver estrogen is in a water-soluble form, given intravenously or intraosseously. Our results have also indicated that lateral fluid percussion brain-injured rats have lessening of their injury when treated with soluble estradiol (E2) or ethinyl estradiol-3-sulfate (EE-3-SO4) was administered one hour after the induction of TBI. These beneficial effects include reduction of cerebral edema and anisotropy as revealed by MRI. Furthermore, invasive measurements demonstrate reduced intracranial pressure, and improved cerebral perfusion pressure and partial brain oxygen pressure. Additionally, longer term benefits in cognition and memory have been shown with improved Morris Water Maze performance.
Our previous study has shown the efficacy of E2 in its sulfate-conjugated form (17 beta-estradiol sulfate, E2-SO4) to treat experimental TBI. E2-SO4 is a naturally-occurring derivative of E2, created by the addition of a sulfate moiety via a steroid sulfotransferase enzyme. The physiological sulfate conjugation process allows excretion of this soluble form of E2 via the kidneys, which regulates the hormone levels. In our studies, the solubility of E2-SO4 enables intravenous delivery of supraphysiological quantities of this hormone, which would not be possible with the highly hydrophobic native E2.
We also used passive infrared (PIR), motion detectors to determine physical activity for three days. Results show that the animal activity was reduced after TBI but was markedly increased in EE-3-SO4 treated animals. Rats were also monitored for weight gain/loss and the weight loss for three days post TBI was significantly less in the EE-3-treated animals than in the vehicle treated animals. Thus, EE-3-SO4 promotes recovery by increasing brain metabolism, resolving edema and improving physiological parameters if administrated one-hour post TBI. These data support the clinical use of EE-3-SO4 for early TBI treatment.
MONITORING AND TREATMENT STRATEGIES FOR PHYSIOPATHOLOGICAL RESPONSES IN SEVERE TBI: A PRIMARY TO CUATERNARY INJURY APPROACH Andres Mariano Rubiano Escobar. Department of Neurosurgery, Neuroscience Research Institute, El Bosque University, Bogota, Colombia
Abstract not available yet.
DEVELOPMENT OF A STRETCH-INDUCED NEUROTRAUMA MODEL FOR MEDIUM-THROUGHPUT SCREENING IN VITRO: IDENTIFICATION OF RIFAMPICIN AS A NEUROPROTECTANT Csaba Szabo1, Isabel Lopez-Garcia1, Domokos Gero1, Bartosz Szczesny1, Kangling Zhang2, Petra Szoleczky1, Gabor Olah1, Katalin Modis1, Douglas DeWitt1, Ping Wu3, Lawrence C. Sowers2, Donald S. Prough1. 1Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA, 2Department of Pharmacology, University of Texas Medical Branch, Galveston, TX, USA, 3Department of Neuroscience and Cell Biology, University of Texas Medical Branch, Galveston, TX, USA
After the initial mechanical insult of traumatic brain injury (TBI), secondary injury develops; it is responsible for a significant part for the TBI-induced neural damage. We developed an in vitro, stretch-based model of neural injury that is applicable to cell-based screening to screen for compounds that may decrease the cellular damage in neurotrauma. We screened three neural cell lines (B35, RN33B, SH-SY5Y) subjected to two differentiation methods. We measured the gene expression changes and tested the sensitivity to stretch-induced secondary damage by measuring the viability changes and selected ATRA-differentiated B35 cells for the screen. Stretch injury, when combined with a low, subthreshold concentration of hydrogen peroxide, induces delayed-onset cell death. The model induced marked alterations in the proteomic signature of the cells; from 4093 proteins, 159 being upregulated and 131 being downregulated. Using this model, we screened a composite library of 3,500 drugs, drug-like compounds and well-annotated pharmacological compounds, with LDH and MTT 24 h end-points. Drugs were applied as a 1-hour post-treatment. Six compounds inhibited both LDH and MTT responses, one of them being the well-known antibiotic rifampicin, which protected in the concentration range of 3–30 μM. Rifampicin maintained its protective effect in a 3-hour post-treatment scenario. It reduced cell necrosis and apoptosis and improved cellular bioenergetics. The mode of action in the current model does not appear to involve protein phosphatase 2A (a previously identified secondary target of rifampicin). Rifampicin was found to also inhibit neuroinjury elicited by high-dose hydrogen peroxide (without stretch). Its protective effect was also confirmed in a secondary model (stretch injury in human stem cell derived neurons); rifampicin attenuated LDH release, protected against the loss of neurite length and maintained neuron-specific class III beta-tubulin immunoreactivity. Taken together, the current model is suitable for medium-throughput screening to identify compounds with neuroprotective potential. Because the neuroprotective concentrations of rifampicin are achievable in vivo, and it has adequate safety profile and CNS uptake, this drug may be a candidate for repurposing for the therapy of TBI.
MILD TRAUMATIC BRAIN INJURY BUT NOT TAIL TRAUMA AUGMENTS PULMONARY CLEARANCE OF BACTERIA VIA RELEASE OF SUBSTANCE P Daniel Remick, Terry Hsieh, Max Vaickus, Thor Stein, Bethany Lussier, Jiyoun Kim, David Stepien, Elizabeth Duffy, Evan Chiswick. Department of Pathology and Laboratory Medicine, Boston University School of Medicine, Boston, MA, USA
Neural input on the immune system can alter its ability to effectively clear pathogens. Patients suffering mild traumatic brain injury (TBI) have shown reduced rates of pneumonia and a murine model replicates these findings, with better overall survival of TBI mice compared to sham injured mice. To further investigate the mechanism of improved host response in TBI mice, this study developed and characterized a mild tail trauma model of similar severity to mild TBI. Both mild tail trauma and TBI induced similar systemic changes that normalized within 48 h. Examination of tissues showed that injuries are limited to the target tissue, i.e. tail in tail trauma, brain in mTBI. Pneumonia challenge showed that mild TBI mice displayed improved immune responses, characterized by increased survival, pulmonary neutrophil recruitment, and bacterial clearance compared to tail trauma. Administration of a neurokinin-1 receptor (NK-1R) antagonist to block substance P signaling eliminated the improved survival of mTBI mice. NK-1R antagonism did not alter pneumonia mortality in tail trauma mice. These data demonstrate that immune benefits of trauma are specific to mTBI and that tail trauma is an appropriate control for future studies aimed at elucidating the mechanisms of improved innate immune responses in mTBI mice.
GUT EPITHELIAL INTEGRITY IN SEPSIS Craig Coopersmith. Emory University, Atlanta, GA, USA
The gut has long been hypothesized to be the motor of multiple organ dysfunction syndrome. Whereas initial theories proposed that worsening gut barrier function resulted in bacterial translocation into the systemic circulation, the reality is significantly more complex than this. Multiple components of the gut including the epithelium, the immune system and the microbiome are altered by critical illness and can, in turn, propagate a pathologic response. This talk will focus on our current understanding of how dysregulated gut epithelial apoptosis, proliferation, and permeability contribute to the development of MODS as well as potentially ways in which gut integrity can be manipulated for therapeutic gain.
NUTRITION MANAGEMENT OF CRITICALLY ILL PATIENTS: EVIDENCES AND PRACTICE Joji Kotani, Atsunori Hashimoto, Kunihiro Shirai, Takahiro Ueda, Michiko Aoyama-Ishikawa, Taihei Yamada, Naomi Mambo, Yasutomi Hirai, Noritomo Fujisaki, Isamu Yamada, Hiroyuki Nakao. Department of Emergency, Disaster and Critical Care Medicine, Hyogo College of Medicine, Hyogo, Japan
It has been well known that eating/feeding is necessary in recovery from severe illness. Since Dr. Dudrick invented total parenteral nutrition, its usefulness and superiority of enteral nutrition, strangely enough, have been emphasized. While stress response of immune system appears to be involved in risk of infection and organ failure, effectiveness and mechanisms of immuno-modulating diets that strengthen or regulate immune system have been also investigated vigorously. In other words, we need to recognize that feeding is no longer only for supply of nutrients but also the therapeutic tools. Nevertheless, we often experience difficulties with practice at the clinical site; hence, we should develop an individual strategy corresponding to each case.
I will explain the evidences of early enteral nutrition, the one of the most important steps in nutritional therapy in critically ill patients, and the methods to initiate enteral nutrition including the technique to place the feeding tube beyond pyloric ring, appropriate amount of enteral nutrition, clinical situation requiring immuno-modulating nutrients, and so on, with the case study.
DYNAMIC CHANGES OF GUT MICROBIOTA IN CRITICALLY ILL PATIENTS Kentaro Shimizu. Department of Traumatology and Acute Critical Medicine, Osaka University Graduate School of Medicine, Osaka, Japan
The gut is an important target organ following severe insult such as sepsis, trauma, burn, shock, bleeding and infection. Severe insult to the gut is considered to have an important role in promoting infectious complications and multiple organ dysfunction syndrome. The gut is the “motor” of multiple organ failure, and gut dysfunction is recognized as a causative factor in the progression of diseases.
Gut microbiota have been related with not only intestinal metabolism but also immune system. We have reported that systemic inflammatory response syndrome (SIRS) patients had 100–10,000 times fewer total anaerobes, including Bifidobacterium and Lactobacillus, and 100 times more Staphylococcus bacteria compared with healthy volunteers. Total organic acids, acetic acid and butyric acid were significantly decreased in the SIRS patients compared with healthy volunteers. Fecal pH was markedly increased in patients with severe SIRS in comparison with healthy volunteers (p < 0.05). These gut microbiota changes occurred in a few days and sustained long time decrease. Especially, the decrease of the number of total obligate anaerobes, which are dominant in human, was associated with mortality and septic complications. These results suggested that the maintenance of gut microbiota had the potential to prevent septic complications in critically ill patients.
Probiotics/synbiotics therapy has been noted as an intestinal therapy which maintained gut microbiota. Postoperative infectious complications decreased in hepatectomy for biliary cancer, liver transplantation, liver cancer, and etc. For critically ill patients such as trauma, sepsis and ventilated patients in the ICU, it has been reported that prophylactic probiotics reduced ventilator associated pneumonia and other septic complications. The mechanism has not been clarified thoroughly, but it may be explained with IgA or defensins secretion via intestinal receptors and probiotics metabolites such as short-chain fatty acids.
Recently, metagenomic analysis of gut microbiota revealed more precise proportions than culture methods. We indicated serial dynamic changes in the percentages of Bacteroidetes and Firmicutes at the phylum level of ICU patients by high-throughput DNA sequencing. A ratio of Bacteroidetes to Firmicutes (B/F ratio), which means extreme dysbiosis in the gut microbiota, was associated with prognosis.
In conclusion, gut microbiota changes dynamically in the acute phase of critically ill patients. These changes could deteriorate host response against injuries. Intervention for these changes such as probiotics/synbiotics could modulate host immune response and prevent septic complications. Further basic and clinical research would delineate the mechanisms and promote intestinal treatments.
APPLICATION OF THERAPEUTIC MEDICAL GAS FOR BOWEL DYSFUNCTION Atsunori Nakao. Department of Emergency and Critical Care Medicine, Okayama University Graduate School of Medicine, Dentistry and Pharmaceutical Sciences, Okayama, Japan
Gastrointestinal motility disorders are common complications in the intensive care setting. Adverse effects of bowel dysfunction include increased abdominal pain, delay in resuming oral intake, poor wound healing, delayed mobilization, increased risk of pulmonary complications and prolonged hospitalization. Several risk factors for the development of gastrointestinal motility problems have been identified and include sepsis, being on mechanical ventilation, and the use of vasopressors, opioids or anticholinergic medications.
Gaseous molecules which have known medical applications include a number of low molecular weight substances such as carbon monoxide and hydrogen. These gases have gained widespread attention as novel therapeutics with anti-inflammatory and antioxidant properties. Medical gas-related pharmacological research is a rapidly emerging field, which is likely to yield a number of therapeutic possibilities for patients in the clinical setting.
The use of medical gases to treat bowel dysfunction is an exciting and evolving therapeutic possibility. Herein, we will present the recent advances and current knowledge pertaining to therapeutic medical gases including carbon monoxide (CO) and hydrogen, which have been reported as a biologically active product. Additionally, we will discuss their clinical applications and therapeutic properties. These gases may be toxic, hazardous or poisonous at a higher concentration, however they are safe and potentially therapeutic at lower concentration. The therapeutic benefit of these gases in human patients remains unclear, and further controlled clinical studies are needed.
TOLL-LIKE RECEPTOR STIMULATION REVERSES ANTIBIOTIC-INDUCED GUT DEFENSE IMPAIRMENT Lee-Wei Chen1,3, Ying-Ying Wu2, Pei-Hsuan Chen1, Ching-Mei Hsu2. 1Department of Surgery, Kaohsiung Veterans General Hospital, Kaohsiung, Taiwan, 2Department of Biological Sciences, National Sun Yat-Sen University, Kaohsiung, Taiwan, 3Institute of Emergency and Critical Care Medicine, National Yang-Ming University, Taipei, Taiwan
Prior antibiotic exposure is associated with increased mortality in gram-negative bacteria-induced sepsis. However, how antibiotic-mediated changes of commensal bacteria promote the spread of enteric pathogenic bacteria in patients remains unclear. In this study, the effects of systemic antibiotic treatment with or without toll-like receptor (TLR) stimulation on bacteria-killing activity, antibacterial protein expression in the intestinal mucosa, and bacterial translocation were examined in mice receiving antibiotics with or without oral supplementation of dead Escherichia coli or Staphylococcus aureus. We developed a systemic ampicillin, vancomycin, and metronidazole treatment protocol to simulate the clinical use of antibiotics. Antibiotic treatment decreased the total number of bacteria including the aerobic Enterobacteriaceae and Enterococcus as well as the anaerobic Lactococcus/Bifidobacterium in the intestinal mucosa and lumen. Antibiotic treatment significantly decreased the bacteria-killing activity of the intestinal mucosa and the expression of non-defensin family proteins, such as RegIIIβ, CRP-ductin, and RELMβ, but not the defensin family proteins, and increased Klebsiella pneumoniae translocation. TLR stimulation after antibiotic treatment increased NF-κB DNA binding activity, non-defensin protein expression, and bacteria-killing activity in the intestinal mucosa and decreased K. pneumoniae translocation. Moreover, germ-free mice showed a significant decrease of non-defensin proteins as well as of intestinal defense against pathogen translocation. Since TLR stimulation induced NF-κB DNA binding activity, TLR4 expression, and mucosa-killing activity in germ-free mice, we conclude that commensal microflora is critical in maintaining intestinal non-defensin protein expression and the intestinal barrier. In turn, we suggest that TLR stimulation induces non-defensin protein expression and reverses antibiotic-induced gut defense impairment.
SURGERY INDUCED SEPTIC COMPLICATIONS IN MYANMAR: A SINGLE CENTER EXPERIENCE Kyaw Htet, Than Than Yee, Than Naing Tun, Hla Myo. Department of GI and HBP Surgery, No. 2 Defence Services General Hospital, Nay Pyi Taw, Myanmar
Sepsis and its complications are the major challenge for the best surgical outcomes. Even in the era of advances molecular technologies and potent antibiotics, there are still high morbidity and mortality in sepsis related surgical complications. The aim of this presentation is to discuss the current situation of major surgery-induced septic complication, single center experience.
Total number of 734 patients underwent major abdominal surgery at No. 2 Defenses Services General Hospital (1000 Bedded), Nay Pyi Taw, Myanmar from January 2011 to December 2015. The most common septic complications are surgical site infection (7.6%) followed by postoperative chest infection and ARDS, anastomosis leak and intra-abdominal abscess. Postoperative chest complications is less noted in laparoscopic operations compare to its counterpart in open procedures. The commonest microorganisms identified were E coli, Klebsiella and Pseudomonas (78.2%). Mortality rate in highest in ARDS patients (52%) and colorectal anastomosis with generalized peritonitis patients (25.1%).
Preoperative optimization, strict aseptic protocol and good surgical techniques play the most important role to reduce sepsis related surgical complications but multidisciplinary team approach is also vital for management of that sort of complications.
SELECTIVE MANAGEMENT OF UNEXPECTED COMPLICATIONS BY DELAYED SURGICAL MANAGEMENT OF OESOPHAGEAL PERFORATION AND THERAPEUTIC ENDOSCOPY Sirikan Limpakan (Yamada). Division Chief of Gastrointestinal Surgery and Endoscopy, Department of Surgery, Chiang Mai University, Chiang Mai, Thailand
The management for delayed oesophageal perforation still be difficult unexpected task for surgeon due to delayed detection and transfer for surgical management, including failure to control sepsis occurred in various stage conditions lead of high morbidity and mortality. The author discuss about 2 cases on delayed oesophageal perforation with long tearing by Senstaken-Blakemore tube that is common used in variceal bleeding patients for temporary stop of exsanguinous bleeding. Another case is foreign body caused thoracic oesophageal perforation with sepsis and renal failure. On the first case, patients was delayed transfer for 10 days with right thoracic tube and low grade fever. We decided to immediate right thoracotomy. Perforation length was 10 cm. We decided to simple repair over t-tube drainage, EG junction banding over nasogastric tube, tube cervical esophagostomy with jejunostomy. Patient could return to eat in 6 weeks after T-tube, and cervical esophagostomy tube, and chest tube withdrawing. The second case was delayed transfer about 2 weeks with severe sepsis and renal failure. After sepsis was controlled, then exploratory laparotomy for jejunostomy and EG junction banding. We placed self-expandable stent (SEM) successfully, but stent migrated out to the pleural cavity, and needed to be removed. The patient developed sepsis again, and he underwent right thoracotomy to decorticate, toilet, and esophageal resection with both blinded end closure left in the thoracic cavity. He recovered in 6 weeks after second operation and awaiting to go on oesophageal substitute operation. The latter was delayed transfer about 2 weeks with sepsis and right thoracic tube drainage. The large fish bone caused perforation at mid to lower thoracic oesophagus, and it was demonstrated at perforation site. We used argon plasma cut the fishbone into piece via endoscopy and it became small piece down into stomach. However, the thoracic collection progressed 3 days later, and relapsed of sepsis. Patient underwent right thoracotomy to adequate drainage and decortication. He was explored laparotomy to band the EG junction over NG tube, and feeding jejunostomy without cervical esophagostomy. Patients recovered with having normal swallowing within 4 weeks, and chest tube was removed without problem.
Conclusion: We may avoid fatal complication of delayed thoracic oesophageal perforation with sepsis by selective management. We agree to do oesophageal repair if possible, but T-tube drainage and EG junction banding with or without tube cervical esophagostomy are recommended. Selective endoscopy procedure, drainage, or salvage control with reconstruction in combination with medication and nutrition management are necessary.
CHEMOKINE-CHEMOKINE RECEPTOR NETWORK AND ESOPHAGECTOMY FOR ESOPHAGEAL SQUAMOUS CELL CARCINOMA Hiroya Takeuchi, Masazumi Inoue, Tomohiko Nishi, Masaharu Ogura, Sachiko Matsuda, Kazumasa Fukuda, Rieko Nakamura, Norihito Wada, Hirofumi Kawakuno, Yuko Kitagawa. Department of Surgery, Keio University School of Medicine, Tokyo, Japan
Background: Esophagectomy is one of the most invasive surgical treatments, and the inflammatory cytokines in blood such as interleukin-8 (IL-8) are increased for several hours after esophagectomy or in patients experiencing postoperative complications including sepsis. IL-8, and its receptor, CXCR2, are expressed in various cancer cells. The IL-8/CXCR2 network is believed to be involved in angiogenesis, tumor cell proliferation and invasion. However, the impact of postoperative complications following esophagectomy on long-term survival is controversial. In this study, we demonstrate the significance of CXCR2 expression and validate the effects of CXCR2 expression and postoperative complications on long-term prognosis of esophageal squamous cell carcinoma (ESCC). Furthermore we investigated the role of the IL-8/CXCR2 network in the tumor microenvironment of ESCC.
Methods: Eighty-two specimens were sectioned from archived, paraffin-embedded tumor tissues obtained from patients with ESCC who underwent esophagectomy and extended lymphadenectomy for complete resection of cancer. Immunohistochemistry was performed using an antibody to CXCR2, and the correlation of stainability with clinicopathological factors and long-term survival was examined. The concentration of IL-8 and CXCR2 were investigated using the human ESCC cell lines. IL-8/CXCR2 signaling was stimulated by IL-8 addition, and suppressed by SB225002, a selective antagonist of CXCR2, and IL-8 specific siRNA in vitro and in vivo.
Result: CXCR2 was expressed in 33 of 82 specimens. In the CXCR2-positive group, the recurrence-free survival and overall survival rates of patients who developed postoperative complications were both significantly lower than those for patients who did not develop any complications. In contrast, in the CXCR2-negative group, there was no significant difference in long-term prognosis. Stimulation or inhibition of the IL-8/CXCR2 network resulted in significant enhancement or suppression of cell proliferation in ESCC cell lines (P < 0.05) in a dose-dependent manner. IL-8 over-expressing TE4 cells showed significantly higher proliferative activity than the parental line (P < 0.05), which was markedly suppressed by inhibition of the IL-8/CXCR2 network using SB225002 (P < 0.05). In vivo, tumor sizes were smaller in the SB225002-treated group than in the untreated group.
Conclusion: Our results demonstrated that stimulation of the IL-8/CXCR2 network clearly enhanced ESCC cell proliferation, while its inhibition obviously suppressed ESCC cell proliferation. The patients with CXCR2-positive esophageal cancer who develop postoperative complications have a poor prognosis and should be carefully followed.
IMMUNOLOGICAL EVALUATION OF THE POSTOPERATIVE COMPLICATIONS AFTER ESOPHAGECTOMY Hironori Tsujimoto1, Shuichi Hiraki1, Satoshi Ono2, Hideki Ueno1. 1Department of Surgery, National Defense Medical College, Saitama, Japan, 2Tokyo Medical University Hachioji Medical Center, Tokyo, Japan
The surgical trauma that an esophagectomy imposes is perhaps the greatest among all general surgical operations. Such operative trauma activates immunocompetent cells, which results in the release of inflammatory cytokines, resulting in the development of systemic inflammatory response syndrome (SIRS). In contrast, SIRS followed by ‘compensatory anti-inflammatory response syndrome’ (CARS) is a conceptual framework to explain the immunological trajectory that patients underwent esophagectomy often traverse, but the mechanisms for persistent immune dysfunction remain unexplained. Here, we present immune-inflammatory status after esophagectomy. In addition, we want to discuss the perspective of the possible strategies for immunomodulation during surgery.
The serum IL-6 levels after esophagectomy were significantly higher than those after gastrectomy. The TNFa production of LPS-stimulated monocytes after esophagectomy significantly increased both immediately postoperative and POD3, and TNFa production of LPS-stimulated monocytes after esophagectomy was significantly higher than that after gastrectomy. The HLA-DR expressions on monocytes and the proportion of Tregs (FoxP3+/CD4+) after esophagectomy significantly reduced than those after gastrectomy.
We demonstrated that the preoperative administration of protease inhibitor (Gabexate Mesilate; GM) reduced postoperative serum IL-6 levels compared to those in the postoperative administration of GM. Duration of SIRS was significantly shorter in patients with preoperative GM administration than those in the postoperative administration of GM.
We also demonstrated that immediate enteral feeding (IEN) after esophagectomy showed a significantly earlier recovery of the total lymphocyte count, and attenuated the serum levels of total bilirubin and C-reactive protein, suggesting that IEN may have beneficial effects on immunological competence in patients following esophagectomy.
Minimally invasive surgery such as video-assisted thoracoscopic surgery for esophageal cancer (VATS-e) have been developed, and we investigated the postoperative clinical course of patients who underwent esophagectomy in terms of SIRS induced by VATS-e (VATS-e group) or open surgery (OS group). VATS-e group had a lesser duration of stay in the ICU, a lesser SIRS duration, a lesser number of positive SIRS criteria, and lesser serum IL-6 levels. In addition, the VATS-e group had less postoperative pneumonia.
In conclusion, an esophagectomy for esophageal cancer extremely influences the pro- and anti-inflammatory responses, which may cause postoperative complications. The immunomodulations as the novel strategies for decreasing excessive surgical stress may thus be recommended in patients with esophageal cancer.
THE PREDICTIVE ROLE OF LIPID MEDIATORS FOR POSTOPERATIVE COMPLICATIONS AFTER GASTROINTESTINAL SURGERY Akihisa Matsuda1, Marina Yamada1, Masao Miyashita1, Eiji Uchida2. 1Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 2Department of Surgery, Nippon Medical School, Tokyo, Japan
Backgrounds: Several lipid mediators derived from the metabolism of membrane phospholipids have been established as regulators in inflammatory states including sepsis and cancer. Lysophosphatidylcholine (LPC), which is generated by the action of phospholipase A2 on phosphatidylcholine (PC), has a pivotal role on modulating immune responses via its anti-inflammatory property. LPC is converted to lysophosphatidic acid (LPA) by the enzymatic activity of autotaxin (ATX). The aim of the current study is to investigate the association between LPC and postoperative complications (POC) after gastrointestinal surgery.
Methods: A total of 43 colorectal cancer patients were included in this study and peripheral blood samples were collected from before, immediately after surgery and on postoperative days (POD) 1, 3, 5, and 7. Patients were divided into non-POC (n = 33) and POC (n = 10) groups. Plasma LPC, IL-6, PC, and ATX levels were measured by specific ELISA.
Results: The postoperative LPC levels were decreased (bottom at POD1) and then gradually recovered in both groups. The POC group had significantly lower LPC levels throughout the perioperative period than the non-POC group. The ratio of pre- and post-operative LPC level inversely correlated with IL-6. The predictive impact for POC was demonstrated by ROC analysis (cut off; 51.2%, AUC; 0.795, Figure) and multivariate analysis (OR 15.1, P = 0.01). The PC levels were decreased after surgery in both group, but the decrease is profound in POC group. No significant change was observed in ATX levels in both groups.
Conclusions: Out results suggested that decreased postoperative LPC is associated with exaggerated postoperative inflammatory response and POC occurrence through an inhibition of the anti-inflammatory property. The decreased supply of PC is one of the mechanisms of postoperative decrease of LPC.
BACKGROUND OF MAJOR HEPATO-BILIARY-PANCREATIC SURGERY-INDUCED SEPSIS Yu Katayose1, Ichiro Ise2, Hiroki Hayashi2, Mitsuhisa Mutoh1, Kei Nakagawa2, Takanori Morikawa2, Fuyuhiko Motoi2, Takeshi Naitoh2, Michiaki Unno2. 1Department of Surgery, Tohoku Rosai Hospital, Miyagi, Japan, 2Department of Surgery, Tohoku University Graduate School of Medicine, Miyagi, Japan
Background: Postoperative sepsis is a major factor of surgical morbidity and mortality. However, the rate and epidemiology of postoperative sepsis after hepato-biliary-pancreatic surgery are unknown.
Objectives: We analyzed postoperative sepsis after elective major hepato-biliary-pancreatic surgery in our department.
Methods: Samples of major hepato-biliary-pancreatic surgery were queried between 2011–2015, and patients who experienced sepsis after elective surgeries were identified retrospectively. The procedures of major hepato-biliary-pancreatic surgery were pancreaticoduodenectomy, total pancreatectomy, two sectionectomy and more, and hemihepatectomy with concomitant pancreaticoduodenectomy.
Result: 477 elective surgical cases were analyzed. Males were 298 cases (62.5%) and Females were 179 cases (37.5%). Median age was 67 year old (19 - 87 y.o.). Resection of liver and pancreas was 169 cases (35.4%) and 308 cases (64.6%), respectively. The incidence sepsis was 11.1% (53 cases) in all cases. Sepsis rate of pancreatectomy and liver resection were 7.7% and 22.5%, respectively. Liver resection showed a high incidence of sepsis (p < 0.05). In the liver resection group, concomitant liver resection with bile duct resection cases showed a high incidence of sepsis.
Conclusions: The incident sepsis of major hepato-biliary-pancreatic surgery was 11.1%. Especially, concomitant liver resection with bile duct resection cases had a high incidence. Now, we are analyzing the background details. This may help to prevent morbidity and mortality related to sepsis.
EARLY DETECTION AND TREATMENT FOR POSTOPERATIVE LIVER FAILURE BASED ON THE MECHANISTICAL INSIGHTS Itaru Endo, Ryusei Matsuyama, Ryutaro Mori, Norifumi Kumamoto, Yuka Arisaka, Yusuke Suwa, Kouki Gotoh, Gakuryu Nakayama. Department of Gastroenterological Surgery, Yokohama City University School of Medicine, Kanagawa, Japan
Although the outcomes after major hepatectomy have been improved by advanced surgical techniques and perioperative management, morbidity and mortality after major hepatectomy remains 30–45% and 3–5%, respectively. Postoperative infectious complication often leads to postohepatectomy liver failure (PHLF). Various predictors which can be obtained preoperatively have been reported in order to prevent PHLF, such as indocyanine green retention rate at 15 minutes, Remnant ICGK value, and acialoglycoprotein accumulation etc. Also some comprehensive algorism like Makuuchi's criteria and the prediction score in Hyogo Medical University are widely adopted. However, PHLF cannot be completely avoided by using these preoperative markers alone because intraoperative factors such as ischemic time and bleeding may affect postoperative liver regeneration. Therefore, new biomarkers which are available immediately after liver surgery is needed to predict PHLF and to start definitive treatment much earlier. Our previous studies indicated that hypercytokinemia and suppressed fibrinolysis in the early postoperative phase after extensive hepatectomy is associated with liver failure. It is suggested that both inflammation and coagulation abnormality is involved in the mechanism of liver failure. Possible mechanism of liver failure based on the investigation of perioperative serial changes of pro- and anti-inflammatory cytokines, coagulation and fibrinolytic factors, and also early intervention to prevent PHLF will be discussed.
RELATIONSHIP BETWEEN SURGICAL STRESS INDUCED HYPERGLYCEMIA AND SEPTIC COMPLICATIONS: WHAT IS OPTIMAL PERIOPERATIVE GLYCEMIC CONTROL? Kazuhiro Hanazaki1, Masaya Munekage1, Tomoaki Yatabe2, Hiroyuki Kitagawa1, Tsutomu Namikawa1. 1Department of Surgery, Kochi Medical School, Kochi, Japan, 2Department of Anesthesiology, Kochi Medical School, Kochi, Japan
Objectives: Entity of surgical diabetes associated with operation is surgical stress induced hyperglycemia which is trigger of postoperative infection (POI) including septic complications. Also hypoglycemia associated with tight glycemic control (TGC) is a common and the most serious complications in critically ill patients. This study aimed to evaluate the influences of surgical diabetes to POI and the effects of surgical diabetes treatment using an artificial pancreas (AP).
Methods: From 2006 to 2015, more than 600 surgical patients underwent perioperative glycemic control using a bed side AP with closed-loop system (STG-22 or STG-55, Nikkiso, Tokyo). More than 90% of all the patients performed TGC including intensive insulin therapy (IIT) targeting blood glucose range of 80–110 mg/dL. Data were collected prospectively and were reviewed or analyzed retrospectively.
Results: In conventional perioperative glycemic control, hyperglycemia continued to be within around 20 hours immediately after major surgery even though non diabetic patients. By using AP, all patients undergoing TGC had no hypoglycemia less than 70 mg/dl. Consecutive 305 patients undergoing IIT had not only no hypoglycemia but also high achievement rate of targeting blood glucose range, approximately 90%. Of note, this novel glycemic control revealed stable continuous blood glucose monitoring with less variability of blood concentrations. Also two prospective randomized controlled trials demonstrated that this novel TGC using an AP had better surgical outcomes after hepatectomy and pancreatectomy including POI compared with conventional glycemic control.
Conclusions: Perioperative TGC using an AP is an effective and a safe surgical diabetes treatment to avoid not only hyperglycemia and hypoglycemia but also variability of blood glucose levels, and it may be reduction of POI including septic complications. In the future perspectives, elucidation of optimal glycemic control in various clinical settings with and/or without diabetes including sepsis will be warranted.
THE POTENTIAL OF NEAR-INFRARED SPECTROSCOPY AS AN OXYGENATION INDEX OF PIVOTAL ORGANS AND TISSUE OXYGENATION IN CRITICALLY ILL PATIENTS Yasuyuki Kakihana, Tomotsugu Yasuda, Keizi Yamaguchi, Hiroaki Furubeppu, Chiaki Kamikokuryo, Takahiro Futatsuki, Shinsaku Terada, Shotaro Miyamoto, Yutaro Madokoro. Department of Emergency and Intensive Care Medicine, Kagoshima University, Kagoshima, Japan
Near-infrared spectroscopy (NIRS) was developed as a way to continuously and non-invasively measure tissue oxygenation status and has been used as a cerebral and peripheral tissue oxygenation monitoring method of intraoperative and postoperative patients. Recently, it has been reported that the regional cerebral oxygen saturation (rSO2) by NIRS at hospital arrival can predict neurological outcome at 90 days after out-of-hospital cardiac arrest (1). However, it is not clear whether the cerebral monitoring by NIRS during chest compressions leads to good neurological outcome after resuscitation.
It has been reported that the simultaneous measurement of brain and splanchnic NIRS was able to discriminate whether or not splanchnic ischemia was present in infants with an acute abdomen (2). On the other hand, Schat et al (3) has reported that NIRS monitoring did not proof useful for distinguishing between definite necrotizing enterocolitis (NEC) and no NEC in preterm infants with clinical signs suspicious of NEC. Therefore, it is not clear whether NIRS provides reliable values on their tissue oxygenation of various organs in abdomen (e.g. the liver as a corresponding site in the splanchnic region, which reacts very sensitively to hemodynamic instability), especially adults.
The time resolved spectroscopy (TRS: a new method of NIRS) is a new optical tissue monitor of NIRS from Hamamatsu Photonics K.K (4, 5). The time domain monitor irradiates tissue with pico-second pulses of NIR light. Times of flights of photons are then measured to form a histogram, and estimate the absolute values of absorption coefficient and scattering coefficient, and subsequently derive hemoglobin concentrations and oxygen saturation of tissue.
In this presentation, we will show the two potential of NIRS (TRS) as an oxygenation index of pivotal organs and tissue oxygenation in critically ill patients. Firstly, from the data of piglet cardiac arrest models, we will present the relationship between the cerebral oxygenation by TRS and the way of resuscitation. Secondly, from animal model and clinical data, we will present whether the hemoglobin concentration in the liver and the changes in hepatic oxygenation can be detected by TRS placed on the skin above the liver surface.
1) Ito N, et al. Resuscitation 2014;85:778–784.
2) Fortune PM, et al. Intensive Care Med 2001;27:1401–7.
3) Schat TE, et al. PLoS One. 2016;11:e0154710.
4) Oda M, et al. Phys Med Biol 1996;41:551–562.
5) Gunadi S, et al. Biomed Opt Express. 2014;5:2896–912.
CAPILLARY RECRUITMENT DURING FLUID RESUSCITATION IS LIMITED IN SEPSIS E. Christiaan Boerma1,2, Gerke Veenstra1, Bart W. Barendrecht1, Can Ince2. 1Department of Intensive Care, Medical Center Leeuwarden, Leeuwarden, The Netherlands, 2Department of Translational Physiology, Academic Medical Center Amsterdam, Amsterdam, The Netherlands
Introduction: In-vivo microscopy at the bedside of critically ill patients has revealed a variety of microcirculatory alterations in sepsis and has contributed substantially to the understanding of heterogeneity in microvascular blood flow. In addition microvascular blood flow characteristics have the potential to serve as endpoints for well-known interventions such as fluid resuscitation. We hypothesized that an optimal total vessel density (TVD) may be identified as a stop-sign for fluid administration in sepsis.
Methods: In a single-center observational study we observed 29 sepsis patients during fluid administration. At the maximum positive fluid balance we performed sublingual Incident Dark Field imaging (Cytocam®). 25 Healthy persons served as controls.
Results: Median APACHE III score 85[55–107]; all patients were mechanically ventilated. Median fluid balance in the sepsis group was 8.6[5.8–11.8] liters. TVD was significantly lower during sepsis in comparison to controls (18.7[16.9–20.5] vs 21.4[18.3–23.9], p = 0.019). Comparison between quartiles of fluid balances revealed that the median maximum TVD was 20.1[15.7–21.5] in the second quartile (Fig.).
Conclusion: TVD in sepsis patients is significantly lower in comparison to controls. However, recruitment of capillaries during fluid resuscitation is limited. The maximum TVD may serve as a stop-sign for fluid administration.
THE ROLE OF ARGININE VASOPRESSIN RECEPTORS IN MICROVASCULAR HYPER-PERMEABILITY DURING SEPTIC SHOCK Perenlei Enkhbaatar. Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA
Sepsis is associated with the highest mortality rate in Intensive Care Units. Severe hypotension and organ tissue edema cause tissue hypo-perfusion leading to multiple organ dysfunctions. Various vasopressors are used to antagonize hypotension; however, it often becomes refractory to vasopressors and the use of their high doses increases the morbidity and mortality of septic patients. To date, there is no specific therapy available for management of microvascular hyper-permeability.
There is ongoing debate around the use of arginine vasopressin (AVP) as adjunct therapy to first line vasopressors, such as norepinephrine in septic patients. As previously described, AVP has multiple receptors and we aimed to separately investigate roles of its main receptors i.e., V1a and V2 receptors in microvascular hyper-permeability and cardiovascular collapse during sepsis and septic shock.
Our findings and results of previous studies indicate that V1a receptor agonist effectively maintains arterial blood pressure and reduces/prevents vascular leakage, while V2 receptor activation promotes vascular hyper-permeability and tissue edema. We found nitric oxide and its byproducts i.e., peroxynitrite and potent permeability factor angiopoietin-2 as main players in modulation of vascular permeability by either V1a or V2 receptor activations.
In conclusion, use of nonselective AVP (especially their high doses) should be avoided; instead the use of selective AVP V1a receptor agonist should be considered as a first line or an adjunct vasopressor. The use of selective AVP V2 receptor antagonist should also be considered as an adjunct vasopressor for treatment of septic patients.
INTEGRIN-MEDIATED CROSS TALK BETWEEN COAGULATION AND INFLAMMATION Motomu Shimaoka1, Eiji Kawamoto1, Takayuki Okamoto2. 1Mie University Graduate School of Medicine, Mie, Japan, 2Shimane University, Shimane, Japan
Integrins represent the major cell adhesion molecules that mediate the interactions of leukocytes with inflamed endothelial cells. In response to chemo-attractant stimuli, integrins undergo global conformational changes to the ligand-competent high-affinity state (1, 2). By binding to the primary endothelial ligand intercellular adhesion molecule-1 (ICAM-1), leukocyte integrins support shear-resistant abrupt arrest to, and subsequent crawling along, the surface of the endothelial cells. This eventually culminates to the transmigration across the endothelial cells, thereby leading to the accumulation of activated leukocytes at the interstitial space. Inflammation augments the ICAM-1 expression, while modifying the expression of coagulation-regulating molecules displayed on the endothelial surface such as thrombomodulin, endothelial protein C-receptor (EPCR), and activated protein C (APC). We, and others, have shown that thrombomodulin (3), EPCR (4), and APC (5) not only regulate blood coagulation but also participate in supporting leukocyte adhesion to endothelial cells. We will present our work that determined the structural basis of leukocyte integrin activation (2) as well as the binding to thrombomodulin (3). While lymphocyte function-associated antigen-1 (LFA-1) integrin binds to the membrane-distal part of ICAM-1 (Domain 1), LFA-1 binds to the membrane-proximal part of thrombomodulin (Domain 3). This contrasting spatial localization of the LFA-1 binding sites of ICAM-1 and thrombomodulin might provide a unique adhesive microenvironment on the endothelial surface. We will also present our work that demonstrates the strategies to exploit the structural biology of integrins towards the development of therapeutic small-molecules (6, 7).
1) Park EJ et al. J Biomed Sci. 2015
2) Shimaoka M et al. Cell. 2003
3) Kawamoto E et al. BBRC. 2016
4) Fink K et al. PLoS One. 2013
5) Elphick GF et al. Blood 2009
6) Shimaoka M et al, Nature Rev Drug Discover. 2003
7) Zhang H et al. FASEB J. 2009
S1P2, A RECEPTOR FOR THE LYSOPHOSPHOLIPID MEDIATOR SPHINGOSINE 1-PHOSPHATE, PROTECTS AGAINST VASCULAR BARRIER DISRUPTION Yasuo Okamoto1,2, Hong Cui1, Kazuaki Yoshioka1, Noriko Takuwa1,3, Toshishige Shibamoto4, Yoh Takuwa1. 1Department of Physiology, Kanazawa University Graduate School of Medical Sciences, Ishikawa, Japan, 2Department of Pharmacology, Kawasaki Medical University, Okayama, Japan, 3Ishikawa Prefectural Nursing University, Ishikawa, Japan, 4Department of Physiology II, Kanazawa Medical University, Ishikawa, Japan
Sphingosine 1-phosphate (S1P) binds to S1P-specific G-protein coupled receptor (S1P1∼S1P5) to induce cellular responses. S1P is released from erythrocytes and vascular endothelial cells (ECs) and enriched in plasma. Previous studies demonstrated that plasma S1P maintains barrier integrity via endothelial S1P1. ECs also express S1P2. However, a role of S1P2 in the regulation of vascular barrier integrity was unknown. We found in S1P2−/− mice that vascular permeability increase elicited by intradermal injections of histamine and PAF was augmented in Miles’ assay. Pulmonary vascular leak induced by intravenous injection of PAF was also enhanced in S1P2−/− mice compared with wild-type (WT) mice although the steady-state pulmonary vascular permeability was not different between WT and S1P2−/− mice. Consistently, hypotension, hypothermia and lethality provoked by PAF injection were all augmented in S1P2−/− mice. Furthermore, vascular leak and lethality in an active anaphylaxis model were also exaggerated in S1P2−/− mice. Mechanistically, PAF injection-induced activation of Akt and eNOS in the aorta and lung was enhanced in S1P2−/− mice compared with WT mice. Either genetic eNOS deletion or pharmacological eNOS blockade protected S1P2−/− mice from exacerbation of barrier disruption after antigen challenge and PAF injection. Stimulation of S1P2−/− ECs with PAF and S1P induced exaggerated NO production and disassembly of adherens junctions through S-nitrosylation of beta-catenin compared with WT ECs. The exaggerated responses in S1P2−/−ECs were restored by pharmacological eNOS inhibition. These observations indicate that augmented eNOS stimulation and NO production in anaphylaxis lead to exaggerated barrier disruption through NO-mediated disassembly of adherens junctions in S1P2−/− mice. Administration of LPS provokes acute lung injury, which is also characterized by vascular barrier disruption. LPS-induced permeability increase, leukocyte infiltration, increases in proinflammatory cytokine mRNA expression and lethality were aggravated in S1P2−/− mice compared with WT mice. Interestingly, administration of a NOS inhibitor inhibited exacerbation of LPS-induced leukocyte infiltration and vascular hyperpermeability in S1P2−/− mice. Thus, S1P regulates vascular barrier integrity via multiple S1P receptor subtypes. S1P1 maintains the steady-state vascular barrier integrity while S1P2 protects against vascular barrier disruption on acute insults to the vasculature. S1P2 mediates a barrier-protective effect through suppressing NOS activity.
GLYCOCALYX, ‘MISSING’ LINK BETWEEN ENDOTHELIUM AND MICROCIRCULATION Christa Boer. Department of Anesthesiology, VU University Medical Center, Amsterdam, The Netherlands
Impairment of microcirculatory perfusion is a hallmark of critical illness, and a predictor for patient outcome. Previous research by our group showed that loss of microcirculatory perfusion in patients undergoing cardiac surgery with cardiopulmonary bypass (CPB) is paralleled by endothelial activation, endothelial barrier dysfunction and inflammation. One of the key structures that may be involved in this deterioration of microcirculatory perfusion in is the glycocalyx.
Although there are indications that the integrity of the glycocalyx influences vascular perfusion patterns, this has mostly been shown in experimental animal studies. In particular, perturbation of the endothelial glycocalyx causes a reduction in the number of functional capillaries, but it is unknown whether this phenomenon is of importance in the clinical setting.
In this lecture it is discussed how the glycocalyx might connect endothelial dysfunction and microcirculatory perfusion disturbances based on clinical studies in critically ill patients or patients exposed to cardiopulmonary bypass. Moreover, therapeutic options will be discussed that might contribute to preservation of glycocalyx integrity and microcirculatory perfusion in critical illness.
BLOCKING LIVER AUTOPHAGY RESULTS IN ADVERSE MORTALITY THROUGH ACCELERATED MITOCHONDRIAL INJURY AND HEPATOCYTE APOPTOSIS IN A MURINE SEPSIS MODEL Takehiko Oami1, Eizo Watanabe1, Masahiko Hatano2, Youichi Teratake2, Lisa Fujimura2, Akemi Sakamoto2, Shigeto Oda1. 1Department of Emergency and Critical Care Medicine, Chiba University, Chiba, Japan, 2Biomedical Research Center, Chiba University Graduate School of Medicine, Chiba, Japan
Objective: Autophagy plays an important role for cell survival, sequestering and degrading a wide variety of substrates. Although an increase of autophagosomes in liver was reported in sepsis patients as well as septic mice, neither definite influence of autophagy on liver injury nor the interaction between autophagy and other types of cell death has been unclear. The aims of this study were to elucidate the contribution of liver autophagy to the pathophysiology of sepsis.
Methods: We performed a cecal ligation and puncture (CLP) on liver-specific autophagy deficient (Alb-CreERT2/Atg5f/f) mice (six to eight-weeks-old male). At 24 hours after the operation, we investigated the expression of p62 (a marker protein for insufficient/inhibition of autophagy flux) by western blot analysis. Mitochondrial homeostasis was evaluated by measuring reactive oxygen species (ROS) and the number of mitochondria by flow cytometry. Serum liver enzymes and cytokine concentrations were measured by enzyme linked immune sorbent assay. Immunohistochemistry and transmission electron microscope (TEM) were performed to evaluate liver injury and cell death morphologically. Furthermore, we analyzed subpopulation of activated macrophages and Kupffer cells in the liver by flow cytometry.
Measurements and Main Results: We found a significant accumulation of p62 in the liver of knockout (KO) mice (p < 0.001). Also, mitochondrial accumulation and increase of ROS were observed. Serum aspartate transaminase (AST) levels were markedly elevated in the KO mice (p = 0.005). Furthermore, we found more cleaved caspase-3 positive liver cells in the KO mice by immunohistochemistry. Additionally, we confirmed significant increase of autophagic vacuolization in the control mice, in contrast, cell shrinkage and nuclear fragmentation, which indicate morphological characteristics of apoptosis, were notably seen in the KO mice by TEM. Severe mitochondrial damages were also markedly observed in the KO mice relative to the control mice. On the other hand, the subpopulation of Kupffer cells in the liver was increased in the KO mice. Serum interleukin-6 levels were significantly increased in the KO mice compared with the control mice (p = 0.020). Consequently, deficiency of autophagy in liver significantly decreased the survival rate in the murine sepsis model (p = 0.025).
Conclusions: We demonstrated that blocking liver autophagy increased mortality in the murine sepsis model, suggesting that liver autophagy plays a protective role for organ failure through degradation of damaged mitochondria as well as prevention of apoptosis.
DOES INTERLEUKIN-18 PARTICIPATE IN HYPERGLYCEMIA IN SEPTIC MICE? Hayato Yamashita1, Michiko Ishikawa2, Taketo Inoue1,2, Makoto Usami1, Atsunori Nakao2, Joji Kotani2. 1Department of Biophysics, Kobe University Graduate School of Health Sciences, Hyogo, Japan, 2Department of Emergency and Critical Care Medicine, Hyogo College of Medicine, Hyogo, Japan
Background: Critical illness commonly induces glucose metabolism dysfunction, such as hyperglycemia accompanied with insulin resistance, which is a cause of poor outcome. Interleukin-18 (IL-18) reported to improve the insulin resistance associated with obesity. However the relationship between IL-18 and glucose metabolism in sepsis was unclear. The purpose of this study was to investigate the influence of IL-18 on hyperglycemia during sepsis.
Methods: Polymicrobial sepsis was induced in wild type (WT), IL-18 knockout (KO) mice, and IL-18 KO mice pretreated with recombinant IL-18 using cecal ligation and puncture (CLP). Blood glucose and plasma insulin, glucagon were measured. The mRNA expressions for gluconeogenic enzymes (g6pc, pck1) and protein activation of insulin signaling were also analyzed. Organ damage was histopathologically assessed in the lung, liver, and kidney at 24 hours after CLP.
Results: WT and IL-18 KO mice showed hyperglycemia for 18 hours after CLP operation. And IL-18 KO mice showed significantly higher blood glucose levels than WT mice, although the plasma insulin or glucagon levels of IL-18 KO mice were same as those of WT mice. Insulin signaling in the liver and skeletal muscle was decreased during hyperglycemia in IL-18 KO mice as compared with WT mice without suppression of hepatic glucose production enzymes. At 24 hour after CLP, the WT mice showed hypoglycemia and higher mortality without higher organ damages (lung, liver, and kidney) than IL-18 KO mice. Pretreatment with IL-18 reduced blood glucose levels in IL-18 KO mice after CLP.
Conclusion: Sepsis-induced hyperglycemia may be attenuated by endogenous IL-18 via modulating insulin signals in the liver and skeletal muscle. IL-18 may be a useful indicator of the need to regulate blood glucose during sepsis.
NEW MOLECULAR BIOMARKERS OF ALVEOLAR DAMAGE IN ARDS IN SEPTIC SHOCK PATIENTS Artem N. Kuzovlev1,2,3, Viktor V. Moroz1,2, Arkady M. Goloubev1,2, Vladimir M. Pisarev1,2, Evgeny A. Tishkov3. 1Society for Shock Studies (Russia), Moscow, Russia, 2V.A. Negovsky Research Institute of General Reanimatology, Moscow, Russia, 3Department of Anesthesia and Intensive Care, A.I. Evdokimov Moscow Medical Dental University, Moscow, Russia
Currently no molecular biomarkers of alveolar damage in septic shock are available. The objective of this study was to investigate into the role of new molecular biomarkers of ARDS in septic shock patients.
This observational study in ICU ventilated septic shock patients (sepsis due to peritonitis (70%), pancreonecrosis (25%) and mediastinitis (5%)) was carried out in 2012 and 2016. ARDS was diagnosed and staged according to the V.A. Negovsky Research Institute criteria and the Berlin definition. Plasma surfactant proteins A and D (SPA, SPD) and Club cell protein (CCP) were measured on ARDS diagnosis (day 1) and days 3 and 5 by the immunoenzyme essay (BioVendor, USA). Patients were treated according to the international guidelines. Data were statistically analyzed by STATISTICA 7.0, ANOVA and presented as median and 25–75 percentiles, ng/ml; p < 0.05 was considered statistically significant. Areas under the receiver operating curves were calculated. Eighty patients (out of 487 screened) were enrolled in the study according to the inclusion/exclusion criteria. Patients were assigned into groups: “ARDS” (n = 50, 47 ± 4.8 y.o., mortality 21.3%) and “no ARDS” (n = 35, 39 ± 4.3 y.o., mortality 17,9%). Groups were comparable in APACHE II and SOFA scores on the baseline. There were no statistically significant differences between groups in CCP plasma concentration detected. Plasma SPD on day 1 > 111.2 ng/ml yielded a sensitivity of 68.2% and specificity of 92.3% (AUC 0.85; 95% CI 0.684–0.945; p < 0.0001) for diagnosing ARDS in septic shock patients. PaO2/FiO2 ratio on day 1 < 280 yielded a sensitivity of 94.1% and specificity of 76.9% (AUC 0.89; 95% CI 0.744–0.952; p < 0.0001) and EVLWI on day 1 >8.3 ml/kg yielded a sensitivity of 94.1% and specificity of 92.3% (AUC 0.92; 95% CI 0.810–0.982; pp < 0.0001) for the diagnosis of ARDS. A complex ROC analysis (for SPD in the group of patients with P/F p < 280 and EVLWI>8.3 ml/kg) yielded a much better diagnostic accuracy of SPD: cut-off >93.7 ng/ml, sensitivity 81.0%, specificity 100.0% (AUC 0.96; 95% CI 0.817–0.998; pp < 0.0001). Plasma SPA on day 1 had a good capacity for prediction of mortality in ARDS patients: SPA on day 1 >18.6 ng/ml yielded a sensitivity of 88.9% and specificity of 72.70% (AUC 0.86; 95% CI 0.685–0.962; p = 0.0021).
Conclusion: A complex approach (P/F<280, EVLWI>8.3 ml/kg, SPD>93,7 ng/ml) presents as a sensitive and highly specific method of diagnosing ARDS in septic shock patients. Plasma SPA level >18,6 ng/ml on day 1 is a sensitive and specific biomarker of mortality prediction in ARDS patients. CCP is not a sensitive/specific biomarker of ARDS in septic shock patients.
IκB KINASE INHIBITOR ATTENUATES SEPSIS-INDUCED CARDIAC DYSFUNCTION IN MICE WITH CKD Jianmin Chen1, Julius E. Kieswich1, Fausto Chiazza2, Amie Moyes1, Thomas Gobbetti1, Gareth SD. Purvis1, Nimesh SA. Patel1, Mauro Perretti1, Adrian J. Hobbs1, Massimo Collino2, Muhammad M. Yaqoob1, Christoph Thiemermann1. 1Queen Mary University of London, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, London, UK, 2Department of Drug Science and Technology, University of Turin, Turin, Italy
Rational: Patients with chronic kidney disease (CKD) requiring dialysis have a higher risk of sepsis and a 100-fold higher mortality than the general population with sepsis. The severity of cardiac dysfunction predicts mortality among septic patients.
Objectives: Here we investigated the roles of pre-existing CKD on the cardiac outcome in mice with sepsis, and whether inhibition of IκB kinase (IKK) reduces the cardiac dysfunction in these animals.
Methods: Male C57BL/6 mice were subjected to 5/6th nephrectomy (SNX) for 8 weeks, and were further subjected to either lipopolysaccharide (LPS; 2 mg/kg) or sepsis by cecum ligation and puncture (CLP). CKD mice with sepsis or endotoxaemia received an IKK inhibitor (IKK 16, 1 mg/kg, 1 h-post CLP or LPS administration). Cardiac function was determined using echocardiography at 18 h after LPS injection or 24 h after CLP. To investigate whether the changes in cardiac function was dependent on mean arterial blood pressure (MABP), MABP was recorded in radiotelemetric transmitters-implanted mice or anesthetised mice.
Results: SNX resulted in significant rises in urea and creatinine, and a small (P < 0.05) reduction in ejection fraction (EF), and increases in the cardiac phosphorylation of IκBα, nuclear translocation of the NF-κB subunit p65, iNOS expression, and phosphorylation of Akt and ERK1/2. When subjected to LPS or CLP, CKD mice exhibited exacerbation of (a) cardiac dysfunction (FigA), (b) lung inflammation and plasma cytokine levels (TNF-α, IL-1β, IL-6, IL-10) and (c) phosphorylation of IKKα/β, phosphorylation of IκBα, nuclear translocation of p65 and iNOS expression (heart). IKK 16 attenuated cardiac dysfunction (FigB), lung inflammation, cytokine formation and cardiac phosphorylation of IKKα/β and activation of NF-κB, and increased gross motor activity in CKD mice with sepsis or endotoxemia. IKK 16 increased MABP in CKD/CLP mice. However, IKK 16 did not affect MABP in anesthetised CKD mice, indicating that the higher MABP in IKK 16-treated CKD/CLP mice might be due to improved cardiac function or increased activity.
Conclusions: Pre-existing CKD aggravates the cardiac dysfunction caused by sepsis or endotoxaemia in mice; this may be due to increased cardiac activation of NF-κB and iNOS expression.
GENETIC DEFICIENCY OF APMKα1 EXACERBATES SYSTEMIC INFLAMMATION AND ORGAN INJURY IN SEPTIC MICE Yu Inata, James O’Connor, Michael O’Connor, Paul Hake, Giovanna Piraino, Patrick Lahni, Basilia Zingarelli. Department of Critical Care Medicine, Cincinnati Children's Hospital Medical Center and College of Medicine, University of Cincinnati, Cincinnati, OH, USA
Introduction: AMP-activated protein kinase (AMPK) is a crucial regulator of energy homeostasis, which controls disposal of mitochondria by autophagy and mitochondrial biogenesis by activation of peroxisome proliferator-activated receptor γ coactivator 1α (PGC-1α). Little is known about AMPK-dependent metabolic pathways and their roles in sepsis-induced organ injury.
Objective: Considering the central role of AMPK in metabolic signaling, we investigated whether genetic deficiency in α1 subunit of AMPK exacerbated organ injury in sepsis by impairing energy homeostasis.
Methods: Polymicrobial sepsis was induced in AMPKα1−/− (KO), +/- (HT), and +/+ (WT) 2–3 months old male mice by cecal ligation and puncture (CLP). Blood and organs were collected at 18 hours after CLP for biochemical assays and histopathological examination.
Results: Microscopic examination of lung and liver demonstrated worse organ injuries in AMPKα1 deficient mice. Myeloperoxidase (MPO) assays corroborated the histopathological findings (Table). MPO activity in the lung increased after CLP in all three groups, but more greatly in AMPKα1 deficient mice. Similar increase in MPO activity was also observed in the liver of KO mice. These severe organ damages in AMPKα1 deficient mice were associated with higher inflammatory cytokines in plasma as well as higher NF-κB activation in the nuclear extracts of lung (Table). Next, we investigated whether the observed worse organ injuries were caused by the impairment of AMPK-dependent metabolic pathways and the resultant energy failure. Tissue ATP level was measured in the liver (Table). AMPKα1 deficiency was associated with lower ATP level at baseline. After CLP, ATP levels decreased in all groups, but more severely in KO mice. Unexpectedly, at Western blot analysis using the liver nuclear extracts, AMPKα1 deficiency did not diminish PGC-1α, an important regulator of mitochondrial biogenesis. However, in KO mice, there was an increased expression of SIRT1 that can also activate PGC-1α, which could be a compensatory mechanism to maintain PGC-1α activity. Finally, autophagy process was assessed by measuring the conversion of LC3B-I to LC3B-II, a marker for autophagosome formation. In KO mice, autophagy activity dramatically increased during sepsis, which could be a result of worse organ injury.
Conclusions: AMPKα1 deficiency was associated with severe organ injury and impaired energy production during sepsis. This is likely a result of diminished anti-inflammatory effect of AMPK via NF-κB, rather than impaired AMPK-dependent metabolic pathways.
A NOVEL ROLE FOR PROGRAMMED CELL DEATH RECEPTOR LIGAND-2 (PD-L2) IN SEPSIS-INDUCED HEPATIC DYSFUNCTION Marilyn Le1,2, Chun-Shiang Chung1,2, Yaping Chen1,2, Anne-Lise Rossi1,2, Alfred Ayala1,2. 1Division of Surgical Research, Rhode Island Hospital, Cranston, RI, USA, 2Department of Molecular Pharmacology and Physiology, Brown University, Province, RI, USA
Sepsis is both a costly and deadly state in which dysregulated host response to infection leads to life-threatening organ dysfunction. The programmed cell death receptor 1 (PD-1) pathway has been implicated in sepsis, but the role of its ligands, PD-L1 and PD-L2, are not well understood. In our studies, based on preliminary data indicating that PD-L2 gene expression was altered in the livers of septic mice, we propose that sepsis-induced change in PD-L2 expression in the liver should contribute to altered liver function and subsequently, altered general septic morbidity/mortality. We aim to determine the effects of PD-L2 gene deficiency on septic outcomes in the liver, with respect to what has been established with wild type and PD-L1 deficiency. We also aim to deduce whether these PD-L2 mediated changes in liver function are facilitated by liver permeability/edema formation.
Polymicrobial sepsis was produced by cecal ligation puncture (CLP) in the murine model. From there, we perfused the liver to remove blood cells, isolated non-parenchymal cells and ran various experiments to determine aspects of liver injury/function and selected cell protein expression.
Our results indicate a low basal gene expression level of PD-L2 in organs, with comparably high expression levels in peritoneal macrophages. However, following CLP, PD-L2 expression changed only in the liver. We saw a decrease in its expression after CLP; as opposed to our published findings that PD-L1 expression is increased following CLP. In our preliminary results, we have seen that PD-L2 is expressed primarily in liver sinusoidal endothelial cells (LSECs), with low expression in Kupffer cells (KCs) and hepatocytes. We found significant changes in PD-L2 expression after CLP in LSECs. Our lab has also shown that in sepsis, elevated markers of liver distress can be seen. One of these markers is an increase in vascular permeability, which may be linked to PD-L1 expression based on our published findings. Thus, we propose that PD-L2 might also be a potential mediator of liver permeability.
Having a clearer understanding of PD-L2's roles and functions could give insight into discerning early markers for liver distress and failure, as well as developing new therapeutic targets to alleviate the high mortality rate and other dismal outcomes associated with sepsis.
THE SYNTHETIC ANTIMICROBIAL PEPTIDE 19-2.5 ATTENUATES SEPTIC CARDIOMYOPATHY IN A MURINE MODEL OF POLYMICROBIAL SEPSIS Lukas B. Martin1,2, Elisabeth Zechendorf1, Jianmin Chen2, Klaus Brandenburg3, Gernot Marx1, Chris Thiemermann2, Tobias Schuerholz1. 1Intensive Care and Intermediate Care, University Hospital RWTH Aachen, Aachen, Germany, 2The William Harvey Research Institute, Queen Mary University of London, Barts and the London School of Medicine and Dentistry, London, UK, 3Division of Biophysics, Forschungszentrum, Borstel, Germany
Objectives: Sepsis-induced cardiomyopathy indicates a worse prognosis with mortality rates up to 70%, however causal therapeutics are still missing. The aim of the present study was to evaluate the effects of the newly designed synthetic antimicrobial peptide 19–2.5 (Pep2.5) on cardiac tissue injury, cardiac dysfunction and survival in a murine model of polymicrobial sepsis.
Methods: The local ethics committee (EK.206.09) and the animal protection authority (LANUV-NRW AZ 84-02.04.2014.4300) approved this study. Murine cardiomyocytes (HL-1 cells) were stimulated with serum of septic shock patients (SsP) in the presence or absence of Pep2.5 and cellular injury was investigated after 24 hours. Mice (N = 48) were subjected to cecal ligation and puncture (CLP) or sham operation 48 hours after central venous catheterization and treated either with Pep2.5 (2.0 μg/hour: i.v.) or vehicle (saline 0.9%) as sepsis-control. Cardiac tissue injury and cardiac dysfunction were investigated 24 hours later and survival of the animals was evaluated up to 96 hours.
Results: Stimulating HL-1 cells with SsP resulted in a significant activation of anti-survival signalling as shown by an increased mRNA expression and cleavage of pro-Caspase 3 (both p < 0.0001). SsP stimulation also caused a significant decrease of cell survival, compared to unstimulated cells (p = 0.0048). Pep2.5 protected the cells against both cell death and Caspase 3 activation (all p < 0.0001). Mice subjected to CLP showed a significant increased mRNA expression and cleavage of pro-Caspase 3 in the heart, compared to sham (both p < 0.0001). Pep2.5 treatment protected the animals against Caspase 3 activation as shown by a significant diminished increase of mRNA expression and cleavage of pro-Caspase 3, compared to sepsis-control (both p < 0.0001). Mice treated with Pep2.5 also showed significantly attenuated cardiac dysfunction as shown by higher cardiac output, higher strokework and a higher first derivate of developed pressure (dp/dtmin), compared to sepsis-control (all p < 0.0001). Pep2.5 treatment significantly improved the survival time of the animals (Figure 1, p < 0.0001).
Conclusions: Continuous intravenous administration of the antimicrobial Pep2.5 is able to reduce sepsis-induced cardiomyopathy and to improve survival time in mice. Thus, Pep2.5 may have the potential for further development as a new additive drug in sepsis-induced cardiomyopathy.
ACTIVATION OF TRPV1 BY 12(S)-HPETE AND 20-HETE RELEASES CGRP AND PROTECTS THE HEART AGAINST THE CARDIAC DYSFUNCTION CAUSED BY LPS Jianmin Chen, Alexander JP. Hamers, Michaela Finsterbusch, Christoph Thiemermann, Amrita Ahluwalia. Queen Mary University of London, William Harvey Research Institute, Barts and The London School of Medicine and Dentistry, London, UK
Rationale: The severity of cardiac dysfunction predicts mortality in patients with sepsis. Activation of transient receptor potential vanilloid receptor type 1 (TRPV1), expressed on sensory neurons innervating the heart, improves outcome in sepsis/endotoxaemia. However, the identity of the endogenous activators of TRPV1 and the role of the channel in the cardiac dysfunction caused by sepsis/endotoxaemia is unknown.
Objectives: We investigated the impact of TRPV1 deletion upon the cardiac dysfunction caused by lipopolysaccharide (LPS; endotoxaemia). In addition we explored the role of the arachidonic acid metabolites 12-(s)-hydroperoxyeicosatetraenoic acid (12(s)-HpETE) and 20-hydroxyeicosatetraenoic acid (20-HETE) (potent ligands of TRPV1) and neuropeptides (downstream mediators of TRPV1 activation) in any effects seen.
Methods: TRPV1−/− and wild-type (WT, C57BL6 background) littermate mice were subjected to low (2 mg/kg, i.p.) or high dose LPS (6 mg/kg + peptidoglycan 0.1 mg/kg, i.p.). After 18 h cardiac function was determined using echocardiography. To investigate mechanisms some mice were treated with the CGRP receptor antagonist CGRP8-37 (150 μg/kg; 30 min prior to LPS and 1 and 2 h after LPS, i.v.), the somatostatin receptor antagonist C-SOM (250 μg/kg; 30 min prior to LPS and 1 and 2 h after LPS, i.v.), the 12-lipoxygenase inhibitor cinnamyl-3, 4-dihydroxy-a-cyanocinnamate (CDC; 0.2 mg/mouse, i.p. to block 12(s)-HpETE synthesis) or the cytochrome P450 inhibitor 17-octadecynoic acid (17ODYA; 0.7 mg/mouse, i.p., 20-HETE synthesis inhibitor).
Results: Whilst low dose LPS did not alter cardiac function in WT, in TRPV1−/− mice a pronounced cardiac dysfunction was evident; an effect similar to that evident in WT mice treated with high dose LPS. Blockade of 12(s)-HpETE or 20-HETE synthesis augmented the cardiac dysfunction caused by low dose LPS in WT mice. Low dose LPS caused increases in plasma CGRP and SP levels but a decrease in somatostatin in WT mice; interestingly 17ODYA or CDC/17ODYA co-administration attenuated the rises in CGRP and somatostatin. CGRP8-37 treatment, but not C-SOM, increased cardiac dysfunction induced by low dose LPS, suggesting that the release of endogenous CGRP mediates the cardioprotective action of TRPV1. Western blotting confirmed an increased phosphorylation of TRPV1 in low dose LPS-treated WT mice relative to vehicle control.
Conclusions: Activation of TRPV1, reflected through increased phosphorylation of the channel, by 12(s)-HpETE and 20-HETE likely leads to the release of CGRP, which protects the heart against the cardiac dysfunction caused by LPS.
IMPAIRED DIRECTIONALITY IN CHEMOTAXIS AND SPONTANEOUS MOTILITY AS NEW ASPECTS OF NEUTROPHIL DYSFUNCTION IN BURN PATIENTS: ROLE OF PROTEIN FARNESYLATION Naohide Kuriyama1,2, Marina Yamada1, Yasuyo Shimomura2, Osamu Nishida2, Edward A. Bittner1, Jeremy Goverman1, Masao Kaneki1. 1Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA, 2Fujita Health University School of Medicine, Aichi, Japan
Introduction: Aberrations in neutrophil (PMN) migration impair effective bacterial clearance and cause or exacerbate unnecessary PMN organ infiltration, leading to multi-organ damage. Previous studies have shown that directional PMN chemotaxis is defective while some PMNs spontaneously move in the absence of chemoattractants in burn patients. Nonetheless, the underlying mechanisms are not known. We have shown that farnesyltransferase inhibitor (FTI) reduced the mortality of septic mice. Here, we studied the effects of in vitro incubation with tipifarnib, a clinically applicable FTI, on defective PMN motility in burn patients and the effects of tipifarnib in vivo in septic mice.
Methods: Chemotaxis toward fMLP and spontaneous motility were evaluated in PMNs from patients with >20% TBSA burn injury (n = 7) and healthy controls (n = 8) in the presence or absence of fMLP (10 nM) and tipifarnib (1 μM). To study the effects of FTI in septic mice, male CD-1 mice at weeks of age were treated with tipifarnib (3 mg/kg, sc) or vehicle alone at 2 h after cecum ligation and puncture.
Results: Sustained directionality index and speed during chemotaxis were significantly decreased in burn patient PMNs compared with healthy controls. In the absence of chemoattractants, however, spontaneous amoeboid motility was increased in burn patient PMNs by 2-fold relative to healthy controls. In vitro treatment with tipifarnib for 30 min significantly ameliorated the defective chemotaxis and reversed the spontaneous motility of burn patient PMNs. In contrast, tipifarnib did not alter these parameters in healthy controls. In septic mice, tipifarnib almost completely blocked PMN infiltration into organs (i.e., liver, heart, kidney), while promoting PMN migration to the peritoneal cavity (the site of infection), which paralleled improved survival and bacterial clearance.
Discussion: These results show that impaired directionality and reduced speed during chemotaxis were associated with spontaneous motility in burn patient PMNs, all of which were reversed or ameliorated by tipifarnib in vitro. Consistently, tipifarnib inhibited PMN organ infiltration and promoted PMN migration to the site of infection in septic mice. Our data indicate that protein farnesylation plays an important role in the defective PMN motility in burn patients. Our findings highlight these aberrations as new aspects of PMN dysfunction that may worsen the clinical trajectory. Overall, our study identifies FTI as a novel, potential strategy to prevent infection, sepsis and multi-organ dysfunction by reversing defective PMN motility in burn patients.
PEROXYNITRITE DECOMPOSITION CATALYST ATTENUATES HYPO-RESPONSIVENESS TO NOREPINEPHRINE IN OVINE MRSA SEPTIC SHOCK Satoshi Fukuda1, Koji Ihara1, Anita Randolph1, Ernesto Lopez1, Robert A. Cox2, Hal K. Hawkins2, Salsbury J. Randy1, David Herndon2, Donald S. Prough1, Perenlei Enkhbaatar1. 1Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA, 2Shriners Hospital for Children, USA
Rationale: Septic shock is a frequent cause of death in ICU. Severe cardiovascular collapse during septic shock is refractory to both aggressive fluid and vasopressors. The goal of this study was to test the hypothesis that reactive nitrogen species (e.g. peroxynitrite) play an important role in septic shock-induced vascular hypo-responsiveness to norepinephrine (NE). We hypothesized that peroxynitrite decomposition catalyst (PDC) improves host response to NE and reduces its dose requirement.
Methods: To test our hypothesis, we used our well-characterized ovine septic shock model. Fourteen sheep were chronically instrumented with Swan-Ganz, femoral arterial, left atrial catheters 5–7 days before the induction of injury. Septic shock was induced by a two-hit injury; cotton smoke inhalation and instillation of live MRSA (1.6–2.5 × 1011 CFU) into lung by bronchoscope under anesthesia. After injury, sheep were awakened, placed on ventilator and randomly allocated to two groups: 1) Control; injured, fluid resuscitated and titrated with NE, n = 7; and 2) PDC; injured, treated with PDC, fluid resuscitated and titrated NE, n = 7. PDC (0.1 mg/kg) was administered as a bolus at 1 h post injury followed by continuous infusion (0.04 mg/kg/hr) for 24 hrs. NE started at 0.05 μg/kg/min when MAP dropped by 10 mmHg from baseline, despite aggressive fluid resuscitation with LR solution, and further titrated to maintain MAP close to baseline. Long acting buprenorphine provided analgesia. Intraluminal diameters of mesenteric arteries isolated from healthy sheep were measured after NE treatment with (59.1 nM) with/without peroxynitrite pretreatment (180 mM).
Results: PDC improved survival of septic sheep (71.4% vs. 28.6%). Control sheep developed severe hypotension refractory to fluid resuscitation and required high dose of NE to maintain MAP. The PDC group required significantly lower NE dose during the 24-study period. Total NE requirement was significantly lower in PDC group, mean of the highest NE dose was significantly lower in treatment group as well. PDC had no adverse effects and it effectively reversed severe hypotension. Changes in diameters of isolated arteries following NE treatment were significantly weaker when the vessels were pretreated with peroxynitrite (3.1%) compared to vehicle (26.7%).
Conclusions: Peroxynitrite causes vascular hypo-responsiveness to NE. Its modulation with PDC reduces NE requirements, maintains MAP and improves survival in a clinically relevant ovine model of refractory septic shock. This novel approach may be an attractive option for management of patients with refractory septic shock. (GM097480/SHC84050/SHC85500)
FEATURES OF NEUTROPHIL EXTRACELLULAR TRAP FORMATION IN OUT-OF-HOSPITAL CARDIAC ARREST PATIENTS WITH HYPERFIBRINOLYSIS Johannes Zipperle1,2, Soheyl Bahrami1,2, Heinz Redl1,2, Herbert Schoechl1,2,3. 1Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Vienna Austria, 2AUVA Trauma Research Center, Vienna, Austria, 3Department of Anaesthesia and Intensive Care Medicine, AUVA Trauma Center, Salzburg, Austria
Hyperfibrinolysis (HF) has been shown to occur in out-of hospital cardiac arrest. HF appears to be driven by 1) ischemia-reperfusion-induced activation of plasmin, 2) a downregulation of the protein C pathway 3) the depletion and inhibition of antifibrinolytic mediators. Severe hypoperfusion triggers an early inflammatory response that has been associated with the release of Neutrophil Extracellular Traps (NETs) in critically ill patients. These NETs bare a number of neutrophil (PMN) serine proteases, including elastase which is suspected to interfere with hemostasis and fibrinolysis.
We investigate whether, aside from classical mechanisms, HF in cardiac arrest patients was also associated with the release of elastase and the activation of non-canonical pathways.
Blood samples of 58 patients who suffered from non-traumatic out-of hospital cardiac arrest were drawn on scene by an emergency physician and were analysed by standard coagulation tests and thromboelastometry (ROTEM). Plasma levels of t-PA, PAI-1, sTM, t-PA-PAI-1-complex and APC-PCI-complex were determined with commercially available ELISAs. Furthermore PMN elastase alpha1-Pi and histonylated (h)DNA fragments were measured as an indicator for NET formation. HF was diagnosed with a ROTEM (maximum lysis >15%) and NETosis was defined as elastase and hDNA levels above median. Patients with HF and NETosis were compared with non HF or non-NETosis, respectively.
Nineteen patients presented with ML>15 and 26 patients met the criteria for NETosis. Interestingly t-PA activity but not antigen levels were significantly higher in HF. This was accompanied by an increased formation of t-PA-PAI-1 complex but not with elevated PAI-1 levels. There was a highly significant correlation between elastase and hDNA levels but no association with HF. APC-PCI complex level were elevated in both HF and NETosis.
Conclusions: Elevated t-PA activity but not antigen level is associated with HF in cardiac arrest patients. Both plasmin and the protein C pathway are tightly interwoven in the pathophysiology of HF. Ischemia and reperfusion in cardiac arrest patients appears to result in a NETosis-associated pattern. The interaction of soluble and NET-immobilized Elastase with the coagulation system should be considered in our understanding of HF.
ZONE I REBOA DEPLOYMENT WITH PARTIAL OCCLUSION USING A SMALLER DEVICE MAY LEAD TO FEWER ACCESS RELATED COMPLICATIONS: DIRECT-IABO REGISTRY IN JAPAN Yosuke Matsumura1, Junichi Matsumoto2, Hiroshi Kondo3, Koji Idoguchi4, Tokiya Ishida5, Yuri Kon6, Keisuke Tomita7, Kenichiro Ishida8, Tomoya Hirose9, Kensuke Umakoshi10, Tomohiro Funabiki11. 1R Adams Cowley Shock Trauma Center, University of Maryland, Baltimore, MD, USA, 2St. Marianna University, Kanagawa, Japan, 3Teikyo University, Tokyo, Japan, 4Senshu Trauma and Critical Care Center, Osaka, Japan, 5Ohta Nishinouchi Hospital, Fukushima, Japan, 6Hachinohe City Hospital, Aomori, Japan, 7Chiba University, Chiba, Japan, 8NHO Osaka National Hospital, Osaka, Japan, 9Osaka University, Osaka, Japan, 10Ehime University, Ehime, Japan, 11Saiseikai Yokohamashi Tobu Hospital, Kanagawa, Japan
Introduction: Resuscitative endovascular occlusion of the aorta (REBOA) has been gaining acceptance as a feasible and less invasive resuscitation procedure alternative to resuscitative thoracotomy. The purpose of this study is to evaluate current REBOA practice in Japan.
Methods: The DIRECT-IABO study group registered hemorrhagic shock patients requiring REBOA from 18 hospitals. Patients’ characteristics, outcome, complications as well as the size of the sheath were collected and analyzed. Sheath size <=8Fr and >=9Fr were respectively defined as small and large.
Results: From Aug 2011–Dec 2015, 142 REBOA patients (Male 64%, median BMI 23) with sites of hemorrhage commonly found in the abdominal (55%), pelvic (50%), chest (21%) and retroperitoneal (13%) regions. Pre-occlusion systolic blood pressure (SBP) was 60 mmHg, and 22% of patients required CPR at REBOA deployment. Following aortic occlusion, SBP improved to 110 mmHg (p < 0.001). Improved hemodynamics with REBOA was observed in 82% of cases and 70% were stabilized (SBP consistently>90 mmHg).
The common REBOA access were percutaneous without imaging (92%), US-guided (2.8%) and femoral cut-down (2.1%). In 77% of cases, imaging (US 15%, plain film 29%, and fluoroscopy 32%) were utilized for guidewire confirmation within the aorta. Zone I deployment was achieved in 95% while balloon migration and rupture occurred in 2.1% and 0.7% respectively. Partial occlusion (titrating the balloon inflation to regulate SBP>90 mmHg) was performed in 68% with a median 12 min duration while total occlusion duration was 50 min.
Seventy six patients survived >24 hours and of these n = 53 had small sheaths placed for 19 hours while n = 25 were placed large sheaths for 7.5 hours (p = 0.02). The remaining 3 patients were placed either exchanged oversized or multiple sheaths.
Upon sheath removal, external manual compression was performed in 94% of the small group and 45% of the large group (p < 0.001). Common femoral artery stenosis resulting from thrombus formation was identified in 1 patient of the large group while 2 cases of lower limb ischemia followed by fasciotomy and amputation were identified in the oversize group.
Conclusion: Japanese REBOA management showed tentative zone I placement with partial occlusion as a substantial option in REBOA practice. Smaller sheath might have fewer complications with external compression even after long placement.
BEWARE OF THE GRAND DELUSION: THE (POLY)TRAUMA HIT REQUIRES FUNCTIONAL VERIFICATION OF ITS IMMUNO-INFLAMMATORY PHENOTYPE AND OF OUTCOME EFFECT UPON THE SECONDARY HIT Susanne Drechsler, Johannes Zipperle, Pia Rademann, Mohammad Jafarmadar, Soheyl Bahrami, Marcin Osuchowski. Department of Intensive Care, Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Vienna, Austria
Polytrauma activates and then severely impairs the immune response, predisposing patients to secondary infections/sepsis. Only the most severe mouse polytrauma models appear to properly reproduce those immuno-inflammatory impairments. New mouse polytrauma models combine various elements (as the 1st hit). Yet, we often fail to verify the true impact of the 1st hit polytrauma-induced phenotype on the 2nd hit sepsis outcome. Thus, we compared the effect of two mouse polytrauma models of increasing severity on 1) the immuno-inflammatory phenotype, 2) phagocytic capacity and 3) 2nd hit sepsis outcome. We also studied the impact of splenectomy as a polytrauma element.
Female Balb/c mice (n = 84) underwent 1) severe polytrauma (SPT) by femur fracture (FF), hemorrhagic shock (HS; 30% blood loss) and splenectomy, and 2) moderate polytrauma (MPT) by FF and HS. Blood was collected at 0, 24 and 48 h post-trauma. Absolute/relative counts of granulocytes, lymphocytes, CD4+, CD8+ and regulatory T-cells, monocyte MHC-2 expression and leukocyte phagocytic capacity were assessed. Additional SPT/MPT mice underwent 2nd-hit cecal ligation and puncture (CLP) 48 h after the 1st-hit.
CLP sepsis alone caused 32% mortality by day 28. The MPT exacerbated the 2nd-hit CLP mortality to 56% (p < 0.05). Remarkably, the severe polytrauma model (i.e. SPT with splenectomy) reduced the 28-day mortality to 8% (p < 0.05). Within 48 h, both SPT and MPT induced a distinct neutrophilia (5 and 2.5-fold increase) and lymphocytosis (1.3 and 2-fold increase; p < 0.05). Only the more severe SPT induced a 1.5-fold increase in the absolute CD4+ and CD8+ T-lymphocyte counts (p < 0.05) at 48 h; contrasted by a 30% decline in the relative CD4+ and CD8+ counts (p < 0.05). In SPT mice at 48 h, both absolute and relative counts of regulatory (immunosuppressive) T-cells (CD4+CD25+CD127-) increased by 1.5-fold (p < 0.05). Monocyte MHC-2 expression increased by 46% within 48 h post-SPT, while MPT had no effect. Similarly, phagocytic capacity of macrophages/granulocytes isolated from the whole blood increased by 54% in SPT by 48 h, but was unchanged in MPT mice.
The greater SPT severity did not exacerbate the post-traumatic sepsis mortality but lowered it, while milder MPT worsened it. Splenectomy caused a strong immunosuppressive T-reg phenotype coinciding with a robust immune system activation, contrasted by far milder effects after MPT. We advise a mandatory end-effect verification of the existing (poly)trauma hits (as the whole and of individual elements) before they are deemed as clinically relevant and used in any 2-hit model system.
By FWF Hertha-Firnberg grant.
A NOVEL SYNTHETIC PEPTIDE PROTECTS AGAINST ORGAN INJURY IN A RAT-MODEL OF SEVERE HAEMORRHAGIC SHOCK Noriaki Yamada1, Lukas Martin1, Regina Sordi1, Klaus Brandenburg2, Tobias Schuerholz3, Christoph Thiemermann1. 1Centre for Translational Medicine and Therapeutics, The William Harvey Research Institute, Queen Mary University of London, London, UK, 2Division of Biophysics, Forschungszentrum, Borstel, Germany, 3Department of Intensive Care and Intermediate Care, University Hospital RWTH, Aachen, Germany
Objectives: Haemorrhagic shock (HS) is still a common cause of death in severely injured patients and is characterized by impairment of organ perfusion, systemic inflammatory response, and multiple organ failure (MOF). Although there are clear clinical guidelines for the management of HS and organ support strategies, there is no specific pharmacological intervention that prevents the MOF associated with HS. Thus, the aim of the present study was evaluate potential therapeutic effects of the newly developed synthetic peptide 19–4LF (Pep-4LF) on haemodynamic and organ injury/dysfunction in rats subjected to HS.
Methods: Male Wistar rats (220–350 g, Charles River, UK) were anesthetized with sodium thiopentone (induction: 120 mg/kg i.p, maintenance: 10 mg/kg i.v.). HS was induced by withdrawal of blood from a catheter placed in the carotid artery to reduce mean arterial blood pressure (MAP) to approximately 30 mmHg for 90 min. At 90 min after initiation of haemorrhage (or when 25% of the shed blood had to be re-injected to sustain target MAP), resuscitation was performed with the (remaining) shed blood over a period of 5 min. The same volume of blood reinfused for maintenance of MAP was replaced by Ringer lactate on resuscitation. Rats received infusions of Pep-4LF low dose; 66 μg/kg × h(Pep4-LFLD group), high dose 333 μg/kg × h(Pep4-LFHD group) or vehicle (saline; HS group) throughout the 4 h resuscitation period. At the end of the experiment, we evaluated biochemical parameters of organ injury and dysfunction.
Results: In rats subjected to HS treated with vehicle, the mean values for MAP fell throughput the resuscitation period. Treatment of HS-rats with low dose Pep-4LF had no significant effect on MAP, but MAP (at 4 h) was higher in HS-rats treated with the high dose of Pep-4LF (P < 0.05). In rats subjected to HS treated with vehicle, there was a significant rise in urea, creatinine and a decline in creatinine clearance and, hence, renal dysfunction. This renal dysfunction was attenuated by the high dose of Pep-4LF (P < 0.05; Figure 1). Similarly, HS resulted in liver injury (rise in AST/ALT), pancreatic injury (rise in lipase) and organ ischemia (rise in lactate), all of which were significantly attenuated by the high dose of Pep-4LF (P < 0.05; Figure 1).
Conclusion: Thus, the high dose of Pep4LF improved hemodynamic (MAP) and reduced the renal dysfunction, liver injury and pancreatic injury associated with HS.
APPLYING ADAPTIVE MACHINE LEARNING TO DYNAMIC MULTI-OUTCOME PREDICTION AFTER CRITICAL INJURY S. Ariane Christie1, Alan Hubbard2, Amanda S. Conroy1, Rachael A. Callcut1, Mitchell J. Cohen1. 1Department of Trauma Surgery, University of California San Francisco, San Francisco, CA, USA, 2Department of Biostatistics, University of California, Berkeley, CA, USA
Introduction: Machine learning techniques are used in non-medical fields to rapidly distill large amounts of data into actionable dynamic prediction. The objectives of this study were to evaluate if big data analytics could 1) be used as a novel tool to rapidly assess risk for key outcomes after severe injury and 2) identify dynamic patient-specific causative variables which could be modified to improve trajectory in critically-ill patients.
Methods: SuperLearner, an ensemble machine-learning algorithm, was applied to 2-million observations of severely-injured patients. A set of >1000 unbiased clinical predictors were used to generate prediction models from multiple candidate learners for outcomes of interest at serial time points post-injury. To prevent overfitting, models were prospectively cross-validated and evaluated with ROC curves. Clinical variables responsible for driving outcomes (variable importance) were extracted from high performance models at each time point and compared across time.
Results: SuperLearner models demonstrated excellent prediction of death, multi-organ failure (MOF), and transfusion across multiple post-injury time points, and good prediction of acute lung injury (ALI) and venous thromboembolism (VTE) [Figure A]. Outcomes with inferior data quality, including coagulopathic trajectory, demonstrated sub-optimal prediction, establishing a ceiling for prediction optimization and highlighting the necessity of high-fidelity data inputs even with machine learning approaches. Variable importance evolved over time, suggesting that prediction is driven by dynamic clinical factors that vary in predictive power across the clinical trajectory. Key clinical predictors included both anticipated and unexpected variables, emphasizing the importance of utilizing an unbiased approach to predictor selection. In particular, non-random missingness of data was identified as a strong predictor of multiple outcomes over time.
Conclusions: SuperLearner is a powerful tool which can help clinicians rapidly integrate the voluminous, evolving data currently available on severely-injured patients into potentially real-time, dynamic decision-making support. It avoids the risk of overfitting seen with traditional statistical approaches used for outcome prediction and holds significant promise for optimizing precision medicine approaches to critical care medicine.
THE PREDICTIVE VALUE OF THE LYMPHOCYTE-TO-MONOCYTE RATIO ON INFECTIOUS COMPLICATIONS AFTER LAPAROSCOPIC COLORECTAL CANCER SURGERY Fumihiko Ando1, Akihisa Matsuda1, Satoshi Matsumoto1, Nobuyuki Sakurazawa1, Kumiko Sekiguchi1, Marina Yamada1, Masao Miyashita1, Eiji Uchida2. 1Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 2Department of Surgery, Nippon Medical School, Tokyo, Japan
Backgrounds: Recently, systemic inflammation-based markers such as NLR, PLR, and mGPS have been well defined as prognostic factors for various cancers. The lymphocyte-to-monocyte ratio (LMR) is relatively newly identified and is reported to have a potential to predict long-term survival. However, the predictive value of LMR for short-term outcomes after cancer surgery is not reported so far. The aim of this study is to explore the predictive potential of the preoperative LMR on postoperative infectious complications (POI) after laparoscopic colorectal cancer surgery.
Methods: From Jan 2014–Aug 2015, consecutive 211 cases were performed laparoscopic colorectal cancer surgeries in the Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital and included retrospectively in this study. The association of clinicopathological variables and systemic inflammation-based markers including NLR, PLR, and LMR with POI were analyzed and then multivariate logistic regression analysis was conducted.
Results: The overall rate of POI is 16.6%. Included patients were divided into non-POI (n = 176) and POI (n = 35) groups. Univariate analyses demonstrated that sex, body mass index, smoking, conversion to open surgery, surgery duration, transfusion, simultaneous resection, blood albumin, and LMR were significantly different between the groups. The lower LMR (cut-off 3.46), longer surgery duration, and smoking were identified as independent prognostic factors for POI by multivariate analysis (LMR: odds ratio 3.97, 95% CI 1.44–10.9, P = 0.008).
Conclusions: The lower LMR was established as a predictive factor for POI after laparoscopic colorectal cancer surgery first time ever. This finding could confer a future treatment plans of preoperative modulations for surgeons to prevent POI.
ESTABLISHING EXTRACORPOREAL MEMBRANE OXYGENATION TEAM PROGRAM IMPROVES CLINICAL RESULTS IN EXTRACORPOREAL LIFE SUPPORT PATIENTS Atthasit A. Komindr1,2, Ryuzo Abe1, Yoshihisa Tateishi1, Yuka Takahashi1, Jun Goto1, Keita Wada1, Yutaka Furukawa1, Atsushi Sugiura1, Taro Imaeda1, Natsumi Suga1, Noriyuki Hattori1, Shigeto Oda1. 1Department of Emergency and Critical Care Medicine, Chiba University Hospital, Chiba, Japan, 2Emergency Unit, King Chulalongkorn Memorial Hospital, Thai Red Cross Society, Bangkok, Thailand
Objective: Extracorporeal life support (ECLS) is a rescue procedure involving use of extracorporeal membrane oxygenation (ECMO) in critical patients. Successful results could be achieved with well-coordinated teamwork even under limited time constraints in stressful situations. We sought to identify clinical effects following the establishment of ECMO team.
Methods: This was a retrospective cohort study performed at a Tertiary care hospital in Japan. Comparisons were made between results in pre-ECMO team cases (PRE) (10/2011-9/2012) and post-ECMO team cases (POST) (10/2014-9/2015). Patients resuscitated by ECLS were included; patients on which ECLS was started by central ECMO were excluded. We collected demographic data, ECMO event, clinical time process, ventilator settings and laboratory results.
Results: Of 65 ECLS patients, median age was 61 years (42-73), ECMO treatments for 61 (94%) were started with veno-arterial fashion and 51 (79%) were subject to extracorporeal cardiopulmonary resuscitation (ECPR). There was a greater number of ECLS cases per year in POST than in PRE (40 vs. 25). Patient's demographic data did not differ between groups except for the median APACHE II score which in POST was lower than PRE, 39 (35–41) vs. 35 (27–39), P = 0.02. The POST had significantly more data record completion compared to the PRE: ECMO process time recorded was 23% and 8%, P < 0.01. Activated clotting time at 6 hours after initiation of ECMO tended to be closer to target of 180 in POST than PRE, 185 (161–201) vs. 202 (179–214), P = 0.06. Survival to discharge among both groups did not differ, 19/40 (48%) vs. 12/25 (48%), P = 0.89. Ventilator days and length of hospital stay among patients who survived were not differ, while both tended to be shorter in POST than PRE, 8 (4–11) vs. 11 (7–20) ventilator days (P = 0.15), and 51 (22–72) vs. 67 (32–125) hospital stay days (P = 0.21). ECMO duration in survived patients were not different between groups, 2 (0–3) days in POST vs. 3 (1–5) days in PRE (P = 0.43).
Conclusions: As demonstrated by comparative data, following establishment of ECMO team the number of ECLS cases increased and clinical data collection improved. However, survival rates, ventilator days and lengths of hospital stay in pre- and post-ECMO team cases were not different. Identification of clinical factors from collected data would be the key to augmenting ECLS survival rates and cost effectiveness. Further study is therefore needed.
DYNAMICS OF PLATELET-LEUKOCYTE AGGREGATION THROUGHOUT HEMORRHAGIC SHOCK AND RESUSCITATION: IMPLICATIONS FOR TRAUMA-INDUCED COAGULOPATHY Johannes Zipperle1,2, Katrin Altenburger1,2, Nikolaus Hofmann1,2, Mostafa Ashmwe1,2, Claudia Keibl1,2, Martin Ponschab1,2, Christoph Schlimp1,2, Soheyl Bahrami1,2, Heinz Redl1,2, Herbert Schoechl1,2. 1Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, Vienna, Austria, 2AUVA Trauma Research Center, Vienna, Austria
Platelets are the main effectors of hemostasis and impaired platelet responsiveness is a feature of trauma-induced coagulopathy. Trauma hemorrhage and resuscitation (THR) result in ischemia-reperfusion injury, recruitment of leukocytes and platelet activation. Aggregation of platelets with leukocytes has been described in a number of conditions, including major trauma, SIRS and sepsis. However, the dynamics of platelet-leukocyte aggregation (PLA) in THR and the effect on platelet hemostatic function has not been studied in detail.
We aimed to investigate 1) whether THR promotes the formation of PLA 2) which selective states of cellular activation favoured the aggregation of platelets with leukocyte subtypes, and/or 3) whether this interaction is associated with an impairment of platelet hemostatic function.
Ten Sprague Dawley rats were subjected to trauma-haemorrhage and were resuscitated with Ringer's lactate. Leukocyte CD11b expression as well as platelet P-Selectin positivity (PLA formation) were determined by flow cytometry. To study PLA formation in vitro, whole blood of 10 healthy human donors was stimulated with selective and general platelet and leukocyte agonists and analyzed by flow cytometry. Leukocyte subsets were identified by morphology and specific antibodies. Platelet-mediated hemostatic function in whole blood was measured by thromboelastometry (ROTEM) and impedance aggregometry (Multiplate). Complete blood counts (CBC) were performed to determine changes in platelet numbers.
We found that PLA formation undergoes changes throughout THR. At the end of shock PLAs decreased significantly but tended to re-emerge when resuscitation measures were taken. Resuscitation was accompanied by the recirculation of activated leukocytes. When different states of cellular activation were simulated in vitro, platelet aggregation with monocytes was associated with a reduced blood clot firmness (MCF) in ROTEM and impaired platelet responsiveness to TRAP and ADP (AUC) in Multiplate. Aggregation with monocytes occurred also when only platelets were activated, whereas the interaction with granulocytes required a profound co-activation of both cell types. None of the observed phenomena was paralleled by a reduction of measurable platelet counts.
Conclusion: PLA formation is a dynamic process that is altered throughout THR. Furthermore, our findings suggest that PLA formation is associated with impaired platelet hemostatic function and does not require the acute up-regulation of leukocyte integrins. PLA formation does not appear to be derivable from platelet numbers in CBC.
IMPAIRMENT OF HEMATOPOIETIC STEM CELL ACTIVATION DURING THE GRANULOPOIETIC RESPONSE TO BACTEREMIA IN MICE WITH CHRONIC BINGE ALCOHOL ADMINISTRATION Ping Zhang1, Xin Shi1, Yuan-Ping Lin2. 1Integrative Medical Sciences, Northeast Ohio Medical University, Rootstown, OH, USA, 2National Defense Medical Center, School of Pharmacy and Graduate Institute of Pharmacy, Taipei City, Taiwan
During bacterial infection, hematopoietic stem cells (HSCs) are activated to enhance their commitment to granulocyte development. This granulopoietic response is critical for reinforcing host defense. Excessive alcohol consumption injures hematopoietic tissue and impairs immune defense. To determine the effect of chronic binge alcohol exposure on the HSC activation during the granulopoietic response, acute alcohol intoxication was introduced [via intraperitoneal injection (i.p.) of 20% alcohol in saline at a dose of 5 g alcohol/kg] to mice fed on the Lieber-DeCarli low fat liquid alcohol diet for 5 weeks. Bacteremia was induced in animals by intravenous injection (i.v.) of Escherichia coli (∼1×106 or ∼5×107 CFUs/mouse) 30 min post i.p. alcohol. Bacteremia caused a remarkable increase in the number of marrow HSCs bearing lin-c-kit+Sca-1+ markers. Activation of cell proliferation supported the increase in marrow HSCs following E. coli infection. Alcohol administration inhibited this increase in marrow HSCs. In the process of granulopoiesis, cell expression of granulocyte differentiation antigen-1 (Gr1) increases with the maturation of granulocytes. In control mice, the bone marrow stored a large amount of mature granulocytes (Gr1hi cells). In response to bacteremia, the bone marrow released granulocytes into the systemic circulation. In the meantime, proliferation of granulocyte precursor cells was enhanced. The ratio of immature Gr1lo cells in the bone marrow was markedly increased following bacteremia. Alcohol administration impaired these activities associated with the granulopoietic response. In vitro culture of primary hematopoietic stem/progenitor cells (HSPCs) bearing lin-c-kit+ markers with lipopolysaccharide (LPS) increased cell proliferation. Alcohol exposure inhibited the LPS-stimulated cell proliferation. Bacteremia caused rapid activation of the ERK-cyclin D1 signal pathway (signaling for hematopoietic cell proliferation). In addition, expression of C/EBPb by marrow lineage negative precursor cells was significantly increased following bacteremia. Systemic infection with E. coli also tended to up-regulate C/EBPa expression by hematopoietic precursor cells. Members of the C/EBP family are master transcription factors for granulopoiesis. Alcohol administration impaired these signaling mechanisms. Our results demonstrate that chronic binge alcohol exposure inhibits the granulopoietic response through disrupting key signal systems regulating primitive hematopoietic precursor cell activation and reprogramming.
Supported by NIH grants R01AA019676 and R01AA022816.
METHANE BIOACTIVITY - AND ITS ROLE IN MAMMALIAN PHYSIOLOGY Mihaly Boros, Andras Meszaros, Petra Hartmann, Gerda Strifler. Institute of Surgical Research, University of Szeged, Szeged, Hungary
Many gases within the human body are biologically active, but contrary to classical pathways of signal transduction, the exact roles of gaseous compounds in the mediation of extra- or intracellular events are still not completely understood. Methane is part of the gaseous environment which maintains the aerobic metabolism within the eukaryote cell. It is widely recognized that large amounts of methane can be produced by the anaerobic methanogenic flora in the gastrointestinal tract of mammals, and methane is present in measurable amounts in the breath of approximately one-third of humans. Nevertheless, as opposed to the previous view, in vitro and in vivo studies have revealed the possibility of non-microbial methane formation in both plants (Keppler et al. Nature 2006) and animals (Tuboly et al. Am J Physiol 2013), and the overall evidence from these findings suggests that the excretion of methane in the breath in mammals reflects not only intestinal bacterial fermentation, but also unidentified nonbacterial generation induced from target cells.
Furthermore, it was also demonstrated that methane can protect the tissues by mitigating the effects of an ischemia-reperfusion insult (Boros et al. Crit Care Med 2012). In this animal model normoxic ventilation with methane decreased the signs of oxidative and nitrosative stress and reduced the structural damage of the mucosa. These data were validated by independent observers; subsequent publications have confirmed the biological effects of methane in various ischemia-reperfusion settings and stress conditions (Ye et al. Shock 2015, Wu et al. BBRC 2015, Chen et al. Free Radic Biol Med 2016, He et al. BBRC 2016), and demonstrated the anti-inflammatory and antiapoptotic properties of methane-based treatments.
Besides, the inhalation of methane-containing normoxic artificial air preserved the mitochondrial oxidative phosphorylation after a period of liver ischemia, and significantly improved the basal mitochondrial respiration state after the onset of reperfusion. In line with this, the ischemia-reperfusion-induced oxygen radical production, cytochrome c activity and hepatocyte apoptosis were also reduced significantly (Strifler et al. PLoS One 2016).
In conclusion, the results presented to date indicate a bioactive role for methane. The immediate challenge is therefore to test new hypotheses on the underlying mechanisms which are still not known with certainty.
Supported by Hungarian Science Research Fund OTKA K104656.
NEUROMUSCULAR ELECTRICAL STIMULATION (NMES) CAN IMPROVE LIPID METABOLISM AND SURVIVAL OF THE ACUTE ENDOTOXIC SHOCK MICE Takayuki Irahara1,2,3, Norio Sato2, Kosuke Otake1,2, Kazuo Inoue3, Kaoru Koike2, Hiroyuki Yokota1. 1Graduate School of Emergency and Critical Care Medicine, Nippon Medical School, Tokyo, Japan, 2Department of Primary Care and Emergency Medicine, Kyoto University, Kyoto, Japan, 3Laboratory of Nutrition Chemistry, Division of Food Science and Biotechnology, Graduate School of Agriculture, Kyoto University, Kyoto, Japan
Objectives: We have revealed that low-intensity exercise in the acute phase of endotoxic shock mice might exert a therapeutic effect such as improving lipid metabolism and survival by stimulating PGC-1α expression (PMID:26953756). In this study, we investigated whether neuromuscular electrical stimulation (NMES) also have a similar effect.
Methods: C57BL/6 mice were given 20 mg/kgBW of lipopolysaccharide (LPS) intraperitoneally and divided into C (control) group and NMES groups (n = 4–8/group). Intermittent neuromuscular electrical stimulation to bilateral gastrocnemius muscle was performed to NMES groups for 1 hour. Frequency and voltage were set to low-frequency(LF = 2 Hz) or high-frequency(HF = 50 Hz) and low-voltage(LV = 10 V) or high-voltage(HV = 50 V). Furthermore, in LF-HV group, NMES was performed twice (NMES*2) to compare with once (NMES*1). The alterations of energy metabolism were measured using indirect calorimetry until 24 hours after NMES. Survival proportions were also measured until 72 hours after LPS administration.
Results: Marked decrease of energy metabolism and worsening of survival by LPS was observed in C group. Almost no change was observed in LF-LV group and HF-LV group compared with C group. In LF-HV group, fatty acid oxidation (FAO), oxygen consumption (VO2) and resting energy expenditure (REE) were slightly increased. In HF-HV group, FAO and VO2 and REE were markedly increased (p < 0.01) and CHO was slightly increased. Nevertheless, survival proportions tended to be worsened in HF-HV group and improved in LF-HV group. Surprisingly, in NMES*2 group, FAO was significantly increased (p < 0.01) and survival was also significantly improved (p < 0.05) compared with C and NMES*1 group.
Conclusions: These results suggest that NMES can alter the energy substrate metabolism and survival of the acute endotoxic shock mice depending on its conditions and exert a therapeutic effect especially when performed twice in low-frequency and high-voltage. These effects may occur through PGC-1α activation like exercise intervention.
AN69ST-CRRT: AN EFFECTIVE BRIDGE THERAPY FOR PEDIATRIC MACROPHAGE ACTIVATION SYNDROME IN SHOCK Nobuyuki Nosaka1,2, Masato Yashiro1, Tomonobu Kikkawa1, Yousuke Fujii1, Kohei Tsukahara2, Emily Knaup2, Toyomu Ugawa2, Atsunori Nakao2, Hirokazu Tsukahara1. 1Department of Pediatrics, Okayama University Hospital, Okayama, Japan, 2Advanced Emergency and Critical Care Center, Okayama University Hospital, Okayama, Japan
We report a case of macrophage activation syndrome (MAS) treated with continuous hemodiafiltration with a cytokine-absorbing hemofilter, AN69ST. The patient was an eight-year-old girl diagnosed with systemic juvenile idiopathic arthritis (s-JIA) four years ago, now under treatment with tocilizumab (TCZ), a humanized anti-human interleukin (IL)-6 receptor antibody. She presented to our hospital with complaints of fever and vomiting. She was alert, but apparently in shock. Respiratory distress and hypovolemic shock were aggressively treated with mechanical ventilation, fluid challenge, antibiotics, vasoactive drugs, and blood transfusion. She was diagnosed with MAS and given 30 mg/kg of intravenous (iv.) methylprednisolone. During the next three hours, the patient's clinical condition remained critical with continuous volume demand, an extremely high ventilator setting, and no diuresis. As a reno-protective and anti-inflammatory strategy, we started continuous renal replacement therapy (CRRT) with an AN69ST hemofilter. During the following three hours, the patient's vital signs progressed with decreased volume demand, improved ventilator setting, and better diuresis. 30 mg/kg of iv. methylprednisolone was completed on day 3. Inotropic drugs were titrated on day 2. Ventilator weaning was successful on day 6. No microorganisms developed. She was discharged from the intensive care unit on day 8 without complications.
We prospectively analyzed the trends of different cytokines and the serum concentration of TCZ using her frozen serum obtained prior to and during AN69ST CRRT. A significant decrease was observed in the level of multiple cytokines with high baseline serum concentrations (IL-1b, IL-6, IL-8, IL-10, IL-18, interferon gamma-induced protein-10, and monocyte chemotactic protein-1). In addition, the concentrations of serum TCZ obtained just before and 16 hours after starting CRRT were 24.2 mcg/ml and 21.6 mcg/ml, respectively.
MAS is a life-threatening disorder caused by uncontrolled proliferation and activation of macrophages and T-lymphocytes. The key treatment strategy is allaying the cytokine storm. Conventionally, immunosuppressive agents are responsible for the treatment; however, they require time to take effect. This report showed that AN69ST-CRRT could be a useful bridge therapy for MAS in shock by supporting renal function and reducing serum cytokine levels. Moreover, AN69ST hemofilter will not affect the serum concentration of TCZ. CRRT with AN69ST membrane would be a promising bridge therapy when s-JIA patients, even treated with TCZ, suffer from distributive shock with a differential diagnosis of septic shock and MAS.
THE IMPACT OF HYPOXIC CONDITION AND PENTOXYFILLINE IN SH-5YSY CELLS Young-Duck Cho1,2, Sung-Hyuk Choi1,2, Ziang Qiuyu2, Ji-Min Park1, Jung-Youn Kim1. 1Department of Emergency Medicine, Korea University, Seoul, Korea, 2Institute of Trauma, College of Medicine, Korea University, Seoul, Korea
Objective: Lots of traumatic patients admit the emergency department. These patients with massive hemorrhage, respiratory failure, and further that the experience can fall into hypovolemic shock. Thereafter, oxygen is important to essential treatment in trauma patients. However, we don’t know whether how much oxygen dose in pre-hospital, hospital treatment. Therefore, we conducted an experiment to apply hypoxia, hyperoxia and other medicine with oxygen for decreasing hypoxic brain damage using to MTT, apoptosis, hydrogen peroxide and hypoxia-inducible factor (HIF) in brain cells.
Methods: The experiments were performed with SH-5YSY cells. First, SH-5YSY cells put through normoxic state, hypoxic state, hyperoxic state each times and measured the MTT, apoptosis, hydrogen preoxide by FACScan, HIF by western-blot method. Second, SH-5YSY cells divided into hypoxia, hypoxia-hyperoxia state, hypoxia-hyperoxia-other medicine each 2 hours based on the MTT, apoptosis, hydrogen peroxide and HIF was measured.
Results: The hypoxic group than in the normoxic decreased MTT and increased apoptosis. However, there were no difference between hypoxia state and hypoxia-hyperoxia state. Steroid, hypertonic saline, PTX with oxygen group a little incrased MTT and decreased apoptosis than in the hypoxic group and there were not statistic difference. However, PTX, hypertonic saline and steroid decreased hydrogen peroxide expression in hypoxic condition, and also PTX and hypertonic salne decreased HIF expression in hypoxic sondition.
Conclusions: The in vitro oxygen supply in SH-5YSY cells may not be difference in hypoxia state in the aspect of MTT, apoptosis. However, PTX, hypertonic saline may have effect in SH-5YSY cells at hypoxia state, especially hydrogen peroxide, HIF.
This manuscript is not previously published and prepare manuscripts according to the instructions published in SHOCK.
CIRCULATING CELL-FREE HEMOGLOBIN CAUSES ACUTE LUNG INJURY AND INCREASES MORTALITY IN MURINE POLYMICROBIAL SEPSIS Lorraine B. Ware, Ciara M. Shaver, Nathan Putz, Jamie Kuck, Melinda Paul, Julie A. Bastarache. Department of Medicine, Vanderbilt University, Nashville, TN, USA
Objective: Cell-free hemoglobin (CFH) levels in the circulation are elevated in sepsis patients and are associated with increased mortality. However, the specific mechanisms of worse clinical outcomes are unknown. We developed a titratable, non-surgical model of polymicrobial intraperitoneal sepsis supplemented with intravenous (IV) CFH to test the hypothesis that circulating CFH causes endothelial injury, organ dysfunction and increased mortality in sepsis.
Methods: Sepsis was induced in C57Bl/6 mice by intraperitoneal (IP) injection of a cecal slurry (CS, 1.4–2.0 mg CS/g mouse) of cecal contents collected from donor mice and suspended at 80 mg/mL in 5% dextrose. Purified human CFH was delivered IV at a dose of 0.15 mg CFH/g at the same time as CS. Control mice received IP 5% dextrose and IV PBS. For survival experiments, mice were monitored every 6 h until moribund. Other endpoints were measured at 24 h. Lung endothelial permeability was assessed by accumulation of a fluorescent-labeled macromolecule (Angiosense, 70 kD) in excised organs. Acute lung injury was assessed by protein concentration in bronchoalveolar lavage (BAL) fluid and whole lung cytokine expression by semi-quantitative PCR. Endothelial dysfunction was assessed by measurement of endothelial biomarkers (E-Selectin, ICAM-1, PECAM-1, P-Selectin, PAI-1, Thrombomodulin). Circulating biomarkers of inflammation were also measured (KC, IFN-gamma, IL-6, IL-10).
Results: CS decreased survival in mice in a dose-dependent manner, demonstrating that the CS provides a titratable model of severe polymicrobial intraperitoneal sepsis that does not require a surgical procedure. Supplemental IV CFH significantly diminished survival compared to CS alone at both the 1.8 and 2.0 mg/g doses (Figure). CFH plus CS significantly augmented lung permeability with increased accumulation of Angiosense in the lung and increased BAL protein as compared to CS alone. CFH plus CS also increased lung mRNA expression of the pro-inflammatory cytokines TNF-alpha, IL-6, IL-10 and KC. Circulating markers of endothelial dysfunction were significantly higher in CFH plus CS compared to other groups as were circulating proinflammatory markers (KC, IFN-gamma, IL-6, IL-10).
Conclusion: Intravenous injection of CFH to mimic the elevated levels of plasma CFH seen in human sepsis augments the severity of illness in a model of polymicrobial intraperitoneal sepsis and increases lung inflammation and permeability. Elevated markers of endothelial dysfunction suggest that one potential mechanism of the effects of CFH is through adverse effects on endothelial function that augment inflammation and organ dysfunction. CFH represents a novel therapeutic target in severe sepsis.
Support: NIH HL103836 and HL126671
NEUTROPHIL EXTRACELLULAR TRAPS INDUCE ORGAN DAMAGE DURING SEPTIC SHOCK: PROTECTIVE EFFECTS OF HEMOPERFUSION WITH A POLYMYXIN B CARTRIDGE Muhammad A. Huq, Yuka Kajita, Yuuki Maruchi, Teruaki Miyake, Yuusuke Kato, Tsuguaki Terajima, Hisatake Mori, Masamitsu Hashiba, Atsutoshi Tomino, Ruri Aoki, Masanobu Tsuda, Hideki Kano, Naoshi Takeyama. Department of Emergency and Critical Care Medicine, Aichi Medical University, Aichi, Japan
Background: Neutrophil extracellular traps (NETs) are formed in response to infection. Although NETs would possess both physiological and pathological roles during sepsis, no comprehensive study elucidating the correlation between septic shock severity and NETs formation has been conducted in septic shock. Hemoperfusion with a polymyxin B cartridge (PMX) is being employed for the treatment of septic shock. The exact mechanism of PMX effect for septic shock is not elucidated. Accordingly, the aims of this study were 1) to determine the contribution of NETs for septic shock, and 2) to evaluate the protective effect of PMX for septic shock by removing circulating NETs component.
Methods: Before and after PMX we took samples from septic shock patients at three time points (days 1, 3 and 7). We measured circulating myeloperoxidase (MPO)-associated DNA by a sandwich ELISA method to quantify NETs. We also evaluated sequential organ failure assessment (SOFA) score, P/F ratio, and mean arterial pressure (MAP) at the time of sample collection and survival.
Results: A total number of 43 septic shock patients were treated in the ICU by PMX. Plasma levels of MPO-DNA were 8.17, 6.34 and 3.68 AU/l for days 1, 3 and 7, respectively. SOFA score at day 7 was positively correlated with MPO-DNA at day 3 (p = 0.01) and 7 (p = 0.017). MPO-DNA at day 3 shows negative correlation with P/F ratio (p = 0.003) and MAP (p = 0.025) at day 7. Higher concentration of MPO-DNA at day 3 showed poorer outcome (p = 0.019).
Comments: Our results indicate that excess NET formation is correlated with sepsis severity which may play some part in the pathogenesis of septic shock. Further studies are needed to clarify the direct effect of PMX for removal of circulating components of NETs.
B-CELLS AND NEUTROPHILS PROMOTE DISEASE TOLERANCE TO MURINE GRAM NEGATIVE SEPSIS Riem Gawish1, Barbara Maier2, Karin Lakovits3, Anastasiya Hladik3, Ana Korosec4, Ildiko Mesteri5, Fiona Oakley6, John Brain6, Irene Lang7, Sylvia Knapp3,4. 1Department of Biomedical Science, Institute of Animal Breeding and Genetics, University of Veterinary Medicine Vienna, Vienna, Austria, 2Icahn School of Medicine at Mount Sinai, Hess Center for Science and Medicine, New York, NY, USA, 3Department of Medicine 1, Lab. of Infection Biology, Medical University of Vienna, Vienna, Austria, 4CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria, 5Pathology Ueberlingen, Baden-Wuerttemberg, Germany, 6Institute of Cellular Medicine, Fibrosis Laboratory, Newcastle University, Newcastle upon Tyne, UK, 7Internal Medicine II, Division of Cardiology, Medical University of Vienna, Vienna, Austria
Despite the availability of antibiotics, sepsis is still a life threatening condition characterized by uncontrolled systemic inflammation, coagulation and subsequent organ failure. Disease tolerance is a previously unrecognized host defense strategy, next to the well-studied strategies of avoidance and resistance, and involves mechanisms aiming to improve the host fitness without directly influencing pathogen burden.
We have established a model of disease tolerance, where mice are long-term (up to 5 weeks) protected from organ damage during Gram-negative sepsis. As the protected group shows the same pathogen burden as the control group, we are now able to study mechanisms of disease tolerance uncoupled from effects that depend on the bacterial load. Using cell-depletion strategies and genetically modified mouse models we want to identify cells that mediate organ-protection during sepsis.
So far we could show that neutrophils and platelets are the main contributors to sepsis-induced organ damage. Disease tolerant mice show dampened early inflammation, less neutrophil recruitment into organs and are subsequently protected from micro thrombus formation and liver failure. This goes along with a profound expansion of splenic B-cells and an increase of total IgM in the serum of tolerant mice, which is also consumed during the subsequent bacterial challenge. Given that the long-lasting protective effect is abolished in Rag2 knockout mice, we propose that IgM, or B-cells directly, might suppress inflammation and neutrophil migration, thereby improving host fitness, while maintaining efficient bacterial clearance. The final identification of distinct cells and potential mediators of disease tolerance might possibly allow for novel therapeutic approaches in patients suffering from sepsis.
RELATIONSHIP BETWEEN INTESTINAL BACTERIAL TRANSLOCATION AND ACTIVATION OF INTESTINAL LAMINA PROPRIA DENDRITIC CELLS AFTER TRAUMA/HEMORRHAGIC SHOCK Yun Zhang1, Wenqiao Yu1, Wei Wu2, Fangfang Huang2, Yi Zhang2. 1The Second Affiliated Hospital of Medical College, Zhejiang University, Hangzhou, China, 2The First Affiliated Hospital of Medical College, Zhejiang University, Hangzhou, China
Introduction: A subset of CD11chiCD11bhi lamina propria dendritic cells (LPDCs) has been identified that expressed Toll-like receptor 5 (TLR5) in the small intestine. After stimulated by flagellin, TLR5+ LPDCs can induce the differentiation of naive CD4+T cells into interleukin 17-producing T helper cells (Th17) and type 1 T helper cells (Th1). In this study, we examined the relationship between intestinal bacterial translocation and the activation of lamina propria dendritic cells (LPDCs).
Methods: B6/C57 mice (wild type or Tlr5 KO) were grouped as: A: T/HS (wt); B: SHAM (wt); C: T/HS (Tlr5−/−); D: SHAM (Tlr5−/−). After an open mid-diaphyseal transverse fracture been created, mice were bled to achieve an MAP of 30 mmHg for 90 min, and finally they were resuscitated with lactated Ringer's solution (see fig.C1-3, D). LPDCs subsets enriched from the small intestine were sorted on the basis of high expression both of CD11c and CD11b with FACS (see fig.C4). After being stimulated by LPDCs from each group, the naive CD4+T cells sorted from intestine were underwent T cell differentiation (Th1/Th17) analysis with flow cytometer (see fig.B). Homogenates of MLNs, livers, spleens and blood were cultured 24hr after intragastric administration with bioluminescent citrobacter, and bacterial colony forming units (CFU) were determined by IVIS Spectrum optical imaging system (see fig.A).
Results: In wild type mice, T/HS can induce a higher ratio of citrobacter translocation to liver and blood, but there is low ratio of citrobacter translocation in TLR5 KO mice with or without T/HS. 48 hours after T/HS LPDCs induced lower differentiation ratio of CD4+T cells towards Th1 and Th17 compared with SHAM group in wild type mice, on the contrary, the differentiation ratio maintain a stable low level in TLR5 KO mice with or without T/HS.
Discussion: We put forward the hypothesis that TLR5+LPDCs may work against bacterial infection by inducing local T helper cell responses through TLR5 stimulation, but they will become fragile when suffering T/HS. On the other hand, in TLR5 KO mice, some compensatory immune mechanisms which may be insensitivity to T/HS have taken effect in preventing bacterial translocation.
THERAPEUTIC POTENTIAL OF CELL SECRETOMES IN TRAUMA & HEMORRHAGE Arian Bahrami1, Carina Penzenstadler1, Mostafa Ashmwe1, Anton Klotz1, Mohammad Jafarmadar1, Mohammad Mahdi Kasiri2, Hendrik Jan Ankersmit2, Heinz Redl1, Soheyl Bahrami1. 1Ludwig Boltzmann Institute for Experimental and Clinical Traumatology, AUVA Research Center, Vienna, Austria, 2Department of Thoracic Surgery, Medical University of Vienna, Vienna, Austria
Background & Aim: Trauma & hemorrhage (TH) is a leading cause of death worldwide and often associated with inflammatory response, cell and organ injury. We evaluated the therapeutic potential of a) a newly developed blood-cell secretome preparation (APOSEC) and b) the secretome obtained by conditioning adipose-derived stem cells (AdSC-Secretome), to ameliorate inflammation, cell and organ injury, and to improve the outcome after TH in rats.
Methods: Animals subjected to TH and a resuscitation protocol, mimicking pre-hospital setting, received APOSEC, AdSC-secretome, or vehicle intravenously 20 minutes after onset of reperfusion.
Results: APOSEC and the AdSC-Secretome both modulated the immune response, reflected by circulating cytokine levels (IL-6, IL-10, MCP-1). Similarly, TH-induced cell injury (LDH, CK) and liver damage (ALT) were significantly ameliorated in both treatment groups. The 28-day mortality (25%) was prevented by APOSEC treatment (100% survival) but was not affected by the AdSC-Secretome.
Conclusion: We show for the first time that supplemented resuscitation with cell-derived secretomes ameliorates TH-related inflammation, cell- and organ-injury. Moreover, APOSEC may prevent TH-induced long-term mortality in rats.
KLOTHO DEFICIENCY EXAGGERATES ENDOTOXEMIC CARDIAC DYSFUNCTION IN AGING MICE Xianzhong Meng, Haipeng Hui, Yufeng Zhai, Lihua Ao, David A. Fullerton. Department of Surgery, University of Colorado Denver, Aurora, CO, USA
Background: Endotoxemia occurs frequently following major surgeries and elder subjects with sterile endotoxemia display worse cardiac dysfunction. However, the mechanism underlying this age-related worse cardiac performance is incompletely understood. Klotho, an anti-aging protein primarily expressed in the kidney, is recently implicated in the modulation of inflammatory responses.
Objective: This study was to test the hypothesis that Klotho deficiency augments the inflammatory responses and thereby contributes to the mechanism underlying the worse cardiac dysfunction in elder subjects.
Methods: Lipopolysaccharide (LPS, 0.5 mg/kg, iv) was injected into adults (4–6 months) and aging (18–20 months) C57/BL6 mice. At 24, 48 or 96 h after injection of LPS, plasma and myocardial tissue were collected following left ventricle (LV) function analysis with a pressure-volume catheter. ELISA was used to analyze cytokines (TNF-alpha, IL-1beta and IL-6) and chemokines (MCP-1 and KC). Immunoblotting was applied to determine Klotho levels in the myocardium, liver and kidney. Recombinant Klotho (0.01 mg/kg, iv) was administered to a group of aging mice 30 min after injection of LPS to evaluate its therapeutic potential.
Results: In comparison to adult mice, aging mice not only had more severe cardiac contractile depression at 24 h, but also displayed impairment of LV function recovery at 48 and 96 h. Such worse cardiac performance in aging endotoxemic mice was associated with augmented mononuclear cell accumulation and cytokine production in the myocardium. Interestingly, Klotho protein was detectable in the heart. The heart and kidney, not the liver, of aging mice had markedly lower levels of Klotho in comparison to adult mice, and endotoxemia caused a further reduction of Klotho levels in aging hearts. Treatment of aging endotoxemic mice with recombinant Klotho reduced myocardial mononuclear cell accumulation and cytokine production, and significantly improved cardiac function at all time points.
Conclusion: The augmented inflammatory responses and worse cardiac dysfunction in aging endotoxemic mice are associated with Klotho deficiency. Post-treatment with recombinant Klotho suppresses myocardial inflammation and improves cardiac function in aging endotoxemic mice. These novel findings suggest that Klotho deficiency contributes to the mechanism underlying the worse outcome of elder subjects with sterile endotoxemia and that recombinant Klotho has therapeutic potential in amelioration of endotoxemic cardiac dysfunction.
THE LOW FALL AS A SURROGATE MARKER OF FRAILTY IS A RISK FACTOR FOR READMISSION IN OLDER TRAUMA PATIENTS: A RETROSPECTIVE COHORT STUDY Ting Hway Wong1, Hai Van Nguyen2, Marcus Eng Hock Ong3, Dennis Chuen Chai Seow4, Ming Terk Chiu5. 1Department of General Surgery, Singapore General Hospital, Singapore, 2Duke-National University of Singapore Graduate Medical School, Singapore, 3Department of Emergency Medicine, Singapore General Hospital, Singapore, 4Department of Geriatric Medicine, Singapore General Hospital, Singapore, 5Department of General Surgery, Tan Tock Seng Hospital, Singapore
Background: Frailty is associated with adverse outcomes including readmissions, mortality and risk of falls. Frail patients who fall comprise a significant proportion of the injured in ageing populations and are likely to have different outcomes compared to non-frail injured patients. We hypothesized that a low fall, from 0.5 metres or less, is a surrogate marker of frailty and predicts readmission rates in older injured patients.
Methods: Our primary outcome was all-cause readmission (planned and unplanned) within six months of the index hospitalization for the injury. Using Singapore National Trauma Registry data for 2011–2013, we analysed a cohort of adults aged over 55, admitted via the emergency department in public hospitals, sustaining blunt injuries with an injury severity score (ISS) of 9 or more, excluding isolated hip fractures from same-level falls in the over 65. We excluded patients who died within six-months of discharge by matching to the death registry. Our registry uses standardized conversion metrics (furniture, steps, buildings) to convert patient histories to fall heights. A variable for each hospital was included in the final model to adjust for inter-hospital variation in detecting and recording readmission.
Patients injured by a low fall were compared to patients injured by high-velocity blunt mechanisms. Logistic regression was used to analyze the six-month readmission rate, adjusting for ISS, revised trauma score (RTS), complications significant on univariate analysis, Charlson co-morbidities score, gender and age.
Results: Of our cohort of 6165 patients, those who were injured after low falls were more likely to be readmitted to hospital within six months, compared to those sustaining injuries by high-velocity blunt trauma and higher fall heights (OR 1.64, 95% CI 1.29–2.09, p < 0.001). On multivariable analysis, after adjusting for ISS, RTS, age, gender and co-morbidities, and complications that were significant on univariate analysis, low fall patients were at increased risk of readmission (OR 1.54, 95% CI 1.15–2.08, p < 0.001, Table 1). Falls that were higher than 0.5m did not increase risk of readmission on univariate and multivariable analysis. Urinary tract infection, ISS and Charlson comorbidity score were also independent predictors of readmission.
Conclusions: The low fall is a risk factor for six-month readmission for older patients after moderate and severe injury, and may be a surrogate marker of frailty.
GLYCOCALYX DAMAGE IN EMERGENCY DEPARTMENT PATIENTS WITH INFECTION, COMPARED TO TRAUMA Lisa Smart1,2, Stephen PJ. MacDonald1,2, Sally Burrows3, Erika Bosio1,2, Glenn Arendts1,2, Daniel M. Fatovich1,2. 1Centre for Clinical Research in Emergency Medicine, Harry Perkins Institute of Medical Research, Perth, WA, Australia, 2Department of Emergency Medicine, University of Western Australia, Perth, WA, Australia, 3Department of Medicine and Pharmacology, University of Western Australia, Perth, WA, Australia
Background: Endothelial glycocalyx damage (EGD) biomarkers are associated with poor outcomes in ICU patients with sepsis but there is little information on behaviour of these biomarkers in Emergency Department (ED) patients with infection.
Objectives: 1) To explore three biomarkers of EGD in ED patients with infection over the first 24 hours, 2) to compare these results to a trauma group, and 3) to explore associations with illness severity, comorbidities and clinical outcomes in patients with infection.
Methods: Banked serum samples were used from patients with simple infection (n = 22), sepsis (43), septic shock (45) and trauma (30) collected at: enrolment in the ED (T0), 1 hour (T1), 3 hours (T3) and 12–24 hours (T24) later. Healthy controls (n = 29) were included for T0 comparisons. Biomarkers syndecan-1 (SYN1), syndecan-4 (SYN4) and hyaluronan (HA) were measured via ELISA. Results were obtained using regression methods including random effects linear, logistic and truncated negative binomial methods. Significance was set at P < 0.05.
Results: SYN1 concentration at T0 was significantly higher in all groups compared to control (median 494pg/ml, [Q1-Q3 382–709]). Septic shock (3064pg/ml, [1645–7116]) and sepsis (2542, [1096–5263]) groups had significantly higher concentrations than simple infection (1207, [695–1647]) and trauma (1404, [1055–2265]). SYN1 increased between T0 and T24 in the simple infection and septic shock groups, which was significantly different to trauma.
SYN4 was not different among groups at T0. A decrease in SYN4 in the simple infection group at T3 was the only between-group difference.
HA concentration was significantly higher in the septic shock group at T0 (128 ng/ml, [62–425]) than simple infection (32, [13–89]), sepsis (62, [22–135]), trauma (45, [15–82]) and healthy control groups (41, [24–60]). HA increased between T0 and T3 in infection groups, whereas HA decreased between T0 and T3 in trauma.
Patients with liver disease or alcohol abuse had significantly higher SYN1 and HA over time, compared to others. Patients with vascular disease failed to show a peak in HA concentration at T3, compared to others. Higher SYN1 and HA at T0 significantly increased the odds of ICU admission and higher sequential organ failure assessment score.
Conclusions: ED patients with sepsis and septic shock had higher levels of EGD biomarkers compared to those with simple infection or trauma. Differing patterns of EGD occurred over the first 24 hours of hospitalisation for infection, compared to trauma. Increased SYN1 and HA was also related to key comorbidities, ICU admission and organ failure.
RELATIONSHIP OF ORGAN DYSFUNCTION AND MORTALITY IN SEPTIC SHOCK PATIENTS USING THE SEPSIS-3 DEFINITIONS Yasuyuki Tsujita1, Takayuki Kato2, Naomi Kitamura1, Tetsunobu Yamane1, Kazunori Fujino1, Ryoichi Mandai1, Mikiko Matsushita2, Takahisa Tabata1, Kan Takahashi3, Kazuhiro Matsumura2, Yutaka Eguchi1. 1Department of Critical and Intensive Care Medicine, Shiga University of Medical Science, Shiga, Japan, 2Department of General Medicine, Shiga University of Medical Science, Shiga, Japan, 3Department of Anesthesiology, Shiga University of Medical Science, Shiga, Japan
Background: The definitions of sepsis and septic shock were revised in the third international consensus definitions (Sepsis-3) and organ dysfunction was represented by an increase of in the Sequential Organ Failure Assessment (SOFA) score of 2 points or more. The relationship between each organ damage and mortality of septic shock patients using Sepsis-3 definitions has not been fully reported.
Methods: We studied all septic shock (Sepsis-3) patients over 16-year-old entered to the ICU of Shiga University of Medical Science Hospital (Otsu, Japan) from April 2011 to March 2015 (n = 179) retrospectively and evaluated the relationship between the organ-specific points of the SOFA score on admission and in-hospital mortality.
Results: The mean age of the patients was 66 ± 15 years and 67% were male. The mean APACHE II and SOFA score on admission were 24 ± 10 and 12 ± 4, respectively. The mean ICU stay was 12 ± 11 days. The 28-day, 90-day, and in-hospital mortality rate was 35%, 43%, and 44%, respectively. The figure shows the relationship of the organ specific point of the SOFA score and in-hospital mortality. In-hospital mortality was gradually increased with the rise of the organ-specific points of the SOFA score in respiration, coagulation, and renal systems. If each organ failure was defined as the organ-specific points of the SOFA score of 2 points or more, the percentage of organ failure was 100% in cardiovascular, 81% in respiration, 64% in coagulation, 45% in renal, 46% in central nervous system (CNS), and 38% in liver system. Multivariate logistic analyzes with the co-variables of age, sex, and each organ failure excepting cardiovascular system revealed that respiration failure (hazard ratio 5.2; 95%CI, 1.3–29; P = 0.02) and renal failure (hazard ratio 4.1; 95%CI, 1.5–12; P = 0.004) were independent predictor of in-hospital death.
Conclusions: The organ-specific points of the SOFA score on admission of 2 points or more in respiration and renal systems were independent predictor of in-hospital mortality in septic shock patients.
THE EFFECT OF ANTITHROMBIN III ON LIPOPOLYSACCHARIDE-INDUCED NEUTROPHIL EXTRACELLULAR TRAPS Michiko Ishikawa1, Hayato Yamashita2, Nobuki Oka1, Noritomo Fujisaki1, Taihei Yamada1, Naomi Manbo1, Takahiro Ueda1, Joji Kotani1. 1Department of Emergency, Disaster and Critical Care Medicine, Hyogo College of Medicine, Hyogo, Japan, 2Department of Biophysics, Kobe University Graduate School of Health Sciences, Hyogo, Japan
Background: Neutrophil extracellular traps (NETs) are the new aspects of neutrophil function of eliminate pathogens. NETs has beneficial role on the early phase of sepsis, however, NETs or their components has some effect on coagulation disorder or organ failure in septic patients. We focused on the effect of antithrombin (AT), which is an anticoagulant that also has anti-inflammatory activities, on NETs.
Methods: Male C57BL/6J mice were injected intraperitoneally with 20 mg/kg lipopolysaccharide (LPS; Escherichia coli O111:B4) and 250 IU/kg ATIII under anesthesia. The ATIII was injected 3 hours after LPS injection. At 18 hour after LPS injection, mice were sacrificed and then obtained lung tissues to make paraffin sections. Four milliliter of whole blood was obtained from healthy volunteers. Neutrophils were isolated using MACSxpress neutrophil isolation kit (Miltenyi Biotech) and stimulated by LPS (E.coli O55:B5, 1 μg/mL). ATIII (10 IU/mL) was added at 1 hour after LPS stimulation. Cells were harvested on collagen coated 8 well chamber slide for 4 hours in 37°C 5% CO2. Then the cells were fixed with 4% paraformaldehyde. Tissue sections and harvested neutrophils were stained immunofluorescence staining using anti-histone H1 antibody and anti-elastase antibody as primary antibody. The quantification of NETs areas were used Image J software.
Results: the NETs areas of mice endotoxemia were significantly ameliorated by AT III administration (LPS group, 2.0 ± 0.95 mm2; LPS+ATIII group, 1.1 ± 0.81 mm2; p < 0.05 by Tukey-Kramer's post hoc test). In healthy volunteers, LPS-induced NETs formation (34.5 ± 31.06 μm2) was significantly ameliorated by ATIII (20.8 ± 13.22 μm2, p < 0.05 by paired t-test).
Conclusion: ATIII decreased LPS-induced NETs formation both in in vivo and in vitro.
A NEW THERAPEUTIC CONCEPT IN ABDOMINAL SEPTIC SHOCK Takuya Ueno1,3, Toshiaki Ikeda1, Takayoshi Yokoyama2, Yu Kihara2, Osamu Konno2, Yuki Yokoyama2, Hitoshi Iwamoto2, Shigeyuki Kawachi2, Tetsunosuke Shimizu3, Martina M. McGrath3, Anil Chandraker3. 1Division of Critical Care and Emergency Medicine, Tokyo Medical University Hachioji Medical Center, Tokyo, Japan, 2Department of Digestive and Transplantation Surgery, Tokyo Medical University Hachioji Medical Center, Tokyo, Japan, 3Transplantation Research Center, Renal Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA, USA
Severe sepsis places a large burden on health care systems, with a short-term mortality of 20% to 25%, reaching up to 50% when shock is present. Therefore, there is an urgent unmet clinical need to develop novel therapies to improve patient outcomes and survival rate.
In Japan, we have access to various Continuous Renal Replacement Therapies (CRRTs) such as PMX-DHP and PMMA-CHDF in patients with abdominal septic shock. In the current study, we investigated whether these CRRTs improve patient outcomes and how…
We measured levels of IL-6, PAI-1, IL-1ra, and HMGB-1 at different time points following PMX-DHP +/- PMMA-CHDF and found that 1. significant reduction of some cytokines/mediators with different manners following CRRTs and 2. employing CRRTs could provide us a new therapeutic concept as a cytokine modulation in abdominal septic shock.
LOW DOSAGE ASPIRIN ATTENUATED MULTIPLE ORGAN FAILURE IN SEPSIS THROUGH VASCULAR ENDOTHELIAL PROTECTION Takahiro Usui1, Hideshi Okada1, Kodai Suzuki1, Chihiro Takada1, Risa Inagawa1, Go Furui1, Hiroki Uekado1, Hiroki Jinno1, Junya Ota1, Yasuaki Hotta2, Taku Tanaka1, Shiho Nakano1, Takahiro Yoshida1, Shozo Yoshida1, Nagisa Miyazaki2, Hiroaki Ushikoshi1, Izumi Toyoda1, Genzou Takemura2, Shinji Ogura1. 1Emergency and Disaster Medicine, Gifu University Graduate School of Medicine, Gifu, Japan, 2Internal Medicine, Asahi University School of Dentistry, Gifu, Japan
Introduction: Sepsis is associated with vascular endothelial disorders, the formation of blood clots in small blood vessels, multiple organ failure and death. Aspirin is typical of Non-Steroidal Anti-Inflammatory Drugs (NSAIDs) and the low dosage aspirin controls the platelet aggregation. Recent report showed that aspirin therapy was reduced risk of acute respiratory distress syndrome mortality.
Objective: To demonstrate that low dosage aspirin attenuated multiple organ failure in sepsis.
Methods & Results: Ten-week-old C57BL6 male mice were given aspirin (2 mg/kg/day, n = 23) intraperitoneally for 5days. As control, mice were given ethanol (2 mg/kg/day, n = 30) in the same manner. On the 3rd day of administration, both groups were given lipopolysaccharide (LPS, 15 mg/kg). Forty-eight hours after LPS administration, the survival ratio was significantly higher in aspirin injected group mice compared with control mice (87 vs. 20%, p < 0.05). Frozen fracture samples were made from kidney, lung and liver obtained from these mice, and these samples were observed using scanning electron microscopy (SEM, HITACHI S-4500). In control mice, the destruction of lung, kidney and liver capillaries, with edematous endothelial walls and fibrin deposition were observed 48 hours after LPS injection in ultrastructural analysis using SEM. In aspirin treated mice, the destruction of lung, liver and kidney capillaries was attenuated compared with control group. To further confirm vascular endothelial injury, western blot analysis and immunohistochemical analysis were performed. Although thrombomodulin was expressed on the surface of healthy endothelial cells, the expression was decreased in injured endothelial cells of control mice. On the other hands, in aspirin group mice, thrombomodulin expression was increased in liver, kidney and lung vessels compared with control mice.
Conclusion: These results suggest that low dosage aspirin therapy is a highly effective treatment strategy for endothelial injury in sepsis.
INFLUENCE OF AUTOPHAGY ON ACUTE KIDNEY INJURY IN MURINE SEPSIS MODEL Satoshi Sunahara1, Eizo Watanabe1, Masahiko Hatano2, Takehiko Oami1, Lisa Fujimura2, Yoichi Teratake2, Shigeto Oda1. 1Department of Emergency and Critical Care Medicine, Chiba University Graduate School of Medicine, Chiba, Japan, 2Department of Biomedical Science, Chiba University Graduate School of Medicine, Chiba, Japan
Objective: To determine the role of autophagy in kidney during sepsis.
Summary Background Data: It is not well understood whether the process of autophagy is accelerated or blocked in the pathophysiology of sepsis, and whether it is beneficial or harmful to the immune defense mechanism over a time course of sepsis.
Methods: We examined the kinetics of autophagy in a murine model of sepsis induced by cecal ligation and punctures (CLP). For biochemical examination of autophagy flux, we performed western blotting analyses on p62 and Rubicon of kidney over time. Sham and CLP mice kidneys were harvested at 6–8 hrs and 24 hrs after operation. We also examined renal function using serum blood urine nitrogen, creatinine, and cystatin C. Furthermore we also investigated the effect of acceleration of autophagy, using rapamycin, on these processes.
Results: As evaluated by western blotting, p62 expression were increased at 24 hrs after CLP compared to that at 6–8 hrs in the kidney (p < 0.05, n = 6). And p62 expression were increased in CLP mouse kidney compared to sham mouse at 24hrs after operation (p < 0.001, n = 6). A similar tendency was recognized in Rubicon. Rubicon protein expression were increased at 24 hrs after CLP compared to that at 6–8 hrs in the kidney (p < 0.05, n = 6). And Rubicon expression were increased in CLP mouse kidney compared to sham mouse at 24hrs after operation (p < 0.05, n = 6). Furthermore, decreased expression of p62 (p < 0.01, n = 4) and Rubicon (p < 0.05, n = 4) in CLP at 24hrs after operation was observed by rapamycin administration, indicating acceleration of autophagy process in the kidney. At the same time, serum cystatin C was decreased (p < 0.05, n = 6) in rapamycin administered mice (at 24 hrs), indicating that rapamycin administration resulted in improvement of renal function.
Conclusions: Autophagy processes were stagnated in the kidney in a murine model of sepsis and acceleration of autophagy mitigated kidney injury. Therefore, autophagy in kidney appears to play a protective role against sepsis.
MICRORNA-125B PRESERVES CARDIADC FUNCTION IN POLYMICROBIAL SEPSIS BY TARGETING TRAF6 AND P53 MEDIATED APOPTOTIC SIGNALING Chuanfu Li, Xiaohui Wang, Tuanzhu Ha, Yuanping Hu, He Ma, John Kalbleisch, Race Kao, David Williams. Department of Surgery, East Tennessee State University, Johnson City, TN, USA
MicroRNAs (miRs) are novel endogenous regulators of gene expression via degradation or translational inhibition of their target mRNAs. MiR-125b is expressed in several organs including lung, heart, spleen, brain and skeletal muscle and it regulates NF-κB activation pathway via a negative feedback mechanism. This study examined the role of miR-125b in polymicrobial sepsis-induced cardiovascular dysfunction. We generated lentivirus expressing miR-125b (LmiR-125b) and transfected them into the myocardium of mice (n = 6). Lentivirus expressing scrambled miR (LmiR-control) served as control (n = 6). Seven days after transfection, mice were subjected to cecal ligation and puncture (CLP). Untreated CLP mice served as sepsis controls. Cardiac function was examined by echocardiography before and 6 hours after CLP. The data show that sepsis-induced cardiac dysfunction was significantly attenuated by LmiR-125b transfection. The values for EF% and %FS in LmiR-125b transfected septic mice were significantly (p < 0.05) higher than in the untreated CLP group. LmiR-125b transfection also markedly improved survival outcome. Immunohistochemistry staining showed that infiltration of macrophages and neutrophils into the myocardium were markedly reduced. The expression of adhesion molecules in the myocardium was also suppressed by LmiR-125b transfection of septic mice. LmiR-125b transfection suppressed TRAF6 expression and NF-κB binding activity and attenuated circulating levels of TNFα and IL-1β in CLP-sepsis. In addition, LmiR-125b transfection attenuated sepsis-induced myocardial apoptosis by suppressing the expression of p53, Bak and Bax in the myocardium. In vitro studies showed that delivery of miR-125b mimics into endothelial cells significantly attenuated endotoxin induced increases in the expression of adhesion molecules and TNFα production. We conclude that miR-125b serves a protective role in sepsis-induced cardiovascular dysfunction by targeting TRAF6 mediated inflammatory responses and p53-mediated apoptotic signaling. Modulation of miR-125b expression may be a useful approach for septic cardiomyopathy.
THE mRNA EXPRESSION OF FATTY ACID AMIDE HYDLASE IN HUMAN WHOLE BLOOD CORRELATES WITH ANANDAMIDE IN PATIENTS WITH SEPSIS Mitsuhide Hamaguchi, Katsuyuki Maruyama, Tomohide Matsushima, Takami Nakao, Takuya Ishibe, Noriko Tsuda, Yoshinori Murao. Critical Care Medical Center, Faculty of Medicine, Kindai University, Osaka, Japan
Introduction: An excessive accumulation of anandamide (N-archidonoylethanolamine, AEA) is associated with septic shock. Results of previous studies have suggested that mRNA coding for the AEA degrading enzyme fatty acid amide hydrolase (FAAH), which converts AEA into arachidonic acid and ethanolamine, might be down-regulated in septic shock.
We used real-time reverse transcription PCR assays to measure relative FAAH mRNA concentrations in the whole blood of 30 healthy donors and eight septic patients to evaluate whether such down-regulation takes place. The levels of the FAAH mRNA were 14.2 ± 7.3 in healthy male and 11.2 ± 3.6 in healthy female (M.Tanaka et al, Journal of Endotoxin Research, 2007).
Method: AEA was measured by liquid chromatography-tandem mass spectrometry. The measurement was done at SHIMADZU TECHNO-RESEARCH, INC. Kyoto. The following parameters, APACHE II, FAAH mRNA and AEA, were measured in septic patients on admission and on day 7.
Results and Discussion: As shown in the table, the levels of APACHE II score were significantly reduced from 19.7 ± 2.2 on admission to 9.5 ± 1.1 at day 7, respectively (P < 0.05). The levels of the FAAH mRNA in patients with sepsis were significantly reduced to 0.21 ± 0.06 on admission (P < 0.01) compared to healthy volunteers. On day 7 after admission, the levels of FAAH mRNA appeared to increase to 0.34 ± 0.06 (P = 0.058) in patients with sepsis. We speculate that AEA increase of immediate mediator to septic shock might contribute to drop manifestation of FAAH mRNA. We speculate that AEA data may influence treatment using continuous hemodiafiltration because of adsorption.
CONNECTION BETWEEN AROMATIC MICROBIAL METABOLITES, HORMONES AND BIOMARKERS IN CRITICALLY ILL PATIENTS WITH INFECTION AND ARTERIAL HYPOTENSION Natalia V. Beloborodova, Ekaterina Chernevskaya, Aleksandra Bedova, Yulia Sarshor, Alexander Sergeev. Negovsky Scientific Research Institute of General Reanimatology, Moscow, Russia
The excessive production of bacterial exometabolites and their accumulation in blood are observed as response to changes in microbial environment in the body of critically ill patients. Previously it was found that high levels of some aromatic microbial metabolites (AMM) are associated with mortality of critically ill patients (1, 2, 3). The development of arterial hypotension in ICU patients with infection leads to a high mortality rate.
Objective: to search the relationship between endogenous hormones, biomarkers and bacterial exometabolites potentially related to hypotension in critically ill patients with infection.
Methods: in prospective study were observed 64 critically ill patients with different diseases at the day of admission to mixed ICU. Patients with hemorrhagic shock, acute and chronic cardiac pathology were excluded. So, 41 patients with infection (27 - severe pneumonia, 14 - peritonitis), 18–80 aged were included. The level of AMM: phenyllactic (PhLA), p-hydroxyphenyllactic (HPhLA), p-hydroxyphenylacetic (HPhAA) acids and sum of three acids (3PhCAs) were measured in serum using gas chromatography (GC-FID). ACTH, TSH, cortisol, PCT, S100b, NT-proBNP were measured using Elecsys-2010. Comparison between two groups of patient - with arterial hypotension (21 pts.) and without (20 pts.) was made. Arterial hypotension (AH) was defined as systolic blood pressure less than 90 mm Hg. Both groups of patients had similar diagnosis and age. Data were analyzed by Mann-Whitney U-test, Spearman correlation coefficient p < 0.05 was considered significant (Statistic 10).
Results: the levels of central regulation hormones (ACTH, TSH) had no difference in the both groups (AH and normal pressure). However, a higher level of S100b in the AH group indicates indirectly the involvement of the brain. The most significant differences between the groups were identified in the levels of AMM. Also higher levels of cortisol and PCT in the AH group were revealed (table). A direct Spearman correlation between PhLA and NT-proBNP was 0.409, p < 0.01; between PhLA, p-HPhLA, 3PhCAs and cortisol 0.464, 0.569, 0.432 respectively, p < 0.01; 3PhCAs and S100b 0.411, p < 0.05. These correlations were more expressive in the group of patients with AH.
Conclusion: the study results indicate potential involving of aromatic bacterial exometabolites in the pathogenesis of arterial hypotension in critically ill patients with infection.
Work was supported by the Russian Science Foundation Grant 15-15-00110.
1. Beloborodova N.V.et al. Shock 2015; 44 (Suppl. 2) 13
2. Beloborodova N.V.et al. Critical Care 2016, 20 (Suppl 1): P3.
3. Rogers A.J. et al. PLoS ONE 2014; 9(1): e87538
THE ROLE OF ASTROCYTES IN THE NERVOUS SYSTEM DYSFUNCTION, SEPSIS INDUCED MEMORY IMPAIRMENT Marina Yamada1,2, Tomohiro Chiba3, Akihisa Matsuda4, Sadakazu Aiso2, Tomohiko Masuno1, Hisashi Mastumoto1, Hiroyuki Yokota1, Masao Miyashita4. 1Department of Emergency and Critical Care Medicine, Nippon Medical School, Chiba, Japan, 2Department of Anatomy, Keio University School of Medicine, Tokyo, Japan, 3Department of Pathology, Kyorin University School of Medicine, Tokyo, Japan, 4Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan
Background: Post intensive care syndrome (PICS) covers both physical and psychiatric dysfunction among patients recovered from intensive care units such as sepsis patients. Chronic presentation of cognitive impairment is becoming one of the most important problems in PICS along with the marked improvement of physical treatment. To overcome PICS-related cognitive dysfunction, we developed and analyzed a murine PICS cognitive impairment model.
Methods: Male ICR mice at the age of 8 weeks were subjected to cecal ligation and puncture (CLP). After CLP, mice were evaluated in behavioral tests (Y-maze), and sacrificed to analyze histological and biochemical changes of inflammatory parameters including cytokines (TNFα, IL-6), inflammatory cells and β-amyloid. Murine primary cultured astrocytes were treated with β-amyloid and LPS. Cultured media at 24 hour after treatment were analyzed biochemically.
Results: CLP-induced sepsis caused chronic impairment of working memory in Y-maze. In consistence, we observed significant increase in inflammatory cells such as astrocytes and microglia together with increase in β-amyloid levels in the cerebral cortex and hippocampus.
Biochemical analyses further revealed that simultaneous treatment of primary astrocytes with β-amyloid and LPS significantly increased secretion of TNFα and IL-6 expression as compared with those treated with either β-amyloid or LPS alone.
Conclusion: We here developed a murine model of PICS-related cognitive impairment. Our data suggest that chronic increase in inflammatory response and β-amyloid is likely to cause chronic cognitive impairment in PICS.
GENDER-DEPENDENT DIFFERENCES OF LUNG INJURY IN LIVER-SPECIFIC AMPKα1 KNOCK OUT MICE AFTER SEPSIS Satoshi Kikuchi1,2, Vivian Wolfe1, Paul W. Hake1, Giovanna Piraino1, Michael O’Connor1, Basilia Zingarelli1. 1Cretical Care Medicine, Cincinnati Children's Hospital, Cincinnati, OH, USA, 2Ehime University Graduate School of Medicine, Ehime, Japan
Alterations in energy homeostasis contribute to sepsis-mediated multiple organ failure. Several studies have suggested female are less susceptible to the adverse outcome of sepsis. The liver is critical to a number of key metabolic functions and participates to the immune and inflammatory responses of sepsis. AMP-activated protein kinase (AMPK) is an important regulator of signaling metabolic pathways. We hypothesized that genetic deficiency of AMPKα1 in the liver would worsen lung injury in male and female mice. Furthermore, we investigated whether the AMPK activator A-769662 would attenuate sepsis-induced inflammation.
Objective: To determine the role of hepatocyte AMPK we developed a specific AMPKα1 hepatocyte-knockout (Cre+/+/AMPKα1−/−) mouse model employing Cre expression under the control of the albumin promoter (Cre-Alb).
Methods: Sepsis was induced by cecal ligation and puncture (CLP) in anesthetized male and female mice (2–3 months old). Mice were randomly divided into Cre-Alb control, Cre+/+/AMPKα1−/− control, Cre-Alb CLP, Cre+/+/AMPKα1−/− CLP, Cre-Alb CLP with A-769662, and Cre+/+/AMPKα1−/− CLP with A-769662 groups. For the treatment groups, A-769662 (10 mg/kg) was administrated intraperitoneally one hour after CLP. Animals were sacrificed 18 hours after CLP. Lung and liver were collected for Western blot and myeloperoxidase assay (MPO, U/100 mg tissue), as an index of neutrophil infiltration.
Results: At 18 hours after CLP, Cre+/+/AMPKα1−/− male mice exhibited higher MPO when compared with Cre-Alb male mice in the lung (336.9 ± 74.3 vs 168.7 ± 19.5, p = 0.003). A-769662 attenuated lung MPO in the Cre-Alb groups (122.1 ± 6.72, p = 0.005), but not in the Cre+/+/AMPKα1−/− male groups (279.6 ± 63.77 p = 0.205). Interestingly, in the female group liver-specific genetic absence of AMPKα1 afforded protective effects in the lung neutrophil infiltration since Cre+/+/AMPKα1−/− mice had lower MPO than Cre-alb mice (57.2 ± 11.2, 125.8 ± 20.77, p = 0.001). Furthermore, in the Cre+/+/AMPKα1−/− female group A-769662 worsened MPO (115.0 ± 21.68, p = 0.003). Western blot analysis showed expression of pAMPKα1 in the lung was increased in both genders Cre-Alb CLP groups after CLP when compared with Cre-Alb control groups. In male Cre+/+/AMPKα1−/− expression of pAMPKα1 decreased in the lung after CLP when compared with Cre+/+/AMPKα1−/− control groups. However, no significant changes were observed in lung pAMPKα1 expression in female Cre+/+/AMPKα1−/− after CLP when compared to control group.
Conclusion: Our study shows AMPKα1-dependent liver metabolic functions are important in modulating inflammation in distant organs, such as the lung. However, the protective effects of AMPKα1 are affected by gender.
IL-15 MEDIATED EFFECTS OF FIBROCYTE TRANSFER IN SEPSIS Jean Nemzek-Hamlin, Christopher Fry. Unit for Laboratory Animal Medicine, University of Michigan, Ann Arbor, MI, USA
The adoptive transfer of fibrocytes improves survival after cecal ligation and puncture (CLP) in mice. In vitro studies have suggested that the improved survival may be due to direct interaction of fibrocytes with T cells mediated through transpresentation of IL-15. To determine the role of IL-15 in fibrocyte interactions in vivo, mice were given fibrocytes derived from either wildtype or IL-15 knockout mice intraperitoneally, at the time of CLP. The mice were observed for 10days. The control group treated with saline had a 10% survival while the group treated with wildtype fibrocytes had a significantly increased survival of 67%. The group treated with fibrocytes derived from IL-15 knockout mice had a 40% survival. This suggests that IL-15 expression by transferred fibrocytes plays a role in the survival advantage conferred by fibrocyte transfer but may not be the exclusive mechanism. In further studies, mice were given either saline or wildtype fibrocytes immediately after CLP. The mice were euthanized at 6 and 24 hours to obtain plasma. At 24 hours, plasma levels of IL-15 were elevated in the mice treated with fibrocytes compared to those given saline. To examine whether fibrocyte interactions could directly contribute to soluble IL-15 concentrations, splenic T cells were incubated under three conditions: monoculture, co-culture with fibrocytes or culture with fibrocytes in a transwell system. At various time points up to one week, media was collected for ELISAs. The IL-15 levels in the fibrocyte/T cell co-cultures were significantly increased by 24 hours as compared to either the monoculture or transwell system. Likewise, the levels of IL-21 were significantly increased by fibrocyte transfer in septic mice and were significantly increased by direct contact between fibrocytes and T cells in vitro. These findings and those from previous studies suggest that the direct contact of fibrocytes and T cells is not only mediated by gamma c cytokines but results in the further production of gamma c cytokines. This may further explain the increased proliferation of both CD4 and CD8 T cells after transfer of fibrocytes in septic mice. However, other mechanisms may also play a role in the dramatic survival benefits seen after fibrocyte transfer in sepsis.
S14G-HUMANIN IMPROVES THE SURVIVAL FROM SEVERE SEPSIS Kumiko Sekiguchi1, Marina Yamada2, Tomohiro Chiba3, Akihisa Matsuda1, Sadakazu Aiso4, Tomohiko Masuno2, Hisashi Matsumoto2, Hiroyuki Yokota2, Masao Miyashita1. 1Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 2Department of Emergency and Critical Care Medicine, Nippon Medical School, Tokyo, Japan, 3Department of Pathology, Kyorin University School of Medicine, Tokyo, Japan, 4Department of Anatomy, Keio University School of Medicine, Tokyo, Japan
Background: Mortality from severe sepsis remains to be high in spite of the recent improvement of intensive care. It is partially caused by dysregulation of the immune system. Humanin (HN), a short neuroprotective peptide for Alzheimer's disease-related neurotoxicity, is also reported to regulate apoptosis, insulin sensitivity and inflammatory response. We here examined the effect of a potent HN analogue S14G-HN (HNG) on survival and inflammatory response in mice severe sepsis model.
Methods: Male ICR mice at the age of 8 weeks were intraperitoneally (i.p.) administrated with saline (vehicle) or HNG (0.75 mg/kg/day for 4 days) after cecal ligation puncture (CLP). The effects of HNG were evaluated by survival study and inflammatory response assessed by serum TNF-alpha and IL-6 levels at 24 h after CLP.
Results: HNG treatment significantly improved survival of CLP mice as compared with vehicle-treated mice (35.2% vs. 56.0%, respectively; p < 0.05). The exaggerated inflammatory response induced by CLP is significantly ameliorated with HNG treatment.
Conclusion: We, here, report the beneficial effects of HNG on severe sepsis. HNG could have a novel therapeutic potential for sepsis via modulation of exaggerated inflammatory response. However, the detailed mechanisms and the effect on sepsis-related psychiatric dysfunction should be addressed in the future experiments.
DIAGNOSTIC POTENTIAL OF NOVEL ENDOTOXIN DETECTION METHOD FOR SEPSIS AND SEPTIC SHOCK Tomoharu Shimizu1, Toru Obata1, Hiromichi Sonoda1, Tohru Miyake1, Hiroya Akabori1, Takahisa Tabata2, Yutaka Eguchi2, Yoshihiro Endo3, Tohru Tani4, Masaji Tani1. 1Department of Surgery, Shiga University of Medical Science, Shiga, Japan, 2Department of Emergency and Critical Care Medicine, Shiga University of Medical Science, Shiga, Japan, 3Department of Clinical Nursing, Shiga University of Medical Science, Shiga, Japan, 4Biomedical Innovation Center, Shiga University of Medical Science, Shiga, Japan
Introduction: The turbidimetric limulus amoebocyte lysate (LAL) assay is usually used for the detection of endotoxin; however, there are often situations where plasma endotoxin levels cannot be detected in plasma samples obtained from patients suspected of having septic shock due to Gram-negative bacterial infections. A novel rapid LAL assay for endotoxin has recently been developed that uses a laser light-scattering particle-counting method, called Endotoxin Scattering Photometry (ESP).
Methods: We compared ESP, standard turbidimetric LAL assay, and procalcitonin assay for the evaluation of sepsis after emergency gastrointestinal surgery. A total of 174 samples were collected from 40 adult patients undergoing emergency gastrointestinal surgery and 10 patients with colorectal cancer undergoing elective surgery as non-septic controls.
Results: Plasma endotoxin and procalcitonin levels increased corresponding to the degree of sepsis. ESP significantly discriminated between patients with or without septic shock: sensitivity, 81.1%; specificity, 76.6%; positive predictive value, 48.4%; negative predictive value, 93.8%; and accuracy, 77.6%. The area under the receiver operating characteristic curve (AUROC) for septic shock with the ESP assay (endotoxin cut-off value, 23.8 pg/mL) was 0.8532 (95% CI, 0.7841 to 0.9030; P < 0.0001). The predictive power of ESP was superior to that of turbidimetric assay (difference, 0.1965; 95% CI, 0.0812 to 0.3117; P = 0.0008). There was no significant difference in predictive power between ESP and procalcitonin assay. ESP also discriminated between patients with and without sepsis. AUROC analysis showed that ESP had the best predictive power for diagnosing sepsis.
Conclusion: ESP appeared to more sensitively detect plasma endotoxin and significantly discriminated between sepsis and septic shock in patients undergoing gastrointestinal emergency surgery compared with turbidimetric LAL assay.
SEPTIC SHOCK CAUSED BY SELF-INJECTION OF METHYLPHENIDATE HYDROCHLORIDE (RITALIN®) Noriko Saito, Makiko Saito, Ahmin Kim, Azusa Ichimaru, Rui Komatsu, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
A 52-year-old man visited the emergency department of our hospital with chief complains of high fever accompanied by chills and general fatigue persisting from the previous night. On arrival, his consciousness was restlessness, body temperature was 39°C, heart rate was 122 beats/min, blood pressure was 88/59 mmHg, and SpO2 was 98%. Indurations of approximately 2 cm with swelling were present on both forearms and the back of the right hand but were not accompanied by burning sensations or erythema. Laboratory findings were as follows: WBC, 12,950/μL,; CRP, 5.12 mg/dL; procalcitonin, 62.66 ng/mL; base excess, −5.6 mmol/L; and lactate, 3.7 mmol/L. Contrast-enhanced computed tomography revealed emphysema, swelling of the pancreatic head, and slight dilation of the pancreatic duct without any evidence of infection. He was admitted to the intensive care unit with a diagnosis of septic shock from unknown cause. Noradrenaline (1.2γ) was required to maintain his blood pressure. He was administered Tazobactam/Piperacillin and intravenous immunoglobulin. His fever decreased the next day, and laboratory values indicating inflammation were reduced. The patient admitted that he had injected himself with methylphenidate hydrochloride (Ritalin®) dissolved in tap water 2 days before admission. Vancomycin was administered on the third day because gram-positive cocci were detected in blood cultures initiated upon admission. Gallium scintigraphy detected no sites of inflammation. His clinical course was benign, and he discharged himself without the hospital's consent on the seventh day. Staphylococcus epidermidis was identified in blood cultures, and ritalinic acid was detected in his urine and serum using GC/MS and LC/MS/MS. A higher morbidity of sepsis is associated with intravenous drug users, because needles and drugs are seldom sterilized. The patient was subsequently admitted to other hospitals as he contracted sepsis due to the self-injection of Ritalin dissolved in tap water. It is difficult to determine the cause of sepsis in intravenous drug users; however, the incidence of sepsis caused by bloodstream infections may be increasing among these people.
LACTOFERRIN PREVENTS INTESTINAL EPITHELIAL CELL DAMAGE INDUCED BY CLOSTRIDIUM DIFFICILE TOXIN B Kosuke Otake1,2, Norio Sato2, Ayako Kitaguchi2, Takayuki Irahara1,2, Satoru Murata2, Yutaka Harima2, Keiji Nakata3, Shigeru Ohtsuru2, Kaoru Koike2, Hiroyuki Yokota1. 1Department of Critical Care and Emergency Medicine, Nippon Medical School, Tokyo, Japan, 2Department of Primary Care and Emergency Medicine, Kyoto University, Kyoto, Japan, 3Department of Modern Sociology, Kobe Gakuin University, Hyogo, Japan
Background: Clostridium difficile infections (CDI) have increased around the world and are known to cause Clostridium difficile-associated diarrhea. Some CDI progress to fulminant and recurrent CDI and are related to high mortality and morbidity. CD mainly produces toxin A and toxin B, which cause mucosal damage. Toxin B is well known to have more cytotoxicity than toxin A. Lactoferrin (LF) is contained in whey protein and it has been known to have multifunctional nutrients including anti-inflammatory functions. However, there are no many studies about LF against CDI. The purpose of this study is to investigate the effect of LF in IEC-6 (rat intestinal epithelial cell) under toxin B stimulation.
Methods: Experiment 1: IEC-6 cells were cultured to confluence then different concentrations of toxin B (from 0 to 1.0 ng/mL) were added with LF (1.0 mg/mL) or pepsin treated lactoferrin (PLF, 1.0 mg/mL). Cytotoxicity was measured in medium by the level of LDH release after 24 hours and mitochondrial function of viable cells was measured by the level of WST-1 after 24 hours. Experiment 2: Wound restitution was measured by a microscope to investigate the effect of LF or PLF under the toxin B stimulation (0.125 ng/mL).
Results 1. LDH was significantly increased dose dependently when different concentrations of toxin B were added. After LF or PLF were added with toxin B, LDH was significantly decreased compared to when toxin B alone was added. On the other hand, WST-1 was significantly decreased dose dependently when different concentrations of toxin B were added. After LF or PLF were added with toxin B, WST-1 showed higher levels of activity compared to control. However, WST-1 in LF group was higher than in PLF group.
Results 2. Wound restitution with toxin B was 0 ± 0.0 μm, which was significantly lower than without toxin B 28.1 ± 3.5 μm. Furthermore, wound restitution addition of LF with toxin B was 20.6 ± 2.5 μm and PLF with toxin B was 17.4 ± 2.6 μm, which were significantly higher than that of the toxin B only group (P < 0.0001).
Conclusions: We demonstrated that LF prevents cytotoxicity of toxin B and this might have a strong potential to control CDI.
CAR PEPTIDE ENHANCES HYDROCORTISONE TO MORE EFFECTIVELY TREAT INJURED ENDOTHELIUM IN SEPSIS Kodai Suzuki1, Hideshi Okada1, Chihiro Takada1, Kazumasa Oda1, Takahiro Yoshida1, Shiho Nakano1, David Mann2, Genzou Takemura3, Masanobu Komatsu4, Shinji Ogura1. 1Department of Emergency and Disaster Medicine, Gifu University Graduate School of Medicine, Gifu, Japan, 2VBS Pharmaceuticals Division, Vascular BioSciences, Goleta, CA, USA, 3Department of Internal Medicine, Asahi University of Dentistry, Gifu, Japan, 4Sanford-Burnham Medical Research Institute at Lake Nona, Orlando, FL, USA
Introduction: Inflammatory cytokines and lipopolysaccharides (LPS) injure the endothelium of multiple organs in sepsis. While some studies have demonstrated that hydrocortisone (HC) has antiinflammatory effects in sepsis, others have concluded that high dose steroids may be potentially harmful and associated with higher mortality, secondary infections, renal and hepatic failure. A novel tissue-penetrating cyclic peptide, CARSKNKDC (CAR), has previously demonstrated the unique ability to facilitate delivery of co-administered drugs to the injured endothelium.
Hypothesis: Co-administration of HC with CAR could prevent endothelial injury and improve sepsis survival.
Methods & Results: Ten-week-old C57BL6 male mice were given LPS (20 mg/kg). We first examined whether CAR homes to septic-injured endothelium. FAM-CAR was injected in the mice 12 hours after LPS admin, and were sacrificed 1 hour later. FAM immunostaining showed that CAR peptide specifically homed to sepsis-injured endothelial cells but not to healthy endothelium. We determined whether CAR bound with HC directly using Isothermal Titration Calorimetry and Biacore analysis. Both assays showed that CAR peptide did not bind to HC. Subsequently, five different in-vivo treatments were initiated: untreated saline only group (Group S, n = 31); CAR peptide-alone administration 500ug/mouse (Group C, n = 20); HC 0.2 mg/kg or 10 mg/kg (Group Low-H or High-H, n = 20, 28 respectively); and CAR co-administered with HC 0.2 mg/kg (Group C+H, n = 20). HC and/or CAR peptide were injected intraperitoneally at 3, 12 and 24 hours after LPS administration. Forty-eight hours after LPS administration, Group C+H showed the best survival ratio compared with all Groups, Group C +H vs Group S (90 vs. 21%, p < 0.05), while Group High-H showed a better survival ratio than Group S (57 vs. 21%, p < 0.05). There were no significant survival difference between Groups C, Low-H and S. Scanning electron microscopy showed that the destruction of kidney, lung and liver capillaries, with edematous endothelial walls and fibrin deposition were observed after LPS injection in Group S. In Group C+H, however, the destruction of kidney, lung and liver capillaries was significantly attenuated compared with Group S.
Conclusion: These results suggest that drug delivery therapy with CAR peptide enhances the effectiveness of low-dose HC to treat injured endothelium and significantly improve survival in sepsis.
EVALUATION WITH PROVISIONAL DIAGNOSTIC CRITERIA OF THE JAPANESE SOCIETY ON THROMBOSIS AND HEMOSTASIS IN PATIENTS WITH SEPSIS-INDUCED DISSEMINATED INTRAVASCULAR COAGULATION Atsushi Murao, Tomoki Tanaka, Kengo Hashimoto, Takayoshi Mizuno, Emi Fujii, Hidemitsu Miyatake, Junji Shimizu, Tetsunobu Yamane, Takahisa Tabata, Yasuyuki Tsujita, Yutaka Eguchi. Department of Critical and Intensive Care Medicine, Shiga University of Medical Science, Shiga, Japan
Disseminated intravascular coagulation (DIC) is a lethal disease, especially in patients with sepsis/septic shock. Early treatment based on an appropriate diagnosis is considered to be very important for improving patients’ prognosis. Several criteria have been proposed, diagnostic criteria includes the Japanese Ministry of Health and Welfare's old DIC diagnostic criteria (JMHW criteria), and the Japanese Association for Acute Medicine's acute-stage DIC diagnostic criteria (JAAM criteria). The JMHW criteria have poor sensitivity, especially in the case of infection, and JAAM criteria has been sensitive but not specific. Provisional diagnostic criteria of the Japanese Society on Thrombosis and Hemostasis (JSTH) include antithrombin (AT) activity and thrombosis related marker. These three different diagnostic criteria were investigated in 30 patients with sepsis/septic shock-induced DIC diagnosed by JAAM criteria from July 2013 to June 2015. Out of 30 patients, 19 patients (63%) were diagnosed both by JMHLW and JSTH criteria. Antithrombin (AT) concentrate and AT concentrate plus recombinant thrombomodulin (rTM) were administrated in 27 and 3 patients, respectively. The over-all 28-day and in-hospital mortality were 13% and 17%, respectively, and the former was 15% and 19%, and the later was 0% and 0%, respectively. DIC recovery rates after administration of AT-concentrate were 33%, 42% and 63% diagnosed by JAAM, JMHW and JSTH criteria and the 28-day mortality were 17%, 16% and 16%, respectively. In conclusion, the JSTH criteria were less sensitive but most specific compared with JAAM and JMHLW criteria. Further investigation is necessary to assess these diagnostic criteria.
DEVELOPMENT OF A NEXT-GENERATION SEQUENCING-BASED METHOD TO DETECT BACTERIAL DNA IN HUMAN SERUM FROM SEPSIS AND PNEUMONIA PATIENTS Nobuo Watanabe, Meiko Takeshita, Junko Takeuchi, Yukiko Kirimura, So Nakagawa, Kirill Kryukov, Miho Sera, Satomi Mitsuhashi, Sadaki Inokuchi, Tadashi Imanishi, Shigeaki Inoue. Deptartment of Emergency and Critical Care Medicine, Tokai University School of Medicine, Kanagawa, Japan
Introduction: Prompt identification of causative pathogenic bacteria is important for the treatment of patients suffering from infectious diseases including sepsis and pneumonia. However, current cultivation-based identification methodologies have the following drawbacks: i) the culture period for some bacteria is up to 1 week, ii) anaerobic bacteria are difficult to culture, iii) poor colony formation from antibiotics-administered blood. To circumvent these problems, we aimed to detect bacterial DNA sequences in patient specimens using next-generation DNA sequencing (NGS). In this presentation, we will present our approach to detecting bacterial DNA in blood by using the Ion PGM NGS system (Life Technologies).
Methods: A pool of known bacteria was mixed with blood from healthy volunteers, and total DNA was purified from the blood using a DNA isolation column (Qiagen). In an attempt to enrich the bacterial DNA, the bacteria-tainted blood was fractionated into plasma and human cells before DNA extraction using different centrifugation steps. The DNA samples were sequenced on an Ion PGM NGS system according to the following two protocols. For direct sequencing, a DNA library was prepared using a library preparation kit (Thermo Fisher) after shredding of the total DNA into ∼400-bp fragments. For amplicon sequencing, several regions of 16S rRNA-DNA in the total DNA samples were amplified by PCR, and the sequencing library was prepared using the Ion 16S™ Metagenomics Kit (Thermo Fisher).
Results: Using a stepwise centrifugation procedure, in which blood cells were first removed by low-speed centrifugation and bacterial cells in the remaining plasma were sedimented in a subsequent high-speed centrifugation, we successfully enriched the bacterial DNA by 10-fold, with low human DNA contamination. In the metagenomic analysis of whole blood samples, no increase in the number of human DNA sequences (reads) owing to bacteria supplementation was observed due to the presence of a significant amount of human DNA in the sample (blood vs. blood + bacteria = 300,000 vs. 300,000). On the other hand, in the 16S amplicon analysis, the number of bacterial reads was reasonably increased by the addition of bacteria into the blood (blood vs. blood + bacteria = 3,000 vs. 500,000). Detailed data analyses for both NGS runs are in progress, and the results will be shown at this meeting.
Conclusions: The present NGS-based method could be a useful new tool for the determination of bacterial infections and - will provide great benefit to early diagnosis of sepsis and pneumonia.
SPORADIC NEUROLOGIC MELIOIDOSIS PRESENTING AS PRIMARY MENINGITIS IN A 65 YEAR OLD DIABETIC MALE: A CASE REPORT Jan Patrick O. Ng, Lena C. Amsua, Generoso Licup. Department of Internal Medicine, West Visayas State University Medical Center, Iloilo City, Philippines
Clinical Presentation: The diabetic patient was confined due to vomiting and body malaise where he eventually developed stiffening of extremities. Seizure recurrence prompted referral. The patient complained of a localized (L) temporal throbbing headache.
Physical Findings: Unremarkable physical and neurologic examination.
Laboratory Work-up: CBC showed leukocytosis at 17.94 x109/L with absolute monocytosis at 0.10. There was mild hyponatremia. Blood gas showed uncompensated respiratory alkalosis with adequate oxygenation. Cranial CT scan was normal. Lumbar puncture showed xanthochromia and WBC count of 126 cells/cummwith neutrophils at 0.51. CSF culture revealed Burkholderia pseudomallei, CALAS and PCR for tuberculosis were negative.
Significance: Melioidosis is a potentially fatal disease with a mortality rate of 14 to 50%. A 20-year Darwin prospective study reported 540 cases, only 3% (14) presented with neurologic symptoms: 10 meningo-encephalitis, 2 myelitis and 2 cerebral abscesses. In 2012, Singapore reports 5 cases of neurologic melioidosis, however all patients presented with cerebral abscesses, profound neurologic deficits and low Glasgow Coma Scale scores. In this case, the patient presented with seizure, unilateral headache, unremarkable physical and neurologic findings and normal CT scan.
Diagnosis: Multi-organ Failure 2′ to Septic Shock 2′ to Hospital Acquired Pneumonia, Late onset on top of Neurologic Melioidosis (Primary Bacterial Meningitis); Generalized Tonic Clonic Seizure 2′; Diabetes Mellitus.
Treatment: Leviteracetam and PRN benzodiazepine were started. Remittent fever and nuchal rigidity with note of decrease in sensorium narrowed Bacterial vs. TB meningitis; Ceftriaxone and HRZE therapy initiated. Sensitivity-guided antimicrobial shifting to Meropenem was done.
Outcome: Patient succumbed.
Recommendations: When considering the etiology of undifferentiated neurologic disease, an awareness of the possibility of neurologic melioidosis is important in disease-endemic regions.
A CASE OF SEPTIC SHOCK CAUSED BY PNEUMOCOCCAL PNEUMONIA WITH CONCOMITANT PRESENTATION OF LUNG ABSCESS Ayumi Kawasaki, Ahmin Kim, Kazuma Shibahara, Reon Nagai, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University School of Medicine, Tokyo, Japan
It is considered to be rare for pneumococcus to be the causative bacteria of pulmonary suppuration in adults. We herein report the case of a diabetic patient who had septic shock as a result of pneumococcal pneumonia combined with pulmonary suppuration. A 67-year-old woman with type ll diabetes presented at the emergency department of our hospital with a chief complaint of coughing was accompanied by right chest pain. She had a history of tuberculosis in her 20 s and hypertension and lipid metabolism disorder after surgery for left-side breast cancer. On arrival at the hospital, the patient was alert with body temperature of 36.4°C, pulse of 107 beats/min, blood pressure of 74/40 mmHg, and SpO2 of 94% (room air). A blood test revealed WBC of 19.04 × 103/μl, CRP of 24.13 mg/dl, and pro-CT of 0.20, and plain X-ray image and CT showed an infiltrative shadow in the right middle lung field. The patient was admitted to the ICU and subsequently started ABPC/SBT 1.5 g x 2/day, ATlll formulation 3000 U/day, and globulin formulation 5 g/day. On Day 2, the patient was changed to PIPC/TAZ due to a concomitant onset of cholecystitis, and also started VCM since gram-positive cocci were detected from blood culture. Although rapid pneumococcal test was negative, Streptococcus pneumoniae was detected with sputum and blood cultures. A fever of more than 38°C persisted thereafter, and CT that was performed on Day 10 to determine the source of fever showed abscess in the right lung. Chest drainage was performed on Day 12. Pleural adhesion was severe and muscle flap was performed to treat lung decortication and fistula on Day 14 at the chest surgery department. Subsequently, inflammatory response showed a decreasing trend, and, in consideration for long-term positive pressure management, tracheotomy was performed on Day 23 and chest drainage was removed on Day 24. On Day 26, the patient was weaned from the ventilator and continued feeding training and physical rehabilitation. Decannulation was performed on Day 67, and the patient was able to walk independently and was discharged to go home on Day 76. Anaerobic bacteria such as Peptostreptococcus spp are the primary causative bacteria of pulmonary suppuration, and Staphylococcus aureus are most frequently isolated aerobic bacteria in this condition. There are very few reports of pulmonary suppuration caused by pneumococcus in adults in Japan. Compromised hosts, exemplified by diabetic patients, have a high risk of developing pulmonary suppuration, and it is therefore necessary to consider the concomitant onset of pulmonary suppuration in patients who develop pneumococcal pneumonia with persisting fever and inflammatory response.
A FATAL CASE OF SEPTIC SHOCK DUE TO UNKNOWN INFECTION SITES WITH PSEUDOMONAS AERUGINOSA Noboru Akizuki, Kentaro Mochiduki, Reon Nagai, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Case: A 51-year-old Japanese woman with a history of depression, hypothyroidism, and migraine was prescribed antibiotics by her family doctor for coughing and chills two days before presenting at our hospital. The next day she had bloody sputum followed by worsening respiratory distress during the night so she called for an ambulance. Vital signs when the emergency services arrived were as follows: level of consciousness, Glasgow Coma Scale (GCS) 15; body temperature, 36.9°C pulse, 120/min, blood pressure, 60/40, respiratory rate, 48 breaths/min: and SpO2, 84%. She was transported to her previous doctor, examined, and diagnosed with respiratory failure due to severe pneumonia and transferred to our critical care center. Vital signs on admission were as follows: level of consciousness, GCS 15; body temperature, 36.4°C pulse 128/min; blood pressure, 89/54; and respiratory rate, 41 breaths/min. Her face was pale and stridor could be heard. Breathing with the accessory muscles of respiration was observed. An unhealed shallow cut caused by self-injurious behavior was present on the left forearm. Testing on admission revealed the following blood counts: leukocytes, 1320/μl platelets, 94000/μl C-reactive protein, 19.89 mg/dl and procalcitonin levels; 38.01 ng/ml, together with coagulopathy (prothrombin time, 18.3 s; international normalized ratio, 1.55; and activated partial thromboplastin time, >150 s) and acute kidney injury (blood urea nitrogen, 26.9 mg/dl and creatinine, 1.45 mg/dl). Arterial blood gas analysis results were pH 7.238, pO2 92.4, pCO2 29.4, HCO3 12.3, BE-13.9, and lactate 7.0 mmol/l indicating metabolic acidemia. Sequential organ failure assessment score on admission was 8. She was diagnosed with septic shock due to pneumonia, disseminated intravascular coagulation (DIC), and multiple organ failure. Non-invasive positive-pressure ventilation was started to treat respiratory failure but her respiratory status continued to worsen and she was intubated and put on artificial ventilation. Antimicrobial, γ-globulin, and noradrenaline treatment was started for the pneumonia and septic shock. Respiratory status did not improve even with FiO2 at 1.0 (PaO2/FiO2 ratio, <100); therefore, extracorporeal membrane oxygenation (ECMO) was started. Fresh frozen plasma, platelet concentrate, and red blood cells were administered for subsequent hemorrhage; however, there was no improvement and bleeding diathesis continued followed and she died. Pseudomonas aeruginosa was later detected in two sets of blood culture.
Conclusion: The present case took a rapidly fatal course. Veno-arterial ECMO should be considered for cardiopulmonary resuscitation.
A CASE OF TOXIC EPIDERMAL NECROSIS (TEN) DUE TO DIURETICS Mayu Kikuchi, Reon Nagai, Miri Kang, Ayumi Kawasaki, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Although the annual incidence of toxic epidermal necrosis (TEN) is quite low at 0.4–1.2 per 1 million individuals, it has a high mortality rate of 17%-25% annually in Japan. Several studies have revealed that antibiotics, antipyretic analgesics, and anticonvulsants cause TEN. Here we report a case of TEN due to diuretics that was treated with multidisciplinary therapy in our intensive care unit (ICU).
A 55-year old woman regularly visited the outpatient clinic for chemotherapy after surgery for breast cancer with metastasis to the liver, bones, lungs, and lymph nodes. During a visit, it was observed that pleural effusion and edema of the lower extremities had developed. She was started on 10 mg/day furosemide and 25 mg/day spironolactone. After 2 days, blister vesicles and erosion formed in her mouth, on both of the lower extremities, on the breech, and on the genital area. Because TEN was diagnosed by skin biopsy.she was admitted to another hospital, and After 7 days, she transferred to our ICU because she required mechanical ventilation and catecholamines On arrival, her level of consciousness was Richmond Agitation-Sedation Scale 0, body temperature was 36.3°C, heart rate was 88/min, and blood pressure was 108/64 mmHg. Blister vesicles and erosion had developed across her face and whole body. Laboratory findings revealed the following findings: WBC count, 30,000 μ/L; Hb level, 10.1 g/dL; platelet count, 11.0 X 104 μ/L; CRP level, 4.41 mg/dL; procalcitonin level, 0.55 ng/mL; BUN level, 107.5 mg/dL; creatinine level, 1.01 mg/dL; Na level, 158 mEq/L; K level, 3.5 mEq/L; Cl level, 121 mEq/L; PT, 13.1 s; APTT, 38.1 s; FDP, 17.2 μg/mL; D-dimer, 15.9 μ/mL; and fibrinogen level, 385 mg/dL. After 10 days, renal replacement therapy was initiated for AKI. TEN improved with IVIG therapy and steroids; however, she developed sepsis due to a catheter-associated blood stream infection, and multiple organ dysfunctions resulted in her death on the 45th day.
TEN is the most critical type of drug eruption with systemic flush and erosion and causes multiple organ dysfunctions. Our case of TEN due to diuretics had a SCORTEN scale score of 4 and a mortality rate of 58%. The patient appeared to recover at one point because of the multidisciplinary therapy. However, the patient had already been suffering from cancer, and complicated sepsis resulted in her death. It is crucial to treat TEN with early multidisciplinary therapy, particularly for a compromised host such as the present case.
DISTAL TO PROXIMAL PROGRESSIVE LEFT HEMIPARESIS IN A 33 YEAR OLD MALE WITH DOUBLE OUTLET RIGHT VENTRICLE: A CASE REPORT Darleen M. Sy, Patricio P. Palmes. Department of Internal Medicine, West Visayas State University Medical Center, Iloilo City, Philippines
Clinical Presentation: A 33 year old male developed distal to proximal weakness of the L upper extremity 6 days prior to admission. Involvement of the L lower extremity prompted admission.
Physical Findings: Grade 3/6holosystolic murmur along the L anterior axillary line and digital clubbing were apparent on physical exam. On Neurologic exam, there was grade 3/5 motor weakness in the Lupper and lower extremities, the rest were intact.
Laboratory Work-up: CBC showed erythrocytosis with hemoglobin of 197 g/L, and RBC count of 6.19 × 1012/L. 2D-echocardiography revealed congenital heart disease, double outlet right ventricle, hypertrophied R ventricle with volume overload, and anterior mitral valve prolapse. Electrocardiogram showed R ventricular hypertrophy. Brain CT and MRI scans showed R parietal rim enhancing mass.
Diagnosis: Intracranial mass, R parietal lobe most probably Inflammatory; Congestive Heart Failure NYHA FC I 2′ to Congenital Heart Disease (Double Outlet Right Ventricle); Hyperviscosity Syndrome 2′ to Secondary Non-decompensated Erythrocytosis.
Treatment: Initially managed as a case of cerebrovascular disease, most probably infarct 2′ to hyperviscosity syndrome. However, an R parietal lobe rim enhancing nodule noted on brain CT scan channeled an inflammatory process, hence IV ceftriaxone, mannitol and dexamethasone were started. Eventually, the patient improved and was advised to complete antibiotics for 6 weeks duration.
Outcome: Patient discharged improved.
Significance: Double-outlet right ventricle is an uncommon congenital cardiac anomaly characterized by the origin of both great arteries from the morphologic right ventricle. It affects 1 – 1.5% of patients with congenital heart diseases with a frequency of 1 in each 10,000 live births.
Recommendations: Close follow up of patients with cardiac problems is essential to prevent unwanted complication.
BRADYCARDIA AND CARDIAC ARREST DURING TRACHEAL SUCTION IN A PATIENT WITH TRAUMATIC SPINAL CORD INJURY Shuji Shimamoto, Noriko Saito, Hideaki Suzuki, Noboru Akiduki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Spinal shock is a common complication in patients with acute spinal cord injury. It involves a transient loss of spinal reflexes for approximately 48 h after the injury. Neurogenic shock is another possible complication, particularly in patients with high complete cervical spinal cord injuries; this usually diminishes within the first 2–6 weeks after the injury, but occurs during the chronic phase. We present the case of a tetraplegic patient who developed cardiac arrest following bradycardia during tracheal suction in the ICU 6 weeks after the injury.
A 72-year-old male fell down the stairs and was transferred to the emergency department with tetraplegia. He was admitted to the intensive care unit with a diagnosis of complete cervical spinal cord injury at C4 and C5. He required mechanical ventilation. A tracheostomy was performed at 14 days, and laminectomy and laminoplasty were performed 24 days after the injury for the dislocation fracture of the cervical vertebrae. Mechanical ventilatory support was still required. Forty-two days after the injury, bradycardia during tracheal suction was accompanied by a decline in SpO2, and cardiac arrest occurred. After 4 min of resuscitation and administration of 1 mg adrenaline, the patient recovered without complications. Although cardiac pacemaker and administration of atropine or aminophylline were considered for preventing further occurrences of bradycardia and cardiac arrest, providing high oxygenation before tracheal suction and shortening the duration of tracheal suction prevented those adverse events. This complication did not occur again in the clinical course of this patient.
There are some reports on cardiopulmonary complications following spinal cord injury regardless of spinal shock. These complications are associated with more severe spinal cord injuries and those at higher levels. In particular, tracheal stimuli such as suction, intubation, and bronchoscopy seem to induce bradycardia and even asystole in the presence of hypoxia due to the vasovagal reflex. In normal conditions, this reflex is controlled by sympathetic activity due to increased breathing; however, this compensatory sympathetic activity is inhibited by cervical cord lesions and does not function in patients with cervical spinal injury. The patient in this case has not had any complication since the cardiac arrest and does not require additional therapy; however, some patients die because of this complication and some require a cardiac pacemaker or maintenance treatment with atropine. Cardiac dysfunctions are usually life-threatening complications which occur during the non-acute phase in patients with severe cervical spinal cord injury.
ADMISSION HEMOGLOBIN LEVELS IN SEVERE TRAUMA PATIENTS WITHOUT PREHOSPITAL FLUID ADMINISTRATION: A SINGLE CENTER STUDY IN JAPAN Yasuyuki Kawai, Hidetada Fukushima, Keisuke Takano, Yuusuke Tada, Hideki Asai, Tomoo Watanabe, Yasuyuki Urisono, Tatsuya Nakamura, Kazuo Okuchi. Department of Emergency and Critical Care Medicine, Nara Medical University, Nara, Japan
Introduction: It has long been considered that blood hemoglobin level or hematocrit remains stable after a bleeding episode due to trauma. Recent studies have reported that compensatory fluid shifts can occur faster than had been previously thought and hematocrit level on arrival at the emergency department is associated with trauma severity and outcome. However, these reports are from western countries where it is almost impossible to eliminate the effect of prehospital intravenous fluid administration. In Japan, emergency medical service crews were not permitted to administer intravenous fluid until Jan. 31st, 2014. The aim of this study is to explore how much blood hemoglobin levels decrease during transportation among severe trauma cases without prehospital intravenous fluid.
Methods: This is a single center observational retrospective study. We included trauma cases with abbreviated injury scores of 3 or above between 2008 and 2014. Exclusion criteria were: less than 18 years-old, cardiac arrest, transferred, and single head trauma cases. We investigated the association of blood hemoglobin levels with time intervals and other physiological variables.
Results: Three hundred forty two cases were included in this study (the median age was 53 and there were 242 male cases). The median time for transportation was 45 minutes. The median hemoglobin level on arrival was 13.1 g/dl and there was no statistical correlation between hemoglobin level and transportation time (R = 0.048, p = 0.37). However, hemoglobin levels were significantly lower in patients over the age of 65, female patients, patients with systolic blood pressure below 90 mmHg and patients who required surgical hemostasis within 24 hours.
Conclusions: This current study focusing on severe trauma cases without prehospital intravenous fluids administration revealed that decreased hemoglobin levels on arrival do not correlate with time intervals, but do correlates with patients’ case profile in other areas such severity of injury.
DAMAGE ASSOCIATED MOLECULAR PATTERNS RELEASING IN A NEW BLUNT LIVER INJURY MODEL IN MICE Takayuki Shibusawa, Jun Namiki, Sayuri Suzuki, Junichi Sasaki. Department of Emergency and Critical Care Medicine, Keio University School of Medicine, Tokyo, Japan
Background: Damage associated molecular patterns (DAMPs) are released into circulating blood stream and induced some immunologic reactions. There was no valuable animal model of blunt abdominal trauma. The aim of this study was to create a new mice model of blunt liver injury and prove DAMPs are in the circulation.
Methods: We anesthetized adult (8 week old) male wild-type C57BL/6J mice with isoflurane. We made a straight midline incision on the skin of the abdomen by scissor and opened the abdominal cavity. After the liver was exposure, we made bored wound on the right medial lobe (1 hole), left medial lobe (1 hole), and left lateral lobe (2 holes) by using Dermapunch. After making liver injuries, we closed the abdomen. Then we allowed to recover from anesthesia. We anesthetized again 2, 24, 96 and 168 hours later after making liver injuries and opened the abdomen. We punctured the posterior vena cava and took a blood sample. Then we separated plasma from whole blood. Asparate aminotransferase (AST), alanine aminotransferase (ALT), Interleukin-6 (IL-6), HMGB-1, total histone H3, total histone H4 and mitochondrial DNA in separated plasma were measured.
Results: AST and ALT levels were significantly higher in the liver injury group than the sham group at 2 and 24 hours after surgery. IL-6 level in the liver injury group was significantly higher at 2 hours after surgery. HMGB-1 level were significantly higher in 2 hours in liver injury group than sham group. There were no significant changes over 24 hours between two groups. Mitochondrial DNA levels in sham and liver injury group didn’t have significant changes. Total Histone H3 and H4 levels were significantly higher in liver injury group than sham group in all phases.
Conclusion: We could create a new valuable mice model of blunt liver injury. In this animal model, DAMPs were released in the circulation.
THE ALTERATIONS OF MILK FAT GLOBULE EGF-FACTOR 8 AND THE ASSOCIATION WITH ORGAN DYSFUNCTION IN TRAUMA ICU PATIENTS Yusuke Konda1, Akihisa Matsuda2, Marina Yamada1, Ami Shibata3, Yoriko Ichikawa1, Nobuyuki Saito1, Takanori Yagi1, Yoshiaki Hara1, Hisashi Matsumoto1, Hiroyuki Yokota4. 1Shock and Trauma Center, Chiba Hokusoh Hospital, Nippon Medical School, Chiba, Japan, 2Department of Gastrointestinal Surgery, Chiba Hokusoh Hospital, Nippon Medical School, Chiba, Japan, 3Department of Critical Care Center, Tama Nagayama Hospital, Nippon Medical School, Tokyo, Japan, 4Department of Emergency and Critical Care Medicine, Nippon Medical School, Tokyo, Japan
Objective: There is a strong relationship between inflammation and organ dysfunction in severe ICU patients. Previous studies showed that Milk fat globule-EGF factor 8 (MFG-E8) plays an important role in promoting phagocytosis and removal of apoptotic cell, attenuating “secondary necrosis” and inflammation. Although experimental studies demonstrated the deteriorative effect of reduced MFG-E8 production on organ function in sepsis, the functional role in human trauma patients is not determined. Therefore, we investigated the alterations of serum MFG-E8 and the association with organ function in trauma ICU patients.
Method: Peripheral serum samples were obtained from 31 trauma patients in our institute on hospital arrival, ICU admission (day 1), 2, 3, 5, 7, and 14. We measured serum MFG-E8 and IL-6 at point of hospital arrival, ICU admission (day 1), day 2, day 3, day 5, day 7, and day 14. We also examined Sequential Organ Failure Assessment (SOFA) score, probability of survival (TRISS), and length of ventilation day.
Result: Serum MFG-E8 levels were significantly suppressed at hospital compared with healthy controls, then turn to increase on day 2 to day 3. After that, MFG-E8 level decreased again on day 7 to day 14 (Fig.). Significant correlations were not observed between MFG-E8 and IL-6, SOFA score, length of ventilation. On day 14, the patients whose predicted survival rate is over 50% had significantly higher levels of MFG-E8 than those of less than 50% of predicted survival rate.
Conclusion: Our results suggested that MFG-E8 may not regulate inflammatory responses and organ function directly in trauma patients. However, the biphasic alterations and the higher values of severe patients on subacute phase indicated multi-functional roles of MFG-E8 such as anti-inflammatory and tissue remodeling effects that was shown in animal model.
COAGULOFIBRINOLYTIC RESPONSES TO TRAUMA HAS THE POTENTIAL TO DISSEMINATED INTRAVASCULAR COAGULATION WITH FIBRINOLYSIS INDEPENDENT FROM PRECEDED HYPERCOAGULATION Hironori Matsumoto, Kensuke Umakoshi, Yuki Nakabayashi, Suguru Annen, Muneaki Ohshita, Jun Takeba, Mayuki Aibiki. Department of Emergency and Critical Care Medicine, Ehime University Hospital, Ehime, Japan
Background: The aim of the current prospective observational study was to investigate coagulofibrinolytic responses after trauma.
Methods: In adult trauma patients admitted to our university hospital, clinical data and examinations were recorded. We measured blood cell counts and biomarkers of coagulation and fibrinolysis.
Results: Mean ISS (Injury Severity Score) of thirty-seven patients enrolled was 20.5. Time course in coagulofibrinolysis: A) Coagulatory parameters: The levels of TAT (thrombin-antithrombin complex) increased to the maximum level just after trauma on day 0, then remarkably decreased over time. B) Anticoagulants: AT (antithrombin) and PC (protein C) slightly decreased on day 1 through 0. TM (soluble thrombomodulin) showed the maximum level on day 2. C) Fibrinolytic parameters: PIC (plasmin-α2-plasmin inhibitor complex) showed the maximum level on day 0, which was followed by a drastical drop to the minimum level on day 2 through 1, then increased again. The time course of FDP (fibrin/fibrinogen degradation product) was similar to that of PIC. PLG (plasminogen) showed the minimum level on day 1 through 0. D) Inhibitors of fibrinolysis: α2-PI (α2-plasmin inhibitor) showed the minimum level on day 1 through 0. tPAI-1 (total plasminogen activator inhibitor) increased on day 1 through 0, that occurred later than increases in TAT and PIC after trauma. After that tPAI-1 dropped on day 2, and then it gradually increased as PIC. Differences between DIC and non-DIC groups: Ten patients were diagnosed with the DIC criteria of the JAAM (Japanese Association for Acute Medicine). IL-6 (interleukin-6) of the DIC group was higher than the non-DIC group, whereas ISS and lactate showed no differences between the groups. There were no differences in TAT on day 0 between the groups, whereas the DIC group showed significantly higher from day 1 to 4 than the non-DIC group. AT and PC of the DIC group showed lower levels than the non-DIC group during the study period. On day 0, PIC of the DIC group was significantly higher than the non-DIC group.
Conclusions: Our data showed that in the early phase of trauma, fibrinolytic activation and shut down were followed by reactivation. The DIC patients showed marked fibrinolytic activation as compared with the non-DIC patients, whereas the activation of coagulation was similarly observed in the both groups. These results indicate that the DIC in the early phase of trauma may occur through fibrinolytic activation. Furthermore our results showed the interaction between the sustained hypercoagulation and the lower levels of anicoagulants in the DIC patients.
SHOCKOMICS: NEW-OMICS SIGNATURES OF CIRCULATORY SHOCK Federico Aletti1, Manuela Ferrario1, Cristina Barlassina2, Giuseppe Baselli1, Eliandre de Oliveira3, Roberta Pastorelli4, Vicent Ribas5, Geert W. Schmid-Schoenbein6, Gabriella Tedeschi2, Bernardo Bollen Pinto7, Antoine Herpain8, Giuseppe Ristagno4, Eduardo Romay Medina9, Karim Bendjelid7. 1Dipartimento di Elettronica, Informazione e Bioingegneria, Politecnico di Milano, Milano, Italy, 2Universita degli Sudi di Milano, Milano, Italy, 3Parc Cientific de Barcelona, Barcelona, Spain, 4Istituto di Ricerche Farmacologiche Mario Negri, Milano, Italy, 5Custom Software & Electronics SL, Barcelona, Spain, 6University of California San Diego, San Diego, CA, USA, 7Hopitaux Universitaires de Geneve, Geneva, Switzerland, 8Universite Libre de Bruxelles, Brussels, Belgium, 9Hospital Universitari Mutua de Terrassa, Terrassa, Spain
The project “ShockOmics: Multiscale Approach to the Identification of Molecular Biomakers in Acute Heart Failure Induced by Shock” (ClinicalTrials.gov identifier NCT02141607) is a multidisciplinary effort which aims to identify novel biomarkers of shock-induced acute heart failure, to achieve a multiscale interpretation of circulatory shock, and to propose innovative therapeutic strategies against shock.
ShockOmics combines an observational clinical study with a complex animal study and in vitro experiments in order to search for -omics signatures of circulatory shock and acute heart failure related to shock. ShockOmics aims at the multiscale integration of data from the transcriptome, peptidome, proteome and metabolome of the plasma of shock patients monitored during a multiple day stay in the intensive care unit. In addition, the chronological -omics signatures of the patients will be related to hemodynamic and echocardiographic data.
The ShockOmics clinical trial has enrolled 66 shock patients, including septic shock and cardiogenic shock, from three intensive care units (Hopitaux Universitaires de Geneve, Geneva, Switzerland; Hopital Erasme, Brussels, Belgium; Hospital Mutua de Terrassa, Terrassa, Spain). Preliminary analyses of the blood samples collected from the patients at three different time points during ICU stays of up to 7 days have been aimed at verifying one of the hypotheses of ShockOmics, which attributes a fundamental injurious role to uncontrolled proteolysis. This hypothesis has been supported by data obtained through the analysis of plasma from hemorrhagic shock rats, which have shown the cleavage in shock of important categories of proteins, such as endogenous inhibitors of serine proteases and proteins involved in important inflammatory and metabolic processes. Preliminary transcriptomic, proteomic and metabolomic analyses of plasma patients have also shown the involvement of pathways related to the function of these families of proteins. Further, complex mathematical methods applied to -omics and hemodynamic data have shown promise as to the ability to cluster important metabolites in function of important clinical outcomes, such as mortality.
SWINE CLOSED HEAD INJURY MODEL INDUCED BY A PNEUMATIC NON-PENETRATING CAPTIVE BOLT DEVICE John R. Salsbury1,2, Christina L. Nelson1,2, Anita C. Randolph1,2, Perenlei Enkhbaatar1,2. 1Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA, 2Shriners Hospital for Children, USA
Background: Traumatic brain injury (TBI) is a common cause of morbidity and mortality with over 1.7 million cases in the U.S. each year. It is a major health concern in the military in both combative and noncombative settings. While there are several large animal models of TBI, the most studied are rodents. Results from rodent studies have translated poorly to clinical trials. Large animal models often require custom devices, which substantially increase costs. Many large animal models use gun powder charges. However, the limits in charge variations in these models can also limit gradation of the injury. We developed a swine closed head injury model, using pneumatically powered non-penetrating captive bolt device that induces a moderate TBI.
Methods: Four female Yorkshire pigs (20–30 kg) were anesthetized with Telazol. Surgical anesthesia was maintained with Isoflurane. Pigs were instrumented with an aortic catheter and a 5 Fr Swan-Ganz thermodilution catheter. The injury occurred in the same region in the right parietal lobe. The location of impact was identified by the intersection of 2 lines bisecting the right parietal bone (from the frontoparietal and parieto-occipital sutures, and the intraparietal suture and the lateral edge of the parietal bone). Midline incisions were made in the skull skin exposing the parietal bones. Periosteum was retracted. For intracranial pressure (ICP), an approximate 10 mm hole was drilled through skull and a catheter was placed in the ventricle. For inducing moderate TBI, impact was produced with a pneumatic injury device with regulated air pressure with 120 psi, producing 20 joules of energy. A 3×32×45 mm steel plate was placed on the site of target hit to spread impact force across plate and attenuate possible skull fracture. Animals were monitored for hemodynamic changes for 1–4 hours post injury in an anesthetized state and euthanized.
Results: All animals survived the study period (1–4 hrs). Mean arterial pressure started to fall (90 vs 106 baseline) 1minute after the injury and with a quick recovery at 5 minutes. Heart rate was increased, with the highest at 15 minutes to 213bpm vs 160 bpm baseline, and it sustained elevated throughout the study period. There was a sharp and sustained increase in ICP throughout the study. The highest ICP was observed (34 mmHg) at 5 minutes following the impact. There were no skull fractures seen in any animals. Subdural hematomas were present in all animals.
A CASE OF TOP OF THE BASILAR SYNDROME COMPLICATED WITH BRADYCARDIA AND ATRIAL FIBRILLATION-INDUCED CARDIOGENIC SHOCK Hideaki Suzuki, Ayumi Kawasaki, Kentaro Mochizuki, Rui Komatsu, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Background: Top of the basilar syndrome (TOBS) is a syndrome proposed by Caplan, and is characterized by a variety of clinical symptoms due to cerebral infarctions distributed above and below the cerebellar tentorium. We report a case of TOBS complicated with bradycardia and atrial fibrillation-induced cardiogenic shock.
Case: The patient was a 50-year-old woman with a history of untreated atrial fibrillation. A visiting friend found the patient in a state of impaired consciousness in her bed at home and called an ambulance. Upon arrival at our hospital, her level of consciousness was E1V3M5 on the Glasgow Coma Scale. Body temperature was 36.9°C, heart rate was 43 bpm with RR irregularity, blood pressure was 174/46, respiratory rate was 18 bpm, SpO2 was 100% (6 L of oxygen). The pupillary light reflex was weak in both eyes, with left and right pupil diameters of 2.0 mm and 3.0 mm, respectively and symptoms of bilateral vertical gaze paralysis and oculomotor nerve palsy. Head imaging done as part of a detailed examination revealed findings of bilateral paramedian thalamic infarction. The patient was consequently diagnosed with TOBS. Administration of heparin and edaravone was commenced on the same day. The bradyarrhythmia waveform was a mixture of atrial fibrillation and escape rhythm, which persistently deteriorated following admission despite several bolus administrations of 0.5 mg atropine sulfate. Bradycardia, reaching a heart rate of about 30 bpm, and hypotension, reaching a systolic blood pressure of 80 mmHg, persisted, and a temporary pacemaker was implanted. This pacemaker was electively replaced with a permanent pacemaker on day 19. Heparin and edaravone, which were administered for the treatment of TOBS in the acute phase, were switched to novel oral anticoagulants from day 31 of hospitalization. With this therapy, her disturbance of consciousness gradually improved and she was lucid by day 36 of hospitalization. Neurologically, the patient has residual eye movement disorders.
Discussion: In the present case, since the patient had a history of atrial fibrillation, cardiogenic cerebral infarction was the suspected etiology based on the mode of onset; however, head magnetic resonance imaging revealed no correlation between diffusion-weighted images and findings of apparent diffusion coefficient, indicating atypical cerebral infarction. More than 60% of cases of cerebral infarction exhibiting TOBS are reported to be embolic in nature, and while the present case was atypical, the impaired blood flow resulting from bradycardia and decrease in blood pressure is thought to have caused.
AUTOPHAGY IS MORE PROBABLE THAN APOPTOSIS IN ACUTE KIDNEY INJURY IN THE HOMORRHAGIC SHOCK MODEL USING IL-10 KNOCKOUT MICE Yoshinori Murao, Takami Nakao, Mitsuhide Hamaguchi, Noriko Tsuda, Ikuo Ota, Katsuyuki Maruyama, Toshifumi Uejima. Department of Emergency and Critical Care Medicine, Kindai University Faculty of Medicine, Osaka, Japan
Introduction: We examined the effects of IL-10 deficiency and HSP70 on acute kidney injury after hemorrhagic shock and resuscitation in mice. Autophagy and apoptosis were also examined using LC-3 or caspase-3.
Method: Male C57BL6/J mice and IL-10 knockout mice (IL-10 KO) weighing 20 to 30 g were anesthetized and the left femoral artery was cannulated. Heparin of 100 units was injected through the catheter. Blood was withdrawn until a mean arterial pressure of 40+/-5 mm Hg was attained. The level of hypotension was maintained for 60 min, and resuscitated with lactated Ringer's solution (2 times the volume of the shed blood,) and SB (shed blood). Kidney samples were harvested for histology section and immunohistochemical analysis. HSP 70, caspase-3 and LC-3 were examined with immunohistochemistry.
Results: Inflammatory cytokines were elevated at 2 h after hemorrhagic shock and resuscitation. On the contrary, histological section showed that renal tubule was more damaged at wild type at 48 h compared to IL-10 knockout type at 48 h (P = 0.053). HSP 70 was expressed more at renal tubule in KO type at 48 h than that of wild type at 48 h (P = 0.076). Caspase-3 (apoptosis) was expressed slightly, however, LC-3 (autophagy) was expressed at 4 h much more than caspase-3 in the renal tubule of the wild type.
Conclusion: These results show that the renal damage developed early after hemorrhagic shock, and it was thought that IL-10 might exacerbate acute kidney injury in this model. It is further suggested that HSP70 participated in the restoration of the renal tubule damage after hemorrhagic shock and resuscitation. Autophagy is more probable than apoptosis in its mechanism.
ELECTRO ACUPUNCTURE REDUCES NEED FOR VASOACTIVE DRUGS AFTER CARDIAC SURGERY Sven Asmussen1,2,3, Dirk M. Maybauer2, Rene Przkora4, Filippo Sanfilippo5, Kristofer Jennings2, John F. Fraser3, Marc O. Maybauer2,3,6. 1University Hospital of Ruhr University Bochum, Bochum, Germany, 2University of Texas Medical Branch, Galveston, TX, USA, 3University of Queensland, Australia, 4University of Florida, Gainesville, FL, USA, 5St. George Hospital, London, UK, 6Manchester University Hospitals, NHS FT, Manchester Heart Centre at Manchester Royal Infirmary and University, Manchester, UK
Objective: Acupuncture has a long history in China for over 2,500 years and is used worldwide as analgesic treatment in patients with acute and chronic pain. Recent studies showed that acupuncture might also be beneficial as a analgesic supplement to general anesthesia (GA). Aim of this study was to perform a systematic review and meta- analysis to assess the level of evidence for the clinical use of electro acupuncture (EA) in addition to GA in patients that underwent open-heart surgery about the need for vasoactive drugs on ICU post surgery and the levels for myocardial tissue marker Troponin I following cardiac surgery.
Methods: The systematic literature search in Medline, Cochrane Library, and Web of Science yielded a total of 220 citations, published between 1965 and December 31st, 2015. No systematic review or meta-analyses on this topic matched our search criteria. Each article of any language was assessed and rated for the methodological quality of the studies, using the recommendation of the Oxford Centre for Evidence Based Medicine (OCEBM).
Results: Three prospective randomized controlled clinical trials with a total of 180 patients matched our search criteria. The need for postoperative vasoactive drugs in cardiac surgery was significantly lower in the group that received EA+GA vs. GA alone: Odds ratio 5.618, lower limit 2.166, upper limit 14.571, Z-Value 3.550, P-Value <0.001. The amounts of cardiac Troponin I measured 24hrs post surgery were significantly lower in EA+GA group vs GA (n = 89 vs n = 91). Std difference in means 0.761, SE 0.256, Lower limit 0.259, upper limit 1.262, P-Value 0.003.
Conclusions: We conclude that electro acupuncture in combination with general anesthesia can be beneficial in regard of reducing postoperative circulatory instability shown by lower need for vasoactive drug support. In the following significantly lower values of cardiac tissue marker Troponin I might represent a less severe myocardial tissue damage. Further prospective investigations are needed to study these effects in regard of length of hospital stay and potential additional economical benefits.
INFLUENCE OF ETHYL PYRUVATE ON ORGAN DAMAGE, INFLAMMATION AND OUTCOME AFTER BLUNT CHEST TRAUMA AND HEMORRHAGIC SHOCK IN VIVO Nils Wagner1, Niklas Franz1, Scott Dieteren1, Katharina Mörs1, Mario Perl2, Ingo Marzi1, Borna Relja1. 1Department of Trauma, Hand and Reconstructive Surgery, University Hospital Frankfurt, Goethe University, Frankfurt, Germany, 2BG-Trauma Center Murnau, Murnau, Germany
Background: Blunt chest trauma (TxT) and hemorrhagic shock with resuscitation (H/R) are closely associated with a strong inflammatory response and impaired organ function. Treatment of patients suffering from TxT+H/R is still challenging. Therefore, here we investigated whether a reperfusion treatment with ethyl pyruvate (EP) influences liver damage, inflammatory changes and outcome after TxT+H/R.
Methods: Female rats received a blunt chest trauma followed by hemorrhagic shock (35 ± 5 mmHg) for 60 minutes and subsequent resuscitation. Animals undergoing surgical procedure were grouped as shams. Resuscitation was performed by own blood transfusion either with lactated Ringer solution (Rl) or RI supplemented with EP. Blood, liver and lung tissue were removed two hours later for analyses. Mortality was assessed after 72 h. Serum alanine aminotransferase (ALT) and liver damage using hematoxylin-eosin (H/E) staining were evaluated. Hepatic gene expression of interleukin (IL)-6 or intercellular adhesion molecule (ICAM)-1, and infiltration with polymorphonuclear leukocytes (PMNL) were assessed. Pulmonary gene expression of surfactant protein A (SP-A) was analyzed. Lipolysaccharide-stimulated tumor necrosis factor (TNF)-α release from peripheral whole blood was detected. p < 0.05 was considered significant (ANOVA).
Results: TxT+H/R-induced ALT increase and liver damage were significantly reduced by EP. Total circulating protein and albumin levels were comparable to sham group, and markedly higher than in Rl group after TxT+H/R. Hepatic IL-6 gene expression was significantly reduced by EP compared to Rl after TxT+H/R and comparable to sham. Consistent with enhanced ICAM-1 gene expression, highest PMNL infiltration of the liver was found in Rl group and significantly blunted by EP after TxT+H/R. SP-A was higher in sham and EP group after TxT+H/R compared to Rl after TxT+H/R. TNF-α release was reduced after TxT+H/R compared with sham, and further increased by EP after TxT+H/R. EP-treated animals showed a not significant but 10% improved survival compared to RI after TxT+H/R.
Conclusions: This study indicates that resuscitation with EP attenuated the local and improved the systemic immune response after blunt chest trauma and hemorrhagic shock indicating at a protective effect on liver and lung.
Supported by DFG RE 3304/5-1 and PE 908/3-1
PIOGLITAZONE RESTORES PHAGOCYTIC ACTIVITY AND AMELIORATES POSTBURN INFECTION IN MICE Hiromi Miyazaki1, Manabu Kinoshita2, Hiroyuki Nakashima2, Masahiro Nakashima2, Soichiro Seno1, Yasumasa Sekine1, Satoshi Tomura1, Shuhji Seki2, Daizoh Saitoh1. 1Division of Traumatology, National Defense Medical College, Saitama, Japan, 2Department of Immunology and Microbiology, National Defense Medical College, Saitama, Japan
Background: Severe burn injury causes the Kupffer cells dysfunction and attenuates the host defense against bacterial infections. Peroxisome proliferator-activated receptor-γ (PPAR-γ) is abundantly expressed in macrophages and plays a critical role for regulation of immune responses during the inflammation. Pioglitazone, a PPAR-γ agonist, exert anti-inflammatory actions in different disease models, including septic shock. We herein investigated the role of PPAR-γ during the inflammatory response in postburn bacterial infection by using the pioglitazone.
Material and Methods: C57BL/6 mice received a 20% full-thickness burn injury. Five days after injury, they were administered with pioglitazone (10 mg/kg, i.p.) or vehicle, followed by E. coli challenge.
Results: Burn injury did not affect the expression of PPAR-γ in hepatic mononuclear cells including Kupffer cells. Although burn injury augmented the elevation of plasma TNF after E. coli challenge in mice, pioglitazone pretreatment significantly suppressed this elevation. Burn injury also down-regulated the phagocytic activity by Kupffer cells and impaired bacterial clearance in the liver after E. coli challenge. Interestingly, pioglitazone pretreated burn-injured mice augmented the phagocytic activity by Kupffer cells and restored bacterial clearance in the liver, thereby improving mouse survival after postburn E. coli infection.
Conclusion: It is concluded that PPAR-γ agonist modulates inflammatory responses following post-burn infection via a mechanism dependent on suppressing proinflammatory cytokine production and enhancing phagocyte function of Kupffer cells/macrophages.
ACUTE VS. SUB-ACUTE ALCOHOL-BINGE END IN DIFFERENTIAL MORTALITY RATES, INFLAMMATORY RESPONSE AND LIVER INTEGRITY AFTER BLUNT CHEST TRAUMA FOLLOWED BY HEMORRHAGE/RESUSCITATION (H/R) IN VIVO Borna Relja1, Nils Wagner1, Niklas Franz1, Scott Dieteren1, Katharina Moers1, Mario Perl2, Ingo Marzi1. 1Trauma, Hand and Reconstructive Surgery, University Hospital, Goethe University Frankfurt, Frankfurt, Germany, 2BG-Trauma Center Murnau, Murnau, Germany
Background: Alcohol-binge is associated with increased incidence of trauma injury and impaired organ integrity. Previously, we have demonstrated improved hepatic integrity and survival rates after sub-acute (12–16 h) alcohol-binge prior H/R. Here, we examined the impact of acute vs. sub-acute alcohol-binge on liver injury, inflammation and mortality in a more severe trauma model consisting of blunt chest trauma (TxT) followed by H/R.
Methods: 12 h (sub-acute) or 2 h (acute) before TxT+H/R, female rats received alcohol (5 g/kg, 30%EtOH) or saline gavage. Then, TxT followed by hemorrhage for 60mins and resuscitation were induced. 2 h later, liver tissue and blood were harvested. Mortality was assessed after 72 h. Hepatic necrosis, local inflammation (infiltration with polymorphonuclear leukocytes, PMNL, tumor necrosis factor (TNF)-alpha protein and gene expressions of interleukin (IL)-6 and intercellular adhesion molecule (ICAM)-1 and systemic TNF-alpha release upon an overnight lipopolysaccharide (LPS) stimulation of peripheral whole blood were evaluated (ANOVA, p < 0.05 was considered significant).
Results: Alcohol decreased the basal mean arterial blood pressure (MABP) and total blood loss required to reach and sustain 35 mmHg. This decrease was more profound in acute vs. sub-acute alcohol-binge group. TxT+H/R-induced increase in hepatic TNF-alpha protein level was significantly reduced to 85% in sub-acute and to baseline in acute-binge group. Similarly, TxT+H/R-induced ICAM-1 gene expression was diminished by sub-acute and acute binge. In contrast, acute alcohol-binge further increased IL-6 gene expression compared to the corresponding control group after TxT+H/R, data that were in line with PMNL infiltration. LPS-stimulated release of TNF-alpha was diminished after TxT-H/R in all groups compared with shams, while sub-acute alcohol-binge suppressed further TNF-alpha release compared to the corresponding TxT-H/R-control group. Liver injury was amplified by alcohol notably in the acute-binge group. Mortality has not worsened in the sub-acute alcohol-binge group, while it was increased in acute alcohol-binge group compared with corresponding control groups after TxT+H/R.
Conclusions: Alcohol-binge results in hemodynamic instability and suppression of inflammatory markers. Acute alcohol-binge has worsened the outcome from TxT+H/R as compared with sub-acute alcohol-binge, indicating at the importance of timing and dose of alcohol-binge for the inflammatory status quo.
Supported by DFG RE 3304/5-1 and PE 908/3-1
FIBRINOGEN γ-CHAIN PEPTIDE-COATED, ADP-ENCAPSULATED LIPOSOMES RESCUE MICE FROM LETHAL BLAST LUNG INJURY VIA ADENOSINE SIGNALING Kohsuke Hagisawa1, Manabu Kinoshita2. 1Department of Physiology, National Defense Medical College, Saitama, Japan, 2Department of Immunology and Microbiology, National Defense Medical College, Saitama, Japan
Objectives: Fibrinogen γ-chain (dodecapeptide HHLGGAKQAGDV, H12)-coated, adenosine diphosphate-encapsulated liposomes [H12-(ADP)-liposomes] can accumulate via H12 interactions at bleeding sites where they release ADP that is rapidly metabolized to adenosine, which has tissue-protective effects. We investigated the efficacy of H12-(ADP)-liposomes to treat blast lung injury, with a focus on adenosine signaling.
Methods: Mice were pretreated with H12-(ADP)-liposomes, H12-(PBS)-liposomes, ADP-liposomes, or PBS-liposomes. Five minutes after treatment the mice received a single laser induced shock wave (LISW, 1.8 J/cm2) that caused lethal blast lung injury, and their survivals and lung injuries were then examined. Post-treatment with H12-(ADP)-liposomes or H12-(PBS)-liposomes was also performed to the mice 1 min after LISW exposure and evaluate the therapeutic effect. To examine the effect of adenosine signaling, adenosine A2A receptor (ZM241385) or adenosine A2B receptor (PSB 1115) antagonists were administered to the mice 1 h before the pre-treatment with H12-ADP-liposomes that was followed by LISW exposure.
Results: Pretreatment as well as post-treatment with H12-(ADP)-liposomes significantly increased mouse survivals [H12-(ADP)-liposomes: 58% survival vs. H12-(PBS)-liposomes: 8%, p < 0.05 (post-treatment)] and mitigated pulmonary tissue damage/hemorrhage and neutrophil accumulation after LISW exposure. H12-(ADP)-liposomes accumulated at pulmonary vessel injury sites after LISW exposure by both pre and post-treatment. Pretreatment with H12-(ADP)-liposomes reduced albumin and macrophage inflammatory protein-2 levels in bronchoalveolar lavage fluid. Although H12-(ADP)-liposome pretreatment did not affect blood coagulation activity in the injured mice, its beneficial effect on blast lung injury were significantly abrogated by adenosine receptor A2A or A2B antagonism [A2A antagonist: 17% survival, A2B antagonist; 33%, vs. DMSO control: 80%, p < 0.05 respectively], suggesting that adenosine signaling improved lung injury.
Conclusions: H12-(ADP)-liposomes may be effective against blast lung injury by promoting tissue-protective adenosine signaling, and could represent a novel controlled-release drug delivery system.
ETHYL PYRUVATE REDUCES LUNG APOPTOSIS AND INFLAMMATION AFTER BLUNT CHEST TRAUMA AND HEMORRHAGIC SHOCK Mario Perl1, Nils Wagner2, Niklas Franz2, Scott Dieteren2, Katharina Moers2, Ingo Marzi2, Borna Relja2. 1BG-Trauma Center Murnau, Murnau, Germany, 2Trauma, Hand and Reconstructive Surgery, University Hospital Goethe University Frankfurt, Frankfurt, Germany
Background: After blunt chest trauma followed by hemorrhagic shock with resuscitation, intense local inflammatory changes are closely associated with apoptotic cell loss and subsequently impaired pulmonary function. Therefore, the treatment of patients suffering from blunt chest Trauma and hemorrhage with resuscitation is still challenging. Here, the early post-traumatic local inflammatory and apoptotic responses were analyzed upon a therapeutic reperfusion with ethyl pyruvate (EtP) after blunt chest trauma followed by hemorrhage and resuscitation.
Methods: Female rats underwent a blunt chest trauma followed by hemorrhagic shock (35 mmHg) for 60 minutes and were subsequently resuscitated. Sham animals underwent surgical procedures only. Resuscitation was performed by own blood transfusion either with lactated Ringer solution (RL) or RL supplemented with EtP. Two hours later, lung tissue was removed for analyses. Pulmonary gene expression of interleukin (IL)-6 was assessed. Total pulmonary protein content of tumor necrosis factor (TNF)-alpha and infiltration with polymorphonuclear leukocytes (PMNL) were determined. Apoptotic changes were evaluated by gene analyses of Fas and Bcl-2. p < 0.05 was considered significant (ANOVA).
Results: Trauma and hemorrhage induced significant increase in IL-6 gene expression and TNF-alpha protein expression in the lung. Compared to sham animals, the infiltration of lungs with PMNLs was significantly increased after trauma and hemorrhage. Resuscitation with EtP significantly reduced this local inflammatory response to trauma and hemorrhage. Fas and Bcl-2 gene expressions were significantly induced after trauma. While EtP treatment further enhanced Bcl-2 expression, Fas expression was reduced upon EtP resuscitation.
Conclusions: Taken together, this study indicates that resuscitation with EtP diminished the local inflammatory and apoptotic changes after blunt chest trauma and hemorrhagic shock. As it has been shown before that Fas expression silencing in the lung was effective against ischemia-reperfusion injury, the possible protective mechanism of EtP may involve a Bcl-2-mediated down-regulation of Fas-induced apoptosis indicating at protective effects on lung after trauma and hemorrhagic shock.
Supported by DFG RE 3304/5-1 and PE 908/3-1
A CASE OF SUCCESSFUL CARDIAC RESUSCITATION WITH EXTRACORPOREAL MEMBRANE OXYGENATION FOR HYPERTROPHIC CARDIOMYOPATHY Ahmin Kim, Azusa Ichimaru, Jaewan Park, Mayu Kikuchi, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Case: A 26-year-old woman
Patient History: hypertrophic cardiomyopathy
Clinical Course: The patient collapsed at a train station on her way to work and a station attendant subsequently called an ambulance and an emergency medical team arrived 6 minutes later. Cardiopulmonary arrest was confirmed and chest compressions were initiated. After confirmation of ventricular fibrillation, direct current defibrillation was performed twice, but the patient progressed into pulseless electrical activity. The patient was still in a state of PEA on arrival at our hospital, and was intubated and given epinephrine. Cardiopulmonary circulation briefly resumed but VF returned almost immediately afterwards. Defibrillation was performed six times and medication weas also administered. However, VF was sustained, and percutaneous veno-arterial extracorporeal membrane oxygenation was introduced. After a steady waveform was confirmed, the patient was admitted to the intensive care unit, and therapeutic hypothermia was started. Both pupils were 2.5 mm and responded to light. Ejection fraction of 15% was observed with bedside echocardiography under continuous administration of epinephrine and dopamine. The patient was unable to urinate and a trend toward exacerbation in acidemia was observed, and continuous hemodiafiltration was introduced later in the day. On Day 4, rewarming concluded without complications. The patient was transferred to coronary care unit for a coronary angiography was performed; however, a significant stenosis of the coronary artery was not observed and then IABP was inserted. On Day 5, V-A ECMO was removed successfully without complications. On Day 6, 27 consecutive beats of nonsustained ventricular tachycardia developed, which was treated with amiodarone. On Day 8, urinary flow returned to normal with continuous administration of loop diuretics, and the CHDF was removed. On Day 9, stabilization in blood pressure was observed even when the intraaortic balloon pump was removed. On Day 10, ejection fraction on echocardiogram recovered to 45%. The patient was weaned from the ventilator on Day 23.
Discussion: The present case of a hypertrophic cardiomyopathy was resuscitated by V-A ECMO. According to the Study of Advanced Life Support for Ventricular Fibrillation with Extracorporeal Circulation in Japan (SAVE-J) study, the neurological prognosis one month after resuscitation was favorable in a significantly greater percentage of the ECMO group compared to the non-ECMO group. In our case, a recovery in consciousness level was also observed within one month, indicating that a proactive introduction of ECMO should be considered in cardiopulmonary resuscitation.
A PRESSURE-GUIDED SHOCK MODEL DID NOT REDUCE INDIVIDUAL RESPONSE VARIABILITY, COMPARED TO A VOLUME-GUIDED MODEL Lisa Smart, Corrin J. Boyd, Anthea L. Raisis, Giselle Hosgood. Comparative Health Research Group, College of Veterinary Medicine, Murdoch University, Murdoch, WA, Australia
Background: Our group previously used a canine haemorrhagic shock model that removed a set volume of blood over a short period of time before comparing resuscitation interventions. We found wide variability in the degree of shock achieved. Removal of blood guided by blood pressure may be more synergistic with the individual's ability to compensate and reduce variability across a group.
Objectives: 1) To assess if a pressure-guided canine shock model (PGM) created more severe shock, as measured by a higher oxygen extraction ratio index (0ER), compared to a volume-guided model (VGM). 2) To determine if the PGM provided less variability in shock parameters. 3) To identify parameters contributing to 0ER variance in the PGM.
Methods: The VGM model included 12 anaesthetised greyhound dogs whereby removal of 48 mL/kg of blood was targeted over 30 minutes. The PGM model included 24 anaesthetised greyhound dogs whereby blood was removed to maintain a mean arterial blood pressure (MAP) <60 mmHg for 60 minutes. Cardiovascular parameters were measured at baseline (T0) and at the end of blood removal (T1). Linear regression was used to compare means. Variance of each parameter was compared at T1 using Levene's test. Regression modelling was used to find the best explanatory variables for the variance in volume of blood removed (VOL)(variables at T0) and T1 OER (variables at T1) in the PGM. Significance was set at P < 0.05.
Results: There was no significant difference in VOL between the PGM (mean 48.8 mL/kg, 95% confidence interval 44.9–52.7) and VGM (47.9, CI 47.5–48.3). At T1, the PGM had a significantly lower mean OER (0.45, CI 0.39–0.50) compared to the VGM (0.60, CI 0.49–0.70). The PGM also had a significantly higher mean heart rate, higher mean MAP, higher mean systemic vascular resistance index (SVR), lower mean oxygen consumption, higher mean lactate and lower mean base excess.
There was no significant difference in the variance of OER, or other parameters, between the two models.
Stroke volume index (SV) explained most of the variance for both VOL and OER. For predicting VOL, a model including T0 variables SV, MAP and central venous pressure provided the highest R2 (0.45) while minimising C(p)(2.0). For predicting OER, the best model included T1 variables SV, VOL and SVR (R2 0.29, C(p) 0.59).
Conclusion: Although there was no difference in mean VOL, the PGM showed evidence of better compensation, including lower OER and other parameters supportive of a sympathetic response. This may be due to the pressure-guided strategy or longer shock period. The change in model to a PGM did not reduce the variability of shock parameters. Individual SV best explained the variation in response to haemorrhage.
THE COMPARISONS OF EV1000 HEMODYNAMIC PARAMETERS MEASUREMENTS BETWEEN THE JUGULAR AND FEMORAL VEIN COLD SALINE INJECTION DURING CONTINUOUS RENAL REPLACEMENT THERAPY Toshikazu Sakai1, Yoshitaka Hara1, Yu Kato1, Tomoyuki Nakamura1, Yasuyo Shimomura1, Takahiro Kawaji1, Yasuyoshi Kurimoto1, Mariko Nagata1, Seiko Hayakawa1, Hidefumi Komura1, Chizuru Yamashita1, Junpei Shibata1, Kazuhiro Moriyama2, Osamu Nishida1. 1Department of Anesthesiology and Critical Care Medicine, Fujita Health University School of Medicine, Aichi, Japan, 2Laboratory for Immune Response and Regulatory Medicine, Fujita Health University School of Medicine, Aichi, Japan
Introduction: Although previous studies suggest that transpulmonary thermodilution (TPTD) measurements are influenced significantly by blood purification (BP), the influence of injection position of cold saline during BP has not yet been studied. We assessed whether the position of the cold saline injection site in the body of the same individual pig.
Methods: Six female crossbred pigs (35 to 40 kg) were studied. EV1000 monitor was used as TPTD. Vascular access for blood purification was placed in the left external jugular vein. A blood circuit without a filter was used to mimic BP with blood flow rate of 150 ml/min. Cold saline was injected through a central venous catheters in the right external jugular vein or the right femoral vein. Cardiac output (CO), global end-diastolic blood volume (GEDV) and extravascular lung water (EVLW) were measured during BP and at the time of cessation. All data were expressed as the median. Statistical analysis was performed using Wilcoxon signed-rank test.
Results: In the jugular vein, the obtained date when off pump vs on pump were; 2.7 vs. 2.9 L/min (P = 0.04)(CO), 403 vs. 438 ml (P = 0.04)(GEDV), 310 vs. 306 ml (P = 0.92)(EVLW), respectively. In the femoral vein, the obtained date when off pump vs on pump were; 2.6 vs. 2.8L/min (P = 0.18)(CO), 497 vs. 500 ml (P = 0.18)(GEDV), and 341 vs. 345 ml (P = 0.44)(EVLW), respectively.
Discussion: CO and GEDV were significantly underestimated when injected through the jugular vein comparing with through the femoral vein. These results might be caused by the difference of mixed sites where cold saline and circulation blood are joined together.
Conclusion: When blood purification is performed in the jugular vein, TPTD measurements obtained by cold saline injection through the jugular vein are susceptible to influence than those obtained through the femoral vein.
THROMBOELASTOMETRIC ANALYSIS ON THE RISK FACTORS OF RETURN OF SPONTANEOUS CIRCULATION IN ADULT PATIENTS WITH OUT-OF-HOSPITAL CARDIAC ARREST Hiroyuki Koami1, Yuichiro Sakamoto1, Tomoya Matsuda2, Junpei Nishi2, Shiki Nakayama2, Kento Nakayama2, Ryota Sakurai2, Miho Ohta2, Hisashi Imahase2, Mayuko Yahata2, Mitsuru Umeka2, Toru Miike2, Futoshi Nagashima2, Takashi Iwamura1, Kosuke C. Yamada2, Satoshi Inoue3. 1Emergency and Critical Care Medicine, Saga University, Saga, Japan, 2Advanced Critical Care Center, Saga University Hospital, Saga, Japan, 3Trauma Surgery and Surgical Critical Care, Saga University, Saga, Japan
Introduction: It is well-known that coagulation and fibrinolytic disorders are observed in patients with out-of-hospital cardiac arrest (OHCA). However, there are few evidences between the presence of coagulopathy and a possibility of return of spontaneous circulation (ROSC) until now. The aim of this study is to identify factors that are associated with ROSC in patients with OHCA by utilizing thromboelastometry (ROTEM).
Methods: Seventy five adult OHCA patients transported to the Saga University Hospital with ROTEM performed during the cardiopulmonary resuscitation in the emergency department (ED) from January 2013 to December 2015 were enrolled in this study. Patients with trauma, pregnancy, and ROSC before admission were excluded. All patients were divided into two groups; ROSC group (n = 23) and non-ROSC group (n = 52). We retrospectively evaluated the demographics, prehospital care, and laboratory tests on admission including ROTEM findings.
Results: ROSC after admission to ED was not significantly related with gender, age, prehospital care, by stander CPR, initial rhythm at scene, medication history, presence of liver cirrhosis and estimated time from onset to blood sampling. Blood tests showed significant improvement in coagulation and fibrinolytic system in ROSC group including higher platelet and fibrinogen value, shortened APTT, lower FDP, lower D dimer and lower lactate. There were statistically significant shorter clotting time, stronger clot firmness, accelerated clot formation and higher lysis index in the EXTEM test in the case of ROSC group. Next, multiple logistic regression analysis revealed that lactate (P = 0.028) and A30 of EXTEM (P = 0.009) were significantly associated with ROSC. The ROC analysis indicated the cut-off values of them as 12.0 mmol/L and 48.0 mm, respectively. A new criteria for ROSC (lactate <= 12.0 mmol/L and A30 of EXTEM >= 48.0 mm) indicated lower sensitivity (40.9%), higher specificity (94.7%) and higher accuracy (75.0%).
Conclusion: A new diagnostic criteria with lactate and ROTEM parameter of clot firmness predicts a possibility of ROSC after admission to the ED in adult patients with OHCA.
FULL RECOVERY OF CARDIAC ARREST IN ACCIDENTAL SEVERE HYPOTHERMIA TREATED BY EXTRACORPOREAL CARDIOPULMONARY RESUSCITATION (ECPR) Kazuma Shibahara, Hideaski Suzuki, Noriko Saito, Mihoko Tsunoda, Noboru Akiduki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Introduction: Extracorporeal cardiopulmonary resuscitation (ECPR) can contribute to both the rewarming and the resuscitation of patients in cardiac arrest after accidental severe hypothermia; however, there are few studies regarding the effectiveness of ECPR in these patients. Because there is insufficient evidence for ECPR in severe hypothermia, it is not routinely used in these cases. We report two patients with cardiac arrest after severe hypothermia who made a full recovery after treatment by ECPR.
Case 1: A 47-year-old man with schizophrenia was transferred to our department in cardiopulmonary arrest (CPA). Defibrillations were attempted thrice for ventricular fibrillation by paramedics before arriving at hospital; however, there was no return of spontaneous circulation (ROSC). His core body temperature was 22.1°C (71.8°F) on arrival at the hospital. Because an emergency tracheotomy had been performed for truisms, venoarterial extracorporeal membrane oxygenation (VA ECMO) was initiated for rewarming and resuscitation. After 50 min, his core body temperature increased to 28.0°C (82.4°F), and the patient was successfully defibrillated. VA ECMO was withdrawn 10 min after ROSC. Hypothermia was maintained at 34°C (93.2°F) for 48 h, and catecholamines were required for 6 days; the ventilator was withdrawn on day 8. He was discharged from the hospital on day 61.
Case 2: A 53-year-old man with an unremarkable medical history was transferred to our department in CPA. His core temperature on arrival at the hospital was 25.8°C (78.4°F), and VA ECMO was initiated. ROSC was obtained after 1 h, once his core temperature had increased to 28.2°C (82.8°F). Hypothermia with VA ECMO was maintained for 48 h, and catecholamines and ventilation were required for 5 days. After the sepsis due to lower extremity gangrene was treated, the patient was discharged on day 21.
Discussion: Rapid rewarming is the most important procedure for cardiac arrest in accidental severe hypothermia. The rate of body temperature increase by ordinary rewarming is 0.5°C–2.0°C (32.9–35.6°F)/h, while the maximum rate by VA ECMO is 9.5°C (49.1°F)/h. ECPR should be more actively used for cardiac arrest in accidental severe hypothermia.
RENAL SYMPATHETIC RESPONSE TO HEMORRHAGIC SHOCK IN ANESTHETIZED MICE Mamoru Tanida, Tao Zhang, Toshishige Shibamoto. Department of Physiology II, Kanazawa Medical University, Ishikawa, Japan
Objectives: Autonomic nervous system is the defense system against fetal status such as circulatory shock. It was reported that renal sympathetic nerve activity (RSNA) and heart rate (HR) were decreased during acute hemorrhagic shock in anesthetized rats (Victor et al., Circ Res. 64:686–94. 1989). However, it has not yet been known whether the same response to hemorrhagic hypotension is observed in mice. In the present study, we determined the responses of RSNA and HR to hemorrhagic hypotension in anesthetized mice. In addition, we explored the mechanism for the hemorrhage-induced decreases in RSNA and HR.
Method: Male C57/BL6J mice were used in this study. All animal care and handling procedures were approved by the Animal Research Committees of Kanazawa Medical University. Under pentobarbital anesthesia, RSNA, systemic arterial pressure (SAP) and HR were continuously measured. Hemorrhagic shock was evoked by removal of blood through the femoral artery so that SAP decreased to 50 mmHg within 2 min and this hypotensive level was sustained for 10 min. We examined the effects of cervical vagotomy alone and combined with carotid sinus denervation, and pretreatment with capsazepine (i.v., 90 μg), the antagonist of transient receptor potential cation channel subfamily V member 1 (TRPV1).
Result: During hemorrhagic hypotension in mice, RSNA initially increased to around 135% of baseline along with a decrease in SAP and then it showed a sustained reduction to around 70% of baseline. HR progressively decreased from the baseline of 450 bpm to 400 bpm. These responses of RSNA and HR were basically similar to that of rats previously reported. Either cervical vagotomy alone or pretreatment with capsazepine significantly eliminated the hemorrhage-induced secondary decreases in RSNA and HR, but not the initial increases. The carotid sinus nerve denervation combined with vagotomy abolished both of the initial increase and secondary decrease of RSNA and HR during hemorrhagic hypotension.
Conclusion: RSNA shows an initial increase, followed by a sustained decrease, and HR a progressive decrease during acute hemorrhagic hypotension in anesthetized mice. This initial sympathoexcitation is mediated by carotid sinus baroreceptor reflex, while the secondary symathoinhibition and bradycardia by the vagal afferents and TRPV1 activation.
THE NERVE GROWTH FACTOR MIMETIC GK-2 PROTECTS NEURONS IN POST-RESUSCITATIVE PERIOD AFTER CARDIAC ARREST IN RATS Maria Sh. Avrushchenko1, Irina V. Ostrova1, Yuriy V. Zarzhetsky1, Tatyana A. Gudasheva2, Sergey B. Seredenin2. 1V.A. Negovsky Research Institute of General Reanimatology, Moscow, Russia, 2V.V. Zakusov Research Institute of Pharmacology, Moscow, Russia
Ischemia-reperfusion causes nerve cell damage in various regions of the brain. There are experimental data of neuroprotective properties of NGF, however it poorly crosses the blood-brain barrier, and produces side effects. The low-molecular-weight analogue of NGF - dipeptide GK-2 - reproduces the therapeutic effects of NGF without producing side effects.
Objective: The purpose of this study was to evaluate an effectiveness of GK-2 for the brain protection in the post-resuscitative period and to elucidate its influence on expression of neurotrophic factors bFGF, NT4 and BDNF.
Methods: Adult white male rats (n = 20) underwent cardiac arrest for 12 minutes by intrathoracic clamping supracardiac bundle of vessels, followed by resuscitation. 10 rats were injected GK-2 (1 mg/kg i/p) at 30 minutes and 48 hours after resuscitation. 10 animals received equivalent doses of saline. The control group consisted of sham-operated animals (n = 10). The neurological state of the resuscitated animals was assessed. Histological and immunohistochemical studies of highly hypoxia-sensitive neuronal populations of hippocampal pyramidal cells (field CA1) and the cerebellar Purkinje cells were performed on day 7 post-resuscitation. The total number of neurons, the number of normal and altered cells, as well as the number of bFGF-, NT4-, BDNF-positive cells per 1 mm of the layer length were determined. Data were statistically processed by Student's t-test or Mann-Whitney U-test.
Results: The use of GK-2 accelerated neurological recovery of the resuscitated animals. It was found degeneration of CA1 pyramidal neurons and loss of Purkinje cells post-resuscitation. Namely NT4-negative, bFGF-negative and BDNF-negative cells died. GK-2 prevented the development of degenerative changes of neurons in the hippocampus and dramatically reduced the neuronal loss in the cerebellum. GK-2 had no effect on the expression of bFGF and NT4, however, promoted an increase in the expression level of BDNF. Initiating the expression of BDNF in neurons that had been not previously producing this factor GK-2 contributed to preserving these neurons. The obtained data allow us to conclude that at least one of the mechanisms of the neuroprotective action of GK-2 is its ability of to activate the expression of BDNF in nerve cells.
Conclusion: The nerve growth factor mimetic GK-2 protects neurons from degenerative changes and death after ischemia-reperfusion as well as accelerates the neurological recovery of resuscitated animals. These data provide rationale for using this drug for the prevention of posthypoxic encephalopathies.
A CASE OF RUPTURED LEFT GASTROEPIPLOIC ARTERY ANEURYSM LEADING TO HEMORRHAGIC SHOCK Azusa Ichimaru, Kazuma Shibahara, Mayu Kikuchi, Jaewan Park, Kentaro Mochizuki, Noboru Akizuki, Mizuho Namiki, Munekazu Takeda, Arino Yaguchi. Department of Critical Care and Emergency Medicine, Tokyo Women's Medical University, Tokyo, Japan
Introduction: Abdominal visceral artery aneurysm is a rare condition and among these aneurysms, left gastroepiploic artery aneurysm occurs infrequently. We show a case of hemorrhagic shock caused by a rupture in the left gastroepiploic artery aneurysm and this patient was successfully treated with transcatheter arterial embolization (TAE).
Case: An 82-year-old man with a history of prostate cancer, hypertension, and Alzheimer's disease complained of dyspnea at his residential facility, and was transported by ambulance. Vitals on arrival were: Body temperature of 35.7°C, pulse of 90 beats/min, blood pressure of 88/65 mmHg, respiratory rate of 27 breaths/min, and SpO 2 99% (10 L/min of oxygen). There were no significant abnormalities on electrocardiogram. Chest X-ray showed a cardiothoracic ratio of 52% and decreased radiolucency in the left lower lung field. Arterial blood gas test (under 10 L/min of oxygen) results were: pH 7.5, pCO 2 30.9 mmHg, pO 2 274.3 mmHg, BE 2.2 mmol/L and lactic acid 2.9 mmol/L. Blood test results were: white blood cells 12,200/μL, Hb 9.5 g/dL, PT(INR) 1.03, APTT 29.5 sec, FDP 9.2 μg/mL, D-dimer 4.2 μg/mL and CRP 4.88 mg/dL. On chest to pelvis contrast-enhanced computed tomography (CT), a small amount of pleural effusion in both lungs, inflammatory scarring of both lower lobes, vascular wall calcification at the aortic arch were observed. Furthermore, bloody ascites was detected from the right subphrenic space into the pelvis and a 19 mm pseudoaneurysm that connects to the left gastric artery was evident, and the patient was diagnosed with ruptured left gastroepiploic artery aneurysm. TAE was performed and the embolization of the left gastroepiploic artery was performed by blowing gelatin sponge from the proximal portion of the aneurysm to the distal branch. On abdominal contrast-enhanced CT performed on Day 4, blood flow was not observed in the aneurysm. The patient resumed eating on Day 4 without symptoms or progression of anemia, and was subsequently transferred.
Conclusion: In abdominal visceral aneurysm, 90% of the cases are diagnosed due to a rupture, and common symptoms include abdominal pain, anemia, and gastrointestinal bleeding. Moreover, while it has been reported that hemostasis with interventional radiology should be attempted first for treatment, there is no consensus specifically for gastroepiploic artery aneurysm. Here, we experienced a case where a patient who developed dyspnea progressed into a shock state, and was diagnosed with ruptured left gastroepiploic artery aneurysm and successfully underwent treatment with TAE.
ASSOCIATIONS OF GLYCOCALYX DAMAGE BIOMARKERS WITH INFLAMMATION AND ENDOTHELIAL ACTIVATION IN INFECTION COMPARED TO TRAUMA Lisa Smart1,2, Stephen PJ. MacDonald1,2, Sally Burrows3, Erika Bosio1,2, Glenn Arendts1,2, Daniel M. Fatovich1,2. 1Centre for Clinical Research in Emergency Medicine, Harry Perkins Institute of Medical Research, Perth, WA, Australia, 2Department of Emergency Medicine, University of Western Australia, Perth, WA, Australia, 3Department of Medicine and Pharmacology, University of Western Australia, Perth, WA, Australia
Background: Ongoing work by our group found differences in endothelial glycocalyx damage (EGD) biomarker expression over time in Emergency Department (ED) patients with infection and trauma. This may be explained by varying levels of inflammation or endothelial activation.
Objectives: To explore associations of three EGD biomarkers with inflammatory and endothelial activation biomarkers in patients with infection or trauma.
Methods: Banked serum samples were used from patients with simple infection (n = 22), sepsis (43), septic shock (45) and trauma (30) collected at: enrolment in the ED (T0), 1 hour (T1), 3 hours (T3) and 12–24 hours (T24). EGD biomarkers included syndecan-1 (SYN1), syndecan-4 (SYN4), and hyaluronan (HA). Inflammatory biomarkers included interleukin-6 (IL6), interleukin-10 (IL10), neutrophil gelatinase-associated lipocalin (NGAL), resistin (RTN) and C-reactive protein (CRP). Endothelial activation biomarkers included intercellular adhesion molecule-1 (ICAM) and vascular cell adhesion molecule-1 (VCAM). Laboratory methods included ELISA and cytometric bead array. Random effects linear regression was used to explore associations over time. Only significant results (P < 0.05) are presented.
Results: SYN1 was positively associated with IL6 (T24 in sepsis, T3 and T24 in trauma). SYN4 was positively associated with IL6 in septic shock (T0, T1 and T3). In contrast, SYN4 was negatively associated with IL6 in simple infection at T24. HA was positively associated with IL6 (T3 in septic shock, T24 in trauma). SYN1 and SYN4 were positively associated with IL10, with no differences between groups or over time. SYN1 and SYN4 were positively associated with RTN and NGAL at multiple time points in the infection groups, however only associations with SYN1 showed change over time. HA was positively associated with RTN at multiple time points (T0 in sepsis, T1, T3 and T24 in septic shock, T3 and T24 in trauma), as well as NGAL (T0, T1 and T24 in sepsis, T24 in septic shock and trauma). SYN4 was positively associated with ICAM at T0, T1 and T3 in trauma, whereas HA had an overall negative association with ICAM and VCAM in trauma.
Conclusions: EGD biomarkers showed positive associations with inflammation in the infection groups, varying with biomarker, time point and illness severity. Associations with inflammation in trauma were later in hospitalisation. Only the trauma group showed an association between EGD and endothelial activation. Further investigation into the mechanism of EGD biomarker release in critically ill patients is required.
HIGH MOBILITY GROUP BOX 1 AS A PRONOCICEPTIVE MEDIATOR THROUGH THE ACTIVATION OF CXCR4 IN MICE WITH ACUTE PANCREATITIS Yuhei Irie1,2, Kota Hoshino1, Yoshito Izutani1, Kentarou Muranishi1, Takeshi Nishida1, Junichi Tanaka1, Astufumi Kawabata2, Mistutoshi Iwaasa1, Hiroyasu Ishikura1. 1Fukuoka University Hospital, Fukuoka, Japan, 2Kindai University School of Pharmacy, Osaka, Japan
Objective: Given recent evidence that high mobility group box 1 (HMGB1), a nuclear protein, is participated in the pain including bladder pain, neuropathic pain. The mechanism for the pain in acute pancreatitis is not known clearly. So we examined the role of HMGB1 derived from macrophage as a mediator in visceral nociceptive processing in acute pancreatic pain in mice.
Methods: Cerulein 50 mg/kg was injected i.p. six times in mice at 1 h intervals and referred hyperalgesia in the upper abdomen was evaluated 30 min after the final dose of cerulein using von Frey filaments. The data are expressed as the total score of responses from 10 challenges with each filament. The establishment of acute pancreatitis was confirmed by measuring plasma amylase activity and pancreatic weight in the mice. HMGB1 and other protein levels in pancreatic tissues were measured using western blotting, respectively. The macrophage increase in cerulein treated pancreatic tissue was confirmed using immunohistochemical analysis.
Results: In conscious mice, repeated doses of cerulean produced pancreatitis accompanied by abdominal hyperalgesia. Pretreatment or posttreatment with anti-HMGB1 neutralizing antibody and recombinant human soluble thrombomodulin (rhsTM), known to delete HMGB1, prevented the referred hyperalgesia, but not the pancreatitis. Furthermore, we examined the effect of macrophage depletion using liposomal clodronate, and found the decrease in the splenic macrophage number by flow cytometry and complete prevention of the referred hyperalgesia accompanying pancreatitis. Immunohistochemical analysis showed the increased number of macrophage in pancreatic tissue after repeated cerulein treatment. Pretreatment or posttreatment of AMD3100, a CXCR4 antagonist, depress the hyperalgesia completely. Pretreatment low weight molecular weight heparin (LMWH), known to inhibit receptor for advanced glycation endproducts (RAGE), or FPS-ZM1, a RAGE antagonist, prevented the referred hyperalgesia, while LPS-RS, a toll-like receptor 4 (TLR4) antagonist, did not prevent. Posttreatment of LMWH showed depressed similar tendency. The protein expression of pancreatic RAGE, CXCL12, CXCR4 agonist and CXCR4 increased after the development of cerulein-induced pancreatitis in mice.
Conclusion: The HMGB1/RAGE or CXCL/XCR4 signaling is considered to be involved in the pancreatic pain accompanying the cerulein-induced pancreatitis, and sequestering/inactivation of HMGB1 with rhsTM and blockade of RAGE with LMWH or CXCR4 with AMD3100 appear to serve as novel therapeutic strategy for treatment of painful pancreatic diseases, especially cerulean induced acute pancreatitis in mice.
METASTATIC DISEASE AFTER TUMOUR RESECTION-THE IMPACT OF ANESTHESIA Edwin Boelke, Christiane Matuschek, Wilfried Budach. Heinrich Heine University, Dusseldorf, Germany
Background: Clarification of the question whether anaesthesia may promote the spread of tumour cells and create local recurrence or metastatic disease during surgery due to side effects.
Patients and Methods: PubMed search using the words general and local anaesthesia, tumour spread, local recurrence, distant metastases, circulating tumour cells and mediator response.
Result: General anaesthesia is able to immunocompromised a patient due to mediator releases (storm) like IL-6, hypovolemia and decrease in body temperature. This could help circulating tumours cells to develop metastatic disease. Local anaesthesia could reduce these effects. However, propofol and Cox-2 inhibitors seem to prevent tumour spread. Blood transfusions are a risk factor due to immunosuppression and helps circulating tumour cells to be active tumours. However, the data situation is still very controversial and there are conflicting trials in pubmed.
Conclusion: There is evidence that general anaesthesia may promote tumour spread compared to local anaesthesia. We need prospective randomized multicenter trials in order to clarify the question if we need new approaches to minimize tumour spread with gentle aesthetic procedures for oncology patients.
ASSISTED ELECTRO ACUPUNCTURE REDUCES INFLAMMATORY RESPONSE AFTER OPEN HEART SURGERY Sven Asmussen1,2,3, Rene Przkora4, Dirk M. Maybauer2, Filippo Sanfilippo5, Kristofer Jennings2, John F. Fraser3, Marc O. Maybauer2,3,6. 1Department of Anaesthesiology, University Hospital of Ruhr University Bochum, Bochum, Germany, 2University of Texas Medical Branch, Galveston, TX, USA, 3University of Queensland, Australia, 4University of Florida, Gainesville, FL, USA, 5St. George Hospital, London, UK, 6Manchester University Hospitals NHS FT, Manchester Heart Centre at Manchester Royal Infirmary and University of Manchester, Manchester, UK
Objective: Acupuncture treatment is well known in China for over 2,500 years and is used worldwide as analgesic treatment in patients with acute and chronic pain. Previous studies showed that acupuncture might be beneficial as a supplement to general anesthesia (GA). Aim of this study was to perform a systematic review and meta-analysis to assess the level of evidence for the clinical use of electro acupuncture (EA) in addition to GA in regard of the response of the inflammatory system in patients undergoing open-heart surgery.
Methods: The systematic literature search in Medline yielded a total of 220 citations, published between 1965 and December 31st, 2015. No systematic review or meta-analyses on this topic matched our search criteria. Each article of any language was assessed and rated for the methodological quality of the studies, using the recommendation of the Oxford Centre for Evidence Based Medicine (OCEBM).
Results: Three prospective randomized controlled clinical trials of evidence level 2 with a total of 120 patients matched our search criteria. Three pro-inflammatory markers Interleukin 2 (IL-2), IL-6, and tumor necrosis factor alpha (TNFa) where investigated as well as one anti-inflammatory cytokine. Each trial studied three of the four markers. IL-10 was examined by three trials, the remaining markers were studied by two trials each. The results for the pro-inflammatory cytokines IL-6 and TNFa showed a tendency towards lower values in patients received EA+GA vs GA, only TNFa reached significant levels: EA+GA 49 patients, Control 51 patients, Standardized difference in means 0.753, SE 0.284, Variance 0.080, lower limit 0.197, upper limit 1.309, Z-Value 2.656, P-Value 0.008. This result was consistent with significantly higher values of anti-inflammatory IL-10 for EA+GA vs GA: EA+GA 59 patients, Control 61 patients, Standardized difference in means −0.893, SE 0.447, Variance 0.196, lower limit −1. 760, upper limit −0.026, Z-Value −2.018, P-Value 0.044. Whereas the level of the IL-2 presented significantly higher values in EA+GA (P < 0.05).
Conclusions: We conclude that electro acupuncture in combination with general anesthesia can be beneficial in regard of reducing the immune response compared to the sole use of general anesthesia in patients undergoing cardiac surgery. Further prospective investigations are needed to study these effects in regard of length of hospital stay and potential additional economical benefits.
PLASMA ATP/ADENOSINE BALANCE AND THE INFLAMMATORY RESPONSE TO ELECTIVE SURGERY Yuka Sumi1, Akihisa Matsuda2, Marina Yamada3, Yoko Kuroda1, Jingping Zhang4, Wolfgang Junger4, Hiroshi Tanaka1. 1Department of Emergency and Critical Care Medicine, Juntendo University Urayasu Hospital, Chiba, Japan, 2Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 3Department of Shock and Trauma Center/Hokusoh HEMS, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 4Department of Surgery, Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, MA, USA
Background: Previously, we have shown that ATP and adenosine (ADO) play an important role in the activation of neutrophils. In a mouse sepsis model and in patients, ATP is increased and ADO decreased in plasma, suggesting unbalanced ATP and adenosine as a mechanism involved in inflammation. The aim of the current study was to evaluate whether the ATP/ADO balance also defines the inflammatory response in elective surgery patients.
Methods: Arterial blood was drawn from 10 elective surgery patients (70 ± 11 years; 7 male, 3 female; body mass index, BMI: 23 ± 4.1 kg/m2) before and after surgery under general anesthesia. ATP and ADO plasma levels were analyzed with HPLC.
Results: Laparoscopic surgery for colon cancer was performed in 7 patients and laparotomy for gastric cancer was carried out in 3 patients. Mean operation time was 211 ± 58 min (130–306 min). No blood transfusion was required. There was no significant difference in ATP and ADO between laparoscopic surgery and laparotomy patients. There is a positive correlation between BMI and ATP after surgery (r = 0.8667, p < 0.01). Neutrophil counts after surgery correlated with BMI(r = 0.6727, p < 0.05) and ATP/ADO balance(r = 0.8424, p < 0.01) before surgery and with operation time(r = 0.7416, p < 0.05) (see Figure 1A&B).
Conclusions: The correlation between ATP and BMI suggests that purinergic pathways in visceral adipose tissue may be involved in inflammation. Our results also suggest that the plasma ATP/ADO balance contributes to the inflammatory response after surgery.
ADVERSE EFFECTS OF DEP AND NANOMATERIALS ON BACTERIA-INDUCED PULMONARY DISORDER Kentaro Misaki1, Hirohisa Takano2, Hiroaki Kanazawa1, Kenichiro Inoue1. 1School of Nursing, University of Shizuoka, Shizuoka, Japan, 2Department of Environmental Engineering, Graduate School of Engineering, Kyoto University, Kyoto, Japan
There have been few researches about enhancement factors for biological response to endotoxin exposure. On the other hand, previous epidemiologic studies have indicated that the influence of airborne particulate matter (PM) is positively related to prevalence and mortality of infectious respiratory disease, and health effects have been indicated especially about diesel exhaust particles (DEPs), occupying much of PM in large cities, nanoparticles and nanomaterials. In the present study, in order to elucidate environmental aggravating factors in LPS-induced pulmonary disorder, the exacerbating potential of DEP and nanomaterials were evaluated in view of pathological finding, expression level of cytokines and coagulation reaction in an animal disease model.
ICR male mouse (6 weeks of age) were divided into four experimental groups as follow: 1) vehicle group, 2) LPS group, 3) each particle group, and 4) each particle plus LPS group. DEP, carbon nanotubes (single-wall carbon nanotube: SWCNT, and multi-wall carbon nanotube: MWCNT), latex nanomaterials and so on were selected for experiments. The animals were studied 24 h after intratracheal administration with differential cell distribution in bronchoalveolar lavage fluids, pulmonary vascular permeability of water, cytokine levels in the lung tissue supernatants and the sera, parameters related to coagulatory system and so on.
Synergistic exacerbation of pulmonary disorder such as neutrophil infiltration, lung edema and alveolar hemorrhage followed by the increase of expression levels in inflammatory cytokines and chemokines was confirmed in DEP plus LPS group compared with LPS group or DEP group. Meanwhile, nanomaterials enhanced additively and/or synergistically LPS-induced pulmonary inflammation and vascular permeability and aggregated the concomitant presence of systemic inflammatory response and blood coagulopathy. These adverse effects were approximately exerted more considerably for particles with smaller size. Furthermore, inflammatory cytokine levels in lung and sera were associated with adverse effects for pathology, and their enhancing production/release was presumed as the cause of the exacerbation.
There is the possibility that the increase of PM in atmospheric environment aggregates LPS-induced pulmonary disorder. DEP and several kinds of nanomaterials exacerbated LPS-induced pulmonary disorder, and CNT aggregated the associated systemic inflammatory response and circulatory pro-coagulation associated with several cytokine levels.
INTERLEUKIN 15 MIMETICS FOR SEPSIS -DISCOVERY AND DEVELOPMENT OF NEW DRUG BASED ON IN SILICO SIMULATION- Bunsei Yamamoto, Nobuo Watanabe, Nanami Mizushima, Shigeaki Inoue, Sadaki Inokuchi. Department of Emergency and Critical Care Medicine, Tokai University, Kanagawa, Japan
Introduction: Interleukin 15 (IL-15) is a pleiotropic anti-apoptotic cytokine that promotes activation and proliferation of CD8+ T and natural killer (NK) cell. IL-15 reverses age-related exhaustion of CD8+ T and NK cells and improves survival of aged septic mice and patients. Although human recombinant IL-15 has a therapeutic potential for sepsis, it has a limitation of therapeutic application because of its expensiveness, short half -life. Therefore, the purpose of the study is to find and develop IL-15 mimetics using in silico simulation.
Methods: We assessed immunocompetent potency of a total of 8 compounds that had been selected in silico to occupy the peptide binging IL-15 receptor. Mean fluorescence intensity (MFI) of STAT5 phosphorylation, a downstream of IL-15/IL-15 receptor signal transduction pathway, was determined as a screening of IL-15 mimetics in vitro 15 min stimulation of KAI-3 NK cell line with IL-2/15 receptors NK cell line.
Results: Out of the 8 test compounds, two that had been selected in the in silico screening showed a potent of NK cell activation, identifying a possible lead compound for subsequent study. Compound X demonstrated 43% increase, compound Y showed 27% decrease of pSTAT5 compared to IL-15. Other 6 compounds had no elevation of MFI in STAT5 phosphorylation compared to IL-15.
Conclusions: The present study demonstrated the effectiveness of in silico screening and the prospect for the establishment of high-throughput drug screening system. Achievement of our goal could lead to a discovery of IL-15 mimetics to reverse impaired immunocompeteince of T cells in the elderly, leading to a new drug for sepsis.
THE IMPACT OF HYPOXIA AND PENTOXIFYLLINE ON THE iNOs AND MACROPHAGE MIGRATION INHIBITORY FACTOR OF THP-1 DERIVED MACROPHAGE Ji-Min Park1, Sung-Hyuk Choi1,2, Tae-Yeon Lim1, Jun-Young Suh1, Ziang Qiuyu2. 1Department of Emergency Medicine, Korea University Guro Hospital, Seoul, Korea, 2Institute of Trauma, College of Medicine, Korea University, Seoul, Korea
Objective: Many patients admit the emergency department due to the trauma. These patients with massive hemorrhage, respiratory failure, and further that the experience can fall into hypovolemic shock. Thereafter, it will be damaged by a mechanism such as acute respiratory distress syndrome (ARDS), homeostasis maintenance impossible, multiple organ failure (MOF), immune function reduction and over-expression of inflammation and recovery it can become difficult.
Therefore, we conducted an experiment to observation hypoxia that is a damage mechanism of cell unit with respiratory failure and shock that can occur at an early stage of trauma, how they affected in iNOs and macrophage migration inhibitory factor (MIF) as an inflammatory cytokine of macrophage.
Methods: The experiments were performed with PMN cells and THP-1 derived macrophage. First, PMN cells put through normoxic state and hypoxic state (O2 1%) and measured the apoptosis, TLR4 by FACScan. Second, macrophage cells were incubated through hypoxic state (O2 1%) and measured iNOs and MIF by western blotting. Third, PMN cells and macrophage divided into two groups with stimulus of lipopolysaccharide (LPS) and oxygen treated PMN cells or pentoxifylline treated macrophage in hypoxic state.
- In all hypoxic groups, apoptosis decreased when cells were exposed to hypoxia. Compared to the normoxic group, apoptosis decreased in the hypoxic and hyperoxic group in the group stimulated with LPS.
- In the group stimulated with LPS, TLR4 expression inceased significantly in all group. TLR4 expression was higher in the hypoxic group than in the normoxic group, regardless of LPS stimulation.
- In hypoxic group, iNOs expression increased when cells were exposed to hypoxia. In the group stimulated LPS, iNOs expression a little increased in hypoxic condition. Pentoxifylline restored iNOs in LPS stimulated macrophage even though hypoxic condition.
- In hypoxic group, macrophage migration inhibitory factor increased when cells were exposed to hypoxia. In the group stimulated LPS, MIF expression a little decreased in hypoxic condition. Pentoxifylline restored iNOs in LPS stimulated macrophage even though hypoxic condition, though there were not statistical difference.
Conclusions: MIF and iNOs increased in hypoxic state rather than normoxic state. However, PTX restored iNOs in in LPS-stimulated macrophage in the hypoxic group.
This manuscript is not previously published and prepare manuscript according to the instructions published in SHOCK.
THE IMMUNOMODULATORY EFFECTS OF PENTOXYFILLINE IN INFLAMMATION REACTION Tae-Yeon Lim1, Sung-Hyuk Choi1,2, Jong-Hak Park1, Young-Duck Cho1, Jun-Young Suh1. 1Department of Emergency Medicine, Korea University Guro Hospital, Seoul, Korea, 2Institute of Trauma, College of Medicine, Korea University, Seoul, Korea
Objective: Immunity is the state of having sufficient biological defences to avoid infection, trauma, or other unwanted biological invasion. T-cells, macrophage play important role in cell mediated immunity. Pentoxyfilline (PTX) is know that decrease proinflammatory cytokine and TNF-a. However, the effect on immune system was not known well. Aim of this study is to investigate the effect of PTX in inflammation.
Methods: THP-1 derived macrophage were incubated with LPS and/or indicated concentration of PTX for 6hr and wash with PBS to eliminate effect of LPS. In this media, T cells were plated into at trans well plate and co-culture was done at 12hr. The T cell viability was measured by MTT and expression of IL-2 was analyzed by RT-PCR.
Results: PTX inhibit concentration of MIF, TLR4 protein level and mRNA expression of TLR4 in macrophage. However, PTX did not restore in the T cell proliferation with PGE2. In the co-culture study, The T cells viability decreased in the macrophage cells stimulated with LPS. The additional PTX restored the T cells viability. In the same manner, IL-2 expression in the macrophage stimulated with LPS restored in the macrophage cells stimulated with LPS and PTX.
Conclusion: LPS stimulated macrophage cells inhibit the T cell viability in hyperinflammation condition. In this state PTX restore the T cells viability to increase IL-2. PTX influence the cell-cell interaction, therefore, have its immunomodulatory effects.
This manuscript is not previously published and prepare manuscript according to the instructions published in SHOCK.
HIGH-DOSE INTERLEUKIN-18 INDUCES MICE LEYDIG CELL APOPTOSIS VIA THE DEATH RECEPTOR PATHWAY Taketo Inoue1,2, Michiko Ishikawa1, Mikiko Uemura2, Hayato Yamashita2, Yuka Koga2, Makoto Usami2, Joji Kotani1. 1Department of Emergency, Disaster and Critical Care Medicine, Hyogo College of Medicine, Hyogo, Japan, 2Department of Biophysics, Kobe University Graduate School of Health Sciences, Hyogo, Japan
Background: Interleukin (IL)-18, which is produced by germ cells, Leydig cells and resident macrophages, is an indispensable cytokine in the maintenance of the homeostasis in testis. The IL-18 levels are increased by inflammatory stimulation such as lipopolysaccharide (LPS) in mouse testis. We reported that the role of endogenous IL-18 in testicular germ cell apoptosis may shift from pro-apoptotic to anti-apoptotic depending on the inflammatory stage in mice (Inoue T. et al. Reprod. 2015) and that IL-18 may be a key factor in the modulation of testicular conditions during acute inflammation. However, the role of IL-18 on sustentacular cells remains unclear.
Methods: Mouse Leydig cell line TM3 incubated with LPS (200 or 1,000 ng/mL) or recombinant IL-18 (rIL-18) (0.1, 1, 10 and 100 ng/mL). LPS-stimulated samples were collected at 0, 1, 6, 12, 24 and 48 h, and rIL-18-stimulated samples were collected at 12 h after stimulation. Cell viabilities were determined by trypan blue staining. Cleaved caspase-3 (CC-3) and tBID was measured by western blotting to evaluate apoptosis and the pathways. Expressions of Il-18, Il-6, Tnf-α, Tnfr1, Fas, Fasl, Fadd, Bax and Bcl-2 mRNA were analyzed by reverse transcriptional real-time PCR. Statistical analysis was performed using the Tukey-Kramer's post hoc test.
Results: LPS and rIL-18 had no influence on the cell viabilities on TM3. LPS significantly increased Tnf-α and Il-6 expression at 1 h (p < 0.01) and decreased them to the baseline by 6 h. LPS also increased CC-3 expressions at 6 h and the increase was continued until 48 h. The Fas expression were increased at 6 and 12 h (p < 0.05), whereas the tBID expression were unchanged by LPS. However, LPS did not increase the expressions of Il-18 in TM3 cells. High-dose (10 or 100 ng/mL) rIL-18-stimulation induced CC-3, Tnf-α and Fas expressions at 12 h. Bcl-2 expression were significantly increased by low-dose (1 ng/mL) rIL-18-stimulation.
Conclusion: Both LPS and rIL-18 induced Leydig cell apoptosis via the death receptor pathway. In acute inflammation, the high-dose IL-18 may be derived from macrophages and may induce Leydig cell apoptosis. Preventing the overexpression of IL-18 could be a new therapeutic target to prevent orchitis during acute inflammation.
AN OLD HYPNOTIC BROMVALERYLUREA SUPPRESSES LPS-INDUCED PROINFLAMMATORY REACTION IN BV2 CELLS BY INHIBITING P38 MAPK-DEPENDENT PATHWAYS Shun Kawasaki, Naoki Abe, Fumito Otake, Mohammed E. Choudhury, Afsana Islam, Hajime Yano, Junya Tanaka. Department of Molecular and Cellular Physiology, Ehime University Graduate School of Medicine, Ehime, Japan
Bromvalerylurea (BU) is an old hypnotic/sedative, which was originally discovered at the beginning of 20th century and is still sold as an over-the-counter drug mainly in Asian countries including Japan. We have found its marked immunosuppressive effects on lipopolysaccharide (LPS)-treated macrophages and ameliorative effects on rat sepsis model by cecum ligation and puncture. In this study, we investigated the mechanisms underlying immunosuppressive effects of BU using immortalized murine microglial cell line BV2. BU significantly suppressed expression of various proinflammatory mediators and nitric oxide (NO) release induced by LPS. NFkB, JAK-STAT and p38 MAPK (p38) pathways are involved in LPS-stimulated proinflammatory reaction. In BV2 cells, BU did not prevent LPS-induced NFkB translocation into nuclei as well as the NFkB-dependent transcription as revealed by luciferase assay. Therefore, anti-inflammatory actions of BU may not be attributable to the inhibition of NFkB-dependent pathways. By contrast, BU suppressed phosphorylation of mitogen and stress-activated kinase 1 (MSK1) and MAPK-activated protein kinase 2 (MK2) that are the substrates for p38 at 10 or 30 min after LPS stimulation, whereas BU did not significantly suppressed phosphorylation of p38. These results suggest that BU inhibit p38 activity not through the suppression of p38 phosphorylation. BU and p38 inhibitor suppressed iNOS mRNA in 60 min after LPS stimulation, which implicated that BU effects may be attributable to the inihibition of p38 activity. Furthermore, BU suppressed phosphorylation of STAT1 3 h after LPS stimulation, which may be the secondary effect of suppressing p38 MAPK pathways. However, knockdown experiments of JAK1 or STAT1 and experiments using a JAK1 inhibitor resulted in significant suppression of LPS-induced NO release by BV2 cells, suggesting that the secondary suppressive effects of BU may at least partly contribute to its anti-inflammatory actions. We compared the anti-inflammatory effects of BU with that of a synthetic glucocorticoid dexamethasone; both iNOS mRNA expression in a short-term incubation with LPS and NO release 24 h after LPS stimulation were more markedly by BU than dexamethasone. In conclusion, BU suppressed LPS-induced expression of proinflammatory mediators and NO release mainly by inhibiting p38 MAPK-dependent pathways. BU has been used clinically for one century and there are no reports describing BU-associated immune deficiency or abnormality of glucose metabolism. BU may be a strong candidate agent to ameliorate prognosis of inflammatory diseases such as sepsis because of its established safety.
DEVELOPMENT OF A SMALL-MOLECULE IMMUNOSUPPRESSIVE AGENT BASED ON INHIBITION OF HLA-EPITOPE PEPTIDE BINDING Nobuo Watanabe, Yusuke Suzuki, Akinori Hirayama, Sadaki Inokuchi, Shigeaki Inoue. Deptartment of Emergency and Critical Care Medicine, Tokai University School of Medicine, Kanagawa, Japan
Introduction: Human leukocyte antigen (HLA) class II molecules are expressed in antigen-presenting cells and present peptides derived from foreign antigens. In some autoimmune diseases, such as rheumatoid arteritis (RA), HLA presents self-peptides, provoking the proliferation of self-reactive CD4 T cells. Therefore, small-molecule compounds that can interfere with the peptide-HLA binding may have therapeutic potential. In this regard, bilirubin, a heme degradation product, has been suggested to bind to the cleft of HLA molecules. In this study, we addressed the therapeutic potential of inhibition of HLA-peptide binding.
- Using the mixed lymphocyte reaction (MLR), we assessed the inhibitory potency of 12 compounds, including bilirubin derivatives and compounds that had been predicted in silico to occupy the peptide-binding groove of the HLA molecule.
- To establish a high-throughput compound screening system, biotinylated myelin basic protein (MBP) or binding immunoglobulin protein (BiP) was incubated with peripheral blood mononuclear cells (PBMCs) from healthy volunteers and peptide binding to HLA on cells was analyzed by flow cytometry.
- In the MLR, expansion of particular T cell clones can be induced by the interaction of T cells with specific HLA on allogeneic antigen-presenting cells. Out of the 12 test compounds, one that had been selected in the in silico screening showed a potent anti-proliferative activity with relatively low cytotoxicity, identifying a possible lead compound for subsequent study.
- The HLA-DRB1*1501 and -DRB1*0405 molecules, the genes of which have been linked to multiple sclerosis and RA, respectively, have been suggested to present peptides derived from MBP and BiP, respectively. Our FACS analysis confirmed that APCs including B cells and monocytes, but not T cells, in PBMCs from healthy volunteers bearing either of the HLA alleles incorporated the respective biotinylated antigen peptide. Based on these results, the DRB1*1501 and DRB1*0405 alleles were cloned for the generation of stably HLA-expressing cells to develop a high-throughput drug-screening system.
Conclusions: The present study demonstrated the effectiveness of in silico screening and a prospect for the establishment of a high-throughput drug-screening system. Achievement of our goal could lead to novel therapeutics specific for individual autoimmune-diseases.
KINASE DEPENDENT- AND INDEPENDENT FUNCTIONS OF TYROSINE KINASE 2 (TYK2) DURING ENDOTOXEMIA Andrea Poelzl1, Michaela Prchal-Murphy2, Riem Gawish1, Rita Rom1, Caroline Lassnig1,3, Mathias Mueller1,3, Birgit Strobl1. 1Institute of Animal Breeding and Genetics, University of Veterinary Medicine Vienna, Vienna, Austria, 2Institute of Pharmacology and Toxicology, University of Veterinary Medicine Vienna, Vienna, Austria, 3Biomodels Austria, University of Veterinary Medicine Vienna, Vienna, Austria
Initiated by the presence of bacteria and their products (e.g. endotoxins), sepsis still represents a major problem in health care due to high mortality rates. TYK2, a member of the Janus kinase (JAK) family, is a key player in essential signal transduction cascades utilized by various cytokines that are crucial for innate and adaptive immune responses. Previous studies using mice expressing a kinase-inactive version of TYK2 (Tyk2K923E) revealed kinase-independent roles of TYK2 in NK-cell maturation and function. Together with the fact that TYK2 knockout (Tyk2−/−) mice are protected from lipopolysaccharide (LPS)-induced shock, we hypothesized that TYK2 might also exert kinase-independent functions during sepsis.
Impaired kinase activity of TYK2 as well as complete absence of TYK2 improves survival during lethal LPS challenge but, interestingly, Tyk2K923E mice show an intermediate phenotype between Tyk2−/− and wild type mice, suggesting a kinase-independent detrimental role for TYK2 during endotoxemia. Importantly, all genotypes initially respond similar to LPS, as indicated by profound hypothermia and loss of bodyweight.
It is well established that TYK2 kinase activity is essential for the responsiveness of cells to type I interferon (IFN) and in line with this, phosphorylation of the signal transducer and activator of transcription (STAT) 1, downstream of TYK2, is similarly impaired in Tyk2−/− and Tyk2K923E mice. Our data now suggest that in addition, enzymatically active TYK2 is involved in the systemic induction of pro-inflammatory cytokines such as IL-1β, IL-17A, IL-6 and IL-18, as their levels are substantially reduced in the blood of Tyk2−/− and Tyk2K923E mice at 6 h post LPS injection as compared to wild types. In contrast to these kinase-dependent effects, the immediate, local production of pro- and anti-inflammatory cytokines in the peritoneum at 2 h post LPS challenge seems to be partially independent of Tyk2 kinase activity. Mice harbouring Tyk2K923E exhibit higher concentrations of IL-6, IL-10, IL-1β and TNFα in the peritoneum than Tyk2−/− mice. Whether this contributes to the higher sensitivity of Tyk2K923E mice to LPS as compared to full knockout mice is currently investigated.
Taken together, our data suggest a kinase-dependent contribution of TYK2 in driving systemic inflammation and reveal a kinase-independent impact of TYK2 on survival and the immediate, local production of cytokines during endotoxemia.
This project is funded by the Austrian Science Fund (FWF, grants P25642-B22 and SFB-F28).
ENTERAL FEEDING WOULD NOT BE SUITABLE AS THE INITIAL NUTRITION FOR THE CRITICAL PROLONGED STARVATION: A CASE REPORT Takahiro Kawaji, Tomoyuki Nakamura, Yoshitaka Hara, Seiko Hayakawa, Kotaro Kawata, Sohta Uchiyama, Satoshi Komatsu, Akio Yanagi, Saori Takagi, Ken Kastuta, Mina Kasai, Chizuru Yamashita, Junpei Shibata, Hidefumi Komura, Osamu Nishida. Department of Anesthesiology and Critical Care Medicine, Fujita Health University School of Medicine, Aichi, Japan
Recently, enteral nutrition (EN) is used widely in critical care. But malnourished patients sometimes have the gastrointestinal dysfunctions, and can’t absorb nutrients sufficiently. We here report a case of child with a severe malnutrition obliged to change parenteral nutrition (PN) from EN.
A 4-year-old girl was admitted to the ICU of our hospital with prolonged starvation. She weighed only 8 kg with sick appearance and dry skin. Her mother suffered from schizophrenia, and didn’t serve meals for her. We very slowly increased the amount of daily calories by diluted EN, while monitoring energy metabolism with indirect calorimeter under mechanical ventilation for the first several days. After confirmation of not occurring refeeding syndrome, she was extubated. On the other hand watery diarrhea had occurred and was getting worse and worse by increasing EN reaching more than 1L/day at last. Moreover, Staphylococcus epidermidis, not normal intestinal bacteria, was detected from stool culture, which showed her intestinal function had been totally lost. Therefore, on the 14th day, we abandoned continuing EN, and started to increase PN. On the 20th day, the patient's general condition was suddenly deteriorated with pericardial fluids, bilateral pleural effusions, and ascites, followed by cardiopulmonary resuscitation. We diagnosed that the late-onset refeeding syndrome occurred by increasing PN relatively rapidly. Finally, closed management of the nutrition by PN improved the patient's condition. What we have learned through this educational case are follows: We should not stick to EN and should consider PN therapy first in accordance with patient's gastrointestinal function especially long lasting starvation like this case. And we should pay attention to the refeeding syndrome at any stage of increasing nutrition with such a case.
INTERLEUKIN-6 LEVELS ACT AS A DIAGNOSTIC MARKER FOR INFECTION AND A PROGNOSTIC MARKER IN PATIENTS WITH ORGAN DYSFUNCTION IN INTENSIVE CARE UNITS Waka Takahashi, Taka-aki Nakada, Shigeto Oda. Department of Emergency and Critical Care Medicine, Chiba University Graduate School of Medicine, Chiba, Japan
Introduction: There are significant unmet requirements for rapid differential diagnosis of infection in patients admitted to intensive care units. Serum levels of interleukin-6 (IL-6), procalcitonin (PCT), presepsin, and C-reactive protein (CRP) are measured in clinical practice; however, their clinical utility in patients with organ dysfunction has not been tested adequately. Thus, we investigated the diagnostic and prognostic value of IL-6, PCT, presepsin, and CRP in critically ill patients who had organ dysfunction with suspicion of infection.
Methods: In 100 consecutive critically ill patients with organ dysfunction and suspected infection, serum levels of IL-6, PCT, presepsin, and CRP were measured upon suspicion of infection and serially every other day up to 7 days (cohort 1). The primary outcome variable was the presence of infections. The diagnostic value of IL-6 was further tested in cohort 2 (n = 72, case-control matched). The secondary outcome variables were the sequential organ failure assessment (SOFA) score, serum creatinine levels, and 28-day mortality.
Results: Among the four biomarkers, serum IL-6 levels had the highest AUC value of 0.824 (95% confidence interval [CI] 0.735–0.913) for diagnosing infection in critically ill patients with organ dysfunction and suspected infection in cohort 1 (AUC [95% CI] for the other biomarkers: PCT, 0.813 [0.714–0.911]; CRP, 0.764 [0.645–0.883]; presepsin, 0.681 [0.513–0.849]). In cohort 2, the sensitivity and specificity of IL-6 for diagnosing infection were 0.861 and 0.806, respectively. The presepsin levels were significantly correlated with the SOFA score and serum creatinine levels upon suspicion of infection (r>0.5), especially serum creatinine levels in the patients without infection (r = 0.789). Serum IL-6 levels were significant predictors of 28-day mortality. The AUC value of serum IL-6 levels for 28-day mortality increased over time; the serum IL-6 levels on Day 7 had the highest AUC value of 0.883 (95% CI, 0.788–0.978) for 28-day mortality.
Conclusion: Among serum IL-6, PCT, presepsin, and CRP levels, serum IL-6 levels had the highest diagnostic value for infection. They were also significant predictors of 28-day mortality. Hence, they may improve diagnosis of infection and prediction of 28-day mortality in critically ill patients with organ dysfunction.
CASE REPORT: SUCCESSFUL TREATMENT WITH INTRAVENOUS LIPID EMULSION FOR LIFE-THREATENING STATUS EPILEPTICUS FOLLOWING LOCAL ANESTHETIC OVERDOSE Seiko Hayakawa, Tomoyuki Nakamura, Minako Fukushima, Ken Katsuta, Eri Isobe, Ayaka Takeda, Yasuyoshi Yamazoe, Shoko Wakako, Ai Otsuki, Akira Ogawa, Takuya Hinoue, Hirofumi Sumi, Yoshitaka Hara, Osamu Nishida. Department of Anesthesiology and Critical Care Medicine, Fujita Health University School of Medicine, Aichi, Japan
Severe local anesthetic toxicity can lead to death by epilepticus or arrhythmia.
Intravenous infusion of fat emulsion (lipid rescue) can treat lethal toxicosis caused by local anesthesia with a high fat affinity. Here we report a case in which lipid rescue successfully treated ropivacaine toxicosis which was caused by an incorrect connection of the infuser pump. A male patient in his twenties with a body weight of 64 kg, underwent thoracoscopic lung partial resection for spontaneous pneumothorax. An epidural catheter was placed in the extrapleural space during surgery for post-operative analgesia. After returning to the ward, he lost consciousness and exhibited generalized tonic seizures which were unresponsive to diazepam. The patient was intubated with propofol infusion and was admitted to the intensive care unit. This toxicosis was thought to have been caused by an incorrect connection of the balloon-injecter pump which caused a rapid infusion of 200 ml of 0.475% ropivacaine within the hour. The tonic spasms continued after a primary dose of 200 ml of 20% fat emulsion which was given in 90 minutes. With an additional infusion of 500 ml in 30 minutes, the tonic seizures ended and the patient improved consciousness. The patients’ general status was stable on the next day and he was discharged from the intensive care unit. Although serum total cholesterol level was temporarily elevated, the patient was discharged without adverse effects of lipid rescue.
OVARIAN SEROUS CYSTADENOCARCINOMA IN A 29-YEAR OLD POST-TAHBSO FEMALE WITH MAYER-ROKITANSKY-KUSTER-HAUSER SYNDROME, HETEROTAXY SYNDROME WITH POLYSPLENIA AND HYPOTHYROIDISM: A CASE REPORT Renato C. Ong Jr1, Patricio P. Palmes2, Tomasito Sy2. 1Department of Internal Medicine, Adventist Medical Center - Bacolod, Bacolod City, Philippines, 2Department of Internal Medicine, West Visayas State University Medical Center, Iloilo City, Philippines
Background: The incidence of MRKH is 1 in every 4,500 female live births, while Heterotaxy syndrome occurs in 4 in every 1 M live births. There was one reported case of MRKH with Hypothyroidism and one case with Hypoplastic R lung. The incidence of MRKH plus Heterotaxy syndrome with polysplenia is still unknown. MKRH with ovarian tumors revealed 5 reported cases but none of these was consistent with ovarian serous cystadenocarcinoma. There is no documented association of Heterotaxy syndrome with polysplenia and ovarian serous cystadenocarcinoma.
Clinical Presentation: A 29-year old female was admitted for an appendectomy with incidental note of an unresectable retroperitoneal mass. Biopsy showed suppurative appendix and an abscess with granuloma formation. She underwent pelvic laparotomy with bilateral salpingooophorectomy at age 16 for primary amenorrhea and an absent vaginal orifice.
Laboratory Work Up: CBC showed anemia and persistent leukocytosis with absolute neutrophilia. There were low Na+ and K+ levels and remittent azotemia with hypoalbuminemia. TSH was high but T3 was low. Karyotyping bared normal 46, XX karyotype. Chest X-ray revealed narrowed right intercostal spaces; hyperlucent left lung with widened intercostal spaces; shifting of the trachea and mediastinum to the right; and elevated right hemidiaphragm. Chest and abdominal CT scan demonstrated structural alterations consistent with Heterotaxy syndrome; a lobulated, solid mass with areas of hypodensities and punctuate calcifications within the pelvic cavity. 2D-Echocardiography showed a dextro-positioned heart. Hormone panel was consistent with menopausal levels. Immunohistochemistry studies showed inflammatory myofibroblastic tumor. Autopsy revealed ovarian serous cystadenocarcinoma.
Diagnoses: Ovarian Serous Cystadenocarcinoma; Mayer-Rokitansky-Kuster-Hauser syndrome, Atypical; Heterotaxy syndrome with Polysplenia; Hypothyroidism; Chronic kidney disease Stage 4 due to Obstructive nephropathy; Undernutrition; S/P Bilateral salpingooophorectomy, Closure of Inguinal Hernia (1997); S/P Appendectomy, Biopsy of Retroperitoneal tumor, Loop colostomy, Enterocutaneous fistula creation (March 2010).
Treatment Outcome: Anemia was corrected, and intravenous antibiotics were started. Patient was discharged improved, however, eventually succumbed after 5 months.
MITOCHONDRIA MEDIATED LIVER FAILURE UPON SIRS Andrey V. Kozlov, Andras Meszaros, Adelheid Weidinger. L. Boltzmann Institute for Experimental and Clinical Traumatology, Vienna, Austria
Acute Liver failure is accompanied by systemic inflammatory response syndrome (SIRS) of both infectious and non-infectious origins. Liver failure induced by inflammation is often associated with mitochondrial dysfunction; however the mitochondrial mechanism linking SIRS and mitochondria mediated liver failure is still the matter of discussions. Several hypotheses assume the impairment of a specific mitochondrial function such as drop in ATP synthesis, opening of mitochondrial permeability transition pore, specific changes in mitochondrial morphology, impaired Ca2+ uptake, generation mitochondrial reactive oxygen species (mtROS), turnover of mitochondria and misbalance in electron supply to the respiratory chain. Our data suggests that the drop in ATP levels has protective rather than deleterious character. The most critical mitochondrial event occurring upon SIRS is the release of mtROS in cytoplasm, which can activate two specific intracellular signaling cascades. The first is the mtROS mediated activation of NADPH-oxidase in liver macrophages and endothelial cells; the second is the acceleration of the expression of inflammatory genes in hepatocytes. The signaling action of mtROS is strictly controlled in mitochondria at two points, at the site of ROS generation at complexes I and III and at the site of mtROS release in cytoplasm via permeability transition pore. The systems controlling mtROS-signaling at these two points include pro- and anti-inflammatory mediators, nitric oxide, Ca2+ and NADPH-oxidase.
AN INDUCIBLE MICROSOMAL PROSTAGLANDIN E SYNTHASE-1 (MPGES-1) CONTRIBUTES TO HEPATIC ISCHEMIA/REPERFUSION INJURY VIA PGE2/EP4 RECEPTORS Hirotoki Ohkubo1, Yoshiya Ito2, Ken Kojo2, Nobuyuki Nishizawa2, Masahiko Watanabe2, Masataka Majima3, Kagami Miyaji1. 1Department of Cardiovascular Surgery, Kitasato University School of Medicine, Kanagawa, Japan, 2Department of Surgery, Kitasato University School of Medicine, Kanagawa, Japan, 3Department of Pharmacology, Kitasato University School of Medicine, Kanagawa, Japan
Liver injury elicited by ischemia and reperfusion is an important clinical problem after liver resection or transplantation. The administration of PGE2 prevents hepatic I/R injury, while cyclooxygenase (COX)-derived PGE2 aggravates the injury. Because an inducible microsomal prostaglandin E synthase-1 (mPGES-1) acts downstream of COX-2 in the synthesis of PGE2, we examined the role of mPGES-1 in hepatic I/R injury. mPGES-1-knockout mice (mPGES-1−/− mice) or their wild-type counterparts (WT mice) were subjected to 45 min of partial (70%) hepatic ischemia followed by reperfusion. Hepatic I/R was associated with enhanced expression of COX-2 and mPGES1 in WT livers. Compared with WT mice, mPGES-1−/− mice exhibited lower levels of serum ALT and necrotic area during hepatic I/R. The hepatic mRNA levels of IL-1, IL-6, and iNOS in WT mice were enhanced as compared with those in mPGES-1−/− mice. Immunoreactive iNOS was detected in CD68-positive cells (macrophages). The numbers of hepatic Gr-1-positive cells (neutrophils) in WT mice were greater than those in mPGES-1−/− mice, which was accompanied by enhanced hepatic expression of CXCL1 and CXCL2. The levels of reactive oxygen species (ROS) produced by neutrophils in WT mice were greater than those in mPGES-1−/− mice. Among the hepatic mRNA expression of PGE2 receptor subtypes (EP1-4), the EP2 and EP4 receptors in WT livers were up-regulated as compared with mPGES-1−/− livers. Treatment of WT mice with a specific antagonist of EP4, but not EP2 attenuated hepatic I/R injury as indicated by reduced ALT levels and necrotic area. Likewise, EP4-knockout mice exhibited lower levels of serum ALT, necrotic area, the mRNA of IL-1, IL-6, and iNOS, and the numbers of hepatic Gr-1-positive cells, as compared with WT counterparts. These results suggest that mPGES-1 enhances hepatic I/R injury through EP4 receptor signaling. The selective inhibition of mPGES-1 may be an effective therapeutic approach for hepatic I/R injury.
ULINASTATIN AMELIORATED ACUTE KIDNEY INJURY IN SEPSIS VIA INHIBITION OF ENDOTHELIAL INJURY Akihiro Uchida1, Hideshi Okada1, Kodai Suzuki1, Chihiro Takada1, Airi Kageyama1, Yuki Kato1, Ayumi Kuroda1, Yoshiaki Ogawa1, Yasuaki Hotta2, Taku Tanaka1, Shiho Nakano1, Takahiro Yoshida1, Shozo Yoshida1, Nagisa Miyazaki2, Hiroaki Ushikoshi1, Izumi Toyoda1, Genzou Takemura2, Shinji Ogura1. 1Department of Emergency & Disaster Medicine, Gifu University Graduate School of Medicine, Gifu, Japan, 2Internal Medicine, Asahi University School of Dentistry, Gifu, Japan
Introduction: Acute kidney injury (AKI) lead to a number of complications, including metabolic acidosis, high potassium levels, uremia, changes in body fluid balance, and effects to other organ systems. Sepsis was associated with multiple organ failure, including AKI, through vascular endothelial disorders. Ulinastatin is a glycoprotein that acts as a trypsin inhibitor, and can be effective in treatment of sepsis.
Objective: To address that ulinastatin ameliorated AKI in sepsis.
Methods & Results: Ten-week-old C57BL6 male mice were given lipopolysaccharide (LPS, 15 mg/kg) intraperitoneally. Sham mice were injected PBS instead of LPS in same manner. Ulinastatin (10,000 IU/kg) was injected intraperitoneally 3, 6, 9, 12 and 24 hours after LPS injection (n = 15). As control, PBS was injected in same manner (n = 21). Forty-eight hours after LPS administration, the survival ratio was significantly higher in ulinastatin injected group mice compared with control mice (73 vs. 29%, p < 0.05). Although Kim-1, which was AKI marker, existed on urine tubule 48 hours after LPS injection, Kim-1 expression was decreased in ulinastatin injected mice compare with control mice in western blot analysis. In immunohistochemistry analysis, thrombomodulin (TM), which was one of the vascular endothelial injury markers, was expressed on the surface of healthy endothelial cells in glomerulus in sham mice. However, 48 hours after LPS injection, TM expression was decreased in glomerulus and increased on urine tubule. In ulinastatin treated mice, TM expression was increased in glomerulus and decreased on urine tubule compared with control mice. Frozen fracture kidney samples were obtained from these mice, and these samples were observed using scanning electron microscopy (SEM, HITACHI S-4500). In control mice, the destruction of kidney capillaries, with edematous endothelial walls and fibrin deposition were observed 48 hours after LPS injection in ultrastructural analysis using SEM. In ulinastatin treated mice, the destruction of kidney capillaries was attenuated compared with control group.
Conclusion: These results suggest that ulinastatin treatment is a highly effective treatment strategy for endothelial injury in sepsis.
BIOELECTRICAL IMPEDANCE PHASE ANGLE AS AN INDICATOR OF FLUID STATUS IN CRITICALLY ILL PATIENTS Osamu Takasu, Keita Tashiro, Hideaki Uzu, Mikinori Kannae, Mariko Moroki, Tsunehisa Hirayu, Toshio Morita, Masakazu Nabeta, Atsuo Nakamura, Norio Yamashita. Department of Emergency and Critical Care Medicine, Kurume University School of Medicine, Fukuoka, Japan
Background/Purpose: Endothelial hyperpermeability is a frequent complication in critically ill patients. Bioelectrical impedance analysis (BIA) is an established and widely used noninvasive technique to assess not only body composition and nutritional status but also fluid status. In BIA, phase angle (PA) is calculated from resistance and reactance, which reflect cellular stability and water distribution in intracellular and extracellular spaces. However, whether PA can explain the capillary leakage in critically ill patients has not been elucidated. The purpose of this study was to assess the usefulness of PA as a biological parameter in critically ill patients.
Methods: This study was conducted with 29 patients who were admitted to the intensive care unit (ICU; age, 71 ± 17 years; Acute Physiology and Chronic Health Evaluation II [APACHE II] score, 27 ± 9. Of the 29 patients, 26 survived. PA and fluid distribution (extracellular water [ECW] and intracellular water [ICW]) were determined by using a bioelectrical impedance analyzer (Scan 920-II, Malton) on days 1–5 in the ICU. APACHE II score, body weight (BW) in relation to the day 1 value (BW ratio), and fluid status were compared between three groups divided according to the minimum PA value (min-PA) at 50 kHz as follows: low min-PA (n = 10), mid min-PA (n = 10), and high min-PA (n = 9). The relationship between the fluid distribution or the min-PA value and two different endovascular markers, namely plasma syndecan 1 (SDC-1) and soluble vascular endothelial cadherin (sVE-cadherin) levels on days 1, 3, and 5 in the ICU, was analyzed.
Results: APACHE II score and serum IL-6 (Log IL-6) levels were significantly higher in low min-PA group than in the other two groups. Logistic regression analysis revealed significant negative correlations between min-PA and APACHE II score (r = −0.64), BW ratio (r = −0.66), and ECW% (r = −0.89). The ECW in the low min-PA group characteristically increased with the increase in ICW. Although SDC-1 levels were increased especially the low min-PA group, no statistically significant relationship was found between PA and SDC-1 level. Almost all the patients with increased ICW had positive plasma sVE-cadherin on day 3 in the ICU.
Conclusion: Our results suggested that PA is a valuable indicator of fluid status and capillary leakage in critically ill patients. In addition, we hypothesized that lower PA indicates not only capillary leakage but also lower cellular membrane integrity with increased ICW.
PROTECTIVE MECHANICAL VENTILATION IN NOSOCOMIAL PNEUMONIA TO PREVENT ACUTE RESPIRATORY DISTRESS SYNDROME Artem N. Kuzovlev1,2,3, Viktor V. Moroz1,2, Arkady M. Goloubev1,2, Tamara V. Smelaya2,3, Evgeny A. Tishkov3. 1Society for Shock Studies (Russia), Moscow, Russia, 2V.A. Negovsky Research Institute of General Reanimatology, Moscow, Russia, 3Department of Anesthesia and Intensive Care, A.I. Evdokimov Moscow Medical Dental University, Moscow, Russia
Acute respiratory distres syndrome (ARDS) is a common (25–33% incidence) complication of nosocomial pneumonia (NP) in septic patients. Diagnosis and treatment of ARDS in NP is well established while preventive approaches are poorly investigated. Protective mechanical ventilation (MV) is the most probable way to prevent ARDS in NP.
Objective: to evaluate the efficiency of protective MV in preventing ARDS in NP in septic patients.
Materials and Methods: This retrospective study was conducted 2013–2015. During a retrospective analysis patients were split into two groups: 1) protective MV - after NP was diagnosed patients were ventilated in a protective mode (tidal volume (TV) 6–8 ml/kg); 2) standard MV - patients were ventilated with TV 8–10 ml/kg. The incidence of ARDS was analyzed as a primary outcome. Secondary outcomes were: duration of MV, length of ICU stay, and 28-day mortality. Data were statistically analyzed with Statistica 7.0 and were expressed as median (25–75 percentiles IQR). The difference at p < 0.05 was considered significant.
Results: Comparison of oxygenation index (OI) and extravascular lung water index (ELWI) showed expected differences between the groups. Standard MV group displayed a decrease in OI and an increase in ELWI at day 7 versus day 1. From day 3 lung compliance was lower in the standard MV group, which was linked to the ARDS development. From day 3 the peak airway pressure was higher in the standard MV group. From this time the plateau airway pressure was higher in the standard MV group than that in the protective MV group. There were significant differences in the ARDS incidence in the groups: ARDS developed in 6 (15.0%) and 20 (67.1%) NP patients who underwent protective and standard MV, respectively (p = 0.0001, Fisher's exacttest). The NP patients who were ventilated in a protective mode had shorter length of MV (14.8 ± 6.2 days) and ICU stay (19.2 ± 6.0 days) than those in the standard MV group (20.0 ± 6.3 and 23.9 ± 7.7 days). There were differences in mortality rates between the groups: 27.5% in protective MV group vs. 46.7% in standard MV group (p = 0.0046, Fisher's exact test).
Conclusion: Protective mechanical ventilation prevents development of ARDS in NP in patients with surgical abdominal infection, which improves treatment outcomes.
LYOPHILIZED EXTRAXT OF THE LARVA LUCILIA SERICATA FOR THE MANAGEMENT OF CHRONIC WOUNDS: AN UPDATE Edwin Boelke, Christiane Matuschek, Peter Arne Gerber, Norman Hoff. Heinrich Heine University, Dusseldorf, Germany
Background: Chronic wounds are a common problem in the elderly population. The so-called biodebridement or maggot-therapy with the use of living larvae of the greenbottle fly Lucilia sericata is an effective therapy to debride chronic wounds and stimulate the generation of fresh granulation tissue.
Objectives: Recently, we have generated a lyophilized extract of the larva of the greenbottle fly Lucilia sericata. Here, we present the results of the first 10 patients with venous leg ulcers treated with Larveel; Moreover, we analyzed the effect of Larveel on wound healing in an organotypic 3D-skin model (Phenion®).
Methods and Results: Nine patients with venous leg ulcers that did not respond sufficiently to Standard wound management strategies were treated with Larveel. Larveel was resuspended with 2 ml of sodium Chloride solution and directly applied on wound surface at dressing changes (3 x/week) and covered with a non-adhesive dressing. Patients were treated with Larveel for 8 weeks. In 8 patients a significant improvement of the wound status was achieved. In those patients we did not observe any relevant adverse effects of Larveel-therapy. In 2 patients Larveel therapy resulted in an increased wound colonization with Pseudomonas aeruginosa and progression of the ulcer. Interestingly, it has been reported that Pseudomonas aeruginosa may significantly impair the growth of Lucilia sericata, eventually killing the living larva. Accordingly, larval therapy is particularly effective in the management of wounds infected with Gram-positive bacteria, like Staphylococcus aureus, but less so for wounds colonized or infected with Gram-negative bacteria, especially Proteus spp. and Pseudomonas spp. strains. Analysis of an organotypic skin model (Phenion®) demonstrated that Larveel; promoted the healing of standardized artificial wounds in vitro in a dose- and time-dependent manner.
Conclusions: Larveel may be a promising new option for the treatment of chronic wounds.
CHRONIC INFLAMMATION AND HPV INFECTION IN PATIENTS WITH HEAD AND NECK CANCER Edwin Boelke1, Christiane Matuschek1, Marion Schneider2, Martijn van Griensven3, Wilfried Budach1. 1Heinrich Heine University, Dusseldorf, Germany, 2University of Ulm, Ulm, Germany, 3University of Munich, Munich, Germany
Background: The causal relationship between cancer of the cervix uteri and infection with human papillomavirus (HPV) has been adequately studied. There are also indications that there is link between HPV infection and squamous cell carcinoma of the head and neck. The aim of this study was to find out the incidence and clinical outcome of HPV infection in head neck cancer patients in a German population.
Patients and Methods: In a retrospective study, the tumour tissue from 164 patients (110 men, 54 women, 62.8 years + 12.7) with head neck cancer (oropharynx = 65 nasopharynx = 10, larynx = 13, hypopharynx = 15, oral cavity = 33 CUP = 5, Other = 22) were tested for HPV infection and the clinical outcome was investigated. p16 as a surrogate marker for HPV infection was determined. Moreover, risk factors such as nicotine, alcohol abuse, resection margin of the tumour tissue, histology, lymph nodes involvement, extracapsular spread, tumour stage, and the treatment of the tumour were analysed for local control and overall survival.
Results: The incidence of HPV infection in oropharynx-carcinoma patients was 33%. Patients with HPV-positive oropharyngeal carcinomas showed a tendency towards longer survival time, (p = 0.76, HR: 2.42, 95% CI 0.91 - 6.44) compared to HPV-negative tumours. All other parameters were except the tumour stage in the uni and multivariate analysis not significant for local control and overall survival. The reason for this might be the small number of cases in our individual subgroups.
Conclusion: The HPV infection in Germany was lower than expected. Also in our investigation it appears that HPV positive patients have a better clinical outcome than HPV negative patients. Vaccination is important to reduce chronic inflammation and cancer.
NON-OCCLUSIVE MESENTERIC ISCHEMIA: PATHOGENESIS, DIAGNOSTIC AND THERAPEUTIC MODALITIES FROM OUR EXPERIENCES Hiroki Masuda1, Akihisa Matsuda1, Satoshi Matsumoto1, Nobuyuki Sakurazawa1, Marina Yamada1, Masao Miyashita1, Eiji Uchida2. 1Department of Surgery, Nippon Medical School Chiba Hokusoh Hospital, Chiba, Japan, 2Department of Surgery, Nippon Medical School, Tokyo, Japan
Backgrounds: Non-occlusive mesenteric ischemia (NOMI) consists of intestinal ischemia/necrosis in the absence of organic obstruction within the main trunk of the mesenteric artery. It is currently recognized to be caused by vasoconstriction in the setting of splanchnic hypoperfusion and the mortality is extremely high as over 50%. Selective angiography is considered the gold standard for the diagnosis. However, debate regarding the diagnostic modality and the therapy for NOMI exists and the role of surgical intervention is controversial due to the small number of presented cases. The aim of this study is to investigate the clinical characteristics of NOMI and the treatment outcomes.
Methods and Results: From Jan 2000–May 2016, a total of 9 cases were diagnosed as NOMI in our department and included retrospectively in this study. The overall mortality is 33% (3/9). The median age was 72 years (52–80) and the majority was male sex (7/9, 78%). Cardiovascular disease was the most frequent comorbidity (5/9, 56%), and four had history of surgery for these diseases. Various symptoms were identified and 78% (7/9) of patients demonstrated abdominal pain and 33% (3/9) presented shock state. Two of three patients with shock state resulted in death. The laboratory markers including WBC, CRP, and lactate showed wide range values from normal to extremely high and were not predictive for mortality. In the CT imaging from 89% (8/9) of patients, all patients had poor enhancement of small intestine and ascites. Surprisingly, only one case out of five with portal vein gas reached mortality. Angiography and simultaneous arterial infusion of vasodilators was performed in five cases (56%) and four of five cases survived. 89% (8/9) of patients were subjected to surgical intervention and intestinal resection was required in three of eight cases (death in two of three).
Conclusions: Due to the miscellaneous disease phase at presentation, to identify predictive factors for clinical outcomes of NOMI is difficult. Therefore, further intensive clinical and basic studies of this dismal disease are warranted to elucidate the background pathophysiology and to overcome in the future.
A NEW BIOMAKER PRESEPSIN IN THE DIAGNOSIS AND EVALUATION OF SEPSIS Yasushi Suzuki. Department of Critical Care and Disaster Medicine, Iwate Medical University, Iwate, Japan
Sepsis is a critical inflammation in whole body and life-threatening organ dysfunction caused by a dysregulated host response to infection. Sepsis occurs in approximately 25% of patients in ICU, and common conditions with high mortality. CD14 is the receptor of lipopolysaccharide-lipopolysaccharide binding protein (LPS-LBP ) complexes and activates a series of signal transduction pathway and inflammatory cascades. Soluble CD14 or soluble CD14 subtype (presepsin) is the free fragment of a glycoprotein expressed on monocytes and macrophages. Elevated concentration of presepsin was observed in patients with sepsis compared to control group. Moreover the concentration of presepsin was positively correlated with APACHE II score and SOFA score. PATHFAST presepsin (LSI Medience Corporation, Tokyo) is the quantitative and rapid measurement of presepsin which is a chemiluminescent enzyme immunoassay in whole blood or plasma. The determination of the presepsin concentration can be used for diagnosis and prognosis of sepsis and also to monitor the course of the disease and responses to therapeutic interventions. We report several cases with sepsis to be detected presepsin concentration in this presentation.
ROLE OF ENDOTOXIN IN SEPTIC AKI Kent Doi. Department of Emergency and Critical Care Medicine, The University of Tokyo, Tokyo, Japan
Sepsis and acute kidney injury (AKI) are the most common severe complications for critically ill patients in ICUs. The prognosis of patients with both sepsis and AKI are extremely poor and their mortality rate is an unacceptably high. Of note, sepsis and AKI synergistically worsen the outcomes of ICU patients.
Endotoxin, a lipopolysaccharide derived from the outer membrane of gram-negative bacteria, is recognized as a key factor in the sepsis cascade because it triggers a series of inflammatory reactions that lead to organ dysfunction including AKI. It is widely known endotoxin can cause AKI by several different pathways such as alternating renal hemodynamics and direct tubular cell injury. Recently, a basic study demonstrated reduction of urine flow rate in the early phase of endotoxemia by using two-photon inravital microscopy. Decreased urine flow rate was dominantly observed around the renal tubules in which endotoxin accumulated via TLR4-dependent uptake. Another study reported the role of macrophage in endotoxin preconditioning. Endotoxin preconditioning is a preventative treatment against subsequent larger exposures to endotoxin. It is assumed that endotoxin preconditioning downregulates the inflammatory response of immune cells. Small amount of endotoxin injection caused clustering of M2 macrophages around S1 proximal tubules. Genetically engineered animal experiments demonstrated that CD14-expressing macrophages were necessary for tubular protection after preconditioning.
Endotoxin adsorption by direct hemoperfusion with polymyxin B-immobilized fiber column (PMX-DHP) has been widely used for sepsis. Although several promising results have been reported so far, currently available evidence does not strongly support PMX-DHP against sepsis. The Diagnostic Procedure Combination (DPC) database is a Japanese nationwide administrative claim database representing approximately half of all inpatient admissions to acute care hospitals. By using the DPC database, we conducted retrospective analysis with propensity score matching and demonstrated that not post-abdominal surgery septic shock but septic shock complicated with severe dialysis-requiring AKI might benefit from PMX.
In conclusion, endotoxin appears to play an important role in sepsis and septic AKI, however many different aspects of endotoxin need to be considered for application of endotoxin removal treatment in human sepsis.
UNIVERSAL PLATELET PRODUCTS USING iPS CELL TECHNOLOGY Koji Eto. Department of Clinical Application Research, Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto; Department of Innovation Stem Cell Therapy, Chiba University Graduate School of Medicine, Chiba, Japan.
Platelet transfusions are usually administered as 10 units per treatment (2 x 1011 platelets, Japan case; 1 unit/3 x 1011 platelets in the US), and are often required repetitively, for instance, twice or more per week in patients with severe thrombocytopenia. Some of them need HLA matched platelets by alloimmunization-dependent refractoriness. Currently transfusion of HLA-identical platelets is the sole effective measure. However, their short supply is a matter of ongoing concern. In order to overcome this problem, we are attempting to produce an HLA-null platelet transfusion product from human iPS cells (hiPSCs). As for the large scale production of platelets, we have recently established iPSC derived immortalized megakaryocytic cell line (imMKCL) system which shows continuous growth for up to 5-6 months, and also is capable of releasing platelets in vitro (Cell Stem Cell, 2014). Furthermore, a newly developed 8L scale bioreactor along with several novel drugs allowed us to prepare 100 billion platelets with transfusion grade quality. Towards the realization of universal platelet product, the HLA class I elimination is simultaneously considered. Now we combined iPS cell technology and CRISPR/Cas9 gene editing to establish the HLA knockout imMKCLs, and successfully induced HLA-null platelets. Taken together, our technologies indicate that the HLA-null platelet products are on the verge as HLA independent platelet products, feasible and applicable even to transfusion refractoriness
THE POTENTIAL USEFULNESS OF REGIONAL CEREBRAL OXYGEN SATURATION (rSO2) MONITORING FOR BETTER CARDIOPULMONARY CEREBRAL RESUSCITATION (CPCR) AND POST-CARDIAC ARREST CARE Tomohiko Orita. Department of Emergency and Critical Care Medicine, Saiseikai Yokohamashi Tobu Hospital, Yokohama, Japan
The chain of survival of cardiopulmonary resuscitation (CPR) for out-of-hospital cardiac arrest (OHCA) in adults continues to evolve in its goal of improving better clinical and neurological outcomes. Many studies have demonstrated that the spreading of the use of AED, hands-only CPR, and induction of therapeutic hypothermia (TH) for post-cardiac arrest syndrome (PCAS) have all improved clinical and neurological outcomes. Resuscitation science in this real world keeps trying to attain more favorable cerebral resuscitation.
Various methods have been developed in an attempt to realize earlier and more effective CPCR, including the proactive introduction of automatic chest compression devices at the prehospital stage, mobilization of a fast medical response car or helicopter emergency medical service to the field, earlier introduction of TH and the use of extracorporeal CPR (E-CPR) for aggressive resuscitation.
However, it is still unclear whether the chest compression that is recommended by guidelines is effective for cerebral blood flow; whether a depth of >5 cm is really required for all adult OHCA patients; whether the compression site of the chest is really correct; whether the timing of mechanical ventilation is correct; whether the setting of body temperature as a targeted temperature management (TTM) therapy is appropriate for each cases; and whether vast medical and human resources and also medical bills should be invested in a given case in the first place.
What is the best method or strategy as a cerebral resuscitation? How we could know the possibility of cerebral resuscitation during or before beginning or during CPR including E-CPR? Some objective assessment indicators are required for CPCR.
One such potential indicator is rSO2, one of near-infrared spectroscopy (NIRS), in the hyperacute phase of OHCA. rSO2 measurement for OHCA has been reported to be useful by Ito, et al (Resuscitation 2014;85:778–784). Consequently, research of NIRS and rSO2 monitoring is progressing rapidly worldwide in a field of cardiopulmonary resuscitation. Those researches have been raveling and demonstrating some specific potential about some of the aforementioned questions.
In this seminar, I would like to introduce our investigation and the potential usefulness of rSO2 in the field of CPCR. I will present current findings and experiments that outline the potential of rSO2 as an indicator for chest compression monitoring and of return of spontaneous circulation and defibrillation, and that describe rSO2 as an indicator not only in the hyperacute phase of cardiac arrest but also in post-cardiac arrest care (e.g., stratification of TTM).