Although first considered a benign infection, recent studies have disclosed severe and potentially lethal inflammatory manifestations of COVID-19 in children. To date, the available information point towards an infectious or post-infectious cytokine storm in the pathogenesis of the multisystem inflammatory syndrome associated with COVID-19 (MIS-C) that, in some patients, may present with shock and life-threatening symptoms. Neurologic, gastrointestinal, respiratory, and mucocutaneous symptoms, sometimes resembling Kawasaki disease, are frequent. Here we report the case of a 4-year-old child treated in our Pediatric Intensive Care Unit (PICU) with MIS-C and Kawasaki-like symptoms, who was admitted with prominent neurologic symptoms, and for whom a cytokine storm and decrease in blood brain-derived neurotrophic factor (BDNF) was well documented.
A previously healthy 4-year-old girl was admitted to PICU for a complaint of high fever (39.5°C) for the preceding 6 days (D0), accompanied by vomiting and a non-pruriginous skin rash starting on D1. On D3, she became lethargic, complaining of severe myalgia, and on D4, her parents noticed swelling of the palpebrae, hands and feet. Of notice, parents reported that 4 weeks earlier she had exhibited flu-like symptoms with low-grade fever that did not require medical attention. On admission (D5), she exhibited signs of non-hypotensive shock, with no respiratory distress and SpO2 97% on room air. The physical examination was remarkable for evident somnolence and mental confusion (Glasgow coma score 11), a mild multiforme skin rash on trunk and legs (Fig. 1A, Cl 1-3), cracked lips (Fig. 1A, Cl4) and swelling of palpebrae, hands, and feet. She was intubated for neurologic and hemodynamic support and started on dobutamine. Laboratory screening showed a mild anemia. marked leukocytosis with neutrophilia (white blood cells 30,190/mm3, 87% neutrophils), and thrombocytopenia (84.000/mm3), blood Urea 74 mg/dl, creatinine 0.48 mg/dl, metabolic acidosis (pH 7.2, HCO3 19 mmol/L), and raised levels of CRP (340 mg/L), Ferritin (582 ng/mL), D-Dimer (5580 ng/mL) and Troponin (25,7 pg/mL). A cerebrospinal fluid (CSF) sample obtained on D7 revealed pleocytosis (25 cells, 94% lymphocytes) and increased protein (102 mg/dl). She was started on antibiotics and Acyclovir. Rapid hemodynamic improvement was observed and dobutamine was tapered within 16 hours. A chest CT performed on D6 showed consolidation and ground-glass attenuation in the right lung (Fig. 1B). Head CT was normal. She was given intravenous immunoglobulin (IVIG) 2 g/Kg. In temporal relation with IVIG administration, the fever receded and a progressive improvement on laboratory markers of inflammation was observed. She was extubated on D8 when an echocardiogram showed normal biventricular function and no coronary dilation. She further developed desquamation of face, trunk and limbs (Fig. 1A, Cl5-7). On D17, she was discharged home, fully recovered (Fig. 1C).
SEROLOGIC AND MOLECULAR TESTS
For serologic analysis, IgG and IgM antibodies specific for SARS-CoV-2 were titled using the viral nucleoprotein as the antigen in an enzyme-linked immunosorbent assay (ELISA) test in-house. The antibody neutralization title was measured with the virus neutralization test, performed in Vero-CCL81 cells with the viral strain SARS-CoV-2/SP02/human/2020/BRA (accession number—MT126808.1). The methodology is described in the Supplementary Appendix, Supplemental Digital Content 1, http://links.lww.com/INF/E88.
Quantitative assays of reverse transcription TaqMan [qRT-polymerase chain reaction (PCR)] were performed to detect 17 respiratory viruses, including SARS-CoV-2 RNA, 5 exantematic viruses, in addition to 3 arboviruses, total of 25 viral pathogens. To guarantee the integrity of the RNA and the quality of the sample, the extract was also tested for the presence of the human RNase P gene by qRT-PCR, which showed a robust RNase P cycle threshold value (Ct) 22, 46 demonstrating the integrity of the collected material. The methodology is described in the Supplementary Appendix, Supplemental Digital Content 1, http://links.lww.com/INF/E88.
SERUM ANALYSIS OF PRO AND ANTI-INFLAMMATORY MEDIATORS
The serum levels of interleukins (IL) IL-1β (DY201), IL-1RA (DY280), IL-6 (DY206), IL-10 (DY217B), tumoral necrosis factor-alpha (TNF-α) (DY210) and BDNF (DY248) were measured using the ELISA DuoSet Kits from R&D Systems according to the manufacturer’s recommendations and read in a multi-plate reader SpectraMax i3 (Molecular Devices, San Jose, CA).
Real-time quantitative PCR for SARS-CoV-2, respiratory viruses, herpesvirus 1 and 2, cytomegalovirus, parvovirus B19, measles, and arboviruses obtained by tracheal aspirate on D7 resulted negative for all the 25 viruses tested. SARS-CoV-2, enterovirus, HSV 1/2, and CMV (HHV-5) on CSF, and bacterial cultures of blood and CSF, resulted negative. Repeated RT-PCR for SARS-CoV-2 on D11 and D12 persisted negative (Fig. 1C).
Serologic analysis of IgG specific for SARS-CoV-2 obtained on D8 showed positive by ELISA assay, with high titers of neutralizing antibodies by virus neutralization test assay (1:640), while IgM and IgA resulted negative.
Analysis of blood pro-inflammatory cytokines revealed raised levels of IL-1β (8.9 pg/mL; range 0.3–1.4 pg/mL), IL-6 (81.755 pg/mL; range 0–5 pg/mL), IL-10 (173.6 pg/mL; range 0–5 pg/mL) and TNF-α (607.37 pg/mL; range 5–27 pg/mL), while IL-1RA (1.11 pg/mL, range 100–400 pg/mL) and BDNF (6.9 pg/mL; range 8.36 ± 32690 pg/mL) were markedly decreased.
We report a 4-year-old child with MIS-C presenting with shock, Kawasaki-like and neurologic dysfunction, for whom a cytokine storm and decreased levels of BDNF were well documented. Although the first studies reported a benign course of COVID-19 in children, further reports of a multisystem inflammatory disease raised the concern about severe and potentially lethal complications associated with COVID-19 in children.1
Multiple clinical manifestations have been reported in children with MIS-C. Besides fever, gastrointestinal symptoms are the most frequently reported, occurring in 80%–100% patients.1,2 Dermatologic or mucocutaneous changes and Kawasaki-like features are also frequent. Neurologic symptoms, mainly headache, occur in 30% of patients.1
Despite the reports of radiologic changes, respiratory manifestations are usually mild, with mechanical ventilation being instituted for hemodynamic support rather than for respiratory failure in most cases.2 Cardiac dysfunction can be severe, with increased circulating levels of troponin, brain natriuretic peptide, prohormone BNP, ventricular hypokinesis, and coronary enhancement, dilation or aneurism.3 Patients may present with hypotensive shock and vasoplegia, requiring inotropic and vasoactive agents and, sometimes, extracorporeal membrane oxygenation. Rapid improvement of systolic dysfunction is reported in most studies, raising the possibility that myocardial edema, instead of direct cardiomyocyte damage, might explain the hemodynamic compromise in children with MIS-C.3
Laboratory findings in children with MISC-C include high levels of circulating reactive C protein, IL-6, procalcitonin, ferritin, triglycerides, troponin, liver enzymes, hyponatremia, decreased albumin, neutrophilia, lymphopenia and thrombocytopenia.2 A hypercoagulation state characterized by high D-Dimer and normal fibrinogen levels is also frequent. Thromboembolic events, however, are uncommon. The laboratory findings in our patient characterized a hyperinflammatory response, with marked increase of pro-inflammatory cytokines, reactive C protein, and ferritin. Interestingly, despite high, the serum levels of IL-1β observed in our patient were lower than usually reported for Kawasaki Disease (KD).4 A possible explanation for this finding is that, as demonstrated in other studies, SARS-CoV-2 can suppress the innate immune response, for which IL-1β plays a central role as a trigger cytokine.5 Similarly, IL-1 RA levels were also lower than observed for KD, where the suppression of the anti-inflammatory response is well documented.6 In contrast, IL-6, IL-10 and TNF-α levels were markedly increased. In children, raised IL-6 levels have been reported as a post-infectious response to COVID-19.7 Interleukin-6 is a pivotal pro-inflammatory cytokine in pathogenesis of KD and is an independent risk for the development of coronary lesions and IVIG resistance.8 Likewise, IL-10 levels have been correlated with resistance to IVIG in children with KD, as well as with the severity of COVID-19 in adults.8 Despite its anti-inflammatory properties, IL-10 also plays a role as an immunoregulatory cytokine and can favor pro-inflammatory responses.9 Finally, the pronounced increase in TNF-α levels draws the whole picture of a cytokine storm in our child. TNF-α is a well-established, potent pro-inflammatory cytokine that participates in both the acute and chronic phases of the inflammatory response.10 In viral infections, TNF-α is released by natural killer cells and macrophages. A growing number of studies have reported increased levels of TNF-α in COVID-19 patients, where it has been correlated with worse prognosis.11 We hypothesize that, in COVID-19, the increased levels of TNF-α and IL-6 may reflect a disturbance in the immune response of the natural killer cells, leading to a down-regulation of cytotoxic anti-viral enzymes, such as perforin and granzyme.12
Noticeably, neurologic symptoms were prominent in this child. Aseptic meningoencephalitis, without evidence of current SARS-CoV-2 infection, reinforce the pro-inflammatory CNS response. This finding can be associated with the marked reduced levels of BDNF, a neuroprotective anti-inflammatory cytokine that is found in many brain areas, including the olfactory bulb, hippocampus, hypothalamus, mesencephalon and cortex.13 It is a neuroprotective factor that plays a role in neuronal survival, growth, maturation and neuroplasticity. Decreased levels of BDNF have been well-documented in neurodegenerative diseases. It is possible, therefore, that SARS-CoV-2 dampens BDNF synthesis and release and can be associated with neurologic symptoms observed in patients with COVID-19.
MIS-C is a severe and potentially fatal condition that has been increasingly reported worldwide. The development of a cytokine storm, either during or following SARS-CoV-2 infection, is strongly suspected as the main feature in the pathogenesis of MIS-C. The causal relationship between reduced BDNF and neurologic symptoms in patients with COVID-19, as well as the long-term consequences on neurocognitive function in children, remains to be elucidated and should be further explored.
We thank Luciano Mathsumia Thomazelli from LVCM-USP for technical support at the diagnosis, Dr Adriano Ferreira da Silva, Juliana Bannwart de Andrade, Daniela Crema and Dr Shieh Huei Hsin from the University Hospital, University of São Paulo for their technical support.
1. Feldstein LR, Rose EB, Horwitz SM, et al. Multisystem inflammatory syndrome in U.S. children and adolescents. N Engl J Med. 2020;383:334–346.
2. Toubiana J, Poirault C, Corsia A, et al. Kawasaki-like
multisystem inflammatory syndrome in children during the COVID-19 pandemic
in Paris, France: prospective observational study. BMJ. 2020;369:m2094.
3. Belhadjer Z, Bonnet D. Acute heart failure in multisystem inflammatory syndrome in children (MIS-C
) in the context of global SARS-CoV-2 pandemic
. Circulation. 2020;142:429–436.
4. Porritt RA, Markman JL, Maruyama D, et al. Interleukin-1 beta-mediated sex differences in Kawasaki disease vasculitis development and response to treatment. Arterioscler Thromb Vasc Biol. 2020;40:802–818.
5. McCrindle BW, Manlhiot C. SARS-CoV-2
-related inflammatory multisystem syndrome in children different or shared etiology and pathophysiology as Kawasaki disease? JAMA. 2020;324:246–248.
6. Gambacorta A, Buonsenso D, De Rosa G, et al. Resolution of giant coronary aneurisms in a child with refractory Kawasaki disease treated with Anakinra. Front Pediatr. 2020;8:195.
7. Waltuch T, Gill P, Zinns LE, et al. Features of COVID-19
post-infectious cytokine release syndrome in children presenting to the emergency department. Am J Emerg Med. 2020;20:30403–30404.
8. Wang Y, Qian SY, Yuan Y, et al. Do cytokines
correlate with refractory Kawasaki disease in children? Clin Chim Acta. 2020;506:222–227.
9. Mühl H. Pro-inflammatory signaling by IL-10 and IL-22: bad habit stirred up by interferons? Front Immunol. 2013;4:18.
10. Costela-Ruiz VJ, Illescas-Montes R, Puerta-Puerta JM, et al. SARS-CoV-2
infection: the role of cytokines
disease. Cytokine Growth Factor Rev. 2020 doi:10.1016/j.cytogfr.2020.06.001. Published online June 2, 2020.
11. Go YY, Kim Y, Cheon S, et al. Clinical features of patients infected with 2019 novel Coronavirus in Wuhan, China. Lancet. 2020;395:497–506.
12. Wang J, Jiang M, Chen X, et al. Cytokine storm and leukocyte changes in mild versus severe SARS-CoV-2
infection: review of 3939 COVID-19
patients in China and emerging pathogenesis and therapy concepts. J Leukoc Biol. 2020;108:17–41.
13. Bathina S, Das UN. Brain-derived neurotrophic factor and its clinical implications. Arch Med Sci. 2015;11:1164–1178.