Ischemia-reperfusion (IR) injury is a major early postoperative complication occurring in up to 20% of lung transplants leading to primary graft failure (1, 2). Primed by exposure to donor brain death–related inflammatory molecules (3) and cold ischemia preservation (4), IR injury is triggered immediately upon reperfusion (5). Mechanisms that cause IR include damage to the alveolar-capillary membrane and neutrophil extravasation, resulting in respiratory failure (6, 7).
Arachidonic acid metabolites and platelet-activating factor (PAF) are putatively involved in the pathogenesis of IR lung injury (8, 9). PAF initiates and amplifies the inflammatory response by promoting pulmonary edema, neutrophil extravasation, expression of adhesion molecules, and cytokine release (10–12). Thromboxanes (TXs) are thought to contribute to vasoconstriction and exacerbate lung edema (13), whereas leukotrienes (LTs) mediate neutrophil chemotaxis and vascular permeability (14, 15).
Phospholipase A2 (PLA2) is a key enzyme that plays an important role in orchestrating the host inflammatory response in lung injury (16). PLA2 enzymes hydrolyze lung surfactant (17), and are reported to up-regulate the expression of cytokines and adhesion molecules (18). The PLA2 family is classified into three main subtypes: secretory PLA2 (sPLA2), cytosolic Ca2+-dependent PLA2 (cPLA2), and intracellular-Ca2+-independent PLA2 (iPLA2) (19). Cytosolic-PLA2 is reported to be the main gatekeeper of the arachidonic acid cascade and also the catalyst of the first step of PAF synthesis (16, 19). Studies on acute lung injury (20) and IR (21, 22) have focused mainly on the sPLA2 form; however, recent reports have highlighted the importance of the cPLA2 in these pathogenic processes (23–26).
Arachidonyl trifluoromethyl ketone (AACOCF3), an analog of arachidonic acid, acts by competing with endogenous phospholipid molecules for the active catalytic site on the PLA2 enzyme. Thus, AACOCF3 inhibits more specifically the cPLA2 than iPLA2 form by forming a stable hemiketal with the active site; however, it exerts no inhibitory effect on the sPLA2 form (27, 28).
In the present study, we hypothesized that cPLA2 activation triggers IR lung injury; therefore, pharmacological interruption of the cPLA2 cascade could attenuate IR lung injury. To test this hypothesis, we chose to use AACOCF3 as an inhibitor of cPLA2 and examined its effects in an isolated rat lung model subjected to 18 h of cold ischemia followed by 2 h of warm reperfusion.
MATERIALS AND METHODS
The experimental protocol conformed to the Guide for the Care and Use of Laboratory Animals was approved by the Institutional Committee for Experimental Research of our University. As the procedure has been reported previously (29), it is briefly outlined here. Male Sprague-Dawley (SD) rats (300 to 350 g) from Sankyo Labo Service, Tokyo, Japan were anesthetized (ketamine hydrochloride 50 mg/kg ip), intubated through a tracheostomy, and mechanically ventilated (volume-limited ventilator, Shinano Ika, Tokyo, Japan) with FiO2 0.95, a TV of 3 ml, a PEEP of 3 cm water, and a rate of 50 breath/min. After an abdominal incision, heparin (1000 U/kg) was injected into the IVC and blood sampled from the abdominal aorta for gasometry (baseline PO2). The main pulmonary artery was cannulated through a midline sternotomy and lungs flushed with 20 ml of low potassium dextram (LPD) solution, harvested, and stored for 18 h at 4°C. Lungs were then mounted in a warm chamber (37°C), ventilated as above, and perfused for 2 h. The rate of perfusion was gradually increased to 8 ml/min in the first 10 min and then kept constant throughout the experiment. Additional heparinized SD rats (1000 U/kg) served as fresh allogeneic blood donors. To avoid excessive hemolysis, heparinized blood was adjusted to a hematocrit of 20% with modified Krebs-Henseleit solution. Drained blood from the lungs was deoxygenated to adjust the PO2 to 40–50 mmHg using a membrane oxygenator device (Senko Ika, Tokyo, Japan) ventilated with 95% N2 + 5% CO2. At the end of reperfusion, lungs were flushed with 20 ml of saline solution.
Three experimental groups were defined (n=6, each): in the vehicle group, lungs were immediately perfused after harvest for 2 h without an ischemic period. In the two ischemic groups, 20 mg/Kg of arachidonyl trifluoromethyl ketone (Cayman Chemical, Ann Arbor, MI) in 0.05% of dimethylsulfoxide (AACOCF3 group) or saline solution + 0.05% dimethylsulfoxide (Control group) was intravenously infused via the external jugular vein 15 min before lung harvest, respectively. Then, lungs were flushed, stored, and reperfused as described above. The utilized dose and timing of the administration of AACOCF3 were applied on the basis of previous reports (30, 31) and preliminary experiments.
Monitoring
During reperfusion, blood samples were evaluated for gas tension at 30, 60, 90, and 120 min (STAT PROFILE pHOx, Nova Biomedical, Waltham, MA). Mean pulmonary artery pressure (mPAP) (LIFE SCOPE 9, NIHON KODEN Co., Tokyo, Japan) and peak airway pressure (pAwP) were recorded continuously.
Lung Wet-to-Dry Weight Ratio
Apical and accessory right lobes were excised and weighed together for the wet weight, desiccated at 70°C for 1 wk and weighed again for the dry weight. The remaining lobes (middle and inferior) were snap frozen and stored at –80°C for further analysis.
Bronchoalveolar Lavage
Left lungs were injected four times with 2.5 ml of 0.01 M phosphate buffer (pH 7.4), with three sets of instillations and withdrawals each. At least 90% of the total injected volume was recovered. The bronchoalveolar lavage fluid (BALF) was centrifuged at 1000 rpm for 10 min and the protein concentration in the BAL supernatant liquid was measured by the bicichonic acid (BCA) method. Results are expressed as mg/ml.
Myeloperoxidase (MPO) Activity
To assess the degree of neutrophil extravasation (30), lungs were homogenized with 0.5% hexadecyltrimethylammonium bromide (HTAB) in 50 mM phosphate buffer (pH 6.0) and centrifuged at 17,000 g for 20 min. The pellet was resuspended in HTAB, freeze-thawed (20 min at –80°C), homogenized for 60 s, sonicated three times for 30 s, and centrifuged at 17,000 g for 20 min. The supernatant fluid was mixed with 100 mM phosphate buffer (pH 6.0) containing 1 mM of o-dianisidine dihydrochloride and 0.005% of H2O2, and absorbance was read at 450 nm. Results are expressed as OD/mg/min.
Histological Examination
Additional experiments were conducted (n=3, per group). Right lungs were fixed with 10% formalin and left lungs embedded with Tissue-Tek compound (Sakura Finetechnical Co, Tokyo, Japan). Formalin fixed-tissues from the apical, middle, and inferior lobes were sliced into 4-μm-thick sections and processed for H&E stain. Counts of intra-alveolar neutrophils were performed in ten randomly selected high-power fields (HPF) at Ă—1,000 magnification.
Lung PLA2 Activity
The activity of cPLA2 and sPLA2 was determined using a PLA2 assay kit (765021, Cayman Chemical). Lungs were homogenated according to the protocol provided by the manufacturer. To separate the sPLA2 forms, the supernatant fluid was transferred to a cellulose membrane filter device with a molecular weight cut-off of 30,000 (Microcon YM-30, Millipore, MA) and centrifuged at 14,000 g for 40 min at 4°C. The reaction of cPLA2 and sPLA2 with arachidonyl thiophosphorylcholine at room temperature during 60 min was determined using Ellman’s reagent and the absorbance read at 405 nm. To avoid any measurement of iPLA2 activity, samples were incubated with 5 μM bromoenol lactone (BEL).
For the determination of the iPLA2 activity, the incubation mixture contained 100 mM Hepes, pH 7.5, 5 mM EDTA, 4 mM Triton X-100, 30% glycerol, 1 mg/ml BSA, and 0.1 mM ATP. Arachidonyl thio-phosphorylcholine (1,5 mM) was used as substrate. Incubation took place for 60 min at 37°C and absorbance was read at 405 nm. Protein was determined as above and results expressed as nmol/mg/min.
Immunohistochemistry
To determine the expression of phosphorylated cPLA2 (active form) and PAF receptor (PAFR) a double or single immunostaining was adopted, respectively. Lung frozen sections (4 μm) were fixed in 1% parafolmaldehyde for 10 min, washed in PBST, and blocked with Protein Block Serum-Free (X0909, DakoCytomation, CA) for 5 min. First antibodies (rabbit polyclonal anti-phospho-cPLA2, Cell Signaling, Beverly, MA or anti-PAF receptor, sc-20732, Santa Cruz, CA) at 1:25 were incubated at room temperature for 2 h. Secondary antibody (anti-rabbit TRITC- or FITC-conjugated from DakoCytomation, respectively) at 1:20 was incubated for 1 h. Slides for cPLA2 detection were further subjected to antigen retrieval in 10 mM citrate buffer (pH 6.0) for 10 min at 100°C and reincubated with goat polyclonal CD68 from Santa Cruz during 1 hr and FITC-conjugated for 30 min at the same dilutions as above, for alveolar macrophage identification. Rabbit serum without primary rat antibody was used as negative control. Mounted slices (S3023, DakoCytomation) were visualized within 2 h on a Zeiss Axiophot microscope (Optikon/Photometrics).
Leukotriene and Thromboxane Measurement
The LTB4 and TXB2 levels in BALF were determined by enzyme immunoassay (EIA) kit (Cayman Chemical). The detection limit of the EIA assay for LTB4 and TXB2 was 3.9 and 11.0 pg/ml, respectively.
iPLA2 Pharmacological Intervention
To determine if the reported weak inhibitory effect of AACOCF3 on the iPLA2 form affects IR lung injury, 5 mg/kg of bromoenol lactone from Cayman Chemical (BEL group), a specific iPLA2 inhibitor, was i.v. administered 15 min before lung harvest in an additional group of rats (n=6). Then, lungs were preserved and reperfused as outlined. At the end of reperfusion, the lung oxygenation capacity (PO2), wet-to-dry weight ratio, total proteins in BALF, and MPO activity were determined as above. The utilized dose was applied on the basis of preliminary experiments.
Data Analysis
Data are expressed as the mean ± SD. Lung functional data (PO2, pAwP, and PAP) were analyzed by repeated measures of ANOVA and remaining data by one-way ANOVA with a parametric or nonparametric method, as appropriate. If ANOVA showed an overall difference, post-hoc comparisons were performed with the Turkey (parametric data) or Dunn test (nonparametric data). P<0.05 was considered statistically significant.
RESULTS
Physiological Evaluation of the Alveolar-Capillary Membrane Alteration
Before lung harvest, the oxygenation capacity (baseline PO2) was not different among the groups. Throughout reperfusion, the PO2 level in the AACOCF3 group remained similar to the vehicle group, whereas the oxygenation capacity in the control group showed the lowest values compared to the other two groups (P<0.001; Fig. 1A). Despite the higher peak airway pressure and mean pulmonary artery pressure in the ischemic groups, no significant difference between the vehicle and AACOCF3 group was observed. Conversely, the pAwP and mPAP in the control group were increased compared to the other two groups (P<0.05, respectively; Table 1).
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FIGURE 1.:
Effects of cPLA2 inhibition on ischemia-reperfusion lung injury. Two groups of rats were treated 15 min before lung harvest with a cPLA2 inhibitor (AACOCF3 group) or saline (Control group). Lungs were stored for 18 h and reperfused for 2h. Lungs perfused for 2 h without ischemia served as the vehicle group, (n=6 for each group). Results are expressed as mean ± SD. (A) Effect of AACOCF3 on PO2 during reperfusion. Values of PO2 were measured with FiO2=0.95. Baseline PO2 was measured before lung harvest (¶ P<0.001 vs. vehicle and AACOCF3 groups, respectively). (B) Lung wet-to-dry weight ratio after reperfusion (¶ P<0.001 vs. vehicle and AACOCF3 groups, respectively; ‡P<0.05 vs. vehicle group). (C) Total protein content in BAL fluid (BALF) (¶ P<0.001 vs. vehicle and AACOCF3 groups, respectively; ‡P<0.05 vs. vehicle group). (D) Neutrophil infiltration assessed by lung myeloperoxidase (MPO) activity (‡P<0.05 vs. vehicle and AACOCF3 groups, respectively).
TABLE 1: Mean pulmonary artery pressure and peak airway pressure of the lungs during reperfusion
The alveolar-capillary membrane integrity was investigated by the determination of the wet-to-dry weight ratio (an index of pulmonary edema) and total protein in BALF (indicating increased microvascular permeability). Despite the significant decreases in the wet-to-dry ratio and total protein in BALF observed after reperfusion in the AACOCF3 group compared to the control group (P<0.001, respectively), these parameters were increased compared to the vehicle group (P<0.05, respectively; Fig. 1, B–C).
Neutrophil Extravasation and Histological Examination
IR increased the MPO activity in the control compared to AACOCF3 group (1.21±0.34 and 0.41±0.05 OD/mg/min, respectively; P<0.01). No significant difference in MPO activity was observed between the AACOCF3 and the vehicle group (0.38±0.09) (Fig. 1D).
Histological examination revealed an increased number of neutrophil in the intra-alveolar space of control lungs after IR compared to the other two groups (P<0.001, respectively) and increased alveolar thickening, whereas AACOCF3 treatment significantly decreased neutrophil extravasation and preserved the alveolar architecture resembling vehicle lungs (Fig. 2).
FIGURE 2.:
Histological examination. Representative specimens from (A) vehicle, (B) control, and (C) AACOCF3 groups (400Ă—) stained with H&E. Yellow bar, 100 μm. Control lungs appear with an increased inflammatory cell infiltration (head arrow) and alveolar thickening. (D) Counts of intra-alveolar neutrophils (PMN) were performed in 10 randomly selected high-power fields at a magnification of 1,000Ă— (n=3 per group) (¶ P<0.001 vs. vehicle and AACOCF3 groups, respectively; †P<0.05 vs. vehicle group).
Phospholipase A2 Study
After ischemia-reperfusion, we observed a significant and marked increment in the activity of cPLA2 and sPLA2 forms in the control group compared to the other two groups, while the treatment with AACOCF3 maintained the cPLA2 activity at a level similar to that observed in the vehicle group. Of note, AACOCF3 treatment also decreased the sPLA2 activity compared to control (P<0.05), although it was increased compared to vehicle (P<0.01). The iPLA2 activity observed in the control was not significantly different compared to the vehicle group, but increased compared to the AACOCF3 group (P<0.01; Table 2).
TABLE 2: Phospholipase A2 (PLA2) activity in lung homogenates and eicosanoids level in bronchoalveolar lavage fluid
The LTB4 and TXB2 levels in BALF markedly increased in the control group compared to the other two groups, whereas AACOCF3 treatment decreased their levels compared to the control group (P<0.05; Table 2). In addition, immunohistochemistry analysis showed an increased expression of phosphorylated cPLA2 and PAF receptor in control lungs compared to vehicle or AACOCF3-treated lungs. The cPLA2 fluorescent signal was mainly localized in alveolar macrophages, whereas the PAF receptor signal was observed mainly in alveolar cells (Fig. 3).
FIGURE 3.:
IR-induced increases in phosphorylated cPLA2 and PAF receptor protein expression in the lung. Lung sections (4 μm) from the (A, B, E, F) vehicle group, (C, G) control group, and (D, H) AACOCF3 group were processed as follows: (B, C, D) specimens were incubated with polyclonal anti phospho-cPLA2 and anti-rabbit IgG TRITC-conjugated antibodies, and the location is depicted in red. Goat Polyclonal anti-CD68 (a cytoplasmic alveolar macrophage marker) reacts with anti-goat IgG FITC-conjugated antibody, and the location is shown in green. Immunofluorescent images obtained from the above observations were used to generate a pixel overlay of images, signal confluence resulting in the yellow color shown. Yellow bar, 50 μm. (F, G, H) Specimens were incubated with rabbit polyclonal anti-PAFR and anti-rabbit FITC-conjugated antibodies, and the location is depicted in green. Yellow bar, 100 μm. (A, E) Specimens incubated with rabbit serum without primary antibody served as negative control. Control lungs showed an increased fluorescent signal for phosphorylated cPLA2 mainly in macrophage cells (head arrow), and for PAF receptor mainly in alveolar cells (arrow).
Conversely to the effects observed in the AACOCF3 group, after IR no significant difference in the oxygenation capacity, wet-to-dry weight ratio, total proteins in BALF, or MPO activity were observed between the control and BEL group (Fig. 4).
FIGURE 4.:
Effects of iPLA2 inhibition on IR lung injury. An additional group of rats (n=6) was treated 15 min before lung harvest with an iPLA2 inhibitor (BEL group) and lungs were stored and reperfused as outlined. Results are expressed as the mean ± SD (A) Effect of BEL on PO2 at 2 h of reperfusion (¶ P<0.01 vs. control and BEL group, respectively). (B) Lung wet-to-dry weight ratio (†P<0.01 vs. control and BEL group, respectively). (C) Total protein content in BAL fluid (†P<0.01 vs. control and BEL group, respectively). (D) Lung myeloperoxidase (MPO) activity (†P<0.01 vs. control).
DISCUSSION
Lung transplantation has become the mainstay therapy for most end-stage lung diseases (1, 2). Lungs selected for transplantation are generally flushed with a preservation solution and hypothermically stored for as short a period as possible, typically between 4 and 8 hr (1, 6). A new preservation solution developed specifically for the lung combines a low potassium concentration and dextran and has been associated with a reduction in the incidence of primary graft failure. It is reported that the ischemic period with low potassium dextran (LPD) solution has been successfully extended up to 12 hr in clinical transplantation (6) and up to 16 hr in a rodent model (32). Preliminary experiments showed a similar oxygenation capacity in isolated lungs perfused without an ischemic period and lungs preserved up to 12 hr with LPD solution; however, with 18 hr or longer ischemic periods the PO2 dropped significantly (29). Based on this preliminary data, the cold ischemic storage time was set at 18 hr for the present study.
Despite improvements in surgical technique and perioperative care and refinements in lung preservation, IR remains a significant cause of early morbidity and mortality following lung transplantation (2–6), suggesting other factors are involved in the pathogenesis of this process. In this study, we have investigated the effect of cytosolic-PLA2 inhibition on preventing lung injury resulting from prolonged cold ischemia in an isolated rat model. The results of the present study show that cPLA2 activation is crucial to the pathogenesis of IR lung injury. Pharmacological inhibition of the cPLA2 significantly attenuated IR, probably through a decreased formation of bioactive lipids. Therefore, pharmacological inhibition of the cPLA2 may be an effective treatment of IR lung injury for which no clinical agents are currently available.
Ischemia-reperfusion resulted in impaired gas exchange and increased pulmonary pressure and edema formation, together with increased levels of thromboxane and leukotriene in BALF and expression of PAFR in lung tissue. These findings were significantly attenuated by AACOCF3 treatment. In terms of the mechanism of action, the better preservation of the alveolar-capillary membrane functional integrity prevented the drop in PO2 in the AACOCF3 treated lungs. Consistent with our findings, Bonventre et al. reported an attenuated brain IR injury in cPLA2 knockout mice (25). In addition, Nagase et al. reported in acute lung injury animal models an improved oxygenation and decreased lung injury in cPLA2 knockout mice or through pharmacological inhibition of the cPLA2 cascade (31, 33). The potential mechanism by which cPLA2 mediates IR-induced lung injury include the release of inflammatory lipid mediators. Arachidonate acid metabolites such as leukotrienes and thromboxanes have been shown to increase in the lung during IR (34). Thromboxanes are thought to contribute to the hypertension associated with IR and exacerbate lung edema (13), while leukotrienes are associated with increase vascular permeability and neutrophils chemotaxis (35).
A fair criticism of this study is that it did not directly measure PAF; however, we did investigate the expression of the PAF receptor in lung tissue. PAF initiates and amplifies the inflammatory response by promoting neutrophil activation and extravasation, cytokine release, increased vascular permeability, and consequently, pulmonary edema (10–12). Nagase et al. have reported that PAF receptor knockout mice developed less severe acute lung injury after acid aspiration, whereas the overexpression in transgenic mice exaggerated the injury (12). For lung transplantation, several groups have reported that administration of PAF receptor antagonists reduces IR injury and improves lung function in animal models (36, 37) and clinical studies (8).
Immediately upon reperfusion, a series of molecular pathways are activated, with those dependent on phospholipases being readily stimulated (7). PLA2 is a key enzyme that plays an important role in orchestrating the host inflammatory response in lung injury through the synthesis of lipid mediators, and is regulated by Ca2+ and phosphorylation at serine residues (16, 19). In our study, the results showed not only an increased activity of the cPLA2 located mainly in alveolar macrophage cells, but also in sPLA2 activity, together with an increased LTs and TXs formation after IR. Of note, AACOCF3 treatment was able to reduce not only the activity of cPLA2, but also the activity of sPLA2, leading to decreased eicosanoids formation. Consistent with our findings, Nakos et al. reported an increased expression and activity of both the cPLA2 and sPLA2 forms in BALF in patients with acute respiratory distress syndrome, together with a positive correlation between mortality and PLA2 activity (23). An interesting feature of the arachidonic acid release process is that cross talk appears to take place between the PLA2 forms. Thus in vitro, the stimulation of macrophages promotes an immediate arachidonate mobilization, where cPLA2 activation precedes and appears to be required for the subsequent action of sPLA2 (38, 39). Therefore, since no inhibitory effect on sPLA2 has been observed with AACOCF3, these results indicate that cPLA2 activation may regulate, at least in part, the activity of sPLA2 in the lung upon ischemia-reperfusion challenge, although the underlying mechanisms remain to be elucidated.
Although iPLA2 is considered a housekeeping enzyme involved in the maintenance of membrane phospholipid composition (16), recent studies have demonstrated that iPLA2 activation is important in IR myocardial injury (40). In our study, we observed no increased activity of the iPLA2 form after reperfusion and our pharmacological intervention with BEL (an iPLA2 inhibitor) failed to attenuate IR-induced lung injury. Taken together, these results along with those of other investigators suggest that the relative contribution of the PLA2 forms to inflammation might differ between tissue and cell types. In the lung, it appears that cPLA2 and sPLA2 are the main forms orchestrating the inflammatory response in lung injury, in which cPLA2 seems to play an important regulatory role.
Alveolar macrophages are reported to be the primary source of PLA2 via local production and secretion (16, 19). Their importance in IR lung injury has been recently identified (41). In lung transplantation, alveolar macrophages are activated in the early phase of reperfusion as stimulated resident cells, which secrete several proinflammatory substances (42). Consistent with the reported literature, using a double immunostaining technique we observed an increased expression of the phosphorylated cPLA2 mainly in alveolar macrophages after IR, indicating the central role of these cells in the regulation of the PLA2 cascade.
A critical step in the amplification and perpetuation of IR-induced injury is the interaction between neutrophils and the endothelium, which progressively infiltrate the transplanted lung after reperfusion (6, 7, 42, 43). Our results show that treatment with AACOCF3 decreased neutrophil extravasation, LTB4 formation, and PAFR expression. Neutrophil infiltration was confirmed by MPO activity assay and histological examination. In the pulmonary vasculature, it has been observed that LTB4 is more potent than PAF in eliciting chemoattractant response of PMNs (14). Recent evidence using transgenic mice has revealed LTB4 to be an important mediator of neutrophil induced lung injury (15). It is also known that PAF is rapidly produced and displayed on the surface of stimulated endothelial cells and acts as a juxtacrine signal for the activation and adhesion of neutrophils (10, 12). Therefore, our results suggest the major mediator of PMN infiltration is a phospholipase A2 product, where LTB4 and PAF may act synergically eliciting this phenomenon.
In the present study, we observed that cPLA2 inhibition improves lung function and decreases IR-induced lung injury. However, IR-induced increases in edema formation, protein leakage, sPLA2 activity, and eicosanoids formation were significantly attenuated but not eliminated by AACOCF3 treatment. These observations indicate that factors other than cPLA2 may also play a role and contribute to pathophysiology of IR, such factors as sPLA2, oxygen radicals, adhesion molecules, and cytokines (6, 7). Although studies in animal models have shown that sPLA2 plays an important role in the pathogenesis of acute lung injury and IR (20, 22), clinical studies using specific sPLA2 inhibitor have failed to improve survival in patients with severe lung injury (44); therefore, the current study could be a clue to improve management of lung injury induced by IR.
Finally, we acknowledge certain limitations and weaknesses in this experimental model and investigation. The model is an ex vivo model with the inherent limitations of not being a physiological system. Therefore, for clinical application, the effect of cPLA2 inhibition observed in the present study should be restudied in an in vivo lung transplantation model. The current observations suggest that cPLA2 products are involved in the pathogenesis IR-induced lung injury; therefore inhibition of cPLA2-initiated pathways might provide a novel therapeutic approach to improve the outcome of lung transplantation.
In summary, our findings point to an important role for cPLA2 in the development of lung injury induced by IR probably through formation of the pathologic downstream lipid mediators. Moreover, pharmacological intervention of the cPLA2 pathway improves gas exchange capacity and attenuates IR-induced lung injury. These results open new therapeutic strategy for the management of IR in the early postoperative period after lung transplantation.
ACKNOWLEDGMENTS
We thank Dr. Motoaki Bessho and Miss Chiharu Saito for technical assistance and Dr. Kevin Boru for editorial assistance.
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