The Role And Mechanism Of Lipoxin A4 And Its Receptor In Fat Embolism Syndrome Cell Model

Objective:This study was designed to investigate the role and mechanism of lipoxin A4(LXA4)and lipoxin A4 receptor(ALX)on inflammatory response in fat embolism syndrome at the cellular level.We apply a cellular model inwhich pulmonary microvascular endothelial cells(PMVECs)from rats will be pretreated with a free fatty acid(FFA)mixture.Methods:Pulmonary microvascular endothelial cells(PMVECs)were from Healthy adult male Sprague-Dawley(SD)rats.PMVECs were cultured and identificated.Fat embolism syndrome cell model was established.The changes of pro-inflammatory cytokines of PMVECs were determined after pretreated with FFA by using ELISA and PMVECs viability will be determined by using CCK-8 assay.The express of ALX was tested by immunofluorescence technique and immunohistochemistry.And ALX mRNA was certified by Real-time PCR.After pretreated with LXA4 or Boc-2(ALX antagonist),pro-inflammatory cytokines of PMVECs were determined by using ELISA.Furthermore,we will explore the potential signaling pathway bywhich lipoxin A4 works on FES in PMVECs.By using agonists and antagonistsof p38 MAPK,ERK1/2 MAPK and JNK MAPKrespectively,the phosphorylationof p38 MAPK,ERK1/2 MAPK and JNK MAPKafter lipoxinA4 exposure will be evaluated.Results:1 Pro-inflammatory cytokines of PMVECs including IL-6,IL-10,TNF-a increased significantly after pretreated with 0.5mM FFA 12 and 24 hours compared to the blank control group.2.Immunofluorescence technique and immunohistochemistry verified the express of ALX on PMVECs.3.ALX and ALX mRNA increased significantly in group pretreated with 0.5mM FFA and pretreated with FFA for 12 hours compared to the blank control group.4 LXA4 pretreatment decreased the levels of pro-inflammatory cytokines.And Boc-2(ALX antagonist)reversed this effect.5 P-p38 protein expression was increased significantly compared with control group by western blotting analysis.LXA4 pretreatment decreased p-p38 protein expression.After SB203580 pretreatment,p-p38 protein expression in LXA4+FFA+SB203580 was increased significantly compared to the LXA4+FFA group.Conclusion:1.Activation of inflammatory response was observed on pulmonary microvascular endothelial cells of rats after pretreated with a free fatty acid(FFA)mixture.2.ALX was expressed on pulmonary microvascular endothelial cells.3.LXA4 and ALX,ALX mRNA were increased in fat embolism syndrome cell model.4.Pro-inflammatory aytokines were decreased andinflammatory response was inhibited after pretreated with FFA.Boc-2 reversed this action.5.The mechanism of anti-inflammatory effect in FES of LXA4 may relevant to the p38 MAPK signaling pathway.