Preliminary Endothelial Cell Membrane Microparticles (EMPs) On Endothelial Cell Function And Apoptosis In Vitro

Background:microcirculation disorder,hypercoagulation state and microvascular occlusion are important pathways in the pathogenesis of Avascular Necrosis of Femoral Head(ANFH).Endothelial microparticles(EMPs) are phospholipid vesicles derived from endothelial cells.It has been reported that EMPs can influence plasma coagulation system and vasodilatation.Also,they are implicated in inflammation and vascular thrombosis associated with endothelial injury.Moreover,it has been shown that EMPs are involved in cell signaling,apoptosis and recruitment of inflammatory cells.Object:To investigate the effect of glucocortoid on EMPs releasing in vitro. To investigate the effect of EMPs on HUVECs,including endothelial function and apoptosis and to study the contribution of EMPs on ANFH.Material and methods:Object:Human Umbilical Vein Endothelial Cells(HUVECs) in vitro(EAhy926).Stimulator:dexamethasone(DEX) and EMPs released by HUVECs.Method:After cultured with EMps,HUVECs were examined under microscope, and then Mps were counted using flowcytometry.The value of bovine serum albumin was determined by Bradford method to evaluate the permeability of HUVECs. Cellular apoptosis were examined using Caspase-3 and Annexin-V FITC kit.The expression of genes associated with apoptosis were detected by reverses transcription-PCR.Results:Flowcytometry analysis showed that EMPs were released at maximum concentration after 24 hours stimulated by 10mM DEX.Moderate and high concentrations of EMPs caused injury to the permeability of HUVECs,while low concentration of EMP didn't.All concentrations of EMPs induced higher activity of Caspase-3 and PS than control did.Moderate and high concentration groups showed increased expression of APAF-1,AIF gene in comparison with control,while low concentration group didn't.Conclusion:DEX(10mM,24h) stimulated HUVECs to release EMPs at maximum concentration.EMPs at the concentration of 7.66*104 and 7.66*103/ml caused injury to the permeability of HUVECs,while 7.66*102/ml did not show significant difference from control.All three concentrations of EMPs induced HUVECs apoptosis with increased activity of Caspase-3 and PS.EMPs at the concentration of 7.66*104 and 7.66*103/ml showed increased expression of APAF-1,AIF gene in comparison with control,while low concentration group didn't.