[Gly 14]-humanin Ameliorates High Glucose-induced Apoptosis By Inhibiting The Expression Of Mir-155 In Endothelial Microparticles From Endothelial Cells

Background:Hyperglycemia can cause endothelial cell damage and vascular endothelial dysfunction,which is an important event in the pathogenesis of diabetes.Endothelial microparticles(EMPs)are submicron vesicles that shed from the plasma membrane in response to activation,injury or apoptosis of endothelial cells.They can participate in the progression of diabetic vascular complications by carrying microRNAs(miRNAs).Humanin has an anti-apoptotic effect.This study aims to clarify the function of EMPs and miR-155 in the mechanism of HNG against high glucose(HG)-induced vascular endothelial cell apoptosis.Materials and Methods:Human umbilical vein endothelial cells(HUVECs)were cultured in vitro,The apoptosis model of HUVECs was established with 30mM glucose for 72h,30mM mannitol was used as the osmotic pressure control,and 1 ?M[Glyl4]-Humanin(HNG)was used for drug intervention.EMPs were extracted by ultrahigh-speed centrifugation.The characteristics and level of EMPs were detected by the transmission electron microscope(TEM),nanoparticle tracking analysis(NTA)and confocal laser scanning microscope(CLSM);qRT-PCR was used to detect the expression level of miR-155 in EMPs released by HUVECs;Flow cytometry was used to detect the apoptosis rate of endothelial cells.Results:(1).TEM showed that EMPs presented goblet vesicle clusters,and the results of NTA results showed that most EMPs were between 0.1 and 1 ?m in size.The presence of EMPs could be observed by CLSM,and EMPs could be uptake by endothelial cells.(2).HG promotes the apoptosis rate of endothelial cells(P<0.05),and HNG reduces endothelial cell apoptosis induced by HG(P<0.05).(3).The number of EMPs increased significantly after the treatment of endothelial cells with HG(P<0.05);after HNG intervention,the expression level of EMPs decreased(P<0.05).Mannitol and HNG alone had no effect on the release of EMPs.(4).The results of qRT-PCR showed that compared with the control group,the expression of miR-155 in EMPs released by HUVECs induced by HG increased significantly(P<0.05),and the expression of miR-155 in EMPs decreased after HNG intervention(P<0.05).Mannitol and HNG alone did not effect the expression of miR-155 in EMPs.(5).After transfection with miR-155 inhibitor to inhibit the expression of miR-155 in cells,the apoptosis rate of endothelial cells was significantly reduced compared with that of NC inhibitor(P<0.05).(6).Under the environment of HG,the inhibition of miR-155 in cells promotes the decrease of the apoptosis rate of endothelial cells(P<0.05);After HNG pretreatment,the inhibition of miR-155 in cells augmented the effects of HNG on endothelial cell apoptosis(P<0.05).Conclusion:Our research shows that Humanin can exert anti-high glucose-induced apoptosis of vascular endothelial cells by regulating the expression of miR-155 in EMPs.