Insulin Protects Cardiac Microvascular Endothelial Cells Against Ischemia/reperfusion Injury： The Key Role Of Survivin

BackgroundRestoration of blood flow to the ischemic heart may paradoxicallyexacerbate tissue injury, which is defined as ischemia/reperfusion injury (I/RI). Akey to pathogenesis of I/RI is dysfunction of endothelial coronary microvessels,within which cardiac microvascular endothelial cells (CMECs) play an importantrole. The apoptosis of CMECs precedes than myocytes during I/RI and maypromote myocyte apoptosis. Our previous study has demonstrated that insulinexert protective effect of myocardial cells against I/RI through up-regulating ofsurvivin (SVV) expression. However, it has not been established whether insulincould protect CMECs against I/RI and whether SVV play a role during protectiveeffect of insulin.The present study was designed to detect the role ofanti-apoptotic protein survivin in the cardioprotective effect of insulin of CMECs against I/RI.MethodsCMECs isolated from the hearts of adult rats were exposed tohypoxia(94％N2,5％CO2,1％O2) for2h followed by3h reoxygenation (95％air,5％CO2). The CMECs were randomized into4groups:(1) control group;(2)simulated ischemia reperfusion (SI/R);(3) simulated ischemia reperfusion+insulin (SI/R+Ins);(4) simulated ischemia reperfusion+Insulin+SVV RNAi(SI/R+Ins+SVV RNAi). The cell viability of CMECs was measured by MTTassay and migration ability of CMECs was detected by cell scratch wound assay.The apoptosis of CMECs was detected by TUNEL method. The expression ofsurvivin protein was analyzed by western blot. Then in order to further detect theroll of SVV in the cardial protective effect of insulin of CMECs against I/RI, weused RNAi to depress the expression of SVV. The CMECs were randomized into3groups:1) Non-trasfected;2) β-actin RNAi;3) SVV RNAi. After transfectingfor48h, each group receives Normal, SI/R and SI/R+Ins. Then apoptosis ofCMECs was detected by TUNEL method.ResultsBoth cell viability and migration ability were impaired after SI/R (P <0.01vs control), and the apoptosis index was increased compared with control group(P <0.01). While administration of insulin during reperfusion dramaticallyattenuate the dysfunction of CMECs, decrease the apoptosis index [(10.68±2.28)%vs (22.92±3.52)%，P<0.01vs SI/R] and up-regulate the expression ofSVV. To further ascertain the role of SVV in insulin induced cardial protectiveeffect, CMECs were transfected by SVV siRNA. While siRNA targeting SVVsignificantly blunted the anti-apoptotic effect of insulin [(24.54±4.27)%vs(10.68±2.28)%，P <0.01vs (SI/R+Ins)]. ConclusionInsulin could exert protective effect of CMECs against ischemia/reperfusioninjury through up-regulating of SVV expression.