Experimental Study Of Transfection Withlentivirus Mediated HGF Gene Suppressesapoptosis Of Vascular Endothelial Cellsin Rabbitclosed Peripheral Vascular Injury

Objective: To reveal the internal relations between HGF and VECs apoptosis pathway,assessed valuation of inhibitory effects of HGF on apoptosis of VECs in theprogress of closed peripheral vascular injury and promoting the repair and angiogenesis,provide a new therapeutic direction for further exploration of closed vascular injury.Methods: This object intends to use the endovascular method to establish the closed peripheral vascular injury animal model,TUNNEL was used to detect the apoptosis index of VECs model,the reliability assessment model.Select the lentivirus as vector mediated HGF gene transfection in rabbit VECs,building Lv-GFP(carrying green fluorescent protein recombinant lentivirus,used to determine the transfection rate and(GFP-Lv-HGF)carrying hepatocyte growth factor gene and green fluorescent protein recombinant lentivirus)two kinds of endothelial cells after transfection by fluorescence observation,the successful expression of transfected cells by GFP microscopy for assessing the effectiveness.45 female New Zealand rabbits were divided into: group A: femoral artery injury GFP-Lv-HGF group(15 rabbits);group B: shares arterial injury Lv-GFP treatment group(15 rabbits);group C: normal control group of 15 rabbits.Group B showed the A,injury of femoral artery,using capillary needles will be successfully transfected by HGF gene in vitro The VECs was injected into the arterial wall and perivascular tissues,respectively in 1,3,5days after operation was taken for each 5 rabbits were isolated from surgical segment of femoral artery.Bax,caspase3,8,9,Ask1,JNK and apoptosis related protein expression was detected by Western blot and immunofluorescence.Results: The model of closed vascular injury was successfully constructed as indicated by severe intimal hyperplasia.The apoptosis rate in model group 1(68.74%)was significantly higher compared with control group(1.02%,P<0.05).VECs were successfully cultivated and transfected with Lenti-GFP-HGF at a transfection efficiency of 80% or higher.Lenti-GFP-HGF induced the proliferation of transfected VECs in a dose-dependent manner,and the proliferation of VECs at day 4 and 5 was significantly higher compared with day 1(P < 0.05).The Fas/Fas L expression in Lenti-GFP-HGF group was significantly reduced compared with control group without transfection(P < 0.05).Conclusion: HGF exhibits anti-apoptotic effects on VECs,which might be mediated through the inhibition of Fas/FasL pathway.Our study may provide a theoretic basis for the application of HGF in the gene therapy of closed peripheral vascular injury.