Protective effect of human insulin-like growth factor 1 gene transfection on rat skeletal myoblasts with ischemic/reperfusion injury

AIM: To observe the protective effect of human insulin-like growth factor 1(hIGF-1) on rat skeletal myoblasts with ischemic/reperfusion injury.METHODS: Myoblasts were isolated from SD rats,cultured,purified,and transfected with plasmid pLghIGF-1SN or pLgGFPSN.The myoblasts were divided into insulin-like growth factor(IGF) group(myoblasts transfected with pLghIGF-1SN),green fluorescent protein(GFP) group(myoblasts transfected with pLgGFPSN),and control group(untransfected myoblasts).The expression of hIGF-1 in myoblasts was investigated by immunocytochemistry,RT-PCR and ELISA.The proliferation rate of myoblasts 14 days after transfection was detected.To observe the protective effect of IGF-1 gene on skeletal myoblasts with ischemic/reperfusion injury 7 days after transfection,the apoptotic myoblasts were detected by the method of in situ TdT-mediated dUTP nick end labeling(TUNEL).The expression of bax and bcl-2 mRNA was detected by RT-PCR,and the expression of caspase-3 was determined by Western blotting.RESULTS: The expression of hIGF-1 in myoblasts transfected with pLghIGF-1SN was detected by immunocytochemistry,RT-PCR and ELISA,but not in myoblasts transfected by pLgGFPSN and untransfected myoblasts.The proliferation rate of myoblasts in IGF group was higher than that in other groups(P0.05).The results of RT-PCR showed that the expression of bax mRNA significantly decreased and bcl-2 mRNA significantly increased in IGF group compared with GFP group(P0.05).The results of Western blotting showed that the expression of caspase-3 significantly decreased in IGF group compared with GFP group(P0.05).CONCLUSION: The transfection of hIGF-1 gene mediated by a retroviral vector produces a protective effect in rat skeletal myoblasts with ischemic/reperfusion injury.The mechanisms may be associated with down-regulating the expression of Bax and caspase-3 and up-regulating Bcl-2 expression.