Hypoxic Preconditioning Inhibits Apoptosis Of MSCs Through Up Regulation Of Pim-1

Objecttives: Upon hypoxic precondition, The expression level of Pim-1, Akt, Bcl-2,Caspase, Bax in MSCs with different culture condition and different processing time length of hypoxic preconditioning, together with Akt, anti-apoptosis and apotosis factors. The ability of anti-apotosis in MSCs were furthered evaluated. In addition, MSCs upon hypoxic preconditioning were injected into the peninfarct zone of myocardial infarction model,survival of MSCs and cardiac function of MI hearts were detected.Methods: In vitro, MSCs were isolated from SD rat by method of differential adhesion and cultivated to the third generation and then were divided into 6 different groups: control group; cell cultured in normal condition; cell cultured in normal medium in hypoxic condition for 6 hours; cell cultured in normal medium in hypoxia condition for 12hours; cell cultured in serum-free medium in hypoxia condition for 12 hours; cell cultured in normal medium in hypoxia condition for 24 hours; cell cultured in serum-free medium in hypoxia condition for 24 hours. After hypoxic precondition, and we detected the expression of Pim-1, Bcl-2, Caspase-3, Bax in each group with q-PCR, and the expression of Pim-1,t-Akt,p-Akt at different time points with Western Bolt in each group. We use flow cytometry analysis to detect the cell apoptosis in each group and use transwell assay to analyze the cell’s migration ability in each group, mitochondrial membrane potential was detected by JC-1 kit. In vivo, 24 SD rats(300g-350g) were divided into three groups randomly and subgelted to the surgery of LAD ligation. One week after the surgery,BMSCs in each group with dil-dye were injected into the infarct zone. One week after the injection, we took out the heart of SD rats in each group and analyze the survival rate of MSCs in infarct area. Besides, we also used echocardiography to observe the curativeeffect of each group.Results: In vitro, q-PCR showed the highest expression of Pim-1 in cells cultured in serum-free medium in hypoxic condition for 12 hours(P<0.05) and the expression of Bcl-2have the same trend with Pim-1 while Caspase-3 and Bax are on the contrary(P<0.05).Western Bolt showed that the expression of t-Akt is irrelevant to hypoxic and medium condition while p-Akt changed consistanty with Pim-1(P<0.05). Flow cytome try analysis and transwell experiment showed that cell’s migration ability and anti-apoptosis ability were enhanced after hypoxic preconditioning. In vivo, the survival rate of MSCs and cardiac functions were highest in hypoxic preconditioning group.Conclusions: Hypoxic preconditioning MSCs can inhibit cell apoptosis and enhance cell migration by activating Akt and up regulating Pim-1 expression; The decrease of mitochondrial membrane potential of MSCs was slowed by hypoxic preconditioning by inducing Pim-1 overexpression; Hypoxic preconditioning increased the survival rate of MSCs by inducing Pim-1 expression and increased the therapeutic effect of myocardial ischemia and enhanced the cardiac function.