恒磁场抑制缺血缺氧条件下大鼠骨髓间充质干细胞凋亡

Constant Magnetic Field Inhibit Bone Mesenchymal Stem Cells Apoptosis under Hypoxia and Serum Deprivation Condition

目的:研究恒磁场对体外缺血缺氧培养条件下大鼠骨髓间充质干细胞(Bone marrow-derived mesenchymal stem cells,BMSCs)凋亡的影响并探讨其作用机制。方法:采取大鼠骨髓,以密度梯度离心分离出单个核细胞(MNCs),于体外培养并由牛垂体提取物(PEX)诱导扩增传代培养出骨髓间充质干细胞(MSCs)。经形态学和流式细胞仪检测MSCs表面标志物鉴定后,将骨髓间充质干细胞(BMSCs)在缺血缺氧条件下培养,通过TUNEL检测比较不同组别细胞的凋亡率和蛋白印迹法(western blot)来观察细胞中特定蛋白质的变化。结果:①经形态学观察和流式细胞仪检测MSCs表面标志物鉴定,提示骨髓间充质干细胞培养成功。②缺血/缺氧组与缺血/缺氧+磁场组比较,缺血缺氧组的凋亡率显著性增加,Akt磷酸化水平显著上升(P<0.05)。提示恒磁场可以使PI3K(Phosphoinositide-3kinase)/Akt(ProteinkinaseB,PKB)信号通路被激活而抑制凋亡的发生。结论:恒磁场通过激活PI3K/Akt信号通路抑制体外缺血缺氧条件下培养的骨髓间充质干细胞的凋亡。

Objective：To investigate the influence of constant magnetic fields on the apoptosis of marrow-derived mesenchymal stem cells under hypoxia and serum deprivation condition and further to explore its mechanism.Methods：Mononuclear cells were isolated from rat bone marrow by density-gradient centrifugation and then they were cultured on fibronectin-coated plates supplied with bovine pituitary extract to get MSCs.BMSCs were identified by Morphology and Surface molecule marker analyzed by FCM.Then the marrow-derived mesenchymal stem cells were cultured under hypoxia and serum deprivation condition（hypoxia and SD）,the proportion of apoptosis and the level of p-Akt in different groups were detected by tunnel analysis and Western bolt respectively.Results：① The marrow-derived mesenchymal stem cells were successfully obtained by identification of morphology and the MSCs specific surface marker.② The proportion of apoptosis is significantly higher in the group under hypoxia and SD than that of group under constant magnetic fields plus hypoxia and SD.The level of p-Akt upgraded significantly,therefore indicated the activation of PI3K/Akt pathway（P0.05）.Conclusion：Constant magnetic field inhibited the apoptosis of marrow-derived mesenchymal stem cells under hypoxia and SD by activating PI3K/Akt.