二氮嗪对低氧环境下骨髓间充质干细胞增殖活力及凋亡的影响

Diazoxide effects on proliferation and apoptosis of bone marrow mesenchymal stem cells under hypoxic conditions

背景:如何有效干预骨髓间充质干细胞进行缺氧预适应使得其移植后发挥应有的疗效是目前急需解决的问题。目的:探讨抗氧自由基药物Mito-K_(ATP)通道激活剂二氮嗪对低氧环境中小鼠骨髓间充质干细胞增殖活力及凋亡的影响。方法:将购买的小鼠骨髓间充质干细胞分为4组:空白对照组、0.16μmol/L二氮嗪组、0.8μmol/L二氮嗪组、4μmol/L二氮嗪组,各组在二氮嗪干预下放置于体积分数为10%O_2培养箱培养。分别干预1,2,4,6,8 d进行MTT检测细胞增殖活性。干预后第14天进行Hoechst 33258染色,观察细胞凋亡情况。结果与结论:(1)建系的小鼠骨髓间充质干细胞纯度较高,形态均一,但增殖分化能力较弱;(2)不同浓度二氮嗪作用下骨髓间充质干细胞活力差异无显著性意义(P>0.05);(3)空白对照组细胞核浓缩,颜色深蓝,核聚集,发现数个类圆形凋亡小体,二氮嗪组偶发现凋亡小体,3个浓度二氮嗪组间无明显差异,3组均低于空白对照组;(4)一定浓度的抗氧自由基Mito KATP通道激活剂二氮嗪在低氧环境下(体积分数为10%O_2)不影响小鼠骨髓间充质干细胞增殖活力但可减少细胞凋亡。

BACKGROUND:It is an urgent problem to effectively make bone marrow mesenchymal stem cells exert proper effects under hypoxic preconditioning.OBJECTIVE:To investigate the effects of diazoxide,a Mito-KATP channel activator,on the proliferation and apoptosis of mouse BMSCs in hypoxic environment.METHODS:Mouse BMSCs were divided into four groups:blank control group,0.16,0.8,4μmol/L diazoxide groups.Cells intervened by diazoxide were cultured in a 10%O2 incubator.MTT assay was performed to detect cell proliferation at 1,2,4,6,8 days after intervention,and Hoechst 33258 staining was performed to observe cell apoptosis at 14 days after intervention.RESULTS AND CONCLUSION:High homogeneity and purity but low proliferation of BMSCs was found.There was no significant difference in the activity of BMSCs among 0.16,0.8,4μmol/L diazoxide groups(P>0.05).In the blank control group,concentrated nuclei were dark blue in color and aggregated,and several round apoptotic bodies were found.In the diazoxide groups,apoptotic bodies were occasionally found,and no significant difference was found among different diazoxide groups.These findings indicate that a certain concentration of diazoxide can reduce cell apoptosis but has no effects on the proliferation of mouse BMSCs under hypoxic environment(10%O2).