黄芩苷对老年人骨髓间充质干细胞增殖能力和凋亡水平的影响及机制

The mechanism and effect of baicalin on the proliferative ability and apoptosis of human bone marrow mesenchymal stem cells in the elderly

目的:探讨黄芩苷对老年人骨髓间充质干细胞(human bone marrow mesenchymal stem cells,hBM-MSCs)增殖能力和凋亡水平的影响及机制。方法:采用全骨髓贴壁法体外分离、培养老年hBM-MSCs。将老年hBM-MSCs用不同浓度的黄芩苷(0、0.5、1、10、50、100、500、1000μM/L)处理3 d,用CCK-8法检测黄芩苷对老年hBM-MSCs的细胞毒性作用,筛选适合的黄芩苷浓度;在适合的浓度范围内(0.5、1、10μM/L),用细胞计数法检测黄芩苷对老年hBM-MSCs增殖能力的影响,并筛选出最适药物浓度。将老年hBM-MSCs分为对照组和黄芩苷组,用Annexin-V和PI染色法检测黄芩苷对老年hBM-MSCs凋亡水平的影响;用Westernblot印迹法检测凋亡相关蛋白caspase 3、bcl-2、Bax蛋白水平的表达,ELISA法检测SOD、MDA的含量。结果:0.5、1、10μM/L黄芩苷对老年hBM-MSCs具有最小毒性,浓度大于50μM/L时,呈现明显细胞毒性表现;与对照组比较,0.5、1μM/L和10μM/L黄芩苷处理的老年hBM-MSCs增殖能力显著提高(P<0.01),1μM/L和10μM/L黄芩苷处理的老年hBM-MSCs增殖能力更强;黄芩苷组细胞Bax、caspase 3的表达水平显著下降,Bcl-2表达升高,MDA含量下降,SOD含量升高(P<0.05)。结论:黄芩苷可提高老年hBM-MSCs的增殖能力,降低凋亡水平,而这些可能是通过抗氧化应激反应实现的。

Objective:To discuss the effect and mechanism of baicalin on proliferation and apoptosis of human bone marrow mesenchymal stem cells(hBM-MSCs)in the elderly.Methods:hBM-MSCs were isolated and cultured by whole bone marrow adherent method.The elderly hBM-MSCs were treated with different concentrations of baicalin(0,0.5,1,10,50,100,500,1000μM/L)for 3 days,then cytotoxicity assay of baicalin on the elderly hBM-MSCs was detected with CCK-8 method,and the appropriate baicalin concentration was screened.In the appropriate concentration range(0.5,1,10μM/L),the effect of baicalin on the proliferative ability of hBM-MSCs in the elderly was detected by cell counting method,and the optimal drug concentration was screened.The elderly hBM-MSCs were divided into control group and baicalin group.Baicalin(1μM/L)was added to the baicalin group.Westernblot was used to detect the expressions of apoptosis-related proteins(caspase 3,Bcl-2 and Bax)and the contents of SOD1 and MOD were detected with ELISA.Results:0.5,1,10μM/L baicalin had the minimum cytotoxicity to hBM-MSCs in the elderly.When the concentration went above 50μM/L,the cell survival rate was significantly lower.The proliferation ability of hBM-MSCs in the elderly treated with 0.5,1μM/L and 10μM/L baicalin was significantly improved compared with that in the 0μM/L baicalin(P<0.01),but the proliferative ability of hBM-MSCs in the elderly treated with 1μM/L and 10μM/L baicalin was higher compared with 0.5μM/L baicalin group(P<0.05),and there was no significant difference between 1μM/L and 10μM/L baicalin group.Compared with the control group,the expression levels of Bax,caspase 3 and MDA content in baicalin group were significantly decreased(P<0.05),while the expression levels of bcl-2 and SOD content were increased(P<0.05).Conclusion:Baicalin can improve the proliferative ability of hBM-MSCs in the elderly and reduce the level of apoptosis,which may be achieved through the antioxidant stress response.