黄芪甲苷提高骨髓间充质干细胞缺氧微环境下耐受性的作用机制研究

Effect of Astragaloside IV in Improving the Survivability of Bone Marrow Mesenchymal Stem Cells Under Hypoxia Microenvironment

关键词:  黄芪甲苷  骨髓间充质干细胞  缺氧  增殖  凋亡

Key words:  astragaloside IV  bone marrow mesenchymal stem cells  hypoxia  proliferation  apoptosis

摘要:

目的 研究黄芪甲苷（astragaloside IV，AS-IV）预处理对提高骨髓间充质干细胞（bone marrow mesenchymal stem cells，BMSCs）在体外缺氧环境下的增殖活力及抗凋亡能力的潜在机制。方法 设正常组、模型组和AS-IV不同浓度组；CCK-8法检测AS-IV对BMSCs细胞增殖活力的影响；通过H/SD诱导BMSCs凋亡细胞模型，TUNEL染色法评价各组细胞凋亡率；并采用Western blotting检测凋亡相关蛋白Bcl-xl、caspase-3的表达水平。结果 通过H/SD成功诱导BMSCs凋亡，不同浓度AS-IV干预组与对照组相比细胞增殖生存活力明显提高（P<0.01）；AS-IV干预72 h与干预24 h相比，在AS-IV （1.28×10^–5，2.56×10^–5，5.12×10^–5 mmol·L^–1）组间有显著性差异（P<0.05），AS-IV （1.02×10^–6 mmol·L^–1）组间干预72 h与24 h相比无显著性差异；与模型组相比，AS-IV干预组随着干预浓度的增加细胞凋亡率逐渐下降（P<0.05）；与模型组比较，AS-IV组Bcl-xl表达显著上调、caspase-3表达显著下调（P<0.01），AS-IV+AG490（JAK2阻断剂）组Bcl-xl、caspase-3的表达无显著性差异。结论 AS-IV能促进BMSCs在缺氧环境下增殖能力和抗凋亡能力的提高，其机制可能是通过调控JAK2通路影响Bcl-xl、caspase-3蛋白的表达。

Abstract:

OBJECTIVE To investigate the potential mechanism of astragaloside IV(AS-IV) preconditioning on the proliferation and anti-apoptotic ability of bone marrow mesenchymal stem cells(BMSCs) under hypoxia in vitro. METHODS Normal group, model group and AS-IV different concentration groups were set; CCK-8 method was used to detect the effect of AS-IV on the proliferation activity of BMSCs; apoptosis model of BMSCs induced by H/SD, the apoptosis rate of BMSCs in each group was evaluated by TUNEL staining; Western blotting was used to detect the expression levels of apoptosis-related proteins Bcl-xl and caspase-3. RESULTS Apoptosis of BMSCs was successfully induced by H/SD, and cell proliferation viability of the AS-IV intervention group with different concentrations was significantly improved compared with the control group(P<0.01). There was a significant difference in AS-IV(1.28×10^–5, 2.56×10^–5, 5.12×10^–5mmol·L^–1) between the groups after intervention for 72 h and 24 h(P<0.05). There was no significant difference between AS-IV (1.02×10^–6mmol·L^–1) groups after intervention for 72 h and 24 h. Compared with the model group, with the intervention concentration increased, the apoptosis rate of the AS-IV intervention group decreased gradually(P<0.05). Compared with the model group, Bcl-xl expression was significantly up-regulated and caspase-3 expression was significantly down-regulated in the AS-IV group(P<0.01), while Bcl-xl and caspase-3 expression were not significantly different in the AS-IV+AG490(JAK2 blocker) group. CONCLUSION AS-IV can promote the proliferation and antiapoptosis ability of BMSCs under hypoxic environment, possibly by regulating the JAK2 pathway and affecting the expression of Bcl-xl and caspase-3.