目的 探讨髓源性生长因子(MYDGF)对高糖环境中大鼠骨髓间充质干细胞(BMSCs)成骨分化的影响及其分子机制。方法 体外分离、培养大鼠BMSCs,流式细胞仪检测BMSCs表面标记物,碱性磷酸酶(ALP)定量检测BMSCs的成骨状态,RT-qPCR检测OSX、RUNX2和COL-I 的mRNA水平,Western blot检测p-GSK3β和β-catenin的蛋白水平。结果 高糖抑制BMSCs的ALP活性及OSX、RUNX2和COL-I的mRNA表达(P＜0.05),添加MYDGF后该抑制作用得以改善(P＜0.05)。MYDGF能够逆转高糖对p-GSK3β和β-catenin蛋白表达的抑制作用[p-GSK3β:(0.85±0.13)vs.(0.15±0.07),β-catenin:(0.79±0.14)vs. (0.37±0.11);P＜0.05];Wnt信号通路拮抗剂DKK1处理后,MYDGF促进p-GSK3β和β-catenin蛋白表达的作用受到抑制[p-GSK3β:(0.43±0.11)vs.(0.85±0.13),β-catenin:(0.20±0.06)vs. (0.79±0.14);P＜0.05]。结论 MYDGF可能通过激活Wnt/β-catenin信号通路进而促进高糖环境中BMSCs的成骨分化。
Abstract
Objective To examine the effect of myeloid-derived growth factors (MYDGF) on the osteogenic differentiation of rat bone marrow mesenchymal stem cells (BMSCs) in high glucose environment and its molecular mechanism. Methods BMSCs were isolated and cultured in vitro, the surface markers of BMSCs were detected by flow cytometry, the osteogenic status of BMSCs was quantitatively detected by alkaline phosphatase (ALP), and the mRNA levels of OSX, RUNX2 and COL-I were detected by RT-qPCR. The protein levels of p-GSK3β and β-catenin were detected by western blot. Results ALP activity and mRNA expression of OSX, RUNX2 and COL-I in BMSCs were inhibited by high glucose (P＜0.05), and the inhibitory effect was improved after adding MYDGF (P＜0.05). The inhibitory effect of high glucose on the expression of p-GSK3β and β-catenin could be reversed by MYDGF [p-GSK3β:(0.85±0.13) vs. (0.15±0.07), β-catenin: (0.79±0.14) vs. (0.37±0.11); P＜0.05], and the promotion of MYDGF on the expression of p-GSK3β and β-catenin were inhibited after DKK1(the inhibitor of Wnt signaling pathway) treatment [p-GSK3β:(0.43±0.11) vs. (0.85±0.13), β-catenin:(0.20±0.06) vs. (0.79±0.14); P＜0.05]. Conclusions MYDGF may promote the osteogenic differentiation of BMSCs in high glucose environment by activating the Wnt/β-catenin signaling pathway.
关键词
髓源性生长因子 /
骨髓间充质干细胞 /
高糖 /
成骨分化 /
Wnt/β-catenin信号通路
Key words
myeloid-derived growth factor /
bone marrow mesenchymal stem cells /
high glucose /
osteogenic differentiation /
Wnt/β-catenin signaling pathway
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基金
博士后科学基金资助项目(2019T120977)
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