目的：研究转化生长因子-β1（transforming growth factor-β1，TGF-β1）诱导骨髓间充质干细胞（bone marrow-derived mesenchymal stem cells，BMSCs）向肌成纤维细胞分化的机制。方法：采用全骨髓贴壁培养法体外培养小鼠原代BMSCs，将对数生长期的P3~P5代BMSCs作为实验细胞，使用不同浓度的TGF-β1诱导BMSCs向肌成纤维细胞分化，并在此基础上观察加入自由基清除剂N-乙酰半胱氨酸（N-acetylcysteine，NAC）对其分化的影响。采用实时荧光定量PCR及Western blot技术检测BMSCs分化指标α-平滑肌肌动蛋白（α-smooth muscle actin，α-SMA）、Ⅰ 型胶原［collagen α1(Ⅰ)，Col α1(Ⅰ)］和Ⅲ 型胶原［collagen α1(Ⅲ), Col α1(Ⅲ)］的表达情况。使用2’，7’-dichlorohydrofluorescein diacetate（DCFH-DA）预孵育BMSCs 15 min，然后加入TGF-β1处理不同时间，检测TGF-β1刺激下BMSCs中活性氧（reactive oxygen species，ROS）的产生，并使用高内涵方法对BMSCs中产生的ROS进行统计分析。结果：TGF-β1可以剂量依赖地诱导BMSCs向肌成纤维细胞分化，上调α-SMA、Col α1(Ⅰ)和Col α1(Ⅲ)的表达。TGF-β1诱导BMSCs分化的作用可以被NAC阻断。TGF-β1在BMSCs中能够引起ROS的产生，且该过程迅速而短暂，当TGF-β1作用30 min时，其在BMSCs中诱发的ROS达到峰值。结论：TGF-β1通过产生ROS介导BMSCs向肌成纤维细胞分化。

Objective：The aim of this study was to investigate the mechanism underlying transforming growth factor-β1 (TGF-β1) induced differentiation of bone marrowderived mesenchymal stem cells (BMSCs)into myofibroblasts.Methods:Primary mouse BMSCs were isolated from bone marrow by flushing the tibias and femurs of mice, and passage 3 to passage 5 of BMSCs were used in the experiments. BMSCs differentiation into myofibroblast was induced by different doses of TGF-β1. In addition, reactive oxygen species (ROS) inhibitor (N-acetylcysteine, NAC) was added to test its effect on the action of TGF-β1. Expressions of BMSCs differentiation parameters, α-smooth muscle actin (α-SMA), collagen α1(Ⅰ) ［Col α1(Ⅰ)］ and collagen α1(Ⅲ) ［Col α1(Ⅲ)］ were measured by real-time quantitative PCR (RT-qPCR) and Western blot analysis. BMSCs were preloaded for 15 min with 2’, 7’-dichlorohydro-fluorescein diacetate (DCFH-DA), then stimulated with TGF-β1 for different times, and fluorescence of ROS was measured using high content analysis. Results:TGF-β1 stimulated differentiation of BMSCs into myofibroblasts and up-regulated expression of α-SMA, Col α1(Ⅰ) and Col α1(Ⅲ) in a dose-dependent manner, which blocked by ROS inhibitor NAC. In addition, TGF-β1 could induce a significant rapid and transient increase in ROS production in BMSCs, and the effect of TGF-β1 on ROS production was peaked at 30 min.Conclusion:TGF-β1 induced differentiation of BMSCs into myofibroblasts via production of ROS.