Transforming growth factor-β1 induces differentiation of bone marrow-derived mesenchymal stem cells into myofibroblasts via production of reactive oxygen species / 北京大学学报(医学版)

ABSTRACT

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), collagenα1(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.