miR-21调控人骨髓间充质干细胞成骨分化的研究

摘要

目的:观察miR-21在人骨髓间充质干细胞( hBMMSCs)成骨分化过程中的表达. 方法:通过转染使hBMMSCs过表达或抑制miR-21后,分别采用ALP染色、茜素红染色观察各组ALP活性和钙化结节的表达形成量;并采用Real time RT-PCR、Western blot检测各组成骨相关基因Runxz、osterixmRNA及其蛋白的表达水平,以观察miR-21对hBMMSCs体外成骨分化的调控作用. 将表达不同水平miR-21的各组hBMMSCs与HA/TCP复合后植入裸鼠皮下,8 周后取材,采用HE染色和Masson三色染色观测各组类骨质的形成量.结果:miR-21在hBMMSCs成骨过程中表达量较对照组明显升高(P<0. 05). 过表达miR-21能促进hBMMSCs体外成骨分化及体内的异位成骨能力;而抑制miR-21则能降低hBMMSCs体外成骨分化及体内的异位成骨能力. 结论:miR-21可外调控hBMMSCs成骨分化,在骨形成过程发挥作用.%AIM:To investigate the expression and the role of miR-21 in osteogenesis of human marrow-derived mesenchymal stem cells ( hBMMSCs) . METHODS:The expression level of miR-21 after transfection of syn-thetic per-miR-21 was confirmed by real time RT-PCR. After 2 days of transfection, hBMMSCs were induced to os-teoblast differentiation. ALP and Alizarin red S staining were used to determine osteogenic ability. Real time RT-PCR and western blot were used to analyze the estrogenic marker gene expression. hBMMSCs transfected with pre-miR-21, anti-miR-21 or miR control were loaded on hydroxyapatite-tricalcium phosphate scaffold and implanted s. c. in NOD/SCID mice. 8 weeks after transplantation HE staining and Masson 's trichrome staining were used to observe the new bone formation. RESULTS:miR-21 was increased during osteogenic differentiation of hBMMSCs. Gain-loss-functional analysis showed that up-regulation of miR-21 promoted osteogenic differentiation of hBMMSCs in vitro and in vivo. On the contrary, silencing of miR-21 inhibited the osteogenic differentiation of hBMMSCs in vitro and in vivo. CONCLU-SION:miR-21 can promote osteogenic differentiation of hBMMSCs in vitro and enhance ectopic bone formation in vivo.