miR-124-3p经Wnt通路因子Axin1调控骨髓间充质干细胞的研究.

Objective To investigate the molecular targeting mechanism of miR-124-3p regulation of BMSCs osteogenic differentiation via factor A (Axin1) by bone marrow mesenchymal stem cells (BMSCs), and its participation in the development of postmenopausal osteoporosis (PMOP). Methods Twenty patients with femoral fractures requiring surgery or exposing the medullary cavity were collected, including 10 postmenopausal women with osteoporosis (PMOP group) and 10 non-osteoporosis postmenopausal women (control group). Bone marrow tissue (3-5 ml) was extracted during the operation. A bone mesenchymal stem cells (BMSCs) cell model was established. Surface marker antigens of BMSCs were detected using flow cytometry. BMSCs were induced to differentiate into osteoblasts. The expression of miR-124-3p was detected using RT-qPCR. The binding sites of A-Axin1 to miR-124-3p was detected with double luciferase reporter gene system. Overexpression vector of miR-124-3p was constructed. miR-124-3p was overexpressed and transfected to PMOP-BMSCs. Osteogenic induction medium was used for osteogenic induction culture. On 7d, 14d, 21d of induction, the cells were collected. Alkaline phosphatase (ALP) and Alizarin red staining were detected. On day 7, RT-qPCR was used to detect Runx2, Osterix, and target factor Axin1 in each group. The relationship between miR-124-3p expression after miR-124-3p transfected in BMSCs and Axin1 was detected using Western blotting. Results ①RT-qPCR was used to detect miR-124-3p expression. The expression in PMOP group was lower than that in control group. ②In cultures of overexpressed transfection and induced osteogenesis of miR-124-3p, the level of alkaline phosphatase and nodules of Alizarin red positive staining increased gradually with the extension of osteogenic induction time. ③RT-qPCR was used to detect the expressions of Runx2 and Osterix on day 7 of osteogenic induction. The expressions of Runx2 and Osterix in BMSC+ osteogenic differentiation group were significantly higher than those in BMSC group, and they were significantly higher in BMSC+ overexpression of miR-124-3p group than in control group. The expression of Axin1 in BMSC+ osteogenic differentiation group was significantly lower than in BMSC group. It was significantly lower in BMSC+ overexpression of miR-124-3p group than in control group. ④Luciferase reporter gene experiments demonstrated the binding of miR-124-3p to Axin1. ⑤Western blotting detection in BMSCs showed that the expression of Axin1 in miR-124-3p overexpressed group decreased significantly while the expression of Axin1 in miR-124-3p inhibitory group increased significantly. Conclusion miR-124-3p regulates BMSCs osteogenic differentiation and participates in PMOP formation via Wnt pathway target factor Axin1. [ABSTRACT FROM AUTHOR]