This study aims to explore the role of lncRNA MSC-AS1/microRNA-140-5p/BMP2 regulatory loop in promoting osteogenic differentiation of BMSCs. BMSCs were isolated from bone marrow of mice. Expression levels of MSC-AS1, microRNA-140-5p and BMP2 during osteogenic differentiation were detected by qRT-PCR. Meanwhile, regulatory effect of MSC-AS1 on osteogenic differentiation was detected through ALP staining and alizarin red staining. The binding sites between microRNA-140-5p and MSC-AS1 as well as between microRNA-140-5p and BMP2 were predicted by TargetScan, which were further confirmed by dual-luciferase reporter gene assay. In addition, protein levels of MSC-AS1/microRNA-140-5p/BMP2 were detected by Western blot. Finally, rescue experiments were conducted to clarify the regulatory effects of MSC-AS1/microRNA-140-5p/BMP2 axis on osteogenic differentiation. MSC-AS1 and BMP2 were found to be remarkably up-regulated during osteogenic differentiation, while microRNA-140-5p was conversely down-regulated. Meanwhile, knockdown of MSC-AS down-regulated expression levels of osteogenesis-associated genes and weakened the mineralization capacity of BMSCs. MicroRNA-140-5p was verified to bind to the 3'UTR of MSC-AS1 and BMP2 genes. Knockdown of MSC-AS1 in BMSCs could reduce the expression of microRNA-140-5p, while knockdown of microRNA-140-5p also down-regulated BMP2 level. In addition, co-silence of MSC-AS1 and microRNA-140-5p reversed the inhibitory effect of MSC-AS1 knockdown on osteogenic differentiation and protein levels of p-Smad1/5/8, RUNX2 and Osterix. MSC-AS1 might promote the osteogenic differentiation of BMSCs through sponging microRNA-140-5p to up-regulate BMP2, thus alleviating the progression of osteoporosis.
Keywords: BMP2; MSC-AS1; MicroRNA-140–5p; osteoporosis.
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