The aim of this study was to reveal the effect of let-7b on osteoporosis (OP). Synthetic let-7b mimics or inhibitors were transfected into MC3T3-E1 cells. The expression of let-7b in MC3T3-E1 and its effect on cell viability, apoptosis, and the apoptosis-related proteins (Bcl-2, Bax, and cleaved caspase-9) were tested by CCK-8 assay, flow cytometry and Western blot, severally. The osteogenic differentiation markers (Runx2 and Osterix) and Wnt/β-catenin pathway related markers (β-catenin and C-myc) were detected by qRT-PCR and Western blot. The relationships between let-7b and cyclin D1 (CCND1) were confirmed by luciferase reporter assay. The differentiation and mineralization of MC3T3-E1 cells were analyzed by alkaline phosphatase (ALP) activity assay and alizarin red staining. The outcomes indicated that overexpression/ablation of let-7b repressed/facilitated MC3T3-E1 cell viability and accelerated/suppressed MC3T3-E1 cell apoptosis. Besides, a remarkable decrease/augment of Bcl-2 protein expression and the distinct fortify/reduction of Bax and cleaved caspase-9 expression levels were observed in let-7b mimics/inhibitors group in MC3T3-E1 cells. Moreover, we discovered that let-7b overexpression/ablation retrained/facilitated the mRNA and protein expression of Runx2 and Osterix. It was confirmed that CCND1 was a downstream target of let-7b and was negatively modulated by let-7b. In addition, high-expression/deficiency of let-7b inhibited/increased the expression levels of β-catenin and C-myc in MC3T3-E1 cells. Taken together, our study revealed that let-7b overexpression/depletion repressed/accelerated MC3T3-E1 cell proliferation, differentiation, and mineralization while promoted/suppressed MC3T3-E1 cell apoptosis through targeting CCND1, which might be adjusted by Wnt/β-catenin pathway. Our findings might offer a basis for developing novel targets for OP treatment.
Keywords: CCND1; Wnt/β-catenin signaling pathway; let-7b; osteoblast differentiation; osteoporosis.
© 2020 The Authors. The Kaohsiung Journal of Medical Sciences published by John Wiley & Sons Australia on behalf of Kaohsiung Medical University.