miR-138–5p negatively regulates osteoblast differentiation through inhibiting β-catenin under simulated microgravity in MC3T3-E1 cells

With the increasing interest in exploring the deep-space environment, the problems of health and safety of astronauts' bone induced by microgravity warrants an investigation. Recent studies have discovered that several microRNAs (miRNAs) have played a vital role in osteoblast differentiation and bone formation under microgravity, whereas the in-depth signaling pathway mechanisms are not yet completely understood. Here, we performed the hind limb unloading (HLU) mice model and 3D clinostat-random position machine (RPM) to simulate the effects of microgravity on bone at animal and cellular level, respectively. Firstly, we screened the different expressed miRNAs under simulated microgravity by miRNA sequencing, we then identified the highest different expressed miRNA (miR-138–5p) that was up-regulated under simulated microgravity and inhibited osteoblast differentiation in MC3T3-E1 cells. Moreover, we analyzed miR-138's targets during osteogenic differentiation and we found that these targets regulated osteogenic differentiation through Wnt/β-catenin signaling, a key signaling pathway in controlling osteoblast differentiation. The association between miR-138–5p and β-catenin activity was determined by a luciferase reporter assay. Further experiments confirmed that miR-138–5p suppressed β-catenin expression and β-catenin activity. Moreover, β-catenin overexpression attenuated osteoblast differentiation reduction induced by increased miR-138–5p level in miR-138–5p overexpression osteoblastic cells. In addition, inhibition of miR-138–5p counteracted the negative effects of simulated microgravity on osteoblast differentiation and β-catenin expression in MC3T3-E1 cells. Our findings demonstrated that a signaling pathway mechanism of miR-138–5p in regulating osteoblast differentiation under simulated microgravity, which may reveal a novel mechanism for astronauts' bone loss induced by microgravity and provide a potential therapeutic target for disused osteoprosis.