MENG Rui, LUO Ming-Zhi, WANG Liang, GAO Xiang, DI Sheng-Meng, SHANG Peng*
（Key Laboratory for Space Biosciences & Biotechnology, Institute of Special Environmental Biophysics, Faculty of Life Sciences, Northwestern Polytechnical University, Xi，an710072, China）
Abstract：Osteocytes are the major mechanosensors in bone tissue. It is important for explaining the mechanism of bone loss to investigate the regulating role of osteocytes on its effector cells under weightlessness. To investigate the regulating effects of conditioned medium of osteocytes on functions of osteoblasts, the 3-D rotation was used to simulate weightlessness. After mouse osteocyte-like cell line MLO-Y4 subjected 3-D rotation for 72 hours, the conditioned medium of rotation (RCM) and conditioned medium of control (CCM) were collected. The effects of RCM and CCM on proliferation, cell cycle, ALP activity and osteogenesis regulated gene expression of MC3T3-E1 cells were analyzed by MTT (methyl thiazolyl tetrazolium), pNPP (p-nitrophenyl phosphate), flow cytometry and RT-PCR, respectively. The results showed that the proliferation of MC3T3-E1 was significantly stimulated by RCM of MLO-Y4 after subjected 3-D rotation for 72 hours. After MC3T3-E1 cells were cultured for 24 and 48 hours, the rates of proliferation in 50% RCM were 1.62 and 1.60-fold than in CCM, respectively. (*P<0.05). After 72 hours, the rate of proliferation in 100% RCM was 1.69-fold than in CCM. The cell cycle assay indicated that the cell cycle arrest of MC3T3-E1 induced by 100% CCM were partly restored by 100% RCM. There was no difference between RCM and CCM on ALP activity of MC3T3-E1 cells. RT-PCR showed that 100% RCM significantly decreased the mRNA expression of ALP, Runx2, OPN and OC（* P<0.05，**P<0.01，*** P<0.00 1）. In summary, the proliferation of osteoblasts were stimulated by conditioned medium of MLO-Y4 after subjected 3-D rotation for 72 hours. However, the osteogenesis regulated gene expression were significantly decreased.
