Fig. 2: OGRU promotes the osteoblast activity and matrix mineralization, and attenuates the osteogenic decline of MC3T3-E1 cells induced by clinorotation unloading.

MC3T3-E1 cells were transfected with pcDNA3.1(+)–OGRU, si-OGRU, or the corresponding controls, and were cultured in osteogenic medium. a The mRNA levels of Alp, Osx, Runx2, and Ocn were measured by qRT-PCR 4 days after osteogenic treatment (n = 3). b The protein levels of Osx, Runx2, and Ocn were measured by western blotting 4 days after osteogenic treatment (n = 3). c The relative Alp activity, 4 days after osteogenic treatment (n = 3). d Representative images of Alp staining in MC3T3-E1 cells 7 days after osteogenic treatment (n = 3). e Representative images of alizarin red staining in MC3T3-E1 cells 21 days after osteogenic treatment are shown, and relative areas of mineralization were quantified by Image J (n = 3). MC3T3-E1 cells were transfected with pcDNA3.1(+) or pcDNA3.1(+)–OGRU and cultured in clinorotation-unloading condition for 48 h, and were then subjected to qRT-PCR analysis of Alp, Osx, Runx2, and Ocn mRNA levels (f), measurement of the relative Alp activity (g), and western blot analysis of Osx, Runx2, and Ocn protein expression (h) (n = 3). All data are the mean ± SD. *P < 0.05, **P < 0.01.