Panx3 promotes osteoblast differentiation. C2C12 cells were transiently transfected with a control pEF1 vector, pEF1/Panx3, a control sh vector, or a Panx3 shRNA vector, and these cells were cultured with BMP2 for various durations, as indicated. Total RNA was extracted each day for 4 d and mRNA levels were analyzed by quantitative RT-PCR. (A) Panx3 overexpression promoted the expression of osteoblast marker genes for osterix, ALP, and Ocn, except that the expression of Runx2 remained the same. (B) shPanx3 suppressed the induction of these genes, except for Runx2. (C) Panx3 overexpression promoted ALP activity, whereas shPanx3 inhibited it. Representative ALP staining (top) and its quantitative data (bottom). C2C12 cells and pEF1/Panx3- and shPanx3-transfected C2C12 cells were cultured with BMP2 for 3 d. (D) Representative Alizarin red S staining (top) and its quantitative data (bottom) of C2C12 and pEF1/panx3- and shPanx3-transfected C2C12 cells cultured with BMP2 for 15 d. **, P < 0.01. Error bars represent the mean ± SD; n = 3.