Figure 3.

Osteo-induced PLA cells express several osteogenic genes and proteins. (A) PLA cells and MSCs were induced for up to 6 wk in OM. Cells were assayed for AP activity and total calcium and normalized with respect to protein. Non-induced PLA cells (control) were analyzed as a negative control. Values were expressed as the mean ± SD. (B) PLA cells were cultured in OM or noninductive control medium for up to 6 wk and analyzed by RT-PCR for the indicated genes. NHOst cells maintained in control medium (Con) or OM for 4 wk (28 d) were analyzed as a positive control. (C) Expression of the genes CBFA-1 and AP was quantitated by real-time PCR in PLA cells induced in control medium and OM for up to 4 wk. Gene expression levels were normalized with respect to endogenous GAPDH and expressed relative to non-induced control levels. (D) PLA cells were cultured in OM or control medium for 7 and 28 d and analyzed by Western blotting for the expression of OP, ON, AP, RARα, the VDR, and CNI. Expression of the TfR and α-actin was assessed as internal controls.