Reduced secretion of osteoclast coupling factors RANKL and m-CSF by Akt1-deficient osteoblasts. (A) Experimental scheme for assessment of secreted factors found in conditioned medium (CM) from MSCs induced to undergo osteoblast differentiation for 7 days. (B to D) Measurement by ELISA for RANKL (B), m-CSF (C), and OPG (D) (mean ± SD of results of three experiments; ∗, P < 0.01 versus Con CM). (E) Steady-state levels of RANKL, m-CSF, OPG, and S17 mRNAs assessed by RT-PCR after 0 to 10 days of osteoblast differentiation in control and Akt1-deficient MSCs. (F) Experimental scheme for assessment of secreted factors found in CM from MEFs induced to undergo osteoblast differentiation for 7 days. (G to I) Measurement by ELISA for RANKL (G), m-CSF (H), and OPG (I) (mean ± SD of results of three experiments; ∗, P < 0.01 Akt1 KO versus WT CM).