Fig. 3

PKA stimulators regulate gene expression for adipogenesis, osteogenesis and osteoclast-inducing factors by suppressing leptin. (A,B) Suppression of leptin secretion and transcription by IBMX, forskolin and Sp-cAMP. Cell treatments were the same as Fig. 1. (A) Leptin concentrations in the conditioned medium of the last 2 days were measured by ELISA and normalized by cell numbers. The figure shows representative data from two independent experiments with three replicates for each condition; data are presented as the meanSD values, and significance was determined by Student's t-test. (**<0.01 versus A; ^#<0.05 versus S) (B) RT-PCR analysis of mRNA for leptin and β-actin. (C) Exogenous leptin inhibits forskolin-induced effects on gene expression of PPARγ2 and Runx2 and the RANKL/OPG gene expression ratio. Cells were cultured in complete growth medium [C] or treated in AIM with 100 μM forskolin [F] in the absence or presence of leptin (0.6, 1.5, 3 μg/ml) for 7 days. RT-PCR analysis for β-actin, PPARγ2, Runx2, RANKL and OPG. Real-Time RT-PCR analysis of the gene expression ratio of PPARγ2/GADPH (D), Runx2/GADPH (E), and RANKL/OPG. (F). Exogenous leptin inhibits the effects of AIM with IBMX (I: AIM with 0.45 mM IBMX) on gene expression of PPARγ2, Runx2 and the RANKL/OPG gene expression ratio.