Fig. 3.

BMP15 induces the Smad and non-Smad pathway. a HGrC1 cells were treated with or without 500 ng/ml BMP15, 10 μM LDN193189, or 10 μM SB203580 for 48 h. BMP15 increased FSHR protein expression. However, LDN193189 suppressed the effect, similar to that of the control. SB203580 suppressed the effect but to a lesser extent as compared to LDN193189. b HGrC1 cells were treated with or without 500 ng/ml BMP15 for 0–90 min. The cells were pretreated with LDN193189 and SB203580 for 2 h. BMP15 increased phosphorylation of Smad 1/5/8. The effect was inhibited by LDN193189, but not by SB203580. c HGrC1 cells were stimulated with or without 250 ng/ml BMP15, 10 μM SB203580, or 10 μM LDN193189 for 24 h. BMP15 significantly increased HAT activity compared to the control. The effect was inhibited by LDN193189, but not by SB203580. Data are shown as mean ± SD of three experiments. * p < 0.05. d BMP15 increased phosphorylation of p38 MAPK, whereas the addition of LDN193189 suppressed this effect. SB203580 addition exerted no effect. e BMP15 increased phosphorylation of USF1, whereas the addition of LDN193189 suppressed this effect. SB addition suppressed BMP15-induced phosphorylation of USF1. In Western blotting, we analyzed the ratio of FSHR to actin, p-Smad 1/5/8 to Smad 1/5/8, p-p38 to p38, and p-USF1 to USF1 by using ImageJ to show densitometry