FIGURE 6.

SMAD2 and SMAD3 are the downstream mediators of the growth differentiation factor 8 (GDF8)-induced upregulation of follistatin-like 3 (FSTL3) expression in HTR8/SVneo cells. (A–D) HTR8/SVneo cells were transfected for 24 h with 20 nM non-targeting control siRNAs (siCtrl), 20 nM siRNAs targeting SMAD2 (siSMAD2) or 20 nM siRNAs targeting SMAD3 (siSMAD3), after which the cells were treated with the vehicle control (Ctrl) or 25 ng/mL GDF8 for 24 h; the mRNA and protein levels of SMAD2 (A,B) and SMAD3 (C,D) were examined using RT-qPCR and western blot analysis, respectively. (E–H) HTR8/SVneo cells were transfected for 24 h with 20 nM siCtrl, 20 nM siSMAD2 (E,F) or 20 nM siSMAD3 (G,H), after which the cells were treated with the vehicle control (Ctrl) or 25 ng/mL GDF8 for 24 h; the mRNA (E,G) and accumulated (F,H) levels of FSTL3 were examined using RT-qPCR and ELISA, respectively. GAPDH used as the loading control and a reference for densitometric analysis for SMAD2 and SMAD3. The results are displayed as the mean ± SEM of at least three independent experiments, and the values with different letters are significantly different (P < 0.05).