Taken together, these results suggest that downregulation of activin A and Smad3, both members of the TGF-β pathway, may offer a mechanistic explanation for the inhibitory effect of a high-dose of genistein on UtLM cells, and might be potential therapeutic targets for treatment of clinical cases of uterine leiomyomas.
Moreover, we found that activin A stimulated the growth of UtLM cells, and the inhibitory effect of genistein was partially abrogated in the presence of activin A. Overexpression of activin A and Smad3 were found in tissue samples of leiomyoma compared to matched myometrium, supporting the contribution of activin A and Smad3 in promoting the growth of UtLM cells.
Using semiquantitative RT-PCR, Western blot analysis, and immunohistochemistry, we demonstrated that leiomyoma, myometrium, LSMC, and MSMC express receptor-activated Smad3, common Smad4, and the inhibitory Smad7 mRNA and protein and showed that TGFbeta1, in a time-dependent manner, transiently induced Smad7 expression, with Smad3 and Smad4 remaining largely unchanged.
Semi-quantitative RT-PCR, Western blotting and immunohistochemistry indicate that leiomyoma and myometrium expresses receptor-activated Smad3, common Smad4 and inhibitory Smad7, with elevated expression of Smad3, Smad4 and phosphorylated Smad3 (pSmad3) as well as TGF-betaR type I and type II in leiomyoma compared to myometrium (P<0.05).