Previously, we found that high doses of genistein show an inhibitory effect on uterine leiomyoma (UtLM) cell proliferation. In this study, using microarray analysis and Ingenuity Pathways Analysis(TM), we identified genes (up- or down-regulated, > or = 1.5 fold, P < or = 0.001), functions and signaling pathways that were altered following treatment with an inhibitory concentration of genistein (50 microg/ml) in UtLM cells. Downregulation of TGF-beta signaling pathway genes, activin A, activin B, Smad3, TGF-beta2 and genes related to cell cycle regulation, with the exception of the upregulation of the CDK inhibitor P15, were identified and validated by real-time RT-PCR studies. Western blot analysis further demonstrated decreased protein expression of activin A and Smad3 in genistein-treated UtLM cells. 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. Taken together, these results suggest that down-regulation of activin A and Smad3, both members of the TGF-beta 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.
Activins/*genetics/metabolism/pharmacology ; Anticarcinogenic Agents/*pharmacology ; Cell Line, Tumor ; Cell Proliferation/drug effects ; Cyclin-Dependent Kinase Inhibitor p15/genetics/metabolism ; Down-Regulation ; Female ; Genistein/*pharmacology ; Humans ; Leiomyoma/*metabolism ; Oligonucleotide Array Sequence Analysis ; Signal Transduction/drug effects ; Smad3 Protein/*genetics/metabolism ; Transforming Growth Factor beta/*genetics/metabolism ; Up-Regulation ; Uterine Neoplasms/*metabolism