Altered expression of the TGF-beta system is recognized to play a central role in various fibrotic disorders, including leiomyoma. In this study we performed microarray analysis to characterize the gene expression profile of leiomyoma and matched myometrial smooth muscle cells (LSMC and MSMC, respectively) in response to the time-dependent action of TGF-beta and, after pretreatment with TGF-beta type II receptor (TGF-beta RII) antisense oligomer-blocking/reducing TGF-beta autocrine/paracrine actions. Unsupervised and supervised assessments of the gene expression values with a false discovery rate selected at P < or = 0.001 identified 310 genes as differentially expressed and regulated in LSMC and MSMC in a cell- and time-dependent manner by TGF-beta. Pretreatment with TGF-beta RII antisense resulted in changes in the expression of many of the 310 genes regulated by TGF-beta, with 54 genes displaying a response to TGF-beta treatment. Comparative analysis of the gene expression profile in TGF-beta RII antisense- and GnRH analog-treated cells indicated that these treatments target the expression of 222 genes in a cell-specific manner. Gene ontology assigned these genes functions as cell cycle regulators, transcription factors, signal transducers, tissue turnover, and apoptosis. We validated the expression and TGF-beta time-dependent regulation of IL-11, TGF-beta-induced factor, TGF-beta-inducible early gene response, early growth response 3, CITED2 (cAMP response element binding protein-binding protein/p300-interacting transactivator with ED-rich tail), Nur77, Runx1, Runx2, p27, p57, growth arrest-specific 1, and G protein-coupled receptor kinase 5 in LSMC and MSMC using real-time PCR. Together, the results provide the first comprehensive assessment of the LSMC and MSMC molecular environment targeted by autocrine/paracrine action of TGF-beta, highlighting potential involvement of specific genes whose products may influence the outcome of leiomyoma growth and fibrotic characteristics by regulating inflammatory response, cell growth, apoptosis, and tissue remodeling.