TGF-Β SIGNALING REGULATES WNT/Β-CATENIN PATHWAY IN UTERINE LEIOMYOMA CELLS

Abstract

Uterine leiomyomas, also known as uterine fibroids, are the most common benign gynecologic tumor of the female reproductive tract. Despite their high prevalence and heavy burden, their pathogenesis is not clear. Several signaling aberrations have been identified, including Wnt/β-catenin and TGF-β. TGF-β pathway is a crucial mediator of extracellular matrix (ECM) deposition and fibrosis and is upregulated in leiomyoma compared to the myometrium. The Wnt/β-catenin pathway is a conserved pathway implicated in proliferation, differentiation and stem cell renewal and has also been implicated in uterine leiomyoma development. Due to the importance of these two pathways in uterine leiomyoma development, we are interested in studying the interaction between these two pathways to better understand the pathogenesis of these tumors. After obtaining informed consent, primary fibroid and myometrial cells were isolated from hysterectomy surgeries at Johns Hopkins Hospital. Primary cells and immortalized fibroid (HuLM) and myometrial (UtSM) were treated with TGF-β3 (10ng/mL) for 48 hours. The effect of TGF-β3 on the Wnt/β-catenin pathway was examined using western blot through assessing the expression of total β-catenin, non-phosphorylated β-catenin, the active form of β-catenin, and Dickkopf-1 (DKK1), an inhibitor of Wnt pathway. The student’s t-test was used to determine statistically significant differences (P<0.05). As expected, TGF-β3 treatment induced the expression of ECM proteins collagen 1A and fibronectin in both primary and immortalized fibroid and myometrial cells. The expression of total β-catenin and non-phosphorylated β-catenin was higher in leiomyoma cells, and TGF-β3 treatment significantly increased the expression of total β-catenin and non-phosphorylated β-catenin in both myometrial by 66% and 59%, respectively and fibroid cells by 58% and 62%, respectively (p-value <0.05). The expression of the Wnt pathway inhibitor, DKK-1, was significantly lower in primary leiomyoma cells compared to myometrial cells, and TGF-β3 treatment resulted in a significant reduction in DKK-1 expression in both myometrial by 85% and fibroid cells by 61%(p-value <0.05). These results indicate that TGF-β3 and Wnt/β-catenin signal transduction pathways interact in uterine leiomyoma. Further analyses are needed to clarify the interaction and the role of DKK-1 in leiomyoma development.