Retinoic acid treatment of human leiomyoma cells transformed the cell phenotype to one strongly resembling myometrial cells

Abstract

Background Uterine leiomyomas are clinically significant tumours that may develop due to an altered differentiation pathway. We have previously identified a dysregulated retinoic acid (RA) pathway that reduced retinoic exposure in human leiomyoma surgical specimens, and have shown that the leiomyoma phenotype was characterized by excessive and disorganized extracellular matrix (ECM). Objective The goal of this study was to determine the impact of RA exposure on the disrupted ECM phenotype of leiomyomas. Design and methods Study of immortalized and molecularly confirmed cells generated from surgical specimens of spontaneous uterine leiomyoma and matched myometrium. Results Immortalized leiomyoma and myometrial cells retained the molecular characteristics of their progenitor tissue. Proliferation of leiomyoma cells was inhibited by all -trans retinoic acid (ATRA). Furthermore, there was a dose-dependent decrease in soluble extracellular collagen protein in ATRA-treated leiomyoma cells. Exposure of leiomyoma cells to ATRA resulted in a dose-dependent inhibition of templates for specific ECM protein production including collagen 1, collagen 4, fibronectin and versican. Notably, expression levels in treated leiomyoma cells approached those found in myometrial cells. These mRNA alterations translated into altered protein. Down-regulation was also observed among the RA pathway genes such as CYP26A1 with exposure to ATRA. Finally, ATRA down-regulated TGF-β3 mRNA expression and the TGF-β regulated genes in leiomyoma cells. Conclusion Exposure of leiomyomas to ATRA down-regulated cell proliferation, ECM formation, RA metabolism and TGF-β regulation, suggesting that RA exposure can alter the leiomyoma phenotype to one that more closely approximates normal myometrium.

Leiomyoma Cells in 3-Dimensional Cultures Demonstrate an Attenuated Response to Fasudil, a Rho-Kinase Inhibitor, When Compared to 2-Dimensional Cultures

“…Immortalized myometrial and leiomyoma cells 38 À7 mol/L to 10 À5 mol/L in CH 10% for 24 hours and 72 hours. Fresh media containing treatment concentrations of fasudil were replaced after 48 hours.…”

Common fibroid-associated genes are differentially expressed in phenotypically dissimilar cell populations isolated from within human fibroids and myometrium

“…Members of the TGFB family can elicit tumour-promoting effects in a paracrine manner by triggering fibroblast differentiation and desmoplastic responses from neighbouring mesenchymal cells within a tumour (reviewed by Elenbaas & Weinberg (2001) and Polanska & Orimo (2013)). Activation of the retinoic acid (RA) pathway significantly reduces growth and extracellular matrix composition of uterine fibroid cells via regulation of the TGFB pathway (Malik et al 2008). In fibroblasts, differential regulation can occur via the RA receptors (RAR), which are cell-type specific with distinctive receptor binding properties between different cell types (Delacroix et al 2010), providing a mechanism for paracrine interaction in the tumour microenvironment.…”

Optimized Culture Conditions for Tissue Explants of Uterine Leiomyoma

“…Hormone receptor content in relation to culture duration As mentioned before, fast dedifferentiation of myoma cells in culture -especially the loss of hormone receptors -is a main problem in studying these tissues [14,18,19]. After optimizing different parameters, we tested for the presence of hormone receptor proteins ER and PR in uterine myoma tissue cultured for 24, 48, and 72 hours and compared these data to native tissue.…”

“…Thus some scientists use cultivated cells from myomas; however to use them as a cell line they have to be immortalised by HPV-16 genes or telomerase-reverse-transcriptase [13,14] due to scanty growth and early senescence under culture conditions [15,16], which may change phenotype of myoma cells [17]. Another barrier to using cell cultures in myoma studies is the problem of fast cell dedifferentiation [14,18,19]. Within hours after beginning of culturing, m-RNA transcripts of ER and PR were decreased, independent of adding estrogens and/or progesterone.…”