Human uterine leiomyoma-derived fibroblasts stimulate uterine leiomyoma cell proliferation and collagen type I production, and activate RTKs and TGF beta receptor signaling in coculture

Abstract

BackgroundUterine leiomyomas (fibroids) are benign smooth muscle tumors that often contain an excessive extracellular matrix (ECM). In the present study, we investigated the interactions between human uterine leiomyoma (UtLM) cells and uterine leiomyoma-derived fibroblasts (FB), and their importance in cell growth and ECM protein production using a coculture system.ResultsWe found enhanced cell proliferation, and elevated levels of ECM collagen type I and insulin-like growth factor-binding protein-3 after coculturing. There was also increased secretion of vascular endothelial growth factor, epidermal growth factor, fibroblast growth factor-2, and platelet derived growth factor A and B in the media of UtLM cells cocultured with FB. Protein arrays revealed increased phosphorylated receptor tyrosine kinases (RTKs) of the above growth factor ligands, and immunoblots showed elevated levels of the RTK downstream effector, phospho-mitogen activated protein kinase 44/42 in cocultured UtLM cells. There was also increased secretion of transforming growth factor-beta 1 and 3, and immunoprecipitated transforming growth factor-beta receptor I from cocultured UtLM cells showed elevated phosphoserine expression. The downstream effectors phospho-small mothers against decapentaplegic -2 and -3 protein (SMAD) levels were also increased in cocultured UtLM cells. However, none of the above effects were seen in normal myometrial cells cocultured with FB. The soluble factors released by tumor-derived fibroblasts and/or UtLM cells, and activation of the growth factor receptors and their pathways stimulated the proliferation of UtLM cells and enhanced the production of ECM proteins.ConclusionsThese data support the importance of interactions between fibroid tumor cells and ECM fibroblasts in vivo, and the role of growth factors, and ECM proteins in the pathogenesis of uterine fibroids.

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

“…Fibroblasts are an integral cellular component of human uterine fibroids and myometrium (Koumas et al 2001, Zaitseva et al 2007, Moore et al 2010, HoldsworthCarson et al 2014. Although fibroids are clonal, significant cellular heterogeneity exists (HoldsworthCarson et al 2014).…”

“…Of relevance to this investigation, it is interesting to note that genes associated with steroid hormone regulation, the TGFB pathway and genes involved in the RA pathway make up a significant portion of genes that are differentially expressed in fibroid tissue compared with myometrium (refer to Table 1 for a brief summary; Sadan et al 1987, Dou et al 1996, Tsibris et al 2002, Skubitz & Skubitz 2003, Weston et al 2003, Shozu et al 2004, Arslan et al 2005, Leppert et al 2006, Zaitseva et al 2006, Roth et al 2007, Litovkin et al 2008, Lee et al 2010, Moore et al 2010, Davis et al 2013, Yin et al 2013.…”

“…Others and we have shown that in addition to SMC, phenotypically dissimilar cells make up a major component of uterine fibroids, including vascular SMC (VSMC) and fibroblasts (Koumas et al 2001, Zaitseva et al 2007, Moore et al 2010, Holdsworth-Carson et al 2014. It is known that the tumour microenvironment greatly impacts on tumour growth and is strongly influenced by paracrine signalling between several different interacting cells and their secretomes; critical in this process are fibroblasts (Elenbaas & Weinberg 2001).…”

A Mathematical Model of the Growth of Uterine Myomas

“…They generally grow in a spherical shape and, on the macroscopic scale, the cells appear to be distributed uniformly throughout. In addition to the smooth muscle component, they also possess a significant extracellular matrix consisting of fibroblasts [26]. They are therefore highly fibrous with the collagen fibrils randomly oriented, a characteristic different from the adjacent myometrium in which the collagen fibrils are aligned in an orderly fashion [20,30].…”

“…In the case of myomas, the presence of oestrogen and progesterone up-regulates expression of a plethora of growth factors both within myocytes and fibroblasts, enhancing cell proliferation and collagen deposit [26], while at the same time the change in homeostasis in myomas accompanied by altered mechanical stresses, suggests the interplay between the chemical and mechanical effects and the role of stress in cell growth and fibroid development [20]. In vitro studies on tumour spheroids have also demonstrated resistance to tumour growth as a result of stress accumulated both within the tumour as well as in the surrounding medium.…”