Abstract
Transformation of epithelial cells into connective tissue cells (epithelial-mesenchymal transition, EMT) is a complex mechanism involved in tumor metastasis, and in normal embryogenesis, while type II EMT is mainly associated with inflammatory events and tissue regenaration.In this study we examined type II EMT at the ultrastructural and molecular level during the inflammatory process induced by Freund's adjuvant treatment in rat mesenteric mesothelial cells. We found that upon the inflammatory stimulus mesothelial cells lost contact with the basal lamina and with each other, and were transformed into spindle-shaped cells. These morphological changes were accompanied by release of interleukins IL-1alpha, -1beta and IL-6 and by secretion of transforming growth factor beta (TGF-β) into the peritoneal cavity. Mesothelial cells also expressed estrogen receptor alpha (ER-α) as shown by immunolabeling at the light and electron microscopical levels, as well as by quantitative RT-PCR. The mRNA level of ER-α showed an inverse correlation with the secretion of TGF-β. At the cellular and subcellular levels ER-α was colocalized with the coat protein caveolin-1 and was found in the plasma membrane of mesothelial cells, in caveolae close to multivesicular bodies (MVBs) or in the membrane of these organelles, suggesting that ER-α is internalized via caveola-mediated endocytosis during inflammation. We found asymmetric, thickened, electron dense areas on the limiting membrane of MVBs (MVB plaques) indicating that these sites may serve as platforms for collecting and organizing regulatory proteins. Our morphological observations and biochemical data can contribute to form a potential model whereby ER-α and its caveola-mediated endocytosis might play role in TGF-β induced type II EMT in vivo.
Highlights
Epithelial-mesenchymal transition (EMT) is a biological process that allows a polarized epithelial cell to undergo several biochemical/morphological changes to gain a mesenchymal phenotype [1]
Since mesothelial cells can serve as one of the sources of activated macrophages upon Freund’s adjuvant treatment and it is known that macrophages express estrogen receptor alpha (ER-a), the question arose whether mesenteric mesothelial cells express the molecule? Our present results show that ER-a is present in mesothelial cells both under steady state and inflammatory conditions
Our results revealed that control mesothelial cells expressed ER-a mRNA and it showed a significant downregulation during the progression of the inflammatory process (Fig. 3E). (We found the same pattern of changes in the mRNA expression levels of ERb and G protein-coupled receptor 30, GPR30 as well (Fig. S1.).)
Summary
Epithelial-mesenchymal transition (EMT) is a biological process that allows a polarized epithelial cell to undergo several biochemical/morphological changes to gain a mesenchymal phenotype [1]. The process was first described by Elisabeth Hay who depicted the basic differences of these cellular actions during embryogenesis and tumorigenesis [2]. It has been demonstrated that upon inflammation many cells (monocytes/macrophages, fibroblasts) can trigger type II EMT through secretion of growth factors such as transforming growth factor-beta (TGF-b) or epidermal growth factor (EGF). Most prominent among these cells are the macrophages and activated resident fibroblasts that accumulate at the site of injury and release these growth factors [1], [2], [5]. TGF-b was first described to induce EMT via Smad 2/3dependent pathway and it became evident that the cellular actions of the cytokine can be modulated and defined by other Smad-independent signaling pathways like the MAP kinase pathways [6]–[8]
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