Integrity of Cell-Cell Contacts Is a Critical Regulator of TGF-β1-Induced Epithelial-to-Myofibroblast Transition

Abstract

Injury of the tubular epithelium and TGF-beta1-induced conversion of epithelial cells to alpha-smooth muscle actin (SMA)-expressing myofibroblasts are key features of kidney fibrosis. Since injury damages intercellular junctions and promotes fibrosis, we hypothesized that cell contacts are critical regulators of TGF-beta 1-triggered epithelial-to-mesenchymal transition (EMT). Here we show that TGF-beta 1 was unable to induce EMT in intact confluent monolayers, but three different models of injury-induced loss of epithelial integrity (subconfluence, wounding, and contact disassembly by Ca(2+)-removal) restored its EMT-inducing effect. This manifested in loss of E-cadherin, increased fibronectin production and SMA expression. TGF-beta 1 or contact disassembly alone only modestly stimulated the SMA promoter in confluent layers, but together exhibited strong synergy. Since beta-catenin is a component of intact adherens junctions, but when liberated from destabilized contacts may act as a transcriptional co-activator, we investigated its role in TGF-beta 1-provoked EMT. Contact disassembly alone induced degradation of E-cadherin and beta-catenin, but TGF-beta1 selectively rescued beta-catenin and stimulated the beta-catenin-driven reporter TopFLASH. Moreover, chelation of free beta-catenin with the N-cadherin cytoplasmic tail suppressed the TGF-beta1 plus contact disassembly-induced SMA promoter activation and protein expression. These results suggest a beta-catenin-dependent two-hit mechanism in which both an initial epithelial injury and TGF-beta 1 are required for EMT.

The role of TGF-β and epithelial-to mesenchymal transition in diabetic nephropathy

“…Since these antagonists are critical in ensuring the regulation of Smadmediated gene transcription, a fine balance must be achieved in order to match cellular demands. It is of no surprise that diminished levels of co-repressors are observed in animal models of obstructive nephropathy and diabetes [67]. Smad ubiquitination regulatory factor-2 (Smurf2) is an ubiquitin ligase that specifically targets certain members of Smad proteins for degradation, including Ski, SnoN and TGIF [63].…”

Salvia Miltiorrhiza Bge.f.alba Ameliorates the Progression of Monocrotaline-Induced Pulmonary Hypertension by Protecting Endothelial Injury in Rats

“…Studies have shown that TGF-β 1 stimulates lung microvascular endothelial cells to produce ET-1, which promotes smooth muscle cell proliferation (Star et al 2008). Furthermore, TGF-β 1 plays an initiation and facilitation role in the process of endothelium-smooth muscle transformation (Masszi et al 2004). In our study, higher TGF-β 1 levels in the lung tissue of the MCT group may be due to elevated shear force, which can cause increased release of TGF-β 1 from the endothelium (Cucina et al 1998).…”

Heterogeneity of Macrophage Populations and Myofibroblasts Appearing in Rat Renal Interstitial Fibrosis

“…Submesothelial stromal cells, ovarian thecal cells, pericytes, hepatic perisinusoidal cells (hepatic stellate cells), pancreatic stellate cells or reactive macrophages have been also proposed as cells capable of expressing myofibroblastic phenotypes under pathological conditions [33][34][35][38][39][40] . Furthermore, it has been reported that renal epithelial cells may transdifferentiate into myofibroblastic cells in renal interstitial fibrosis [41][42][43][44] . It thus appears that so-called "myofibroblastic cells" in fibrotic lesions are generated from heterogeneous cell populations 45 .…”

“…2); these cytoskeletons represent useful markers of myofibroblastic differentiation 33,35 . It has been recently postulated that EMT, in which renal epithelial cells acquire myofibroblastic phenotypes, may contribute to the progressive renal fibrosis [41][42][43][44]60 . In the chronic model, it has been found that a number of cuboidal or flattened, single-or multi-layered epithelia in the fibrotic areas show a strong immunoexpression to α-SMA (Fig.…”

“…TGF-β1 is the most effective factor for EMT 41,42 . NRK-52E, an epithelial type cell line isolated from rat normal kidneys, has been used for the demonstration of EMT in vitro.…”