Connexin43 Controls the Myofibroblastic Differentiation of Bronchial Fibroblasts from Patients with Asthma

Abstract

Pathologic accumulation of myofibroblasts in asthmatic bronchi is regulated by extrinsic stimuli and by the intrinsic susceptibility of bronchial fibroblasts to transforming growth factor-β (TGF-β). The specific function of gap junctions and connexins in this process has remained unknown. Here, we investigated the role of connexin43 (Cx43) in TGF-β-induced myofibroblastic differentiation of fibroblasts derived from bronchoscopic biopsy specimens of patients with asthma and donors without asthma. Asthmatic fibroblasts expressed considerably higher levels of Cx43 and were more susceptible to TGF-β-induced myofibroblastic differentiation than were their nonasthmatic counterparts. TGF-β efficiently up-regulated Cx43 levels and activated the canonical Smad pathway in asthmatic cells. Ectopic Cx43 expression in nonasthmatic (Cx43) fibroblasts increased their predilection to TGF-β-induced Smad2 activation and fibroblast-myofibroblast transition. Transient Cx43 silencing in asthmatic (Cx43) fibroblasts by Cx43 small interfering RNA attenuated the TGF-β-triggered Smad2 activation and myofibroblast formation. Direct interactions of Smad2 and Cx43 with β-tubulin were demonstrated by co-immunoprecipitation assay, whereas the sensitivity of these interactions to TGF-β signaling was confirmed by Förster Resonance Energy Transfer analyses. Furthermore, inhibition of the TGF-β/Smad pathway attenuated TGF-β-triggered Cx43 up-regulation and myofibroblast differentiation of asthmatic fibroblasts. Chemical inhibition of gap junctional intercellular communication with 18 α-glycyrrhetinic acid did not affect the initiation of fibroblast-myofibroblast transition in asthmatic fibroblasts but interfered with the maintenance of their myofibroblastic phenotype. Collectively, our data identified Cx43 as a new player in the feedback mechanism regulating TGF-β/Smad-dependent differentiation of bronchial fibroblasts. Thus, our observations point to Cx43 as a novel profibrotic factor in asthma progression.

Helicobacter pylori‐activated gastric fibroblasts induce epithelial‐mesenchymal transition of gastric epithelial cells in vitro in a TGF‐β‐dependent manner

“…Close associations of GC with the growth factor‐producing activity of organ‐specific fibroblasts have already been reported . Numerous reports indicate that this kind of reactions is common for cancer‐predisposed fibroblasts . Notably, in our experimental model the expression of Snail mRNA was upregulated in Hp ‐infected fibroblasts.…”

Responsiveness of human bronchial fibroblasts and epithelial cells from asthmatic and non-asthmatic donors to the transforming growth factor-β1 in epithelial-mesenchymal trophic unit model

“…However, only NA HBFs in the mixed TGF-β1 treated EMTU cultures showed a significant protective effect of HBFs on the epithelial layer permeability (measured as the TEER value). A similar protective effect of fibroblasts on the TEER values was described previously in co-cultures of HBECs with MRC5 cells in the EMTU model compared to equivalent HBEC monocultures[49].Longitudinal studies on the TGF-β1-induced FMT potential (a key event in asthmatic subepithelial fibrosis) revealed a facilitated ability of asthmatic HBF populations in standard '2D' cultures for phenotypic shifts[23,25,27,28,50], that may indicate the involvement of intrinsic properties of these cells on the regulation of FMT. In this study, we also observed the enhanced FMT potential defined by a relatively high level of ACTA2 (encodes α-SMA), TAGLN…”

“…Our longitudinal studies clearly show that populations of the AS and NA HBF cultured in the identical conditions in a serum-free medium display different features affecting their sensitivity to the external TGF-β1 and FMT potential. In standard in vitro cultures the AS HBFs in response to the TGF-β1 develop the myofibroblastic phenotype characterized by a relatively high level of α-SMA incorporated into stress fibers more efficiently than the NA counterparts [23- 25,27,28]. To clarify whether this phenomenon is pronounced in the EMTU co-cultures, we firstly checked in this study if the medium dedicated for the air-liquid-interface (ALI) and EMTU cultures significantly affects the FMT potential of the TGF-β1-treated HBFs.…”

“…It suggests that mesenchymal cells, especially bronchial fibroblasts, may play more important role in induction or progression of remodeling than it was previously thought. Our recent studies indicate that bronchial fibroblasts isolated from asthmatic patients reveal different features than their nonasthmatic counterparts [23][24][25][26][27][28]. Therefore, our present study aims to assess the effect of human bronchial fibroblasts (HBFs) and their intrinsic properties on the responsiveness of human bronchial epithelial cells (HBECs) to the TGF-β1 using in vitro EMTU model.…”

Crosstalk between TGF-β and WNT signalling pathways during cardiac fibrogenesis

“…The ultimate consequence of the TGF-β signalling is transcriptional dysregulation of target genes which control cell proliferation and production of structural and ECM proteins, such as collagens, laminins, fibronectin and many others (Branton & Kopp, 1999). TGF-β-dependent mechanisms contribute to fibrotic processes not only in the heart, but also in other organs, including kidney, liver, lung or skeletal muscle (Biernacka et al, 2011;Dooley & ten Dijke, 2012;Germano et al, 2009;Leask, 2007;Meng et al, 2015;Paw et al, 2017;Tatler & Jenkins, 2012, Li et al, 2004.…”