Dataset Information

TGF-? Induced CTGF Expression in Human Lung Epithelial Cells through ERK, ADAM17, RSK1, and C/EBP? Pathways.

ABSTRACT: Background: Lung epithelial cells play critical roles in idiopathic pulmonary fibrosis.

Methods: In the present study, we investigated whether transforming growth factor-? (TGF-?)-induced expression of connective tissue growth factor (CTGF) was regulated by the extracellular signal-regulated kinase (ERK)/a disintegrin and metalloproteinase 17 (ADAM17)/ribosomal S6 kinases 1 (RSK1)/CCAAT/enhancer-binding protein ? (C/EBP?) signaling pathway in human lung epithelial cells (A549).

Results: Our results revealed that TGF-?-induced CTGF expression was weakened by ADAM17 small interfering RNA (ADAM17 siRNA), TNF-? processing inhibitor-0 (TAPI-0, an ADAM17 inhibitor), U0126 (an ERK inhibitor), RSK1 siRNA, and C/EBP? siRNA. TGF-?-induced ERK phosphorylation as well as ADAM17 phosphorylation was attenuated by U0126. The TGF-?-induced increase in RSK1 phosphorylation was inhibited by TAPI-0 and U0126. TGF-?-induced C/EBP? phosphorylation was weakened by U0126, ADAM17 siRNA, and RSK1 siRNA. In addition, TGF-? increased the recruitment of C/EBP? to the CTGF promoter. Furthermore, TGF-? enhanced fibronectin (FN), an epithelial-mesenchymal transition (EMT) marker, and CTGF mRNA levels and reduced E-cadherin mRNA levels. Moreover, TGF-?-stimulated FN protein expression was reduced by ADAM17 siRNA and CTGF siRNA.

Conclusion: The results suggested that TGF-? induces CTGF expression through the ERK/ADAM17/RSK1/C/EBP? signaling pathway. Moreover, ADAM17 and CTGF participate in TGF-?-induced FN expression in human lung epithelial cells.

SUBMITTER: Ou SC

PROVIDER: S-EPMC7731197 | biostudies-literature | 2020 Nov

REPOSITORIES: biostudies-literature

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TGF-β Induced CTGF Expression in Human Lung Epithelial Cells through ERK, ADAM17, RSK1, and C/EBPβ Pathways.

Ou Shu-Ching SC Bai Kuan-Jen KJ Cheng Wun-Hao WH Chen Jing-Yun JY Lin Chien-Huang CH Wen Heng-Ching HC Chen Bing-Chang BC

International journal of molecular sciences 20201129 23

<h4>Background</h4>Lung epithelial cells play critical roles in idiopathic pulmonary fibrosis.<h4>Methods</h4>In the present study, we investigated whether transforming growth factor-β (TGF-β)-induced expression of connective tissue growth factor (CTGF) was regulated by the extracellular signal-regulated kinase (ERK)/a disintegrin and metalloproteinase 17 (ADAM17)/ribosomal S6 kinases 1 (RSK1)/CCAAT/enhancer-binding protein β (C/EBPβ) signaling pathway in human lung epithelial cells (A549).<h4>R ...[more]

PMID: 33260349

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Project description:Caffeine administration is an important part of the therapeutic treatment of bronchopulmonary dysplasia (BPD) in preterm infants. However, caffeine mediated effects on airway remodelling are still undefined. The TGF-β/Smad signalling pathway is one of the key pathways involved in airway remodelling. Connective tissue growth factor (CTGF), a downstream mediator of TGF-β, and transgelin, a binding and stabilising protein of the cytoskeleton, are both regulated by TGF-β1 and play an important role in airway remodelling. Both have also been implicated in the pathogenesis of BPD. The aim of the present study was to clarify whether caffeine, an unspecific phosphodiesterase (PDE) inhibitor, and rolipram, a prototypical PDE-4 selective inhibitor, were both able to affect TGF-β1-induced Smad signalling and CTGF/transgelin expression in lung epithelial cells. Furthermore, the effect of transgelin knock-down on Smad signalling was studied. The pharmacological effect of caffeine and rolipram on Smad signalling was investigated by means of a luciferase assay via transfection of a TGF-β1-inducible reporter plasmid in A549 cells. The regulation of CTGF and transgelin expression by caffeine and rolipram were studied by promoter analysis, real-time PCR and Western blot. Endogenous transgelin expression was down-regulated by lentiviral transduction mediating transgelin-specific shRNA expression. The addition of caffeine and rolipram inhibited TGF-β1 induced reporter gene activity in a concentration-related manner. They also antagonized the TGF-β1 induced up-regulation of CTGF and transgelin on the promoter-, the mRNA-, and the protein-level. Functional analysis showed that transgelin silencing reduced TGF-β1 induced Smad-signalling and CTGF induction in lung epithelial cells. The present study highlights possible new molecular mechanisms of caffeine and rolipram including an inhibition of Smad signalling and of TGF-β1 regulated genes involved in airway remodelling. An understanding of these mechanisms might help to explain the protective effects of caffeine in prevention of BPD and suggests rolipram to be a potent replacement for caffeine.

Project description:BackgroundAlthough caffeine and glucocorticoids are frequently used to treat chronic lung disease in preterm neonates, potential interactions are largely unknown. While anti-inflammatory effects of glucocorticoids are well defined, their impact on airway remodeling is less characterized. Caffeine has been ascribed to positive effects on airway inflammation as well as remodeling. Connective tissue growth factor (CTGF, CCN2) plays a key role in airway remodeling and has been implicated in the pathogenesis of chronic lung diseases such as bronchopulmonary dysplasia (BPD) in preterm infants. The current study addressed the impact of glucocorticoids on the regulation of CTGF in the presence of caffeine using human lung epithelial and fibroblast cells.MethodsThe human airway epithelial cell line H441 and the fetal lung fibroblast strain IMR-90 were exposed to different glucocorticoids (dexamethasone, budesonide, betamethasone, prednisolone, hydrocortisone) and caffeine. mRNA and protein expression of CTGF, TGF-β1-3, and TNF-α were determined by means of quantitative real-time PCR and immunoblotting. H441 cells were additionally treated with cAMP, the adenylyl cyclase activator forskolin, and the selective phosphodiesterase (PDE)-4 inhibitor cilomilast to mimic caffeine-mediated PDE inhibition.ResultsTreatment with different glucocorticoids (1 μM) significantly increased CTGF mRNA levels in H441 (p < 0.0001) and IMR-90 cells (p < 0.01). Upon simultaneous exposure to caffeine (10 mM), both glucocorticoid-induced mRNA and protein expression were significantly reduced in IMR-90 cells (p < 0.0001). Of note, 24 h exposure to caffeine alone significantly suppressed basal expression of CTGF mRNA and protein in IMR-90 cells. Caffeine-induced reduction of CTGF mRNA expression seemed to be independent of cAMP levels, adenylyl cyclase activation, or PDE-4 inhibition. While dexamethasone or caffeine treatment did not affect TGF-β1 mRNA in H441 cells, increased expression of TGF-β2 and TGF-β3 mRNA was detected upon exposure to dexamethasone or dexamethasone and caffeine, respectively. Moreover, caffeine increased TNF-α mRNA in H441 cells (6.5 ± 2.2-fold, p < 0.05) which has been described as potent inhibitor of CTGF expression.ConclusionsIn addition to well-known anti-inflammatory features, glucocorticoids may have adverse effects on long-term remodeling by TGF-β1-independent induction of CTGF in lung cells. Simultaneous treatment with caffeine may attenuate glucocorticoid-induced expression of CTGF, thereby promoting restoration of lung homeostasis.

Dataset Information

TGF-? Induced CTGF Expression in Human Lung Epithelial Cells through ERK, ADAM17, RSK1, and C/EBP? Pathways.

Publications

TGF-β Induced CTGF Expression in Human Lung Epithelial Cells through ERK, ADAM17, RSK1, and C/EBPβ Pathways.

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