Epigallocatechin-3-gallate prevents TGF-β1-induced epithelial-mesenchymal transition and fibrotic changes of renal cells via GSK-3β/β-catenin/Snail1 and Nrf2 pathways.

Several lines of evidence have demonstrated anti-fibrotic property of epigallocatechin-3-gallate (EGCG) in many tissues/organs but with unclear mechanisms. This study thus aimed to define cellular mechanisms underlying such protective effect of EGCG. HK-2 renal cells were treated with 5 ng/ml TGF-β1 for 24 h with/without pretreatment by 5 μM EGCG for 1 h. The cells were then evaluated by morphological examination, immunofluorescence study, semi-quantitative RT-PCR, Western blotting, and atomic force microscopy (AFM). The results showed that TGF-β1-treated cells underwent epithelial mesenchymal transition (EMT) as evidenced by morphological change into fibroblast-like and increases in spindle index, mesenchymal markers (Snail1 and vimentin), extracellular matrix (fibronectin), cell stiffness (by AFM measurement) and actin stress fibers, whereas the epithelial markers (E-cadherin and ZO-1) were decreased. All of these features were abolished by EGCG pretreatment. Functional studies revealed that the anti-fibrotic property of EGCG was, at least in part, due to de-activation/stabilization of GSK-3β/β-catenin/Snail1 (EMT-triggering) signaling pathway that was activated by TGF-β1 as shown by maintaining phosphorylated GSK-3β, β-catenin and Snail1 to their basal levels. Additionally, Nrf2 knockdown by small interfering RNA could abolish the EGCG effect on β-catenin expression. These data indicate that EGCG attenuates TGF-β1-induced EMT in renal tubular cells through GSK-3β/β-catenin/Snail1 and Nrf2 pathways.

Kanlaya R ，Peerapen P ，Nilnumkhum A ，Plumworasawat S ，Sueksakit K ，Thongboonkerd V ... -  《-》

Induction of mesenchymal-epithelial transition (MET) by epigallocatechin-3-gallate to reverse epithelial-mesenchymal transition (EMT) in SNAI1-overexpressed renal cells: A potential anti-fibrotic strategy.

Dynamic transdifferentiation of epithelial cells from epithelial-mesenchymal transition (EMT) to its reverse process, mesenchymal-epithelial transition (MET), has gained wide attention for management of cancers and tissue fibrosis. In this study, we addressed beneficial effects of epigallocatechin-3-gallate (EGCG) on EMT-MET reversion using an in vitro EMT model by overexpressing SNAI1 gene encoding Snail1, an EMT-inducing transcription factor, into renal tubular epithelial cells (pcDNA6.2-SNAI1 cells). The cells transfected with empty vector (pcDNA6.2 cells) served as the control. Titrating EGCG concentrations revealed its optimal dose at 25 µM for 24-h, which was used throughout. pcDNA6.2-SNAI1 cells had increased spindle index and typical morphology of EMT, whereas EGCG could restore the normal index and morphology. Increased nuclear Snail1 and β-catenin; increased cytoplasmic Snail1, p-GSK-3β, vimentin, fibronectin and F-actin; and decreased occludin, ZO-1, transepithelial resistance (TER), E-cadherin and cell cluster size were observed in the pcDNA6.2-SNAI1 cells. These pcDNA6.2-SNAI1 cells also had increased migrating activity associated with increased forward but decreased non-forward α-tubulin filaments, G0/G1 cell cycle escape, and increased matrix metalloproteinase-2 (MMP-2) and MMP-9. All of these EMT features were successfully abolished by EGCG (partially, completely, or overly). Collectively, our data have demonstrated that EGCG can reverse EMT to MET processes in renal cells. Therefore, EGCG may have the therapeutic potential as one of the promising anti-fibrotic agents to reverse the fibrotic kidney.

Kanlaya R ，Kapincharanon C ，Fong-Ngern K ，Thongboonkerd V ... -  《-》

Epigallocatechin-3-gallate attenuates transforming growth factor-β1 induced epithelial-mesenchymal transition via Nrf2 regulation in renal tubular epithelial cells.

Transforming growth factor-β1 (TGF-β1) induced epithelial-mesenchymal transition (EMT) plays an important role in renal fibrotic process regulation. Epigallocatechin-3-gallate (EGCG) exerts a protective effect against acute renal damage through its anti-oxidative effect by activating the Nrf2 signaling pathway. This study aims to investigate whether EGCG prevents TGF-β1 induced EMT and whether this effect acts via the Nrf2-mediated suppression of TGF-β1 signaling. MTT was used for cytotoxicity of EGCG examination and Western blotting and immunofluorescence were used for protein expression analysis. Results showed that EGCG prevented TGF-β1 mediated EMT and Smad 2 and Smad 3 phosphorylation in a dose dependent manner in NRK-52E cells. In addition, EGCG increased Nrf2 nuclear accumulation. Overexpression of Nrf2 blocked the phosphorylation of Smad 2 and Smad 3 mediated by TGF-β1 and decreased protein expression of plasminogen activator inhibitor 1 (PAI-1) and α-smooth muscle actin (α-SMA). Furthermore, siRNA-mediated knockdown of Nrf2 gene completely blocked the effects of EGCG, indicated by the reduced expressions of type I collagen (Col-I) and α-SMA were restored. In summary, EGCG inhibits TGF-β1 induced EMT and fibrotic proteins expression by Nrf2 activation. This study reveals a possible underlying mechanism of the renal protective effects of EGCG, and may provide a potential candidate to renal fibrosis therapy.

Wang Y ，Liu N ，Su X ，Zhou G ，Sun G ，Du F ，Bian X ，Wang B ... -  《-》

Protective effect of epigallocatechin-3-gallate (EGCG) via Nrf2 pathway against oxalate-induced epithelial mesenchymal transition (EMT) of renal tubular cells.

Kanlaya R ，Khamchun S ，Kapincharanon C ，Thongboonkerd V ... -  《Scientific Reports》

Troglitazone ameliorates high glucose-induced EMT and dysfunction of SGLTs through PI3K/Akt, GSK-3β, Snail1, and β-catenin in renal proximal tubule cells.

Lee YJ ，Han HJ 《-》