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Background/Aims: The epithelial-to-mesenchymal transition (EMT) of peritoneal mesothelial cells (PMCs) is a crucial event in the induction of peritoneal fibrosis (PF), in which canonical Wnt/β-catenin signaling participates. Smads signaling is reported to interact with β-catenin and synergistically regulates EMT. This study was aimed to reveal the effect of Astragalus on β-catenin in EMT of PMCs. Methods: To obtain the role of β-catenin in EMT, gene transfer into HMrSV5 cell line and rats has been achieved. After Astragalus treatment, EMT markers and signaling pathway-related indicators were detected by western blotting, immunofluorescence, immunohistochemistry, immunoprecipitation and real time-PCR. Results: β-catenin knockdown suppressed EMT of HMrSV5 cells. Astragalus alleviated EMT of PMCs characterized by increased E-cadherin and decreased α-SMA and Vimentin. In rat model of peritoneal dialysis (PD), Astragalus attenuated peritoneal thickening and fibrosis. Astragalus down-regulated β-catenin by stabilizing the Glycogen synthase kinase-3β (GSK-3β)/β-catenin complex and further inhibited the nuclear translocation of β-catenin. Meanwhile, Astragalus down-regulated β-catenin by enhancing Smad7 expression. Silencing Smad7 antagonized the EMT-inhibitory effect of Astragalus. Conclusion: Astragalus inhibits EMT of PMCs by down-regulating β-catenin. The modulation of β-catenin in peritoneum can be a novel tool to prevent PF. [ABSTRACT FROM AUTHOR]

Background/Aims: Peritoneal fibrosis is a frequent complication of peritoneal dialysis that follows inflammation. It is recognized that epithelial-mesenchymal transition (EMT) of peritoneal mesothelial cells (PMCs), plays a key role in fibrogenesis. However, the relationship between inflammatory macrophages and PMCs remains elusive. In this study, we investigated the effects of different polarized macrophages on EMT of HMrSV5 PMCs. Methods: Monocytes were polarized to M1/M2 macrophages before being added to HMrSV5 in direct or indirect contact. Morphological changes of HMrSV5 were observed and toll-like receptors 4 (TLR4) on macrophage surfaces was detected using flow cytometry. EMT markers and intracellular signals of HMrSV5 cells were assessed using real time-PCR and WB. Results: The typical epithelial cell morphology of HMrSV5 disappeared after co-culture with M1 macrophages and was accompanied by decreased E-cadherin and increased α-SMA, suggesting HMrSV5 undergo EMT. These effects depended on direct contact between the two cells, as indirect contact or co-culture with M2 macrophages had no effect. Intriguingly, we found TLR4 surface receptors were activated on sorted M1 cells in co-culture, and related signal adaptors, such as TRIF, were obviously upregulated. Conclusion: Direct contact with M1 macrophages induces EMT of PMCs, during which TRIF-dependent TLR4 signaling pathway was activated. [ABSTRACT FROM AUTHOR]

Background/Aims: To investigate the effect of Astragaloside IV (AS-IV) on the regulation of the TGF-β1/Smad signaling pathway in peritoneal mesothelial cells with an epithelial-to-mesenchymal transition (EMT). Methods: EMT of human peritoneal mesothelial cells (HMrSV5) was induced using 2 ng/ml TGF-β1. Cells were randomly divided into a vehicle group, a vehicle group with AS-IV, a TGF-β1 treated group, and a TGF-β1 treated group receiving varied doses of AS-IV or NAC. Real-time quantitative PCR and western blot were used to detect the expression of genes and proteins associated with the TGF-β1/Smad signaling pathway and EMT. DCFH-DA was used to detect the generation of ROS in HMrSV5 cells, and a transwell migration assay was used to verify the capacity of AS-IV to inhibit EMT in HMrSV5 cells. Lentiviruses were used as carriers for the overexpression or knockdown of the Smad7 gene. Results: Expression levels of E-cadherin (epithelial marker) was decreased and vimentin, α-SMA (EMT markers) and collagen I (extracellular matrix protein) phospho-Smad2/3, Snail1 and Snail2 was increased significantly in the TGF-β1-treated HMrSV5 cells. AS-IV was associated with downregulated expression of vimentin and phospho-Smad2/3 in a dose-dependent manner, while the expression of Smad7 increased. Silenced or forced expression of Smad7 verified its role in the inhibitory effect of AS-IV on TGF-β1-induced EMT in HMrSV5 cells. Conclusion: AS-IV effectively promotes the upregulation of Smad7 in the TGF-β1/Smad signaling pathway during the EMT of HMrSV5 cells, indicating its potential therapeutic effect for the control of PF. © 2015 S. Karger AG, Basel [ABSTRACT FROM AUTHOR]