Hirudin, a thrombin inhibitor, attenuates TGF-β-induced fibrosis in renal proximal tubular epithelial cells by inhibition of protease-activated receptor 1 expression via S1P/S1PR2/S1PR3 signaling

Lin,Qiang; Long,Chunli; Wang,Zhengang; Wang,Ronghui; Shi,Wei; Qiu,Jiwei; Mo,Junlin; Xie,Yongxiang

doi:10.3892/etm.2021.10924

Hirudin, a thrombin inhibitor, attenuates TGF-β-induced fibrosis in renal proximal tubular epithelial cells by inhibition of protease-activated receptor 1 expression via S1P/S1PR2/S1PR3 signaling

Authors:
- Qiang Lin
- Chunli Long
- Zhengang Wang
- Ronghui Wang
- Wei Shi
- Jiwei Qiu
- Junlin Mo
- Yongxiang Xie
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Affiliations: Department of Nephrology, The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning, Guangxi 530023, P.R. China, Department of Health Maintenance, Faculty of Preclinical Medicine, Guangxi University of Chinese Medicine, Nanning, Guangxi 530001, P.R. China, Department of Endocrinology, The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning, Guangxi 530023, P.R. China, Department of Critical Care Medicine, The First Affiliated Hospital of Guangxi University of Chinese Medicine, Nanning, Guangxi 530023, P.R. China
Published online on: October 26, 2021 https://doi.org/10.3892/etm.2021.10924
Article Number: 3
Copyright: © Lin et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Renal interstitial fibrosis (RIF) is the final common outcome of numerous chronic kidney diseases, contributing to end‑stage renal disease. Hirudin, a thrombin inhibitor, has attracted increased attention as a potential treatment approach for renal fibrosis. The present study aimed to investigate the molecular mechanism underlying the effect of hirudin on fibrosis in renal proximal tubular epithelial cells. An in vivo mouse RIF model established using unilateral ureteral obstruction (UUO) and an in vitro of RIF using the renal tubular epithelial cell line HK‑2 treated with TGF‑β were used. Expressions of sphingosine‑1‑phosphate (S1P) receptors (S1PR)1‑4 and protease‑activated receptor 1 (PAR1) were measured by reverse transcription‑quantitative PCR and western blotting in mice with UUO and TGF‑β induced HK‑2 cells. Western blotting was used to detect the expression of N‑cadherin, Slug, E‑cadherin, Collagen IV, fibronectin, MMP9 and monocyte chemoattractant protein‑1. Immunofluorescence staining was conducted to measure α‑SMA level expression. The results demonstrated that the expression levels of S1PR1, S1PR2, S1PR3, S1PR4 and PAR1 were upregulated in both TGF‑β‑induced HK‑2 cells and renal tissues from mice with unilateral ureteral ligation. Notably, hirudin inhibited TGF‑β‑induced PAR1, S1PR2 and S1PR3 upregulation in both HK‑2 cells and renal tissues. Additionally, the inhibition of S1PR2 and S1PR3 resulted in PAR1 downregulation. Furthermore, treatment with S1P and PAR1 agonists abolished the effect of hirudin on the expression of EMT, fibrosis‑related proteins and monocyte chemoattractant protein 1. In conclusion, hirudin attenuated TGF‑β‑induced fibrosis in proximal renal tubular epithelial HK‑2 cells by inhibiting PAR1 expression via the S1P/S1PR2/S1PR3 signaling pathway. Therefore, hirudin may be considered as a promising therapeutic agent for RIF.

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