A Rho kinase inhibitor (Fasudil) suppresses TGF-β mediated autophagy in urethra fibroblasts to attenuate traumatic urethral stricture (TUS) through re-activating Akt/mTOR pathway: An in vitro study.

Huan Feng, Xiaobing Huang, Weihua Fu, Xingyou Dong, Fengxia Yang, Longkun Li, Lingling Chu

Life sciences 2021 Feb 15

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Transforming growth factor-β (TGF-β) mediated super-activation of urethra fibroblasts contributes to the progression of traumatic urethral stricture (TUS), and the Rho-associated kinase inhibitors, Fasudil, might be a novel therapeutic agent for TUS, but the underlying mechanisms had not been studied. The primary urethral fibroblasts (PUFs) were isolated from rabbit urethral scar tissues and cultured in vitro, and the PUFs were subsequently treated with TGF-β (10 μg/L) to simulate the realistic conditions of TUS pathogenesis. Next, the PUFs were exposed to Fasudil (50 μM) and autophagy inhibitor 3-methyladenine (3-MA) treatment. Genes expression was examined by Western Blot and immunofluorescence staining, and cellular functions were determined by MTT assay and Transwell assay. TGF-β promoted cell proliferation, migration, autophagy, and secretion of extracellular matrix (ECM), including collagen I and collagen III, which were reversed by co-treating cells with both Fasudil and 3-MA. In addition, TGF-β treatment decreased the expression levels of phosphorylated Akt (p-Akt) and mTOR (p-mTOR) to inactivate the Akt/mTOR pathway in the PUFs, which could be re-activated by Fasudil. Then, the fibroblasts were treated with the Pan-Akt inhibitor (GDC-0068), and we surprisingly found that GDC-0068 abrogated the inhibiting effects of Fasudil on cell autophagy and proliferation in the PUFs treated with TGF-β. Fasudil regulated Akt/mTOR pathway mediated autophagy to hamper TGF-β-mediated super-activation in PUFs, which supported that Fasudil might be an ideal candidate therapeutic agent for TUS treatment for clinical utilization. Copyright © 2020 Elsevier Inc. All rights reserved.

Citation

Huan Feng, Xiaobing Huang, Weihua Fu, Xingyou Dong, Fengxia Yang, Longkun Li, Lingling Chu. A Rho kinase inhibitor (Fasudil) suppresses TGF-β mediated autophagy in urethra fibroblasts to attenuate traumatic urethral stricture (TUS) through re-activating Akt/mTOR pathway: An in vitro study. Life sciences. 2021 Feb 15;267:118960