PMID- 30536599 OWN - NLM STAT- MEDLINE DCOM- 20191217 LR - 20191217 IS - 1464-410X (Electronic) IS - 1464-4096 (Linking) VI - 123 IP - 4 DP - 2019 Apr TI - Simvastatin and the Rho-kinase inhibitor Y-27632 prevent myofibroblast transformation in Peyronie's disease-derived fibroblasts via inhibition of YAP/TAZ nuclear translocation. PG - 703-715 LID - 10.1111/bju.14638 [doi] AB - OBJECTIVES: To uncover the anti-myofibroblast (MFB) properties of Rho-kinase inhibitor (compound Y-27632) and simvastatin in an in vitro model of Peyronie's disease (PD), a sexually debilitating disease caused by an irreversible fibrotic plaque in the penile tunica albuginea (TA). MATERIALS AND METHODS: Primary human fibroblasts (FBs) were isolated from surgically obtained TA tissue from patients with PD. To induce MFB status, cells were stimulated with 3 ng/mL transforming growth factor-beta1 (TGF-beta1). Increasing doses of Y-27632 and simvastatin were added. Real-time quantitative PCR was used to assess mRNA expression of alpha-smooth muscle actin (alpha-SMA), collagen III, elastin and connective tissue growth factor (CTGF) after 72 h. Western blot analysis was used to quantify alpha-SMA protein contents, and immunofluorescence (IF) was used to visualize MFB differentiation by staining for alpha-SMA after 72 h. A resazurin-based assay was used to assess cell viability to ensure the anti-MFB effect of the drugs. A mechanistic study was performed using IF staining for YAP/TAZ nuclear translocation. RESULTS: After 72 h of stimulation with TGF-beta1, a six- to 10-fold upregulation of alpha-SMA could be observed. When treated with Y-27632 or simvastatin, the alpha-SMA, collagen III, elastin and CTGF mRNA expression was impeded. Additionally, TGF-beta1 stimulation showed a twofold increase in alpha-SMA protein expression, which was reversed to non-stimulated levels after treatment with Y-27632 and simvastatin. Using IF, stimulated cells were identified as MFB (alpha-SMA+, Vim+) as opposed to the non-stimulated, Y-27632- and simvastatin-treated cells (alpha-SMA-, Vim+). The resazurin-based assay confirmed that the cell viability was not compromised by the administered drugs. On stimulation with TGF-beta1, nuclear translocation of YAP/TAZ could be observed, which was prevented by adding the aforementioned compounds. CONCLUSION: Transformation of FBs into the contractile and extracellular matrix-producing MFBs occurs after TGF-beta1 stimulation. In our experiments, Rho-kinase inhibition and simvastatin treatment were shown to prevent this in TGF-beta1-stimulated cells on an RNA and protein level through the inhibition of YAP/TAZ nuclear translocation. Y-27632 and simvastatin could become a novel treatment option in the early treatment of PD. CI - (c) 2018 The Authors BJU International (c) 2018 BJU International Published by John Wiley & Sons Ltd. FAU - Milenkovic, Uros AU - Milenkovic U AUID- ORCID: 0000-0002-0740-2534 AD - Laboratory of Experimental Urology, Department of Development and Regeneration, KU Leuven, Leuven, Belgium. AD - Department of Urology, University Hospitals Leuven, Leuven, Belgium. FAU - Ilg, Marcus M AU - Ilg MM AD - Faculty of Health, Education, Medicine and Social Care, Medical Technology Research Centre, Anglia Ruskin University, Chelmsford, UK. FAU - Zuccato, Carola AU - Zuccato C AD - Laboratory of Experimental Urology, Department of Development and Regeneration, KU Leuven, Leuven, Belgium. AD - Faculty of Medicine and Surgery, University of Padua, Padua, Italy. FAU - Ramazani, Yasaman AU - Ramazani Y AD - Department of Pediatric Nephrology and Growth and Regeneration, University Hospitals Leuven and KU Leuven, Leuven, Belgium. FAU - De Ridder, Dirk AU - De Ridder D AD - Laboratory of Experimental Urology, Department of Development and Regeneration, KU Leuven, Leuven, Belgium. AD - Department of Urology, University Hospitals Leuven, Leuven, Belgium. FAU - Albersen, Maarten AU - Albersen M AD - Laboratory of Experimental Urology, Department of Development and Regeneration, KU Leuven, Leuven, Belgium. AD - Department of Urology, University Hospitals Leuven, Leuven, Belgium. LA - eng PT - Journal Article PT - Research Support, Non-U.S. Gov't DEP - 20190106 PL - England TA - BJU Int JT - BJU international JID - 100886721 RN - 0 (Amides) RN - 0 (Anticholesteremic Agents) RN - 0 (Intracellular Signaling Peptides and Proteins) RN - 0 (Pyridines) RN - 138381-45-0 (Y 27632) RN - AGG2FN16EV (Simvastatin) RN - EC 2.7.11.1 (rho-Associated Kinases) SB - IM MH - Amides/*pharmacology MH - Anticholesteremic Agents/*pharmacology MH - Cells, Cultured MH - Humans MH - Intracellular Signaling Peptides and Proteins MH - Male MH - Myofibroblasts/*pathology MH - Penile Induration/*pathology MH - Pyridines/*pharmacology MH - Real-Time Polymerase Chain Reaction MH - Simvastatin/*pharmacology MH - rho-Associated Kinases/*pharmacology OTO - NOTNLM OT - #Andrology OT - #Peyronies OT - Peyronie's disease OT - Rho-kinase inhibition OT - statins OT - transforming growth factor-beta EDAT- 2018/12/12 06:00 MHDA- 2019/12/18 06:00 CRDT- 2018/12/12 06:00 PHST- 2018/12/12 06:00 [pubmed] PHST- 2019/12/18 06:00 [medline] PHST- 2018/12/12 06:00 [entrez] AID - 10.1111/bju.14638 [doi] PST - ppublish SO - BJU Int. 2019 Apr;123(4):703-715. doi: 10.1111/bju.14638. Epub 2019 Jan 6.