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U. Milenkovic, M. Ilg, C. Zuccato, Y. Ramazani, D. De Ridder, M. Albersen, HP-01-001 Simvastatin and the Rho-kinase Inhibitor Y-27632 Prevent Myofibroblast Transformation in Peyronie’s Disease-derived Fibroblasts via Inhibition of YAP/TAZ Nuclear Translocation., The Journal of Sexual Medicine, Volume 16, Issue Supplement_2, May 2019, Page S30, https://doi.org/10.1016/j.jsxm.2019.03.116
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Objective
To uncover anti-myofibroblast (MFB) properties of Rho-kinase inhibitor (compound Y-27632) and simvastatin in an in vitro Peyronie’s disease model. It is a sexually debilitating disease caused by an irreversible fibrotic plaque in the penile tunica albuginea (TA). Treatment is limited to surgically restoring anatomical shape. Evidence for effective medical treatment is lacking.
Methods
Human fibroblasts were isolated from surgically obtained TA-tissue from PD-patients. To induce MFB status, cells where stimulated with 3ng/mL TGF-β1. Increasing doses of Y-27632 and simvastatin were added. RT-qPCR was used to assess mRNA expression of alpha-smooth muscle actin (α-SMA), collagenIII, elastin and CTGF after 72 h. WB was used to quantify α-SMA protein contents and IF visualized MFB differentiation by staining for α-SMA after 72 h. Resazurin-based assay was performed to assess cell viability. A mechanistic study was performed using IF staining for YAP/TAZ nuclear translocation.
Results
After 72 h of stimulation a 6-to10-fold upregulation of α-SMA could be observed. When treated with Y-27632 and simvastatin, the α-SMA, collagen III, elastin and CTGF mRNA expression was impeded. Additionally, it showed a twofold increase in α-SMA protein expression, which was reversed to non-stimulated levels after both treatments. Using IF, stimulated cells were identified as MFB (α-SMA+, Vim+) as opposed to the non-stimulated, Y-27632- and simvastatin-treated cells (α-SMA-, Vim+). The resazurin-based assay confirmed that cell viability was uncompromised when administering the drugs. Upon stimulation with TGF-β1 there was a nuclear translocation of YAP/TAZ, which was prevented by adding respective compounds.
