Ligand-activated PPARδ upregulates α-smooth muscle actin expression in human dermal fibroblasts: A potential role for PPARδ in wound healing

Background: The phenotypic changes that accompany differentiation of resident fibroblasts into myofibroblasts are important aspects of the wound healing process. Recent studies showed that peroxisome proliferator-activated receptor (PPAR) delta plays a critical role in wound healing. Objective: To determine whether the nuclear receptor PPAR delta can modulate the differentiation of human dermal fibroblasts (HDFs) into myofibroblasts. Methods: These studies were undertaken in primary HDFs using Western blot analyses, small interfering (si)RNA-mediated gene silencing, reporter gene assays, chromatin immunoprecipitation (Chip), migration assays, collagen gel contraction assays, and real-time PCR. Results: Activation of PPAR delta by GW501516, a specific ligand of PPAR delta, specifically upregulated the myofibroblast marker alpha-smooth muscle actin (alpha-SMA) in a time- and concentration-dependent manner. This induction was significantly inhibited by the presence of siRNA against PPAR delta, indicating that PPAR delta is involved in myofibroblast transdifferentiation of HDFs. Ligand-activated PPAR delta increased alpha-SMA promoter activity in a dual mode by directly binding a direct repeat-1 (DR1) site in the alpha-SMA promoter, and by inducing expression of transforming growth factor (TGF)-beta, whose downstream effector Smad3 interacts with a Smad-binding element (SBE) in another region of the promoter. Mutations in these cis-elements totally abrogated transcriptional activation of the alpha-SMA gene by the PPAR delta ligand; thus both sites represent novel types of PPAR delta response elements. GW501516-activated PPAR delta also increased the migration and contractile properties of HDFs, as demonstrated by Transwell and collagen lattice contraction assays, respectively. In addition, PPAR delta-mediated upregulation of alpha-SMA was correlated with elevated expression of myofibroblast markers such as collagen I and fibronectin, with a concomitant reduction in expression of the epithelial marker E-cadherin. Conclusion: PPAR delta plays pivotal roles in wound healing by promoting fibroblast-to-myofibroblast differentiation via TGF-beta/Smad3 signaling. (C) 2015 Japanese Society for Investigative Dermatology. Published by Elsevier Ireland Ltd. All rights reserved.