title: Electrophilic PPARγ ligands inhibit corneal fibroblast to myofibroblast differentiation in vitro: a potentially novel therapy for corneal scarring.
abstract: A critical component of corneal scarring is the TGFβ-induced differentiation of corneal keratocytes into myofibroblasts. Inhibitors of this differentiation are potentially therapeutic for corneal scarring. In this study, we tested the relative effectiveness and mechanisms of action of two electrophilic peroxisome proliferator-activated receptor gamma (PPARγ) ligands: cyano-3,12-dioxolean-1,9-dien-28-oic acid-methyl ester (CDDO-Me) and 15-deoxy-Δ(-12,14)-prostaglandin J(2) (15d-PGJ(2)) for inhibiting TGFβ-induced myofibroblast differentiation in vitro. TGFβ was used to induce myofibroblast differentiation in cultured, primary human corneal fibroblasts. CDDO-Me and 15d-PGJ(2) were added to cultures to test their ability to inhibit this process. Myofibroblast differentiation was assessed by measuring the expression of myofibroblast-specific proteins (αSMA, collagen I, and fibronectin) and mRNA (αSMA and collagen III). The role of PPARγ in the inhibition of myofibroblast differentiation by these agents was tested in genetically and pharmacologically manipulated cells. Finally, we assayed the importance of electrophilicity in the actions of these agents on TGFβ-induced αSMA expression via Western blotting and immunofluorescence. Both electrophilic PPARγ ligands (CDDO-Me and 15d-PGJ(2)) potently inhibited TGFβ-induced myofibroblast differentiation, but PPARγ was only partially required for inhibition of myofibroblast differentiation by either agent. Electrophilic PPARγ ligands were able to inhibit myofibroblast differentiation more potently than non-electrophilic PPARγ ligands, suggesting an important role of electrophilicity in this process. CDDO-Me and 15d-PGJ(2) are strong inhibitors of TGFβ-induced corneal fibroblast to myofibroblast differentiation in vitro, suggesting this class of agents as potential novel therapies for corneal scarring warranting further study in pre-clinical animal models.
authors.affiliation: University of Rochester School of Medicine and Dentistry, University of Rochester, Rochester, NY, United States.
authors.forename: A E
authors.initials: AE
authors.lastname: Kuriyan
authors.forename: G M
authors.initials: GM
authors.lastname: Lehmann
authors.forename: A A
authors.initials: AA
authors.lastname: Kulkarni
authors.forename: C F
authors.initials: CF
authors.lastname: Woeller
authors.forename: S E
authors.initials: SE
authors.lastname: Feldon
authors.forename: H B
authors.initials: HB
authors.lastname: Hindman
authors.forename: P J
authors.initials: PJ
authors.lastname: Sime
authors.forename: K R
authors.initials: KR
authors.lastname: Huxlin
authors.forename: R P
authors.initials: RP
authors.lastname: Phipps
mesh_terms: D000199:Actins / Q000235:genetics / Q000378:metabolism; D015415:Biomarkers / Q000378:metabolism; D015153:Blotting, Western; D054338:Cell Transdifferentiation / Q000187:drug effects*; D002478:Cells, Cultured; D024042:Collagen Type I / Q000235:genetics / Q000378:metabolism; D003315:Cornea / Q000166:cytology* / Q000378:metabolism; D005347:Fibroblasts / Q000166:cytology* / Q000378:metabolism; D005353:Fibronectins / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D008024:Ligands; D058628:Myofibroblasts / Q000166:cytology* / Q000378:metabolism; D009828:Oleanolic Acid / Q000031:analogs & derivatives* / Q000494:pharmacology; D047495:PPAR gamma / Q000378:metabolism*; D015230:Prostaglandin D2 / Q000031:analogs & derivatives* / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D016212:Transforming Growth Factor beta / Q000494:pharmacology
chemical_list: C477819:15-deoxyprostaglandin J2; C541116:ACTA2 protein, human; D000199:Actins; D015415:Biomarkers; D024042:Collagen Type I; D005353:Fibronectins; D008024:Ligands; D047495:PPAR gamma; D012333:RNA, Messenger; D016212:Transforming Growth Factor beta; D009828:Oleanolic Acid; C445068:bardoxolone methyl; D015230:Prostaglandin D2