title: Dasatinib affects focal adhesion and myosin regulation to inhibit matrix contraction by Müller cells.
abstract: Epiretinal membrane (ERM) contraction is associated with a variety of ocular diseases that cause macular dysfunction. Trans-differentiated Müller cells have been identified in ERMs, and have been implicated to be involved in the contractile process. In this study, we tested the effect of dasatinib, an FDA-approved tyrosine kinase inhibitor, on matrix contraction caused by Müller cells, and examined molecular mechanism of action. Type I collagen matrix contraction assays were used to examine the effect of drugs on matrix contraction by trans-differentiated Müller cells. Fluophore-conjugated phalloidin was used for the detection of actin cytoskeleton, and Western-blot analyses were carried out to examine protein expression and phosphorylation status. Dasatinib inhibited collagen matrix contraction by trans-differentiated Müller cells that was associated with decreased cell spreading and reduction of actomyosin stress fibers. Concomitantly, dasatinib-treated Müller cells had reduced phosphorylation of Src family kinase, paxillin, as well as myosin II light chain. Specific inhibitors of Rho/ROCK and myosin II confirmed the critical role played by this pathway in Müller cell contraction. Our data demonstrate that dasatinib significantly reduced matrix contraction by Müller cells via inhibition of focal adhesion, as well as actomyosin contraction.
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Louisville, 301 E. Muhammad Ali Blvd., Louisville, KY 40202, USA; Department of Ophthalmology, Tokyo Medical University, 6-7-1 Nishi Shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Rintaro
authors.initials: R
authors.lastname: Tsukahara
authors.affiliation: Department of Ophthalmology, Tokyo Medical University, 6-7-1 Nishi Shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Kazuhiko
authors.initials: K
authors.lastname: Umazume
authors.affiliation: Department of Ophthalmology, Tokyo Medical University, 6-7-1 Nishi Shinjuku, Shinjuku, Tokyo 160-0023, Japan.
authors.forename: Naoyuki
authors.initials: N
authors.lastname: Yamakawa
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Louisville, 301 E. Muhammad Ali Blvd., Louisville, KY 40202, USA.
authors.forename: Kevin
authors.initials: K
authors.lastname: McDonald
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Louisville, 301 E. Muhammad Ali Blvd., Louisville, KY 40202, USA.
authors.forename: Henry J
authors.initials: HJ
authors.lastname: Kaplan
authors.affiliation: Department of Ophthalmology and Visual Sciences, University of Louisville, 301 E. Muhammad Ali Blvd., Louisville, KY 40202, USA; Department of Biochemistry and Molecular Genetics, University of Louisville, 319 Abraham Flexner Way, Louisville, KY 40202, USA. Electronic address: shigeo.tamiya@louisville.edu.
authors.forename: Shigeo
authors.initials: S
authors.lastname: Tamiya
mesh_terms: D000818:Animals; D017209:Apoptosis; D002448:Cell Adhesion / Q000187:drug effects; D000069439:Dasatinib / Q000494:pharmacology*; D004195:Disease Models, Animal; D063928:Ependymoglial Cells / Q000378:metabolism* / Q000473:pathology; D005109:Extracellular Matrix / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D015151:Immunoblotting; D020287:In Situ Nick-End Labeling; D008268:Macular Degeneration / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D009218:Myosins / Q000096:biosynthesis / Q000235:genetics*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects; D022502:Stress Fibers / Q000187:drug effects / Q000378:metabolism; D013552:Swine
chemical_list: D047428:Protein Kinase Inhibitors; D009218:Myosins; D000069439:Dasatinib
keywords: Contraction; Dasatinib; Epiretinal membrane; Focal adhesion; Müller cells; Stress fibers