title: Regulation of Cdc42 expression and signaling is critical for promoting corneal epithelial wound healing.
abstract: OBJECTIVE
  Cdc42, a member of Rho GTPases (guanosine triphosphatases), participates in cytokine- and growth factor-controlled biological functions in mammalian tissues. Here, we examined Cdc42 role in corneal epithelial wound healing and the influence of hepatocyte, keratinocyte, and epidermal growth factor (HGF, KGF, and EGF)-mediated signaling on Cdc42.
  
  
  METHODS
  Epithelial wounds were created on the corneas of live rabbits by complete debridement and in rabbit corneal epithelial primary cultures through scratch injury. Cdc42 expression in cultures was suppressed using Cdc42 siRNA. Cdc42 activation was determined by pull-down assays with PAK-agarose beads. Cdc42 expression was analyzed by immunoblotting and immunofluorescence. Association of Cdc42 with cell-cycle proteins was identified by immunoprecipitation.
  
  
  RESULTS
  In rabbit corneas, significant increase in Cdc42 expression that occurred 2 to 4 days after the injury coincided with wound closure, and by 8 days the expression reached near basal levels. Silencing of Cdc42 expression in cultures caused inhibition of wound closure as a result of 60% to 75% decrease in epithelial migration and growth. HGF, KGF, and EGF increased Cdc2 expression, activation, and its phosphorylation on ser71. Inhibition of growth factor-mediated PI-3K signaling resulted in the downregulation of Cdc42 expression and its phosphorylation. Increased association of cell-cycle proteins p27(kip) and cyclin-dependent kinase 4 (CDK4) with Cdc42; and phosphorylated Cdc42 with plasma membrane leading edges was also observed in the presence of growth factors.
  
  
  CONCLUSIONS
  Cdc42 is an important regulator of corneal epithelial wound repair. To promote healing, Cdc42 may interact with receptor tyrosine kinase-activated signaling cascades that participate in cell migration and cell-cycle progression.
authors.affiliation: Department of Pharmaceutical Sciences, College of Pharmacy, South Dakota State University, Brookings, South Dakota 57007, USA.
authors.forename: Swetha
authors.initials: S
authors.lastname: Pothula
authors.forename: Haydee E P
authors.initials: HE
authors.lastname: Bazan
authors.forename: Gudiseva
authors.initials: G
authors.lastname: Chandrasekher
mesh_terms: D000818:Animals; D015153:Blotting, Western; D002465:Cell Movement; D002478:Cells, Cultured; D004591:Electrophoresis, Polyacrylamide Gel; D019573:Epithelium, Corneal / Q000201:enzymology / Q000293:injuries* / Q000473:pathology; D005131:Eye Injuries / Q000201:enzymology* / Q000235:genetics / Q000473:pathology; D005786:Gene Expression Regulation; D034741:RNA, Small Interfering / Q000235:genetics; D011817:Rabbits; D015398:Signal Transduction / Q000235:genetics; D014945:Wound Healing / Q000235:genetics*; D020764:cdc42 GTP-Binding Protein / Q000096:biosynthesis / Q000235:genetics*
chemical_list: D034741:RNA, Small Interfering; D020764:cdc42 GTP-Binding Protein
keywords: Cdc42; HGF; KGF; Rho-GTPases