June 2013
Volume 54, Issue 15
ARVO Annual Meeting Abstract  |   June 2013
Cdc42 expression increases during corneal epithelial wound healing in vivo and suppression of its expression delays wound closure
Author Affiliations & Notes
  • Swetha Pothula
    Pharmaceutical Sciences, South Dakota State University, Brookings, SD
  • Gudiseva Chandrasekher
    Pharmaceutical Sciences, South Dakota State University, Brookings, SD
    Internal Medicine, Sanford School of Medicine of University of South Dakota, Sioux Falls, SD
  • Haydee Bazan
    Neuroscience and Eye Center, LSU Health Sciences Center, New Orleans, LA
  • Footnotes
    Commercial Relationships Swetha Pothula, None; Gudiseva Chandrasekher, None; Haydee Bazan, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 2562. doi:
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      Swetha Pothula, Gudiseva Chandrasekher, Haydee Bazan; Cdc42 expression increases during corneal epithelial wound healing in vivo and suppression of its expression delays wound closure. Invest. Ophthalmol. Vis. Sci. 2013;54(15):2562.

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      © ARVO (1962-2015); The Authors (2016-present)

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Purpose: Growth factors as well as extracellular matrix (ECM) proteins secreted after injury to the corneal epithelium play a major role in regulating wound repair. Cdc42, a Rho family GTPase member is known to control wound healing-promoting cellular processes such as migration, cytokinesis and growth. Previous studies from our laboratory showed that HGF, KGF, EGF and ECM proteins modulate Cdc42-mediated signaling in corneal epithelial cells (ARVO meeting 2012). Here, we report the involvement of Cdc42 in wound repair.

Methods: Corneal epithelial wounds were made in rabbits by scraping the epithelium (complete de-epithelialization). Epithelium regenerated at 1, 2, 4 and 8 day post injury was harvested. Epithelial primary cultures from rabbit corneas were grown in DMEM-F12/10% FCS were transfected with Cdc42 siRNA. Scratch wounds to the cultures were made 24 hours after transfection with siRNA. For some studies epithelial cultures were treated with HGF, KGF or EGF (20ng/ml). Immunoprecipiatation and immunofluorescence were employed to determine Cdc42 association with cell cycle proteins and plasma membrane.

Results: Significant elevation in the expression of Cdc42 was observed in regenerating epithelium at 2-4 days post- injury. By 8 days after the injury Cdc42 expression reached to the basal levels. Suppression of Cdc42 levels by silencing its gene expression produced almost 80-90% inhibition in wound closure in scratch wounded cell cultures. Further, association of cell cycle controlling proteins cyclin A, CDK2 and CDK4 with Cdc42 was observed in epithelial cells. Interestingly, HGF, KGF and EGF caused increased association of cell cycle inhibitor p27kip with Cdc42. Growth factors also induced phosphorylation of Cdc42 on ser71 and the phosphorylated Cdc42 was found to be associated with the plasma membrane.

Conclusions: Increase in Cdc42 levels during corneal epithelial wound repair and suppression of wound closure upon silencing Cd42 expression indicate the requirement of Cdc42-mediated signaling for the promotion of wound healing. Phosphorylation of Cdc42 at ser71 and its translocation to the cell membrane could trigger cell migration. Further, growth factor promoted association of p27kip with Cdc42 could relieve the effect of p27kip on cell cycle arrest and sustain cell cycle progression which is essential for rapid new cell regeneration following injury.

Keywords: 482 cornea: epithelium • 765 wound healing • 543 growth factors/growth factor receptors  

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