MOLECULAR ASPECTS OF CORNEAL EPITHELIAL MIGRATION

Summary

Principal Investigator: MARY STEPP
Affiliation: George Washington University
Country: USA
Abstract: The corneal epithelium has a variety of cell:cell and cell:substrate contacts which maintain much of its integrity even as it migrates in response to injury. Among the cell:substrate junctions, the hemidesmosomes (HDs) are responsible for most of the adhesion of the epithelium to its substrate. HDs contain the alpha6beta4 integrin as one of their components and other integrin heterodimers are present in cell:cell boundaries of the corneal epithelium. Aim 1 of this proposal is to determine whether the expression and structure of beta1, beta4, alpha3, alpha5 and alpha6 integrin mRNAs are altered during migration by a) amplifying rat integrin cDNAs from a rat corneal epithelial cell library, b) determining if the expression of rat integrin mRNAs is altered during migration, c) determining the relative abundance of the cytoplasmic integrin mRNA alternative splicing variants and whether there are changes during migration, and d) correlating the data on integrin mRNA expression with data on integrin protein synthesis. Aim 2 is to determine if the delay in corneal epithelial migration induced by addition of extracts from rat polymorphonuclear leukocytes (PMNs) to debridement wounded corneal organ cultures involves alterations in the amounts or the localization of integrins by a) using immunoblotting, immunoprecipitation, and mRNA quantitation to discover if the PMN extract added to debridement wounds affects protein synthesis in the epithelium by determining the level of integrins, vinculin, alpha-actin, alpha- enolase, and ICAM-1 and b) determining if the addition of purified inflammatory cytokines to corneal organ cultures with epithelial debridement wounds results in a delay in epithelial migration by a mechanism similar to PMN extract. Aim 3 is to determine whether the disassembly of hemidesmosomes and migration of the corneal epithelium involves phosphorylation and/or proteolytic cleavage of the beta4 subunit by a) determining whether the HD alpha6 and beta4 subunits are phosphorylated on tyrosine in the unwounded cornea and if their phosphorylation state is altered during migration, b) developing polyclonal antisera with specificity against either the entire extracellular domain or the entire cytoplasmic portion of the beta4 molecule, and c) using the antisera to determine if the beta4 molecule undergoes cleavage during epithelial cell migration. Aim 4 is to determine if integrins at regions of cell:cell interaction are functionally important in maintaining cell:cell contacts in the corneal epithelium by a) establishing cell culture conditions for bovine corneal epithelial cells, b) determining the state of assembly of desmosomes, adherins junctions, and alphav- and the beta1-containing cell:cell junctions in cells cultured in low calcium and after shifting cells to high calcium medium, c) determining the effect of adhesion blocking integrin peptides and antibodies on the ability of cultured corneal epithelial cells to maintain cell:cell contact in low and in high calcium media, and d) determining whether addition of PMN extract to cultured cells disrupts cell:cell contacts and the cell:cell localization of alphav and the beta1 integrins in low and high calcium media. These experiments will help accomplish our goal of obtaining a better understanding, at the molecular level, of the role of integrins in the normal cornea and in epithelial cell migration during wound healing.
Funding Period: 1992-07-01 - 1998-04-30
more information: NIH RePORT

Top Publications

  1. ncbi Effect of wound type on Smad 2 and 4 translocation
    Audrey E K Hutcheon
    Schepens Eye Research Institute and Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts 02114, USA
    Invest Ophthalmol Vis Sci 46:2362-8. 2005
  2. pmc Regulation by P2X7: epithelial migration and stromal organization in the cornea
    Courtney Mayo
    Departments of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118, USA
    Invest Ophthalmol Vis Sci 49:4384-91. 2008
  3. pmc BALB/c and C57BL6 mouse strains vary in their ability to heal corneal epithelial debridement wounds
    Sonali Pal-Ghosh
    Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, Washington, DC 20037, USA
    Exp Eye Res 87:478-86. 2008
  4. pmc Primary dermal fibroblasts derived from sdc-1 deficient mice migrate faster and have altered alphav integrin function
    Rosalyn A Jurjus
    Department of Anatomy and Regenerative Biology, George Washington University Medical School, 2300 I Street NW, Washington, DC 20037, USA
    Wound Repair Regen 16:649-60. 2008
  5. pmc Cytokine deposition alters leukocyte morphology and initial recruitment of monocytes and γδT cells after corneal injury
    Sonali Pal-Ghosh
    Department of Anatomy and Regenerative Biology and Department of Ophthalmology, The George Washington University Medical School, Washington, DC, United States
    Invest Ophthalmol Vis Sci 55:2757-65. 2014
  6. pmc Wounding the cornea to learn how it heals
    Mary Ann Stepp
    Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, Washington, DC 20037, USA Department of Ophthalmology, The George Washington University Medical Center, Washington, DC 20037, USA Electronic address
    Exp Eye Res 121:178-93. 2014
  7. pmc A central role for vimentin in regulating repair function during healing of the lens epithelium
    A S Menko
    Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, PA 19107 Wills Vision Research Center at Jefferson, Philadelphia, PA 19107 Department of Anatomy and Regenerative Biology, George Washington University, Washington, DC 20037
    Mol Biol Cell 25:776-90. 2014
  8. pmc Corneal goblet cells and their niche: implications for corneal stem cell deficiency
    Ahdeah Pajoohesh-Ganji
    Department of Anatomy and Regenerative Biology, The George Washington University Medical School, Washington, District of Columbia 20037, USA
    Stem Cells 30:2032-43. 2012
  9. pmc Spontaneous skin erosions and reduced skin and corneal wound healing characterize CLIC4(NULL) mice
    V C Padmakumar
    Laboratory of Cancer Biology and Genetics, National Cancer Institute, Bethesda, MD 20892, USA
    Am J Pathol 181:74-84. 2012
  10. pmc Removal of the basement membrane enhances corneal wound healing
    Sonali Pal-Ghosh
    The George Washington University Medical Center, Department of Anatomy and Regenerative Biology, Washington, DC 20037, USA
    Exp Eye Res 93:927-36. 2011

Scientific Experts

  • MARY STEPP
  • Sonali Pal-Ghosh
  • Gauri Tadvalkar
  • Ahdeah Pajoohesh-Ganji
  • Rosalyn A Jurjus
  • James D Zieske
  • A S Menko
  • V C Padmakumar
  • Brajendra K Tripathi
  • Courtney Mayo
  • Audrey E K Hutcheon
  • Hye Young Oh
  • Daniel R Saban
  • L Zhang
  • A Sue Menko
  • B M Bleaken
  • J L Walker
  • A A Libowitz
  • Katelyn E Masiuk
  • Andrew Ryscavage
  • Kelsey Speer
  • Lino Tessarollo
  • Vincenzo Coppola
  • Shelly Hwang
  • John C Edwards
  • Samuel L Dengler
  • Stuart H Yuspa
  • Tomas Blanco
  • Arpitha Parthasarathy
  • Vickery Trinkaus-Randall
  • Ruiyi Ren
  • Chun Y Gao
  • Yueyuan Liu
  • Celeste Rich
  • Peggy S Zelenka
  • Xiaoqing Q Guo
  • Paul H Weinreb
  • Kenneth J Simon
  • Shelia M Violette

Detail Information

Publications15

  1. ncbi Effect of wound type on Smad 2 and 4 translocation
    Audrey E K Hutcheon
    Schepens Eye Research Institute and Department of Ophthalmology, Harvard Medical School, Boston, Massachusetts 02114, USA
    Invest Ophthalmol Vis Sci 46:2362-8. 2005
    ..The present study was undertaken to examine whether Smads 2 and 4 translocate into the nucleus during wound repair and whether the wound type affects the extent of translocation...
  2. pmc Regulation by P2X7: epithelial migration and stromal organization in the cornea
    Courtney Mayo
    Departments of Biochemistry, Boston University School of Medicine, Boston, Massachusetts 02118, USA
    Invest Ophthalmol Vis Sci 49:4384-91. 2008
    ..The goal here was to characterize the role of the P2X(7) receptor in the repair of in vivo corneal epithelial debridement wounds and in the structural organization of the corneal stroma...
  3. pmc BALB/c and C57BL6 mouse strains vary in their ability to heal corneal epithelial debridement wounds
    Sonali Pal-Ghosh
    Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, Washington, DC 20037, USA
    Exp Eye Res 87:478-86. 2008
    ..These data prove that strain-specific differences in cell migration rate in vivo are present in the cornea and are accompanied by differences in the frequencies of recurrent erosions and corneal epithelial stem cell deficiency...
  4. pmc Primary dermal fibroblasts derived from sdc-1 deficient mice migrate faster and have altered alphav integrin function
    Rosalyn A Jurjus
    Department of Anatomy and Regenerative Biology, George Washington University Medical School, 2300 I Street NW, Washington, DC 20037, USA
    Wound Repair Regen 16:649-60. 2008
    ....
  5. pmc Cytokine deposition alters leukocyte morphology and initial recruitment of monocytes and γδT cells after corneal injury
    Sonali Pal-Ghosh
    Department of Anatomy and Regenerative Biology and Department of Ophthalmology, The George Washington University Medical School, Washington, DC, United States
    Invest Ophthalmol Vis Sci 55:2757-65. 2014
    ..Here, we characterize differences in cytokine deposition and changes in leukocytes between 0 and 6 hours after dulled-blade and rotating-burr wounding...
  6. pmc Wounding the cornea to learn how it heals
    Mary Ann Stepp
    Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, Washington, DC 20037, USA Department of Ophthalmology, The George Washington University Medical Center, Washington, DC 20037, USA Electronic address
    Exp Eye Res 121:178-93. 2014
    ..The subject of corneal wound healing is broad and includes chemical and mechanical wound models. This review focuses on mechanical injury models involving debridement and keratectomy wounds to reflect the authors' expertise. ..
  7. pmc A central role for vimentin in regulating repair function during healing of the lens epithelium
    A S Menko
    Department of Pathology, Anatomy and Cell Biology, Thomas Jefferson University, Philadelphia, PA 19107 Wills Vision Research Center at Jefferson, Philadelphia, PA 19107 Department of Anatomy and Regenerative Biology, George Washington University, Washington, DC 20037
    Mol Biol Cell 25:776-90. 2014
    ..These studies reveal a critical role for vimentin in repair cell function in regulating the collective movement of the epithelium in response to wounding. ..
  8. pmc Corneal goblet cells and their niche: implications for corneal stem cell deficiency
    Ahdeah Pajoohesh-Ganji
    Department of Anatomy and Regenerative Biology, The George Washington University Medical School, Washington, District of Columbia 20037, USA
    Stem Cells 30:2032-43. 2012
    ..This study is the first description of compound niches and corneal goblet cells and demonstration of a role for these cells in the pathology typically associated with corneal stem cell deficiency...
  9. pmc Spontaneous skin erosions and reduced skin and corneal wound healing characterize CLIC4(NULL) mice
    V C Padmakumar
    Laboratory of Cancer Biology and Genetics, National Cancer Institute, Bethesda, MD 20892, USA
    Am J Pathol 181:74-84. 2012
    ..These results indicate that CLIC4 participates in skin healing and corneal wound reepithelialization through enhancement of epithelial migration by a mechanism that may involve a compromised TGF-β pathway...
  10. pmc Removal of the basement membrane enhances corneal wound healing
    Sonali Pal-Ghosh
    The George Washington University Medical Center, Department of Anatomy and Regenerative Biology, Washington, DC 20037, USA
    Exp Eye Res 93:927-36. 2011
    ..Despite the fact that rotating-burr wounds do more damage to the cornea, fewer immune cells are recruited and significantly more wounds resolve completely...
  11. pmc MMP9 cleavage of the β4 integrin ectodomain leads to recurrent epithelial erosions in mice
    Sonali Pal-Ghosh
    The George Washington University Medical Center, Department of Anatomy and Regenerative Biology, Washington, DC 20037, USA
    J Cell Sci 124:2666-75. 2011
    ..This report is the first to show that β4 integrin associates with MMP9 and that its ectodomain is a target for cleavage by MMP9 in vivo under pathological conditions...
  12. pmc Syndecan-1 regulates cell migration and fibronectin fibril assembly
    Mary Ann Stepp
    Department of Anatomy and Regenerative Biology, George Washington University Medical Center, Washington DC 20037, USA
    Exp Cell Res 316:2322-39. 2010
    ..A better understanding of how different cell types regulate FN fibril formation via syndecans and integrins will lead to better treatments for scarring and fibrosis...
  13. pmc Loss of syndecan-1 is associated with malignant conversion in skin carcinogenesis
    Mary Ann Stepp
    Department of Anatomy and Regenerative Biology, The George Washington University Medical Center, Washington, District of Columbia, USA
    Mol Carcinog 49:363-73. 2010
    ..These data indicate that sdc-1 is important both early in the development of skin tumors and in progression of skin cancers suggesting that reduced expression of sdc-1 could be a useful marker for progression in neoplastic skin lesions...
  14. pmc The Cdk5 inhibitor olomoucine promotes corneal debridement wound closure in vivo
    Brajendra K Tripathi
    Laboratory of Molecular and Developmental Biology, National Eye Institute, National Institutes of Health, Bethesda, MD 20892, USA
    Mol Vis 14:542-9. 2008
    ..To investigate the effect of the Cdk5 inhibitor olomoucine on corneal debridement wound healing in vivo...
  15. ncbi Reduced migration, altered matrix and enhanced TGFbeta1 signaling are signatures of mouse keratinocytes lacking Sdc1
    Mary Ann Stepp
    Department of Anatomy and Cell Biology, George Washington University Medical School, Washington, DC 20037, USA
    J Cell Sci 120:2851-63. 2007
    ..Thus, our results identify TGFbeta1 signaling and Sdc1 expression as important factors regulating integrin surface expression, activity and migration in keratinocyte and provide new insight into the functions regulated by Sdc1...