Regular Article
Kinetics of Keratocyte Proliferation in Response to Epithelial Debridement

https://doi.org/10.1006/exer.2000.0926Get rights and content

Abstract

Over the past 40 years, several groups have shown that epithelial debridement results in the death of keratocytes subjacent to the wound area. More recently this cell death has been shown to involve apoptosis. The purpose of this project was to examine the proliferative response of the normally quiescent keratocytes to repopulate the apoptotic area. Three mm wounds were made in the central cornea of adult rats and allowed to heal 4 hr to 14 days. Cryostat sections were stained with propidium iodide to mark the nuclei of all cells. Actively proliferating cells were identified with anti-Ki67, a marker of the late G1-M phase of the cell cycle. Anti-α-smooth muscle actin was used to determine if myofibroblasts were present. In unwounded corneas, keratocytes were uniformly spread throughout the stroma, and less than one proliferating cell per mm was observed. By 4 hr after wounding, the anterior one-half to three-fourths of the stroma subjacent to the wound was devoid of cells. No increase in Ki67-expressing cells was observed in the stroma until 24 hr after wounding (3.9 ± 0.5 and 6.3 ± 0.5 mm−1in the wound center and edge, respectively). The number of Ki67-expressing cells steadily increased, peaking 44 hr after debridement (41.2 ± 1.7 and 39.6 ± 1.0). These cells were confined to a narrow zone adjacent to the area of cell death. No change in the number of cells expressing Ki67 was observed in the keratocytes distal to the original debridement. Ki67 levels did not return to control levels until 7 days after wounding. No α-smooth muscle actin was detected at any time point. This study indicates that epithelial debridement stimulates a synchronous increase in keratocyte proliferation. This stimulation is specific for cells immediately adjacent to the area of cell death. This activation does not involve the transformation of the stromal cells to a myofibroblast phenotype.

References (34)

  • R. Garrana et al.

    Radial keratotomy. II. Role of myofibroblast in corneal wound contraction

    Invest. Ophthalmol. Vis. Sci.

    (1992)
  • J. Gerdes et al.

    Cell cycle analysis of a cell proliferation-associated nuclear antigen that is defined by monoclonal antibody Ki-67

    J. Immunol.

    (1984)
  • J. Gerdes et al.

    Immunobiochemical and molecular biologic characterization of the cell proliferation-associated nuclear antigen that is defined by monoclonal antibody Ki-67

    Am. J. Pathol.

    (1991)
  • G.J. Gores et al.

    Extracellular acidosis delays onset of cell death in ATP-depleted hepatocytes

    Am. J. Physiol.

    (1988)
  • K.D. Hanna et al.

    Corneal stromal wound healing in rabbits after 193 nm excimer laser surface ablation

    Arch. Ophthalmol.

    (1989)
  • M.C. Helena et al.

    Keratocyte apoptosis after corneal surgery

    Invest. Ophthalmol. Vis. Sci.

    (1998)
  • Cited by (158)

    • The role of topical N-acetylcysteine in ocular therapeutics

      2022, Survey of Ophthalmology
      Citation Excerpt :

      This cytokine is responsible for upregulating Fas ligand, hepatocyte growth factor, keratocyte growth factor, platelet derived growth factor (PDGF), interleukin-6 (IL-6) and interleukin-8 (IL-8), as well as tumor necrosis factor alpha (TNF- α).40,58,59 When injury to the cornea disrupts the epithelial barrier, these secondary mediators have direct access to the stroma where they can bind to their respective receptors located on corneal stromal keratocytes to initiate the healing cascade.101,105 In summary, NAC modulates its effects through various mechanisms (Table 1) including its:

    View all citing articles on Scopus
    f1

    Address correspondence to: James D. Zieske, Schepens Eye Research Institute and Department of Ophthalmology, Harvard Medical School, 20 Staniford Street, Boston, MA, U.S.A. E-mail: [email protected]

    View full text