Keratocyte apoptosis may be key to corneal wound healing

Corneal wound healing research is concentrating on blocking the initial keratocyte apoptosis response.

DALLAS — A decade’s worth of research and observation has culminated in the possibility of a prophylactic pharmacological approach to corneal wound healing modulation.

“There has been explosive growth in our understanding of the corneal wound healing response over the past ten years. I believe the day will come, in the not-too-distant future, where pharmacologic application of drugs to control the wound healing response will be as important as the excimer laser or the microkeratome,” said Steven E. Wilson, MD, here at the annual American Academy of Ophthalmology meeting.

“We get amazing accuracy with the results of LASIK; however, we always have some that are either overcorrected or undercorrected, and I believe that a large proportion of this variability is due to the variability in the wound healing response.

“The cornea’s response to laser refractive surgery has revealed itself in incremental steps over a decade,” he said.

“Any source of epithelial injury is associated with keratocyte apoptosis. The epithelial injury can extend from viral infections to scrape injuries to photorefractive keratectomy (PRK) to pressure of a contact lens on the ocular surface. This keratocyte apoptosis response starts immediately after the injury to the epithelium,” Dr. Wilson explained.

Keratocyte apoptosis response

The keratocyte apoptosis response is thought to be mediated by cytokines released from the injured epithelial cells, such as IL-1 alpha. “In the case of LASIK, it is thought that the microkeratome blade tracks the epithelial debris and associated cytokines into the interface, or provides a route for their passage into the cornea after surgery,” Dr. Wilson said.

Once these keratocytes have undergone apoptosis, there is a proliferative response. The keratocytes that remain in the tissue proliferate to restore the keratocytes initially with myofibroblast cells. Interestingly, there is a high variability in the level of the keratocyte apoptosis response between two eyes performed with identical procedures.

“I’d also like to point out that the thickness of the flap is important. In patients who have identical keratocyte apoptosis responses, there can be a marked difference in wound healing depending on how thin the flap is,” Dr. Wilson said.

“The interaction between the overlying epithelium and the keratocyte cells, and the continuing ‘communication’ [between the two], are really the key factors in what leads to the final wound healing response,” he explained.

Because the corneal wound healing response is so complex, it is difficult to intervene at one of the later steps in the process, so Dr. Wilson and colleagues are concentrating most of their efforts on blocking the initial keratocyte apoptosis response. He is pursuing pharmacologic mediation of keratocyte apoptosis response that can be used both in PRK and LASIK patients. “The day will come when we will have medication we can apply to the cornea before the procedure to transiently block this apoptosis response and thus diminish the subsequent wound healing response,” Dr. Wilson said.

Assay for apoptosis in PRK (left) and in LASIK (right). Both are rabbit corneas.

Area of relevance

An increasing area of relevance in corneal wound healing is the inflammatory cell infiltration that occurs about 24 hours after “injury” in cases like PRK and LASIK. “When we use a monoclonal antibody for monocytes and macrophages, we see a large number of inflammatory cells that are not detected well with stains such as H and E staining,” Dr. Wilson said.

“The mechanism for drawing these cells in is also being elucidated,” he reported. One example is monocyte chemotactic and activating factor (MCAF).

“Recent studies in my laboratory have demonstrated that IL-1 released from the epithelium stimulates keratocyte cells to produce this factor MCAF associated with the drawing in of these monocytes and macrophages into the corneal stroma. As the myofibroblasts migrate back into the anterior stroma, they produce cytokines, such as HGF and KGF, which then regulate the overlying epithelial wound healing response. The migration and proliferation of these epithelial cells leads to closure of the epithelial defect,” Dr. Wilson said.

These same cytokines are also released from the lacrimal gland and can appear in the tear film, he reported. “The myofibroblasts that migrate back in also produce irregular collagen, glycosaminoglycans and other substances associated with stromal remodeling, and in some cases, haze, such as in high level PRK cases.”

Dr. Wilson and colleagues are also looking into gene transfer type technologies where they can transiently put a gene into the keratocytes, with the goal of blocking the apoptosis response for a few days surrounding the surgical procedure.

For Your Information:

• Steven E. Wilson, MD, can be reached at University of Washington, Ophthalmology, Box 356485, Seattle, WA 98195; (206) 598-2020; fax: (206) 598-3740. Dr. Wilson has no direct financial interest in the products mentioned in this article, nor is he a paid consultant for any companies mentioned.