The role of MMPS and TIMPS in the feline ocular surface system during corneal epithelial wound healing

Abstract

The ocular surface system is a complex microenvironment including the cornea, conjunctiva, lacrimal gland, meibomian glands and tear film. Matrix metalloproteinases (MMPs), a family of zinc- and calcium-dependent proteinases involved in maintaining and remodelling extracellular matrix and inflammation, have been found in ocular tissues and tears. The analysis of tears has been used to identify potential biomarkers for eye diseases. Contributions of individual ocular surface components to tear proteins are not fully understood. This study investigated the expression of MMPs and tissue inhibitors of MMPs (TIMPs) (particularly MMP-9 and -2) in tears and ocular tissues following corneal epithelial wounding (debridement) at predetermined time points using a feline model. Preliminary work validated the use of eye-flush tears (collected after instillation of saline) and procedural control eyes (same treatment as wounded eyes) as controls for the tear collection study. Gelatin zymography and immunohistochemistry were used to detect MMPs and TIMPs in tears and ocular surface tissues, and data was then analysed using statistical methods. During corneal healing, expression of MMP-9 was detected in all ocular surface components with significant increase in tears, the corneal and limbal epithelium, keratocytes and conjunctival epithelium. In contrast, MMP-2 was found significantly increased only in tears and the corneal and conjunctival epithelium. Analysis of tears from wounded and procedural control eyes suggested a minimal contribution of the healing corneal epithelium to tear-derived MMP-9 and -2. Other possible contributors to increased tear MMP-9 levels included the lacrimal gland, inflammatory cells in conjunctival tissue including conjunctival-associated lymphoid tissue (CALT). Expression of MMP-1, TIMP-1 and -2 in ocular tissues in response to wounding were also investigated. MMP-1 was increased in the healing corneal epithelium, keratocytes and limbal epithelium. TIMP-1 and -2 immunoreactivity paralleled that of MMP-9 in the cornea and lacrimal gland. The current study revealed the complexity of interactions in the corneal epithelial wound healing processes, as exemplified for MMP-9 and -2 with simultaneous increases in tears and ocular surface tissues in response to corneal wounding, and subsequent regulation during wound healing. These observations highlight the importance of considering the ocular surface microenvironment in its entirety for future studies