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SOMSHUVRA BHATTACHARYA, Gudiseva Chandrasekher; Kinetics of extracellular matrix (ECM) protein release from cornea following epithelial injury and its relevance to wound repair. Invest. Ophthalmol. Vis. Sci. 2016;57(12):4352.
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© ARVO (1962-2015); The Authors (2016-present)
Corneal epithelial injury induces significant changes in cellular physiology of stromal layer that results in the release of growth promoting factors to facilitate the healing. Wound repair is a complex process that requires the participation of different ECM proteins for cell adhesion, migration and proliferation. To attribute a distinct function for different ECM proteins during epithelial regeneration we analyzed the pattern of their release after epithelial debridement.
Porcine corneas subjected to injury by complete epithelial debridement were maintained in organ culture (DMEM/F12 medium at 37 oC for 0- 60 hours). The culture medium was removed periodically and replaced with fresh medium. The presence of different ECM proteins in the medium was analyzed by SDS-PAGE/immunoblotting. A preparation of epithelial primary cultures and ECMs (collagen I and fibronectin), and a pectin-Ca based scaffold embedded with cornea primary epithelial cultures and ECMs were evaluated for their ability to regenerate epithelial layer on deepithelialized corneas and in cell culture respectively.
Presence of collagen I, collagen IV and fibronectin was identified in the medium in which the corneas were incubated. As compared to uninjured corneas, deepithelialized corneas released significantly higher amount of ECM proteins. Rate of release of these proteins varied with time. While collagen I release mostly remained steady throughout the experimental conditions, fibronectin release was higher in first four hours of incubation and collagen IV levels increased in 8-12 hours. Release of all three proteins decreased gradually, but sustained beyond 60 hours. Topical administration of a mixture of epithelial cells and ECM proteins promoted the regeneration of epithelium on epithelial debrided cornea surface. A scaffold embedded with ECM proteins and epithelial cells produced the growth of epithelial layer in cell culture.
Fibronectin released early from injured cornea may form a provisional basement membrane to initiate epithelial adhesion. Delayed release of collagen IV could facilitate the formation of permanent basement membrane. Prolonged release of ECM proteins indicates their necessity for epithelial cell regeneration. Engineering of ECM-based therapeutic devices for regeneration of damaged ocular surface is worthy of consideration.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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