KERATOCYTE-POPULATED COLLAGEN GEL AS AN IN-VITRO MODEL OF EXCIMER-LASER KERATECTOMY

BACKGROUND: To develop an in vitro model to study the effects of excim er laser keratectomy on corneal stromal cells, we evaluated two types of collagen gel populated with keratocytes. METHODS: Keratocyte-popula ted collagen gels were prepared with type I collagen in 6-well plates or in culture plate inserts, the bottom of which consisted of a nitroc ellulose membrane, contained within 6-well plates. The gels were ablat ed by the 193-nm excimer laser, set to ablate 50, 100, or 200 mu m dee p, and was observed under a phase contrast microscope for 2 days. RESU LTS: Keratocytes cultured in collagen gel developed cytoplasmic proces ses and formed networks of interconnected cells. Cells within the abla ted area in the 6-well plates began to lose their cytoplasmic processe s and became round approximately 3 hours after excimer laser ablation. These cellular changes were more prominent in the gels ablated to a d epth of 200 mu m. Cells outside of the ablation zones in the 6-well pl ates and the culture plate inserts remained intact. CONCLUSIONS: These results suggest the use of keratocyte-populated collagen gel as an in vitro model of cellular response to excimer laser keratectomy and als o suggest that gel prepared in culture plate inserts is the preferred method.