REQUIREMENT FOR VITAMIN-A IN LONG-TERM CULTURE OF HUMAN CORNEA

Purpose. To develop an in vitro model for the study of vitamin A funct ions in the cornea by determining the morphologic responses to its abs ence and to the addition of varying concentrations in long-term (21-da y) human organ culture. Methods. Donor corneas were cultured for 21 da ys in an air-liquid environment designed to maintain normal corneal mo rphology in long-term culture. Corneas were graded by light microscopy and were further analyzed by transmission electron microscopy. Result s. Epithelium of human corneas cultured in the absence of vitamin A fo r 21 days showed characteristics of vitamin A deficiency: multilayerin g of squamous cells, bundling of tonofilaments to form keratofibrils, increased desmosomes, and loss of microplicae on the surface cells. Th e cells responded to the addition of vitamin A in concentrations rangi ng from 5 X 10(-9) to 5 X 10(-6) M. At the lowest concentration, cell differentiation and morphology appeared normal. Cells developed secret ory morphology at higher concentrations of vitamin A, with intracellul ar vesicles, basement membrane overproduction, and reduced cell-cell a nd cell-substrate attachments. Keratocytes and endothelial cells showe d increased intracellular vacuoles in the presence of higher concentra tions of vitamin A. Conclusions. Cultured adult human corneas showed p rimary, dose-related responses to vitamin A in the absence of inflamma tory, neuronal, or humoral factors. The epithelium was most sensitive to the absence of vitamin A and showed a dose-related response to addi tional vitamin A. All corneal cell layers showed morphologic changes a t the highest concentration of vitamin A used.