DETECTION OF NEURON-SPECIFIC ENOLASE IN LONG-TERM CULTURES OF HUMAN CORNEAL ENDOTHELIUM

Background: Human corneal endothelial cells cultivated in monolayer cu lture for protracted periods undergo morphological dedifferentiation, whereby they assume a more fibroblast-like appearance. These cultures may also become overgrown with contaminating stromal fibroblasts and/o r with keratocytes, when non-selective media are employed, thus render ing identification of actual endothelial cells difficult on a strictly morphological basis. Methods: The endothelium of the human cornea sta ins for neurone-specific enolase (NSE) in situ, and we therefore: wish ed to study the expression of this marker in primary and long-term mon olayer cultures of these cells. Ten such cultures were established, si x being stained for NSE at the primary and first-passage stage, the ot her four for 6, 8, 10 and 12 months. The NSE-staining pattern manifest ed in co-cultures of corneal endothelium and fibroblasts or keratocyte s (first to fifth passage cultures) was also investigated, and co-cult ures established from each of the latter two cell types served as cont rols. Results: In monolayers of corneal endothelium which had retained their cobblestone-like morphology, NSE could be demonstrated even aft er more than 20 passages, which amounted to 1 year in culture. Dediffe rentiated or degenerating endothelial cells stained poorly and inhomog eneously. Control cultures of fibroblasts or keratocytes were consiste ntly NSE-negative, and when each of these cell types was co-cultured s eparately with corneal endothelium, only the latter expressed the mark er protein. Conclusion: Since antibodies against NSE are commercially available, practical use may be made of this marker protein for confir ming corneal endothelial status in longterm cultures.