IMMORTALIZATION OF RABBIT CORNEAL EPITHELIAL-CELLS BY A RECOMBINANT SV40-ADENOVIRUS VECTOR

Purpose. Cultured corneal epithelial cell is detrimental because of its short life span and its heterogeneity. We have tried to establish an immortalized epithelial cell line. Methods. Primary cultured rabbit corneal epithelial cells were infected with a recombinant SV40-adenovirus vector and were cloned three times. Results. The immortalized cell continued to grow by more than 400 generations through 100 passages. SV40-associated large T antigen was demonstrable on the nuclear membrane of these immortalized cells by immunofluorescence technique. This cell line exhibited a similar cobblestone-like appearance as normal corneal epithelial cells. Transmission electron microscopy showed a line of evidence for stratification, including desmosome formation and microvilli development at the superficial cell layer. As the culture grew, these cells began to express cornea-specific 64 kD cytokeratins. In contrast to cultured normal corneal epithelial cells, this cell line had a good proliferative ability after a long-term storage in liquid nitrogen. Conclusions. Because this particular cell line shares properties consistent with normal corneal epithelial cells and is easy to handle in vitro, it may serve as a useful tool in corneal epithelial research.