[Immortalization of rat corneal epithelial cells by SV40-adenovirus recombinant vector]

Using a SV40-adenovirus recombinant vector, we have successfully established a rat corneal epithelial cell line (RatCE) and studied its biological characteristics. RatCE continued to grow for more than 400 generations. It proliferated centrifugally in the early phase of the culture (1-3 days in culture) and had a cobblestone-like appearance in confluency. Desmosomes and microvilli were clearly seen under a transmission electron microscope. RatCE could be stored in liquid nitrogen and its biological characteristics were: doubling time, 18.3 hrs, colony forming ability, 36%, and growth ability in soft agar, 2%. When the insoluble extract from RatCE was electrophoresed, insoluble proteins were seen at 36 kD, 40 kD, 44 kD, 48 kD, 56 kD, and 64 kD. Anti-64 kD cytokeratin antibody strongly reacted with numerous filaments in the cytoplasm of RatCE. Hence, RatCE possessed 64 kD corneal specific keratin. A large amount of fibronectin was also assessed at focal contact by immunohistochemistry. Thus, RatCE retains several kinds of epithelial characteristics, is derived from one clone, and is immortalized. RatCE will be a useful tool in studies of the corneal epithelium.