Development and characterization of a rabbit corneal endothelial cell line.

Abstract

To develop a rabbit corneal endothelial cell line by transducing human papilloma virus (HPV) type 16 E6 and E7 oncogenes, and to characterize the inherent biological properties of the established cell line. Primary rabbit corneal endothelial cells were infected with a recombinant retrovirus harboring HPV E6 and E7, and the transformed cells were clonally selected by G418. Among the total of eight independent clones, one cell line (Clone no. A3) cultured over 40 passages was chosen to further characterize its inherent biological properties. Typical cell doubling time for these cells was 51 h, and the mean cell density of a flask culture was 1140 cells/mm(2). The various genes that are important for corneal endothelial functions were expressed at a level comparable to that of their normal counterparts. Furthermore, Na(+)/K(+) ATPase activity was maintained throughout an extended period, and the measured value at passage 30 was about 10 nmol inorganic phosphate/mg protein per minute. These results suggest that the rabbit corneal endothelial cell lines obtained here maintain normal corneal endothelial characteristics, and could be used not only for biological studies on corneal endothelial cells but also for such applications as the reconstruction of the ocular surface with an artificial cornea.