Although primary cultures of human lens epithelial (HLE) cells provide important information concerning the role of epithelium in normal lens and cataract formation, the lack of a cell line precludes a broad range of studies on the metabolism and molecular biology of these cells. We have, therefore developed an HLE cell line. Primary cultures of HLE cells were transfected with plasmid vector DNA containing a large T antigen of SV40. The immortalized cells were characterized with regard to morphology, growth rate, karyotype, and expression of crystallins, aldose reductase and other enzymes. A single clone of the immortalized cells, SRA 01/04, formed a monolayer and grew constantly over 130 passages. Isozyme phenotype showed that SRA 01/04 was of human origin, and the chromosome counts were in the hypotetraploid range. Western blot analysis showed that the cells expressed a very low level of crystallins (alphaA and betaB2) and aldose reductase. Messenger RNA (mRNA) for both alpha and beta crystallins was detected by reverse transcription polymerase chain reaction (RT-PCR) in both early and late passages. Sequence analysis of the PCR products, corresponding to alphaA and betaB2 crystallins in the cell line and in primary cultures of HLE, revealed a 100% match with published human alphaA and betaB2 sequences. These characteristics were unchanged in the cell line in early and late passages. This is the first report of the presence of alphaA and transcripts of mRNA for both alphaA and betaB2 in an established human cell line. This new HLE cell line makes it possible to undertake many future studies on the role of epithelium in lens and cataract formation.
Copyright 1998 Academic Press.