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Originally published In Press as doi:10.1167/iovs.08-2239 on September 29, 2008
 (Investigative Ophthalmology and Visual Science. 2009;50:604-613.)
© 2009 by The Association for Research in Vision and Ophthalmology, Inc.
doi:10.1167/iovs.08-2239
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Genomic Aberrations and Cellular Heterogeneity in SV40-Immortalized Human Corneal Epithelial Cells

Kenta Yamasaki,1 Satoshi Kawasaki,1 Robert D. Young,2 Hideki Fukuoka,1 Hidetoshi Tanioka,1 Mina Nakatsukasa,1 Andrew J. Quantock,2 and Shigeru Kinoshita1

1From the Department of Ophthalmology, Kyoto Prefectural University of Medicine, Kyoto, Japan; and the 2School of Optometry and Vision Sciences, Cardiff University, Cardiff, United Kingdom.

PURPOSE. Simian virus (SV)40–immortalized human corneal epithelial (HCE-T) cells have been widely used as an in vitro model of human corneal epithelial cells. The nature of this cell line was assessed for genomic aberrations and cellular heterogeneity.

METHODS. For the quantitative measurement of genomic aberrations, array-based comparative genomic hybridization (CGH) analysis was performed. For identification of cellular heterogeneity, cell morphology, growth kinetics, transepithelial electrical resistance, and transfection/transcriptional efficiency were analyzed. Real-time PCR and chromosomal fluorescent in situ hybridization (cFISH) against some gained or lost loci were performed, to assess genomic heterogeneity. Expressed sequence tags (ESTs) for this cell line were collected to assess differences in the gene expression profiles between HCE-T cells and normal corneal epithelial cells. Southern blot analysis and inverse PCR analyses were used to determine the genomic integration site of the SV40 large T antigen gene (LTAG).

RESULTS. Array CGH analysis demonstrated that the genomic content of HCE-T cells is different from the normal healthy genome. The results from cellular functional assays, real-time PCR, and cFISH strongly indicated that HCE-T cells consist of a significant number of heterogeneous cell populations. The genomic integration site of the SV40 large T antigen was at p22.1 of chromosome 9.

CONCLUSIONS. The results indicate that HCE-T cells have an altered genomic content and that they are composed of heterogeneous cell populations. This should be considered when conducting experiments or interpreting the results of studies that use this cell line.






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