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Retention of low copy number human papillomavirus DNA in cultured cutaneous and mucosal wart keratinocytes

¹ Department of Experimental Dermatology,
² Department of Virology
³ Department of Genito-Urinary Medicine
and⁴ Department of Oral Pathology, The London Hospital Medical College, London E1 2AD
⁵ Department of Cell Biology, Wellcome Research Laboratories, Beckenham, Kent
⁶ International Center for Genetic Engineering and Biotechnology, Padriciano 99, I-34012 Trieste, Italy
and⁷ Department of Genito-Urinary Medicine, St Mary's Hospital, London, U.K.

Cultured wart keratinocytes have previously been described as having a limited proclivity to maintain episomal human papillomavirus (HPV) DNA. To investigate the nature of episome loss, and to determine keratinocyte-specific factors involved in it, we have examined a large series of anogenital and oral wart keratinocyte cultures, tracing episomal copy number with culture passage. We report that a higher proportion of oral wart keratinocytes maintain episomal HPV DNA to first passage (70% compared with 37% of anogenital wart cultures) when screened by slot blot hybridization. Furthermore, oral wart keratinocytes maintain episomal HPV copy through a greater number of passages (60% positive at passage 2 compared with 2% of anogenital wart cultures) with this technique. When anogenital cultures were examined at first passage for HPV infection by PCR with Southern blot hybridization of the product, a further 34% were found to be HPV-positive. To determine the mechanism of loss of episomal DNA from these cultures we examined the relative HPV copy number in cells which adhered to the culture vessel following passage and in those which did not adhere. Those which remained floating contained episomal HPV at high copy number whereas those which adhered were negative by slot blotting. The adherent cells, however, remained positive by PCR at subsequent passages until senescence. We conclude that a subpopulation of HPV-positive keratinocytes may be maintained in culture through serial passage until senescence.

Received 9 June 1993; accepted 1 October 1993.