HPV-DNA is not detectable in outgrowing cells from explant cultures of skin lesions established at the air-liquid-interface

Keratinocyte cultures established from HPV containing skin cancers were des cribed earlier to lose their HPV DNA after passaging in vitro. A different approach was therefore used in this study. Explant cultures were generated by depositing small pieces of various benign and (pre)malignant skin specim ens of renal transplant recipients and non-immunosuppressed patients on fib roblast-populated collagen lattices or on de-epidermized dermis. Subsequent ly, the cultures were maintained at the air-liquid interface. At various ti me points, samples were collected for both HPV analysis, using a nested PCR approach, and morphology. The outgrowing keratinocytes developed into mult ilayered epithelial structures showing terminal differentiation. No histolo gical differences were observed between cultures established from HPV posit ive and negative lesions. Eighteen biopsy specimens were tested for their H PV content before and after culture. Before culture 11 out of these skin sp ecimens contained DNA of the Epidermodysplasia Verruciformis-related HPV ty pes (EV-HPV). Comparison of the HPV types detected in two different parts o f the same skin specimen before culture was strongly suggestive for a non-h omogeneous distribution of EV-HPV in the lesions. From the explant cultures derived from the 11 HPV-positive biopsies, 31 samples from the originally explanted pieces of tissue and 38 samples from the outgrowing multilayered epithelial sections were collected. HPV DNA was detected in 10 of the 31 an d in 3 of the 38 samples (Chi-square test, P = 0.01), respectively. These r esults indicate that EV-HPV positive keratinocytes do not efficiently proli ferate or lose their HPV DNA in this culture system or EV-HPV DNA is presen t in only a few basal cells, making it improbable that these cells are loca ted at the outgrowing margins. (C) 2000 Wiley-Liss, Inc.