QUANTITATION BY COMPETITIVE PCR OF HIV-1 PROVIRAL DNA IN EPIDERMAL LANGERHANS CELLS OF HIV-INFECTED PATIENTS

Langerhans cells (LC) belong to the dendritic cell family and represen t the principal antigen presenting cells populating squamous epithelia . We have reported the presence of human immunodeficiency virus Type 1 (HIV-1) proviral DNA and RNA in purified LC from the epidermis of ser opositive patients. The aim of this study was to quantify HIV-1 provir al DNA in LC of infected patients using a competitive polymerase chain reaction (PCR) assay. Bulk epidermal cell (EC) suspensions were obtai ned from the skin of nine AIDS patients and six seronegative subjects. Purified LC and LC-depleted EC were prepared by immunomagnetic separa tion using an anti-CD1a monoclonal antibody. LC preparations did not c ontain T cells, as assessed by reverse transcription PCR analysis of t he T cell receptor beta-chain gene (C region). In addition, no CD14(+) cells could be detected in LC fractions by immunostaining of cytospin preparations. To quantify HIV-1 DNA, a new competitive PCR system was devised using SK145/150 as primers (gag) and a competitor plasmid DNA with a modified sequence (209 instead of 142 bp). The number of HIV-1 DNA copies found in the LC of AIDS patients ranged from 107 to 3,645/ 10(5) LC. In contrast, LC-depleted EC from the same subjects were all negative. The results indicate that in AIDS patients the frequency of infected LC is comparable to that reported for peripheral blood CD4(+) T cells.