Fluorescent detection of HIV particles in human tissue explant cultures and the rhesus macaque genital tract: model to understand sexual transmission

OA07-01 Antiviral Therapy 2007; 12 Suppl 4:OA07-01 Fluorescent detection of HIV particles in human tissue explant cultures and the rhesus macaque genital tract: model to understand sexual transmission S McCoombe, N Byers, M McRaven, S Shukair, M Dinh and TJ Hope Northwestern University, Feinberg School of Medicine, Chicago, IL, USA Objective: Sexual transmission accounts for over 80% of all HIV infections. How HIV breaches the genital epithelium of men and women remains poorly defined. The objective of this study is to define the interaction of HIV with human and macaque genital tissue to gain insights into the mechanism(s) of sexual transmission of HIV. Methods: Previous attempts to observe HIV interactions with tissue explants and primate models have been hindered by high background. To study single virion interactions with genital tissues, we developed a new technology utilizing photoactivatable GFP (PA-GFP)-labelled HIV and fluorescent deconvolution microscopy. Photoactivation of PA-GFP is achieved by exposure to 400–430 nm light. Using this system, tissue autofluorescence is defined, the sample is activated and newly detected signal is labelled HIV. To study initial HIV interactions with the genitalia, we incubated human foreskin and cervical explants for 4–24 h with PA-GFP-labelled HIV. These experiments were replicated in the rhesus macaque non-human primate model. Following inoculation, genital samples were frozen, sectioned and stained, enabling obser- vation of tissue structure, HIV target cells and PA-GFP-labelled virions. Results: Within 4 h HIV penetrates the superficial squamous epithelia of the ectocervix, in some cases deeper than 30 microns. Penetrating virions are observed between differenti- ated squamous epithelial cells. We also observe HIV penetration of the columnar endocervical epithelium with virions observed in the subbasal milieu. Penetrating virions regularly colocalize with both Langerhans cells and exposed CD4+ T-cells. In cervical explants, mucus harbours numerous virions, and when present reduces the incidence of epithelial penetration. Complimentary results were obtained using the rhesus macaque model. Conclusion: HIV can clearly penetrate the intact squamous and columnar epithelium of the genital tract in both human explants and living macaques gaining access to underlying target cells. The presence of mucus decreases access of virus to the columnar epithelium. Breaks in the epithelial barrier facilitate the association of virus with tissue resident T-cells. Observing early HIV interactions with genital tissues has provided important insight into HIV transmission across epithelial barriers. A better understanding of the initial events of HIV sexual transmission is essential for the development of a successful vaccine.