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Fig. 6. Dendritic cell (DC) maturation in the presence of HIV-infected and noninfected CD4⁺ cell blasts. CD83-phycoerythrin (PE) (Immunotech, Luminy, France ) labeling of DC [gated using forward scatter (FSC) and side scatter (SSC) parameters and CD11c staining] after overnight culture with UV-irradiated (UV CD4⁺) or nonirradiated Z-VAD (N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone)-treated (ZVAD CD4⁺) CD4⁺ T cell blasts. HIV⁺, infected with 100 ng of p24 per ml of HIV-1_lai. Note that CD83 up-regulation does not depend on T cell apoptosis. Results are representative of two experiments.

Fig. 7. Acquisition of material from live H9-HIV cells by DC. (Left) Acquisition of PKH67 (green) from H9-HIV cells into the membrane and into intracellular vesicles of CD11c-labeled DC (red) after 15 min coculture. (Right) DC and H9-HIV cells were cocultured for 2 h, then labeled using anti-CD1a (in red) and anti–CD3 (green) antibodies. The resulting surface labeling shows acquisition of CD3 by a CD1a⁺ DC. Data are representative of at least two experiments.

Fig. 8. Despite passage of particles through transwell membrane, cross-presentation from live H9-HIV cells is not obtained without cell:cell contact. (A) Cross-presentation from live, Z-VAD-treated H9-HIV cells was tested as in Fig. 5A with or without transwells (TW) using an anti- RT_476-84 CD8⁺ T line and a DC:T ratio of 1:3. (B) Microparticles smaller than 0.45 m m reached the lower compartment. H9 cells were cultured for 2 days. The culture supernatant was filtered through a 0.45-m m filter, then ultracentrifuged at 100,000 ´ g at 4°C for 18 h (1). Pellet was resuspended in PBS, added to the upper compartment of transwells overnight with PBS in the lower chamber, and then analyzed by flow cytometry. For size comparison purposes, calibrated latex beads (Sigma) were used.

1. Gluschankof, P., Mondor, I., Gelderblom, H. R. & Sattentau, Q. J.

(1997) Virology 230, 125-33.