FIG.2.

HIV replication in IPC requires CD40L activation. IPC and MDC were enriched with immunomagnetic beads and were then separated by FACS based on their differing expression of CD123 and CD11c. IPC and MDC were incubated with HIV for 2 h, washed, and then cultured in parallel in 96 wells at a concentration of 0.2 × 10⁶ cells/ml (40,000 DC/well). (a) Following incubation of IPC and MDC with either HIV_LAI/IIIB (triangles) or HIV_BaL (circles), triplicate cultures were assessed weekly for viral replication on the basis of RT activity. (b) IPC and MDC were also stimulated with CD40LT following exposure to virus and were assessed weekly for RT activity. (c) IPC exposed or unexposed to HIV_BaL and then stimulated with CD40LT were also assessed by flow cytometry for intracellular HIV p24 following 1 week of culture. The histogram on the right represents infected cells stained with an irrelevant isotype-matched control antibody. (d) IPC cultures were also assessed for T-cell contamination and for viability by annexin V and PI staining at day 14. Numbers represent the percentage of cells present in the indicated gates. (e) IPC were incubated with either RANTES or SDF-1α at a concentration of 1 μg/ml for 1 h prior to and during exposure to medium alone (black bars), HIV_BaL (gray bars), or HIV_LAI/IIIB (black bars). Cultures were then assessed for p24 production at 1 week by ELISA. Error bars represent standard deviations from triplicate wells. Data shown are representative of three experiments.