FIG. 6.

A fraction of resting CD4⁺ T cells bearing integrated HIV-1 DNA was latently infected. In each plot, the percentage of events in the boxed region is shown. The experiments were performed in the presence of the protease inhibitor saquinavir to prevent spreading infection. (A) IL-7 alone. Uninfected cells treated with IL-7 showed minimal background staining with anti-Gag antibodies. Uninfected resting CD4⁺ T cells were cultured for 3 days and then stimulated with IL-7 for an additional 3 days. This was a control for background staining with the anti-Gag monoclonal antibody. Isotype control antibodies gave less background staining than anti-Gag antibodies. (B) HIV alone. Unstimulated HIV-inoculated cells stained minimally with anti-Gag antibodies. Resting cells were spinoculated with HIV and 6 days later were permeabilized and stained for intracellular Gag. (C) HIV plus IL-7. IL-7 stimulated 3.0% of HIV-inoculated cells to produce Gag. Resting cells were spinoculated with HIV, 3 days later they were stimulated with IL-7, and 3 days after IL-7 addition they were permeabilized and stained for intracellular Gag. Gag was measured 3 days after IL-7 stimulation because Gag production peaked at that time. (D) HIV plus CD3 and CD28 beads. Anti-CD3 and -CD28 beads stimulated 4.5% of HIV-inoculated cells to produce Gag. Resting cells were spinoculated with HIV, 3 days later they were stimulated with anti-CD3 and -CD28 beads in the presence of the integrase inhibitor L870,810, and 24 h later they were permeabilized and stained for intracellular Gag. Gag was measured 24 h after bead stimulation because Gag production peaked at that time. We demonstrated that our integrase inhibitor was effective by inoculating resting CD4⁺ T cells and culturing them in the presence of the integrase inhibitor L870,810 at 100 nM. Under these conditions, we saw no reduction in the frequency of reverse transcripts, but we found that the frequency of proviruses/cell was reduced by 96.7%.