Figure 6. Fungus-loaded MDDCs facilitate the activation of resting CD4⁺ T cells and promote HIV-1 infection by activating more permissive cell targets.

MDDCs were pulsed separately with heat-killed C. albicans, PMy, PMm or medium control for 2 h, then were co-cultured with same numbers of allogeneic resting CD4⁺ T cells. Transwell plates with membrane pore sizes of 0.4 µm were used to separate MDDCs from T cells. The activation of resting T cells directly stimulated with fungi was also checked. After additional 48 h incubation, T cells were gated by a CD3⁺ cell population to detect CD69 expression by flow cytometry (A). CD4⁺ T cells were purified with magnetic micro-beads, pulsed with 5 ng p24^gag amounts of pseudotyped HIV-Luc/NL4-3 for 2 h, and HIV-1 infection was detected 3 days later by measuring the luciferase activity in cell lysates (B, C). Asterisks indicate significantly enhanced HIV-1 replication in T cells activated by fungus-stimulated MDDCs during co-culture, relative to that in medium-treated MDDCs (**P <0.01, ***P <0.001, paired t test). Direct treatment of heat-killed fungi neither activated resting T cells nor enhanced T-cell susceptibility to HIV-1 infection. PHA-P was used as a positive control for non-specific activation of resting T cells, and the positive percentages of CD69 are shown in (A) and (C). Results of one representative experiment out of three are shown. Data are mean ± SD.