FIG. 5.

MVA-HIV-infected cells are cross-presented by DCs to HS CTLs. (A) Cross-presentation experimental procedure. HLA-A2⁻ donor cells were mock treated or infected with MVA-HIV (MOI = 1, 1 h), washed, seeded for 5 h to allow Gag expression, and cocultured with HLA-A2⁺ DCs for 1 h. DCs were then harvested and incubated for at least 8 h with HS T cells. (B) HS T-cell activation monitored by IFN-γ ELISPOT assay. As a positive control, HLA-A2⁺ DCs were directly infected with MVA-HIV (MOI = 1) or loaded with the SL9 peptide (1 μg/ml). Importantly, in the absence of DCs, HLA-A2⁻ donor cells infected with MVA-HIV (MOI = 1) cannot stimulate HS CTLs. Mock-treated or MVA-infected donor cell-to-DC ratios are indicated (donor cell/DC). Background IFN-γ production induced by uninfected cells and IFN-γ production by target cells alone (at least three times lower than that with HS CTLs) were subtracted. For each panel, data are means ± SD for duplicates or triplicates and are representative of 3 independent experiments.