Fig. 7

Proliferation of CD4⁺ T-cells expressing IL-17A versus IFN-γ in response to low TCR triggering. Memory CD4⁺ T-cells were isolated from PBMCs of four different HIV-uninfected donors by negative selection using magnetic beads. Cells were loaded with CFSE (0.5 µM) and cultured in the presence of different doses of immobilized CD3 and soluble CD28 Abs (0.1, 0.25, or 0.5 µg/ml) for 3 days. Cells were then stimulated with PMA/Ionomycin in the presence of Brefeldin A for 18 h. Intracellular staining was performed with IL-17A and IFN-γ Abs. Based on the expression of IL-17A and/or IFN-γ, three cell subsets were identified as follows: Th1 (IL-17A⁻IFN-γ⁺), Th17 (IL-17A⁺IFN-γ⁻), and Th1Th17 (IL-17A⁺IFN-γ⁺). The frequency of proliferating cells (CFSE^low) was analyzed in each of the three cytokine-expressing subsets. a Shown are flow cytometry results in one representative donor out of four. Shown are statistical analyses of absolute (b) and relative (Th1Th17 proliferation levels were considered 100 %) c proliferation (CFSE^low) levels in Th1, Th17, and Th1Th17 cells from four different donors. Paired t test p values are indicated on the graphs