LDGs inhibit T cell IFN-γ production in an H2O2-dependent manner. (A) MLN cells were activated with anti-CD3 mAb for 15 hours. Activated MLN cells were cultured alone (Co) or with HDGs, LDGs, or LDGs plus catalase (1000 U/mL) and stimulated with PMA, ionomycin, and monensin for the last 5 hours. Cells were stained with anti–IFN-γ and anti-CD5 to assess IFN-γ–producing T cells (CD5 gated). Results are expressed as percentage of control response. The percentage of IFN-γ–producing cells in the controls ranges from 9% to 17% of all T cells. Numbers in parentheses indicate the number of experiments. Bars represent standard error. *P < .05. (B) MLNs cultured or not cultured with LDGs, as in panel A, were assessed for viability with annexin V and PI staining. Data presented correspond to lymphocytes gated by SSC and FSC parameters. (C) Hydrogen peroxide intracellular content measured with DHR on LDGs (top panel) or lymphocytes (bottom panel). LDGs were cultured in the absence (stim) or presence of catalase (stim/cat) or were left unstained (control). Anti-CD4/CD8 gated, preactivated MLN cells were cultured alone (control), in the presence of LDGs without (LDG) or with (LDG/cat) catalase. Shown are histograms for LDGs alone or lymphocytes. Note that lymphocytes fluoresce with DHR only in the presence of LDGs.
