TLR9 engagement decreases sensitivity to IR-induced apoptosis via the TLR9-MyD88–signaling transduction pathway. (A) CD4 T cells from WT or MyD88^–/– mice were activated and treated with TLR9 ligand or control ligand as described in Figure 1. Twenty-four hours after TLR9 treatment, T cells were irradiated using various doses of γ-radiation, and apoptosis was determined by flow cytometry. (B-D) CD4 T lymphocytes from WT, TLR9^–/–, or MyD88^–/– BL/6 mice were activated with plate-bound anti-CD3/CD28 Ab and treated or not with the indicated concentrations of TLR9 ligand before exposure to IR (300 cGy). Irradiated T cells were allowed 30 minutes to repair DNA before damage was measured using comet assay. Data shown are representative of several experiments, each showing similar trends; data are the means (± SD, n > 150 cells per condition). (E) CD4 T cells from CD45.1 mice were mixed at a 1:1 ratio with MyD88^–/– CD4 T lymphocytes (which express CD45.2) and activated with plate-bound CD3/CD28 Ab and treated with TLR9 ligand as described. Twenty-four hours after TLR9 treatment, T cells were or were not irradiated (300 cGy) and the levels of apoptosis from each group were measured by flow cytometry 24 hours after irradiation. The Student t test was used to determine P values between the indicated groups; *P < .05; ns indicates not significant; data are the means (± SD).