Cell-intrinsic regulation of sex differences in T-cell function. A, Schematics depicting in vitro generation of exhausted T cells. B, Exhaustion markers and cytokine expression were measured on day 5 by flow cytometry after polyclonal stimulation with a stimulation cocktail for 4 hours. Data are shown as mean ± SD (n = 3) and are representative of three independent experiments. Two-way ANOVA with Tukey multiple comparisons test (*, P < 0.05; **, P < 0.01; ***, P < 0.001). gMFI, geometric mean fluorescence intensity. C, qPCR analysis on exhausted T cells. Relative expression levels normalized to exhausted T cells from one male are shown (n = 4). An unpaired t test was performed. D, Male and female in vitro exhausted T cells were cultured with SB28-OVA cells for 24 hours, and viability of tumor cells was measured by flow cytometry. Multiple t test (*, P < 0.05; **, P < 0.01). E, A schematic depicting the adoptive transfer model. F, Kaplan–Meier curves depicting survival of male and female RAG1−/− mice bearing SB28-OVA tumor cells after adoptive transfer of OT-I cells. Data shown are combined from three independent experiments and n = 10–14 each group. Statistical significance was determined by the log-rank test (*, P < 0.05; ***, P < 0.001). d, days.