Fig. 4: Co-expressed PD-1 protects TIM-3+ T cells from Gal-9-induced cell death. | Nature Communications

Fig. 4: Co-expressed PD-1 protects TIM-3+ T cells from Gal-9-induced cell death.

From: Galectin-9 interacts with PD-1 and TIM-3 to regulate T cell death and is a target for cancer immunotherapy

Fig. 4

a–c Jurkat cells transduced with indicated proteins individually or in combinations were treated with or without Gal-9 for two days and stained with PD-1/TIM-3 antibodies. Cell survival of relevant PD-1/TIM-3 subsets was determined by flow cytometry with counting beads. a Cells were gated based on FSC/SSC parameters and 7-AAD staining. b, c Viable single cells equivalent to 3000 counting beads are shown in plots for each sample (10,000 beads were added to each sample just prior to data acquisition). Numbers in plots indicate cell count in corresponding gates. Data (mean values ± SD) from three independent experiments are shown (c). Two-tailed unpaired t-test. NS, not significant (P > 0.05); **P < 0.01; ***P < 0.001. Control vs TIM-3, P = 0.0003; control vs TIM-3 + PD-1, P = 0.9382; control vs TIM-3 + PD-1(N116Q), P = 0.0040; TIM-3 + PD-1 vs TIM-3 + PD-1(N116Q), P = 0.0084. d, e Human CD8 T cells were incubated in ImmunoCult-XF T Cell Expansion Medium with or without Gal-9 in the presence of IL-2 and ImmunoCult Human CD3/CD28/CD2 T Cell Activator for 2 days and analyzed by flow cytometry with counting beads as described above for the survival of different PD-1/TIM-3 subsets. Numbers in plots indicate cell counts in corresponding quadrants. Data (mean values ± SD) are representative of three independent experiments. Two-tailed unpaired t-test. *P < 0.05. PD-1–TIM-3+ vs PD-1+TIM-3+, P = 0.0212; PD-1–TIM-3+ vs PD-1–TIM-3–, P = 0.0164. Source data are provided as a Source data file.

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