Fig. 3: Lnc-Tim3 suppresses CD8 T cell function and protects from galectin-9–mediated cell death.

a, b Tim-3⁺CD8 T cells isolated from tumor-infiltrating lymphocytes (TILs) of HCC patients with Lnc-Tim3 high expression (Lnc-Tim3^highTim-3⁺CD8 T; cut-off, -6.53) were transfected with indicated lentiviral particles (Lnc-Tim3 scramble or shRNA). Intracellular expression of IFN-γ (a, left) and IL-2 (b) in CD8 T cells after treated with anti-TCR/anti-CD28 beads (bead-to-cell ratio of 1:1), IL-12 (100 ng/ml), and peptide cocktail (AFP_158-166, GPC_3144-152, and NY-ESO-1_157-165) were assessed by flow cytometry. The percentage of IFN-γ⁺CD8 T cells was showed in the right panel. (n = 10). c Concentration of IFN-γ in culture supernatant 48 h of the above groups. (n = 10). d, e Tim-3⁺CD8 T cells isolated from TILs of HCC patients with Lnc-Tim3 low expression (Lnc-Tim3^lowTim-3⁺CD8 T; cut-off, −6.53) were transfected with indicated lentiviral particles (Lnc-Tim3 wild type or Δ226-277). Intracellular expression of IFN-γ (d, left) and IL-2 (e) in CD8 T cells after treated with anti-TCR/anti-CD28 beads (bead-to-cell ratio of 1:1), IL-12 (100 ng/ml), and peptide cocktail (AFP_158-166, GPC_3144-152, and NY-ESO-1_157-165) were assessed by flow cytometry. The percentage of IFN-γ⁺CD8 T cells was showed in the right panel. (n = 10). f Concentration of IFN-γ in culture supernatant 48 h of the above groups. (n = 10). g Jurkat T cells were transfected with indicated lentiviral particles (Tim-3–HA, Lnc-Tim3 wild type or Δ226-277). Frequency of apoptosis were assessed by 7-aminoactinomycin D (7-AAD) after cells treated with galectin-9 for 3 h. Numbers contained in the fluorescence-activated cell sorting (FACS) plots represent the frequency of 7-AAD⁺ cells. SSC, side scatter. Data in the bar graph (right) represent the percentage of 7-AAD⁺ cells after galectin-9 treatment divided by the percentage of 7-AAD⁺ cells before treatment. Data are presented as means ± S.E.M. (*P < 0.05)