Platelets from mice with sepsis directly induce antigen-specific CD8⁺ T-cell suppression in vitro. (A) Platelets isolated from B2M^+/+ or B2M^f/f-Pf4Cre mice on day 4 after CLP were pulsed with SIIN for 2 hours, washed, and cocultured with antigen-specific CD8⁺ T cells isolated from Rag2^−/− OT-I mice for 60 hours at 37°C in uncoated cell culture plates. (B) Representative flow plots for CD8⁺ T-cell proliferation at baseline. CD8⁺ T cells were stained with the proliferation dye V450 before coincubation. (C-D) A representative flow plot and quantitation demonstrating decreased CD8⁺ T-cell proliferation when CD8⁺ T cells were incubated with platelets isolated from C57BL/6 mice after CLP compared with sham. Proliferation was normalized to CD8⁺ T cells incubated with platelets from sham mice. (E-F) A representative flow plot and quantitation demonstrating suppressed CD8⁺ T-cell proliferation during sepsis, when CD8⁺ T cells were incubated with platelets isolated from B2M^+/+ mice compared with platelets from B2M^f/f-Pf4Cre mice. Proliferation was normalized to CD8⁺ T cells incubated with platelets from B2M^f/f-Pf4Cre mice after CLP. All plots are representative of 3 independent experiments. An unpaired Student t test and Mann-Whitney test were used. *P < .05. FSC, forward scatter; NIR, near infrared region; SSC, side scatter.