Sepsis-induced antigen-specific CD4⁺ T-cell dysfunction is corrected by anti-GITR treatment. DO11.10 mice were administered 300 μg anti-GITR or control antibody and were immunized 30 minutes later with Ova_323-339 in alum at the time of cecal ligation and puncture or sham surgery. Five days later, lymph node cells were harvested and 2.5 × 10⁴ CD4⁺ cells were cultured with 2.5 × 10⁵ irradiated APCs with Ova peptide for 72 hours. (A) Proliferation of CD4⁺ T cells was measured as the incorporation of ³H-thymidine during the last 18 hours of culture. Media was harvested at the time of ³H-thymidine addition to assess cytokine production. (B) IL-2, (C) IFN-γ, (D) IL-4, and (E) IL-10 concentrations were assessed by Luminex multiplex analysis. (F) Representative examples of flow plots demonstrating the expansion of DO11.10 T cells (KJI-26⁺CD4⁺) from lymph nodes of Balb/c mice that were injected intravenously with 5 × 10⁶ DO11.10 CD4⁺ T cells 3 days before sham (left), CLP with control antibody (middle), or CLP with anti-GITR antibody (right) treatment at the time of immunization with Ova_323-337 peptide in alum. (G) The calculated percentage of living (Sytox Blue⁻), nondebris DO11.10 T cells (KJI26⁺CD4⁺) expanded in the peripheral lymph nodes of the Balb/c mice. (H) The calculated total number of live (Sytox Blue⁻), nondebris DO11.10 T cells using cell counts obtained with a hemacytometer. P values indicate differences between groups after post hoc analysis using the Fisher LSD method.