Use of Intracellular Cytokine Staining and Bacterial Superantigen to Document Suppression of the Adaptive Immune System in Injured Patients

Abstract

Objective To determine the percentages of major T lymphocyte subsets in the circulating peripheral blood mononuclear cell population in patients with major traumatic injury at early and late time points and to determine the expression of coreceptors and cytokine production by these T cell subsets. Summary Background Data Prior studies suggest that serious injury in humans suppresses the adaptive immune system as revealed by diminished proliferation and altered cytokine production in response to polyclonal T cell activation. However, the contribution of individual cell types to this immune dysfunction has not been well characterized. Methods The percentage of circulating CD4⁺ and CD8⁺ T cells and the relative density of CD4 and CD8 coreceptor expression was determined by flow cytometry in 17 consecutive trauma patients (injury severity score > 20) within 24 hours of injury and at day 7. Intracellular expression of the cytokines interleukin 2 (IL-2), interferon gamma (IFNγ), IL-4, and IL-10 were also studied after stimulation with bacterial superantigen (SEB). Patients were compared with age- and sex-matched controls and to themselves for differences between early and late cytokine expression. Results The percentage of circulating CD4⁺ and CD8⁺ T cells was decreased versus controls at day 1 and further decreased by day 7 following injury. CD4 and CD8 cell surface expression was also decreased at days 1 and 7. CD4⁺ T cells in injured patients responded to SEB activation with decreased expression of IFNγ and IL-2 on day 1 versus controls (P < 0.05) and of all 4 cytokines by day 7 (P < 0.05), while CD8⁺ T cells showed diminished expression of IFNγ and IL-2 only at both time points. When day 1 and day 7 cytokine expression results were compared in the same patients, CD4⁺ T cells showed diminished expression of IFNγ, IL-2, and IL-4 by day 7 (P < 0.05), but maintained expression of IL-10. CD8 T cells showed diminished expression of IFNγ only. Conclusions Severe injury induces a loss of circulating CD4⁺ and CD8⁺ T lymphocytes and diminished coreceptor expression by these cells. Both T cell subsets show progressive loss of immunostimulatory cytokine production with maintenance of potentially suppressive IL-10 production. These events may have negative consequences for host defense.