The role of innate lymphocyte metabolism in sepsis

Abstract

Sepsis is the result of a dysregulated systemic immune response to microbial infection that leads to organ failure and death in about 30% of affected individuals. The antimicrobial response is biphasic with an overwhelming inflammatory response preceding a period of profoundimmune suppression. We hypothesized that altered metabolism in immune cells during sepsis may affect their functions and contribute to the clinical phenotypes of the disease. Natural killer (NK) cells and Vδ2+γδ T cells from patients in late stages of sepsis and healthy controls were assessed for functional and metabolic readouts. Total peripheral blood NK cells were found at normal frequencies in the sepsis patients, whereas the CD56bright subset was found at significantly lower frequencies compared to those from healthy donors. The expression of the activation marker CD69, the transferrin receptor CD71, and the amino acid transporter CD98 at baseline and in response to cytokine stimulation, were similar on NK cells from sepsis patients compared to controls. The frequencies of Vδ2 T cells were lower in sepsis patients compared to controls. Although Vδ2 T cells from sepsis patients had reduced CD69 expression, they had significantly higher basal expression of CD98 compared to healthy donors. Vδ2 T cells from sepsis patients and controls subjects activated with cytokines or the antigenic ligand (E)-4-Hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP) upregulated CD69 and CD71 expression. Stimulated NK cells and Vδ2 T cells from sepsis patients produced interferon-γ (IFN-γ) at lower frequencies compared to the same cells from healthy donors. Furthermore, the metabolic regulator mTORC1 exhibited reduced phosphorylation in NK cells from sepsis patients possibly explaining the functional defects in the production of IFN-γ. Mitochondrial investigation indicated that NK cells and Vδ2 T cells from sepsis patients had similar mitochondrial structure and function (mitochondrial mass, ATP synthase and reactive oxygen species production) to those from healthy donors. Thus, NK cells and Vδ2 T cells from sepsis patients display similar functional and metabolic profiles to those from healthy donors, except for a selective defect in mTORC1 activity in NK cells and IFN-γ production in NK vicells and Vδ2 T cells. A thorough investigation into Vδ2 T cell metabolism and function showed that HMB-PP or zoledronate can be used to expand and generate high purity Vδ2 T cell lines from peripheral blood mononuclear cells of healthy donors. Further activation of expanded Vδ2 T cells withHMB-PP or PMA/ionomycin resulted in increased IFN-γ production, whereas stimulation with zoledronate, IL-12/IL-15, or anti-CD3/CD28 mAb failed to induce IFN-γ, IL-4 or IL-17 production. Hence, zoledronate might be optimal for expanding Vδ2 T cells, whereas HMB-PP is optimal for inducing cytokine production. Metabolic analysis using Seahorse technology demonstrated that resting and activated Vδ2 T cells from healthy donors predominantly use glycolysis to generate energy. HMB-PP stimulated Vδ2 T cells had higher levels of glycolysis, with increased rates of basal glycolysis and glycolytic capacity compared to resting Vẟ2 T cells. In addition, HMB-PP stimulated and zoledronate stimulated Vẟ2 T cells consumed oxygen at higher rates than compared to resting Vẟ2 T cells, indicating that activation induces oxidative phosphorylation. Our data support that Vδ2 T cells undergo metabolic changes in response to activation. These finding provide the first evidence that Vδ2 T cells may be targeted for the treatment of sepsis and that that the therapeutic activities of Vδ2 T cells may be regulated by modulating their metabolism.