The contribution of signals 1, 2 & 3 on the metabolic phenotype of CD4+ T cells

T cell metabolism is differentially regulated in order to support their activation, replication and effector functions. For example, T cells preferentially upregulate glycolysis following activation and this is a crucial determining factor at the Th17/Treg axis. Productive T cell activation requires several signals namely 1) antigen, 2) co-stimulation, and 3) cytokines but if, and how, these influence the metabolic changes associated T cell activation is largely unknown. To investigate this we assessed the metabolic phenotypes of CD4+ T cells following activation with; 1) variable strengths of stimulation through antigen-T cell receptor (TCR) engagement, 2) incubation with CHO cells transfected with individual costimulatory molecules CD80 or CD86, 3) exposure to the inflammatory cytokine IL-6. Metabolism was assessed using NMR spectroscopy-based metabolomics and metabolic flux analysis to assess the balance between the major pathways of glycolysis and oxidative phosphorylation. We optimised the techniques for determining human CD4+ T cell metabolic phenotype and have observed quantitative and qualitative differences associated with variations in the activating signals 1 to 3. In particular, we found that exposure to IL-6 prior to TCR stimulation leads to an increase in glycolysis, a response which might prepare T cells for the metabolic burden following activation. Subsequent flow cytometry analysis demonstrates that IL-6 exposed cells have an increased proliferative phenotype, greater mitochondrial mass and markedly upregulate GLUT1, however, these changes were only observed in naïve cells and not memory. Abnormal T cell responses and systemically measurable dysregulated metabolism are hallmarks of autoimmune and inflammatory diseases such as rheumatoid arthritis. An improved understanding of T cell metabolism has implications for the treatment of these diseases and the mechanisms of action of biological therapies such as abatacept and tocilizumab which target T cell differentiation and function