The role of altered immune metabolism in the inflammatory CD4+ T-cell response in equine recurrent uveitis

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Equine recurrent uveitis (ERU) is a disease of great importance as it occurs worldwide with a prevalence of 10% and can lead to blindness. Because the disease occurs spontaneously, it has also been shown to be a model for the transmission of autoimmune uveitis in humans. As pathologic immune responses and inflammation of the eye are increasingly recognized as an essential component of many ocular diseases previously thought to be purely neurodegenerative, it is becoming increasingly important to understand such abnormal immune responses in this immune-privileged organ. Recently, it has become known that immune cell metabolism and immune cell effector functions are intimately linked, and the role of immune metabolism in pathological immune cell functions is increasingly being investigated. The major drivers of ERU are CD4+ T cells.Through our preliminary work, we know that the immune metabolism of these cells is significantly altered. By differential proteomic analysis of the mitochondria of CD4+ T cells, we have demonstrated selective alteration of proteins responsible for complex I respiratory chain function in mitochondria. We have also identified differentially abundant proteins associated with mitochondrial fission and fusion. In this project, we will analyze the mitochondrial morphology of live CD4+ T cells from the blood of control horses and from animals with ERU and compare them with CD4+ T cells from the vitreous that migrate in large numbers into the eye in ERU. For this purpose, we will use high-resolution STORM microscopy and 3D microscopy, which will allow detailed study of mitochondria localization and morphology. The influence of cell metabolism on the inflammatory responses of these cells will be investigated using complex I inhibitors (rotenone and metformin) to understand the selective activation of this complex in the CD4+ T cells of ERU animals. Of this complex, NDUFAB1 was particularly abundant in CD4+ T cells from ERU animals, so we characterize its function in detail here, as well as that of MUL1, a protein that has been associated with aberrant immune responses in other models. In addition to spontaneously diseased animals, we are also using cells from biobanks of horses immunized with IRBP 20 years ago, from which we have PBMC from throughout the course of induced disease. Finally, we are investigating the metabolic flexibility of CD4+ T cells from ERU cases to determine whether selective blockade of a metabolic pathway could be used to therapeutically influence inflammation or whether the cells then switch to other nutrients and energy production pathways.