Project description:Recent findings suggest that undifferentiated, stem-like, antigen specific T cells serve as an important long-term reservoir for autoimmune CD8 T cell responses. However, it is still unclear whether CD4 T cells exhibit a similar differentiation trajectory culminating in terminal differentiation, acquisition of an exhausted phenotype, and loss of stemness and function. We analyzed islet infiltrating T cells by scRNAseq and flow cytometry and found that while CD4 T cells in autoimmune diabetes share many features of exhaustion with CD8 T cells, expression patterns of inhibitory receptors are distinct in autoimmune T cells compared to T cells in chronic LCMV infection.
Project description:Recent findings suggest that undifferentiated, stem-like, antigen specific T cells serve as an important long-term reservoir for autoimmune CD8 T cell responses. However, it is still unclear whether CD4 T cells exhibit a similar differentiation trajectory culminating in terminal differentiation, acquisition of an exhausted phenotype, and loss of stemness and function. We analyzed islet infiltrating T cells in 8- and 16-week old NOD mice by scRNAseq and flow cytometry and found that while CD4 T cells in autoimmune diabetes share many features of exhaustion with CD8 T cells, expression patterns of inhibitory receptors are distinct in autoimmune T cells compared to T cells in chronic LCMV infection.
Project description:Seropositivity for autoantibodies against islet autoantigens is associated with the development of autoimmune type 1 diabetes and B cell targeted therapies are effective in both mouse models and in patients who are affected by or at risk for autoimmune type 1 diabetes. The role of B cell receptor affinity in autoimmune type 1 diabetes is unclear. Here, we employed single cell RNA sequencing to define the relationship between B cell receptor affinity for insulin and B cell phenotype during disease development using immunoglobulin heavy chain (VH125) transgenic mouse model (VH125.NOD) in which insulin binding B cells lose self-tolerance, becoming activated during development of autoimmmun type 1 diabetes.
Project description:Antigen-specific regulatory T cells (Tregs) suppress pathogenic autoreactivity and are potential therapeutic candidates for autoimmune diseases such as systemic lupus erythematosus (SLE). Lupus nephritis is associated with autoreactivity to the Smith (Sm) autoantigen and the human leucocyte antigen (HLA)-DR15 haplotype; hence, we investigated the potential of Sm-specific Tregs (Sm-Tregs) to suppress disease. We identified a novel HLA-DR15 restricted immunodominant Sm T cell epitope using biophysical affinity binding assays, then identified high-affinity Sm-specific T cell receptors (TCRs) using high-throughput single-cell sequencing. Using lentiviral vectors, we transduced our lead Sm-specific TCR onto Tregs derived from patients with SLE who were anti-Sm and HLA-DR15 positive. Compared with polyclonal mock-transduced Tregs, Sm-Tregs potently suppressed Sm-specific pro-inflammatory responses in vitro and suppressed disease progression in a humanized mouse model of lupus nephritis. These results show that Sm-Tregs are a promising therapy for SLE.
Project description:A new Treg-specific, FoxP3-GFP-hCre BAC transgenic was crossed to a conditional Dicer knock-out mouse strain to analyze the role of microRNAs (miRNA) in the development and function of regulatory T cells (Tregs). Although thymic Tregs developed normally in this setting, the cells showed evidence of altered differentiation and dysfunction in the periphery. Dicer-deficient Treg lineage cells failed to remain stable as a subset of cells down-regulated the Treg-specific transcription factor, FoxP3, while the majority expressed altered levels of multiple genes and proteins (including Neuropilin 1, GITR and CTLA-4) associated with the Treg fingerprint. In fact, a significant percentage of the Treg lineage cells took on a Th memory phenotype including increased levels of CD127, IL-4, and interferon-g. Importantly, Dicer-deficient Tregs lost suppression activity in vivo; the mice rapidly developed fatal systemic autoimmune disease resembling the FoxP3 knockout phenotype. These results support a central role for miRNAs in maintaining the stability of differentiated Treg function in vivo and homeostasis of the adaptive immune system. Experiment Overall Design: Lymph node CD4+YFP+ T cells from FoxP3-GFP-hCre x ROSA26R-YFP Dicerwt/lox (Het) and FoxP3-GFP-hCre x ROSA26R-YFP Dicerlox/lox (KO) mice were isolated by flow cytometry. Triplicate GeneChips were used for each T cell population.
Project description:A new Treg-specific, FoxP3-GFP-hCre BAC transgenic was crossed to a conditional Dicer knock-out mouse strain to analyze the role of microRNAs (miRNA) in the development and function of regulatory T cells (Tregs). Although thymic Tregs developed normally in this setting, the cells showed evidence of altered differentiation and dysfunction in the periphery. Dicer-deficient Treg lineage cells failed to remain stable as a subset of cells down-regulated the Treg-specific transcription factor, FoxP3, while the majority expressed altered levels of multiple genes and proteins (including Neuropilin 1, GITR and CTLA-4) associated with the Treg fingerprint. In fact, a significant percentage of the Treg lineage cells took on a Th memory phenotype including increased levels of CD127, IL-4, and interferon-g. Importantly, Dicer-deficient Tregs lost suppression activity in vivo; the mice rapidly developed fatal systemic autoimmune disease resembling the FoxP3 knockout phenotype. These results support a central role for miRNAs in maintaining the stability of differentiated Treg function in vivo and homeostasis of the adaptive immune system.