Atherosclerosis is a lipid-driven chronic inflammatory disease; however, whether it can be classified as an autoimmune disease remains unclear. In this study, we applied single-cell T cell receptor seqencing (scTCR-seq) on human carotid artery plaques and matched peripheral blood mononuclear cell samples to assess the extent of TCR clonality and antigen-specific activation within the various T cell subsets. We observed the highest degree of plaque-specific clonal expansion in effector CD4+ T cells, and these clonally expanded T cells expressed genes such as CD69, FOS and FOSB, indicative of recent TCR engagement, suggesting antigen-specific stimulation. CellChat analysis suggested multiple potential interactions of these effector CD4+ T cells with foam cells. Finally, we integrated a published scTCR-seq dataset of the autoimmune disease psoriatic arthritis, and we report various commonalities between the two diseases. In conclusion, our data suggest that atherosclerosis has an autoimmune compondent driven by autoreactive CD4+ T cells.

Aims Aging is a dominant driver of atherosclerosis and induces a series of immunological alterations, called immunosenescence. Given the demographic shift towards elderly, elucidating the unknown impact of aging on the immunological landscape in atherosclerosis is highly relevant. While the young Western diet-fed Ldlr-deficient (Ldlr-/-) mouse is a widely used model to study atherosclerosis, it does not reflect the gradual plaque progression in the context of an aging immune system as occurs in humans. Methods and Results Here, we show that aging promotes advanced atherosclerosis in chow diet-fed Ldlr-/- mice, with increased incidence of calcification and cholesterol crystals. We observed systemic immunosenescence, including myeloid skewing and T-cells with more extreme effector phenotypes. Using a combination of single-cell RNA-sequencing and flow cytometry on aortic leukocytes of young versus aged Ldlr-/- mice, we show age-related shifts in expression of genes involved in atherogenic processes, such as cellular activation and cytokine production. We identified age-associated cells with pro-inflammatory features, including GzmK+CD8+ T-cells and previously in atherosclerosis undefined CD11b+CD11c+T-bet+ age-associated B-cells (ABCs). ABCs of Ldlr-/- mice showed high expression of genes involved in plasma cell differentiation, co-stimulation, and antigen presentation. In vitro studies supported that ABCs are highly potent antigen-presenting cells. In cardiovascular disease patients, we confirmed the presence of these age-associated T- and B-cells in atherosclerotic plaques and blood. Conclusions Collectively, we are the first to provide comprehensive profiling of aged immunity in atherosclerotic mice and reveal the emergence of age-associated T- and B-cells in the atherosclerotic aorta. Further research into age-associated immunity may contribute to novel diagnostic and therapeutic tools to combat cardiovascular disease.

Aims CD8+ T-cells can differentiate into subpopulations that are characterized by a specific cytokine profile, such as the Tc17 population that produces IL-17. The role of this CD8+ T-cell subset in atherosclerosis remains elusive. In this study, we therefore investigated the contribution of Tc17 cells to the development of atherosclerosis. Methods and Results Flow cytometry analysis of atherosclerotic lesions from apoE-/- mice revealed a pronounced increase in RORγt+CD8+ T-cells compared to the spleen, indicating a lesion-specific increase in Tc17 cells. To study whether and how the Tc17 subset affects atherosclerosis, we performed an adoptive transfer of Tc17 cells or undifferentiated Tc0 cells into CD8-/-LDLr-/- mice fed a Western-type diet. Using flow cytometry, we showed that Tc17 cells retained a high level of IL-17A production in vivo. Moreover, Tc17 cells produced lower levels of IFN-γ than their Tc0 counterparts. Analysis of the aortic root revealed that the transfer of Tc17 cells did not increase atherosclerotic lesion size, in contrast to Tc0-treated mice. Conclusion These findings demonstrate a lesion-localized increase in Tc17 cells in an atherosclerotic mouse model. Tc17 cells appeared to be non-atherogenic, in contrast to their Tc0 counterpart. Translational perspective CD8+ T-cells are present in high numbers in human atherosclerotic plaques, however their role in inflammation and the pathogenesis of atherosclerosis remains elusive. Our results indicate that the majority of CD8+ T-cells in atherosclerotic plaques of mice have lost their ability to produce the pro-inflammatory cytokine IFN-γ and gain traits of IL-17-producing CD8+ T-cells (Tc17 cells). We show that this subset of CD8+ T-cells is less atherogenic then IFN-γ producing Tc1 cells.