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Background:Coronary artery disease is an incurable, life-threatening disease that was once considered primarily a disorder of lipid deposition. Coronary artery disease is now also characterized by chronic inflammation‚ notable for the buildup of atherosclerotic plaques containing immune cells in various states of activation and differentiation. Understanding how these immune cells contribute to disease progression may lead to the development of novel therapeutic strategies.Methods:We used single-cell technology and in vitro assays to interrogate the immune microenvironment of human coronary atherosclerotic plaque at different stages of maturity.Results:In addition to macrophages, we found a high proportion of αβ T cells in the coronary plaques. Most of these T cells lack high expression ofCCR7andL-selectin, indicating that they are primarily antigen-experienced memory cells. Notably, nearly one-third of these cells express theHLA-DRAsurface marker, signifying activation through their TCRs (T-cell receptors). Consistent with this, TCR repertoire analysis confirmed the presence of activated αβ T cells (CD4HLA-DRA, 2 clusters expressed a proinflammatory and cytolytic signature characteristic of CD8 cells, while the other expressed AREG (amphiregulin), which promotes smooth muscle cell proliferation and fibrosis, and, thus, contributes to plaque progression.Conclusions:Taken together, these findings demonstrate that plaque T cells are clonally expanded potentially by antigen engagement, are potentially reactive to self-epitopes, and may interact with smooth muscle cells and macrophages in the plaque microenvironment.

Thrombosis has long been reported as a potentially deadly complication of respiratory viral infections and has recently received much attention during the global coronavirus disease 2019 pandemic. Increased risk of myocardial infarction has been reported during active infections with respiratory viruses, including influenza and severe acute respiratory syndrome coronavirus 2, which persists even after the virus has cleared. These clinical observations suggest an ongoing interaction between these respiratory viruses with the host’s coagulation and immune systems that is initiated at the time of infection but may continue long after the virus has been cleared. In this review, we discuss the epidemiology of viral-associated myocardial infarction, highlight recent clinical studies supporting a causal connection, and detail how the virus’ interaction with the host’s coagulation and immune systems can potentially mediate arterial thrombosis.

Background: Vascular thrombosis has been described in patients actively infected with influenza and, more recently, with the coronavirus. The risk may persist during the convalescence period. The underlying mechanisms mediating these events remain unclear. Objective: To determine whether human coronary plaque T cells reactive to influenza virus are also activated by self-proteins. Methods/Results: Using single cell targetRNAseq, we mapped the T cell repertoire in 19 patients with a range of coronary artery disease severity. To determine the potential peptides activating coronary plaque T cell receptors (TCRs), we matched TCR alpha and beta identity with HLA restriction to a database of known specificities. We found that plaque TCRs were potentially activated by viral epitopes, including those from the influenza M1 protein. We used a computational algorithm to identify self-proteins in the endothelial cells and vascular smooth muscle cells that may have peptides with similar homologies to viral epitopes predicted to bind our plaque TCRs. We then introduced plaque TCRs into the Jurkat T cells, exposed these cells to T2 (HLA-2 antigen presenting cells), viral peptides (e.g., M1) and self-epitopes (e.g., TSPAN17; ZIP9), and demonstrated T cell cross reactivity in vitro ( Figure 1 ). TSPAN17 is a transmembrane protein that regulates VE-cadherin expression and promotes T cell transmigration. Zip9, in contrast, is a protein that regulates zinc homeostasis, serves as an androgen receptor on endothelial cells, and has been shown to promote endothelial proliferation. Conclusion: Taken together, our findings suggest that T cell cross reactivity may be a potential mediator of viral-induced arterial thrombosis.

Background: The atherosclerotic plaque is a complex niche composed of immune cells including T cells and macrophages as well as vascular smooth muscle cells. How T cells interact with the plaque microenvironment is not well understood. Objective: To map the crosstalk between T cells, macrophages, and smooth muscle cells. Methods/Results: We performed single cell RNAseq using the 10x transcriptomics platform in coronary plaque samples from 12 patients, who underwent heart transplantation. We found a high proportion of macrophages as well as T cells, many of which display a memory phenotype that was confirmed by immunohistochemistry. Ligand-receptor computational analysis demonstrated significant cross talk between T cells, macrophages, and smooth muscle cells. One cytokine of interest is amphiregulin (AREG), which has been shown to mediate tissue healing and fibrosis. Of note, AREG expression appears to increase as coronary plaques progress from early lipid-rich stages to more advanced stages. To elucidate the effects of AREG on smooth muscle cells, we treated human coronary artery smooth muscle cells (hCASMCs) in vitro with recombinant AREG and found increased cell proliferation compared to control (P Conclusion: Taken together, our findings suggest that cross talk between T cells and smooth muscle cells, partially through AREG production by T cells, may contribute to plaque progression.

Background: Atherosclerosis is considered a chronic inflammatory disorder characterized by the presence of T cells and macrophages in the plaque. Given that T cells are the master regulators of the immune system, the possible role of T cells in mediating disease warrants further investigation. Objective: To map the T cell transcriptome in coronary artery plaque. Methods/Results: Using single cell RNAseq, we mapped the transcriptome of immune cells within the coronary atherosclerotic plaque in 12 patients with various disease stages. In addition to macrophages, we find an abundance of plaque T cells that appear to be enriched in plaque compared to blood ( Figure 1A ). Two plaque-enriched CD8 effector memory clusters (CD8 CTL Tem1 and CD8 CTL Tem2) display the highest proportion of activated cells (e.g., HLA-DRA+). These clusters are characterized by high expression of the pro-inflammatory cytokines (e.g., CCL3, CCL4, CCL5, and IFNG) and cytolytic markers (e.g.,GZMA, GZMH, GZMM, NKG7). CD8 CTL Tem1, however, has higher expression of GZMK, a cytolytic marker recently associated with inflammaging. CD8 CTL Tem2, on the other hand, has a higher expression of perforin and granulysin, two pore-forming proteins that mediate cell killing by enabling granzyme entry. Notably, CD8 CTL Tem2 appears to track with plaque progression, increasing as plaques mature from lipid-rich to more complex lesions then declining as plaques became more stable and calcified post-rupture. Specifically, this cluster displayed two-fold enrichment in complex plaques compared to other plaque phenotypes (12.1% vs. 6.9%, p=0.0069) ( Figure 1B ). This pro-inflammatory and cytolytic signature is also more apparent in the activated compared to nonactivated cells in this cluster. Conclusion: Taken together, our findings suggest that a pro-inflammatory CD8 T cell cluster characterized by a trifecta of cytolytic enzymes distinguishes complex plaque and deserves further investigation.