Monaco, Claudia

Cardiovascular disease is the global #1 cause of mortality and morbidity. The majority of cardiovascular diseases is caused by atherosclerosis, a lipid-driven, inflammatory disease of the middle- and large-sized arteries. The disease is characterized by the formation of atherosclerotic plaques throughout the arterial tree. Over the years, insights into the pathogenesis of atherosclerosis have shifted from a ‘lipid-driven’ model to a ‘response-to-injury’ perspective, and more recently to a ‘lipid-driven inflammatory disease’ viewpoint. We are now aware that a network of multiple immune cell-types and -subsets of the innate and adaptive immune system inhabit our arteries. Intricate interactions between these immune cellsubsets, non-immune cells, and local environmental substances such as lipids, cell debris and calcium cause a fluidic balance of pro-inflammatory and regulatory responses. A dysregulation of this balance towards a pro-inflammatory milieu drives atherosclerotic disease progression. Although we have acknowledged that atherosclerosis is an inflammatory disease, state-of-theart treatments are still based on lipid lowering, anti-hypertensive and lifestyle-changing strategies. In the past decade, clinical phase I, II and III trials targeting the immune system revealed that patients tolerate immunotherapy, show decreased inflammation and/or had a reduction in cardiovascular endpoints. However, the search for novel immunotherapeutic targets and treatment regimens as well as stratification of patients who would benefit from such treatments to combat atherosclerotic cardiovascular disease is only just beginning. In this review article, we will highlight the newest insights on the different cell subsets and components of the immune system in atherosclerosis and elaborate on current and future immunotherapeutics to treat atherosclerotic cardiovascular disease.

Abstract Aims Recent studies have revealed a close connection between cellular metabolism and the chronic inflammatory process of atherosclerosis. While the link between systemic metabolism and atherosclerosis is well established, the implications of altered metabolism in the artery wall are less understood. Pyruvate dehydrogenase kinase (PDK)-dependent inhibition of pyruvate dehydrogenase (PDH) has been identified as a major metabolic step regulating inflammation. Whether the PDK/PDH axis plays a role in vascular inflammation and atherosclerotic cardiovascular disease remains unclear. Methods and results Gene profiling of human atherosclerotic plaques revealed a strong correlation between PDK1 and PDK4 transcript levels and the expression of pro-inflammatory and destabilizing genes. Remarkably, the PDK1 and PDK4 expression correlated with a more vulnerable plaque phenotype, and PDK1 expression was found to predict future major adverse cardiovascular events. Using the small-molecule PDK inhibitor dichloroacetate (DCA) that restores arterial PDH activity, we demonstrated that the PDK/PDH axis is a major immunometabolic pathway, regulating immune cell polarization, plaque development, and fibrous cap formation in Apoe−/− mice. Surprisingly, we discovered that DCA regulates succinate release and mitigates its GPR91-dependent signals promoting NLRP3 inflammasome activation and IL-1β secretion by macrophages in the plaque. Conclusions We have demonstrated for the first time that the PDK/PDH axis is associated with vascular inflammation in humans and particularly that the PDK1 isozyme is associated with more severe disease and could predict secondary cardiovascular events. Moreover, we demonstrate that targeting the PDK/PDH axis with DCA skews the immune system, inhibits vascular inflammation and atherogenesis, and promotes plaque stability features in Apoe−/− mice. These results point toward a promising treatment to combat atherosclerosis.

Abstract The advent of single-cell biology opens a new chapter for understanding human biological processes and for diagnosing, monitoring, and treating disease. This revolution now reaches the field of cardiovascular disease (CVD). New technologies to interrogate CVD samples at single-cell resolution are allowing the identification of novel cell communities that are important in shaping disease development and direct towards new therapeutic strategies. These approaches have begun to revolutionize atherosclerosis pathology and redraw our understanding of disease development. This review discusses the state-of-the-art of single-cell analysis of atherosclerotic plaques, with a particular focus on human lesions, and presents the current resolution of cellular subpopulations and their heterogeneity and plasticity in relation to clinically relevant features. Opportunities and pitfalls of current technologies as well as the clinical impact of single-cell technologies in CVD patient care are highlighted, advocating for multidisciplinary and international collaborative efforts to join the cellular dots of CVD.

Background Adolescents with chronic disease are often exposed to inflammatory, metabolic, and hemodynamic risk factors for early atherosclerosis. Since postmortem studies have shown that atherogenesis starts in the aorta, the CDACD (Cardiovascular Disease in Adolescents with Chronic Disease) study investigated preclinical aortic atherosclerosis in these adolescents. Methods and Results The cross‐sectional CDACD study enrolled 114 adolescents 12 to 18 years old with chronic disorders including juvenile idiopathic arthritis, cystic fibrosis, obesity, corrected coarctation of the aorta, and healthy controls with a corrected atrial septal defect. Cardiovascular magnetic resonance was used to assess aortic pulse wave velocity and aortic wall thickness, as established aortic measures of preclinical atherosclerosis. Cardiovascular magnetic resonance showed a higher aortic pulse wave velocity, which reflects aortic stiffness, and higher aortic wall thickness in all adolescent chronic disease groups, compared with controls ( P <0.05). Age (β=0.253), heart rate (β=0.236), systolic blood pressure (β=−0.264), and diastolic blood pressure (β=0.365) were identified as significant predictors for aortic pulse wave velocity, using multivariable linear regression analysis. Aortic wall thickness was predicted by body mass index (β=0.248) and fasting glucose (β=0.242), next to aortic lumen area (β=0.340). Carotid intima‐media thickness was assessed using ultrasonography, and was only higher in adolescents with coarctation of the aorta, compared with controls ( P <0.001). Conclusions Adolescents with chronic disease showed enhanced aortic stiffness and wall thickness compared with controls. The enhanced aortic pulse wave velocity and aortic wall thickness in adolescents with chronic disease could indicate accelerated atherogenesis. Our findings underscore the importance of the aorta for assessment of early atherosclerosis, and the need for tailored cardiovascular follow‐up of children with chronic disease.

Abstract Aims CD40 and its ligand, CD40L, play a critical role in driving atherosclerotic plaque development. Disrupted CD40-signalling reduces experimental atherosclerosis and induces a favourable stable plaque phenotype. We recently showed that small molecule-based inhibition of CD40-tumour necrosis factor receptor associated factor-6 interactions attenuates atherosclerosis in hyperlipidaemic mice via macrophage-driven mechanisms. The present study aims to detail the function of myeloid CD40 in atherosclerosis using myeloid-specific CD40-deficient mice. Method and Results Cd40flox/flox and LysM-cre Cd40flox/flox mice on an Apoe−/− background were generated (CD40wt and CD40mac−/−, respectively). Atherosclerotic lesion size, as well as plaque macrophage content, was reduced in CD40mac−/− compared to CD40wt mice, and their plaques displayed a reduction in necrotic core size. Transcriptomics analysis of the CD40mac−/− atherosclerotic aorta revealed downregulated pathways of immune pathways and inflammatory responses. Loss of CD40 in macrophages changed the representation of aortic macrophage subsets. Mass cytometry analysis revealed a higher content of a subset of alternative or resident-like CD206+CD209b− macrophages in the atherosclerotic aorta of CD40mac−/− compared to CD40wt mice. RNA-sequencing of bone marrow-derived macrophages of CD40mac−/− mice demonstrated upregulation of genes associated with alternatively activated macrophages (including Folr2, Thbs1, Sdc1, and Tns1). Conclusions We here show that absence of CD40 signalling in myeloid cells reduces atherosclerosis and limits systemic inflammation by preventing a shift in macrophage polarization towards pro-inflammatory states. Our study confirms the merit of macrophage-targeted inhibition of CD40 as a valuable therapeutic strategy to combat atherosclerosis.