Summary – Project 2 Cardiovascular disease remains the leading cause of death worldwide, with atherosclerosis being a major contributor. The unremitting rise in obesity and its co-morbidities, including diabetes and non-alcoholic fatty liver diseases, further increase the risk of atherosclerosis. While such cardiometabolic diseases were once attributed primarily to dysregulations of lipid metabolism, it is now appreciated that the immune response to excess lipid in tissues (artery, adipose tissue, liver) shapes cardiometabolic disease progression and its complications. Determining the mechanisms underlying this damaging metabolic inflammation, and identifying therapeutic approaches to quench it, are major focuses of our Program Project Grant (PPG). In its first cycle, our PPG unveiled key pathways through which alterations in macrophage (Mø) metabolism, trafficking, and tissue-specific molecular reprogramming drive the chronic inflammation that fuels atherosclerosis, and the metabolic disorders that accelerate it (e.g., obesity and non-alcoholic steatohepatitis; NASH). Through examination of mouse models and human tissues from patients with these disorders, Project 2 studies point to pathogenic roles for the neuroimmune guidance molecule netrin-1 in directing non-resolving inflammation and lipid accumulation in metabolic tissues. We hypothesize that netrin-1 contributes to dysregulation of Mø metabolism and trafficking, and molecular re-programming of inflammation, and that these processes are driven by tissue- and environment- specific stimuli that contribute to cardiometabolic disease. Key Project 2 discoveries in cycle 1 of the PPG include: (1) Deletion of netrin-1 in myeloid cells protects mice from high fat diet-induced obesity and hepatic steatosis, and regresses advanced atherosclerosis; (2) Netrin-1 expression alters the functional trajectory of Mø in obese adipose tissue and atherosclerotic plaques; and (3) Distinct isoforms of netrin-1 accumulate in Mø during metabolic inflammation that can provoke both receptor-dependent and -independent signaling. In this proposal, we will investigate how Mø-derived netrin-1 promotes atherosclerosis, obesity and NASH through both Mø-intrinsic and -extrinsic mechanisms, including cross-talk with other immune cells and parenchymal cells, and how interventions targeting netrin-1 can be leveraged toward our goal of mitigating cardiometabolic disease. Together with Projects 1 and 3, we will probe how netrin-1 contributes to intra- and inter-organ communications through shared tools, strategies and bioinformatics approaches, and test therapeutically relevant approaches to block its detrimental actions. Fortified by complementary examinations in human tissues and transcriptome databases, we will employ state-of-the-art RNA sequencing, coupled with spatial transcriptomics, to generate and “visualize” a comprehensive map of the putative interactome and upstream transcriptional regulators that regulate intra- a...