IMMUNE-RESPONSIVE GENE 1 REGULATES ATHEROSCLEROTIC PLAQUE INFLAMMATION II

Atherosclerosis is the major underlying cause of cardiovascular disease (CVD) mortality and is fueled by a failure to resolve inflammation within the vessel wall1. The mechanisms underpinning sustained inflammation within this microenvironment are not fully elucidated. A more comprehensive understanding of the immunoregulatory pathways modulating inflammation in this context holds the potential to inform the development of more specific therapies. Immune responsive gene 1 (Irg1) is an enzyme that diverts cis-aconitate from the TCA cycle and produces the metabolite itaconate, which elicits immunological tolerance in the context of microbial infection2–4. We hypothesized that the IRG1-itaconate axis restricts inflammation in the atherosclerotic plaque. Overall design: Formation of atherslerotic plaque was induced in mice by placing them on Western Diet. Aorta's were removed and single-cell suspensions of the aortic arch were made and CD45+immune cells were isolated by staining (anti-CD45 PerCp-Cy5.5, BioLegend) and flow sorting viable (fixable viability dye e780, eBioscience) single cells on a Facs Aria II cytometer (BD) equipped with a 100 µm nozzle. Next scRNA seq was performed and analyzed. ADT/HTO Barcode sequence hashtagged samples CMO301 ATGAGGAATTCCTGC WT_mouse_1 CMO302 CATGCCAATAGAGCG WT_mouse_2 CMO303 CCGTCGTCCAAGCAT WT_mouse_3 CMO304 AACGTTAATCACTCA WT_mouse_4 CMO305 CGCGATATGGTCGGA WT_mouse_5 CMO306 AAGATGAGGTCTGTG KO_mouse_1 CMO307 AAGCTCGTTGGAAGA KO_mouse_2 CMO308 CGGATTCCACATCAT KO_mouse_3 CMO309 GTTGATCTATAACAG KO_mouse_4 CMO310 GCAGGAGGTATCAAT KO_mouse_5 CMO301 ATGAGGAATTCCTGC WT+DMSO_mouse_1 CMO302 CATGCCAATAGAGCG WT+DMSO_mouse_2 CMO303 CCGTCGTCCAAGCAT WT+DMSO_mouse_3 CMO304 AACGTTAATCACTCA WT+DMSO_mouse_4 CMO308 CGGATTCCACATCAT KO+DMSO_mouse_1 CMO309 GTTGATCTATAACAG KO+DMSO_mouse_2 CMO310 GCAGGAGGTATCAAT KO+DMSO_mouse_3 CMO311 GAATCGTGATTCTTC KO+DMSO_mouse_4 CMO312 ACATGGTCAACGCTG KO+DMSO_mouse_5 CMO305 CGCGATATGGTCGGA KO+4OI_mouse_1 CMO306 AAGATGAGGTCTGTG KO+4OI_mouse_2 CMO308 CGGATTCCACATCAT KO+4OI_mouse_3 CMO309 GTTGATCTATAACAG KO+4OI_mouse_4