Abstract 2051: Monocyte Recruitment And Fate Specification Within Atherosclerotic Lesions

Recruited monocytes that subsequently differentiate into macrophages within atherosclerotic lesions are a primary driver of plaque progression and a risk factor for rupture. Advances in single cell RNA sequencing have expanded our understanding of macrophage diversity within plaques. However, limited analysis has explored regulators of monocyte lineage commitment and spatial localization in atherosclerotic plaque. Examining scRNAseq gene expression data of plaque-associated monocyte and macrophage lineages, pseudotime trajectory analysis predicted a binary fate decision by monocytes differentiating toward either an inflammatory or lipid-loaded “foamy” differentiation state. Foamy state was associated with lipid processing genes ( Fabp5, Lgals3, Trem2 ), whereas inflammatory state was associated with inflammasome genes ( Il1b, Nlrp3 ) and chemokines ( Ccl3, Ccl4 ). To test the in silico analysis, we established a monocyte fate mapping approach using a tamoxifen-inducible reporter mouse (CCR2 creERT2 Rosa26 lsl-tdTomato ) and performed kinetic analysis of tdTomato+ cells within plaque. tdTomato-labeled cells could be found shortly after tamoxifen treatment with small, rounded morphology, consistent with newly recruited monocytes. Within days, small, rounded cells were absent and labeled cells appeared to be lipid loaded with bloated morphology. Immunostaining revealed inflammasome activation in a large subset of monocytes immediately following recruitment to lesions. However, IL-1b hi cells were no longer detectable days after recruitment. Together, these data support a model where recruited monocytes commit toward either inflammatory or foamy lineage immediately upon plaque entry. Further, data support that inflammatory cells are short lived in atherosclerotic plaques, whereas foamy cells persist for longer periods of time. Ongoing work in our lab will address the role of known foamy macrophage regulators, including Trem2, in modulating fate commitment of monocytes entering early and late stage atherosclerotic lesions. Understanding the spatial-temporal dynamics of monocyte recruitment and differentiation will illuminate new mechanisms of atherosclerosis progression and potential therapeutic targets.