Unveiling the Impact of Arsenic Toxicity on Immune Cells in Atherosclerotic Plaques: Insights from Single-Cell Multi-Omics Profiling
Arsenic exposure is a global health problem and is correlated with adverse atherosclerotic outcomes in humans and in mouse models. However, the mechanisms of action are unclear. Since an atherosclerotic plaque is an amalgam of diverse immune and non-immune cells working in consortium, we set out to understand the mechanisms of arsenic-enhanced atherosclerosis in the CD45+ immune cells of atherosclerotic plaques in apoE-/- mouse model. To do this, we employ single-cell RNA- and single-cell-ATAC sequencings to understand both the genomic and epigenomic landscapes of these plaques. Overall design: 5-week-old apoe-/- male mice were started on a high-fat diet and either left as control or given 200 ppb arsenic (which is a concentration known to exacerbate atherosclerosis in these mice) for 13 weeks. At the end of the timepoint, mice were injected intravenously with anti-CD45 antibody to stain circulating leucocytes. Within 10/15 minutes of this injection, mice were asphyxiated, blood was drawn through cardiac puncture, followed by a cardiac perfusion with ice-cold PBS. At this point, most of the blood should be removed from the heart and aorta and what remains should be stained with the CD45 antibody. Next, the plaques from the aortic arch and the aortic sinus were harvested, minced and then chemically digested with an enzyme cocktail at 37 degrees for 1 hour. Then the sample was stained with CD45 antibody conjugated with a fluorophore different from the one used for IV injection and a L/D stain. Lastly, the live CD45 plaque-resident cells were sorted to eliminate circulating leucocytes and were sent for single-cell sequencing.