Abstract 435: Macrophage Rac1 As A Transcriptional Co-factor For NF-kb-driven IL-1β Expression And Consequent Atherosclerotic Calcification

Coronary artery disease caused by atherosclerosis is a leading cause of morbidity and mortality. Vascular calcification, influenced by inflammation, is associated with atherosclerotic disease burden and increased risk of cardiovascular events. The small GTPase Rac1 is an important signal transducer of inflammatory cytokine expression. We sought to define the mechanisms whereby Rac1 and its activating guanine nucleotide exchange factor (GEF), Tiam1, regulate macrophage expression of the potent inflammatory cytokine, IL-1β, and consequent calcific atherosclerotic vascular disease. To define the role of macrophage Rac1 expression on IL-1β expression and atherosclerotic calcification, we developed mouse models on the atherosclerosis-prone background, including inducible myeloid Rac1 -deletion (mR1KO) ( CSF1 mcm Rac1 fl/fl ApoE -/-) , and global Tiam1- deletion ( Tiam1 -/- ApoE -/- ). Both Rac1-deleted and Tiam1-deleted macrophages demonstrated reduced IL-1β expression in an inflammasome-stimulation assay. Macrophage Rac1 -deletion led to decreased nuclear NF-κB activity and reduced NF-κB and GTP-Rac1 binding to the IL-1β promoter by chromatin immunoprecipitation assay (ChIP). Using an inflammatory stimulus, we found that Rac1 and NF-κB form a protein complex by Proximity Ligation Assay, which is disrupted by Rac1- deletion. Gain-of-function mutations for Rac1 rescued IL-1β expression in Rac1-deleted macrophages, and mutations in the hypervariable region of Rac1 confirmed that the nuclear localization of Rac1 is required for the rescue of IL-1β expression. This recent preliminary data demonstrated that the small GTPase, Rac1, acts as a transcriptional co-factor, helping to activate NF-κB, facilitating NF-κB’s translocation to the nucleus, and binding the IL-1β promoter. In a hyperlipidemia model of experimental atherosclerosis, macrophage Rac1- deletion reduced systemic IL-1β levels and decreased plaque calcification. Targeted inhibition of the macrophage Tiam1-Rac1 signaling complex may have potential as a therapeutic strategy to mitigate inflammatory-driven vascular calcification, given that small molecule inhibitors of Tiam1-Rac1 already exist.