Rationale: Cardiovascular disease (CVD) is the leading cause of death worldwide. While advances in CVD therapeutics have been made, there remain unmet needs for better approaches to prevent and treat various forms of CVD such as coronary atherosclerosis and peripheral artery disease (PAD). The McNamara lab has identified the helix-loop-helix transcription factor Inhibitor of differentiation 3 (Id3) as a regulator of atherosclerosis development in both mice and humans. In my studies, I sought to identify cell type-specific roles for Id3 in regulating atherosclerosis development and adaptive neovascularization in the setting of skeletal muscle ischemia as a model of PAD. I hypothesized that Id3 expression in endothelial cells (ECs) and macrophages plays a protective role during atherosclerosis development and B cell-specific Id3 expression promotes angiogenesis during skeletal muscle ischemia to restore blood flow to distal limbs.

Approach: To address these questions, a number of cell-specific transgenic murine models were utilized and generated. B cell-, macrophage-, and EC-specific Id3 knockout lines were used to investigate whether Id3 in these respective cell types regulated vascular density and perfusion during hind limb ischemia (HLI). Macrophage- and EC-specific Id3 knockout lines were used to investigate whether Id3 inhibited progression of plaque development of the aorta in hyperlipidemic mice (a model of atherosclerosis). Additional assays including, but not limited to, flow cytometry, ELISA, immunofluorescence, and cell culture were applied to quantify cell populations, Ig production, plaque size and characteristics, and EC proliferation where relevant.

Results: HLI experiments reveal that B cell-specific Id3 KO, but not macrophage- or EC-specific Id3 KO, have reduced blood flow during HLI. These results correlate with increased presence of B-1b numbers and IgM levels both within ischemic skeletal muscle as well as globally. Preliminary atherosclerosis experiments suggest that EC-specific Id3 KO, but not macrophage-specific Id3 KO mice, have increased atherosclerosis development compared to WT littermates. Macrophage-specific Id3 KO mice demonstrate increased B cell numbers in aortic perivascular adipose tissue (PVAT).

Conclusion: These studies are the first to investigate EC-, macrophage-, and B cell-specific roles of Id3 in atherosclerosis and a murine model of PAD. EC-specific Id3 is atheroprotective, implicating a new molecular mediator through which ECs regulate lesion formation in the setting of hyperlipidemia. B cell-specific Id3 promotes blood flow recovery during ischemia, implicating a potential novel cell type (B cells) in neovascular adaptation to ischemia during PAD. These findings reveal novel cellular mechanisms regulating vascular disease progression and potential new therapeutic targets.