Loss of Id3 Increases VCAM-1 Expression, Macrophage Accumulation, and Atherogenesis in Ldlr^−/− Mice

Objective—Inhibitor of differention-3 (Id3) promotes B cells homing to the aorta and atheroprotection in Apoe^−/− mice. We sought to determine the impact of loss of Id3 in the Ldlr^−/− mouse model of diet-induced atherosclerosis and identify novel Id3 targets in the vessel wall.

Methods and Results—Ex vivo optical imaging confirmed that Id3^−/− Ldlr^−/− mice have significantly fewer aortic B cells than Id3^+/+ Ldlr^−/− mice. After 8 and 16 weeks of Western diet, Id3^−/− Ldlr^−/− mice developed significantly more atherosclerosis than Id3^+/+ Ldlr^−/− mice, with Id3^+/− Ldlr^−/− mice demonstrating an intermediate phenotype. There were no differences in serum lipid levels between genotypes. Immunostaining demonstrated that aortas from Id3^−/− Ldlr^−/− mice had greater intimal macrophage density and CCL 20 and vascular cell adhesion molecule 1 (VCAM-1) expression compared with Id3^+/+ Ldlr^−/− mice. Real-time polymerase chain reaction demonstrated increased VCAM-1 mRNA levels in the aortas of Id3^−/− Ldlr^−/− mice. Primary vascular smooth muscle cells from Id3^−/− mice expressed greater amounts of VCAM-1 protein compared with control. Gain and loss of function studies in primary vascular smooth muscle cells identified a role for Id3 in repressing VCAM-1 promoter activation. Chromatin immunoprecipitation demonstrated interaction of E12 with the VCAM-1 promoter, which is inhibited by Id3.

Conclusion—Id3 is an atheroprotective transcription regulator with targets in both B cells and vessel wall cells leading to reduced macrophages accumulation and reduced atherosclerosis formation.