PROJECT SUMMARY: Complications resulting from rupture of unstable atherosclerotic lesions, including myocardial infarction and stroke, are the leading cause of death worldwide. Despite decades of research, the mechanisms and factors leading to plaque rupture remain poorly understood. It is generally accepted that lesions with a high smooth muscle cell (SMC) to macrophage (Mϕ) ratio are less likely to rupture. However in the setting of atherosclerosis, SMC down-regulate their characteristic markers and express markers of other cells types and thus are undetectable using traditional markers. Indeed, rigorous SMC lineage tracing studies by our lab using Myh11 ERT2Cre eYFP ApoE-/- mice have shown that >80% of SMC within lesions lack expression of the markers previously used for their identification and that nearly 30% of these cells have activated multiple markers of Mϕ. Therefore, the number of SMC within lesions has not only been grossly underestimated, but many cells thought to be Mϕ are actually SMC-derived. Of even greater significance, we showed that SMC-specific conditional knockout of the stem cell pluripotency genes Klf4 and Oct4 had a profound impact on the pathogenesis of lesions including alterations in multiple indices of plaque stability. However, contrary to the dogma that SMC are always atheroprotective, we showed they can be either atheroprotective or -promoting depending on the nature of their phenotypic transitions. Taken together, studies highlight the importance of identifying factors and mechanisms that promote beneficial changes in SMC phenotype. An ongoing clinical trial is investigating neutralization of interleukin-1 in high-risk cardiovascular patients. The overarching hypothesis is that inflammation drives atherosclerosis, and that inhibition of inflammation will improve patient survival. However, there is a lack of preclinical evidence that neutralization of IL1 will confer beneficial effects in the setting of advanced atherosclerosis. Indeed, recent studies from our lab, which included the applicant, showed that treatment of our Myh11 ERT2Cre eYFP ApoE-/- mice with the Novartis IL1-neutralizing antibody after the establishment of advanced atherosclerosis resulted in multiple changes consistent with reduced plaque stability including marked reductions in the number of SMC-derived eYFP+ cells within the fibrous cap, and replacement of these cells with Mϕ. Studies in this proposal will test the hypothesis that IL1 signaling in SMC is critical for maintenance of plaque stability in late-stage atherosclerosis. Aim 1 test the hypothesis that the detrimental effects of anti-IL1 Ab treatment on late-stage atherosclerosis are primarily mediated through IL1R1 signaling in SMC and that this results in deleterious phenotypic transitions in lesion SMC. Aim 2 will test the hypothesis that increased production of interleukin-4 (IL4) following anti-IL1 Ab treatment of Myh11 ERT2Cre eYFP ApoE-/- mice with advanced lesions contribu...