Endothelial cell (EC) activation and dysfunction increases with age and is linked to a variety of chronic vascular inflammatory disease states including atherosclerosis—the major cause of morbidity and mortality in Western Societies. Vascular senescence induced by the DNA damage response (DDR) promotes chronic inflammation in atherosclerotic lesions. Senescence-associated proinflammatory cytokines and proatherogenic risk factors (acquired or inherited) such as hyperlipidemia activate key signaling pathways that increase expression of adhesion molecules, chemokines on several cell types, including the vascular endothelium. Therefore, suppressing the senescence-associated inflammatory response in the vascular endothelium may provide a novel therapeutic approach to limit atherosclerosis. MicroRNAs (miRNAs) are small, single-stranded, evolutionary conserved non-coding RNAs that suppress the expression of target genes at the post-transcriptional level and participates in a variety of pathophysiological processes including the regulation of inflammatory responses. Our group provided the initial link implicating miR-181b in suppressing endothelial cell inflammation. During the last grant period, we identified miR-181b as a nodal regulator of endothelial cell quiescence through its regulatory effects on two major signaling pathways – NF-κB and AKT/eNOS. Consequently, endogenous miR-181b was found to function as a key determinant of the inflammatory response in vivo, findings that correlate with human inflammatory states including established coronary artery disease. We now identify endothelial miR-181b as a critical determinant of systemic vascular inflammation and atherosclerosis by controlling vascular senescence and the DNA damage response. Furthermore, we demonstrate that the adenosine-A3AR signaling pathway, a translationally relevant target that suppresses endothelial activation, functions in a miR-181b-dependent manner. These observations provide the foundation for the central hypothesis that endothelial miR-181b, via inhibitory effects on the DNA damage response, regulates senescence-associated vascular inflammation and atherosclerosis. To better understand the precise role of miR-181b in regulating vascular senescence, inflammation, and atherosclerosis, we propose 3 aims. In Aim1, we will determine the molecular basis for miR- 181b to regulate the DNA damage response and vascular senescence in response to diverse stimuli. In Aim2, we will explore the effect of altering miR-181b expression on senescence-associated secretory phenotype and atherosclerotic progression and regression in young and aged mice. In Aim3, we will determine whether the anti-senescent effects of adenosine in the vascular endothelium depend on miR-181b. This multi-disciplinary team in the fields of non-coding RNA biology, molecular imaging, nanomedicine, bioinformatics, and atherosclerosis research will establish an unprecedented molecular view of this miRNA in lesions that ...