Summary Heart disease in type 2 diabetes mellitus (T2DM) is directly related to the severity of coronary artery disease, which results in impaired coronary blood flow and increased risk of myocardial infarction (MI). T2DM patients are 2-4 times more likely to experience MI than non-diabetic patients. Although endothelial dysfunction and atherosclerosis are known to decrease coronary blood flow (CBF) in T2DM, the underlying mechanisms are not known. Adverse coronary resistance microvessel (CRM) remodeling is a critical mediator of T2DM-induced cardiovascular disease. Published studies from the Trask lab show that structural remodeling of CRMs occurs early in T2DM (db/db) mice and pigs with metabolic syndrome, leading to impaired CBF. While the mechanisms that govern CRM remodeling are unclear, our preliminary data point to impaired Jagged1 (Jag1)/Notch signaling. The Notch signaling pathway is known to facilitate communication between endothelial cells (ECs) and vascular smooth muscle cells (VSMCs). We found that expression of Jag1 and Notch3 in CRMs of T2DM (db/db) mice is reduced, and that mice lacking EC-expressed Jag1, or Notch3-null mice had impaired CBF similar to T2DM (db/db) mice. Furthermore, EC-Jag1-deficient mice have evidence of inward CRM remodeling. In addition, we have shown a significant reduction in fenestrae in the of CRMs of diabetic mice and pigs, and in EC-Jag1- deficient mice, suggesting that heterocellular EC-VSMC communication at the myoendothelial junction (MEJ) maybe be disrupted in all models. Our recent studies identified Notch3 and Jag1 at the MEJ of normal CRMs. Based upon our collective preliminary data, our overall hypothesis is that reduced coronary blood flow in T2DM is driven by impaired Jag1/Notch signaling between ECs and VSMCs. We will test this hypothesis using an integrative approach, combining genetically-deficient mouse models, EC-VSMC co-culture techniques, single- cell transcriptomics, and transgenic/drug rescue experiments: The Specific Aims are: Aim1. Test the hypothesis that T2DM causes a disruption of Jag1/Notch signaling in CRMs and define this mechanism. Aim 2. Determine if aberrant Jag1/Notch signaling contributes to adverse coronary microvascular remodeling in T2DM using genetic and transcriptomic approaches. Aim 3. Test the hypothesis that activation of Jag1/Notch signaling can prevent and/or treat T2DM-induced coronary microvascular remodeling. These studies will be the first to demonstrate Notch signaling is central to CRM remodeling and impaired CBF, and that correction of this pathway may reverse adverse CRM remodeling and reduced CBF in T2DM.