PROJECT SUMMARY Obesity is a primary driver of cardiometabolic disease and a major contributor to American mortality. One potential mechanism of this is elevated ectopic lipid deposition in highly metabolic organs such as the liver and skeletal muscle (SKM) leading to local and eventually systemic metabolic dysregulation. Myosteatosis is defined as the infiltration of fat into skeletal muscle and is characterized by loss of muscle mass and accumulation of intracellular lipids. These exogenously sourced fats contribute to an elevated lipogenic/lipolytic state whereby lipid metabolites accumulate and impair insulin signaling, contributing to the development of systemic insulin resistance and eventually cardiometabolic syndrome (CMS). Exercise is known to reverse insulin resistance and ameliorate CMS independent of weight loss, though the mechanisms remain incompletely understood. Moreover, while benefits of exercise are recognized, patient compliance and aging impede exercise utility, suggesting the need for pharmacological interventions. Because SKM is the principal site of glucose disposal and the most immediate effector of exercise-mediated adaptation, changes in its physiology provide the most obvious mechanism for exercise-induced improvements to cardiovascular health seen in obese patients. Preliminary data from this application establishes that obesity provides potent restraint on the growth of new blood vessels in the face of ischemia, which is reversed with increased SKM mass due to deletion of MSTN in mice, suggesting an important in vivo consequence of our prior studies showing profound endothelial dysfunction. In novel data we show that GAL3 and NOX1 expression in endothelial cells isolated from these mice are regulated by deletion of MSTN, suggesting that they are also mechanisms of impaired angiogenesis. The impact of GAL3 and NOX1 deletion on obesity-related vascular diseases in vivo is largely unexplored. In this application, we will explore the concept that obesity-driven ectopic fat accumulation in SKM drives vascular dysfunction and impaired angiogenesis in a GAL-3 and NOX1 dependent manner. This goal will be met in two specific aims. The first will use novel models uniquely developed in our lab to identify potential mechanistic drivers (GAL3 and NOX1) of impaired vascular function in obesity that have been identified as improved in obese hypermuscular mice. The second will use a newly made and novel knockout mouse design to resolve steatosis in a genetic model of obesity and measure cardiovascular and metabolic outcomes to determine if removing ectopic fat could serve as a viable therapeutic. The project will be directed under the mentorship of Dr. David Stepp in the Vascular Biology Center at the Medical College of Georgia at Augusta University, which has a rich history of successful pre- and post-doctoral training. The proposed project is for 3 years of funding with the planned aims divided amongst the 3 years of funding, conc...