Abstract Body: The prevalence of overweight/obesity continues to increase in the United States. With increased adiposity there is a shift in the phenotype and biology of adipose tissue resulting in an adipocyte secretome with proatherogenic consequences. Clinical interest in adipocyte-derived extracellular vesicles (Ad-EVs) has intensified due to their potential as systemic mediators of overweight- and obesity-related cardiometabolic dysfunction. While there are experimental data to suggest that Ad-EVs contribute to cardiovascular risk, to date there are no clinical data establishing a link between circulating Ad-EVs and vascular endothelial function. The purpose of this ongoing study is to determine: 1) if circulating Ad-EV levels are elevated in adults with overweight and obesity, independent of other cardiometabolic risk factors; and if so, 2) whether circulating Ad-EVs are associated with adiposity-related endothelial vasodilator dysfunction. Twenty-four sedentary, mid-life adults (45-66 years) were studied: 8 normal weight (4M/4F; age: 56+3 yr; BMI: 21.2+0.7 kg/m²); 8 overweight (4M/4F; 56+2 yr; 28.4+0.3 kg/m²); and 8 obese (4M/4F; 57+2 yr; 31.5+0.8 kg/m²). All subjects were free of overt cardiometabolic disease. Ad-EV identification (perilipin A⁺ vesicles) and concentration in peripheral blood were determined by flow cytometry. Forearm blood flow (FBF: via plethysmography) was assessed in response to intra-arterial infusions of acetylcholine (4.0, 8.0 and 16.0 μg/100 mL tissue/min) and sodium nitroprusside (1.0, 2.0 and 4.0 μg/100 mL tissue/min). Circulating Ad-EVs were markedly higher (P<0.001) in overweight (236+35 Ad-EV/µL) and obese (417±57 Ad-EV/µL) compared with normal weight (107±11 Ad-Ev/µL) adults. Additionally, Ad-EVs were higher (P<0.004) in the obese vs overweight adults. FBF response to acetylcholine was significantly lower (~35%) in the overweight (from 4.5±0.3 to 11.5±0.8 mL/100 mL tissue/min) and obese (from 4.7±0.3 to 9.7±0.8 mL/100 mL tissue/min) vs normal weight group (from 5.0±0.4 to 16.1±1.1 mL/100 mL tissue/min). Ad-EVs were strongly and inversely associated with the vasodilator response to acetylcholine (r=-0.74; P<0.001). In conclusion, increased adiposity, independent of other cardiometabolic risk factors, is associated with elevated circulating levels of Ad-EVs. Moreover, these circulating Ad-EVs may serve as a biomarker of, and possible contributor to, adiposity-related endothelial dysfunction and vascular disease risk.