Endothelial dysfunction precedes atherosclerosis and represents an important link between obesity and cardiovascular events. Strategies designed to prevent endothelial dysfunction may therefore reduce the cardiovascular complications triggered by obesity. We tested the hypothesis that deficiency of P-selectin glycoprotein ligand-1 (Psgl-1) would improve the endothelial dysfunction associated with obesity. Psgl-1-deficient (Psgl-1(-/-)) and wild-type (Psgl-1(+/+)) mice were fed standard chow or a high-fat, high-sucrose diet (diet-induced obesity [DIO]) for 10 weeks. DIO increased mesenteric perivascular adipose tissue (mPVAT) macrophage content and vascular oxidative stress in Psgl-1(+/+) mice but not in Psgl-1(-/-) mice. Pressure myography using mesenteric arteries demonstrated that relaxation responses to acetylcholine were significantly impaired in DIO Psgl-1(+/+) mice, whereas DIO Psgl-1(-/-) mice were protected from endothelial dysfunction with similar relaxation responses to Psgl-1(+/+) or Psgl-1(-/-) mice fed standard chow. The superoxide scavenger 4-hydroxy-2,2,6,6-tetramethylpiperidinyloxy (TEMPOL) partially recovered impaired endothelial function induced by DIO. A neutralizing Psgl-1 antibody was also effective in preventing endothelial dysfunction and reducing mPVAT macrophage content induced by DIO. These results indicate that obesity in mice leads to PVAT inflammation and endothelial dysfunction that is prevented by Psgl-1 deficiency. Psgl-1 inhibition may be a useful treatment strategy for targeting vascular disease associated with obesity.