Adiponectin, an adipocyte-derived protein, exerts anti-atherosclerotic effects on the vascular endothelium. Recently adiponectin protein has been reported in murine vascular endothelial cells, however, whether adiponectin is present in human vascular endothelial cells remains unexplored. We sought to examine 1) adiponectin protein in vascular endothelial cells collected from older adults free of overt cardiovascular disease; 2) the relation between endothelial cell adiponectin and in vivo vascular endothelial function; and 3) the relation between endothelial cell adiponectin, circulating (plasma) adiponectin and related factors. We measured vascular endothelial function (brachial artery flow-mediated dilation using ultrasonography), vascular endothelial cell adiponectin (biopsy coupled with quantitative immunofluorescence) and circulating adiponectin (Mercodia, ELISA) in older, sedentary, non-smoking, men and women (55 – 79 years). We found that higher endothelial cell adiponectin was related with greater flow-mediated dilation (r=0.43, P<0.05) and greater flow-mediated dilation normalized for shear stress (r=0.56, P<0.01), but was not related with vascular smooth muscle responsiveness to nitric oxide (r=0.04, P=0.9). Vascular endothelial cell adiponectin was not related with circulating adiponectin (r=−0.14, P=0.6). Endothelial cell and circulating adiponectin were differentially associated with adiposity, metabolic and other factors, but both were inversely associated with renal function (r=0.44 to 0.62, P ≤ 0.04). In conclusion, higher endothelial cell adiponectin levels are associated with higher vascular endothelial function, independent of circulating adiponectin levels in older adults.