BACKGROUND: Cardiovascular disease is highly prevalent in patients with chronic kidney disease. In hemodialysis patients, the protein-bound uremic retention solute p-cresol is independently associated with cardiovascular disease. The underlying mechanisms have not been elucidated. STUDY DESIGN: (1) Prospective observational study of humans and (2) in vitro study in human umbilical vein endothelial cells. SETTING: Hemodialysis patients. FACTOR: p-Cresol and its main derivative p-cresyl sulfate. OUTCOMES: Endothelial dysfunction. MEASUREMENTS: We studied: (1) the relation between p-cresol and blood markers of endothelial dysfunction, including soluble P-selectin and endothelial microparticles; and (2) direct effects of p-cresol and p-cresyl sulfate on endothelial cell cultures. RESULTS: (1) In a cohort of 100 maintenance hemodialysis patients, free serum p-cresol concentrations (median, 11.7 μmol/L; interquartile range, 15.2) were directly associated with circulating endothelial microparticles (P = 0.007), but not with soluble P-selectin (mean, 37.7 ± 14.4 [SD] pg/mL). Other independent determinants of the degree of circulating microparticles were greater serum phosphorus (mean, 4.8 ± 1.5 mg/dL; P = 0.008) and serum calcium concentrations (mean, 9.3 ± 0.8 mg/dL; P = 0.03), whereas treatment with active vitamin D (P = 0.008) and vintage (median, 25 months; P = 0.04) were inversely associated. (2) In vitro, p-cresyl sulfate induced a dose-dependent increase in the shedding of endothelial microparticles (P < 0.001) by human umbilical vein endothelial cells. Shedding was reduced, but not completely aborted, in the presence of albumin, whereas the selective Rho kinase inhibitor Y-27632 abrogated the p-cresyl sulfate–induced generation of endothelial microparticles. LIMITATIONS: The relationship between p-cresyl sulfate and shedding of endothelial microparticles in vivo was not mechanistically explored. CONCLUSION: p-Cresyl sulfate induces shedding of endothelial microparticles in the absence of overt endothelial damage in vitro and is independently associated with the number of endothelial microparticles in hemodialysis patients. These findings suggest that p-cresyl sulfate alters endothelial function in hemodialysis patients.