Exocytosis from Weibel-Palade bodies, the secretory granules of vascular endothelial cells, causes the rapid release of von Willebrand factor (vWF), an adhesive glycoprotein involved in primary hemostasis, and cell surface expression of P-selectin, a membrane protein involved in neutrophil binding. Thus, exocytosis may represent a link between hemostasis and inflammation. We investigated the effect of reactive oxygen intermediates (ROIs) on vWF secretion. Incubation of cultured endothelial cells with xanthine oxidase (XO), which generates superoxide anions (O2-), induces a potent, rapid secretory response. However, vWF release was not observed in response to H2O2. Extracellular, subendothelial vWF deposits typically seen after exocytosis from Weibel-Palade bodies were observed after exposure to XO. XO caused a rapid, sustained increase in intracellular free calcium concentration ([Ca2+]i). vWF secretion was markedly inhibited by BAPTA-AM, a cell-permeant calcium chelator. Removal of extracellular calcium did not inhibit vWF release, although the sustained phase of the [Ca2+]i increase was suppressed. These results suggest that XO-induced vWF release is mediated by the initial increase in [Ca2+]i which is caused by calcium mobilization from intracellular stores rather than by calcium influx. Exocytosis from Weibel-Palade bodies may contribute to the pathogenic effect of ROIs in atherosclerosis and inflammation.