Previous studies have shown that polymorphonuclear leukocytes (PMNs) release a stable factor that inhibits endothelium-dependent relaxation. In the present studies, the effects of the factor on Ca2+ signaling and on ultrastructure of endothelial cells were investigated. In the cultured endothelial cells, the PMN-derived factor induced an increase in [Ca2+]i in a pattern of oscillations. The frequency of the Ca2+ oscillations was less than 3 spikes/10 minutes. Removal of extracellular Ca2+ by perfusion with Ca(2+)-free Krebs' solution abolished the spikes. The results of electron microscopy showed that this factor induced an increase in vesicle formation on the luminal surface of the rat aortic endothelium. The increased vesicle formation may represent exocytosis. The structure of the smooth muscle cells was not changed. In conclusion, the PMN-derived factor induces a Ca2+ influx and In conclusion, the PMN-derived factor induces a Ca2+ influx and possible exocytosis, suggesting that the factor may have other biological functions besides the inhibition of the vascular relaxation.