Autoantibodies directed against cytoplasmic antigens of neutrophils (ANCA), especially those with specificity for proteinase 3 (PR-3) and myeloperoxidase, are valuable markers for differential diagnosis and monitoring of disease activity in Wegener's granulomatosis (WG) and other vasculitides. Till now, several concepts concerning a direct role of antibodies against PR-3 in the pathogenesis of WG have been discussed. Recently we were able to show that these antibodies recognize PR-3 translocated into the membrane of human endothelial cells. The aim of this study was to investigate putative cytotoxic effects of antibodies to PR-3 on human endothelial cells. Antibodies were obtained by affinity purification of sera from patients with active WG, Purified antibodies to Ro (SS-A), La (SS-B) and RNP served as controls. Purified antibodies to PR-3 displayed a lytic activity against endothelial cells treated with tumour necrosis factor-alpha (TNF-α) with the help of cytokine-primed neutrophils as measured in a Cr-release assay. About 100% specific cytotoxicity occurred after 4 h and was independent of complement, Cytotoxic effects were inhibited by coincubation with unprimed ncutrophils or preincubation of PR-3 antibodies with purified antigen. Antibodies to Ro (SS-A), La (SS-B) or RNP had no cytotoxic effect. In summary, PR-3 antibody-induced cytotoxicity required (i) expression of PR-3 on the surface of TNF-α-treated endothelial cells: and (ii) co-cultivation of cytokine-primed neutrophils. This is to the best of our knowledge the first report on direct cytotoxic effects of PR-3 antibodies on vascular endothelium. Our data give a hint at a PR-3 antibody-mediated mechanism of endothelial injury via antibody-dependent cellular cytotoxicity in WG and other ANCA-related vasculitides.