BINDING OF PROTEINASE-3 AND MYELOPEROXIDASE TO ENDOTHELIAL-CELLS - ANCA-MEDIATED ENDOTHELIAL DAMAGE THROUGH ADCC

Binding of both proteinase 3 (PR3) and myeloperoxidase (MPG) to endoth elial cells (EC) has been suggested to be involved in the vascular dam age seen in patients with Wegener's granulomatosis or microscopic poly angiitis. In the present study we investigated in detail the interacti on of MPO and PR3 with cultured human umbilical vein endothelial cells (HUVEC) and its matrix products. In addition, we investigated whether interaction of PR3 or MPO with HUVEC monolayers also resulted in anti body-dependent cell-mediated cytotoxicity (ADCC) mediated by anti-neut rophil cytoplasmic antibody (ANCA)-positive patient sera or rabbit IgG anti-PR3 or anti-MPO. Preincubation of HUVEC monolayers with PR3 or M PO resulted in a dose-dependent binding of both PR3 and MPO. However, HUVEC, preincubated with PR3 or MPO, followed by ANCA or by rabbit ant i-PR3 or anti-MPO, were not susceptible to ADCC. Detailed analysis of the binding of PR3 to HUVEC monolayers showed that PR3 binds primarily to the extracellular matrix of endothelial cells, and to a very limit ed extent to the cells themselves. For MPO it was shown that it binds both to the extracellular matrix and to the endothelial cells themselv es. However, after binding to HUVEC cultures, MPO was not detectable b y polyclonal rabbit or human antibodies specific for MPO, probably bec ause MPO is bound to sites not accessible for immunoglobulins. Binding of PR3 to HUVEC cultures (cells + matrix) was inhibited by fetal calf serum and by alpha(1)-antitrypsin, but inactivation of enzymatic acti vity of PR3 by PMSF did not influence binding of PR3 to HUVEC cultures . Binding of MPO to HUVEC cultures was not influenced by fetal calf se rum.