INDUCTION OF INTERLEUKIN-1 AND SUBSEQUENT TISSUE FACTOR EXPRESSION BYANTIPROTEINASE-3 ANTIBODIES IN HUMAN UMBILICAL VEIN ENDOTHELIAL-CELLS

Objective. To assess the ability of anti-proteinase 3 (anti-PR3) class ic antineutrophil cytoplasmic antibodies (cANCA) to stimulate endothel ial expression of tissue factor (TF), which is the main initiator of t he coagulation cascade that can lead to endothelial injury and thrombo sis in patients with Wegener's granulomatosis. Methods. Human umbilica l vein endothelial cells (HUVEC) were grown to confluence and stimulat ed with affinity-purified anti-PR3 antibodies, Igs from healthy subjec ts, and endotoxin (lipopolysaccharide) as positive control. Results. T F activity was generated in anti-PR3-stimulated cells, as shown by a c hromogenic test, This activity was inhibited by specific anti-TF antib odies, TP messenger RNA (mRNA) was found in anti-PR3-stimulated cells, as detected by reverse transcriptase-polymerase chain reaction, but n ot in cells stimulated with irrelevant human Igs or Igs from normal co ntrol sera, TF expression reached maximum levels 12 hours after exposu re to the anti-PR3 cANCA, and did not require complement, TF mRNA expr ession was inhibited by cycloheximide, suggesting a requirement for pr otein synthesis. When added to the incubation medium, interleukin-1 (I L-1) receptor antagonist inhibited the induced TF mRNA expression, sug gesting that cANCA-stimulated cells initiate IL-1 synthesis, Moreover, cANCA induced IL-1 alpha mRNA before TF mRNA. Conclusion. This study showed that anti-PR3 treatment of HUVEC induces sequential expression of IL-1 alpha mRNA and TF mRNA, as well as their corresponding protein s, Both proteins could have pathogenic roles in the vasculitic process , since TF is the main initiator of the coagulation cascade.