MECHANISMS OF OXYRADICAL PRODUCTION IN SUBSTANCE-P STIMULATED RHEUMATOID SYNOVIAL-CELLS

We examined the intracellular mechanisms of substance P induced oxyrad ical production in rheumatoid synovial cells by the luminol-dependent chemiluminescence method. After stimulation with substance P (30 mu M) , single synovial A (macrophage-like) or B (fibroblast-like) cells rel eased oxyradicals such as superoxide anions (O-2(-)) and/or hypochloro us anions (OCl-) under a microscope equipped with an ultrasensitive ph otonic image intensifier. The substance P induced oxyradical productio n was blocked by a tachykinin NK1 (NK1) receptor antagonist, GR82334, GTP-binding protein (G-protein) inactivators, GDP beta S and islet-act ivating protein (IAP), and a phospholipase C (PLC) inhibitor, U-73122. Substance P (30 mu M) also induced a transient increase in the intrac ellular Ca2+ concentration ([Ca2+](i)) in both synovial A and B cells as measured by a Ca2+ indicator, fura 2. BAPTA-AM and an inositol-1,4- 5-triphosphate (IP3) receptor antagonist, heparin, inhibited the subst ance P induced increase in [Ca2+](i), but they had no effects on oxyra dical production. In contrast to the effects of BAPTA-AM and heparin, protein kinase C (PKC) inhibitors, H-7 and calphostin C, completely ou t any significant effects on oxyradical production with. increase, The se findings suggest that the NK1 receptor/PLC-linked diacylglycerol (D AG) formation with the resulting activation of PKC is the main signal transduction pathway for substance P stimulated oxyradical production in synovial cells.