Mechanisms of oxyradical production in substance P stimulated rheumatoid synovial cells.
We examined the intracellular mechanisms of substance P induced oxyradical production in rheumatoid synovial cells by the luminol-dependent chemiluminescence method. After stimulation with substance P (30 microM), single synovial A (macrophage-like) or B (fibroblast-like) cells released oxyradicals such as superoxide anions (O2-) and/or hypochlorous anions (OCl-) under a microscope equipped with an ultrasensitive photonic image intensifier. The substance P induced oxyradical production was blocked by a tachykinin NK1 (NK1) receptor antagonist, GR82334, GTP-binding protein (G-protein) inactivators, GDP beta S and islet-activating protein (IAP), and a phospholipase C (PLC) inhibitor, U-73122. Substance P (30 microM) also induced a transient increase in the intracellular 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 substance P induced increase in [Ca2+]i, but they had no effects on oxyradical production. In contrast to the effects of BAPTA-AM and heparin, protein kinase C (PKC) inhibitors, H-7 and calphostin C, completely inhibited substance P induced oxyradical production without any significant effects on [Ca2+]i increase. These findings suggest that the NK1 receptor/PLC-linked diacylglycerol (DAG) formation with the resulting activation of PKC is the main signal transduction pathway for substance P stimulated oxyradical production in synovial cells.
Tanabe
T
T
Department of Pharmacology, Kansai Medical University, Osaka, Japan.
Otani
H
H
Mishima
K
K
Ogawa
R
R
Inagaki
C
C
D001172:Arthritis, Rheumatoid / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D005260:Female; D005609:Free Radicals; D006801:Humans; D008297:Male; D013373:Substance P / Q000494:pharmacology*; D013583:Synovial Membrane / Q000378:metabolism*
D005609:Free Radicals; D013373:Substance P; D002118:Calcium