Artocarpol A stimulation of superoxide anion generation in neutrophils involved the activation of PLC, PKC and p38 mitogen‐activated PK signaling pathways

1 Artocarpol A (ART), a natural phenolic compound isolated from Artocarpus rigida , stimulated a slow onset and long‐lasting superoxide anion generation in rat neutrophils, whereas only slightly activated the NADPH oxidase in a cell‐free system. 2 Pretreatment of neutrophils with pertussis toxin (1  μ g ml −1 ), 50  μ M 2′‐amino‐3′‐methoxyflavone (PD 98059), or 1  μ M 1,4‐diamino‐2,3‐dicyano‐1,4‐bis(2‐aminophenylthio)butadiene (U0126) had no effect on ART‐stimulated superoxide anion generation. ART (30  μ M ) did not induce extracellular signal‐regulated kinase (ERK) phosphorylation. 3 4‐(4‐Fluorophenyl)‐2‐(4‐methylsulfinylphenyl)‐5‐(4‐pyridyl)‐1 H ‐imidazole (SB 203580) markedly attenuated the ART‐stimulated superoxide anion generation (IC 50 value of 4.3±0.3  μ M ). Moreover, ART induced p38 mitogen‐activated PK (MAPK) phosphorylation and activation. 4 The superoxide anion generation in response to ART was also substantially inhibited in a Ca 2+ ‐free medium, and by pretreatment with 1  μ M 1‐[6‐((17 β ‐3‐methoxyestra‐1,3,5(10)‐trien‐17‐yl)amino)hexyl]‐1 H ‐pyrrole‐2,5‐dione (U‐73122) and 100  μ M 2‐aminoethyldiphenyl borate (2‐APB). ART (30  μ M ) stimulated the [Ca 2+ ] i elevation in the presence or absence of external Ca 2+ , and also increased the D ‐ myo ‐inositol 1,4,5‐trisphosphate formation. 5 2‐[1‐(3‐Dimethylaminopropyl)‐1 H ‐indol‐3‐yl]‐3‐(1 H ‐indol‐3‐yl)‐maleimide (GF 109203X) greatly inhibited the ART‐stimulated superoxide anion generation (IC 50 value of 7.8±1.0 n M ). ART increased the recruitment of PKC‐ α , ‐ β I, and ‐ β II to the plasma membrane of neutrophils, and stimulated Ca 2+ ‐dependent PKC activation in the cytosol preparation. 6 ART induced the phosphorylation of p47 phox , which was attenuated by GF 109203X. Moreover, ART evoked the membrane association of p47 phox , which was inhibited by GF 109203X and SB 203580. 7 These results indicate that the ART stimulation of superoxide anion generation involved the activation of p38 MAPK, PLC/Ca 2+ , and PKC signaling pathways in rat neutrophils.British Journal of Pharmacology (2005) 145 , 460–468. doi: 10.1038/sj.bjp.0706205