Pharmacological comparison of LTB₄‐induced NADPH oxidase activation in adherent and non‐adherent guinea‐pig eosinophils

Leukotriene B 4 (LTB 4 ) stimulation of guinea‐pig peritoneal eosinophils, induced a biphasic activation of the NADPH oxidase composed of a rapid (<3 min) phase mediated by non‐adherent cells and a sustained (3 – 120 min) phase mediated by CD11b/CD18 adherent eosinophils. Studies were undertaken to compare the intracellular mechanism that mediate these responses. SB 203580 and PP1, inhibitors of p38 mitogen‐activated protein (MAP) kinase and the src‐family protein tyrosine kinases, respectively caused concentration‐dependent attenuation of both the rapid (SB203580: pD 2 =−6.31; PP1: pD 2 =−5.50) and sustained (SB203580: pD 2 =−6.50; PP1: pD 2 =−5.73) phases. Similarly, the MAP kinase kinase‐1 inhibitor, PD098059 produced partial inhibition of the both phases of superoxide generation. The protein kinase C (PKC) inhibitors Ro‐31 8220, GF 109203X and Gö 6976 attenuated the rapid NADPH oxidase response (pD 2 s=−6.10, −6.72, −6.15 respectively) and, to a lesser extent, (pD 2 s=−5.54, −6.02, −6.51 respectively) the sustained phase. An inhibitor of phosphatidylinositol 3‐kinase (PtdIns 3‐kinase), wortmannin caused concentration dependent attenuation of the sustained (pD 2 =−8.68) but not rapid phase of superoxide generation. In contrast, the syk kinase inhibitor, piceatannol abolished the rapid (pD 2 =−6.43) but not sustained respiratory responses. This study demonstrates that LTB 4 ‐induced superoxide generation from adherent and non‐adherent eosinophils is mediated via both common (p38 MAP kinase, MEK‐1, PKC and the src kinases) and divergent intracellular pathways (syk kinases and PtdIns 3‐kinase). This suggests the possibility of therapeutic intervention to selective attenuate activation of adherent tissue eosinophils. British Journal of Pharmacology (2001) 134 , 797–806; doi: 10.1038/sj.bjp.0704314