Vol. 297, Issue 1, 388-394, April 2001

Rebamipide Suppresses Formyl-methionyl-leucyl-phenylalanine (fMLP)-Induced Superoxide Production by Inhibiting fMLP-Receptor Binding in Human Neutrophils

Chifumi Nagano, Atsushi Azuma, Hironobu Ishiyama, Kazuo Sekiguchi, Kenichi Imagawa and Mikio Kikuchi

Third Institute of New Drug Research, Otsuka Pharmaceutical Co., Ltd., Kawauchi-cho, Tokushima, Japan

The purpose of the present work was to investigate the mechanism underlying the inhibitory action of rebamipide on superoxide anion (O₂) production induced by the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (fMLP) in human neutrophils. Phosphatidylinositol 3,4,5-trisphosphate (PIP₃), a product of phosphoinositide 3-OH-kinase (PI 3-kinase) accumulated in response to fMLP and this accumulation was well correlated with O₂ production in human neutrophils. Rebamipide inhibited PIP₃ production in parallel with the inhibition of fMLP-induced O₂ production. PI 3-kinase activity in anti-PI 3-kinase p85 immunoprecipitates was not affected by the presence of rebamipide, therefore rebamipide did not have a direct inhibitory action on PI 3-kinase activity. Since rebamipide had no inhibitory effect on O₂ production induced by NaF, a direct activator of G protein, the target of the inhibitory action of rebamipide appears to be a component of the signal transduction pathway upstream of G protein. Scatchard analysis of [³H]fMLP binding to human neutrophil membrane revealed that rebamipide increased the K_D value of [³H]fMLP without altering the number of [³H]fMLP binding sites, suggesting that rebamipide has a competitive antagonistic action against the fMLP-receptor. The competitive antagonistic action was further confirmed by the finding that rebamipide caused a parallel shift to the right in the dose-response curve of O₂ production induced by fMLP. These results provide evidence that the competitive inhibitory action of rebamipide on the fMLP-receptor plays a main role in its inhibitory action on fMLP-induced O₂ production.