Inhibition of leukotriene B4-induced increase in intracellular calcium ion level of human peripheral blood polymorphonuclear leukocytes by Y-24180, an antagonist of platelet-activating factor receptor.
Y-24180 ((+/-)-4-(2-chlorophenyl)-2-[2-(4-isobutylphenyl)ethyl]-6,9-dim eth yl-6H-thieno[3,2-f] [1,2,4]triazolo[4,3-a] [1,4] diazepine), an antagonist of platelet-activating factor (PAF) receptor, has already been reported to inhibit leukotriene B4 (LTB4)-induced activation of polymorphonuclear leukocytes. In this article, to clarify the mechanism of inhibition of LTB4-induced activation by Y-24180, we examined the effect of Y-24180 on LTB4-induced increase in intracellular calcium ion ([Ca2+]i) level of human polymorphonuclear leukocytes at a single cell level using a laser scanning confocal microscope. Preincubation with Y-24180 significantly inhibited the increase in [Ca2+]i level at 0.3-3 microM, while WEB 2086 (4-[3-[4-(2-chlorophenyl)-9-methyl-6H-thieno[3,2-f][1,2,4]triazolo[4,3-a ][1,4]diazepin-2-yl]propionyl]morpholine), another PAF receptor antagonist, did not show the inhibitory effect at 3-30 microM, indicating that the difference in potency between these compounds was more than 100-fold. The LTB4-induced increase in [Ca2+]i level was suppressed by microinjection of anti-PAF antibody, but not control antibody, into leukocytes. Microinjection of Y-24180 at 0.003-0.03 microM or WEB 2086 at 0.03-0.3 microM into leukocytes also inhibited the LTB4-induced increase. The microinjection of WEB 2086 at the 10-fold greater concentrations than those of Y-24180 inhibited to the almost equal level to that obtained by Y-24180. In addition, microinjection of PAF into leukocytes induced the increase in [Ca2+]i level. Preincubation with Y-24180 at 1 or 3 microM significantly attenuated the PAF microinjection-induced increase, but WEB 2086 showed little effect at 3-30 microM. These results indicate that Y-24180 inhibits LTB4-induced activation of leukocytes by suppressing the increase in [Ca2+]i level and the inhibitory effect is mediated by antagonistic action against intracellular PAF-induced up-regulation of [Ca2+]i level. The difference in inhibitory activity for the increase in [Ca2+]i level between Y-24180 and WEB 2086 when these are added in the culture medium may depend upon their aptitude for the transmembrane influx.
Komatsu
H
H
Yoshitomi Pharmaceutical Industries, Ltd, Fukuoka, Japan. komatsu@yoshitomi.jp
Amano
M
M
D001381:Azepines / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D006801:Humans; D007975:Leukotriene B4 / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008845:Microinjections; D018613:Microscopy, Confocal / Q000379:methods; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D010972:Platelet Activating Factor / Q000494:pharmacology; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors*; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D014230:Triazoles / Q000494:pharmacology*
D001381:Azepines; D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D014230:Triazoles; C039351:platelet activating factor receptor; C052518:WEB 2086; C068035:4-(2-chlorophenyl)-2-(2-(4-isobutylphenyl)ethyl)-6,9-dimethyl-6H-thieno(3,2-f)(1,2,4)triazolo(4,3-a)(1,4)diazepine; D007975:Leukotriene B4; D002118:Calcium