Inhibitory effects on human eosinophil chemotaxis in vitro by BAY 41-2272, an activator of nitric oxide-independent site of soluble guanylate cyclase

Abstract

This study was designed to investigate the effects of the 5-cyclopropyl-2-[1-(2-fluoro-benzyl)-1 H-pyrazolo[3,4- b]pyridin-3-yl]-pyrimidin-4-ylamine (BAY 41-2272) on formyl-methionyl-leucyl-phenylalanine (fMLP; 10 −7 M)-induced human eosinophil chemotaxis, cyclic guanosine-3′,5′-monophosphate (cGMP) and cyclic adenosine-3′,5′-monophosphate (cAMP) levels. Human eosinophils were pretreated or not with 3-isobutyl-l-methyl-xanthine (IBMX; 500 μM), and then exposed to BAY 41-2272 (0.1–10.0 μM) for either short (10 min) or prolonged (90 min) time periods. Exposition of eosinophils with BAY 41-2272 for either 10 min or 90 min markedly inhibited the eosinophil chemotaxis, independently of IBMX pretreatment. Inhibition of fMLP-induced eosinophil chemotaxis by BAY 41-2272 (in absence of prior treatment with IBMX) was about of the same irrespective if cells were exposed for 10 min or 90 min with this compound. In IBMX-pretreated eosinophils, the inhibition of fMLP-induced chemotaxis by BAY 41-2272 in the 10-min exposure protocols was even higher in comparison with the 90-min protocols. Incubation of IBMX-treated eosinophils for 90 min with BAY 41-2272 resulted in 2.0–2.5 times higher levels of cGMP and cAMP compared with the 10-min protocols. The BAY 41-2272-induced cGMP increases were abolished by pre-incubation of eosinophils with the soluble guanylate cyclase inhibitor 1 H-[1,2,4]-oxidiazolo[4,3- a] quinoxalin-1-one (ODQ). No eosinophil toxicity was observed in any experimental condition, according to 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) assay. Our findings show that inhibitory effects of fMLP-induced human eosinophil chemotaxis by BAY 41-2272 at short-term or prolonged exposition time are accompanied by significant elevations of cGMP and cAMP, but we could not detect a clear correlation between chemotaxis inhibition and elevation of cyclic nucleotide levels.