Relationship between calcium release and NADPH oxidase inhibition in human neutrophils

The aim of this study was to investigate the possible relationship between NADPH oxidase activity and changes in cytosolic Ca2+ in response to different agonists. Treatment of neutrophils with leukotriene B4 (LTB4) demonstrated characteristic changes to cytoslic Ca2+ yielding an EC50 of 4 nM. The pA2 values for the specific LTB4 receptor (BLT) antagonists, U-75302 and LY-255283 were 6.32 and 6.38, respectively. Similarly, neutrophils treated with N-formyl-l-methionyl-l-leucyl-l-phenylalanine (FMLP) and platelet activating factor (PAF) exhibited changes in cytoslic Ca2+ in a dose dependant manner with pD2 values of 9.0 and 9.9, respectively. The phorbol ester PMA prevented elevations in cytosolic Ca2+ in response to LTB4, FMLP and PAF with IC50 values of 5.88, 1.44 and 5.71 nM, respectively. In addition, potent NADPH oxidase inhibitors apocynin and diphenyleneiodonium (DPI) inhibited FMLP mediated cytosolic Ca2+ release. These results demonstrate that inhibition of the NADPH oxidase suppresses cytosolic Ca2+ release in FMLP activated human neutrophils.