Potential roles of Mg2+ and Ca2+ in NADPH oxidase dependent superoxide anion synthesis by human neutrophils.

Recent studies have shown that, in cell free systems, Mg2+, but not Ca2+, is absolutely required for the generation of reducing equivalents by neutrophil NADPH oxidase. Apparently Mg2+ is required for binding of cytosolic protein(s) to the plasma membrane in forming a functional NADPH oxidase complex. Using intact neutrophils made permeable to Mg2+ and Ca2+ by A23187 it was found that Mg2+ was required for the generation of reducing equivalents by PMA stimulated cells while Ca2+ inhibited cytochrome c reduction. On the other hand, FMLP-induced neutrophils were unable to generate reducing equivalents unless both Ca2+ and Mg2+ were present in the media. Sequential addition of these cations indicated that Ca2+ was required for transduction of the FMLP-receptor generated signal to a point in the pathway where Mg2(+)-dependent synthesis of reducing equivalents occurred. This step, presumably, is the Mg2(+)-dependent completion of the NADPH oxidase complex. Finally, approximately 50% of the neutrophil's 2.19 fmoles (6.32 mM) of Mg2+ was found to be in a readily exchangeable pool and that pretreatment of neutrophils with either FMLP or PMA increased this pool by only 4-5%. This suggests there is an adequate pool of Mg2+ available to support completion of NADPH oxidase complex and that a stimulus-induced rise in free Mg2+ is not required to assist in the formation of the complex.