Fine Regulation of Neutrophil Oxidative Status and Apoptosis by Ceruloplasmin and Its Derivatives

Timely neutrophil apoptosis is an essential part of the resolution phase of acute inflammation. Ceruloplasmin, an acute-phase protein, which is the predominant copper-carrying protein in the blood, has been suggested to have a marked effect on neutrophil life span. The present work is a comparative study on the effects of intact holo-ceruloplasmin, its copper-free (apo-) and partially proteolyzed forms, and synthetic free peptides RPYLKVFNPR (883-892) and RRPYLKVFNPRR (882-893) on polymorphonuclear leukocyte (PMNL, neutrophil) oxidant status and apoptosis. The most pronounced effect on both investigated parameters was found with copper-containing samples, namely, intact and proteolyzed proteins. Both effectively reduced spontaneous and tumor necrosis factor-alpha(TNF-alpha)-induced extracellular and intracellular accumulation of superoxide radicals, but induced a sharp increase in the oxidation of intracellular 2',7'-dichlorofluorescein upon short exposure. Therefore, intact and proteolyzed ceruloplasmin have both anti- and pro-oxidant effects on PMNLs wherein the latter effect is diminished by TNF-alpha and lactoferrin. Additionally, all compounds investigated were determined to be inhibitors of delayed spontaneous apoptosis. Intact enzyme retained its pro-survival activity, whereas proteolytic degradation converts ceruloplasmin from a mild inhibitor to a potent activator of TNF-alpha-induced neutrophil apoptosis.