Exposure to monosodium urate crystals (MSU) stimulated the respiratory burst of human neutrophils as measured by increased O2 consumption and the generation of superoxide radicals (027. From the comparison of data derived from nitroblue tetrazolium and cytochrome C reduction (two methods of detecting 0 z r release), it appears that OZT production in response to MSU may be compartmentalized, i.e., occur predominantly in the intracellular space. After exposure to MSU, neutrophils from patients with chronic granulomatous disease lost viability at the normal rate; thus products of the respiratory burst are not likely to be responsible for cell death.The oxidative killing mechanisms of phagocytic cells have been shown to serve a fundamental role in the host defenses against microorganisms (1). Central to this scheme is the generation of superoxide ( 0 2 radicals formed by the univalent reduction of molecular oxygen (2,3). Superoxide and its reaction products hydrogen peroxide, hydroxyl radical, and singlet oxygen may contribute to phagocytic bactericidal capacity (1-3) and mediate cytotoxicity in vitro (43). Activation of the 02T generating system of the neutrophil has been shown to occur after phagocytosis of particulate stimuli [such as bacteria, latex spheres, and opsonized zymosan (2,3)], as well as after exposure of the cells to such diverse soluble stimuli as chemotactic factors, immune complexes, ionophores, lectins, and detergents (2,3). In this study, we compared the respiratory burst of cells from normal individuals and from chronic granulomatous disease (CGD) patients after stimulation by monosodium urate crystals (MSU). This crystal model permitted us to test hypotheses about the intracellular release of 02-and its potential cytotoxic role.
MATERIALS AND METHODSMedium. The medium used throughout this study was a modified Hanks' balanced salt solution supplemented with glucose (1 mg/ml) and bovine serum albumin (1 mg/ml), pH 7.40.Neutrophils. Human peripheral neutrophils were isolated from heparinized blood by sequential dextran sedimentation and Ficoll-Hypaque gradient centrifugation, as previously described (4).In addition to studies performed with cells from normal donors, neutrophils were obtained from 2 male patients with CGD. The diagnosis in both patients (ages 8 and 10) was based on the recurrence of pyogenic infections since early childhood and the absence of all components of respiratory burst activity when neutrophils were subjected to a variety of stimulating agents (3).Reagents and chemicals. The following reagents were purchased from Sigma Chemical Company: cytochalasin B,