MEMBRANE FLUIDITY CHANGES ACCOMPANYING PHAGOCYTOSIS IN NORMAL AND IN CHRONIC GRANULOMATOUS-DISEASE POLYMORPHONUCLEAR LEUKOCYTES

Abstract

Membrane fluidity changes in polymorphonuclear leukocytes (PMN) during phagocytosis were studied. Membrane fluidity was assessed by ESR using a nitroxide-substituted stearic acid analog (5DS) as a spin probe. PMN from normal subjects and from 3 CGD [chronic granulomatous disease] patients (2 males, 1 female) were incubated in Kreb''s Ringers phosphate with or without opsonized zymosan. ESR spectra were obtained and the order parameter (S), which is inversely related to membrane fluidity, was calculated. Without zymosan addition, S for normal (0.638) and for CGD (0.635) were not significantly different (P > 0.35). The S values indicate that under resting conditions the molecular environment of the CGD membrane is similar to that of normal PMN membranes. With addition of opsonized zymosan, the normal, but not the CGD, PMN showed a significant increase (CGD, S = 0.638; normal, S = 0.647; P < 0.001). This change in S for the normals is consistent with a more restricted movement of 5DS. Treatment of normal PMN with a mixture of scavengers specific for H2O2 (catalase, 1600 U/ml), .**GRAPHIC**. (superoxide dismutase, 100 .mu.g/ml) and for HO., (sodium benzoate, 1mM) during zymosan stimulation gave S values similar to those of resting cells. Catalase alone also lowered S value, suggesting that H2O2 was instrumental in causing the initial S value increase. This idea was supported by studies in which CGD cells were incubated with zymosan in the presence of glucose oxidase, an enzyme that catalyzes glucose oxidation resulting in the direct reduction of molecular oxygen to H2O2. Reduced O2 by-products, particularly H2O2, may cause altered biophysical properties of PMN membrane during phagocytosis.