Auranofin affects early events in human polymorphonuclear neutrophil activation by receptor-mediated stimuli

Auranofin, a new oral antirheumatic gold compound, in concentrations achieved therapeutically, inhibits neutrophil phagocytosis, chemotaxis, chemiluminescence, reduction of cytochrome c, and release of lysosomal enzymes. To further characterize the mechanism by which auranofin affects neutrophils, we studied the effects of auranofin on unstimulated properties and functions of neutrophils as well as on rapidly stimulated functions. When examined by electron microscopy, 4 micrograms/ml of auranofin significantly decreased the number of visualized centriole-associated microtubules in resting cells. Furthermore, auranofin inhibited neutrophil spreading on glass and caused a decrease in negative surface charge (electrophoretic mobility). In addition, auranofin inhibited several fmet-leu-phe-stimulated responses such as shape change, increases in centriole-associated microtubules, decreases in surface charge, and elicited membrane potential changes (di-O-C5(3) dye response). Auranofin (1 micrograms/ml) inhibited fmet-leu-phe-stimulated superoxide and hydrogen peroxide production by 80% (p less than 0.05), and also increased the affinity of receptors for fmet-leu-phe (from Ka 0.035 to Ka 0.48, p less than 0.001). Auranofin also affected neutrophil responses to phorbol myristic acetate (PMA). The total amount of PMA-stimulated superoxide production was suppressed by as little as 0.4 micrograms/ml of auranofin, but the lag time for activation was shortened by low concentrations of auranofin (0.5 to 1 microgram/ml). Four micrograms per milliliter of auranofin suppressed the decrease in surface charge induced by PMA. However, auranofin did not influence superoxide production elicited by the ionophore A23187. The results indicate that auranofin affects the earliest detected responses in neutrophil activation by certain receptor-mediated stimuli.     

Neutrophil-stimulating activity of staphylococci based on reactive chemiluminescence data

The neutrophil-stimulating properties of 38 S. aureus strains and 32 S. epidermidis strains were studied in the reaction of luminol-mediated chemiluminescence. All S. aureus strains and 29 S. epidermidis strains were found to possess neutrophil-stimulating activity, the mean activity index for S. aureus being significantly higher. The stimulating activity of the strains varied within a wide range (the variation coefficient was 120.0 +/- 21.9%) and did not correlate with the content of protein A in bacterial cells and the degree of their hydrophoby. The opsonization of staphylococci with normal human serum enhanced the neutrophil reaction 1.5- to 100-fold and simultaneously leveled out the chemiluminescence indices in experiments with different strains (the variation coefficient was 8.0 +/- 1.5%). The nature of the neutrophil-stimulating effect of staphylococci and its relationship to the exploratory reactions of phagocytes are discussed.     

DIFFERENTIAL INCORPORATION OF [3H]LYSINE INTO VISUAL CORTEX PROTEIN FRACTIONS DURING FIRST EXPOSURE TO LIGHT

—A resolution of the enhancement of protein synthesis in the visual cortex of rats during first exposure to light (Richardson and Rose , 1972) was achieved by polyacrylamide gel electrophoresis using a double-labelling technique. Differential incorporation of lysine was established between exposed and control animals in two fractions of the soluble proteins and seven fractions of the insoluble proteins. This suggests that exposure to a new experience of this type involves a specific effect on protein synthesis, rather than a general stimulation across all fractions.     

Determination of plasma leukotrienes in antigen-induced bronchoconstrictive guinea pigs.

Convenient extraction and radioimmunoassay methods for measurement of leukotrienes C4 and D4 (LTC4 and LTD4) in biological fluids are described. LTC4 or LTD4 in plasma was extracted with acetonitrile, and the extract was washed with dichloromethane then adjusted to pH 3.5 or 6.0, respectively. Each leukotriene was partially purified by using a C18-bonded silica cartridge and quantitated by radioimmunoassay. Amounts of LTC4 and LTD4 in the range of 0.025-1.6 ng could be assayed in plasma. This procedure was employed to examine the increase in plasma LTC4 (0.249 +/- 0.036 ng/ml) and LTD4 (1.399 +/- 0.235 ng/ml) of guinea pigs during intravenous challenge-induced anaphylactic bronchoconstriction, and the suppression of the increase of bronchoconstriction and leukotrienes by the administration of 5-lipoxygenase inhibitors such as E6080 (6-hydroxy-2-(4-sulfamoylbenzyl-amino)- 4,5,7-trimethylbenzothiazole hydrochloride), AA861 (2,3,5-trimethyl-6-(12-hydroxy-5,10-dodecadiynyl)-1,4-benzoquinone ) and phenidone. On the other hand, LTC4 and LTD4 were not detected in plasma after an inhaled challenge, though significant bronchoconstriction was provoked. It was concluded that the present study validates a new technique for quantitating plasma leukotrienes on the basis of pH and a suitable method for evaluating the pharmacological efficacy of 5-lipoxygenase inhibitors.