Inhibition Of Neutrophil Function Research Articles

Neutrophil sorbitol production impairs oxidative killing in diabetes.

The mechanism whereby hyperglycaemia inhibits neutrophil function is unclear. We have examined neutrophil killing of Candida albicans in the presence of increased concentrations of glucose and galactose. Killing was abolished in 50 mmol/l glucose and 10 and 50 mmol/l galactose. The oxidative phase of killing was examined using lucigenin-enhanced chemiluminescence. An increase in glucose concentration from 5 to 50 mmol/l produced a fall in chemiluminescence output from 128.5 +/- 16.8 (mean +/- SE) to 82.2 +/- 9.8 mV min (a reduction of 36%). These data suggest the existence of aldose reductase activity in neutrophils and using gas chromatography we have demonstrated the presence of sorbitol in extracts of diabetic neutrophils. As oxidative killing and sorbitol production are both NADPH-dependent the inhibition of killing is likely to be due to competition for this electron donor. This abnormality of neutrophil function may aggravate various infections in the patient with diabetes.

Effects of human serum on bacterial competition with neutrophils for molecular oxygen.

A dialyzable factor(s) in human serum is known to stimulate gonococcal oxygen consumption. Its effect on other human pathogens was investigated. A 10% serum solution increased peak O2 consumption for Escherichia coli and Staphylococcus aureus to 157% (P less than 0.05) and 199% (P less than 0.02), respectively, of their O2 consumption when suspended in Hanks balanced salt solution, compared with a 356% increase for Neisseria gonorrhoeae with serum. Dialyzed serum lacked stimulatory capacity. Bacteria, serum, and neutrophils are often incubated to evaluate neutrophil bactericidal activity. Samples of 10(8) N. gonorrhoeae, S. aureus, and E. coli turned resazurin colorless (anaerobic conditions, Eh less than -42 mV) after 7.4, 13.3, and 15.1 min, respectively. Because neutrophil formation of reactive oxygen intermediates requires ambient O2, the effect of live bacteria and serum on this process was explored. After 5 min of incubation of 10(8) N. gonorrhoeae or S. aureus in 10% normal or dialyzed serum, 10(5) neutrophils were added. Phorbol myristate acetate was then added to assure neutrophil stimulation, and luminol-dependent luminescence was measured. N. gonorrhoeae and S. aureus incubation in normal serum decreased peak LDL 91.7 and 88.6%, respectively, relative to incubation in dialyzed serum. A sample of 10(8) E. coli totally eliminated LDL. A sample of 10(8) E. coli incubated in Hanks balanced salt solution for 5 min also eliminated phorbol myristate acetate induced neutrophil H2O2 production. LDL inhibition increased in proportion to bacterial concentration and time of incubation and was prevented by inclusion of KCN. Increasing the concentration of neutrophils to 10(8) (1:1 particle-to-cell ratio) only partially reversed LDL inhibition. Re-aeration of the system allowed brief LDL which persisted only if KCN was added. Addition of KCN after bacterial incubation also permitted LDL, arguing against depletion of other factors from the media or accumulation of bacterially derived inhibitory substances. A dynamic competition for O2 occurs between bacteria and neutrophils. Serum stimulation of bacterial O2 utilization may contribute to virulence by increasing bacterial capacity to inhibit neutrophil function.

Preferential inhibition of primary granule release from bovine neutrophils by a Brucella abortus extract.

A low-molecular-weight Brucella abortus extract (a nucleotidelike material) inhibited zymosan-elicited neutrophil degranulation and trichloroacetic acid-precipitable protein iodination (a measure of myeloperoxidase and H2O2 release from neutrophilic leukocytes). Inhibition of neutrophil function was directly related to the concentration of the Brucella extract. The extract preferentially inhibited degranulation of primary (azurophilic or peroxidase positive) granules and had limited inhibition of secondary (specific or peroxidase negative) granule release but did not inhibit opsonized zymosan ingestion. Inhibition of protein iodination closely paralleled that of primary granule release but was unrelated to inhibition of secondary granule release. These results suggest that B. abortus has a component which is capable of inhibiting release of myeloperoxidase by dose-dependent preferential inhibition of primary granule release from bovine neutrophilic leukocytes.

A Bacteroides by-product inhibits human polymorphonuclear leukocyte function.

We have previously demonstrated that Bacteroides fragilis enhanced Escherichia coli-induced lethality in the rat fibrin-clot peritonitis model. As a possible mechanism for this phenomenon, it was hypothesized that B fragilis inhibited host defense mechanisms, allowing the E coli to flourish and kill the animal. Culture filtrates of three Bacteroides species were tested in vitro for their effect on human polymorphonuclear leukocyte (PMN) chemotaxis and random migration. Two of these, B fragilis and Bacteroides distasonis, impaired PMN migration. The other, Bacteroides thetaiotaomicron, caused variable inhibition of neutrophil chemotaxis. The ability of the culture filtrates to inhibit neutrophil function appeared to depend on two factors: (1) adequate growth of the Bacteroides culture, permitting production of the leukotoxic factor, and (2) reduction of the culture pH to a level at which the putative toxin could exert its effect. Further studies revealed that the factor was heat stable, had a molecular weight less than 500, and that its effect on PMNs was only partially reversed by multiple washings. This supports the concept that Bacteroides species may contribute to the pathogenicity of mixed infections by producing a factor that inhibits host neutrophil function.

Inhibition of neutrophil phospholipase A2 by p-bromophenylacyl bromide, nordihydroguaiaretic acid, 5,8,11,14-eicosatetraynoic acid and quercetin.

The lipoxygenase and/or cyclooxygenase inhibitors nordihydroguaiaretic acid (NDGA), 4,8,11,14-eicosatetraynoic acid (ETYA) and the bioflavonoid, quercetin, also inhibit phospholipase A2 (phosphatidase 2-acyl hydrolase; EC 3.1.1.4) activity of neutrophil acid extracts and sonicates. The IC50 are 13 microM for NDGA, 22 microM for ETYA, and 100 microM for quercetin when measured on the neutrophil acid extracts; the IC50 obtained with the sonicates are 11 microM, 12 microM and 57 microM, respectively. p-Bromophenylacyl bromide (BPB) inhibits the phospholipase A2 activity of neutrophil acid extracts with an IC50 of 10 microM. In contrast, intact neutrophils incubated for up to 1 h with BPB, washed to remove the drug, and sonicated to expose the phospholipase A2, lose less than 20% of their activity. This strongly suggests that BPB does not inhibit neutrophil function by preventing phospholipase action.

Adenosine in vitro inhibits neutrophil functions by a calcium involving mechanism

Adenosine in vitro inhibits neutrophil functions by a calcium involving mechanism

Interaction of Pseudomonas aeruginosa alkaline protease and elastase with human polymorphonuclear leukocytes in vitro.

Little is known about the interaction of Pseudomonas aeruginosa extracellular products and human polymorphonuclear leukocytes. The present study was designed to examine the effect of alkaline protease and elastase purified from P. aeruginosa on human neutrophil function. Neutrophil chemotaxis, oxygen consumption, glucose oxidation, superoxide production, and nitro blue tetrazolium reduction were studied. It was found that alkaline protease and elastase at fairly low concentrations (0.05 and 0.0025 micrograms/ml, respectively) inhibited chemotaxis. The inhibitory effect of both enzymes was increased at higher concentrations. The chemotaxis of preincubated and washed cells was also inhibited. Alkaline protease but not elastase inhibited opsonized zymosan-stimulated neutrophil oxygen consumption, whereas neither of the enzymes had any effect on glucose oxidation and nitro blue tetrazolium-reducing activity of stimulated neutrophils. The data on superoxide production ability of the cells indicated that the cells preincubated with enzyme and washed were capable of producing superoxide equal to the amount produced by untreated cells when they were stimulated with phorbol myristate acetate or zymosan. However, when elastase was present in the reaction mixture, the reduction of cytochrome c as a measure of superoxide production was inhibited. Inhibition of neutrophil function, particularly chemotaxis, will have important bearing on the escape of the microorganism from the phagocytic defense system of the host. The role of these products in localized infections and avascular areas such as skin burns, cornea, and, at least initially, in chronic lung colonization in cystic fibrosis patients becomes important.

Inhibition of neutrophil function by fluid phase C3b of complement.

A high-molecular-weight fragment of C3 was isolated from normal human serum by column chromatography, was generated by incubation of serum at 37 degrees C with inulin, and was produced from highly purified C3 by limited digestion with trypsin. This product was shown to inhibit the antibacterial function of neutrophils by using Escherichia coli O75 as the main test organism. The inhibitor reacted with anti-C3b and anti-C3c, but not with anti-C3B (anti-native C3) or anti-C3a. The manner of preparation of the inhibitor, the sodium dodecyl sulfate-polyacrylamide gel electrophoresis pattern, and the amino acid composition of the inhibitor indicated that it was fluid phase C3b. The inhibitor of neutrophil function (fluid phase C3b) was shown to bind to C3b receptors or acceptors on sheep erythrocytes in a model system.

Inhibition of neutrophil function by a degradation product of C3

Inhibition of neutrophil function by a degradation product of C3

Inhibition of neutrophil chemotaxis by soluble bacterial products.

BACTERIAL-NEUTROPHIL INTERACTIONS may be critical determinants of virulence in periodontal diseases. This study was undertaken to examine the ability of major bacterial species from the human oral cavity to inhibit (1) peripheral blood neutrophil chemotaxis, (2) chemotactic formylmethionyl peptide binding, and (3) phagocytosis. Included were cultured supernatants and sonic extracts obtained from strains of Capnocytophaga, Bacteroides gingivalis, Fusobacterium nucleatum, Bacteroides asaccharolyticus, Capnocytophaga species, Actinobacillus actinomycetemcomitans, Neisseria, Actinomyces viscosus, Bacterionema matruchotii, Streptococcus mitis, Streptococcus mutans, and Streptococcus sanguis. Chemotaxis was measured using Boyden chambers; phagocytosis was determined using Staphylococcus aureus as the indicator organism and radioactive chemotactic peptide binding was assessed by a rapid filtration assay. None of the test organisms were cytotoxic to neutrophils or inhibited neutrophil phagocytosis. Capnocytophaga species., Bacteroides species., A. actinomycetemcomitans, and F. nucleatum produced factors which specifically inhibited neutrophil chemotaxis. Activity was lost after dialysis. Extracts of Bacteroides species, A. actinomycetemcomitans and F. nucleatum, which were not chemotactic by themselves, inhibited binding of chemotactic peptide suggesting that in vitro chemotaxis inhibition was mediated by nonchemotactic components that compete for the chemotactic factor receptor on the neutrophil. The exception was Capnocytophaga which appeared to inhibit chemotaxis by inhibition of a post-binding event. Such chemotactic inhibitors from periodontopathic organisms that inhibit neutrophil function may be important determinants of virulence.

Stimulation of neutrophil chemotaxis, adhesiveness, phagocytosis, and hexose monophosphate shunt activity by N-(2-mercaptopropionyl)glycine.

The effects of the sulfydryl donor drug N-(2-mercaptopropionyl)-glycine (MPG) on neutrophil chemotaxis, adhesiveness, phagocytosis, and hexose monophosphate shunt activity were investigated in vitro. The drug significantly enhanced all the neutrophil functions tested when used at appropriate concentrations. The results, which are in accord with the well known inhibition of neutrophil function by sulfydryl-blocking agents, suggest the possible therapeutic usefulness of the drug in clinical conditions with defective neutrophil function.

Inhibition of the neutrophil oxidative burst and degranulation by phenothiazines

Trifluoperazine inhibits superoxide production and O 2 uptake by guinea pig neutrophils incubated with arachidonic acid, N-formylmethionylphenylalanine, digitonin or ionophore A23187, with IC 50 values of 7–37uM. Since this inhibition is produced by concentrations of trifluoperazine which inhibit interaction of calmodulin with proteins, we examined the effects of two other phenothiazines which interact less effectively with calmodulin. Chlorpromazine, promethazine and trifluoperazine all inhibit N-formylmethionylphenylalanine-stimulated superoxide production with similar efficiency. Furthermore, degranulation stimulated by A23187 or N-formylmethionylphenylalanine is inhibited similarly by all three phenothiazines with IC 50 values of 18–45 uM. These results are consistent with the suggestion that phenothiazines may inhibit neutrophil function as a result of non-specific interactions with the cells' membranes rather than by specific interaction with calmodulin.