Sodium butyrate, which directly affects chromatin structure and function in many cells, is as effective as granulocyte-macrophage colony-stimulating factor (GM-CSF) in delaying apoptosis in human neutrophils. Both butyrate and GM-CSF preserved the ability of neutrophils cultured for 22 h in vitro to generate reactive oxidants and express receptors such as CD16. They also delayed apoptotic morphology and DNA fragmentation, and stimulated de novo biosynthesis: newly-labelled polypeptides detected by 2D-polyacrylamide gel electrophoresis after treatment with GM-CSF and butyrate were very similar. Cycloheximide abrogated the effects of both GM-CSF and butyrate. Exposure to butyrate for 1 h did not prime oxidant production and did not up-regulate expression of CD11b. Hence, unlike GM-CSF, butyrate does not stimulate translocation of granules to the plasma membrane. These data suggest that active gene expression is involved in the regulation of neutrophil apoptosis and changes in chromatin structure and function may control apopotosis in these cells.

Apoptosis↗

PubMed1996

Neutrophil apoptosis is delayed by the diadenosine polyphosphates, Ap5A and Ap6A: synergism with granulocyte-macrophage colony-stimulating factor.

In addition to ATP, platelets and other cell types can secrete high quantities of diadenosine polyphosphates Ap3A, Ap4A, Ap5A and Ap6A. There is increasing evidence to show that these molecules can function as novel modulators of cell function. For this report we have measured the effects of the diadenosine polyphosphates Ap5A and Ap6A on neutrophil apoptosis. These molecules can themselves delay neutrophil apoptosis (as assessed by morphology, function. CD16 expression and chromatin integrity), and are as effective on a molar basis as ATP, Ap3A and Ap4A. Moreover, these dinucleotides act synergistically with granulocyte-macrophage colony-stimulating factor (GM-CSF) to delay neutrophil apoptosis. Thus, diadenosine polyphosphates may act, in concert with cytokines, as novel modulators of neutrophil function and survival in certain types of inflammatory conditions.

Apoptosis↗

PubMed1995

Potentiation of the respiratory burst of human neutrophils by cycloheximide: regulation of reactive oxidant production by a protein(s) with rapid turnover?

Incubation of human neutrophils with the protein biosynthesis inhibitor cycloheximide for > or = 90 min results in a decreased ability to generate reactive oxidants during the respiratory burst. This implies that active protein biosynthesis is required to sustain the ability of these cells to generate reactive oxidants. However, short term incubation of neutrophils (40-60 min) with either cycloheximide or puromycin results in a significant increase in oxidase activity stimulated by either fMet-Leu-Phe (> 60%) or by leukotriene B4 (> 30%). However, after incubation for 40-60 min with these inhibitors of protein biosynthesis, the respiratory burst stimulated by PMA was unaffected whilst that stimulated by the particulate stimuli opsonised zymosan or latex beads was significantly inhibited. The enhanced oxidase activity stimulated by the soluble agonists was not explained by changes in receptor expression, alterations in intracellular Ca2+ levels or by enhanced degranulation. These results suggest that oxidase activity stimulated by soluble agonists in neutrophils is normally regulated by a short-lived, actively-synthesised protein(s).

Amino Acid Sequence↗

PubMed1995

The cell biology of phagocytes.

Explore the source record for details and available documents.

Chemotaxis, Leukocyte↗

PubMed1995

Cell signalling by integrins and immunoglobulin receptors in primed neutrophils.

Neutrophils use a variety of cell-surface receptors for attachment to surfaces, such as the endothelium or opsonized pathogens. During inflammation or infection, chemoattractants or cytokines bind to neutrophils and upregulate or 'prime' their responsiveness. Priming can increase the number of some receptors expressed on the cell surface but can also change receptor function, so that the receptor has altered ligand-binding properties or becomes linked to new intracellular signalling systems.

Integrins↗

PubMed1995

Interleukin-1 expression by neutrophils in rheumatoid arthritis.

OBJECTIVE: To determine if neutrophils from blood and synovial fluid of patients with rheumatoid arthritis and other joint arthropathies express interleukin-1 beta mRNA. METHODS: RNA was isolated from neutrophils from patient and control blood, and synovial fluid of patients, probed in northern blots, and quantified by densitometry. It was also isolated and analysed from control blood neutrophils after incubation in vitro with granulocyte macrophage colony stimulating factor (GM-CSF). RESULTS: Neutrophils from the synovial fluid of patients with rheumatoid arthritis contained low levels of mRNA for interleukin-1 beta--between 0.1 and 2% of those observed during stimulation of control neutrophils with GM-CSF for one hour. Higher levels (4-40% of the maximal GM-CSF values) were observed in blood neutrophils from patients with rheumatoid arthritis. CONCLUSIONS: Neutrophils contribute to the cytokine network in rheumatoid arthritis. In some circumstances, activation of transcription may occur within the circulation of these patients.

Arthritis, Rheumatoid↗

PubMed1995

Neutrophils from preterm neonates and adults show similar cell surface receptor expression: analysis using a whole blood assay.

Previous work has shown that Fc gamma RIII expression in isolated neonate neutrophils is defective. We have re-examined this phenomenon in view of the facts that (1) the receptor is present on mobilisable subcellular stores and (2) commonly used isolation procedures can affect receptor expression in suspensions of isolated neutrophils. Receptor expression was measured by fluorescence-activated cell sorter analysis of neutrophils in unfractionated whole blood. Examination of receptor expression in preterm, term and adult neutrophils indicated small but significantly decreased expression of CR1 and CR3 in preterm neutrophils compared with term neutrophils (p < 0.01). A small decrease in expression was found for Fc gamma RI and Fc gamma RIII (p < 0.05). No significant difference in expression of Fc gamma RII was observed in all groups analysed. These data suggest that isolated preterm neonate neutrophils have greatly decreased expression of Fc gamma RIII because of impaired composition or mobilisation of the subcellular stores of this receptor and/or increased lability of the surface receptor which leads to its shedding during purification.

Adult↗

PubMed1995

Impaired neutrophil phagocytosis in preterm neonates: lack of correlation with expression of immunoglobulin or complement receptors.

Preterm neonates are vulnerable to infection as a result of a compromised immune system. The function of neutrophils from 'well', 'stressed', and 'maturing' preterm neonates was compared with term neonate and adult neutrophils using a whole-blood phagocytosis assay. Cell surface expression of complement receptors and immunoglobulin G receptors was measured on neutrophils in whole blood from the same samples. Fewer actively phagocytosing neutrophils were found in all preterm neonate samples, especially in maturing neonates. Phagocytic rates were slower, and the number of Escherichia coli ingested was smaller in preterm neonate than in term neonate neutrophils. Expression of immunoglobulin G receptors and complement receptor 3 on neutrophils was not directly related to phagocytic activity.

Adult↗

PubMed1995

Edwards inventory of emotions: assessing emotions in athletes and nonathletes.

A self-report inventory was created on which respondents indicated the frequency of occurrence of 40 basic emotions using a 5-point rating scale. The inventory was administered to two matched, independent college-age samples (ns = 562 and 414) and the factorial validity was tested. Factor 1 was a general factor reflecting over-all Positive Affect. Factor 2 was a more specific factor reflecting Profound Negative Affect. Factor 3 was also a specific factor reflecting Moderate Negative Affect. Subsequent analyses gave significantly greater scores for athletes over nonathletes and men over women on the Profound Negative Affect subscale. Women had significantly higher Positive Affect scores. It was concluded that the questionnaire had sufficient technical merit for use in research.

Adult↗

PubMed1994

Priming of the respiratory burst of human neutrophils by the diadenosine polyphosphates, AP4A and AP3A: role of intracellular calcium.

The diadenosine polyphosphates, Ap3A and Ap4A, prime the respiratory burst of human neutrophils after stimulation with fMet-Leu-Phe. Maximal priming of oxidase activity occurred at 600-800 microM Ap3A and Ap4A, compared with maximal priming observed at 200 microM ATP. The time course of priming of the oxidase by all 3 nucleotides was very rapid, being detectable if added within 10 s of fMet-Leu-Phe. All 3 nucleotides also elicited increases in intracellular Ca2+ levels and there was a close concentration-dependency between the extent of priming and the increase in intracellular Ca2+. However, at low concentrations of nucleotides (< 50 microM Ap3A and Ap4A and < 0.1 microM ATP) priming of the oxidase was observed without detectable increases in intracellular Ca2+. These observations indicate that diadenosine polyphosphates may be novel regulators of neutrophil function and that priming of oxidase activity may occur via mechanisms that are either dependent or independent of increases in intracellular Ca2+.

Adenosine Triphosphate↗

PubMed1994

Phospholipase D-dependent and -independent activation of the neutrophil NADPH oxidase.

Stimulation of the respiratory burst of human neutrophils by fMet-Leu-Phe (in the absence of cytochalasin B) is largely unaffected when the activities of protein kinase C and phospholipase D are inhibited. This has been confirmed using three separate assays to measure the respiratory burst. However, whilst these enzymes are not required for the initiation or maximal rate of oxidant generation, they are required to sustain oxidase activity. In contrast, in the presence of cytochalasin B, fMet-Leu-Phe stimulated oxidase activity is much more dependent on phospholipase D activity. It is proposed that (in the absence of cytochalasin B) activation of the NADPH oxidase utilises cytochrome b molecules that are already present on the plasma membrane and activation occurs independently of phospholipase D and protein kinase C. Once these complexes are inactivated, then new cytochrome b molecules must be recruited from sub-cellular stores. This translocation and/or activation of these molecules is phospholipase D dependent. Some support for this model comes from the finding that the translocation of CD11b (which co-localises with cytochrome b) onto the cell surface is phospholipase D dependent.

1-Butanol↗

PubMed1994

Identification of a subgroup of myelodysplastic patients with a neutrophil stimulation-signalling defect.

f-Met-Leu-Phe-stimulated luminol-enhanced chemiluminescence was found to be repeatedly defective in some MDS patients. This defect was not attributed to myeloperoxidase deficiency, nor to a defect in NADPH oxidase function, because PMA chemiluminescence was found to be normal in these individuals. An arbitrary value of 7 mV (half the mean control value) was chosen to subdivide the group: MDS patients with values < 7 mV had a mean f-Met-Leu-Phe chemiluminescence response of 2.5 +/- 0.5 compared to MDS patients with values > 7 mV who had a mean response of 15.6 +/- 1.6 mV, P < 0.01 (healthy controls 14 +/- 2 mV). The characteristics of the f-Met-Leu-Phe receptor and initial calcium flux results suggested that the receptor itself was normal in number and function in low f-Met-Leu-Phe responders. The rate of superoxide generation, which is calcium-dependent, was also found to be in the normal range in low f-Met-Leu-Phe responders, although total superoxide production was reduced in some of these patients. When MDS neutrophils with a low f-Met-Leu-Phe response were stimulated with PMA, chemiluminescence was normal, suggesting normal activity of the NADPH-oxidase complex. Furthermore, myeloperoxidase activity was reduced in only three out of the 11 low f-Met-Leu-Phe responders. Following priming with GM-CSF, f-Met-Leu-Phe chemiluminescence was 27 +/- 1.6 mV in low f-Met-Leu-Phe responders compared to controls (87.7 +/- 11 mV, P < 0.005). Thus, although responses were improved, they were not as marked as in control neutrophils. These data suggest that a subgroup of MDS patients have a low f-Met-Leu-Phe chemiluminescence response which is not due to a defect in the f-Met-Leu-Phe receptor or oxidase activity, and in the majority of cases MPO activity is normal. Initial patient survival data suggest that these patients may have an increased risk of infective mortality. It is proposed that defective f-Met-Leu-Phe chemiluminescence results from a putative defect in cell-signalling mechanism upstream of PKC, and GM-CSF priming only partially improves responsiveness.

Blood Bactericidal Activity↗

PubMed1994

Cytokine expression by inflammatory neutrophils.

Bloodstream neutrophils do not express mRNA for interleukin-1 beta (IL-1 beta), but transcripts for this cytokine are rapidly induced following exposure to recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) in vitro. Levels of IL-1 beta mRNA reach maximal values 1 h after exposure to rGM-CSF and then decline to near basal levels by 4 h. Similarly, rGM-CSF treatment of blood neutrophils in vitro induced increases in levels of mRNA for IL-6 and tumour necrosis factor-alpha (TNF-alpha). RNA extracted from neutrophils isolated from the synovial fluid of patients with rheumatoid arthritis expressed low, but significant levels of IL-1 beta mRNA that were between 0.5 and 3% of the levels that could be maximally induced by rGM-CSF treatment of blood neutrophils. However, transcripts for TNF-alpha and IL-6 were not detected in these synovial fluid neutrophils. mRNA for transforming growth factor-beta (TGF-beta) was constitutively expressed in blood and synovial fluid neutrophils and transcripts for this cytokine were not altered by rGM-CSF exposure. Because of the transient nature of IL-1 beta expression by activated neutrophils, we propose that the low levels of expression of mRNA for this cytokine in the synovial fluid neutrophils represents expression by a small, perhaps newly-recruited and activated, sub-population of cells. IL-1 beta expression by this sub-population may thus contribute to the pathogenesis of rheumatoid disease.

Arthritis, Rheumatoid↗

PubMed1994

Stimulation of reactive oxidant production in neutrophils by soluble and insoluble immune complexes occurs via different receptors/signal transduction systems.

Cell-free synovial fluid from patients with rheumatoid arthritis contains soluble and insoluble IgG-containing immune complexes which activate reactive oxidant production in human neutrophils. In this report we have measured the effects of inhibitors of signal transduction pathways on neutrophil activation by these complexes and also following activation by synthetic soluble and insoluble immune complexes made from human serum albumin (HSA) and anti-(HSA) antibodies. In all aspects studied, the soluble rheumatoid complexes and the soluble synthetic complexes were indistinguishable in the ways in which they activated neutrophils. Activation of reactive oxidant production in response to these soluble complexes was completely inhibited by pertussis toxin (indicating G-protein coupling of receptor occupancy), completely insensitive to staurosporine (indicating that oxidant production did not require protein kinase C activity), only marginally (< 30%) inhibited by butanol (indicating that dependence upon activity of phospholipase D was minimal), and completely inhibited by chloracysine, an inhibitor of phospholipase A2. In contrast, activation of reactive oxidant production in response to the insoluble rheumatoid or insoluble synthetic immune complexes was largely pertussis toxin insensitive, inhibited by > 50% by staurosporine, inhibited by > 50% by butanol, and completely inhibited by chloracysine. These results show that the receptor-mediated signal transduction systems activated by the soluble and insoluble immune complexes are different. Because the soluble complexes activate a transient burst of reactive oxidant secretion from primed neutrophils, the mechanisms regulating either the release or the intracellular production of oxidants within rheumatoid joints are distinct and hence may be pharmacologically modified independently of each other.

Alkaloids↗