Biomedical subjects

E J Bates

Publications and source records attributed to E J Bates.

At least 37 records · Page 2Linked to original sources

PubMed1991

Characterization of the major neutrophil-stimulating activity present in culture medium conditioned by Staphylococcus aureus-stimulated mononuclear leucocytes.

Culture medium conditioned by stimulating human mononuclear leucocytes (MNL) with killed Staphylococcus aureus (Scm) was found to contain a substantial amount of tumour necrosis factor-alpha (TNF-alpha) but no detectable tumour necrosis factor-beta (TNF-beta). Culture medium conditioned by MNL in the absence of bacteria contained no TNF-alpha activity. When Scm was fractionated by high-performance liquid chromatography (HPLC) on Bio-Sil TSK 250, TNF-alpha co-eluted with neutrophil-stimulating activity measured by chemiluminescence. Similarly, the ability of neutrophils to kill opsonized S. aureus was enhanced in fractions that contained this neutrophil-stimulating activity. The stimulating activity could be almost completely removed by pretreatment of the Scm with a TNF-alpha-specific monoclonal antibody (mAb). The ability of neutrophils to kill S. aureus in response to Scm was also substantially reduced by mAb to TNF-alpha. These results demonstrate that bacterial interaction with MNL leads to the release of neutrophil-stimulating activity that consists predominantly of TNF-alpha.

Antigens, Bacterial↗

PubMed1990

Neutrophil-mediated cartilage injury in vitro is enhanced by tumour necrosis factor alpha.

Neutrophil functions relevant to tissue damage are altered by cytokines such as tumour necrosis factor alpha (cachectin, TNF alpha), known to be present in inflammatory foci. In this study we examined the effect of TNF alpha on neutrophil-mediated cartilage damage in vitro. Human neutrophils were able to injure both human and bovine articular cartilage slices by degrading proteoglycan and inhibiting its synthesis. Recombinant human TNF alpha enhanced neutrophil-mediated degradation of proteoglycan, even when neutrophils were preincubated with TNF alpha and washed before incubating with cartilage. TNF alpha alone degraded proteoglycan and inhibited its synthesis. Neutrophil-mediated inhibition of proteoglycan biosynthesis was increased after incubating cartilage together with neutrophils and TNF alpha, but was unaltered when neutrophils were preincubated with TNF alpha. We conclude that TNF alpha enhances neutrophil injury to articular cartilage.

Adolescent↗

PubMed1990

Degradation of human cartilage by monocytes.

The effect of human peripheral blood monocytes on the degradation of human articular cartilage was studied in vitro using a radiometric assay to detect proteoglycan breakdown. The results showed that proteoglycan breakdown was increased by 60% after a 20 h exposure to monocytes (p less than 0.001).

Cartilage, Articular↗

PubMed1990

Augmentation of the human monocyte/macrophage chemiluminescence response during short-term exposure to interferon-gamma and tumour necrosis factor-alpha.

The effects of short-term (30 min) pre-incubation of human monocytes and macrophages (3-day cultured monocytes) with leucocyte-derived human interferon-gamma (IFN-gamma) and recombinant human tumour necrosis factor-alpha (rTNF-alpha) were examined. Pre-incubation of either monocytes or macrophages with rTNF-alpha or IFN-gamma (100 U/5 x 10(5) cells) augmented their respiratory burst to formyl-L-methionyl-L-leucyl-L-phenylalanine (fMLP), measured by the luminol- and lucigenin-dependent chemiluminescence assay. In addition, both cell types showed a burst of respiratory activity in the presence of rTNF-alpha or IFN-gamma only. The effects of IFN-gamma were removed by adsorption with an anti-IFN-gamma monoclonal antibody and those of rTNF-alpha were abolished by heating at 100 degrees C, or by the addition of anti-TNF-alpha monoclonal antibody. The results demonstrate that both IFN-gamma and rTNF-alpha are stimulators of monocytes and macrophages, and rapidly alter the capacity of the cells to respond to fMLP, which binds to cell surface receptors.

Cells, Cultured↗

PubMed1989

Mechanisms of human neutrophil-mediated cartilage damage in vitro: the role of lysosomal enzymes, hydrogen peroxide and hypochlorous acid.

Cartilage is a focal point of attack by cellular and molecular elements of the inflammatory response which occurs in arthritic diseases. Neutrophils damage articular cartilage by degrading matrix components and inhibiting their synthesis. The aim of this study was to elucidate mechanisms of this damage. Human neutrophils were isolated from blood by centrifuging through Ficoll-Hypaque and granule extract prepared from them. Articular cartilage from adult humans and cattle was maintained in organ culture. Cartilage degradation (release of 35S-labelled proteoglycan) or synthesis (incorporation of 35S into proteoglycan) was determined after various treatments. Human neutrophils and neutrophil granule extract degraded proteoglycan and inhibited proteoglycan synthesis. The specific leucocyte elastase inhibitor N-methoxysuccinyl-(ala)2-pro-val-chloromethylketone (MAAPVCMK) partially reversed these effects. H2O2, a product of the neutrophil respiratory burst, when added directly at 10(-6)mol/L, or generated by glucose oxidase (GO)/glucose inhibited proteoglycan synthesis but had no effect on degradation. Hypochlorous acid (OHCl), a product of the myeloperoxidase (MPO)/H2O2/Cl system at 50 mumol/L degraded proteoglycan and inhibited its synthesis. OHCl produced by granule extract (as a source of MPO) + GO-generated H2O2 + Cl- degraded proteoglycan. The results indicate that neutrophil-mediated proteoglycan degradation and inhibition of synthesis is largely attributable to elastase and secondarily to OHCl, whereas H2O2 impairs synthesis without affecting degradation of proteoglycan.

Animals↗

PubMed1989

Staphylococcus aureus-stimulated mononuclear leucocyte-conditioned medium increases the neutrophil bactericidal activity, and augments oxygen radical production and degranulation in response to the bacteria.

Conditioned medium produced by human mononuclear leucocytes (MNL) stimulated with formalin-fixed, heat-killed Staphylococcus aureus enhanced the killing of opsonized S. aureus by human neutrophils. This enhancement was not seen when conditioned medium from non-stimulated MNL or medium cultured in the absence of both bacteria and MNL was used. There was a five-fold decrease in survival of bacteria when neutrophils were treated with conditioned medium. Conditioned medium-treated neutrophils showed increased initial rate of killing but after 20-30 min the rate of killing was similar to that of non-conditioned medium-treated leucocytes. The neutrophils treated with conditioned medium showed increased production of chemiluminescence, superoxide and lysosomal enzyme release (from both azurophilic and specific granules) in response to opsonized S. aureus. The results demonstrate that bacterial interaction with MNL leads to the release of mediators (cytokines) which potentiate the anti-bacterial function of neutrophils.

Blood Bactericidal Activity↗

PubMed1988

Neutrophil stimulating activity released by Staphylococcus-stimulated mononuclear leukocyte conditioned medium. Further characterization and partial purification.

Culture medium conditioned by human mononuclear leukocytes (MNL) stimulated with formalin fixed heat-killed Staphylococcus aureus induces a small respiratory burst in human neutrophils, and dramatically increases the response of neutrophils to stimuli such as N-formyl-L-methionyl-L-leucyl-L-phenylalanine. The data presented show that the activity is not unique to Staphylococcus aureus. Similar neutrophil modulating activities were produced by medium conditioned by MNL cultured in the presence of Streptococcus pneumonia, and Group B streptococcus. The activity was relatively resistant to heating; significant reduction of activity was observed only when 80 degrees C was reached. Neutrophil stimulating activity production by stimulated MNL was dependent on protein and RNA synthesis and the activity appeared to be released by the non-adherent fraction of the MNL, suggesting that it is not of macrophage origin. The activity was not sensitive to soya bean trypsin inhibitor, but was sensitive to trypsin and was not removed when stimulated conditioned medium was depleted of immunoglobulin and albumin by affinity chromatography. Purification by gel filtration on Sephadex G-100 and high-performance liquid chromatography with Bio-Sil TSK250 columns showed that the major activity had an apparent molecular weight of 35,000-43,000 under conditions in which ionic interactions and association with albumin were reduced; by using polyethylene glycol or high salt (0.46 M Na+) in the elution buffer.

Cell Adhesion↗

PubMed1988

Tumour necrosis factor-beta modulates human neutrophil-mediated cartilage damage.

Human neutrophils, when cultured with human articular cartilage coated with heat-aggregated immunoglobulin G, degraded proteoglycan and inhibited its synthesis. Neutrophil-mediated degradation of cartilage was potentiated by recombinant human tumour necrosis factor-beta (TNF beta), although TNF beta alone did not alter proteoglycan degradation. This effect was seen when TNF beta, neutrophils, and cartilage were incubated together, and also when neutrophils were preincubated with TNF beta and washed before being added to cartilage. Similar results were obtained with living and killed cartilage. In contrast, pretreatment of neutrophils with TNF beta abrogated the neutrophil-mediated inhibition of proteoglycan biosynthesis. There was no effect of TNF beta alone on synthesis of proteoglycan.

Cartilage↗

PubMed1988

Elastase in the pathogenic free-living amoebae Naegleria and Acanthamoeba spp.

The data showed that pathogenic free-living amoebae contain the proteolytic enzyme elastase. The levels of enzyme were similar in Naegleria fowleri, N. australianis italica, and Acanthamoeba culbertsoni A-1. No difference was found between elastase levels in a highly pathogenic N. fowleri and those in the same organism which had lost pathogenicity as a result of long-term axenic maintenance.

Acanthamoeba↗

PubMed1988

Conditioned medium from stimulated mononuclear leucocytes potentiates the ability of human neutrophils to damage human articular cartilage.

Human neutrophils were able to degrade proteoglycan and inhibit its synthesis when incubated with human articular cartilage coated with heat aggregated immunoglobulin G. These effects were potentiated when culture medium conditioned by mononuclear leucocytes stimulated with killed Staphylococcus aureus was also present during the incubations. Neutrophils preincubated with this conditioned medium and washed before incubation with cartilage also showed an increased ability to degrade proteoglycan and inhibit its synthesis. The percentage of neutrophils binding to cartilage was significantly increased in the presence of this conditioned medium.

Adolescent↗

PubMed1988

Stimulation of human neutrophil degranulation by mefloquine.

The effect of the antimalarial drug mefloquine on human neutrophil degranulation, chemiluminescence, superoxide production and viability was examined in vitro. Mefloquine was found to significantly stimulate the release of lysozyme, beta-glucuronidase and myeloperoxide at a concentration of 10 micrograms/ml (2.5 X 10(-5) M) without loss of cell viability. At 40 micrograms/ml mefloquine (1 X 10(-4) M) cell viability was significantly decreased. Mefloquine at 10 micrograms/ml also significantly increased the release of lysozyme and beta-glucuronidase but not myeloperoxidase when neutrophils were stimulated with opsonized zymosan. At a lower zymosan concentration myeloperoxidase release was also increased. Enzyme activity was not directly stimulated by mefloquine. Mefloquine at 10 micrograms/ml significantly increased luminol-dependent chemiluminescence but significantly inhibited lucigenin-dependent chemiluminescence when neutrophils were stimulated with opsonized zymosan. Under these conditions superoxide release, measured by cytochrome c reduction, was inhibited to a lesser degree. These results are discussed with reference to our previous report that mefloquine inhibits the neutrophil iodination reaction [Immunology 58: 125-130, 1986] and the use of mefloquine as an anti-inflammatory drug.

Cell Survival↗

PubMed1988

Tumour necrosis factor beta (lymphotoxin) inhibits locomotion and stimulates the respiratory burst and degranulation of neutrophils.

The data presented here demonstrate that recombinant human tumour necrosis factor beta (rHuTNF beta; lymphotoxin) is a neutrophil modulator. The lymphokine inhibited the locomotion of neutrophils and augmented the neutrophil oxygen-dependent respiratory burst in response to N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP) and phorbol myristate acetate (PMA), as measured by their capacity to produce chemiluminescence, H2O2 and superoxide. The effects on the respiratory burst occurred at a tenth of the concentration of TNF beta required to inhibit locomotion. After incubation with TNF beta, the neutrophils could be washed without any reduction in their capacity to show augmented responses. The TNF beta enhanced granule enzyme (lysozyme and beta-glucuronidase) release of neutrophils stimulated with cytochalasin B-FMLP.

Cell Movement↗

PubMed1987

Staphylococcus aureus-stimulated human mononuclear leucocyte-conditioned medium augments the basal and stimuli-induced neutrophil respiratory burst and degranulation.

Culture medium conditioned by mononuclear leucocytes (MNL) stimulated by formalin-fixed heat-killed Staphylococcus aureus (sCM) modulated a number of neutrophil functions. The sCM inhibited the locomotion of human neutrophils in both the presence and absence of a chemotactic gradient generated with N-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP). It also stimulated the oxygen-dependent respiratory burst as assessed by its ability to stimulate basal H2O2, superoxide and chemiluminescence production by neutrophils. Neutrophils treated with sCM also showed increased release of lysozyme but not beta-glucuronidase. In addition, the sCM-treated neutrophils showed a potentiated response to stimuli that bind surface receptors, i.e. FMLP and opsonized zymosan. The effects of sCM and either of the stimuli were synergistic. Examination of lysosomal enzyme release showed that sCM enhanced the release of lysozyme and beta-glucuronidase induced by either FMLP/cytochalasin B or zymosan. The response to phorbol myristate acetate (PMA), which bypasses the surface receptor, was also stimulated but compared poorly with the FMLP response. The sCM effects on the FMLP-induced chemiluminescence response occurred even when FMLP addition was delayed for 4 hr. Cells treated with sCM and washed retained the ability to show an enhanced FMLP response. The neutrophil-modulating activity was not produced by MNL cultured in the absence of bacteria.

Antigens, Bacterial↗

PubMed1986

Inhibition of human monocyte respiratory burst, degranulation, phospholipid methylation and bactericidal activity by pneumolysin.

The interaction between the pneumococcal toxin pneumolysin and human monocytes was examined. At non-cytotoxic concentrations (0.5-2.5 HU/10(6) cells) pneumolysin depressed the oxygen-dependent respiratory burst in monocytes, induced by opsonized zymosan or phorbol myristate acetate (PMA). This included depressed hexose-monophosphate shunt activity and hydrogen peroxide production. The toxin also depressed the ability of monocytes to degranulate (measured by release of lysozyme) in response to the above stimuli. Phospholipid transmethylation was also markedly decreased by pretreating monocytes with pneumolysin. These effects on monocyte functions were accompanied by a decreased ability of pneumolysin-treated monocytes to kill Streptococcus pneumoniae, the organism that produces the toxin. Cholesterol, which inhibits the haemolytic activity of the toxin, was shown to abrogate the effects of pneumolysin on monocytes.

Bacterial Proteins↗

PubMed1985

Hyaluronic acid synthesis in articular cartilage: an inhibition by hydrogen peroxide.

The synthesis of hyaluronic acid by bovine articular cartilage in culture was inhibited after treatment with xanthine oxidase and hypoxanthine. Through the use of catalase, superoxide dismutase and the specific iron chelator diethylenetriaminepentaacetic acid, the active species responsible for inhibition was shown to be hydrogen peroxide. Hydrogen peroxide generated by glucose oxidase was also inhibitory. Some recovery of hyaluronic synthesis was evident after a further period of culturing. Proteoglycan synthesis was inhibited in parallel with hyaluronic acid synthesis.

Animals↗

PubMed1985

Inhibition of proteoglycan synthesis by hydrogen peroxide in cultured bovine articular cartilage.

Oxygen-derived reactive species, generated enzymatically by the action of xanthine oxidase upon hypoxanthine, significantly inhibit proteoglycan synthesis by cultured bovine articular cartilage (Bates, E.J., Lowther, D.A. and Handley, C.J. (1984) Ann. Rheum. Dis. 43, 462-469). Here we extend these investigations and show, through the use of catalase and the specific iron chelator diethylenetriaminepentaacetic acid, that the active species involved is H2O2 and not the hydroxyl radical. Incubations of cartilage with H2O2 at concentrations of 1 X 10(-4) M and above are also inhibitory to proteoglycan synthesis. Subsequent recovery of the tissue is dependent upon the initial dose of xanthine oxidase or H2O2. Xanthine oxidase at 84 mU per incubation results in a prolonged inhibition of proteoglycan synthesis which is still apparent after 14 days in culture. Lower concentrations of xanthine oxidase (21-66 mU) are inhibitory to proteoglycan synthesis, but the tissue is able to synthesise proteoglycans at near normal rates after 3 days in culture. The inhibition of proteoglycan synthesis by 1 X 10(-4) M H2O2 is completely reversed after 5 days in culture, whereas 1 X 10(-3) M H2O2 results in a more prolonged inhibition. The synthesis of the proteoglycan core protein is inhibited, but the ability of the newly formed proteoglycans to aggregate with hyaluronic acid is unimpaired.

Animals↗

PubMed1984

Oxygen free-radicals mediate an inhibition of proteoglycan synthesis in cultured articular cartilage.

Superoxide radical, generated enzymatically by the action of xanthine oxidase on hypoxanthine, significantly inhibited proteoglycan synthesis by cultured bovine articular cartilage. This inhibition was not due to the generation of uric acid or to the generation of superoxide per se. It was immediate in onset and still evident after six days in culture. The inhibition was similar for both 35S-sulphate and 3H-acetate incorporation into glycosaminoglycans and could not be reversed by addition of beta-D-benzyl xyloside. Protein synthesis was also inhibited.

Animals↗

PubMed1984

Effect of oxygen-derived reactive species on cartilage proteoglycan-hyaluronate aggregates.

Proteoglycan-hyaluronate aggregates were incubated with oxygen-derived reactive species generated enzymatically by the action of xanthine oxidase upon hypoxanthine. Analysis of the products of the incubation by caesium sulphate zonal sedimentation revealed that degradation of aggregate had occurred. This effect was reversed by inclusion of superoxide dismutase, catalase or diethylenetriaminepentaacetic acid in the incubations suggesting that hydroxyl radicals were the active species. Separate analysis by gel filtration chromatography on Sepharose CL-2B of proteoglycan monomer subjected to a similar treatment indicated that the molecule is minimally degraded. These results are discussed with reference to the well established degradation of hyaluronate by oxygen-derived reactive species.

Animals↗