Microbial pathogenicity and host defense in burned patients--the role of inflammatory mediators.
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B-Lymphocytes↗
Biomedical subjects
W König
Publications and source records attributed to W König.
Effects of cefaclor, cefetamet and Ro 40-6890 on inflammatory responses of human granulocytes.
The effect of three cephalosporins (cefetamet, cefaclor and Ro 40-6890) upon human granulocytes and their ability to modulate the chemiluminescence response, phagocytose, kill bacteria and generate leukotrienes was studied. In the presence of the cephalosporins there was a significant increase in phagocytosis of Escherichia coli. The bactericidal activity of human granulocytes for several other bacteria was also enhanced. Cefetamet and cefaclor increased the chemiluminescence response of human neutrophils to Pseudomonas aeruginosa and Proteus mirabilis in contrast to Ro 40-68790, which decreased the chemiluminescence response. The cephalosporins decreased the synthesis of leukotrienes from human neutrophils after stimulation with Escherichia coli and Staphylococcus aureus. These data emphasize the immunomodulatory functions of various cephalosporins on cells involved in host defence.
Cefaclor↗
Leukotriene generation in patients with multiple injuries.
We determined the generation and metabolism of lipoxygenase products in isolated granulocyte fractions of patients with multiple trauma (n = 9) and compared the results with those of healthy volunteers (n = 8). The supernatants of stimulated cells were analyzed by high-performance liquid chromatography. During the first week after injury a significantly reduced capacity to generate LTB4 and an increased metabolism of LTB4 into omega-oxidated products (20-OH-LTB4 and 20-COOH-LTB4) were observed after stimulation of the granulocytes with Ca ionophore. The depressed leukotriene production could be partly abrogated by the addition of arachidonic acid. These findings are comparable with alterations previously described in severely burned patients with postburn sepsis. Additionally, an elevated production of LTC4 by peripheral granulocyte fractions was observed in two patients suffering from adult respiratory distress syndrome (ARDS) as well as an increased number of eosinophils during the time of lung dysfunction. Analysis of bronchoalveolar lavage in patients with multiple trauma (11 patients with ARDS and 11 patients without ARDS) by a specific radio-immunoassay confirmed an elevated production of cysteinyl-leukotrienes.(ABSTRACT TRUNCATED AT 250 WORDS)
Adolescent↗
Induction of inflammatory mediator release (serotonin and 12-hydroxyeicosatetraenoic acid) from human platelets by Pseudomonas aeruginosa glycolipid.
Purified glycolipid from Pseudomonas aeruginosa induced the generation of significant amounts of 12-hydroxyeicosatetraenoic acid (12-HETE) and serotonin release from human platelets. The release of serotonin was first observed 2 min after addition of the glycolipid and increased with time. Significant serotonin release was obtained at glycolipid concentrations above 5 micrograms/ml and increased dose-dependently up to 100% at glycolipid concentrations above 40 micrograms/ml. Glycolipid induced 12-HETE in a time- and dose-dependent manner. 12-HETE formation was first measured after 10 min of incubation and increased with time. Optimal 12-HETE formation was obtained at a glycolipid concentration of 50 micrograms/ml; higher concentrations of glycolipid led to a decrease in 12-HETE formation, indicating a cytotoxic effect. Stimulation of platelets with glycolipid (12-HETE formation and serotonin release) was accompanied by calcium influx, translocation of protein kinase C, activation of guanylylimidodiphosphate binding, and increased GTPase activity in platelet membranes within the same concentration range.
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
Effect of antigens from Nippostrongylus brasiliensis and cytokines on the ongoing IgE synthesis in vitro.
The modulation of an ongoing IgE-mediated immune response induced by the infection of mice with the nematode Nippostrongylus brasiliensis (N.b.) was investigated in vitro. In previous experiments antigens derived from the homogenate of adult worms (WH) and third stage larvae (LH) were characterized by immunoblotting. Our results demonstrate that only antigens of the WH were recognized by IgE antibodies. The effect of worm antigens and cytokines (IL-2, IL-4, IL-5, IFN gamma) on the IgE and IgG synthesis in different culture systems was studied. The IgG synthesis of B cells was stimulated by WH or WH/cytokines. The IgE production of B cells was enhanced only by WH/cytokines or when T cells were present. Individual antigen fractions (WH 5, 8, 9, 10, 13) increased the IgE production, which was enhanced in the presence of IL-4 2- to 3.5-fold) but had no significant effects on the IgG production.
Animals↗
Differential increase in 12-HETE release and CD29/CD49f expression of platelets from normal donors and from patients with atopic dermatitis by Staphylococcus aureus.
The generation of the arachidonic acid-derived inflammatory mediator 12-hydroeicosatetraenoic acid (HETE) and the expression of CD29 as well as CD49f from unstimulated and stimulated platelets has been studied in patients with atopic dermatitis (AD) as well as in healthy volunteers. Heat-killed clinical isolates of Staphylococcus aureus served as stimuli. Unstimulated platelets from patients with AD produced higher amounts of 12-HETE compared to platelets from normal donors. The absolute 12-HETE release from platelets of patients with AD was significantly higher compared to the control group after stimulation with heat-killed S. aureus, whereas the relative increase remained. The expression of CD29 and CD49f on unstimulated platelets of patients with AD was markedly enhanced compared to platelets from normal donors. Stimulation with S. aureus led to similar results as to the CD29 expression on normal and atopic platelets or to a markedly higher expression of CD49f on platelets from normal donors. As compared to platelets from normal donors the CD49f expression on atopic platelets was slightly enhanced by S. aureus. Our data emphasize that platelets may play an important role in the pathogenesis of AD by an increased preactivation and by an enhanced responsiveness to S. aureus which colonizes permanently the skin of patients with AD.
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
Alginate--its role in neutrophil responses and signal transduction towards mucoid Pseudomonas aeruginosa bacteria.
Mucoid Pseudomonas aeruginosa bacteria impaired neutrophil functions, e.g. chemiluminescence response, and leukotriene formation to a significantly higher degree as compared to nonmucoid P. aeruginosa bacteria. To study the cell biological requirements for the different cellular response pattern by mucoid and nonmucoid (NM) P. aeruginosa bacteria, further experiments were performed with purified alginate, the mucoid exopolysaccharide of P. aeruginosa (MEP). In this regard the MEP (alginate) significantly reduced the zymosan-induced leukotriene B4 (LTB4) formation (from 40 +/- 7 to 2 +/- 4 ng). The chemiluminescence response induced by NM bacteria was abolished when the bacteria were precoated with the MEP. Mucoid and NM P. aeruginosa bacteria interacted with components of the cellular signal transduction pathway to a different degree. Mucoid bacteria induced a 2-fold enhanced GTPase activity but activated the protein kinase C (PKC) to a lesser degree than NM P. aeruginosa bacteria. Prior exposure of neutrophils to the MEP increased the sodium fluoride (NaF)-induced GTPase activity and guanylylimidodiphosphate binding [Gpp(NH)p] by approximately 60 and 30%, respectively. The phorbol myristic acid-induced PKC activation was inhibited by 30-40% in the presence of the MEP. However, the MEP by itself was inactive in all assay systems. Our results indicate that the MEP represents an important component which modulates neutrophil responses of mucoid as compared to NM P. aeruginosa bacteria, e.g. the chemiluminescence response, LTB4 generation, and the interaction with components (G proteins, PKC) of the signal transduction pathway.
Alginates↗
Analysis of IgE-binding factors (sCD23) within newborn sera.
Factors are discussed as potential diagnostic parameters of atopic disorders. The amounts of sCD23 in sera from newborn children (n = 4,329) were determined by radioimmunoassay with monoclonal antibodies specific for CD23. The sCD23 levels ranged between 0 and 81.5 ng/ml of CD23-specific mAb. Furthermore, the sera of newborns with more than 5 ng/ml (n = 45) were analyzed by SDS/PAGE and subsequent autoradiography using 125I-labeled IgE (PS). These experiments indicate that newborn sera with normal sCD23 amounts contain an IgE-binding activity with a molecular weight of 25 kD; this component was not observed within sera containing elevated amounts (> 5 ng/ml). In addition, a 60-kD IgE-binding component was detected within most of all newborn sera (76.4%). The data show that the IgE-binding pattern of newborn children and the pattern of adult donors are different. Our data suggest that the measurement of quantitative sCD23 amounts combined with the analysis of the molecular weight pattern of the IgE-binding factors might be a helpful diagnostic parameter with regard to IgE-associated diseases.
Antibodies, Monoclonal↗
Cytokine-induced (interleukins-3, -6 and -8 and tumour necrosis factor-beta) activation and deactivation of human neutrophils.
The effect of various cytokines [interleukin-3(IL-3), IL-6, IL-8, tumour necrosis factor-beta (TNF-beta)] on human neutrophils (PMN) was analysed with regard to the generation of leukotrienes and the involvement of guanosine triphosphate (GTP)-binding proteins (G proteins). Incubation of cytochalasin B-pretreated PMN with cytokines alone did not lead to a generation of leukotrienes. However, the cytokines affected the formyl-methionyl-leucyl-phenylalanine-(FMLP)-induced formation of leukotrienes in a time-dependent manner. Preincubation of the cells with the different cytokines for short periods (15 seconds at 37 degrees) enhanced the subsequent FMLP-induced leukotriene generation, whereas preincubation for prolonged times resulted in a reduced formation of leukotrienes. These results correlated with the respective G protein-associated guanosine triphosphatase (GTPase) activities within isolated membrane fractions. The present study indicates a modulation of the FMLP-induced leukotriene formation by diverse cytokines via interaction with the GTP-binding proteins.
Cells, Cultured↗
Effects of mucoid and non-mucoid Pseudomonas aeruginosa isolates from cystic fibrosis patients on inflammatory mediator release from human polymorphonuclear granulocytes and rat mast cells.
Mucoid Pseudomonas aeruginosa causing chronic bronchopulmonary infection in cystic fibrosis (CF) patients may interfere with host defence mechanisms. We investigated 13 P. aeruginosa strains isolated from sputa of CF patients with regard to the induction or modulation of inflammatory mediator release from human neutrophils (PMN) and rat mast cells. The effects of mucoid as compared to non-mucoid bacteria were studied using a mucoid strain and its non-mucoid revertant. The release of leukotrienes (LT) and histamine in response to the majority of the CF strains was insignificant. However, preincubation of PMN with P. aeruginosa caused a dose-dependent decrease (50-95%) of LTB4 and LTC4 generation and LTB4 metabolism induced by the Ca(2+)-ionophore A23187 or opsonized zymosan (ZX) (P less than 0.001). The mucoid strains caused a three- to 10-fold higher impairment of LTB4 release (P less than 0.05) and a concomitant down-regulation of LTB4 receptors on neutrophils. Inhibitory effects were also obtained for mucoid and non-mucoid bacteria when the phorbol-ester or the Ca(2+)-ionophore induced luminol enhanced chemiluminescence response (P less than 0.001) or the histamine release from rat peritoneal mast cells (P less than 0.01) was studied. The bacteria-cell contact with non-mucoid strains was associated with an increased Ca2+ influx into PMN, whereas mucoid bacteria had no effect. In addition, a protein kinase C-dependent decrease of the C3bi receptor was suppressed by the mucoid--and less effectively--by the non-mucoid strain. The results suggest that the impairment of the phagocytic and inflammatory system may contribute to the pathogenesis and persistence of mucoid P. aeruginosa infection in CF.
Animals↗
Modulation of leukotriene generation by invasive bacteria.
The effect of invasive bacteria on the release of proinflammatory mediators (oxygen radicals, leukotriene release) from human polymorphonuclear neutrophils was studied. Bacterial stimuli were used including genetically cloned invasive Yersinia enterocolitica strains 108-P (bearing the phagocytosis-resistance plasmid) and 108-C (plasmidless variant), Listeria monocytogenes [SLCC 5779 (inv-) and NCTC 7973 (inv+)] as well as an Escherichia coli K 12 strain (pRI 203) in which the inv gene of Y. pseudotuberculosis was cloned. When human polymorphonuclear granulocytes were studied as target cells the inv+ as well as the inv- strains were phagocytosed to a comparable amount with the exception of the L. monocytogenes strain (inv+). Among the invasive strains E. coli HB 101 (pRI 203) was the most active to trigger polymorphonuclear leucocytes (PMN) for oxygen radical production. Preincubation of the cells with bacteria and subsequent stimulation with the Ca ionophore A23187 or opsonized zymosan suppressed the chemiluminescence response to a different degree. The various bacterial strains did not induce leukotriene release from endogenous arachidonic acid. Subsequent stimulation of the infected cells with Ca ionophore or opsonized zymosan led to an altered pattern of the combined amounts of leukotriene B4 (LTB4), 20-OH- and 20-COOH-LTB4 as well as the ratio of LTB4 versus 20-OH and 20-COOH-LTB4. Infection of the cells also reduced strain dependently the number of LTB4-receptor sites. Our data suggest that bacterial uptake modulates the inflammatory response of granulocytes (e.g. chemiluminescence response, leukotriene generation).
Bacterial Adhesion↗
Leukotriene A4 modulates generation of leukotriene B4 and sulphidopeptide leukotrienes by human neutrophils.
We investigated the influence of exogenous leukotriene A4 (LTA4) on the reactivity of polymorphonuclear leucocytes (PMN). PMN were either prestimulated with LTA4 or incubated simultaneously with LTA4 and the Ca ionophore A23187 or sodium fluoride (NaF). The Ca ionophore A23187 and NaF induced generation of LTB4 from PMN was significantly diminished in the presence of LTA4 while the formation of LTC4 was enhanced. In contrast, preincubation of cells with LTA4 followed by subsequent stimulation with NaF synergistically increased the LTB4 generation from PMN. LTA4, either alone or in combination with the calcium ionophore A23187 or NaF, decreases GTPase activity in human PMN. This decrease was abolished when LTA4 pretreated cells were subsequently stimulated with NaF, but not with calcium ionophore A23187, suggesting a regulatory role of LTA4 on G-proteins. The results demonstrate dual functions of LTA4: it serves as a substrate for the generation of leukotrienes and also regulates the susceptibility of human PMN for subsequent response.
Calcimycin↗
[LDL cholesterol apheresis by adsorption to dextran sulfate].
Extracorporeal LDL cholesterol elimination may be the sole successful treatment in familial hypercholesterolemia. By treatment of 41 plasma both total cholesterol and LDL cholesterol were lowered by 69 +/- 1% and 78 +/- 1%, respectively. HDL cholesterol was decreased by 24 +/- 3%. This could be explained both by hemodilution (hematocrit was decreased by 9.3 +/- 2.8%) and unspecific adsorption of various plasma proteins (-15.3 +/- 3.7 g/l, i.e. -22 +/- 4%). Protein electrophoresis showed different affinities of the protein fractions. These data suggest that LDL apheresis by dextran sulfate is an effective method for the elimination of LDL cholesterol. However, other proteins besides apolipoprotein B are adsorbed to dextran sulfate.
Adsorption↗
Priming mechanisms and induction of heat shock proteins in human polymorphonuclear granulocytes induced by eicosanoids and cytokines.
Priming of human polymorphonuclear granulocytes (PMNs) with cytokines (IL-3, IL-6, TNF-alpha) followed by a subsequent stimulation (FMLP) led to an enhanced polymerization of actin and GTPase-activity which correlated to a loss of ras immunoreactivity and an increased expression of Rab proteins. Furthermore TNF-alpha and 12-HETE induced the heat shock proteins (hsp 70 family) in PMNs as was demonstrated by metabolic radiolabeling and Western blotting (anti-hsp 72). This activation of the stress response exerted a protective function towards a subsequent lytic attack by a bacterial cytolysin (leukocidin).
12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
Effects of Staphylococcus aureus cell wall products (teichoic acid, peptidoglycan) and enterotoxin B on immunoglobulin (IgE, IgA, IgG) synthesis and CD23 expression in patients with atopic dermatitis.
The influence of staphylococcal cell wall products (teichoic acid, peptidoglycan) and enterotoxin B on peripheral blood lymphocytes (PBL) from patients with atopic dermatitis (AD) was investigated. The parameters studied were spontaneous and interleukin-inducible immunoglobulin (IgA, IgE, IgG) synthesis and CD23 expression. PBL from non-atopic donors served as controls. Teichoic acid and peptidoglycan induced an enhanced synthesis of IgA and IgG in normal donors. However, IgA and IgG synthesis in PBL from patients with AD was significantly suppressed by teichoic acid and enterotoxin B. The incubation of PBL from normal donors with enterotoxin B and interleukin-4 (IL-4) or IL-5 led to a significant suppression of IgA and IgG synthesis. Co-stimulation of PBL with teichoic acid or peptidoglycan and IL-4 led to a pronounced increase in IgE synthesis and CD23 expression in patients with AD. Our data indicate that cell wall products and toxins of staphylococci modulate the cytokine-dependent humoral immunity in patients with AD and may be responsible for allergic skin reactions in AD.
Antigens, Bacterial↗
Neomycin induces stimulatory and inhibitory effects on leukotriene generation, guanine triphosphatase activity, and actin polymerization within human neutrophils.
The effects of neomycin on human neutrophils (PMN) were studied with respect to the generation of leukotrienes, the involvement of guanine triphosphate binding proteins (G proteins) and the polymerization of actin. Incubation of neutrophils with neomycin induced the generation of low amounts of leukotrienes. Co-incubation of neutrophils with neomycin and the direct G-protein activator sodium fluoride (NaF) resulted in an enhanced leukotriene formation at 0.5 mM neomycin and an inhibition at a concentration of 10 mM. Simultaneous incubation with neomycin and formyl-methionyl-leucyl-phenylalanine (FLMP) did not affect the FMLP-induced leukotriene formation. However, pretreatment of neutrophils with 10 mM neomycin followed by the addition of NaF or FMLP resulted in an enhanced generation of leukotrienes. Crude membrane fractions of PMN incubated with neomycin at different concentrations showed an enhanced (0.5 mM) as well as a reduced (10 mM) guanine triphosphatase activity. Furthermore, incubation of neutrophils with neomycin above a concentration of 0.5 mM led to the depolymerization of actin. The presented results show inhibitory and stimulatory effects of neomycin on various cell functions, which may reflect differences in transmembrane signalling.
Actins↗
Analysis of leukotriene B4 metabolism in human promyelocytic HL-60 cells.
We previously reported that human alveolar macrophages rapidly metabolize the chemotactic active lipid mediator leukotriene B4 (LTB4) into the dihydro-LTB4 by reduction of one of the conjugated double bonds. We herein report that human HL-60 cells (a myeloid precursor which can be differentiated into granulocyte- as well as monocyte-like cells by dimethyl sulphoxide or phorbol myristate acetate) express a highly active LTB4 reductase in the undifferentiated state. Differentiation by dimethyl sulphoxide (1.3%) along the granulocyte lineage, as confirmed by light microscopy, conversion of NitroBlue Tetrazolium into formazan, failed to induce a substantial capacity for omega-oxidation of LTB4; this reaction is exclusively found in mature granulocytes. Studies with the cell homogenate of undifferentiated HL-60 cells indicated that the activity of the enzyme depends on the presence of NADPH, Ca2+ and Mg2+, with a pH optimum of 7.5 at 37 degrees C. The enzyme was not released into the supernatant after stimulation of HL-60 cells with phorbol myristate acetate (100 ng) or Ca2+ ionophore (7.5 microM). Subcellular fractionation revealed evidence that the LTB4 reductase is located within the membrane fraction. Purification of the enzyme by gel filtration and gel electrophoresis suggests an apparent molecular mass of 40 kDa.
Chromatography, Gel↗
Bacterial toxins induce heat shock proteins in human neutrophils.
We studied the influence of different bacterial toxins (alveolysin; toxic shock syndrome toxin 1, TSST-1 and erythrogenic toxin A, ETA) on the expression of heat shock proteins (hsps) in isolated human polymorphonuclear granulocytes (PMNs). As was shown by Western blotting (anti-hsp72) ETA and TSST-1 were potent inducers of hsps at low toxin concentrations (10 ng/ml). Alveolysin led to the expression of hsps at hemolytic concentrations (1 HU; 700 ng/ml) whereas at subhemolytic concentrations (7 ng/ml) no heat shock response was observed. The induction of heat shock proteins was also accompanied by increased mRNA levels for hsp70 as was determined by PCR-analysis.
Bacterial Proteins↗