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1.
We studied the oxidation of alanine and methionine by human neutrophils. Phagocytosis enhanced the decarboxylation of amino acids by human neutrophils. Decarboxylation of amino acids was dependent on the myeloper-oxidase system (MPO-H2O2-Cl–). This was further confirmed using purified canine MPO. Human neutrophils and the MPO system were about 10 times more efficient in decarboxylating alanine than methionine. They also oxidized methionine to methionine sulfoxide. The fraction of methionine decarboxylated by human neutrophils or the MPO system was small compared to the fraction which was oxidized to methionine sulfoxide. Thus methionine was preferentially oxidized to methionine sulfoxide by the MPO system. However, once methionine was oxidized to methionine sulfoxide, it was readily decarboxylated by the MPO system. The results suggest that the thio group of methionine prevents its carboxylic group from being decarboxylated. 相似文献
2.
Role of the Phagocyte in Host-Parasite Interactions XXVII. Myeloperoxidase-H(2)O(2)-Cl-Mediated Aldehyde Formation and Its Relationship to Antimicrobial Activity
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Evidence is presented which suggests that the mechanism of action of the myeloperoxidase-H(2)O(2)-Cl(-) antimicrobial system in the phagocyte is by the formation of aldehydes. Aldehyde production resulting from myeloperoxidase-mediated decarboxylation and deamination of alanine was quantitated with 20,000-g granules from guinea pig polymorphonuclear leukocytes serving as the enzyme. Equimolar quantities of acetaldehyde and CO(2) were obtained. There was an absolute requirement for both H(2)O(2) and Cl(-) for decarboxylation by the myeloperoxidase-containing granules. The myeloperoxidase-H(2)O(2)-Cl(-) system decarboxylated both d- or l-alanine equally and had a pH optimum of 5.3. Decarboxylation of l-alanine by intact guinea pig polymorphonuclear leukocytes was increased 2.5-fold by phagocytosis. Guaiacol peroxidation by the granules was inhibited 90% in the presence of Cl(-) at acid pH. Under these conditions, decarboxylation and deamination of amino acids by myeloperoxidase were significantly stimulated, resulting in aldehyde production. Taurine, a competitive inhibitor of amino acid decarboxylation, inhibited bactericidal activity of the myeloperoxidase-H(2)O(2)-Cl(-) system but had no effect on the myeloperoxidase-H(2)O(2)-I(-) bactericidal system. Since the myeloperoxidase-H(2)O(2)-I(-) system does not participate in amino acid decarboxylation, its mechanism of antimicrobial action appears to be different from that found with Cl(-). 相似文献
3.
Li H Cao Z Moore DR Jackson PL Barnes S Lambeth JD Thannickal VJ Cheng G 《Infection and immunity》2012,80(7):2528-2537
Members of the heme peroxidase family play an important role in host defense. Myeloperoxidase (MPO) is expressed in phagocytes and is the only animal heme peroxidase previously reported to be capable of using chloride ion as a substrate to form the highly microbicidal species hypochlorous acid (HOCl) at neutral pH. Despite the potent bacterial killing activity of HOCl, individuals who fail to express MPO typically show only a modest increase in some fungal infections. This may point to the existence of redundant host defense mechanisms. Vascular peroxidase 1 (VPO1) is newly discovered member of the heme peroxidase family. VPO1 is expressed in cells of the cardiovascular system and is secreted into the bloodstream. In the present study, we investigate whether VPO1 is capable of generating HOCl and its role in host defense. Like MPO, VPO1 in the presence of H2O2 and chloride generates HOCl. VPO1-dependent HOCl generation was demonstrated by chlorination of taurine and tyrosine using mass spectrometry. In addition, the VPO1/H2O2/Cl− system can cause the chlorination of monochlorodimedone and the oxidation of 5-thio-2-nitrobenzoic acid. Purified VPO1 and VPO1 in plasma mediate bacterial killing that is dependent on chloride and H2O2; killing is inhibited by peroxidase inhibitors and by the H2O2 scavenger catalase. In the presence of erythrocytes, bacterial killing by VPO1 is slightly reduced. Thus, VPO1, in addition to MPO, is the second member of the heme peroxidase family capable of generating HOCl under physiological conditions. VPO1 is likely to participate in host defense, with bactericidal activity mediated through the generation of HOCl. 相似文献
4.
Summary Analysis of three inherited defects of granulocyte function (Chediak-Higashi Syndrome, CHS; Chronic Granulomatous Disease, CGD; Myeloperoxidase Deficiency, MPO) has highlighted critical events for the antimicrobial function of these cells and placed others in perspective. Prompt phagosomal fusion may be more important for digestion of organisms rather than killing as indicated by the mild bactericidal defects in the CHS. The formation of O2- and H2O2 during the phagocytic respiratory burst is central for the broad antimicrobial activity of granulocytes. MPO, on the other hand, while perhaps normally participating in granulocyte antimicrobial action, appears to be essential only for the effective killing of eukaryotic organisms such as certain fungal strains. While the non-oxidative killing mechanism of neutrophils have stimulated much recent interest and were the first to be defined no specific inherited defects have been discovered which are clinically important.Genetic disorders of macrophage effector function remain to be clearly defined as do those of eosinophils. The lessons learned from the study of the granulocyte defects discussed have provided both the technology and approach to the analysis of the antimicrobial and cytocidal mechanisms of these important phagocytic cells.Supported in part by USPHS No. 5 ROI AI13251 相似文献
5.
A. A. Jacobs I. E. Low B. B. Paul R. R. Strauss A. J. Sbarra 《Infection and immunity》1972,5(1):127-131
A mycoplasmacidal system consisting of myeloperoxidase (MPO)-containing granules, H(2)O(2), and a halide is described. In all parameters measured, it appears to be identical to the MPO-H(2)O(2)-halide bactericidal system previously reported. It has a pH optimum of approximately 5.5 and an optimal MPO:H(2)O(2) ratio of 1:25. The halide requirement can be satisfied by either chloride or iodide. Through the use of taurine or horseradish peroxidase substitution, chloride-mediated killing can be distinguished from iodide-mediated killing. The relationship of this mycoplasmacidal system to other mycoplasmacidal systems and to host surveillance of mycoplasma is discussed. 相似文献
6.
Function of h(2)o(2), myeloperoxidase, and hexose monophosphate shunt enzymes in phagocytizing cells from different species 总被引:6,自引:9,他引:6
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The effect of phagocytosis on the H2O2 production and myeloperoxidase (MPO) activities of leukocytes from various species was investigated. The intracellular distribution of MPO, reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase (6PGDH) of resting and phagocytizing guinea pig polymorphonuclear leukocytes has also been studied. Phagocytizing cells produce more H2O2 than the corresponding resting cells. This has been found to be true for human peripheral polymorphonuclear leukocytes, mouse peritoneal macrophage, and guinea pig and rat peritoneal polymorphonuclear leukocytes. All of these cells, except rabbit alveolar macrophages, have significant MPO activity. Generally an increased activity is noted with phagocytizing cells. Homogenization and differential centrifugation of guinea pig peritoneal polymorphonuclear leukocytes indicate that the whole homogenate and its fractions from phagocytizing cells have significantly higher MPO and NADPH oxidase activities, when compared to the corresponding fractions from the resting cells. The 27,000 × g supernatant fluid from phagocytizing cells has 6-fold more MPO and 2.5-fold more NADPH oxidase activity than similar supernatant fractions from resting cells. The enzyme 6PGDH was unaffected by phagocytosis. The relationship of these stimulated activities to the intracellular bactericidal function of the phagocytes has been discussed. 相似文献
7.
Interaction of Aspergillus fumigatus Spores with Human Leukocytes and Serum 总被引:13,自引:7,他引:6
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Serum was necessary for optimal phagocytosis of Aspergillus fumigatus spores by human leukocytes, and its opsonic capacity was greatly diminished by heat inactivation (56 C, 30 min). A germination assay, described in this report, was developed to study the fate of phagocytized spores. After incubation for 3 hr with normal leukocytes and serum, spores ingested by peripheral blood neutrophils and monocytes remained viable. Since we had previously found that myeloperoxidase (MPO), a lysosomal enzyme of human neutrophils and monocytes, exerted fungicidal activity against Candida albicans when combined with H2O2 and chloride or iodide, the effects of these substances on A. fumigatus spores were examined. Spore viability was not impaired by MPO alone, H2O2 alone, or KI alone, but high concentrations of KI and H2O2 in combination caused marked inhibition of subsequent germination. MPO imparted fungicidal activity to concentrations of KI and H2O2 that lacked any effect in its absence. NaCl, in combination with MPO and H2O2, was far less effective than the iodide salt against A. fumigatus. The relative ineffectiveness of chloride in this system could underly the apparent inability of human neutrophils to kill ingested A. fumigatus spores, despite their competence to kill C. albicans. 相似文献
8.
Min-Fu Tsan 《Infection and immunity》1982,36(1):136-141
The myeloperoxidase (MPO)-mediated decarboxylation of amino acids and the MPO-mediated oxidation of methionine, two potential bactericidal mechanisms, were compared. In the presence of the MPO system (MPO, 50 mU/ml; H(2)O(2), 0.1 mM; Cl(-), 75 mM), 50% of alanine (0.1 mM) was decarboxylated, whereas only 5% of methionine (0.1 mM) was decarboxylated. In contrast, under similar conditions, 80% of methionine was oxidized to methionine sulfoxide. Once methionine was oxidized to methionine sulfoxide, it was decarboxylated (75%) by the MPO system. Methionine at 0.1 mM completely inhibited the decarboxylation of alanine, whereas alanine at a concentration 200 times that of methionine had no effect on the MPO-mediated oxidation of methionine. Sodium azide, an MPO inhibitor, inhibited the decarboxylation of alanine and the oxidation of methionine to the same extent. Tryptophan markedly inhibited the oxidation of methionine, whereas histidine stimulated it. Alanine, glycine, and taurine had no effect. In contrast, all of these amino acids and taurine markedly inhibited the MPO-mediated decarboxylation of alanine. NaN(3), tryptophan, and methionine, which inhibited the MPO-mediated oxidation of methionine, also inhibited the killing of Staphylococcus aureus or Klebsiella pneumoniae by the MPO system; whereas histidine, alanine, and glycine, which did not inhibit the oxidation of methionine, had less or no effect on the killing of these two bacteria by the MPO system. Results suggest that methionine is preferentially oxidized to methionine sulfoxide by the MPO system. Once methionine is oxidized to methionine sulfoxide, it is then readily decarboxylated by the MPO system. The agent responsible for the oxidation of methionine may play an important role in the MPO-mediated killing of bacteria. 相似文献
9.
Ratnam J. Selvaraj Benoy B. Paul Robert R. Strauss A. Alice Jacobs Anthony J. Sbarra 《Infection and immunity》1974,9(2):255-260
The antimicrobial activities of the myeloperoxidase-H(2)O(2)-halide system have received considerable attention recently. The precise mechanism by which this system exerts its lethal activity is presently not clear. In an effort to learn more regarding a possible mechanism of action, the susceptibility of protein-bound amino acids to enzymatic attack by myeloperoxidase (MPO) in the presence of chloride ions was investigated. [1, 7-(14)C]diaminopimelic acid (DAP) was incorporated into Escherichia coli W-7 proteins with little randomization of the radioactivity. Under appropriate conditions, it was observed that the MPO-H(2)O(2)-halide system released approximately 94% of the radioactivity from labeled bacteria. This would indicate that, in addition to decarboxylation, peptide bonds are also split during this reaction. The oxidative decarboxylation of DAP-labeled bacteria by MPO (i) is Cl(-) dependent, (ii) has an acid pH optimum, (iii) requires a specific concentration of H(2)O(2) for activity, (iv) reaches a plateau by 25 min, and (v) is markedly inhibited by taurine. These properties are similar to those observed with free amino acids. It appears from these data that MPO can not only decarboxylate free and bound amino acids, yielding aldehydes, but also it can actively participate in oxidative peptide cleavage. Both of those activities may play a critical role in the microbicidal action of the leukocyte. 相似文献
10.
Gary Pekoe Knox Van Dyke Henry Mengoli David Peden Denis English 《Inflammation research》1982,12(1-2):232-238
The interaction of myeloperoxidase (MPO) with H2O2 and Cl– provides a potent antimicrobial/cytotoxic system for polymorphonuclear leukocytes (PMNs). MPO-related cytotoxicity may be associated with the formation of toxic oxidant MPO intermediates, HOCl, or both. MPO itself is able to oxidize drugs and cellular components. Non-steroidal anti-inflammatory drugs (NSAIDs) able to act as antioxidant free radical scavengers have recently been shown to inhibit luminol-enhanced chemiluminescence (CL) which results from the MPO–H2O2–Cl– reaction. CL is a measure of the activity of this reaction. At that time it was not clear whether the source of CL which these NSAIDs affected was HOCl or components of the initial MPO–H2O2–Cl– reaction. A NSAID antioxidant mechanism could affect MPO oxidant intermediates and HOCl.This study compares the effects of antioxidant NSAIDs, methylprednisone and free radical scavengers against MPO-based and NaOCl-based luminol-enhanced CL. Most NSAIDs which affected both MPO and NaOCl-CL appeared to share similar mechanisms, suggesting that MPO oxidant internediates and HOCl are susceptible to NSAID effects. However, most NSAIDs were more effective against MPO-CL. The effect of these NSAIDs against MPO-CL followed the profile of NSAIDs effective in previous studies against PMN-CL. One exception to this was methylprednisone, which has no effect on PMN or MPO-CL, yet inhibited NaOCl-CL. This and other data suggest that MPO and not HOCl-related reactions are a major source of PMN-CL. Less effective NSAIDs affected NaOCl-CL better than MPO-CL. While both HOCl and MPO oxidant intermediates may be affected by NSAIDs, it appears that MPO oxidant intermediates or MPO itself are the primary target for NSAID antioxidant free radical scavenging mechanisms. These antioxidant effects impair the major killing system of the PMN and may be NSAIDs' primary anti-inflammatory mechanism. Although our data suggests the production of superoxide anion and hydroxyl radical from the MPO–H2O2–Cl– reaction, the actual presence or involvement of these free radical species is not confirmed herein. 相似文献
11.
Giuliano Zabucchi Renzo Menegazzi Lucia Roncelli Paolo Bertoncin Francesco Tedesco Pierluigi Patriarca 《Inflammation》1990,14(1):41-53
This study investigated the interaction between neutrophil myeloperoxidase (MPO) and the C1q component of the complement system. Using a dot-spot assay, MPO was found to bind to C1q in a dose-dependent manner. The specificity of this reaction was proved by the inhibitory effect of F(ab')2 antibodies to C1q and by the inability of MPO to bind to Clr, Cls and IgG. The interaction between MPO and C1q did not influence the enzymatic activity of the peroxidase but resulted in a more stable C1q as assessed by hemolytic assay for C1q. The protective effect of MPO on C1q did not require the presence of H2O2 in the reaction mixture nor was it inhibited by sodium azide, whereas it was abolished by heating the peroxidase. Lactoferrin and lysozyme, unlike MPO, were ineffective in protecting C1q from functional decay. Addition of H2O2 and chloride to MPO and C1q led to a complete inactivation of C1q, which could not be induced by H2O2 alone. The hypochlorite, which is known to be generated during the reaction of MPO with H2O2 and chloride, exhibited a similar inactivating effect on C1q, which was prevented by an external source of methionine. 相似文献
12.
We have previously shown that deuterium oxide (D2O) enhances the oxidation of methionine, a myeloperoxidase (MPO)-mediated reaction, by human neutrophils during phagocytosis. However, D2O has no effect on the oxidation of methionine by the purified MPO-H2O2-Cl– system. To explain this observation, we studied the effect of D2O on the oxidative metabolism, phagocytosis, and lysosomal enzyme release by human neutrophils. D2O stimulated the hexose monophosphate shunt (HMS) activity of resting neutrophils in a dose-response fashion. In the presence of latex particles or phorbol myristate acetate (PMA), D2O brought about an exaggerated stimulation of the HMS activity. This enhancement of the HMS activity by D2O was markedly reduced when neutrophils form two patients with X-linked chronic granulomatous disease (CGD) were used, either in the presence or absence of latex particles or PMA. Superoxide and H2O2 production by neutrophils in the presence of latex particles or PMA were also stimulated by D2O. In contrast, D2O inhibited the ingestion of latex particles. D2O enhanced the extracellular release of MPO, but not lactate dehydrogenase, by neutrophils only in the simultaneous presence of cytochalasin B and latex particles. The enhancement of HMS activity and MPO release by D2O was partially inhibited by colchicine. Our results suggest that enhancement of neutrophil oxidative metabolism by D2O may in part explain the stimulation of methionine oxidation by phagocytosing neutrophils. 相似文献
13.
Gary Pekoe Knox Van Dyke David Peden Henry Mengoli Denis English 《Inflammation research》1982,12(3):371-376
The action of non-steroidal anti-inflammatory drugs (NSAIDS) has been ascribed to their ability to block the reaction of arachidonate with cyclooxygenase/peroxidase, thus inhibiting the cellular production of inflammation mediators such as prostaglandins and leukotrienes. However, this and other polymorphonuclear leukocyte (PMN) peroxidases such as myeloperoxidase (MPO) would still be capable of producing destructive oxidants which contribute to inflammation. Sulindac sulfide (Clinoril® sulfide) has recently been shown to scavenge oxidant products of prostaglandin cyclooxygenase/peroxidase and MPO. The MPO–H2O2–Cl– reaction is a potent antimicrobial/cytotoxic system which produces HOCl, a strong oxidant. MPO itself has the ability to oxidize drugs and cellular components, and may be the main oxidant in PMN defenses. An antioxidant/free radical scavenger action of NSAIDs against the MPO system could be a primary mechanism of their anti-inflammatory effects. Other antioxidant/free radical scavengers have anti-inflammatory effects.MPO activity has previously been quantified using chemiluminescence (CL). In this study, NSAIDs from various classes were tested for their ability to inhibit luminol-enhanced CL from MPO. The most potent NSAIDs against MPO-CL were BW755C, phenylbutazone, indomethacin and sulindac sulfide. Salicylates and arylacetic acid derivatives, such as naproxen, also decreased MPO-CL. These drugs are also effective against CL from PMNs, of which MPO may be a main source. This effect of NSAIDs on MPO suggests that NSAIDs may impair the killing mechanism of the PMN, preventing cell destruction and release of inflammation mediators. PMN MPO appears to be a target for the antioxidant/free radical scavenging effects of NSAIDs. 相似文献
14.
Spleen cell suspensions from AKR and CD-1 mice contain peroxidase activity as determined by guaiacol oxidation. This activity is found predominately in the 20,000 × g pellet fraction of spleen cell homogenates. In the presence of H2O2 and chloride ion at acidic pH, splenic peroxidase mediates the oxidation of d- or l-alanine to CO2, NH3, and acetaldehyde. The same reaction mixture without added amino acid can kill both gram-positive and gram-negative bacteria. The conditions for both reactions are similar. Both have an absolute requirement for H2O2 and chloride ion, neither is active at neutral or alkaline pH, and both are inhibited by the sulfonic amino acid taurine. In these aspects, splenic peroxidase is qualitatively similar in its activity to myeloperoxidase (MPO) from polymorphonuclear leukocytes. It is quantitatively different from MPO in that the latter is more potent on a per guaiacol unit basis with respect to both amino acid oxidation and bactericidal activity. Still another quantitative difference is that splenic peroxidase requires 0.1 m NaCl for activity, whereas MPO functions with as little as 0.005 m NaCl. Splenic peroxidase and MPO both appear to differ qualitatively from horseradish peroxidase in that the latter enzyme does not mediate amino acid oxidation. 相似文献
15.
Nanduri R. Prabhakar 《Respiratory physiology & neurobiology》2012,184(2):165-169
Carotid bodies are sensory organs for monitoring arterial blood oxygen (O2) levels, and the ensuing reflexes maintain cardio-respiratory homeostasis during hypoxia. This article provides a brief update of the role of carbon monoxide (CO) and hydrogen sulfide (H2S) in hypoxic sensing by the carotid body. Glomus cells, the primary site of O2 sensing in the carotid body express heme oxygenase-2 (HO-2), a CO catalyzing enzyme. HO-2 is a heme containing enzyme and has high affinity for O2. Hypoxia inhibits HO-2 activity and reduces CO generation. Pharmacological and genetic approaches suggest that CO inhibits carotid body sensory activity. Stimulation of carotid body activity by hypoxia may reflect reduced formation of CO. Glomus cells also express cystathionine γ-lyase (CSE), an H2S generating enzyme. Exogenous application of H2S donors, like hypoxia, stimulate the carotid body activity and CSE knockout mice exhibit severely impaired sensory excitation by hypoxia, suggesting that CSE catalyzed H2S is an excitatory gas messenger. Hypoxia increases H2S generation in the carotid body, and this response was attenuated or absent in CSE knockout mice. HO inhibitor increased and CO donor inhibited H2S generation. It is proposed that carotid body response to hypoxia requires interactions between HO-2–CO and CSE–H2S systems. 相似文献
16.
Studies of phagocytosis in chronic granulomatous disease 总被引:4,自引:0,他引:4
Abnormal phagocyte function in chronic granulomatous disease (CGD) is associated with decreased bactericidal activity. Ingestion of serum-opsonized organisms is reported to be normal in these patients. We previously showed that in CGD the expression of C3b receptors (CR1) on polymorphonuclear leukocytes (PMNs) is significantly depressed. In this study, we compared the phagocytic activity of the PMNs from normal healthy controls with that of CGD patients and one individual with myeloperoxidase (MPO) deficiency. The ingestion of sheep erythrocytes (E) by PMNs adherent to a glass surface was examined; the E were coated either with excess IgG (E-IgG) or with C3b plus limited IgG (EAC3b-IgG). The PMNs, both in CGD and in MPO deficiency, ingested E-IgG and EAC3b-IgG at levels markedly above normal. C3b-coated erythrocytes were not phagocytosed. Preincubating the PMNs with sodium azide, which blocks MPO, or catalase, a scavenger of H2O2, caused a marked increase in phagocytosis by normal PMNs. Azide had a variable effect on PMN activity in CGD and no effect on the activity in the subject with MPO deficiency. Even in the presence of azide, the ingestion of EAC3b-IgG by the PMNs from the CGD patients was significantly greater than that seen in paired normals [mean phagocytic index (PI), 2.13 for CGD vs. 1.48 for normals;P < 0.05 by the paired samplet test]. Similar results were obtained with ingestion of E-IgG. Notably, ingestion of serum-opsonizedCandida organisms (relatively nondegradable particles) was markedly above normal with CGD PMNs and, in normal PMNs, azide treatment also evoked an increase. In addition, rosette formation of the adhered PMNs with E-IgG was enhanced with CGD and the azide-treated normal PMNs. We demonstrated that this increased activity was not the result of increased Fc receptor (FcR) number, as determined from the binding of a monoclonal anti-FcR antibody. Both the E-IgG rosette formation and the ingestion by CGD PMNs were abrogated in the presence of an H2O2-generating system. In contrast, the phagocytic activity of MPO-deficient PMNs was not altered by exogenous H2O2. These findings suggest that cellular products generated by the H2O2-MPO-halide system down-regulate the rosette-forming and phagocytic activity of PMNs from normal healthy individuals, but not that from CGD and MPO-deficient patients. 相似文献
17.
Addition of thiourea (TU) or dimethylthiourea (DMTU) decreased killing ofStaphylococcus aureus, 502A, and decreased concentrations of hydrogen peroxide (H2O2), and hydroxyl radical (·OH), but not Superoxide anion (O2
–) or lysozyme concentrations, in mixtures containing human neutrophilsin vitro. Addition of TU or DMTU also decreased concentrations of H2O2, · OH, or hypochlorous acid (HOCl) in neutrophil-free mixtures exposed to-D-glucose and glucose oxidase, gamma irradiation, or HOCl, respectively. Our results suggest that TU or DMTU can decrease neutrophil-mediated killing of bacteria by inhibiting O2 metabolite-dependent bactericidal mechanisms. 相似文献
18.
Painter RG Bonvillain RW Valentine VG Lombard GA LaPlace SG Nauseef WM Wang G 《Journal of leukocyte biology》2008,83(6):1345-1353
Chloride anion is essential for myeloperoxidase (MPO) to produce hypochlorous acid (HOCl) in polymorphonuclear neutrophils (PMNs). To define whether chloride availability to PMNs affects their HOCl production and microbicidal capacity, we examined how extracellular chloride concentration affects killing of Pseudomonas aeruginosa (PsA) by normal neutrophils. PMN-mediated bacterial killing was strongly dependent on extracellular chloride concentration. Neutrophils in a chloride-deficient medium killed PsA poorly. However, as the chloride level was raised, the killing efficiency increased in a dose-dependent manner. By using specific inhibitors to selectively block NADPH oxidase, MPO, and cystic fibrosis transmembrane conductance regulator (CFTR) functions, neutrophil-mediated killing of PsA could be attributed to three distinct mechanisms: CFTR-dependent and oxidant-dependent; chloride-dependent but not CFTR- and oxidant-dependent; and independent of any of the tested factors. Therefore, chloride anion is involved in oxidant- and nonoxidant-mediated bacterial killing. We previously reported that neutrophils from CF patients are defective in chlorination of ingested bacteria, suggesting that the chloride channel defect might impair the MPO-hydrogen peroxide-chloride microbicidal function. Here, we compared the competence of killing PsA by neutrophils from normal donors and CF patients. The data demonstrate that the killing rate by CF neutrophils was significantly lower than that by normal neutrophils. CF neutrophils in a chloride-deficient environment had only one-third of the bactericidal capacity of normal neutrophils in a physiological chloride environment. These results suggest that CFTR-dependent chloride anion transport contributes significantly to killing PsA by normal neutrophils and when defective as in CF, may compromise the ability to clear PsA. 相似文献
19.
David J. Vigerust John A. Lincoln Rod W. Stuart Brian Gnade Doris L. Lefkowitz Stanley S. Lefkowitz Robert C. Allen 《Inflammation》2000,24(3):251-263
It is known that a peroxidase, H2O2, and a halide form a "cytotoxic triad." As a result of the interactions of the components of the triad, reactive oxygen intermediates (ROI) are formed that help to destroy various invading pathogens including Candida. The present study was undertaken to determine if equivalent units of peroxidase induced equivalent levels of macrophage-mediated killing of Candida. Murine peritoneal macrophages were exposed to various concentrations of eosinophil peroxidase (EPO), myeloperoxidase (MPO), and horseradish peroxidase (HRP). Luminol-dependent chemiluminescence studies showed that equivalent units of peroxidase, as determined by oxidation of guaiacol, demonstrated a hierarchical pattern of ROI production. Macrophage phagocytosis and candidicidal activity, as measured by a fluorescence acridine orange assay, also demonstrated the same hierarchical pattern of EPO > MPO > HRP. Therefore, enzymatically equivalent peroxidases do not demonstrate equivalent candidicidal activity. These data indicate a distinct order of peroxidases relative to their ability to stimulate chemiluminescence and macrophage-mediated killing. 相似文献
20.
Myeloperoxidase (MPO) staining has been important for the cytomorphological diagnosis and classification of leukemia. A novel staining method for MPO and its clinical application are presented in the report. Pyronine B (PyB), serving as a chromogenic reagent, was pre-oxidized to obtain stable oxidized Pyronine B solution. The MPO working solution for oxidized pyronine B method consisted of phosphate buffer solution, potassium iodide (KI) solution, and oxidized Pyronine B solution. The positive products of the oxidized Pyronine B method of MPO staining were vibrant red particles located in cytoplasm and the nucleus was stained bluish green. Bone marrow smears from 229 patients with acute leukemia or with grossly normal bone marrow were stained by both oxidized Pyronine B method and the conventional Washburn benzidine staining and a comparison revealed no significant difference in the positive detection rate between the two techniques. The new method eliminates the influence of the varying amount of H2O2 on MPO staining. With this method, the reagents were more stable and the staining procedure was simple and time-saving. This MPO staining technique is a better alternative than the conventional benzidine-based methods. 相似文献