scholarly journals Production of the superoxide adduct of myeloperoxidase (compound III) by stimulated human neutrophils and its reactivity with hydrogen peroxide and chloride

1985 ◽  
Vol 228 (3) ◽  
pp. 583-592 ◽  
Author(s):  
C C Winterbourn ◽  
R C Garcia ◽  
A W Segal
Keyword(s):  
Hydrogen Peroxide ◽  
Superoxide Dismutase ◽  
Phorbol Myristate Acetate ◽  
Catalase Activity ◽  
Human Neutrophils ◽  
H2o2 Concentration ◽  
Myristate Acetate ◽  
Similar Product ◽  
Form Compound ◽  
Compound Ii

Examination of the spectra of phagocytosing neutrophils and of myeloperoxidase present in the medium of neutrophils stimulated with phorbol myristate acetate has shown that superoxide generated by the cells converts both intravacuolar and exogenous myeloperoxidase into the superoxo-ferric or oxyferrous form (compound III or MPO2). A similar product was observed with myeloperoxidase in the presence of hypoxanthine, xanthine oxidase and Cl-. Both transformations were inhibited by superoxide dismutase. Thus it appears that myeloperoxidase in the neutrophil must function predominantly as this superoxide derivative. MPO2 autoxidized slowly (t 1/2 = 12 min at 25 degrees C) to the ferric enzyme. It did not react directly with H2O2 or Cl-, but did react with compound II (MP2+ X H2O2). MPO2 catalysed hypochlorite formation from H2O2 and Cl- at approximately the same rate as the ferric enzyme, and both reactions showed the same H2O2-dependence. This suggests that MPO2 can enter the main peroxidation pathway, possibly via its reaction with compound II. Both ferric myeloperoxidase and MPO2 showed catalase activity, in the presence or absence of Cl-, which predominated over chlorination at H2O2 concentrations above 200 microM. Thus, although the reaction of neutrophil myeloperoxidase with superoxide does not appear to impair its chlorinating ability, the H2O2 concentration in its environment will determine whether the enzyme acts primarily as a catalase or peroxidase.

Factors which affect DNA strand breakage in human leukocytes exposed to a tumor promoter, phorbol myristate acetate

1982 ◽  
Vol 60 (11) ◽  
pp. 1359-1366 ◽  
Author(s):  
H. C. Birnboim
Keyword(s):  
Hydrogen Peroxide ◽  
Dna Damage ◽  
Phorbol Myristate Acetate ◽  
Respiratory Burst ◽  
Superoxide Anion ◽  
Tumor Promoter ◽  
Myristate Acetate ◽  
Strand Breakage ◽  
Dna Strand ◽  
Dna Strand Breakage

We have recently reported that phorbol myristate acetate (PMA) induces extensive DNA strand break damage in human peripheral blood leukocytes. The mechanism of action involves superoxide anion and hydrogen peroxide which are generated by phagocytes during the "respiratory burst." In this report, we describe the effect of various inhibitors and scavengers on PMA-induced DNA damage. Azide and cyanide greatly increased the level of damage; sulfhydryl compounds (glutathione, cysteine, and cysteamine) and ascorbate markedly decreased the level of damage. Hydroxyl radical scavengers such as dimethyl sulfoxide (DMSO) and glycerol also decreased the level of damage but apparently did so by inhibiting the respiratory burst. Diethyldithiocarbamate (DDC) increased the level of DNA damage at low concentrations (<1 mM), but decreased DNA damage at ≥1 mM. The results are consistent with a mechanism involving superoxide anion and hydrogen peroxide, but the precise reaction (free radical or enzymatic) responsible for DNA strand breakage has not been determined. The PMA-stimulated phagocyte is an interesting model system for looking at "active oxygen" mediated DNA damage and factors which influence it.

scholarly journals Regulation of CD18 expression in human neutrophils as related to shape changes

1993 ◽  
Vol 106 (2) ◽  
pp. 493-501
Author(s):  
A. Volz
Keyword(s):  
Phorbol Myristate Acetate ◽  
Surface Expression ◽  
Cytochalasin D ◽  
Human Neutrophils ◽  
Chemotactic Peptide ◽  
Myristate Acetate ◽  
Surface Distribution ◽  
Quantitative Expression ◽  
High Concentrations ◽  
Shape Changes

The study analyses the distribution and quantitative expression of surface CD18 of neutrophils exposed to distinct stimuli that produce different types of continuous shape changes, including types that are associated with locomotion and others that are not. The chemotactic peptide N-formyl-L-norleucyl-L-leucyl-L-phenylalanine, colchicine and nocodazole were used to induce a polarized locomotor morphology, phorbol myristate acetate, 1,2-dioctanoylglycerol and 1-oleoyl-2-acetyl-glycerol to induce non-polar motile cells ruffling all over the surface and 2H2O to induce non-polar cells performing circus movements as have been previously described. Except for colchicine and nocodazole, these stimuli increased surface expression of CD18. Thus, stimulated shape changes are frequently, though not always, associated with increased surface expression of CD18. High concentrations (10(−7) to 10(−5) M) of phorbol myristate acetate but not of chemotactic peptide induced down-regulation of surface CD18. Cytochalasin D (10(−4) M) stimulated CD18 expression even though it inhibited shape changes. The surface distribution of CD18 determined by light microscopy was uniform in unstimulated cells or in various forms of stimulation except for cells treated with 10(−5) M cytochalasin D. Cytochalasin D (10(−5) M) produced CD18 accumulation at the pole opposite the F-actin cap. Experiments with colchicine, nocodazole, 2H2O and cytochalasin D suggest that microtubules as well as microfilaments modulate surface expression of CD18. The results suggest that protein kinase C and phosphatases play a role in regulating surface expression of CD18 in neutrophils.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals Evidence for production of oxidizing radicals by the particulate O-2- forming system from human neutrophils

Blood ◽  
1979 ◽  
Vol 53 (4) ◽  
pp. 666-676
Author(s):  
AI Tauber ◽  
TG Gabig ◽  
BM Babior
Keyword(s):  
Hydrogen Peroxide ◽  
Superoxide Dismutase ◽  
Ethylene Production ◽  
Human Neutrophils ◽  
Peroxy Acid ◽  
Radical Scavengers ◽  
Alkyl Peroxide ◽  
Acyl Peroxide

The particulate O-2-forming system from human neutrophils was found to oxidize methional and 2-keto-4-methylthiobutyric acid (KMB) to ethylene, indicating the formation by this system of strongly oxidizing radicals. Conforming this interpretation was the observation that ethylene production was inhibited by the radical scavengers benzoate, ethanol, and mannitol. Ethylene production was also sharply reduced by superoxide dismutase, implicatin O-2 as a precursor of oxidizing radicals. In our system catalase only partially inhibited ethylene generation from either methional or KMB, suggesting that oxidizing radicals are generated at least in part by the reacton of O-2 with compounds other than H2O2. We propose that in neutrophils oxidizing radicals are formed in a reaction between O-2 and a peroxide according to the following equation: O-2 + ROOH leads to RO . + OH- + O2, in which ROOH may be hydrogen peroxide, an alkyl peroxide, or an acyl peroxide (i.e., a peroxy acid).

Lecithinized superoxide dismutase attenuates phorbol myristate acetate-induced injury in isolated dog lung

1998 ◽  
Vol 344 (2-3) ◽  
pp. 231-239 ◽  
Author(s):  
Takashige Miyahara ◽  
Toshishige Shibamoto ◽  
Hong-Gang Wang ◽  
Tomonobu Koizumi ◽  
Takayuki Honda ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Phorbol Myristate Acetate ◽  
Myristate Acetate

scholarly journals The chlorinating potential of the human monocyte

Blood ◽  
1983 ◽  
Vol 62 (3) ◽  
pp. 645-651 ◽  
Author(s):  
MB Lampert ◽  
SJ Weiss
Keyword(s):  
Phorbol Myristate Acetate ◽  
Cell Concentration ◽  
Human Monocyte ◽  
Human Neutrophils ◽  
Human Monocytes ◽  
Taurine Chloramine ◽  
Myristate Acetate ◽  
Opsonized Zymosan ◽  
Nitrobenzoic Acid ◽  
Characteristic Absorption Peak

Abstract Human monocytes incubated with phorbol myristate acetate (PMA) or opsonized zymosan particles can chlorinate the beta-amino acid taurine to its monochloramine derivative. Taurine monochloramine can then be quantitated by its ability to oxidize 5-thio-2-nitrobenzoic acid to its disulfide or by its characteristic absorption peak at 252 nm. Stimulated, but not resting, monocytes chlorinated taurine by a process dependent on time, cell concentration, and pH. The formation of taurine chloramine by stimulated monocytes could be inhibited by catalase, azide, or cyanide, was unaffected by superoxide dismutase, and was stimulated by exogenous myeloperoxidase. Thus, taurine chloramine generation by human monocytes appeared dependent on both H2O2 and myeloperoxidase. Compared to human neutrophils, the monocyte could generate similar amounts of chloramine when stimulated with phorbol myristate acetate, but far less if opsonized zymosan particles were used as the trigger. Based on the known ability of the H2O2- myeloperoxidase-Cl- system to generate free HOCl, it would seem that this oxidant is the most likely species responsible for the monocyte- mediated chlorination reactions. Thus, we have used a simple quantitative assay to demonstrate the ability of the human monocyte to generate large quantities of a highly reactive and toxic oxygen metabolite.

scholarly journals Superoxide modulates the activity of myeloperoxidase and optimizes the production of hypochlorous acid

1988 ◽  
Vol 252 (2) ◽  
pp. 529-536 ◽  
Author(s):  
A J Kettle ◽  
C C Winterbourn
Keyword(s):  
Superoxide Dismutase ◽  
Xanthine Oxidase ◽  
Hypochlorous Acid ◽  
Direct Reaction ◽  
Chlorination Reaction ◽  
Inactive Compound ◽  
Physiological Conditions ◽  
Oxidant Damage ◽  
Form Compound ◽  
Compound Ii

Myeloperoxidase catalyses the conversion of H2O2 and Cl- to hypochlorous acid (HOCl). It also reacts with O2- to form the oxy adduct (compound III). To determine how O2- affects the formation of HOCl, chlorination of monochlorodimedon by myeloperoxidase was investigated using xanthine oxidase and hypoxanthine as a source of O2- and H2O2. Myeloperoxidase was mostly converted to compound III, and H2O2 was essential for chlorination. At pH 5.4, superoxide dismutase (SOD) enhanced chlorination and prevented formation of compound III. However, at pH 7.8, SOD inhibited chlorination and promoted formation of the ferrous peroxide adduct (compound II) instead of compound III. We present spectral evidence for a direct reaction between compound III and H2O2 to form compound II, and for the reduction of compound II by O2- to regenerate native myeloperoxidase. These reactions enable compound III and compound II to participate in the chlorination reaction. Myeloperoxidase catalytically inhibited O2- –dependent reduction of Nitro Blue Tetrazolium. This inhibition is explained by myeloperoxidase undergoing a cycle of reactions with O2-, H2O2 and O2-, with compounds III and II as intermediates, i.e., by myeloperoxidase acting as a combined SOD/catalase enzyme. By preventing the accumulation of inactive compound II, O2- enhances the activity of myeloperoxidase. We propose that, under physiological conditions, this optimizes the production of HOCl and may potentiate oxidant damage by stimulated neutrophils.

Contrasting effects of inflammatory stimuli on neutrophil and monocyte adherence to endothelial cells

1989 ◽  
Vol 67 (2) ◽  
pp. 556-562 ◽  
Author(s):  
D. W. Kamp ◽  
K. D. Bauer ◽  
A. Knap ◽  
M. M. Dunn
Keyword(s):  
Hydrogen Peroxide ◽  
Endothelial Cells ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Myristate Acetate ◽  
Superoxide Anion ◽  
Leukocyte Adherence ◽  
Myristate Acetate ◽  
Kinase C ◽  
Monocyte Adherence

Leukocyte adherence to endothelial cells (EC) is an important early event in inflammatory responses, which are often characterized by a predominance of either neutrophils (PMN) or monocytes. However, there is little information concerning the molecular events important in leukocyte adherence to EC. Intracellular activation of protein kinase C and the calcium-second messenger system leads to the stimulation of a number of important functions in PMN and monocytes. We compared the effects of members of these pathways on human PMN and monocyte adherence to cultured bovine aortic EC. We observed that phorbol myristate acetate, phorbol, 12,13-dibutyrate, L-alpha-1-oleoyl-2-acetoyl-sn-3-glycerol, and ionomycin each induced significant dose-dependent increases in PMN adherence to EC monolayers. In contrast, similar concentrations of each of these agents induced significant decreases in EC adherence of monocytes enriched by countercurrent centrifugal elutriation. Separate experiments determined that the differences in PMN and monocyte adherence to EC were not related to differences in oxidant production because 1) phorbol myristate acetate and L-alpha-1-oleoyl-2-acetoyl-sn-3-glycerol caused similar marked increases in both PMN and monocyte superoxide anion and hydrogen peroxide production and 2) ionomycin, which had opposing effects on PMN and monocyte adherence, had no effect on PMN and monocyte superoxide anion or hydrogen peroxide release. We conclude that activators of protein kinase C and the Ca-second messenger pathway have opposite effects on PMN and monocyte adherence to EC and that these effects are mediated by O2 radical-independent mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)

Comparison of 1-oleoyl-2-acetyl-glycerol and phorbol myristate acetate as secretagogues for human neutrophils

1986 ◽  
Vol 64 (8) ◽  
pp. 1149-1152 ◽  
Author(s):  
Kenneth Wong ◽  
Christine Chew
Keyword(s):  
Phorbol Myristate Acetate ◽  
Cellular Responses ◽  
Current Evidence ◽  
Human Neutrophils ◽  
Myristate Acetate ◽  
First Order ◽  
Regulatory Constraints ◽  
Kinase C ◽  
Rate Of Decay ◽  
The Stability

Cellular responses induced in human neutrophils by the synthetic diacylglycerol, 1-oleoyl-2-acetyl-glycerol (OAG), paralleled those induced by phorbol myristate acetate (PMA). Like PMA, OAG caused the preferential release of enzymes from specific granules and promoted superoxide (O2−) generation. The efficacy of OAG was similar to that for PMA, but its potency was lower by four orders of magnitude. First derivative kinetic analysis showed that rates of O2− generation elicited by PMA decayed exponentially in a first order manner; the half life was found to be 21 ± 6 min. Results obtained in studies carried out with high OAG concentrations were similar except that after 40 min, the rate of decay increased and became complex order. This difference was attributed to the greater susceptibility of OAG to metabolic alteration, and was reflected in the NADPH oxidase activity of granule rich membrane fractions (GRF) of cells stimulated for 90 min with PMA or OAG. It was found that the O2− generating activity of the PMA treated GRF was significantly greater than that for the OAG treated fraction. Current evidence indicates that cellular responses arise from direct activation of protein kinase C by PMA-OAG. The stability of this complex and the bypassing of normal regulatory constraints may account for the relative longevity of the PMA–OAG O2− respiratory burst.

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