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Oral Diseases 11/2012; · 2.49 Impact Factor
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ABSTRACT: It has been suggested that the very low incidence of atherosclerosis in glycogen storage disease type Ia (GSD Ia) subjects might be attributed to elevated levels of uric acid, one of the potent low molecular- weight antioxidants found in plasma. The present communication describes a use of two analytical methods-cyclic voltammetry and ferric reducing ability of plasma-and also two chemiluminescence methods to evaluate the total oxidant-scavenging capacities (TOSC) of plasma from GSD Ia patients. Our results verified the elevation of TOSC in GSD Ia patients and we propose the inclusion of luminescence and cyclic voltammetry assays as reliable methods for estimating TOSC in a variety of clinical disorders. Our findings with the cyclic voltammetry method add support to the assumption that the elevated uric acid levels might be the main contributor to plasma antioxidant capacity and possible protection against atherosclerosis.
Journal of Inherited Metabolic Disease 09/2009; 32(5):651-9. · 3.58 Impact Factor
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I Ginsburg
Inflammation 03/2001; 25(1):1-6. · 1.75 Impact Factor
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I Ginsburg
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ABSTRACT: The possible role played by streptolysin S (SLS) of group A streptococci in the pathophysiology of streptococcal infections and in post-streptococcal sequelae is discussed. The following properties of SLS justify its definition as a distinct virulence factor: 1) its presence on the streptococcus surface in a cell-bound form, 2) its continuous and prolonged synthesis by resting streptococci, 3) its non-immunogenicity, 4) its extractability by serum proteins (albumin, alpha lipoprotein), 5) its ability to become transferred directly to target cells while being protected from inhibitory agents in the milieu of inflammation, 6) its ability to bore holes in the membrane phospholipids in a large variety of mammalian cells, 7) its ability to synergize with oxidants, proteolytic enzymes, and with additional host-derived proinflammatory agonists, and 8) its absence in streptococcal mutants associated with a lower pathogenicity for animals. Because tissue damage in streptococcal and post-streptococcal sequelae might be the end result of a distinct synergism between streptococcal and host-derived proinflammatory agonists it is proposed that only cocktails of anti-inflammatory agents including distinct inhibitors of SLS (phospholipids), gamma globulin, inhibitors of reactive oxygen species, proteinases, cationic proteins cytokines etc., will be effective in inhibiting the multiple synergistic interactions which lead to fasciitis, myositis and the flesh-eating syndromes, and often develop into sepsis, septic shock and multiple organ failure. The creation of mutants deficient in SLS and in proteases will help shed light on the specific role played by SLS in the virulence of group A hemolytic streptococci.
Apmis 01/2000; 107(12):1051-9. · 1.99 Impact Factor
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ABSTRACT: The purpose of this review-hypothesis is to discuss the literature which had proposed the concept that the mechanisms by which infectious and inflammatory processes induce cell and tissue injury, in vivo, might paradoxically involve a deleterious synergistic 'cross-talk', among microbial- and host-derived pro-inflammatory agonists. This argument is based on studies of the mechanisms of tissue damage caused by catalase-negative group A hemolytic streptococci and also on a large body of evidence describing synergistic interactions among a multiplicity of agonists leading to cell and tissue damage in inflammatory and infectious processes. A very rapid cell damage (necrosis), accompanied by the release of large amounts of arachidonic acid and metabolites, could be induced when subtoxic amounts of oxidants (superoxide, oxidants generated by xanthine-xanthine oxidase, HOCl, NO), synergized with subtoxic amounts of a large series of membrane-perforating agents (streptococcal and other bacterial-derived hemolysins, phospholipases A2 and C, lysophosphatides, cationic proteins, fatty acids, xenobiotics, the attack complex of complement and certain cytokines). Subtoxic amounts of proteinases (elastase, cathepsin G, plasmin, trypsin) very dramatically further enhanced cell damage induced by combinations between oxidants and the membrane perforators. Thus, irrespective of the source of agonists, whether derived from microorganisms or from the hosts, a triad comprised of an oxidant, a membrane perforator, and a proteinase constitutes a potent cytolytic cocktail the activity of which may be further enhanced by certain cytokines. The role played by non-biodegradable microbial cell wall components (lipopolysaccharide, lipoteichoic acid, peptidoglycan) released following polycation- and antibiotic-induced bacteriolysis in the activation of macrophages to release oxidants, cytolytic cytokines and NO is also discussed in relation to the pathophysiology of granulomatous inflammation and sepsis. The recent failures to prevent septic shock by the administration of only single antagonists is disconcerting. It suggests, however, that since tissue damage in post-infectious syndromes is caused by synergistic interactions among a multiplicity of agents, only cocktails of appropriate antagonists, if administered at the early phase of infection and to patients at high risk, might prevent the development of post-infectious syndromes.
FEMS Immunology & Medical Microbiology 10/1999; 25(4):325-38. · 2.44 Impact Factor
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ABSTRACT: The objective of the present communication is to describe the role played by combinations between diethydithiocarbamate (DDC) and divalent metals in hemolysis of human RBC. RBC which had been treated with DDC (10-50 microM) were moderately hemolyzed (about 50%) upon the addition of subtoxic amounts of Cu2+ (50 microM). However, a much stronger and a faster hemolysis occurred either if mixtures of RBC-DDC were immediately treated either by Co2+ (50 microM) or by a premixture of Cu2+ and Co2+ (Cu:Co) (50 microM). While Fe2+ and Ni2+, at 50 microM, initiated 30-50% hemolysis when combined with DDC (50 microM), on a molar basis, Cd2+ was at least 50 fold more efficient than any of the other metals in the initiation of hemolysis by DDC. On the other hand, neither Mn2+ nor Zn2+, had any hemolysis-initiating effects. Co2+ was the only metal which totally blocked hemolysis if added to DDC prior to the addition of the other metals. Hemolysis by mixtures of DDC + (Cu:Co) was strongly inhibited by anaerobiosis (flushing with nitrogen gas), by the reducing agents glutathione, N-acetyl cysteine, mercaptosuccinate, ascorbate, TEMPO, and alpha-tocopherol, by the PLA2 inhibitorbromophenacylbromide (BrPACBr), by tetracycline as well as by phosphatidyl choline, cholesterol and by trypan blue. However, TEMPO, BrPACBr and PC were the only agents which inhibited hemolysis induced by DDC: Cd2+ complexes. On the other hand, none of the classical scavengers of reactive oxygen species (ROS) employed e.g dimethylthiourea, catalase, histidine, mannitol, sodium benzoate, nor the metal chelators desferal and phenanthroline, had any appreciable inhibitory effects on hemolysis induced by DDC + (Cu:Co). DDC oxidized by H2O2 lost its capacity to act in concert either with Cu2+ or with Cd2+ to hemolyze RBC. While either heating RBC to temperatures greater than 37 degrees C or exposure of the cells to glucose-oxidase-generated peroxide diminished their susceptibility to hemolysis, exposure to the peroxyl radical from AAPH, enhanced hemolysis by DDC + (Cu:Co). The cyclovoltammetry patterns of DDC were drastically changed either by Cu2+, Co2+ or by Cd2+ suggesting a strong interaction of the metals with DDC. Also, while the absorbance spectrum of DDC at 280 nm was decreased by 50% either by Co2+, Cd2+ or by H2O2, a 90% reduction in absorbance occurred if DDC + H2O2 mixtures were treated either by Cu2+ or by Co2+, but not by Cd2+. Taken together, it is suggested that DDC-metal chelates can induce hemolysis by affecting the stability and the integrity of the RBC membrane, and possibly also of the cytoskeleton and the role played by reducing agents as inhibitors might be related to their ability to deplete oxygen which is also supported by the inhibitory effects of anaeobiosis.
Free Radical Research 09/1999; 31(2):79-91. · 2.88 Impact Factor
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I Ginsburg
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ABSTRACT: The paper discusses the principal evidence that supports the concept that cell and tissue injury in infectious and post-infectious and inflammatory sequelae might involve a deleterious synergistic interaction among microbial- and host-derived pro-inflammatory agonists. Experimental models had proposed that a rapid cell and tissue injury might be induced by combinations among subtoxic amounts of three major groups of agonists generated both by microorganisms and by the host's own defense systems. These include: (1) oxidants: Superoxide, H(2)O(2), OH', oxidants generated by xanthine-xanthine-oxidase, ROO; HOC1, NO, OONO'-, (2) the membrane-injuring and perforating agents, microbial hemolysins, phospholipases A(2) and C, lysophosphatides, bactericidal cationic proteins, fatty acids, bile salts and the attack complex of complement a, certain xenobics and (3) the highly cationic proteinases, elastase and cathepsin G, as well as collagenase, plasmin, trypsin and a variety of microbial proteinases. Cell killing by combinations among the various agonists also results in the release of membrane-associated arachidonate and metabolites. Cell damage might be further enhanced by certain cytokines either acting directly on targets or through their capacity to prime phagocytes to generate excessive amounts of oxidants. The microbial cell wall components, lipoteichoic acid (LTA), lipopolysaccharides (LPS) and peptidoglycan (PPG), released following bacteriolysis, induced either by cationic proteins from neutrophils and eosinophils or by beta lactam antibiotics, are potent activators of macrophages which can release oxidants, cytolytic cytokines and NO. The microbial cell wall components can also activate the cascades of coagulation, complement and fibrinolysis. All these cascades might further synergize with microbial toxins and metabolites and with phagocyte-derived agonsits to amplify tissue damage and to induce septic shock, multiple organ failure, 'flesh-eating' syndromes, etc. The long persistence of non-biodegradable bacterial cell wall components within activated macrophages in granulomatous inflammation might be the result of the inactivation by oxidants and proteinases of bacterial autolytic wall enzymes (muramidases). The unsuccessful attempts in recent clinical trials to prevent septic shock by the administration of single antagonists is disconcerting. It does suggest however that, since tissue damage in post-infectious syndromes is most probably the end result of synergistic interactions among a multiplicity of agents, only agents which might depress bacteriolysis in vivo and 'cocktails' of appropriate antagonists, but not single antagonists, if administered at the early phases of infection especially to patients at high risk, might help to control the development of post-infectious syndromes. However, the use of adequate predictive markers for sepsis and other post-infectious complications is highly desirable. Although it is conceivable that anti-inflammatory strategies might also be counter-productive as they might act as 'double-edge swords', intensive investigations to devise combination therapies are warranted. The present review also lists the major anti-inflammatory agents and strategies and combinations among them which have been proposed in the last few years for clinical treatments of sepsis and other post-infectious complications.
Inflammopharmacology 02/1999; 7(3):207-17.
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I Ginsburg,
M Sadovnik,
S Sallon,
I Milo-Goldzweig,
R Mechoulam,
A Breuer,
D Gibbs,
J Varani,
S Roberts,
E Cleator,
N Singh
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ABSTRACT: Both aqueous and methanolic fractions derived from the Tibetan preparation PADMA-28 (a mixture of 22 plants) used as an anti-atherosclerotic agent, and which is non-cytolytic to a variety of mammalian cells, were found to strongly inhibit (1) the killing of epithelial cells in culture induced by 'cocktails' comprising oxidants, membrane perforating agents and proteinases; (2) the generation of luminol-dependent chemiluminescence in human neutrophils stimulated by opsonized bacteria; (3) the peroxidation of intralipid (a preparation rich in phopholipids) induced in the presence of copper; and (4) the activity of neutrophil elastase. It is proposed that PADMA-28 might prove beneficial for the prevention of cell damage induced by synergism among pro-inflammatory agonists which is central in the initiation of tissue destruction in inflammatory and infectious conditions.
Inflammopharmacology 02/1999; 7(1):47-62.
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ABSTRACT: An in vitro model was employed to study the potential role of streptococcal extra-cellular products, rich in streptolysin O, in cellular injury as related to streptococcal infections and post-streptococcal sequelae. Extra-cellular products (EXPA) rich in streptolysin O were isolated from type 4, group A hemolytic streptococci grown in a chemostat, in a synthetic medium. EXPA induced moderate cytopathogenic changes in monkey kidney epithelial cells and in rat heart cells pre-labeled with 3H-arachidonate. However very strong toxic effects were induced when EXP was combined with oxidants (glucose oxides generated H2O2, AAPH-induced peroxyl radical (ROO.), NO generated by sodium nitroprusside) and proteinases (plasmin, trypsin). Cell killing was distinctly synergistic in nature. Cell damage induced by the multi-component cocktails was strongly inhibited either by micromolar amounts of gamma globulin, and Evan's blue which neutralized SLO activity, by tetracycline, trasylol (aprotinin), epsilon amino caproic acid and by soybean trypsin inhibitor, all proteinase inhibitors as well as by a non-penetrating PLA2 inhibitor A. The results suggest that fasciitis, myositis and sepsis resulting from infections with hemolytic streptococci might be caused by a coordinated 'cross-talk' among microbial, leukocyte and additional host-derived pro-inflammatory agents. Since attempts to prolong lives of septic patients by the exclusive administration of single antagonists invariably failed, it is proposed that the administration of 'cocktails' of putative inhibitors against major pro-inflammatory agonizes generated in inflammation and infection might protect against the deleterious effects caused by the biochemical and pharmacological cascades which are known to be activated in sepsis.
FEMS Immunology & Medical Microbiology 12/1998; 22(3):247-56. · 2.44 Impact Factor
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I Ginsburg
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ABSTRACT: The mechanisms of cellular damage caused by infectious and inflammatory processes are complex and are still not fully understood. There is, however, a consensus that reactive oxygen species (ROS) generated by phagocytes migrating to injured tissues might be the main agents responsible for cellular damage in inflammatory processes. However, because both activated phagocytes and catalase-negative, peroxide-producing, toxigenic bacteria (Streptococci, Clostridiae) secrete a near-identical array of proinflammatory agonists, including reactive oxygen species (ROS), and because these microbial species might kill their targets by a synergism among several of their secreted enzymes (a multicomponent system), we postulated that activated phagocytes might also function in the same way. Using radiolabeled targets, in culture, we demonstrated that subtoxic amounts of a variety of oxidants (H2O2, radicals produced by xanthine-xanthine-oxidase, peroxyl radical, NO) acted synergistically with subtoxic amounts of a large series of membrane-perforating agents (microbial hemolysins, phospholipases, fatty acids, cationic proteins, proteinases, bile salts, the attack complex of complement, the xenobiotics, lindane, ethanol, methanol) to kill cells in culture and to release large amounts of arachidonic acid and metabolites. Membrane perforators might act primarily to overcome the potent antioxidant systems present in all mammalian cells and scavengers of ROS and inhibitors of the additional agonists might act to abolish the synergism among ROS and the membrane-damaging agents. It is also proposed that protection against tissue damage in vivo should also include 'cocktails' of appropriate antagonists. It is enigmatic that those publications which do describe both in-vitro and in-vivo models proposing that a synergism among a multiplicity of agonists might truly represent the mechanisms by which tissues are injured, in vivo, are hardly ever quoted in the current literature.
Medical Hypotheses 11/1998; 51(4):337-46. · 1.39 Impact Factor
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ABSTRACT: The effect of diethyldithiocarbamate (DDC) and sodium nitroprusside (SNP) on the killing of endothelial cells and on the release of arachidonate by mixtures of oxidants and membrane-damaging agents was studied in a tissue culture model employing bovine aortic endothelial cells labeled either with 51Chromium or 3arachidonic acid. While exposure to low, subtoxic concentrations of oxidants (reagent H2O2, glucose-oxidase generated peroxide, xanthine xanthine oxidase, AAPH-generated peroxyl radical, menadione-generated oxidants) did not result either in cell death or in the loss of membrane-associated arachidonic acid, the addition of subtoxic amounts of a variety of membrane-damaging agents (streptolysin S, PLA2, histone, taurocholate, wheatgerm agglutinin) resulted in a synergistic cell death. However, no significant amounts of arachidonate were released unless proteinases were also present. The addition to these reaction mixtures of subtoxic amounts of DDC (an SOD inhibitor and a copper chelator) not only very markedly enhanced cell death but also resulted in the release of large amounts of arachidonate (in the complete absence of added proteinases). Furthermore, the inclusion in DDC-containing reaction mixtures of subtoxic amounts of SNP, a generator of NO, further enhanced, in a synergistic manner, both cell killing and the release of arachidonate. Cell killing and the release of arachidonate induced by the DDC and SNP-containing mixtures of agonists were strongly inhibited by catalase, glutathione, N-acetyl cysteine, vitamin A, and by a nonpenetrating PLA2 inhibitor as well as by tetracyclines. A partial inhibition of cell killing was also obtained by 1,10-phenanthroline and by antimycin. It is suggested that DDC might amplify cell damage by forming intracellular, loosely-bound complexes with copper and probably also by depleting antioxidant thiols. It is also suggested that "cocktails" containing oxidants, membrane-damaging agents, DDC, and SNP might be beneficial for killing of tumor cells in vivo and for the assessment of the toxicity of xenobiotics in vitro.
Free Radical Research 09/1997; 27(2):143-64. · 2.88 Impact Factor
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ABSTRACT: Phospholipase A2 (PLA2) and H2O2, secreted from activated inflammatory cells, play a central role in the tissue damage occurring in inflammatory processes. However, while exogenous PLA2 alone does not cause cell lysis, it readily does so when acting with H2O2. We have found that H2O2 degrades cell surface proteoglycans, thus rendering the membrane PL accessible to hydrolysis by exogenous PLA2. This novel mechanism introduces a role for cell surface proteoglycans in protection of cells from damage by pro-inflammatory agents, and may assign a central role for the combined action of H2O2 and PLA2 in inflammatory and bacteriocidal processes.
FEBS Letters 04/1996; 383(1-2):75-8. · 3.54 Impact Factor
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ABSTRACT: Oxygen-derived radicals are implicated in the pathogenesis of tissue damage and ulcerogenesis. This study aimed to examine the effect of manganese, glycine, and carotene, oxygen radical scavengers, on ethanol-induced gastric lesions in the rat and on ethanol cytotoxicity in epithelial cell culture.
MnCl2 + glycine (12.5-50 mg/rat) were injected subcutaneously up to 6 h before oral administration of 1 ml of 96% ethanol, and 0.5 ml carrot juice or beta-carotene was given orally 30 min before the ethanol. Mucosal injury was evaluated 1 h later by gross and microscopic scoring. The effect of Mn2+ and carrot juice was also tested in monolayers of radiolabeled epithelial cells exposed to H2O2 + ethanol injury as expressed by the extent of the isotope leakage.
Mn2+ and glycine pretreatment dose-dependently reduced ethanol-induced gastric lesion formation. Protection was maximal when treatment was applied 4 h before the insult. Gross damage was also markedly prevented by pretreatment with carotenes and dimethylthiourea (DMTU, 75 mg/kg intraperitoneally) but not by allopurinol. Mixtures of subtoxic concentrations of ethanol and H2O2 were highly lethal for epithelial cell monolayers. In this model, cell death was markedly attenuated by catalase, DMTU, Mn2+, and carrot juice.
Ethanol-induced gastric mucosal damage may involve generation of oxygen-derived radicals, independent of the xanthine oxidase system. By acting as oxygen radical scavengers, Mn2+, glycine, and carotenes, like catalase and DMTU, provide significant gastroprotection.
Scandinavian Journal of Gastroenterology 10/1995; 30(9):854-60. · 2.02 Impact Factor
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Free Radical Research 07/1995; 22(6):489-517. · 2.88 Impact Factor
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ABSTRACT: The assumption that cellular injury induced in infectious and in inflammatory sites might be the result of a well-orchestrated, synergistic "cross-talk" among oxidants, membrane-damaging agents, proteinases, and xenobiotics was further investigated in a tissue culture model employing monkey kidney epithelial cells (BGM) labeled either with 51 chromium or [3H]arachidonate. The cells could be killed in a synergistic manner following exposure to combinations among H2O2 and the following membrane-damaging agents: streptolysins S (SLS) and O (SLO), poly-D-lysine, arachidonic acid, eicosapentanoic acid, arachidic acid, lysophosphatidylcholine, lysophosphatidylinositol, lysophosphatidylglycerol, ethanol, and sodium taurocholate. Peroxyl radical (ROO) generated by azobisdiamidinopropane dihydrochloride (AAPH) further enhanced cell killing induced by SLS, SLO, and nitroprusside when combined with H2O2 and trypsin. BGM cells labeled either with chromium or with tritiated arachidonate, which had been treated with increasing concentrations of sodium nitroprusside (a donor of NO) and with subtoxic amounts of SLS and H2O2, were also killed in a synergistic manner and also lost a substantial amounts of their arachidonate label. Both cell killing and the release of membrane lipids were totally inhibited by hemoglobin (an NO scavenger) but not by methylene blue, an antagonist of NO2-BGM cells that had been treated with increasing concentrations of taurocholic acid were killed in a synergistic manner by a mixture of subtoxic amounts of ethanol, H2O2, and crystalline trypsin (quadruple synergism). Normal human serum possessing IgM complement-dependent cytotoxic antibodies against Ehrlich ascites tumor cells were killed in a dose-dependent fashion. Cell killing was doubled by the addition of H2O2. Cell killing and the release of membrane lipids by all the mixture of agonists tested were both strongly inhibited by the antioxidants catalase, Mn2+, vitamin A, and by fresh carrot juice. It appears that in order to overcome the antioxidant capacities of the epithelial cells, a variety of membrane-damaging agents had to be present in the reaction mixtures. Taken together, it might be speculated that the killing of mammalian cells in infectious and in inflammatory sites is a synergistic phenomenon that might be inhibited by antagonizing the cross-talk among the various proinflammatory agonists generated by microorganisms by activated phagocytes or by combinations among these agents. Our studies might also open up new approaches to the assessment of the toxicity of xenobiotics and of safe drugs to mammalian cells by employing tissue culture techniques.
Inflammation 03/1995; 19(1):101-18. · 1.75 Impact Factor
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ABSTRACT: Cell-impermeable inhibitors of phospholipase A2 were prepared by linking inhibiting molecules to macromolecular carriers which prevent the inhibitor's internalization. These preparations inhibit the release of oxygen reactive species from neutrophils and cell death induced by inflammatory agents, as well as bleomycin-induced lung injury.
Agents and actions. Supplements 02/1995; 46:77-84.
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I Ginsburg
Comparative biochemistry and physiology. Part C, Pharmacology, toxicology & endocrinology 11/1994; 109(2):147-58.
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ABSTRACT: Monkey kidney epithelial cells, labeled with chromium and grown in culture, were killed in a synergistic manner when subtoxic amounts of ethanol were combined either with subtoxic amounts of glucose oxidase-generated hydrogen peroxide, or with mixtures of peroxide and with 2,2'-Azo-bis (2-amidinopropane)HCl (AAPH)-generated peroxyl radical. A further enhancement of cytotoxicity occurred when subtoxic amounts of trypsin were added to mixtures of all three agents. While ethanol alone caused shrinkage of the monolayers and cell rounding, no visible cytotoxic changes were observed. Hydrogen peroxide at the concentrations used (about 1 mM), caused only some cell rounding. On the other hand, cells exposed simultaneously to ethanol and to H2O2 developed extensive membrane damage characterized by the formation of large polar blebs, which is compatible with altered membrane permeability. The presence of trypsin markedly enhanced cellular cytotoxicity induced by mixtures of peroxide, peroxyl radical, and ethanol. This could markedly be depressed by catalase and by dimethylthiourea. The tissue culture model described might serve to further investigate the role played by synergy among oxidants and a variety of membrane-damaging agents, and by xenobiotics in tissue damage induced by inflammatory processes.
Free Radical Biology and Medicine 03/1994; 16(2):263-9. · 5.42 Impact Factor
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ABSTRACT: Human neutrophils (PMNs) suspended in Hanks' balanced salt solution (HBSS), which are stimulated either by polycation-opsonized streptococci or by phorbol myristate acetate (PMA), generate nonamplified (CL), luminol-dependent (LDCL), and lucigenin-dependent chemiluminescence (LUCDCL). Treatment of activated PMNs with azide yielded a very intense CL response, but only a small LDCL or LUCDCL responses, when horse radish peroxidase (HRP) was added. Both CL and LDCL depend on the generation of superoxide and on myeloperoxidase (MPO). Treatment of PMNs with azide followed either by dimethylthiourea (DMTU), deferoxamine, EDTA, or detapac generated very little CL upon addition of HRP, suggesting that CL is the result of the interaction among H2O2, a peroxidase, and trace metals. In a cell-free system practically no CL was generated when H2O2 was mixed with HRP in distilled water (DW). On the other hand significant CL was generated when either HBSS or RPMI media was employed. In both cases CL was markedly depressed either by deferoxamine or by EDTA, suggesting that these media might be contaminated by trace metals, which catalyzed a Fenton-driven reaction. Both HEPES and Tris buffers, when added to DW, failed to support significant HRP-induced CL. Nitrilotriacetate (NTA) chelates of Mn2+, Fe2+, Cu2+, and Co2+ very markedly enhanced CL induced by mixtures of H2O2 and HRP when distilled water was the supporting medium. Both HEPES and Tris buffer when added to DW strongly quenced NTA-metal-catalyzed CL. None of the NTA-metal chelates could boost CL generation by activated PMNs, because the salts in HBSS and RPMI interfered with the activity of the added metals. CL and LDCL of activated PMNs was enhanced by aminotriazole, but strongly inhibited by diphenylene iodonium (an inhibitor of NADPH oxidase) by azide, sodium cyanide (CN), cimetidine, histidine, benzoate, DMTU and moderately by superoxide dismutase (SOD) and by deferoxamine LUCDCL was markedly inhibited only by SOD but was boosted by CN. Taken together, it is suggested that CL generated by stimulated PMNs might be the result of the interactions among, NADPH oxidase, (inhibitable by diphenylene iodonium), MPO (inhibitable by sodium azide), H2O2 probably of intracellular origin (inhibitable by DMTU but not by catalase), and trace metals that contaminate salt solutions. The nature of the salt solutions employed to measure CL in activated PMNs is critical.
Inflammation 07/1993; 17(3):227-43. · 1.75 Impact Factor
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ABSTRACT: 51Chromium-labeled rat pulmonary artery endothelial cells (EC) cultivated in MEM medium were killed, in a synergistic manner, by mixtures of subtoxic amounts of glucose oxidase-generated H2O2 and subtoxic amounts of the following agents: the cationic substances, nuclear histone, defensins, lysozyme, poly-L-arginine, spermine, pancreatic ribonuclease, polymyxin B, chlorhexidine, cetyltrimethyl ammonium bromide, as well as by the membrane-damaging agents phospholipases A2 (PLA2) and C (PLC), lysolecithin (LL), and by streptolysin S (SLS) of group A streptococci. Cytotoxicity induced by such mixtures was further enhanced by subtoxic amounts either of trypsin or of elastase. Glucose-oxidase cationized by complexing to poly-L-histidine proved an excellent deliverer of membrane-directed H2O2 capable of enhancing EC killing by other agonists. EC treated with rabbit anti-streptococcal IgG were also killed, in a synergistic manner, by H2O2, suggesting the presence in the IgG preparation of cross-reactive antibodies. Killing of EC by the various mixtures of agonists was strongly inhibited by scavengers of hydrogen peroxide (catalase, dimethylthiourea, MnCl2), by soybean trypsin inhibitor, by polyanions, as well as by putative inhibitors of phospholipases. Strong inhibition of cell killing was also observed with tannic acid and by extracts of tea, but less so by serum. On the other hand, neither deferoxamine, HClO, TNF, nor GTP gamma S had any modulating effects on the synergistic cell killing. EC exposed either to 6-deoxyglucose, puromycin, or triflupromazin became highly susceptible to killing by mixtures of hydrogen peroxide with several of the membrane-damaging agents. While maximal synergistic EC killing was achieved by mixtures of H2O2 with either PLA2, PLC, LL, or with SLS, a very substantial release of [3H]arachidonic acid (AA), PGE2, and 6-keto-PGF occurred only if a proteinase was also added to the mixture of agonists. The release of AA from EC was markedly inhibited either by scavengers of H2O2, by proteinase inhibitors, by cationic agents, by HClO, by tannic acid, and by quinacrin. We suggest that cellular injury induced in inflammatory and infectious sites might be the result of synergistic effects among leukocyte-derived oxidants, lysosomal hydrolases, cytotoxic cationic polypeptides, proteinases, and microbial toxins, which might be present in exudates. These "cocktails" not only kill cells, but also solubilize AA and several of its metabolites. However, AA release by the various agonists can be also achieved following attack by leukocyte-derived agonists on dead cells. It is proposed that treatment by "cocktails" of adequate antagonists might be beneficial to protect against cellular injury in vivo.
Inflammation 07/1993; 17(3):295-319. · 1.75 Impact Factor
