Oxidant membrane injury by the neutrophil myeloperoxidase system. II. Injury by stimulated neutrophils and protection by lipid-soluble antioxidants

S. M. Sepe, Robert A Clark

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43 Citations (Scopus)

Abstract

The stimulated human neutrophil can damage a variety of target cells, and in some models, a mechanism involving secretion of myeloperoxidase and H2O2 has been demonstrated. We explored the characteristics of this cell-cell interaction by using neutrophils and our recently described liposome model target cell system. Exposure of 51Cr-labeled liposomes to phorbol myristate acetate-stimulated human neutrophils resulted in release of 25 to 30% of the radioactivity. 51Cr release was abrogated by omission of the neutrophils, the phorbol ester on halide (iodide), replacement of the phorbol by an inactive congener, or addition of azide, cyanide, or catalase. Neutrophils from patients with hereditary absence of myeloperoxidase (MPO) or a failure of H2O2 formation (chronic granulomatous disease) did not cause liposome lysis unless purified MPO or a source of H2O2, respectively, was added. These data indicate that 51Cr release from liposomes is a consequence of the secretion of MPO and H2O2, which combine with extracellular halides to form a membrane lytic system. The influence of liposome composition on injury was then examined, with a focus on physiologically relevant lipid soluble antioxidants. Liposomes containing either α-tocopherol (0.33 to 1.67% of molar fraction of lipid) or β-carotene (1.67% of molar fraction of lipid) were markedly resistant to lysis by the cellfree MPO-H2O2-chloride system. When the major structural lipid phosphatidyl choline was replaced by dipalmitoyl phosphatidyl choline, a synthetic phospholipid with no oxidizable double bonds, the resultant liposomes were totally resistant to lysis by the MPO-H2O2-chloride system. The addition of iodide to this system (i.e., both chloride and iodide present) changed the pattern of protection dramatically in that α-tocopherol and β-carotene were no longer protective and the resistance of dipalmitoyl phosphatidyl choline liposomes was partial rather than complete. In contrast to iodide, the addition of bromide or thiocyanate did not have a major effect on the protection by antioxidants. Finally, we demonstrated protection by α-tocopherol or dipalmitoyl phosphatidyl choline against liposome lysis by phorbol-activated neutrophils. These studies illustrate the use of model phospholipid membranes in the characterization of oxygen-dependent cell-mediated cytotoxicity. Activated neutrophils lyse liposome targets through a MPO-dependent mechanism. Target properties, especially the content of lipid-soluble antioxidants, have a marked influence on susceptibility to lysis. These effects are in turn subject to modulation on the basis of such factors as the availability of various halide cofactors. This model system offers an opportunity to dissect the biochemical basis of effector cell-target cell interactions.

Original languageEnglish (US)
Pages (from-to)1896-1901
Number of pages6
JournalJournal of Immunology
Volume134
Issue number3
StatePublished - 1985
Externally publishedYes

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Oxidants
Liposomes
Peroxidase
Neutrophils
Antioxidants
Lipids
Membranes
Wounds and Injuries
Iodides
Phosphatidylcholines
Tocopherols
Chlorides
Carotenoids
Cell Communication
Phospholipids
Chronic Granulomatous Disease
Azides
Cyanides
Phorbol Esters
Tetradecanoylphorbol Acetate

ASJC Scopus subject areas

  • Immunology

Cite this

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title = "Oxidant membrane injury by the neutrophil myeloperoxidase system. II. Injury by stimulated neutrophils and protection by lipid-soluble antioxidants",
abstract = "The stimulated human neutrophil can damage a variety of target cells, and in some models, a mechanism involving secretion of myeloperoxidase and H2O2 has been demonstrated. We explored the characteristics of this cell-cell interaction by using neutrophils and our recently described liposome model target cell system. Exposure of 51Cr-labeled liposomes to phorbol myristate acetate-stimulated human neutrophils resulted in release of 25 to 30{\%} of the radioactivity. 51Cr release was abrogated by omission of the neutrophils, the phorbol ester on halide (iodide), replacement of the phorbol by an inactive congener, or addition of azide, cyanide, or catalase. Neutrophils from patients with hereditary absence of myeloperoxidase (MPO) or a failure of H2O2 formation (chronic granulomatous disease) did not cause liposome lysis unless purified MPO or a source of H2O2, respectively, was added. These data indicate that 51Cr release from liposomes is a consequence of the secretion of MPO and H2O2, which combine with extracellular halides to form a membrane lytic system. The influence of liposome composition on injury was then examined, with a focus on physiologically relevant lipid soluble antioxidants. Liposomes containing either α-tocopherol (0.33 to 1.67{\%} of molar fraction of lipid) or β-carotene (1.67{\%} of molar fraction of lipid) were markedly resistant to lysis by the cellfree MPO-H2O2-chloride system. When the major structural lipid phosphatidyl choline was replaced by dipalmitoyl phosphatidyl choline, a synthetic phospholipid with no oxidizable double bonds, the resultant liposomes were totally resistant to lysis by the MPO-H2O2-chloride system. The addition of iodide to this system (i.e., both chloride and iodide present) changed the pattern of protection dramatically in that α-tocopherol and β-carotene were no longer protective and the resistance of dipalmitoyl phosphatidyl choline liposomes was partial rather than complete. In contrast to iodide, the addition of bromide or thiocyanate did not have a major effect on the protection by antioxidants. Finally, we demonstrated protection by α-tocopherol or dipalmitoyl phosphatidyl choline against liposome lysis by phorbol-activated neutrophils. These studies illustrate the use of model phospholipid membranes in the characterization of oxygen-dependent cell-mediated cytotoxicity. Activated neutrophils lyse liposome targets through a MPO-dependent mechanism. Target properties, especially the content of lipid-soluble antioxidants, have a marked influence on susceptibility to lysis. These effects are in turn subject to modulation on the basis of such factors as the availability of various halide cofactors. This model system offers an opportunity to dissect the biochemical basis of effector cell-target cell interactions.",
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T1 - Oxidant membrane injury by the neutrophil myeloperoxidase system. II. Injury by stimulated neutrophils and protection by lipid-soluble antioxidants

AU - Sepe, S. M.

AU - Clark, Robert A

PY - 1985

Y1 - 1985

N2 - The stimulated human neutrophil can damage a variety of target cells, and in some models, a mechanism involving secretion of myeloperoxidase and H2O2 has been demonstrated. We explored the characteristics of this cell-cell interaction by using neutrophils and our recently described liposome model target cell system. Exposure of 51Cr-labeled liposomes to phorbol myristate acetate-stimulated human neutrophils resulted in release of 25 to 30% of the radioactivity. 51Cr release was abrogated by omission of the neutrophils, the phorbol ester on halide (iodide), replacement of the phorbol by an inactive congener, or addition of azide, cyanide, or catalase. Neutrophils from patients with hereditary absence of myeloperoxidase (MPO) or a failure of H2O2 formation (chronic granulomatous disease) did not cause liposome lysis unless purified MPO or a source of H2O2, respectively, was added. These data indicate that 51Cr release from liposomes is a consequence of the secretion of MPO and H2O2, which combine with extracellular halides to form a membrane lytic system. The influence of liposome composition on injury was then examined, with a focus on physiologically relevant lipid soluble antioxidants. Liposomes containing either α-tocopherol (0.33 to 1.67% of molar fraction of lipid) or β-carotene (1.67% of molar fraction of lipid) were markedly resistant to lysis by the cellfree MPO-H2O2-chloride system. When the major structural lipid phosphatidyl choline was replaced by dipalmitoyl phosphatidyl choline, a synthetic phospholipid with no oxidizable double bonds, the resultant liposomes were totally resistant to lysis by the MPO-H2O2-chloride system. The addition of iodide to this system (i.e., both chloride and iodide present) changed the pattern of protection dramatically in that α-tocopherol and β-carotene were no longer protective and the resistance of dipalmitoyl phosphatidyl choline liposomes was partial rather than complete. In contrast to iodide, the addition of bromide or thiocyanate did not have a major effect on the protection by antioxidants. Finally, we demonstrated protection by α-tocopherol or dipalmitoyl phosphatidyl choline against liposome lysis by phorbol-activated neutrophils. These studies illustrate the use of model phospholipid membranes in the characterization of oxygen-dependent cell-mediated cytotoxicity. Activated neutrophils lyse liposome targets through a MPO-dependent mechanism. Target properties, especially the content of lipid-soluble antioxidants, have a marked influence on susceptibility to lysis. These effects are in turn subject to modulation on the basis of such factors as the availability of various halide cofactors. This model system offers an opportunity to dissect the biochemical basis of effector cell-target cell interactions.

AB - The stimulated human neutrophil can damage a variety of target cells, and in some models, a mechanism involving secretion of myeloperoxidase and H2O2 has been demonstrated. We explored the characteristics of this cell-cell interaction by using neutrophils and our recently described liposome model target cell system. Exposure of 51Cr-labeled liposomes to phorbol myristate acetate-stimulated human neutrophils resulted in release of 25 to 30% of the radioactivity. 51Cr release was abrogated by omission of the neutrophils, the phorbol ester on halide (iodide), replacement of the phorbol by an inactive congener, or addition of azide, cyanide, or catalase. Neutrophils from patients with hereditary absence of myeloperoxidase (MPO) or a failure of H2O2 formation (chronic granulomatous disease) did not cause liposome lysis unless purified MPO or a source of H2O2, respectively, was added. These data indicate that 51Cr release from liposomes is a consequence of the secretion of MPO and H2O2, which combine with extracellular halides to form a membrane lytic system. The influence of liposome composition on injury was then examined, with a focus on physiologically relevant lipid soluble antioxidants. Liposomes containing either α-tocopherol (0.33 to 1.67% of molar fraction of lipid) or β-carotene (1.67% of molar fraction of lipid) were markedly resistant to lysis by the cellfree MPO-H2O2-chloride system. When the major structural lipid phosphatidyl choline was replaced by dipalmitoyl phosphatidyl choline, a synthetic phospholipid with no oxidizable double bonds, the resultant liposomes were totally resistant to lysis by the MPO-H2O2-chloride system. The addition of iodide to this system (i.e., both chloride and iodide present) changed the pattern of protection dramatically in that α-tocopherol and β-carotene were no longer protective and the resistance of dipalmitoyl phosphatidyl choline liposomes was partial rather than complete. In contrast to iodide, the addition of bromide or thiocyanate did not have a major effect on the protection by antioxidants. Finally, we demonstrated protection by α-tocopherol or dipalmitoyl phosphatidyl choline against liposome lysis by phorbol-activated neutrophils. These studies illustrate the use of model phospholipid membranes in the characterization of oxygen-dependent cell-mediated cytotoxicity. Activated neutrophils lyse liposome targets through a MPO-dependent mechanism. Target properties, especially the content of lipid-soluble antioxidants, have a marked influence on susceptibility to lysis. These effects are in turn subject to modulation on the basis of such factors as the availability of various halide cofactors. This model system offers an opportunity to dissect the biochemical basis of effector cell-target cell interactions.

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