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N Hogg

Publications and source records attributed to N Hogg.

At least 109 records · Page 6Linked to original sources
PubMed1994

Physical and chemical interactions between nitric oxide and nitroxides.

The physical and chemical interaction of nitric oxide (NO) with stable nitroxides have been studied in both aqueous and membrane environments. The ESR spectrum of 3-carbamoyl-2,2,5,5-tetramethyl-3-pyrroline-1- yloxy (CTPO) was observed to broaden upon exposure to NO. This effect can be explained by invoking Heisenberg spin exchange as has been previously reported for molecular oxygen. No loss of total spin was observed negating the possibility of a chemical reaction between NO and CTPO. The extent of signal broadening was proportional to the concentration of NO and can thus be used to monitor NO concentration. We have used this method to observe the partitioning of NO into model membranes. We also report the use of multiquantum ESR to detect directly the effects of NO on the membrane bound spin label 12-doxylstearic acid. This methodology may prove useful for detecting NO in both aqueous and lipid environments and for examining the physical properties of NO within biological membranes.

Cyclic N-Oxides↗
PubMed1994

The oxidation of cytochrome-c oxidase vesicles by hemoglobin.

Human hemoglobin has been used as a pro-oxidant for artificial unilamellar phospholipid vesicles, containing cytochrome-c oxidase inserted into the bilayer. This experimental system was suitable to follow directly the kinetics of lipid oxidation and the effects on both the vesicle membrane permeability and the functional state of cytochrome-c oxidase. Following mixing of vesicles with hemoglobin, an oxygen dependent, peroxyl radical mediated, rapid oxidation (taking a few minutes) of the lipid was found to occur. On a similar time scale the membrane became ion-leaky and cytochrome-c oxidase damaged. The pro-oxidant effects of hemoglobin in various oxidation and ligation states were studied and a mechanism, based on a ferric/ferryl redox cycle of the heme-iron is proposed to account for these observations.

Cell Membrane Permeability↗
PubMed1994

Neutrophil apoptosis is associated with a reduction in CD16 (Fc gamma RIII) expression.

Resolution of inflammation involves removal of recruited neutrophils from inflamed sites via a noninflammatory mechanism, possibly involving neutrophil apoptosis and engulfment/phagocytosis by macrophages. In this study, we describe the reduction in surface expression (> 90%) of the neutrophil molecule Fc gamma RIII (CD16) during in vitro culture at 37 degrees C, which was found to be temporally associated with the appearance of neutrophils with apoptotic morphology during in vitro culture and inhibitable by granulocyte-macrophage colony-stimulating factor (GM-CSF), which postpones apoptosis in the neutrophil. By using dual fluorescence analysis, CD16 "low" expressing neutrophils showed reduced staining with the DNA-binding dye propidium iodide, suggesting that CD16 low expressing neutrophils were apoptotic. Separation of CD16 "high" and CD16 "low" expressing neutrophils by fluorescence-activated cell sorting revealed that morphologically apoptotic cells exhibited the CD16 low phenotype. We did not observe similar marked changes in expression of other neutrophil surface molecules (including other phosphatidylinositol (PI)-linked molecules), indicating that generalized loss of surface molecules does not occur during apoptosis. We believe this to be the first reported cell type-specific membrane alteration in a surface glycoprotein associated with apoptosis, suggesting that the program of cell death in the neutrophil, in addition to morphologic and nuclear changes, includes alterations in expression of surface receptors.

Apoptosis↗
PubMed1994

I domain of beta 2 integrin lymphocyte function-associated antigen-1 contains a binding site for ligand intercellular adhesion molecule-1.

Lymphocyte function-associated antigen-1 (LFA-1) is a beta 2 integrin that participates in a broad range of leukocyte functions through binding to its ligand intercellular adhesion molecule-1 (ICAM-1). The location of the ICAM-1 binding site on LFA-1 is not known. A approximately 200-amino acid "inserted" or "I" domain, which is part of the beta 2 integrin alpha subunit, is homologous to the "A" domains found in the adhesive protein von Willebrand factor and in a number of other proteins. In von Willebrand factor, the A domains are involved in ligand binding, but their function in the other proteins is still unclear. In this report, we show that the LFA-1 I domain contains a binding site for ICAM-1, which can be expressed as an isolated functional unit. The I domain contains the epitopes for 18 out of 20 anti-LFA-1 monoclonal antibodies, many of which interfere with the interaction between LFA-1 and ICAM-1. The I domain binds directly to purified recombinant ICAM-1 and also inhibits LFA-1-dependent T cell adhesion to ICAM-1. This report establishes the I domain as an ICAM-1 binding region in LFA-1 and the first ligand binding site to be identified in a beta 2 integrin.

Antibodies, Monoclonal↗
PubMed1994

Integrin LFA-1 alpha subunit contains an ICAM-1 binding site in domains V and VI.

In order to identify a binding site for ligand intercellular adhesion molecule-1 (ICAM-1) on the beta 2 integrin lymphocyte function-associated antigen-1 (LFA-1), protein fragments of LFA-1 were made by in vitro translation of a series of constructs which featured domain-sized deletions starting from the N-terminus of the alpha subunit of LFA-1. Monoclonal antibodies and ICAM-1 were tested for their ability to bind to these protein fragments. Results show that the putative divalent cation binding domains V and VI contain an ICAM-1 binding site. A series of consecutive peptides covering these domains indicated two discontinuous areas as specific contact sites: residues 458-467 in domain V and residues 497-516 in domain VI. A three-dimensional model of these domains of LFA-1 was constructed based on the sequence similarity to known EF hands. The two regions critical for the interaction of LFA-1 with ICAM-1 lie adjacent to each other, the first next to the non-functional EF hand in domain V and the second coinciding with the potential divalent cation binding loop in domain VI. The binding of ICAM-1 with the domain V and VI region in solution was not sensitive to divalent cation chelation. In short, a critical motif for ICAM-1 binding to the alpha subunit of LFA-1 is shared between two regions of domains V and VI.

Amino Acid Sequence↗
PubMed1994

The role of lipid hydroperoxides in the myoglobin-dependent oxidation of LDL.

It has previously been reported that mb in both the iron-oxo ferryl and the ferric oxidation states can promote lipid peroxidation and lead to oxidative modification of low-density lipoprotein. The mechanism of these oxidation reactions is unclear and could involve either lipid hydroperoxide-dependent or independent reactions. In order to ascertain which of the afore-mentioned mechanisms predominates, the effects of exogenous lipid hydroperoxides on the ability of Mb, in its various oxidation states, to oxidize low-density lipoprotein has been investigated. The results suggest that oxidation proceeds through a one-electron redox cycle between met and ferryl myoglobin and that the reactions of both redox forms are at least partially dependent on lipid hydroperoxides within the LDL particle.

Electrophoresis↗
PubMed1994

The oxidation of alpha-tocopherol and trolox by peroxynitrite.

Peroxynitrite reacts rapidly with alpha-tocopherol to generate a mixture of species. The predominant products are 8a-methoxytocopherone in methanol and alpha-tocopherylquinone in acetonitrile. Only a small fraction (about 2% of original alpha-tocopherol) was detected as alpha-tocopheroxyl radical in either solvent. We propose that peroxynitrite oxidizes alpha-tocopherol in a two-electron process yielding the alpha-tocopherone cation. The two-electron oxidation may be either concerted or sequential. The fate of the alpha-tocopherone cation is solvent dependent. In acetonitrile it undergoes hydrolysis, in the presence of trace amounts of water, to form alpha-tocopherylquinone. In methanol it undergoes nucleophilic addition to yield 8a-methoxytocopherone. Our data suggest that two-electron oxidation of alpha-tocopherol by peroxynitrite represents the major pathway, whereas one-electron oxidation to generate alpha-tocopheroxyl radical is a minor pathway. The biological consequences of two-electron oxidation of alpha-tocopherol are discussed.

Acetonitriles↗
PubMed1994

The sticking point: How integrins bind to their ligands.

The integrin adhesion receptors are alpha beta heterodimers that exist in different ligand-binding states. Because of their large size and conformational lability, it has been difficult to determine how they interact with their ligands. Ligand-binding sites have been identified in the beta subunit, and now more recently in the 'I' domain and EF-hand-like domains V and VI of the alpha subunit. We speculate here about how these various sites might operate together to bind ligand in a stable manner.

Journal Article↗
PubMed1994

Involvement of the "I" domain of LFA-1 in selective binding to ligands ICAM-1 and ICAM-3.

To analyze the binding requirements of LFA-1 for its two most homologous ligands, ICAM-1 and ICAM-3, we compared the effects of various LFA-1 activation regimes and a panel of anti-LFA-1 mAbs in T cell binding assays to ICAM-1 or ICAM-3 coated on plastic. These studies demonstrated that T cell binding to ICAM-3 was inducible both from the exterior of the cell by Mn2+ and from the interior by an agonist of the "inside-out" signaling pathway. T cells bound both ICAM ligands with comparable avidity. A screen of 29 anti-LFA-1 mAbs led to the identification of two mAbs specific for the alpha subunit of LFA-1 which selectively blocked adhesion of T cells to ICAM-3 but not ICAM-1. These two mAbs, YTH81.5 and 122.2A5, exhibited identical blocking properties in a more defined adhesion assay using LFA-1 transfected COS cells binding to immobilized ligand. Blocking was not due to a steric interference between anti-LFA-1 mAbs and N-linked carbohydrate residues present on ICAM-3 but not ICAM-1. The epitopes of mAbs YTH81.5 and 122.2A5 were shown to map to the I domain of the LFA-1 alpha subunit. A third I domain mAb, MEM-83, has been previously reported to uniquely activate LFA-1 to bind ICAM-1 (Landis, R. C., R. I. Bennett, and N. Hogg. 1993. J. Cell Biol. 120:1519-1527). We now show that mAb MEM-83 is not able to stimulate binding of T cells to ICAM-3 over a wide concentration range. Failure to induce ICAM-3 binding by mAb MEM-83 was not due to a blockade of the ICAM-3 binding site on LFA-1. This study has demonstrated that two sets of functionally distinct mAbs recognizing epitopes in the I domain of LFA-1 are able to exert differential effects on the binding of LFA-1 to its ligands ICAM-1, and ICAM-3. These results suggest for the first time that LFA-1 is capable of binding these two highly homologous ligands in a selective manner and that the I domain plays a role in this process.

Antibodies, Monoclonal↗
PubMed1993

Inhibition of low-density lipoprotein oxidation by nitric oxide. Potential role in atherogenesis.

The effects of nitric oxide (.NO) and nitrovasodilators on the oxidation of low-density lipoprotein (LDL) have been studied. S-Nitroso-N-acetylpenicillamine (SNAP) and sodium nitroprusside (SNP) inhibited Cu(2+)- and 2,2'-azobis-2-amidinopropane hydrochloride-dependent oxidation of LDL as monitored by oxygen consumption and the formation of thiobarbituric acid-reactive substances, conjugated dienes, and lipid hydroperoxides. In the case of SNP, inhibition of LDL oxidation occurred only when the incubation mixture was irradiated with visible light. SNAP, however, exerted a dose-dependent inhibition of Cu(2+)-catalyzed oxidation of LDL even in the dark. Addition of .NO dissolved in deoxygenated buffer also inhibited the progression of LDL oxidation. Mouse peritoneal macrophages were less able to degrade LDL that had been oxidized in the presence of SNAP. Using an .NO electrode, it was estimated that a continuous production of .NO (< or = 760 nM/min) could retard the progression of LDL oxidation. We propose that .NO can inhibit LDL oxidation by acting as a chain-breaking antioxidant that is capable of scavenging carbon-centered and peroxyl radicals. Biological implications of this novel .NO antioxidant property are discussed in relation to atherogenesis and contrasted to the prooxidant property of .NO when generated in the presence of superoxide.

Animals↗
PubMed1993

Peroxynitrite modification of low-density lipoprotein leads to recognition by the macrophage scavenger receptor.

Peroxynitrite is an oxidant which could be formed in the vasculature by the reaction of superoxide with nitric oxide. It is capable of modifying amino acid residues and of initiating lipid peroxidation. In the present study we have shown that peroxynitrite converts low density lipoprotein to a form recognized by the macrophage scavenger receptor and that this process is associated with modification of the protein and lipid, and with the oxidation of alpha-tocopherol to alpha-tocopherol quinone.

Cell Line↗
PubMed1993

The oxidation of alpha-tocopherol in human low-density lipoprotein by the simultaneous generation of superoxide and nitric oxide.

Peroxynitrite is the product of the reaction between nitric oxide and superoxide. It is an oxidant which can also decompose to form the hydroxyl radical and nitrogen dioxide. In this report we show that a powerful oxidant with reactivity similar to that of the hydroxyl radical is formed from the generation of superoxide from xanthine oxidase and nitric oxide from S-nitroso-n-acetylpenicillamine (SNAP). Simultaneous generation of these two radicals by either xanthine oxidase/SNAP or the sydnonimine SIN-1 in the presence of low-density lipoprotein (LDL) results in the depletion of alpha-tocopherol and formation of its oxidised product alpha-tocopheroquinone. The mechanism of oxidation required both the formation of nitric oxide and superoxide. In contrast to the promotion of LDL oxidation by transition metals the oxidation of LDL by SIN-1 was not sensitive to the addition of exogenous lipid hydroperoxide.

Acetaldehyde↗
PubMed1993

Ligand intercellular adhesion molecule 1 has a necessary role in activation of integrin lymphocyte function-associated molecule 1.

The signaling that causes the leukocyte integrin lymphocyte function-associated molecule (LFA-1) to bind firmly to its ligand intercellular adhesion molecule 1 (ICAM-1) is transduced indirectly through other T-cell receptors and is termed inside-out signaling. We show here that the high-affinity state of LFA-1 is characterized by expression of the LFA-1 epitope detected by monoclonal antibody 24. This epitope is expressed not in response to the initial agonist-mediated signal but when LFA-1 binds to ICAM-1, indicating that ligand binding induces an alteration in LFA-1. As would be predicted, the monoclonal antibody 24 epitope is confined to the LFA-1, which is located at the site of contact between T cells and ICAM-1-expressing transfectants. When a fixation protocol for "freezing" receptors is used, only T cells that are fixed after prior exposure to ICAM-1 bind firmly to ICAM-1 a second time. This suggests that, in addition to the inside-out signaling, a previously unrecognized requirement for full activation of the leukocyte integrin LFA-1 is the initial interaction with its ligand ICAM-1. Thus, activation of LFA-1 is in part achieved by an induced fit imposed from without by interaction with ligand.

Animals↗
PubMed1993

Coordinate expression of beta 1 and beta 2 integrin "activation" epitopes during T cell responses in secondary lymphoid tissue.

The monoclonal antibodies (mAb) 15/7 and 24 recognize unique activation-dependent, conformational epitopes on beta 1 and beta 2-integrins, respectively. The expression of both of these epitopes closely correlates with the ligand binding ability of their respective integrins, and thus serves as indicators of functional integrin "activation". Here, we have used six-parameter flow cytometry to examine the expression of these epitopes and conventional beta 1- and beta 2-integrin epitopes during human T cell activation in secondary lymphoid tissues in vivo, focusing particularly on the virgin to memory/effector cell transition. Fresh tonsil lymphocytes were stained with mAb against conventional or activation-dependent integrin epitopes, followed by staining with mAb against CD3, CD45RA, and CD45RO, thus allowing the determination of integrin epitope expression on virgin (CD3+) T cells (CD45RA+/RO-to+/-), memory/effector (CD45RA-/RO++) T cells, and T cells undergoing the virgin to memory/effector transition: transition region-1 (T1; CD45RA+to++/RO+); -2 (T2; CD45RA++/RO++); and -3 (T3; CD45RA+/RO++). Conventional beta 1- and beta 2-integrin epitopes progressively increase during the virgin to T3 stages of the transition in tonsil, in keeping with the generally higher levels of these adhesion molecules on memory/effector vs. virgin T cells. Expression of both the beta 1 (15/7)- and beta 2 (24)-integrin activation epitopes first appears on transitional T cells, and is maintained on a relatively constant number of cells (averaging 25-30%) throughout the T1-T3 stages. These epitopes are also noted on a subset of activated memory/effector T cells. Importantly, on both transitional and activated memory/effector T cell subsets, the expression patterns of the 15/7 and 24 epitopes vs. a variety of T cell activation antigens are identical, and the expression of these epitopes relative to each other is linearly correlated, findings strongly supporting the coordinate activation of beta 1 and beta 2 integrins during T cell activation in vivo. These results provide the first evidence of integrin activation during an in vivo immunologic response, and demonstrate the usefulness of mAb recognizing conformational epitopes and multiparameter flow cytometry in delineating the dynamic interplay of adhesion molecules during complex physiologic processes.

Adolescent↗
PubMed1993

Adhesion molecules in cell interactions.

During a successful immune response, several families of adhesion molecules participate in a cascade of binding events that lead to the binding of leukocytes, both to each other and to cell types such as the endothelium and epithelium. A central theme emerging from recent studies is that the function of an adhesion receptor cannot be inferred from its expression alone; rather, adhesion receptors are 'selected' to perform distinct effector functions based on their cell-background and factors present in the local microenvironment. Thus, adhesion receptors expressed on different cell-types may find themselves in different states of 'activation-readiness' and may be further selected by prevailing conditions in the microenvironment to bind tissue-specific ligands and mediate leukocyte effector functions such as homing or transendothelial migration.

Animals↗
PubMed1993

A novel LFA-1 activation epitope maps to the I domain.

A panel of 21 alpha-subunit (CD11a) and 10 beta-subunit (CD18) anti-LFA-1 mAbs was screened for ability to activate LFA-1. A single anti-CD11a mAb, MEM-83, was identified which was able to directly induce the binding of T cells to purified ICAM-1 immobilized on plastic. This ICAM-1 binding could be achieved by monovalent Fab fragments of mAb MEM-83 at concentrations equivalent to whole antibody, was associated with appearance of the "activation reporter" epitope detected by mAb 24, and was completely inhibited by anti-ICAM-1 and LFA-1 blocking mAbs. The epitope recognized by mAb MEM-83 was distinct from that recognized by mAb NKI-L16, an anti-CD11a mAb previously reported to induce LFA-1 activation, in that it was constitutively present on freshly isolated peripheral blood mononuclear cells and was not divalent cation dependent for expression. The ICAM-1 binding activity induced by mAb MEM-83 was, however, dependent on the presence of Mg2+ divalent cations. Using an in vitro-translated CD11a cDNA deletion series, we have mapped the MEM-83 activation epitope to the "I" domain of the LFA-1 alpha subunit. These studies have therefore identified a novel LFA-1 activation epitope mapping to the I domain of LFA-1, thereby implicating this domain in the regulation of LFA-1 binding to ICAM-1.

Antibodies, Monoclonal↗