Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Azides”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,459 records · Page 81Linked to original sources
PubMed1997

Orthovanadate induces cell death in rat dentate gyrus primary culture.

The aim of this study was to define the effects of a potent inhibitor of tyrosine phosphatases, sodium orthovanadate (0.1-100 microM for up to 48 h), on dentate gyrus cells (DGC) in culture. Treatment with 100 microM orthovanadate evoked a delayed form of cell death. To examine the possible involvement of apoptosis in orthovanadate-induced cell death, biochemical and morphological alterations were compared with those of necrotic death induced by sodium azide. Phase-contrast microscopy and nuclear condensation analysis showed that orthovanadate and azide each evoked cell death by distinct pathways. TUNEL assay was positive in both cases. Application of a protein synthesis inhibitor, cycloheximide, did not prevent cytotoxicity caused by either orthovanadate or azide and potentiated the effects of vanadate. We conclude that orthovanadate-induced death of DGC bears features of apoptosis.

Animals↗
PubMed1977

The reduction by dithionite of Fe(III) myoglobin derivatives with different ligands attached to the iron atom. A study by rapid-wavelength-scanning stopped-flow spectrophotometry.

1. The reductions of a number of sperm whale Fe(III) myoglobin-ligand complexes by sodium dithionite in a phosphate buffer pH 6.4, were investigated by using rapid-wavelength-scanning stopped-flow spectrophotometry. The ligands were azide, cyanide, fluoride, imidazole, thiocyanate and water. 2. The reduction of Fe(III) myoglobin cyanide led to the transient formation of Fe(II) myoglobin cyanide but no intermediate species were observable during the reductions of the other derivatives. The final product of the reaction in all cases was unliganded Fe(II)myoglobin. 3. Invesigation of the effect of dithionite concentration on the rate of reduction indicated that the SO2- radical ion was the active species in reducing the azide, cyanide, fluoride and thiocyanate derivatives. 4. Comparison of the observed rates of reduction at different ligand concentrations with those predicted for a pathway of reduction involving prior dissociation of the ligand, allowed us to estimate the rate of reduction with the ligand in position (outer-sphere reduction). There was a large variation in the relative rates of outer-sphere reduction in the order imidazole greater than CN- greater than SCN- greater than N3- greater than F-. The fluoride derivative was so resistant to outer-sphere reduction that the reaction with SO2- proceeded only by a pathway involving dissociation of F- before reduction. It was calculated that any direct reduction of this complex was at least 100 times slower than that of the azide derivative. 5. The results are discussed in terms of the possible rôle of the axial ligands in haem proteins and it is suggested that the pathway of the electron to the Fe(III) centre may be via the ppi orbitals of these ligands.

Animals↗
PubMed1995

The pH-dependent changes of the enzymic activity and spectroscopic properties of iron-substituted manganese superoxide dismutase. A study on the metal-specific activity of Mn-containing superoxide dismutase.

Manganese-containing superoxide dismutases (Mn-SODs) and iron-containing superoxide dismutases (Fe-SODs) from aerobic bacteria often show high metal specificity for their enzymic activities by a standard assay system using xanthine-xanthine oxidase and cytochrome c. In this study, we have attempted to characterize the structural basis of the metal specificity of manganese-containing SOD (Mn-SOD) using Fe-substituted Mn-SOD prepared from apo-Mn-SOD from Serratia marcescens. The Fe3+ content of the Fe-substituted enzyme was 1.71 +/- 0.14 mol/mol dimer and the specific activity was 34.8 +/- 4.8 units.mg protein-1.mol Fe3+(-1).mol subunit-1. Fe-substituted Mn-SOD was found to react with the superoxide anion at pH 8.1 with a second-order rate constant of 6 x 10(6) M-1 s-1, which is approximately 1% of that of native Mn-SOD at the same pH. However, the rate constant increased with decreasing pH to approximately 10% (5 x 10(7) M-1 s-1) that of native Mn-SOD at pH 6.0 with a pK of 7.0. The visible absorption spectrum and EPR spectrum of Fe-substituted Mn-SOD also showed pH-dependent changes with pK values of 6.6 and 7.2, respectively. Similarly, the affinity of the azide ion, an analog of the superoxide ion, for iron of Fe-substituted Mn-SOD increased with decreasing pH, with a pK value of 7.0 (e.g. Kd = 0.1 mM at pH 6.2 and 0.9 mM at pH 8.2). The similarity of these pK values suggests that the activity, the spectral changes and the affinity of the azide ion for iron are derived from the same change in the metal environment. After comparison with the reported pK values (around 9) of similar pH-dependent changes in the spectra, the enzymic activity and the affinity of azide for iron of Fe-SOD from Escherichia coli, we proposed that the difference in the pK values of a hydroxide ion binding to iron between Fe-substituted Mn-SOD and Fe-SOD may cause the different pH dependencies of these changes in each SOD.

Azides↗
PubMed1996

Kinetic evidence for the existence of a rate-limiting step in the reaction of ferric hemoproteins with anionic ligands.

The kinetics of azide and fluroide binding to various monomeric and tetrameric ferric hemoproteins (sperm whale Mb, isolated alpha and beta chains of human Hb reacted with p-chloromercuribenzoate, dromeday, ox and human Hb) has been investigated (at pH 6.5 and 20 degrees C over a large range (20 microM to 2 M) of ligand concentration. It has been observed that the pseuo-first-order rate constant for azide binding to the hemoproteins investigated does not increase linearly with ligand concentration, but tends to level off toward an asymptomatic concentration-independent value typical for each hemoprotein. This behavior, which has been detected only by an investigation covering an unusually large range of ligand concentrations appears to be independent of the ionic strength, and it underlies the existence of a rate-limiting step in the dynamic pathway of azide binding to ferric hemoproteins, which is detectable whenever the observed pseudo- first-order rate constant becomes faster than a given value characteristic of the specific hemoprotein. Such a behavior is not observed in the case of fluroide binding probably because the pesudo- first-order rate constant for this ligand is much slower and never attains a value faster than that of the rate-limiting step. In general terms, this feature should involve a conformational equilibrium between at least two forms (possibly related to the interaction of H2O with distal histidine and its exchange with the bulk solvent) which modulates the access of the anionic ligand into the heme pocket and its reaction with the ferric iron.

Animals↗
PubMed1977

Steady-state oxygen kinetics of terminal oxidases in Trypanosoma mega.

Steady-state oxygen kinetics of Trypanosoma mega reveal the presence of 3 oxidases. These include an oxidase which is sensitive to salicylhydroxamic acid (SHAM) but insensitive to sodium azide. This oxidase could be the L-alpha glycerophosphate oxidase present in bloodstream trypanosomes. In addition, and oxidase is present wthich is azide-sensitive but SHAM-insensitive. This oxidase is inhibited by CO and is probably cytochrome aa3. A 3rd oxidase is insensitive to both azide and SHAM but is inhibited by CO and is possibly cytochrome o. Reciprocal plots of T. mega reveal the presence of 2 oxidases that are inhibited by CO. These results are discussed in the light of previous evidence suggesting the presence of several oxidases and a branched electron transport system in T. mega.

Animals↗
PubMed1995

Exploratory photochemistry of 5-azido-8-alkoxy-substituted psoralens free and bound to DNA.

5-Azido-8-alkoxy psoralens were synthesized. Laser flash photolysis (LFP: XeF, 351 nm, 55 mJ, 17 ns) of the azides in acetonitrile or benzene solution produces the triplet nitrene and a small amount of ketenimine. Laser flash photolysis of the azides in methanol or aqueous solution cleanly produces the triplet nitrene. In aqueous solution containing highly polymerized calf thymus DNA, LFP produces a mixture of triplet nitrene and ketenimine corresponding to photolysis of free and bound psoralen, respectively. The two transients decay slowly but at different rates. Assignment of the transient spectra were secured by matrix EPR and UV-visible spectroscopy. The triplet nitrene lifetime is the same in buffer and in the presence and absence of calf thymus DNA. The results explain why psoralen azides are unable to efficiently nick plasmid DNA pBR322 upon UV activation.

Animals↗
PubMed1997

Singlet oxygen-mediated inactivation of acetylcholinesterase: a comparison of purified enzyme in solution and enzyme bound to K562 leukemia cells.

We have compared the singlet oxygen-mediated inactivation of acetylcholinesterase (ACE) in solution with the inactivation of ACE on the surface of K562 leukemia cells. In solution, the actions of the singlet-oxygen quenchers, methionine, azide, disodium [N,N'-ethylenebis (5-sulfosalicylideneimminato)]nickelate(II) (Ni-chelate 1) and disodium [(N,N'-2,3-propionic acid)bis(5-sulfosal-icylideneimminato)] nickelate(II) (Ni-chelate 2) could be explained quantitatively by assuming their only mechanism of action was to quench singlet oxygen. The singlet oxygen quenchers, azide, Ni-chelate 1 and Ni-chelate 2, caused smaller inhibitions in the rate of singlet oxygen-mediated inactivation of ACE on K562 cells than ACE in solution. The effects of these quenchers and of deuterium oxide were interpreted using a mathematical model of singlet-oxygen quenching and diffusion to estimate the lifetime of singlet oxygen near the cell surface. The azide quenching data and the deuterium-oxide data gave lifetimes of 0.9 +/- 0.2 microsecond and 0.45 +/- 0.15 microsecond, respectively. The increases in ACE inactivation lifetime caused by the nickel chelates were anomalously large. The unexpectedly large quenching due to the nickel chelates may have been due to a nonuniform distribution of the chelates in the cytoplasm with a large concentration of the chelate near the cell membrane.

Acetylcholinesterase↗
PubMed1970

Myeloperoxidase: contribution to the microbicidal activity of intact leukocytes.

Azide and, to a lesser extent, cyanide inhibit the microbicidal activity of myeloperoxidase and of intact normal leukocytes, but they have little or no effect on peroxidase-negative leukocytes. The contribution of the azide-sensitive (peroxidase-dependent?) systems to the total microbicidal activity of normal leukocytes is considerable. The azide-insensitive antimicrobial systems are more highly developed in peroxidase-negative leukocytes than in normal leukocytes, thus suggesting an adaptation.

Azides↗
PubMed2003

Relationships between respiration and susceptibility to azole antifungals in Candida glabrata.

Over the past two decades, the incidence of infections due to Candida glabrata, a yeast with intrinsic low susceptibility to azole antifungals, has increased markedly. Respiratory deficiency due to mutations in mitochondrial DNA (mtDNA) associated with resistance to azoles frequently occurs in vitro in this species. In order to specify the relationships between respiration and azole susceptibility, the effects of respiratory chain inhibitors on a wild-type isolate of C. glabrata were evaluated. Respiration of blastoconidia was immediately blocked after extemporaneous addition of potassium cyanide, whereas a 4-h preincubation was required for sodium azide. Antifungal susceptibility determined by a disk diffusion method on Casitone agar containing sodium azide showed a significant decrease in the susceptibility to azoles. Biweekly subculturing on Casitone agar supplemented with sodium azide was therefore performed. This resulted after 40 passages in the isolation of a respiration-deficient mutant, as suggested by its lack of growth on glycerol-containing agar. This respiratory deficiency was confirmed by flow cytometric analysis of blastoconidia stained with rhodamine 123 and by oxygraphy. Moreover, transmission electron microscopy and restriction endonuclease analysis of the mtDNA of mutant cells demonstrated the mitochondrial origin of the respiratory deficiency. Finally, this mutant exhibited cross-resistance to all the azoles tested. In conclusion, blockage of respiration in C. glabrata induces decreased susceptibility to azoles, culminating in azole resistance due to the deletion of mtDNA. This mechanism could explain the induction of petite mutations by azole antifungals which have been demonstrated to act directly on the mitochondrial respiratory chain.

Antifungal Agents↗
PubMed1996

Involvement of superoxide and myeloperoxidase in oxygen-dependent killing of Staphylococcus aureus by neutrophils.

We have used a quantitative assay that measures independent rate constants for phagocytosis and killing of Staphylococcus aureus to investigate the involvement of superoxide and myeloperoxidase in bacterial killing by human neutrophils. To inhibit superoxide-dependent processes, superoxide dismutase was cross-linked to immunoglobulin G and the conjugate was attached to the surface of S. aureus via protein A in its cell wall. Myeloperoxidase was inhibited with azide, and myeloperoxidase-deficient neutrophils were used. Adding the NADPH oxidase inhibitor diphenyleneiodonium, to prevent superoxide production, decreased the killing rate to 25%, indicating that oxidative killing mechanisms predominate in this system. The rate constant for killing of S. aureus with superoxide dismutase attached was 70% of that for control bacteria linked to inactivated enzyme. Superoxide dismutase had no effect in the presence of diphenyleneiodonium. The rate of killing was decreased to 33% in the presence of azide and to 40% with myeloperoxidase-deficient neutrophils. Superoxide dismutase had no effect in the presence of azide. On the assumption that the oxidative and nonoxidative components of killing can be considered separately, the oxidative rate was decreased by almost half by superoxide dismutase and was about six times lower when myeloperoxidase was inactive. We conclude that myeloperoxidase-dependent processes are strongly favored by human neutrophils as their prime mechanism of oxidative killing of S. aureus and that superoxide makes a direct contribution to killing. Our results also suggest that superoxide acts in conjunction with a myeloperoxidase-dependent pathway.

Azides↗
PubMed1973

Studies on beta-galactoside transport in a Proteus mirabilis merodiploid carrying an Escherichia coli lactose operon.

Merodiploid derivatives bearing an F-linked lac operon (i(+), o(+), z(+), y(+), a(+)) from Escherichia coli were prepared from a Proteus mirabilis strain unable to utilize lactose and from a lac deletion strain of E. coli. A suitable growth medium was found in which the episomal element in the P. mirabilis derivative was sufficiently stable to allow induction of the episome-borne lac operon and thus to permit a comparison of the activities and properties of E. coli lac products in the intracellular environments of P. mirabilis and E. coli. In both derivatives the episomal lac operon was shown to be repressed in the absence of inducer. Kinetics of induction with gratuitous inducer (isopropyl-1-thio-beta-d-galactoside) were similar for both beta-galactosidase activity (beta-d-galactoside galactohydrolase, EC 3.4.1.23) and beta-galactoside transport activity in both derivatives, although the ratio of galactoside transport to beta-galactosidase activity was approximately 1.6-fold higher in the E. coli derivative. Comparison of beta-galactosidase and M-protein (lac y gene product)-specific activities indicated coordinate expression of the induced lac operon in both derivatives. Quantitatively, the maximal beta-galactosidase specific activity was two or three times higher for the E. coli derivative. A significant sodium azide inhibition (65% inhibition by 10 mM sodium azide) of lactose permease-mediated transport of o-nitrophenyl-beta-galactoside from an outside region of high concentration to an inside region of very low concentration ("downhill transport") was observed for the P. mirabilis derivative. Identical conditions for the E. coli derivative yielded only about 15% inhibition. Active transport of thiomethyl-beta-galactoside was similar for both derivatives, the major difference being that active transport was more sensitive to azide poisoning in the P. mirabilis derivative. Preliminary examination of the thiomethyl-beta-galactoside derivatives following active transport did not demonstrate the accumulation of a phosphorylated product in either strain but did reveal an unidentified derivative present in the P. mirabilis merodiploid extract which was not detectable in the E. coli merodiploid.

Azides↗
PubMed1991

Reconstitution of the ethanol oxidase respiratory chain in membranes of quinoprotein alcohol dehydrogenase-deficient Gluconobacter suboxydans subsp. alpha strains.

The ethanol oxidase respiratory chain of Gluconobacter suboxydan was characterized by using G. suboxydans subsp. alpha, a variant species of G. suboxydans incapable of oxidizing ethanol. The membranes of G. suboxydans subsp. alpha exhibited neither alcohol dehydrogenase, ethanol oxidase, nor glucose-ferricyanide oxidoreductase activity. Furthermore, the respiratory chain of the organism exhibited an extremely diminished amount of cytochrome c and an increased sensitivity of the respiratory activity for cyanide or azide when compared with G. suboxydans. The first-subunit quinohemoprotein and the second-subunit cytochrome c of alcohol dehydrogenase complex in the membranes of G. suboxydans subsp. alpha were shown to be reduced and deficient, respectively, by using heme-staining and immunoblotting methods. Ethanol oxidase activity, lacking in G. suboxydans subsp. alpha, was entirely restored by reconstituting alcohol dehydrogenase purified from G. suboxydans to the membranes of G. suboxydans subsp. alpha; this also led to restoration of the cyanide or azide insensitivity and the glucose-ferricyanide oxidoreductase activity in the respiratory chain without affecting other respiratory activities such as glucose and sorbitol oxidases. Ethanol oxidase activity was also reconstituted with only the second-subunit cytochrome c of the enzyme complex. The results indicate that the second-subunit cytochrome c of the alcohol dehydrogenase complex is essential in ethanol oxidase respiratory chain and may be involved in the cyanide- or azide-insensitive respiratory chain bypass of G. suboxydans.

Acetobacter↗
PubMed1980

Localization of membrane-associated proteins in vesicular stomatitis virus by use of hydrophobic membrane probes and cross-linking reagents.

The location of membrane-associated proteins of vesicular stomatitis virus was investigated by using two monofunctional and three bifunctional probes that differ in the degree to which they partition into membranes and in their specific group reactivity. Two hydrophobic aryl azide probes, [(125)I]5-iodonaphthyl-1-azide and [(3)H]pyrenesulfonylazide, readily partitioned into virion membrane and, when activated to nitrenes by UV irradiation, formed stable covalent adducts to membrane constituents. Both of these monofunctional probes labeled the glyco-protein G and matrix M proteins, but [(125)I]5-iodonaphthyl-1-azide also labeled the nucleocapsid N protein and an unidentified low-molecular-weight component. Protein labeling of intact virions was unaffected by the presence of cytochrome c or glutathione, but disruption of membrane by sodium dodecyl sulfate greatly enhanced the labeling of all viral proteins except G. Labeling of G protein was essentially restricted to the membrane-embedded, thermolysin-resistant tail fragment. Three bifunctional reagents, tartryl diazide, dimethylsuberimidate, and 4,4'-dithiobisphenylazide, were tested for their capacity to cross-link proteins to membrane phospholipids of virions grown in the presence of [(3)H]palmitate. Only G and M proteins of intact virions were labeled with (3)H-phospholipid by these cross-linkers; the reactions were not affected by cytochrome c but were abolished by disruption of virus with sodium dodecyl sulfate. Dimethylsuberimidate, which reacts with free amino groups, cross-linked (3)H-phospholipid to both G and M protein. In contrast, the hydrophilic tartryl diazide cross-linked phospholipid primarily to the M protein, whereas the hydrophobic 4,4'-dithiobisphenylazide cross-linked phospholipid primarily to the intrinsic G protein. These data support the hypothesis that the G protein traverses the virion membrane and that the M protein is membrane associated but does not penetrate very deeply, if at all.

Azides↗
PubMed2005

Role of 5'-AMP-activated protein kinase in stimulation of glucose transport in response to inhibition of oxidative phosphorylation.

Glucose transport is stimulated in a variety of cells and tissues in response to inhibition of oxidative phosphorylation. However, the underlying mechanisms and mediating steps remain largely unknown. In the present study we first tested whether a decrease in the redox state of the cell per se and the resultant increase in generation of reactive oxygen species (ROS) lead to stimulation of glucose transport. Clone 9 cells (expressing the Glut1 isoform of facilitative glucose transporters) were exposed to azide, lactate, and ethanol for 1 h. Although all three agents stimulated glucose transport and increased cell NADH-to-NAD(+) ratio and phospho-ERK1/2, signifying increased ROS generation, the response to the stimuli was not blocked by N-acetyl-l-cysteine (an agent that counteracts ROS); moreover, the response to azide was not blocked by diamide (an intracellular sulfhydryl oxidizing agent). We then found that cell AMP-to-ATP and ADP-to-ATP ratios were increased and 5'-AMP-activated protein kinase (AMPK) was stimulated by all three agents, as evidenced by increased phosphorylation of AMPK and acetyl-CoA carboxylase. We conclude that although azide, lactate, and ethanol increase NADH-to-NAD(+) ratios and ROS production, their stimulatory effect on glucose transport is not mediated by increased ROS generation. However, all three agents increased cell AMP-to-ATP ratio and stimulated AMPK, making it likely that the latter pathway plays an important role in the glucose transport response.

AMP-Activated Protein Kinases↗
PubMed1992

Enhancement of glucose transport in response to inhibition of oxidative metabolism: pre- and posttranslational mechanisms.

Addition of 5 mM sodium azide to Clone 9 cells, a rat liver cell line characterized by intracellular glucose concentrations of less than 10% that of the external medium and limited glycogen stores, results in a 50-80% reduction in cell ATP content within 20 min which then recovers to near-basal levels within 1 h and is subsequently maintained at normal levels for 24 h despite continuing the presence of the inhibitor. Associated with this adaptive response is a striking stimulation of facilitated glucose transport, mediated by the GLUT-1 transporter, that exhibits "early" and "late" phases that appear to be mechanistically different. During the early phase of the response (0-2 h), glucose transport rate is enhanced 12-fold in the absence of any change in cell GLUT-1 or GLUT-1 mRNA content. In contrast, the late phase of the response (8-24 h) is characterized by a further large stimulation of glucose transport (to 1.6 times the 2-h value) that is associated with 2- to 3- and 6- to 10-fold increments in cell GLUT-1 and GLUT-1 mRNA content, respectively. In time course studies an increase in GLUT-1 mRNA content was observed at 4 h and preceded the increment in GLUT-1 which became detectable after 8 h of exposure to azide. A marked induction of GLUT-1 mRNA by azide was also demonstrable in cells incubated in medium containing higher concentrations of glucose (10.6 mM), although the increment was approximately 20% less than when cells were incubated in standard medium (containing 5.6 mM glucose).(ABSTRACT TRUNCATED AT 250 WORDS)

3-O-Methylglucose↗
PubMed2005

Role of adenosine A1 and A3 receptors in regulation of cardiomyocyte homeostasis after mitochondrial respiratory chain injury.

Activation of either the A(1) or the A(3) adenosine receptor (A(1)R or A(3)R, respectively) elicits delayed cardioprotection against infarction, ischemia, and hypoxia. Mitochondrial contribution to the progression of cardiomyocyte injury is well known; however, the protective effects of adenosine receptor activation in cardiac cells with a respiratory chain deficiency are poorly elucidated. The aim of our study was to further define the role of A(1)R and A(3)R activation on functional tolerance after inhibition of the terminal link of the mitochondrial respiratory chain with sodium azide, in a state of normoxia or hypoxia, compared with the effects of the mitochondrial ATP-sensitive K(+) channel opener diazoxide. Treatment with 10 mM sodium azide for 2 h in normoxia caused a considerable decrease in the total ATP level; however, activation of adenosine receptors significantly attenuated this decrease. Diazoxide (100 muM) was less effective in protection. During treatment of cultured cardiomyocytes with hypoxia in the presence of 1 mM sodium azide, the A(1)R agonist 2-chloro-N(6)-cyclopentyladenosine was ineffective, whereas the A(3)R agonist 2-chloro-N(6)-iodobenzyl-5'-N-methylcarboxamidoadenosine (Cl-IB-MECA) attenuated the decrease in ATP level and prevented cell injury. Cl-IB-MECA delayed the dissipation in the mitochondrial membrane potential during hypoxia in cells impaired in the mitochondrial respiratory chain. In cells with elevated intracellular Ca(2+) concentration after hypoxia and treatment with NaN(3) or after application of high doses of NaN(3), Cl-IB-MECA immediately decreased the elevated intracellular Ca(2+) concentration toward the diastolic control level. The A(1)R agonist was ineffective. This may be especially important for the development of effective pharmacological agents, because mitochondrial dysfunction is a leading factor in the pathophysiological cascade of heart disease.

Adenosine A1 Receptor Agonists↗
PubMed1997

Pulmonary endothelial thiazine uptake: separation of cell surface reduction from intracellular reoxidation.

The objective of this study was to further evaluate the hypothesis that the accumulation of thiazine dyes, such as methylene blue, by cultured bovine pulmonary arterial endothelial cells involves reduction on the cell surface, followed by diffusion of the lipophilic reduced form of the dye into the cells and intracellular reoxidation to the relatively membrane-impermeant hydrophilic form. The specific question addressed was whether inhibition of methylene blue uptake by cyanide and azide is via inhibition of extracellular reduction or inhibition of intracellular reoxidation. We used the cell membrane-impermeant ferricyanide ion as a secondary electron acceptor to measure the extracellular reduction of methylene blue independently from its uptake by the cells. In addition, toluidine blue O, incorporated into an acrylamide polymer so that it could not permeate the cells in either its reduced or oxidized forms, was used to examine the effects of cyanide and azide on the extracellular reduction. Microscopic observations of the effect of the inhibitors on the intracellular accumulation of methylene blue were also made. The results indicate that the reduction and intracellular sequestration are separate processes and that, in doses that inhibited intracellular reoxidation, and therefore uptake and sequestration, neither cyanide nor azide had an inhibitory effect on extracellular reduction. The intracellular distribution of the observable oxidized form of the dye was consistent with oxidation of the reduced dye within subcellular organelles. The demonstration that extracellular reduction and intracellular sequestration are separate events is consistent with the hypothesized sequence of events.

Acrylic Resins↗
PubMed1977

H2O2 release from human granulocytes during phagocytosis. Relationship to superoxide anion formation and cellular catabolism of H2O2: studies with normal and cytochalasin B-treated cells.

Normal and cytochalasin B-treated human granulocytes have been studied to determine some of the interrelationships between phagocytosis-induced respiration and superoxide and hydrogen peroxide formation and release into the extracellular medium by intact cells. By using the scopoletin fluorescent assay to continuously monitor extracellular hydrogen peroxide concentrations during contact of cells with opsonized staphylococci, it was demonstrated that the superoxide scavengers ferricytochrome c and nitroblue tetrazolium significantly reduced the amount of H(2)O(2) released with time from normal cells but did not abolish it. This inhibitory effect was reversed by the simultaneous addition of superoxide dismutase (SOD), whereas the addition of SOD alone increased the amount of detectable H(2)O(2) in the medium. The addition of sodium azide markedly inhibited myeloperoxidase-H(2)O(2)-dependent protein iodination and more than doubled H(2)O(2) release, including the residual amount remaining after exposure of the cells to ferricytochrome c, suggesting its origin from an intracellular pool shared by several pathways for H(2)O(2) catabolism. When cells were pretreated with cytochalasin B and opsonized bacteria added, reduced oxygen consumption was observed, but this was in parallel to a reduction in specific binding of organisms to the cells when compared to normal. Under the influence of inhibited phagosome formation by cytochalasin B, the cells released an increased amount of superoxide and peroxide into the extracellular medium relative to oxygen consumption, and all detectable peroxide release could be inhibited by the addition of ferricytochrome c. Decreased H(2)O(2) production in the presence of this compound could not be ascribed to diminished bacterial binding, decreased oxidase activity, or increased H(2)O(2) catabolism and was reversed by the simultaneous addition of SOD. Furthermore, SOD and ferricytochrome c had similar effects on another H(2)O(2)-dependent reaction, protein iodination, in both normal and cytochalasin B cells. When oxygen consumption, O(2.) (-), and H(2)O(2) release were compared in the presence of azide under identical incubation conditions, the molar relationships for normal cells were 1.00:0.34:0.51 and for cytochalasin B-treated cells 1.00:0.99:0.40, respectively. Nonopsonized, or opsonized but disrupted, bacteria did not stimulate any of these metabolic functions. The results indicate that with normal cells approximately 50% of H(2)O(2) released during phagocytosis is derived directly from O(2.) (-) by dismutation, the remainder appearing from an (intra)cellular source shared by azide-inhibitable heme enzymes. With cytochalasin B treatment the evidence is consistent with the derivation of all H(2)O(2) from an O(2.) (-) precursor which is released from the cell surface. Furthermore, when activated by phagocytic particle binding, the neutrophil O(2.) (-) generating system appears to make more of this compound than can be accounted for by dismutation to H(2)O(2). This establishes conditions for the direct participation of both compounds in the microbicidal and cytocidal activity of these cells.

Azides↗