scholarly journals Cytosolic Fe-superoxide dismutase safeguardsTrypanosoma cruzifrom macrophage-derived superoxide radical

2019 ◽  
Vol 116 (18) ◽  
pp. 8879-8888 ◽  
Author(s):  
Alejandra Martínez ◽  
Carolina Prolo ◽  
Damián Estrada ◽  
Natalia Rios ◽  
María Noel Alvarez ◽  
...  
Keyword(s):  
Superoxide Radical ◽  
Parasite Burden ◽  
Anion Channel ◽  
Protonated Form ◽  
Superoxide Dismutases ◽  
Wild Type ◽  
Ph Values ◽  
Perhydroxyl Radical ◽  
Intracellular Proliferation

Trypanosoma cruzi, the causative agent of Chagas disease (CD), contains exclusively Fe-dependent superoxide dismutases (Fe-SODs). DuringT. cruziinvasion to macrophages, superoxide radical (O2•−) is produced at the phagosomal compartment toward the internalized parasite via NOX-2 (gp91-phox) activation. In this work,T. cruzicytosolic Fe-SODB overexpressers (pRIBOTEX–Fe-SODB) exhibited higher resistance to macrophage-dependent killing and enhanced intracellular proliferation compared with wild-type (WT) parasites. The higher infectivity of Fe-SODB overexpressers compared with WT parasites was lost in gp91-phox−/−macrophages, underscoring the role of O2•−in parasite killing. Herein, we studied the entrance of O2•−and its protonated form, perhydroxyl radical [(HO2•); pKa= 4.8], toT. cruziat the phagosome compartment. At the acidic pH values of the phagosome lumen (pH 5.3 ± 0.1), high steady-state concentrations of O2•−and HO2•were estimated (∼28 and 8 µM, respectively). Phagosomal acidification was crucial for O2•−permeation, because inhibition of the macrophage H+-ATPase proton pump significantly decreased O2•−detection in the internalized parasite. Importantly, O2•−detection, aconitase inactivation, and peroxynitrite generation were lower in Fe-SODB than in WT parasites exposed to external fluxes of O2•−or during macrophage infections. Other mechanisms of O2•−entrance participate at neutral pH values, because the anion channel inhibitor 5-nitro-2-(3-phenylpropylamino) benzoic acid decreased O2•−detection. Finally, parasitemia and tissue parasite burden in mice were higher in Fe-SODB–overexpressing parasites, supporting the role of the cytosolic O2•−-catabolizing enzyme as a virulence factor for CD.

scholarly journals Experimental Infection withSchistosoma mansoniin CCR5-Deficient Mice Is Associated with Increased Disease Severity, as CCR5 Plays a Role in Controlling Granulomatous Inflammation

2011 ◽  
Vol 79 (4) ◽  
pp. 1741-1749 ◽  
Author(s):  
Adriano L. S. Souza ◽  
Patrícia R. S. Souza ◽  
Cíntia A. Pereira ◽  
Adriana Fernandes ◽  
Rodrigo Guabiraba ◽  
...  
Keyword(s):  
Experimental Infection ◽  
Worm Burden ◽  
Granulomatous Inflammation ◽  
Parasite Burden ◽  
Mortality Rates ◽  
Wild Type ◽  
Schistosomiasis Mansoni ◽  
Cc Chemokines ◽  
Deficient Mice

ABSTRACTThe plasma level of the chemokine CCL3 is elevated in patients with chronic severe schistosomiasis mansoni. We have previously shown that CCL3−/−mice with experimental infection showed diminished pathology and worm burden compared to those of wild-type (WT) mice. To elucidate further the role of CC chemokines during schistosomiasis mansoni infection, we evaluated the course of infection in C57BL/6J mice deficient in CCR5, one of the receptors for CCL3. The CCR5 deficiency proved to be remarkably deleterious to the host, since mortality rates reached 70% at 14 weeks postinfection in CCR5−/−mice and 19% in WT mice. The increased lethality was not associated with an increased parasite burden, since similar numbers of eggs and adult worms were found in mice from both groups. Liver granulomas of chronically infected CCR5−/−mice were larger and showed greater numbers of cells and collagen deposition than liver granulomas from WT mice. This was associated with higher levels of production of intereleukin-5 (IL-5), IL-13, CCL3, and CCL5 in infected CCR5−/−mice than in infected WT mice. Moreover, at 8 weeks after infection, just before changes in pathology and mortality, the numbers of FoxP3-positive cells were lower in liver granulomas of CCR5−/−mice than in WT mice. In conclusion, the CCR5 deletion is deleterious to mice infected withSchistosoma mansoni, and this is associated with enhanced fibrosis and granulomatous inflammation.

Role of different Escherichia coli hydrogenases in H+ efflux and F1Fo-ATPase activity during glycerol fermentation at different pH values

2011 ◽  
Vol 31 (3) ◽  
pp. 179-184 ◽  
Author(s):  
Syuzanna Blbulyan ◽  
Arev Avagyan ◽  
Anna Poladyan ◽  
Armen Trchounian
Keyword(s):  
Escherichia Coli ◽  
Atpase Activity ◽  
Membrane Vesicle ◽  
Wild Type ◽  
Glucose Fermentation ◽  
Glycerol Fermentation ◽  
Whole Cells ◽  
Ph Values ◽  
F1fo Atpase

Escherichia coli is able to ferment glycerol and produce H2 by different Hyds (hydrogenases). Wild-type whole cells were shown to extrude H+ through the F1Fo-ATPase and by other means with a lower rate compared with that under glucose fermentation. At pH 7.5, H+ efflux was stimulated in fhlA mutant (with defective transcriptional activator of Hyd-3 or Hyd-4) and was lowered in hyaB or hybC mutants (with defective Hyd-1 or Hyd-2) and hyaB hybC double mutant; DCCD (dicyclohexylcarbodi-imide)-sensitive H+ efflux was observed. At pH 5.5, H+ efflux in wild-type was lower compared with that at pH 7.5; it was increased in fhlA mutant and absent in hyaB hybC mutant. Membrane vesicle ATPase activity was lower in wild-type glycerol-fermented cells at pH 7.5 compared with that in glucose-fermented cells; 100 mM K+ did not stimulate ATPase activity. The latter at pH 7.5, compared with that in wild–type, was lower in hyaB and less in hybC mutants, stimulated in the hyaB hybC mutant and suppressed in the fhlA mutant; DCCD inhibited ATPase activity. At pH 5.5, the ATPase activities of hyaB and hybC mutants had similar values and were higher compared with that in wild-type; ATPase activity was suppressed in hyaB hybC and fhlA mutants. The results indicate that during glycerol fermentation, H+ was expelled also via F1Fo. At pH 7.5 Hyd-1 and Hyd-2 but not FhlA or Hyd-4 might be related to F1Fo or have their own H+-translocating ability. At pH 5.5, both Hyd-1 and Hyd-2 more than F1Fo might be involved in H+ efflux.

scholarly journals Revisiting the fundamentals of p-nitrophenol analysis for its application in the quantification of lipases activity. A graphical update.

Uniciencia ◽  
2020 ◽  
Vol 34 (2) ◽  
pp. 31-43
Author(s):  
Randall Syedd-León ◽  
Manuel Sandoval-Barrantes ◽  
Humberto Trimiño-Vásquez ◽  
Luis Roberto Villegas-Peñaranda ◽  
Gerardo Rodríguez-Rodríguez
Keyword(s):  
Lipase Activity ◽  
Ph Value ◽  
Protonated Form ◽  
Spectrophotometric Analysis ◽  
Spectrophotometric Methods ◽  
Ph Values ◽  
Reaction Media ◽  
Activity Quantification

p-Nitrophenol (pNP) is a widely used compound for analytical determinations of several esterases (EC. 3.1.1.X), including lipases (E.C. 3.1.1.3). Most enzymatic measurements employ pNP derivatives such as esters, which are broken down by enzymatic hydrolysis, releasing pNP that is quantified by its absorbance at 410 nm. Although this type of methods was developed a few decades ago, the spectrophotometric analysis of pNP requires analytical measurements of pH and temperature to achieve reliable determinations. The aim of this paper is to offer a graphical update of how pH and temperature affect the p-nitrophenol absorbance at different wavelengths in lipase emulsified media, due to its relevance for the quantitative determination of lipase activity using spectrophotometric methods. To highlight the importance of each variable involved in this analysis, we dissolved pNP in emulsified media (for lipase activity quantification) at several pH values from 4.00 to 11.00, and measured its absorbance in a range of 270 nm – 500 nm and at several temperatures from 25°C to 50°C. The absorption patterns of pNP under the established conditions were graphed in 3D plots. The constructed 3D plots showed that, regardless of the temperature, below pH 6.00, pNP predominantly absorbs at 317 nm, due to the greater abundance of its protonated form, which is completely predominant at pH 3.50 and below. On the other hand, at pH 10.0 and above, the major absorption occurs at about 401 nm, confirming that the equilibrium is completely shifted to the pNP anionic form. These results also indicate that close to neutral pH value pNP, it displays a temperature dependence effect, increasing absorbance to 410 nm at higher temperatures. Due to many analytical determinations of enzymatic activities, the release of pNP is carried around pH 7.00. It is necessary to consider the determinant role of both pH and temperature over these measurements, how these variables must be strictly controlled, and how the calibration curves and blanks should take the reaction media pH and temperature into account.

The direct synthesis of phospho enol pyruvate from pyruvate by Escherichia coli

1967 ◽  
Vol 168 (1012) ◽  
pp. 263-280 ◽  
Keyword(s):  
Escherichia Coli ◽  
Inorganic Phosphate ◽  
Direct Synthesis ◽  
Physiological Role ◽  
Wild Type ◽  
Pyruvate Kinase Activity ◽  
Ph Values ◽  
Direct Formation ◽  
Pyruvate Synthase

Extracts of Escherichia coli are shown to contain an enzyme system which in the presence of Mg 2+ catalyses the direct formation of phospho enol pyruvate from pyruvate and ATP with concomitant formation of AMP and inorganic phosphate. This enzyme, which has been designated 'phospho enol pyruvate synthase' ( PEP -synthase) has been purified 80-fold and is free of pyruvate kinase activity; PEP synthesis proceeded most rapidly at pH 8 to 8.5. At pH values between 6.2 and 7.5 the enzyme can catalyse the formation of ATP and pyruvate from PEP , AMP and inorganic phosphate; if arsenate is used instead of phosphate, pyruvate and ADP are produced instead. Studies of the enzymic formation of PEP with ATP specifically labelled with 32 P, and of the reverse reaction with [U -14 C] AMP , suggest that the PEP -synthase reaction involves the transfer of a pyrophosphoryl-group. The physiological role of PEP -synthase has been demonstrated with mutants of E. coli devoid of the enzyme: in contrast to wild-type organisms, such mutants neither grow on pyruvate, lactate or alanine, nor form glycogen from lactate. It is thus concluded that PEP -synthase plays an important role in the anaplerotic and the biosynthetic reactions which enable the organisms to grow on pyruvate as sole carbon source.

scholarly journals Ablation of PLB exacerbates ischemic injury to a lesser extent in female than male mice: protective role of NO

2003 ◽  
Vol 284 (2) ◽  
pp. H683-H690 ◽  
Author(s):  
Heather R. Cross ◽  
Evangelia G. Kranias ◽  
Elizabeth Murphy ◽  
Charles Steenbergen
Keyword(s):  
Ischemic Injury ◽  
Protective Role ◽  
Ischemia And Reperfusion ◽  
Wild Type ◽  
No Donor ◽  
Ph Values ◽  
Nos Inhibitor ◽  
Males And Females ◽  
Phospholamban Phosphorylation

Recent studies suggest a role for phospholamban phosphorylation during ischemia and reperfusion. The role of phospholamban in ischemia was studied by subjecting hearts from male and female wild-type (MWT/FWT) and phospholamban-knockout (MKO/FKO) mice to 20 min of ischemia-40 min of reperfusion while 31P NMR spectra were acquired. ATP and pH values fell lower during ischemia, and postischemic contractility was less, in MKO and FKO versus WT hearts. After shorter ischemia (15 min), recoveries of contraction, ATP, and pH were greater in FKO than MKO hearts. To examine the role of nitric oxide (NO) synthases (NOS) in the protection in FKO versus MKO hearts, we utilized 1 μMl-NAME, a NOS inhibitor, or 100 μM S-nitroso- N-acetylpenicillamine (SNAP), an NO donor. Recoveries of function, ATP, and pH were less inl-NAME-treated FKO than untreated FKO hearts and greater in SNAP-treated MKO than untreated MKO hearts. In conclusion, phospholamban ablation increased ischemic injury in both males and females; however, female hearts were less susceptible than male hearts. Protection in females was decreased by a NOS inhibitor and mimicked in males by an NO donor, implying that protection was NOS mediated.

scholarly journals Periplasmic Superoxide Dismutase in Meningococcal Pathogenicity

1998 ◽  
Vol 66 (1) ◽  
pp. 213-217 ◽  
Author(s):  
Kathryn E. Wilks ◽  
Kate L. R. Dunn ◽  
Jayne L. Farrant ◽  
Karen M. Reddin ◽  
Andrew R. Gorringe ◽  
...  
Keyword(s):  
Superoxide Dismutase ◽  
Free Radicals ◽  
Superoxide Radical ◽  
Infection Model ◽  
Wild Type ◽  
Superoxide Radical Anion ◽  
Mouse Infection Model ◽  
Increased Sensitivity ◽  
Hydroxyl Free Radicals

ABSTRACT Meningococcal sodC encodes periplasmic copper- and zinc-cofactored superoxide dismutase (Cu,Zn SOD) which catalyzes the conversion of the superoxide radical anion to hydrogen peroxide, preventing a sequence of reactions leading to production of toxic hydroxyl free radicals. From its periplasmic location, Cu,Zn SOD was inferred to acquire its substrate from outside the bacterial cell and was speculated to play a role in preserving meningococci from the action of microbicidal oxygen free radicals produced in the context of host defense. A sodC mutant was constructed by allelic exchange and was used to investigate the role of Cu,Zn SOD in pathogenicity. Wild-type and mutant meningococci grew at comparable rates and survived equally long in aerobic liquid culture. The mutant showed no increased sensitivity to paraquat, which generates superoxide within the cytosol, but was approximately 1,000-fold more sensitive to the toxicity of superoxide generated in solution by the xanthine/xanthine oxidase system. These data support a role for meningococcal Cu,Zn SOD in protection against exogenous superoxide. In experiments to translate this into a role in pathogenicity, wild-type and mutant organisms were used in an intraperitoneal mouse infection model. The sodC mutant was significantly less virulent. We conclude that periplasmic Cu,Zn SOD contributes to the virulence ofNeisseria meningitidis, most likely by reducing the effectiveness of toxic oxygen host defenses.

Differential Role of Components of the Fibrinolytic System in the Formation and Removal of Thrombus Induced by Endothelial Injury

1999 ◽  
Vol 81 (04) ◽  
pp. 601-604 ◽  
Author(s):  
Hiroyuki Matsuno ◽  
Osamu Kozawa ◽  
Masayuki Niwa ◽  
Shigeru Ueshima ◽  
Osamu Matsuo ◽  
...  
Keyword(s):  
Plasminogen Activator ◽  
Thrombus Formation ◽  
Endothelial Injury ◽  
Fibrinolytic System ◽  
Tissue Type ◽  
Clot Lysis ◽  
Wild Type ◽  
Type Plasminogen Activator ◽  
Pai 1

SummaryThe role of fibrinolytic system components in thrombus formation and removal in vivo was investigated in groups of six mice deficient in urokinase-type plasminogen activator (u-PA), tissue-type plasminogen activator (t-PA), or plasminogen activator inhibitor-1 (PAI-1) (u-PA-/-, t-PA-/- or PAI-1-/-, respectively) or of their wild type controls (u-PA+/+, t-PA+/+ or PAI-1+/+). Thrombus was induced in the murine carotid artery by endothelial injury using the photochemical reaction between rose bengal and green light (540 nm). Blood flow was continuously monitored for 90 min on day 0 and for 20 min on days 1, 2 and 3. The times to occlusion after the initiation of endothelial injury in u-PA+/+, t-PA+/+ or PAI-1+/+ mice were 9.4 ± 1.3, 9.8 ± 1.1 or 9.7 ± 1.6 min, respectively. u-PA-/- and t-PA-/- mice were indistinguishable from controls, whereas that of PAI-1-/- mice were significantly prolonged (18.4 ± 3.7 min). Occlusion persisted for the initial 90 min observation period in 10 of 18 wild type mice and was followed by cyclic reflow and reocclusion in the remaining 8 mice. At day 1, persistent occlusion was observed in 1 wild type mouse, 8 mice had cyclic reflow and reocclusion and 9 mice had persistent reflow. At day 2, all injured arteries had persistent reflow. Persistent occlusion for 90 min on day 0 was observed in 3 u-PA-/-, in all t-PA-/- mice at day 1 and in 2 of the t-PA-/-mice at day 2 (p <0.01 versus wild type mice). Persistent patency was observed in all PAI-1-/- mice at day 1 and in 5 of the 6 u-PA-/- mice at day 2 (both p <0.05 versus wild type mice). In conclusion, t-PA increases the rate of clot lysis after endothelial injury, PAI-1 reduces the time to occlusion and delays clot lysis, whereas u-PA has little effect on thrombus formation and spontaneous lysis.

The Role of Amplified Wild-Type Neu in the Etiology of Breast Cancer

2003 ◽  
Author(s):  
Michael N. Gould
Keyword(s):  
Breast Cancer ◽  
Wild Type

An ESR study of the peroxidase reaction catalyzed by human methaemoglobin and methaemoglobin-haptoglobin complex

1991 ◽  
Vol 56 (4) ◽  
pp. 923-932
Author(s):  
Jana Stejskalová ◽  
Pavel Stopka ◽  
Zdeněk Pavlíček
Keyword(s):  
Hydrogen Peroxide ◽  
Ascorbic Acid ◽  
Amino Acid ◽  
Peroxidase Activity ◽  
Amino Acid Analysis ◽  
Superoxide Radical ◽  
Esr Spectra ◽  
The Other ◽  
Delocalized Electron

The ESR spectra of peroxidase systems of methaemoglobin-ascorbic acid-hydrogen peroxide and methaemoglobin-haptoglobin complex-ascorbic acid-hydrogen peroxide have been measured in the acetate buffer of pH 4.5. For the system with methaemoglobin an asymmetrical signal with g ~ 2 has been observed which is interpreted as the perpendicular region of anisotropic spectrum of superoxide radical. On the other hand, for the system with methaemoglobin-haptoglobin complex the observed signal with g ~ 2 is symmetrical and is interpreted as a signal of delocalized electron. After realization of three repeatedly induced peroxidase processes the ESR signal of the perpendicular part of anisotropic spectrum of superoxide radical is distinctly diminished, whereas the signal of delocalized electron remains practically unchanged. An amino acid analysis of methaemoglobin along with results of the ESR measurements make it possible to derive a hypothesis about the role of haptoglobin in increasing of the peroxidase activity of methaemoglobin.

scholarly journals ERK activation increases nitroprusside induced apoptosis in  human melanoma  cells  irrespective of p53 status: Role of  superoxide dismutases

2009 ◽  
Vol 8 (12) ◽  
pp. 1173-1182 ◽  
Author(s):  
Luis Alberto Gomez–Sarosi ◽  
Mary Strasberg–Rieber ◽  
Manuel Rieber
Keyword(s):  
Melanoma Cells ◽  
Human Melanoma ◽  
Superoxide Dismutases ◽  
Erk Activation ◽  
Human Melanoma Cells ◽  
P53 Status ◽  
Induced Apoptosis
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