The Experts below are selected from a list of 1410 Experts worldwide ranked by ideXlab platform
Jacek Zielonka - One of the best experts on this subject based on the ideXlab platform.
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HPLC-Based Monitoring of Oxidation of Hydroethidine for the Detection of NADPH Oxidase-Derived Superoxide Radical Anion.
Methods in Molecular Biology, 2019Co-Authors: Jacek Zielonka, Monika Zielonka, Balaraman KalyanaramanAbstract:Hydroethidine is a fluorogenic probe that in the presence of the superoxide radical anion is oxidized to a red fluorescent product, 2-hydroxyethidium. In cells, Hydroethidine is also oxidized to other products, including red fluorescent ethidium. Thus, selective monitoring of 2-hydroxyethidium is required for specific detection of the superoxide radical anion. Here, we provide protocols for HPLC- and LC-MS-based quantitation of 2-hydroxyethidium, among other oxidation products. Also, a protocol for continuous sampling for real-time monitoring of superoxide production using rapid HPLC measurements of 2-hydroxyethidium is described.
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the pulse radiolytic study of one electron oxidation of hydropropidine and Hydroethidine implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radoslaw Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Andrzej Marcinek, Jakub Pieta, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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The pulse radiolytic study of one-electron oxidation of hydropropidine and Hydroethidine – implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radosław Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Jakub Pięta, Andrzej Marcinek, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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N,N.N′,N′-tetramethylHydroethidine (TMHE) - in search for better probes for the detection of superoxide radical anion
Free Radical Biology and Medicine, 2017Co-Authors: Adam Sikora, Micael Hardy, Jacek Zielonka, Olivier Ouari, Radosław Michalski, Andrzej Marcinek, Jan Adamus, Balaraman KalyanaramanAbstract:The superoxide radical anion is an important reactive species produced by one-electron reduction of molecular oxygen. The main sources of superoxide radical anion in vivo are the enzymes NADPH oxidases (NOX) and mitochondria (mitochondrial electron transport chain). Superoxide radical anion reacts rapidly with nitric oxide with the formation of peroxynitrite (ONOO−). It can also undergo spontaneous or SOD-catalyzed dismutation with the formation of hydrogen peroxide (H2O2) and thus it is a precursor of other biologically relevant oxidants playing an important role in various pathologies. Due to its reactivity and short lifetime in vivo, its detection and quantitation is difficult and demands special and sensitive techniques. One of the approaches is the use of fluorogenic probes, the compounds which themselves are not fluorescent but in the reaction with superoxide are oxidized to the fluorescent products that can be directly detected. Among various profluorescent probes available Hydroethidine (HE) seems to be a gold standard for detection of superoxide in biological systems. In the presence of superoxide radical anion, HE undergoes oxidative transformation into 2-hydroxyethidium (2-OH-E+), a specific marker of superoxide radical anion production. In the reaction with other radical, one-electron oxidants, HE is oxidatively transformed into ethidine and dimeric products. Here we present a spectroscopic and chemical characterization of new analogue of Hydroethidine - N,N,N,N-tetramethylHydroethidine (TMHE).
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A Critical Review of Methodologies to Detect Reactive Oxygen and Nitrogen Species Stimulated by NADPH Oxidase Enzymes: Implications in Pesticide Toxicity
Current Pharmacology Reports, 2016Co-Authors: Balaraman Kalyanaraman, Micael Hardy, Jacek ZielonkaAbstract:In this review, potential fluorescent probe applications for detecting reactive oxygen and nitrogen species (ROS/RNS) generated from NADPH oxidases (e.g., Nox2) and nitric oxide synthase enzymes are discussed in the context of pesticide toxicology. Identification of the specific marker products derived from the interaction between ROS/RNS and the fluorescent probes (e.g., Hydroethidine and coumarin boronate) is critical. Due to the complex nature of reactions between the probes and ROS/RNS, we suggest avoiding the use of fluorescence microscopy for detecting oxidizing/nitrating species. We also critically examined the viability of using radiolabeling or positron emission tomography (PET) for ROS/RNS detection. Although these techniques differ in sensitivity and detection modalities, the chemical mechanism governing the reaction between these probes and ROS/RNS should remain the same. To unequivocally detect superoxide with these probes (i.e., radiolabeled and PET-labeled Hydroethidine analogs), the products should be isolated and characterized by LC-MS/MS or HPLC using an appropriate standard.
Balaraman Kalyanaraman - One of the best experts on this subject based on the ideXlab platform.
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HPLC-Based Monitoring of Oxidation of Hydroethidine for the Detection of NADPH Oxidase-Derived Superoxide Radical Anion.
Methods in Molecular Biology, 2019Co-Authors: Jacek Zielonka, Monika Zielonka, Balaraman KalyanaramanAbstract:Hydroethidine is a fluorogenic probe that in the presence of the superoxide radical anion is oxidized to a red fluorescent product, 2-hydroxyethidium. In cells, Hydroethidine is also oxidized to other products, including red fluorescent ethidium. Thus, selective monitoring of 2-hydroxyethidium is required for specific detection of the superoxide radical anion. Here, we provide protocols for HPLC- and LC-MS-based quantitation of 2-hydroxyethidium, among other oxidation products. Also, a protocol for continuous sampling for real-time monitoring of superoxide production using rapid HPLC measurements of 2-hydroxyethidium is described.
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the pulse radiolytic study of one electron oxidation of hydropropidine and Hydroethidine implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radoslaw Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Andrzej Marcinek, Jakub Pieta, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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The pulse radiolytic study of one-electron oxidation of hydropropidine and Hydroethidine – implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radosław Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Jakub Pięta, Andrzej Marcinek, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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N,N.N′,N′-tetramethylHydroethidine (TMHE) - in search for better probes for the detection of superoxide radical anion
Free Radical Biology and Medicine, 2017Co-Authors: Adam Sikora, Micael Hardy, Jacek Zielonka, Olivier Ouari, Radosław Michalski, Andrzej Marcinek, Jan Adamus, Balaraman KalyanaramanAbstract:The superoxide radical anion is an important reactive species produced by one-electron reduction of molecular oxygen. The main sources of superoxide radical anion in vivo are the enzymes NADPH oxidases (NOX) and mitochondria (mitochondrial electron transport chain). Superoxide radical anion reacts rapidly with nitric oxide with the formation of peroxynitrite (ONOO−). It can also undergo spontaneous or SOD-catalyzed dismutation with the formation of hydrogen peroxide (H2O2) and thus it is a precursor of other biologically relevant oxidants playing an important role in various pathologies. Due to its reactivity and short lifetime in vivo, its detection and quantitation is difficult and demands special and sensitive techniques. One of the approaches is the use of fluorogenic probes, the compounds which themselves are not fluorescent but in the reaction with superoxide are oxidized to the fluorescent products that can be directly detected. Among various profluorescent probes available Hydroethidine (HE) seems to be a gold standard for detection of superoxide in biological systems. In the presence of superoxide radical anion, HE undergoes oxidative transformation into 2-hydroxyethidium (2-OH-E+), a specific marker of superoxide radical anion production. In the reaction with other radical, one-electron oxidants, HE is oxidatively transformed into ethidine and dimeric products. Here we present a spectroscopic and chemical characterization of new analogue of Hydroethidine - N,N,N,N-tetramethylHydroethidine (TMHE).
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A Critical Review of Methodologies to Detect Reactive Oxygen and Nitrogen Species Stimulated by NADPH Oxidase Enzymes: Implications in Pesticide Toxicity
Current Pharmacology Reports, 2016Co-Authors: Balaraman Kalyanaraman, Micael Hardy, Jacek ZielonkaAbstract:In this review, potential fluorescent probe applications for detecting reactive oxygen and nitrogen species (ROS/RNS) generated from NADPH oxidases (e.g., Nox2) and nitric oxide synthase enzymes are discussed in the context of pesticide toxicology. Identification of the specific marker products derived from the interaction between ROS/RNS and the fluorescent probes (e.g., Hydroethidine and coumarin boronate) is critical. Due to the complex nature of reactions between the probes and ROS/RNS, we suggest avoiding the use of fluorescence microscopy for detecting oxidizing/nitrating species. We also critically examined the viability of using radiolabeling or positron emission tomography (PET) for ROS/RNS detection. Although these techniques differ in sensitivity and detection modalities, the chemical mechanism governing the reaction between these probes and ROS/RNS should remain the same. To unequivocally detect superoxide with these probes (i.e., radiolabeled and PET-labeled Hydroethidine analogs), the products should be isolated and characterized by LC-MS/MS or HPLC using an appropriate standard.
Adam Sikora - One of the best experts on this subject based on the ideXlab platform.
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the pulse radiolytic study of one electron oxidation of hydropropidine and Hydroethidine implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radoslaw Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Andrzej Marcinek, Jakub Pieta, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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The pulse radiolytic study of one-electron oxidation of hydropropidine and Hydroethidine – implications for the detection of superoxide radical anion in biological systems
Free Radical Biology and Medicine, 2018Co-Authors: Radosław Michalski, Jacek Zielonka, Balaraman Kalyanaraman, Bartosz Michalowski, Jakub Pięta, Andrzej Marcinek, Adam SikoraAbstract:Hydroethidine (HE) is a fluorogenic probe widely used for the detection of superoxide radical anion in cellular studies. Recently, we have synthesized cell-impermeant analogue of HE called hydropropidine (HPr +). HPr+ is a water-soluble molecule, possessing highly localized positive charge on the nitrogen atom of the alkyl group, that prevents its cellular uptake. Analogously to its precursor, HPr+ reacts with superoxide forming a specific fluorescent product 2-hydroxypropidine (2-OH-Pr++). According to the proposed mechanism of superoxide-mediated oxidation of HE and HPr+, the probe undergoes one-electron oxidation to the radical cation, and next, in the presence of superoxide, is converted to the appropriate hydroxylation product. Here, we show the spectroscopic characetrization of radical cation of HPr+ and HE by pulse radiolysis technique. The reactivity of both probes toward biologically relevant radicals, e.g. nitrogen dioxide (•NO2) and carbonate (CO3•−) radicals (products of peroxynitrite decomposition), as well as glutathionyl radicals (GS•), will be presented. These results will be discussed in relation to the mechanism of oxidation of Hydroethidine based probes in biological milieu.
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N,N.N′,N′-tetramethylHydroethidine (TMHE) - in search for better probes for the detection of superoxide radical anion
Free Radical Biology and Medicine, 2017Co-Authors: Adam Sikora, Micael Hardy, Jacek Zielonka, Olivier Ouari, Radosław Michalski, Andrzej Marcinek, Jan Adamus, Balaraman KalyanaramanAbstract:The superoxide radical anion is an important reactive species produced by one-electron reduction of molecular oxygen. The main sources of superoxide radical anion in vivo are the enzymes NADPH oxidases (NOX) and mitochondria (mitochondrial electron transport chain). Superoxide radical anion reacts rapidly with nitric oxide with the formation of peroxynitrite (ONOO−). It can also undergo spontaneous or SOD-catalyzed dismutation with the formation of hydrogen peroxide (H2O2) and thus it is a precursor of other biologically relevant oxidants playing an important role in various pathologies. Due to its reactivity and short lifetime in vivo, its detection and quantitation is difficult and demands special and sensitive techniques. One of the approaches is the use of fluorogenic probes, the compounds which themselves are not fluorescent but in the reaction with superoxide are oxidized to the fluorescent products that can be directly detected. Among various profluorescent probes available Hydroethidine (HE) seems to be a gold standard for detection of superoxide in biological systems. In the presence of superoxide radical anion, HE undergoes oxidative transformation into 2-hydroxyethidium (2-OH-E+), a specific marker of superoxide radical anion production. In the reaction with other radical, one-electron oxidants, HE is oxidatively transformed into ethidine and dimeric products. Here we present a spectroscopic and chemical characterization of new analogue of Hydroethidine - N,N,N,N-tetramethylHydroethidine (TMHE).
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Mitigation of NADPH Oxidase 2 Activity as a Strategy to Inhibit Peroxynitrite Formation
Journal of Biological Chemistry, 2016Co-Authors: Jacek Zielonka, Micael Hardy, Monika Zielonka, Olivier Ouari, Gang Cheng, Adam Sikora, Lynn Verplank, Mehmet Menaf Ayhan, Radosław Podsiadły, J LambethAbstract:Using high throughput screening-compatible assays for superoxide and hydrogen peroxide, we identified potential inhibitors of the NADPH oxidase (Nox2) isoform from a small library of bioactive compounds. By using multiple probes (Hydroethidine, hydropropidine, Amplex Red, and coumarin boronate) with well defined redox chemistry that form highly diagnostic marker products upon reaction with superoxide (O2 ()), hydrogen peroxide (H2O2), and peroxynitrite (ONOO(-)), the number of false positives was greatly decreased. Selected hits for Nox2 were further screened for their ability to inhibit ONOO(-)formation in activated macrophages. A new diagnostic marker product for ONOO(-)is reported. We conclude that the newly developed high throughput screening/reactive oxygen species assays could also be used to identify potential inhibitors of ONOO(-)formed from Nox2-derived O2 ()and nitric oxide synthase-derived nitric oxide.
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Mitigation of NADPH Oxidase 2 Activity as a Strategy to Inhibit Peroxynitrite Formation
Journal of Biological Chemistry, 2016Co-Authors: Jacek Zielonka, Micael Hardy, Monika Zielonka, Olivier Ouari, Gang Cheng, Adam Sikora, Lynn Verplank, Mehmet Menaf Ayhan, Radosław Podsiadły, J LambethAbstract:Using a high-throughput screening (HTS)-compatible assays for superoxide and hydrogen peroxide, we identified potential inhibitors of NADPH oxidase (Nox2) isoform from a small library of bioactive compounds. By using multiple probes (Hydroethidine, hydropropidine, Amplex Red, and coumarin boronate) with well-defined redox chemistry that form highly diagnostic marker products upon reaction with superoxide (O 2 • –), hydrogen peroxide (H 2 O 2), and peroxynitrite (ONOO –), the number of false positives was greatly decreased. Selected hits for Nox2 were further screened for their ability to inhibit ONOO – formation in activated macrophages. New diagnostic marker product for ONOO – is reported. We conclude that the newly developed HTS/ROS assays could also be used to identify potential inhibitors of ONOO – formed from Nox2-derived O 2 • – and nitric oxide synthase (NOS)-derived nitric oxide.
Yi Xiao - One of the best experts on this subject based on the ideXlab platform.
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are Hydroethidine based probes reliable for reactive oxygen species detection
Antioxidants & Redox Signaling, 2019Co-Authors: Yi Xiao, David MeierhoferAbstract:Detection and quantification of the highly reactive and short-lived superoxide (O•2-) can be challenging. Here, we present a new mass spectrometry (MS)-based method to detect and quantify O•2- using three fluorogenic Hydroethidine probes: Hydroethidine (HE), mito-Hydroethidine (mito-HE), and hydropropidine (HPr+), which measure cytosolic, mitochondrial, and extracellular O•2-, respectively. The probes and their oxidation products were simultaneously quantified by applying multiple reaction monitoring (MRM) with MS that allowed the specific measurement of reactive oxygen species (ROS) distribution within the cell. The advantage of this liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is that coeluting compounds can be precisely distinguished using specific precursor and fragment masses. This method overcomes limitations from spectral overlap of O•2--specific and nonspecific products in fluorescence spectra or the low specificity associated with chromatography-based approaches. However, our experiments showed that these HE probes can be prone to autoxidation during incubation at 37°C in Hank's solution. Cell treatments with strong oxidants did not significantly increase levels of the O•2- radical. Thus, subtle changes in ROS levels in cell culture experiments might not be quantifiable. Our findings raise the question of whether HE-based probes can be used for the reliable detection of O•2- radicals in cell culture. Antioxid. Redox Signal. 00, 000-000.
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Are Hydroethidine-based probes reliable for ROS detection?
Antioxidants & Redox Signaling, 2018Co-Authors: Yi Xiao, David MeierhoferAbstract:Detection and quantification of the highly reactive and short-lived superoxide (O•2-) can be challenging. Here, we present a new mass spectrometry (MS)-based method to detect and quantify O•2- using three fluorogenic Hydroethidine probes: Hydroethidine (HE), mito-Hydroethidine (mito-HE), and hydropropidine (HPr+), which measure cytosolic, mitochondrial, and extracellular O•2-, respectively. The probes and their oxidation products were simultaneously quantified by applying multiple reaction monitoring (MRM) with MS that allowed the specific measurement of reactive oxygen species (ROS) distribution within the cell. The advantage of this liquid chromatography-tandem mass spectrometry (LC-MS/MS) method is that coeluting compounds can be precisely distinguished using specific precursor and fragment masses. This method overcomes limitations from spectral overlap of O•2--specific and nonspecific products in fluorescence spectra or the low specificity associated with chromatography-based approaches. However, our experiments showed that these HE probes can be prone to autoxidation during incubation at 37°C in Hank's solution. Cell treatments with strong oxidants did not significantly increase levels of the O•2- radical. Thus, subtle changes in ROS levels in cell culture experiments might not be quantifiable. Our findings raise the question of whether HE-based probes can be used for the reliable detection of O•2- radicals in cell culture. Antioxid. Redox Signal. 00, 000-000.
Costantino Iadecola - One of the best experts on this subject based on the ideXlab platform.
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aβ induced vascular oxidative stress and attenuation of functional hyperemia in mouse somatosensory cortex
Journal of Cerebral Blood Flow and Metabolism, 2004Co-Authors: Laibaik Park, Josef Anrather, Colleen L Forster, Ken Kazama, George A Carlson, Costantino IadecolaAbstract:We investigated the role of vascular oxidative stress in the mechanisms of the impairment in cerebrovascular regulation produced by the amyloid-β peptide (Aβ). In particular, we sought to provide evidence of vascular oxidative stress in mice overexpressing the amyloid precursor protein (APP) and to determine whether the Aβ-induced attenuation in functional hyperemia is mediated by free radical overproduction. Oxidative/nitrosative stress was assessed by 3-nitrotyrosine immunoreactivity, while free radical production was determined in cerebral microvessels by Hydroethidine microfluorography. To study functional hyperemia the somatosensory cortex was activated by whisker stimulation while local blood flow was monitored by laser-Doppler flowmetry. It was found that APP mice show signs of oxidative/nitrosative stress in pial and intracerebral blood vessels well before they develop oxidative stress in neurons and glia or amyloid plaques. Treatment of cerebral microvessels isolated from wild-type mice with Aβ (1 μM) increased free radical production as assessed by the Hydroethidine technique. The Aβ-induced attenuation of the increase in somatosensory cortex blood flow produced by whisker stimulation was prevented by treatment with the free radical scavengers MnTBAP or tiron. These data provide evidence that in APP mice vascular oxidative stress precedes the development of parenchymal oxidative stress, and that Aβ-produced vascular reactive oxygen species are involved in the attendant attenuation in functional hyperemia. Thus, vascular oxidative stress is an early event in the course of the brain dysfunction produced by APP overexpression and Aβ, and, as such, could be the target of early therapeutic interventions based on antioxidants.
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Aβ-Induced Vascular Oxidative Stress and Attenuation of Functional Hyperemia in Mouse Somatosensory Cortex
Journal of Cerebral Blood Flow & Metabolism, 2004Co-Authors: Laibaik Park, Josef Anrather, Ken Kazama, George A Carlson, Colleen Forster, Costantino IadecolaAbstract:We investigated the role of vascular oxidative stress in the mechanisms of the impairment in cerebrovascular regulation produced by the amyloid-beta peptide (Abeta). In particular, we sought to provide evidence of vascular oxidative stress in mice overexpressing the amyloid precursor protein (APP) and to determine whether the Abeta-induced attenuation in functional hyperemia is mediated by free radical overproduction. Oxidative/nitrosative stress was assessed by 3-nitrotyrosine immunoreactivity, while free radical production was determined in cerebral microvessels by Hydroethidine microfluorography. To study functional hyperemia the somatosensory cortex was activated by whisker stimulation while local blood flow was monitored by laser-Doppler flowmetry. It was found that APP mice show signs of oxidative/nitrosative stress in pial and intracerebral blood vessels well before they develop oxidative stress in neurons and glia or amyloid plaques. Treatment of cerebral microvessels isolated from wild-type mice with Abeta (1 microM) increased free radical production as assessed by the Hydroethidine technique. The Abeta-induced attenuation of the increase in somatosensory cortex blood flow produced by whisker stimulation was prevented by treatment with the free radical scavengers MnTBAP or tiron. These data provide evidence that in APP mice vascular oxidative stress precedes the development of parenchymal oxidative stress, and that Abeta-produced vascular reactive oxygen species are involved in the attendant attenuation in functional hyperemia. Thus, vascular oxidative stress is an early event in the course of the brain dysfunction produced by APP overexpression and Abeta, and, as such, could be the target of early therapeutic interventions based on antioxidants.
