What does the commonly used DCF test for oxidative stress really show?

2010 ◽  
Vol 428 (2) ◽  
pp. 183-190 ◽  
Author(s):  
Markus Karlsson ◽  
Tino Kurz ◽  
Ulf T. Brunk ◽  
Sven E. Nilsson ◽  
Christina I. Frennesson
Keyword(s):  
Oxidative Stress ◽  
Cytochrome C ◽  
Enzymatic Oxidation ◽  
Short Exposure ◽  
Outer Side ◽  
Inner Mitochondrial Membrane ◽  
Fluorescent Compound ◽  
Redox Active ◽  
J774 Cells ◽  
Active Transition

H2DCF-DA (dihydrodichlorofluorescein diacetate) is widely used to evaluate ‘cellular oxidative stress’. After passing through the plasma membrane, this lipophilic and non-fluorescent compound is de-esterified to a hydrophilic alcohol [H2DCF (dihydrodichlorofluorescein)] that may be oxidized to fluorescent DCF (2′,7′-dichlorofluorescein) by a process usually considered to involve ROS (reactive oxygen species). It is, however, not always recognized that, being a hydrophilic molecule, H2DCF does not cross membranes, except for the outer fenestrated mitochondrial ones. It is also not generally realized that oxidation of H2DCF is dependent either on Fenton-type reactions or on unspecific enzymatic oxidation by cytochrome c, for neither superoxide, nor H2O2, directly oxidizes H2DCF. Consequently, oxidation of H2DCF requires the presence of either cytochrome c or of both redox-active transition metals and H2O2. Redox-active metals exist mainly within lysosomes, whereas cytochrome c resides bound to the outer side of the inner mitochondrial membrane. Following exposure to H2DCF-DA, weak mitochondrial fluorescence was found in both the oxidation-resistant ARPE-19 cells and the much more sensitive J774 cells. This fluorescence was only marginally enhanced following short exposure to H2O2, showing that by itself it is unable to oxidize H2DCF. Cells that were either exposed to the lysosomotropic detergent MSDH (O-methylserine dodecylamide hydrochloride), exposed to prolonged oxidative stress, or spontaneously apoptotic showed lysosomal permeabilization and strong DCF-induced fluorescence. The results suggest that DCF-dependent fluorescence largely reflects relocation to the cytosol of lysosomal iron and/or mitochondrial cytochrome c.

scholarly journals Alzheimer Disease and Oxidative Stress

2002 ◽  
Vol 2 (3) ◽  
pp. 120-123 ◽  
Author(s):  
George Perry ◽  
Adam D. Cash ◽  
Mark A. Smith
Keyword(s):  
Oxidative Stress ◽  
Alzheimer Disease ◽  
Senile Plaques ◽  
Compelling Evidence ◽  
Cellular Changes ◽  
Redox Active ◽  
Oxidative Processes ◽  
Active Transition ◽  
Initial Source ◽  
And Oxidative Stress

Research in Alzheimer disease has recently demonstrated compelling evidence on the importance of oxidative processes in its pathogenesis. Cellular changes show that oxidative stress is an event that precedes the appearance of the hallmark pathologies of the disease, neurofibrillary tangles, and senile plaques. While it is still unclear what the initial source of the oxidative stress is in Alzheimer disease, it is likely that the process is highly dependent on redox-active transition metals such as iron and copper. Further investigation into the role that oxidative stress mechanisms seem to play in the pathogenesis of Alzheimer disease may lead to novel clinical interventions.

scholarly journals Spatial Organization of the Redox Active Centers in the Bovine Heart Ubiquinol-cytochrome c Oxidoreductase

1989 ◽  
Vol 264 (2) ◽  
pp. 735-744
Author(s):  
T Ohnishi ◽  
H Schägger ◽  
S W Meinhardt ◽  
R LoBrutto ◽  
T A Link ◽  
...  
Keyword(s):  
Cytochrome C ◽  
Spatial Organization ◽  
Active Centers ◽  
Bovine Heart ◽  
Redox Active

Redox-Active Quinones and Ascorbate: An Innovative Cancer Therapy That Exploits the Vulnerability of Cancer Cells to Oxidative Stress

2011 ◽  
Vol 11 (2) ◽  
pp. 213-221 ◽  
Author(s):  
Julien Verrax ◽  
Raphael Beck ◽  
Nicolas Dejeans ◽  
Christophe Glorieux ◽  
Brice Sid ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cancer Therapy ◽  
Cancer Cells ◽  
Redox Active

In vitro study on the effects of iron sucrose, ferric gluconate and iron dextran on redox-active iron and oxidative stress

2011 ◽  
Vol 60 (07) ◽  
pp. 459-465
Author(s):  
Brigitte Sturm ◽  
Hannes Steinkellner ◽  
Nina Ternes ◽  
Hans Goldenberg ◽  
Barbara Scheiber-Mojdehkar
Keyword(s):  
Oxidative Stress ◽  
In Vitro Study ◽  
Iron Sucrose ◽  
Vitro Study ◽  
Iron Dextran ◽  
Active Iron ◽  
Redox Active ◽  
And Oxidative Stress

Cell wall-associated transition metals improve alkaline-oxidative pretreatment in diverse hardwoods

2016 ◽  
Vol 18 (5) ◽  
pp. 1405-1415 ◽  
Author(s):  
Namita Bansal ◽  
Aditya Bhalla ◽  
Sivakumar Pattathil ◽  
Sara L. Adelman ◽  
Michael G. Hahn ◽  
...  
Keyword(s):  
Cell Wall ◽  
Transition Metals ◽  
Critical Role ◽  
Redox Active ◽  
Active Transition ◽  
Oxidative Delignification

Cell wall-associated, redox-active transition metals play a critical role in the efficacy of oxidative delignification.

scholarly journals Endogenous formaldehyde scavenges cellular glutathione resulting in cytotoxic redox disruption

2020 ◽  
Author(s):  
Carla Umansky ◽  
Agustín Morellato ◽  
Marco Scheidegger ◽  
Matthias Rieckher ◽  
Manuela R. Martinefski ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Repair ◽  
Fanconi Anemia ◽  
Redox Homeostasis ◽  
Thiol Group ◽  
Cellular Damage ◽  
Repair Pathway ◽  
Cellular Redox ◽  
Redox Active ◽  
Development And Aging

AbstractFormaldehyde (FA) is a ubiquitous endogenous and environmental metabolite that is thought to exert cytotoxicity through DNA and DNA-protein crosslinking. We show here that FA can cause cellular damage beyond genotoxicity by triggering oxidative stress, which is prevented by the enzyme alcohol dehydrogenase 5 (ADH5/GSNOR). Mechanistically, we determine that endogenous FA reacts with the redox-active thiol group of glutathione (GSH) forming S-hydroxymethyl-GSH, which is metabolized by ADH5 yielding reduced GSH thus preventing redox disruption. We identify the ADH5-ortholog gene in Caenorhabditis elegans and show that oxidative stress also underlies FA toxicity in nematodes. Moreover, we show that endogenous GSH can protect cells lacking the Fanconi Anemia DNA repair pathway from FA, which might have broad implications for Fanconi Anemia patients and for healthy BRCA2-mutation carriers. We thus establish a highly conserved mechanism through which endogenous FA disrupts the GSH-regulated cellular redox homeostasis that is critical during development and aging.

Cytochrome c limits oxidative stress and decreases acidosis in a rat model of hemorrhagic shock and reperfusion injury

2017 ◽  
Vol 82 (1) ◽  
pp. 35-41 ◽  
Author(s):  
Rebecca D. Powell ◽  
Donna A. Goodenow ◽  
Hannah V. Mixer ◽  
Iain H. Mckillop ◽  
Susan L. Evans
Keyword(s):  
Oxidative Stress ◽  
Cytochrome C ◽  
Hemorrhagic Shock ◽  
Reperfusion Injury ◽  
Rat Model

Abstract 371: Mitochondrial Antiviral Signaling (MAVS) Protects Cardiomyocytes Under Oxidative Stress by Interfering with Bax-Mediated Cell Death

2012 ◽  
Vol 111 (suppl_1) ◽  
Author(s):  
Toshitaka Yajima ◽  
Stanley Park ◽  
Hanbing Zhou ◽  
Michinari Nakamura ◽  
Mitsuyo Machida ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Cell Death ◽  
Membrane Potential ◽  
Cytochrome C ◽  
Mitochondrial Membrane Potential ◽  
Mitochondrial Membrane ◽  
Cytochrome C Release ◽  
Antiviral Signaling ◽  
Cell Death Pathway ◽  
The Impact

MAVS is a mitochondrial outer membrane protein that activates innate antiviral signaling by recognizing cytosolic viral RNAs and DNAs. While the discovery of MAVS is the first molecular evidence that links mitochondria to innate immune mechanisms, it is still unclear whether MAVS affects mitochondrial cell death as a member of caspase activation and recruitment domain (CARD)-containing proteins. We found that MAVS interacts with Bax through CARD by Yeast two-hybrid and a series of immunoprecipitation (IP) assay, which led us to hypothesize that MAVS functions not only in the innate antiviral mechanisms but also in the mitochondrial cell death pathway. Methods: 1) We examined molecular interaction between MAVS and Bax under oxidative stress by IP using isolated myocytes with H2O2 stimulation and the heart post ischemia-reperfusion (I/R). 2) We evaluated the effect of MAVS on mitochondrial membrane potential and apoptosis under H2O2 stimulation using isolated myocytes with adenoviral MAVS knockdown. 3) We investigated the impact of MAVS on %myocardial infarction (%MI) post I/R using cardiac-specific MAVS knockout (cKO) and transgenic (cTg) mice which we have originally generated. 4) We examined the effect of MAVS on recombinant Bax (rBax)-mediated cytochrome c release using isolated mitochondria from wild type (WT) and MAVS KO mice. Results: 1) The amount of Bax pulled down with MAVS was significantly increased in isolated myocytes with 0.2 mM H2O2 compared to those without stimulation (mean±SD; 1.808±0.14, n=5, p<0.001) and in the heart post I/R compared to sham (2.2±1.19, n=3, p=0.0081). 2) Myocytes with MAVS knockdown showed clear abnormalities in mitochondrial membrane potential and caspace-3 cleavage with 0.2 mM H2O2 compared to control cardiomyocytes. 3) MAVS cKO had significantly larger %MI than WT (81.9 ± 5.8% vs. 42.6 ± 13.6%, n=8, p=0.0008). In contrast, MAVS cTg had significantly smaller %MI that WT (30.0 ± 4.8% vs. 49.2 ± 4.8%, n=10, p=0.0113). 4) Mitochondria from MAVS KO exhibited cytochrome c release after incubation with 2.5 μ g of rBax while those from WT required 10 μ g of rBax. Conclusion: These results demonstrate that MAVS protects cardiomyocyte under oxidative stress by interfering with Bax-mediated cytochrome c release from mitochondria.

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