Quantifying cellular oxidative stress by dichlorofluorescein assay using microplate reader11Mention of a trade name, proprietary product, or specific equipment does not constitute a guarantee by the United States Department of Agriculture and does not imply its approval to the exclusion of other products that may be suitable.

Wang, Hong; Joseph, James A.

doi:10.1016/s0891-5849(99)00107-0

Cited by 2,081 publications

(461 citation statements)

References 19 publications

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“…Oxygen consumption, ATP content, and mitochondrial membrane potential were measured as previously described (6) using a Clark-type electrode (Hansatech; Norfolk, England, UK), a luciferase-based commercial kit (Sigma-Aldrich), and the fluorescent dye rhodamine 123 (SigmaAldrich), respectively. Intracellular quantification of ROS accumulation in cultured cells was performed as previously described (16,22) by fluorescence-based quantification of the oxidation of the intracellular dye dihydrodichloro-fluorescein (Molecular Probes/Invitrogen).…”

Section: Methodsmentioning

confidence: 99%

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

Schulz

Thierbach

Voigt

et al. 2006

Journal of Biological Chemistry

187

150

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More than 80 years ago Otto Warburg suggested that cancer might be caused by a decrease in mitochondrial energy metabolism paralleled by an increase in glycolytic flux. In later years, it was shown that cancer cells exhibit multiple alterations in mitochondrial content, structure, function, and activity. We have stably overexpressed the Friedreich ataxia-associated protein frataxin in several colon cancer cell lines. These cells have increased oxidative metabolism, as shown by concurrent increases in aconitase activity, mitochondrial membrane potential, cellular respiration, and ATP content. Consistent with Warburg's hypothesis, we found that frataxin-overexpressing cells also have decreased growth rates and increased population doubling times, show inhibited colony formation capacity in soft agar assays, and exhibit a reduced capacity for tumor formation when injected into nude mice. Furthermore, overexpression of frataxin leads to an increased phosphorylation of the tumor suppressor p38 mitogen-activated protein kinase, as well as decreased phosphorylation of extracellular signalregulated kinase. Taken together, these results support the view that an increase in oxidative metabolism induced by mitochondrial frataxin may inhibit cancer growth in mammals.Friedreich ataxia is an inherited neurodegenerative disorder (1) caused by the reduced expression of mitochondrial frataxin protein (2) leading to premature death due to cardiac failure (1), diabetes mellitus and insulin resistance (3), as well as impaired ATP synthesis in humans (4, 5). Concurrently it was shown that frataxin promotes oxidative metabolism and ATP synthesis when overexpressed in fibroblasts (6), possibly by a direct interaction with the respiratory chain (7). Although the primary function of frataxin is still a matter of debate (8), some evidence suggests that this protein directs the intramitochondrial synthesis of Fe/S clusters (9 -12). Individuals suffering from Friedreich ataxia have a reduced life expectancy of 38 years on average (1) and show increased oxidative stress (13-15). Overexpression of frataxin has been shown to reduce intracellular accumulation of reactive oxygen species (ROS) 3 and to prevent menadione-induced malignant transformation of fibroblasts (16). Furthermore, disruption of the frataxin homologue in yeast has been shown to cause increased sensitivity to oxidants and promote oxidative damage to both nuclear (17), as well as mitochondrial, DNA (18). In addition, fibroblasts from Friedreich ataxia patients exhibit increased sensitivity against ionizing radiation and show an increased frequency of transforming events (19). Although malignant disease is not considered a typical feature of the disorder, Friedreich ataxia patients exhibit various types of cancer atypical for their young age (reviewed in Refs. 3 and 16). Lastly, targeted disruption of frataxin in murine hepatocytes causes decreased life span and liver tumor formation in mice. 4 In parallel to these latter findings in rodents, we questioned whether frataxin migh...

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Section: Methodsmentioning

confidence: 99%

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

Schulz

Thierbach

Voigt

et al. 2006

Journal of Biological Chemistry

187

150

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“…The intracellular ROS level was detected by using 5-(and-6)-carboxy-2Ј,7Ј-dichlorodihydrofluorescein diacetate (carboxy-H 2 DCFDA) (Molecular Probes) as reported (9). Carboxy-H 2 DCFDA is a cell-permeable indicator for ROS that is nonfluorescent until the acetate groups are removed by intracellular esterases and oxidation occurs within the cell.…”

Section: Methodsmentioning

confidence: 99%

A role of reactive oxygen species in apoptotic activation of volume-sensitive Cl^-channel

Shimizu

Numata

Okada

2004

Proc. Natl. Acad. Sci. U.S.A.

214

245

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“…This non-fluorescent reagent diffuses passively through the plasma membrane into the cell, where acetate groups are cleaved by intracellular esterases. The compound can then be oxidized by ROS formed within the cell (particularly by hydroxyl radicals, OH) to form fluorescent dichlorofluorescein, while the fluorescence intensity is proportional to the ROS level (24).…”

Section: Determination Of Cellular Reactive Oxygen Species (Ros) Formmentioning

confidence: 99%

Effect of selected catechins on doxorubicin antiproliferative efficacy and hepatotoxicity in vitro

et al. 2014

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Catechins may influence both desirable and undesirable effects of many drugs. In this study, the in vitro effect of (+)-catechin, (-)-epicatechin, (-)-epigallocatechin, (-)-epicatechin gallate, and (-)-epigallocatechin gallate (EGCG) on the efficacy of anticancer drug doxorubicin (DOX) was studied in HCT-8 cancer cells. Rat hepatocytes were used to study the influence of EGCG on DOX hepatotoxicity. Cell proliferation and viability were studied by 3-[4,5-dimethylthiazol- 2-yl]-2,5-diphenyl tetrazolium bromide and neutral red uptake test assays. Formation of reactive oxygen species (ROS) was determined using the dichlorofluorescein assay. All of the studied catechins (1-25 μmol L-1) had no effect on the proliferation of intestinal cancer cells and did not affect the antiproliferative effect of DOX (1-8 μmol L-1) in these cells. Moreover, EGCG at 25 μmol L-1 increased the viability of isolated hepatocytes and significantly protected these cells against DOX-induced toxicity and ROS production. Consumption of EGCG during DOX therapy seems to be safe and beneficial, since EGCG does not decrease DOX anticancer efficacy and could ameliorate DOX hepatotoxicity

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Cited by 2,081 publications

References 19 publications

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

A role of reactive oxygen species in apoptotic activation of volume-sensitive Cl^-channel

Effect of selected catechins on doxorubicin antiproliferative efficacy and hepatotoxicity in vitro

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Cited by 2,081 publications

References 19 publications

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

Induction of Oxidative Metabolism by Mitochondrial Frataxin Inhibits Cancer Growth

A role of reactive oxygen species in apoptotic activation of volume-sensitive Cl-channel

Effect of selected catechins on doxorubicin antiproliferative efficacy and hepatotoxicity in vitro

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A role of reactive oxygen species in apoptotic activation of volume-sensitive Cl^-channel