Measuring Mitochondrial Reactive Oxygen Species (ROS) Production

Starkov, Anatoly A.

doi:10.1007/978-3-642-30018-9_8

Cited by 3 publications

(3 citation statements)

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“…After treatment and incubation, the cells were transferred in the dark in 96-well plates and suspended in 50 μl cell culture medium containing 65 μl Tris HCl 10 mmol/l and 10 μl DCFH-DA. The plates were maintained for 30 min in a cell culture incubator (16). After incubation, fluorescence was measured using a spectrofluorometer (SpectraMax ® i3x -Molecular Devices).…”

Section: Analysis Of Ros Production With 2'7'-dichlorofluorescein Dia...mentioning

confidence: 99%

Activity of Free and Liposomal Antimony Trioxide in the Acute Promyelocytic Leukemia Cell Line NB4

et al. 2021

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Background/Aim: Antimony is a chemical element used in the therapy of parasitic diseases with a promising anticancer potential. The aim of this study was to evaluate in vitro activity of free or liposomal vesicle-packed antimony trioxide (AT or LAT) in the t(15;17)(q22;q21) translocationpositive acute promyelocytic leukemia (APL) cell line NB4. Materials and Methods: Cytotoxicity was analysed with trypan blue exclusion, the MTT assay and neutral red exclusion assay; cell proliferation with PicoGreen ® ; and reactive oxygen species (ROS) production with DCFDA. Results: Liposomal particles did not change the pH of the cell culture medium and entered the cells. Both formulations resulted in a time-and concentration-dependent cytotoxicity and production of ROS. LAT showed higher toxicity at lower concentrations compared to AT. Conclusion: LAT may be used to decrease drug dosage and maintain high anti-tumoral effects on APL cells.Cancer is a multifactorial disease, and therefore its treatment needs to address different targets to achieve better results. New therapeutic strategies are investigated and repositioning of drugs is a promising alternative. Antimony is used in the therapy of leishmaniasis and schistosomiasis. The main serious adverse effects are associated to cardiotoxicity in about 9% of the patients and pancreatitis, more commonly seen in co-infections (HIV and visceral leishmaniasis) (1). Reactive oxygen species (ROS) production and oxidative stress damage are the main mechanisms of cell damage caused by antimony (2). Studies have shown that autophagy induced by antimony is triggered by the production of ROS and not by the activation of the mTOR pathway (3). In the present study, we used liposomes, which have low biological impact (4). Nanoparticles can improve drug delivery and efficiency at a lower dosage, reducing toxic effects, which is highly needed in cancer treatment (5). Therefore, we compared the effects of free antimony trioxide (AT) and AT encapsulated in liposomes (LAT) on the acute promyelocytic leukemia (APL) cell line NB4.APL is caused by the fusion protein PML/RARA and was the first leukemia treated with the specific molecular targeting drug all-trans-retinoic acid (ATRA) (6). Despite the high efficiency of the treatment, 5 to 20% of the patients develop the ATRA syndrome, a life-threatening condition associated with organ infiltration by myeloid cells, cytokine production, high expression of CD13 in leukemic cells, and hyperleukocytosis, leading to endothelial inflammation and bleeding in the affected organs (6). Arsenic Trioxide (ATO) is an efficient drug for APL treatment, and in association with ATRA, decreases relapse in about 90% of cases ( 7), but its use is limited by the risk of toxicity and potential carcinogenicity (8).The free antimony trioxide (Sb 2 O 3 ) shows chemical structure and mechanistic properties similar to ATO against 6061

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Section: Analysis Of Ros Production With 2'7'-dichlorofluorescein Dia...mentioning

confidence: 99%

Activity of Free and Liposomal Antimony Trioxide in the Acute Promyelocytic Leukemia Cell Line NB4

et al. 2021

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“…More stable and membrane permeable hydrogen peroxide can be readily measured using H 2 O 2 -specific enzyme HRP and a variety of its fluorogenic substrates [79]. It should be noted that measurement of ROS generation in these assays requires internal calibration [80].…”

Section: Novel Approaches To Evaluate Spatiotemporal Levels Of H2o2 Imentioning

confidence: 99%

“…ROS removal can be measured separately [24] or determined from the same internal calibration. Currently, Amplex Red (10-acetyl-3,7-dihydroxyphenoxazine) and its recent, more stable derivative Amplex UltraRed are the compounds of choice in HRP-based H 2 O 2 assays [79, 81]. Another very popular compound that is used to measure ROS is DCF-DA.…”

Section: Novel Approaches To Evaluate Spatiotemporal Levels Of H2o2 Imentioning

confidence: 99%

Mitochondrial ROS metabolism: 10 Years later

Andreyev

Kushnareva

Murphy

et al. 2015

Biochemistry Moscow

160

115

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The role of mitochondria in oxidative stress is well recognized, but many questions are still to be answered. This article is intended to update our comprehensive review in 2005 by highlighting the progress in understanding of mitochondrial reactive oxygen species (ROS) metabolism over the past 10 years. We review the recently identified or re-appraised sources of ROS generation in mitochondria, such as p66shc protein, succinate dehydrogenase, and recently discovered properties of the mitochondrial antioxidant system. We also reflect upon some controversies, disputes, and misconceptions that confound the field.

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