Measuring reactive species and oxidative damage<i>in vivo</i>and in cell culture: how should you do it and what do the results mean?

Halliwell, Barry; Whiteman, Matthew

doi:10.1038/sj.bjp.0705776

Cited by 2,026 publications

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References 385 publications

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“…We opted to utilize the triple mutant over the single / double mutants due to its documented hypersensitivity to metal exposure (Hughes et al, 2009). The DCFH-DA assay has previously been applied to detect the formation of metalinduced free radical compounds (Halliwell and Whiteman, 2004) but has also been criticized for its low specificity (Cohn et al, 2008;Karlsson et al, 2010). This assay provided quantitative evidence that the relative base-line levels of ROS generation are comparable between the wildtype and the mutant nematodes.…”

Section: Resultsmentioning

confidence: 99%

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Polak

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Jurkschat

et al. 2014

Comparative Biochemistry and Physiology Part C: Toxicology &

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Spurgeon, David J.; Sturzenbaum, Stephen R.. 2014. Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegansContact CEH NORA team at noraceh@ceh.ac.ukThe NERC and CEH trademarks and logos ('the Trademarks') are registered trademarks of NERC in the UK and other countries, and may not be used without the prior written consent of the Trademark owner. suggesting that the nominal exposure to pure ZnONPs represents in vivo, at best, a mixture exposure of ionic zinc and nanoparticles. Nevertheless, the analyzes provided evidence that the metallothioneins (mtl-1 and mtl-2), the pytochelatin synthase (pcs-1) and an apoptotic marker (cep-1) were transcriptionally activated. In addition, the DCFH-DA assay provided in vitro evidence of the oxidative potential of ZnONPs in the metal exposure sensitive triple mutant.Raman spectroscopy highlighted that the biomolecular phenotype changes significantly in the metal sensitive knockout worm upon ZnONP exposure, suggesting that these metalloproteins are instrumental in the protection against cytotoxic damage. Finally, ZnONP exposure was shown to decrease growth and development, reproductive capacity and lifespan, effects which were amplified in the triple knockout. By combining diverse toxicological strategies, we identified that individuals (genotypes) housing mutations in key metalloproteins are more susceptible to ZnONP exposure, which underlines the importance of fully functional metalloproteins to minimize ZnONP induced toxicity.

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Section: Resultsmentioning

confidence: 99%

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Polak

Read

Jurkschat

et al. 2014

Comparative Biochemistry and Physiology Part C: Toxicology &

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“…In summary, by the turn of the century, the undue fear for OS injury in human health started waning and the appreciation for their contribution in cellular physiology began growing [10, 37-41, 44, 45, 140]. Obviously, an opposite view became progressively louder for AO [52,53,56,133,[141][142][143][144].…”

Section: Reactive Oxygen Species and Reactive Nitrogen Species Transimentioning

confidence: 99%

“…Depending on the conditions, they damage or destroy these molecules [7,9,143,144]. They can easily deshape or mutate DNA which clearly gives inkling that they can cause carcinogenesis.…”

Section: Smoke After Firementioning

confidence: 99%

Reconvene and Reconnect the Antioxidant Hypothesis in Human Health and Disease

Singh

Chandra

Mahdi

et al. 2010

Indian J Clin Biochem

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The antioxidants are essential molecules in human system but are not miracle molecules. They are neither performance enhancers nor can prevent or cure diseases when taken in excess. Their supplemental value is debateable. In fact, many high quality clinical trials on antioxidant supplement have shown no effect or adverse outcomes ranging from morbidity to all cause mortality. Several Chochrane Meta-analysis and Markov Model techniques, which are presently best available statistical models to derive conclusive answers for comparing large number of trials, support these claims. Nevertheless none of these statistical techniques are flawless. Hence, more efforts are needed to develop perfect statistical model to analyze the pooled data and further double blind, placebo controlled interventional clinical trials, which are gold standard, should be implicitly conducted to get explicit answers. Superoxide dismutase (SOD), glutathione peroxidase and catalase are termed as primary antioxidants as these scavenge superoxide anion and hydrogen peroxide. All these three enzymes are inducible enzymes, thereby inherently meaning that body increases or decreases their activity as per requirement. Hence there is no need to attempt to manipulate their activity nor have such efforts been clinically useful. SOD administration has been tried in some conditions especially in cancer and myocardial infarction but has largely failed, probably because SOD is a large molecule and can not cross cell membrane. The dietary antioxidants, including nutrient antioxidants are chain breaking antioxidants and in tandem with enzyme antioxidants temper the reactive oxygen species (ROS) and reactive nitrogen species (RNS) within physiological limits. Since body is able to regulate its own requirements of enzyme antioxidants, the diet must provide adequate quantity of non-enzymic antioxidants to meet the normal requirements and provide protection in exigent condition. So far, there is no evidence that human tissues ever experience the torrent of reactive species and that in chronic conditions with mildly enhanced generation of reactive species, the body can meet them squarely if antioxidants defense system in tissues is biochemically optimized. We are not yet certain about optimal levels of antioxidants in tissues. Two ways have been used to assess them: first by dietary intake and second by measuring plasma levels. Lately determination of plasma/serum level of antioxidants is considered better index for diagnostic and prognostic purposes. The recommended levels for vitamin A, E and C and beta carotene are 2.2-2.8 lmol/l; 27.5-30 lmol/l; 40-50 lmol/l and 0.4-0.5 lmol/l, respectively. The requirement and recommended blood levels of other dietary antioxidants are not established. The resolved issues are (1) essential to scavenge excess of radical species (2) participants in redox homeostasis (3) selective antioxidants activity against radical species (4) there is no universal antioxidant and 5) therapeutic value in case of deficiency. The overar...

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“…Oxidative stress results from a disruption in the prooxidant and antioxidant balance and induces damages to organic compounds such as lipids, proteins, and nucleic acids [9]. So, either a reduction or antioxidant levels or/and an increase in ROS lead to oxidative stress [10]. Enhanced oxidative stress was evidenced in diabetes pathology [3].…”

Section: Pre-laboratory Preparationmentioning

confidence: 99%

Oxidative damage in diabetics: Insights from a graduate study in La Reunion University

Girard¹,

Rondeau²,

Catan³

et al. 2014

Biochem Molecular Bio Educ

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Due to the growing incidence of diabetes in developed nations, there is a compelling case to be made for teaching graduate students more deeply about mechanisms underlying this disease. Diabetes is associated with enhanced oxidative stress and protein glycation via the covalent binding of glucose molecules. Albumin represents the major plasmatic protein and undergoes enhanced glycoxidative modifications in diabetic condition. La Reunion Island, a French department located in the Indian Ocean exhibit a growing incidence of diabetes. At the University of La Reunion, our research group named GEICO (Groupe d'Etude sur l'Inflammation Chronique et l'Obesite) participated to foster research and training in diabetes context and focuses on the impact of glycated albumin mediated oxidative stress on cell physiopathology. A laboratory course was designed by our group to introduce graduate students to cutting edge techniques in redox biology while providing insights into scientific processes and methods. This two weeks research laboratory training took place at CYROI, a local biotechnology center that provides advanced facilities for research, business, and education. Using histochemistry, molecular biology, biochemical techniques, student investigated oxidative damages in liver from leptin receptor deficient diabetic mice compared to control littermates. In addition, they used an in vitro model by assaying oxidative impact of glycated albumin on hepatoma carcinoma HepG2 cells. This article gives an overview of the organization and protocol used by the students during their two weeks training in the laboratory. Therefore, it may be helpful for teaching graduate students techniques used in research laboratory working on redox biology. V C 2014 by The International Union of Biochemistry and Molecular Biology, 42(5): 435-442, 2014.

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Measuring reactive species and oxidative damagein vivoand in cell culture: how should you do it and what do the results mean?

Cited by 2,026 publications

References 385 publications

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Metalloproteins and phytochelatin synthase may confer protection against zinc oxide nanoparticle induced toxicity in Caenorhabditis elegans

Reconvene and Reconnect the Antioxidant Hypothesis in Human Health and Disease

Oxidative damage in diabetics: Insights from a graduate study in La Reunion University

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