Svf1 inhibits reactive oxygen species generation and promotes survival under conditions of oxidative stress in Saccharomyces cerevisiae

Aberrant regulation of apoptosis, or programmed cell death, contributes to the aetiology of several diseases, including cancers, immunodeficiencies and neurodegenerative illnesses. We hypothesized that key features of mammalian cell death regulation may be conserved in single celled organisms such as the budding yeast Saccharomyces cerevisiae. We previously identified the yeast gene SVF1 in a screen for mutations that could be functionally complemented by exogenous expression of the human anti-apoptotic gene Bcl-x(L). Anti-apoptotic Bcl-2 family members have been shown to promote redox stability through upregulation of antioxidant pathways in mammalian cells. Here we demonstrate that the Svf1 protein is required for yeast survival under conditions of oxidative stress, including cold stress. Cells lacking SVF1 are hypersensitive to conditions associated with increased reactive oxygen species (ROS) generation and to direct chemical precursors of ROS, and demonstrate increased levels of ROS under these conditions. Hypersensitivity to oxidative stress can be reversed by treatment with the antioxidant N-acetylcysteine or expression of exogenous SVF1, although exogenous expression of Bcl-x(L) did not protect cells from cold stress. Exogenous SVF1 expression in mammalian cells confers resistance to H(2)O(2) exposure. Our data are consistent with previous observations suggesting a key role of oxidative stress response in mammalian apoptotic regulation and validate the use of S. cerevisiae as a model for studying programmed cell death.     

Aerosol-borne quinones and reactive oxygen species generation by particulate matter extracts

The mass loadings of quinones and their ability to generate reactive oxygen species (ROS) were investigated in total suspended particulate samples collected in Fresno, CA, over a 12-month period. Particles were collected on Teflon filters and were analyzed for the presence of 12 quinones containing one to four aromatic rings by gas chromatography with mass spectrometry. Measured levels are generally greater than mass loadings reported at other locations. The mass loadings were highest during winter months and were strongly anticorrelated with temperature. ROS generation was investigated by measuring the rate of hydrogen peroxide production from the reaction of laboratory standards and ambient samples with dithiothreitol (DTT). ROS generation from ambient samples shows a strong positive correlation with the mass loadings of the three most reactive quinones and may account for all of the ROS formed in the DTT test.     

自由基、活性氧与疾病

该文主要介绍自由基、活性氧与疾病关系,并简要提出抑制自由基方法。     

Effect of cis-urocanic acid on bovine neutrophil generation of reactive oxygen species

Neutrophils play a fundamental role in the host innate immune response during mastitis and other bacterial-mediated diseases of cattle. One of the critical mechanisms by which neutrophils contribute to host innate immune defenses is through their ability to phagocytose and kill bacteria. The ability of neutrophils to kill bacteria is mediated through the generation of reactive oxygen species (ROS). However, the extracellular release of ROS can be deleterious to the host because ROS induce tissue injury. Thus, in diseases such as mastitis that are accompanied by the influx of neutrophils, the generation of large quantities of ROS may result in significant injury to the mammary epithelium. cis-Urocanic acid (cis-UCA), which is formed from the UV photoisomerization of the trans isoform found naturally in human and animal skin, is an immunosuppressive molecule with anti-inflammatory properties. Little is known about the effect of cis-UCA on neutrophils, although one report demonstrated that it inhibits human neutrophil respiratory burst activity. However, the nature of this inhibition remains unknown. Because of the potential therapeutic use that a molecule such as cis-UCA may have in blocking excessive respiratory burst activity that may be deleterious to the host, the ability of cis-UCA to inhibit bovine neutrophil production of ROS was studied. Further, because neutrophil generation of ROS is necessary for optimal neutrophil bactericidal activity, a response which is critical for the host innate immune defense against infection, the effects of cis-UCA on bovine neutrophil phagocytosis and bacterial killing were assayed. cis-Urocanic acid dose-dependently inhibited the respiratory burst activity of bovine neutrophils as measured by luminol chemiluminescence. Subsequently, the effect of cis-UCA on the production of specific oxygen radicals was investigated using more selective assays. Using 2 distinct assays, we established that cis-UCA inhibited the generation of extracellular superoxide. In contrast, cis-UCA had no effect on the generation of intracellular levels of superoxide or other ROS. At concentrations that inhibited generation of extracellular superoxide, bovine neutrophil phagocytosis and bacterial activity remained intact. Together, these data suggest that cis-UCA inhibits the tissue-damaging generation of extracellular ROS while preserving neutrophil bactericidal activity.     

番茄红素纳米分散体体外清除活性氧自由基研究

采用乳化-蒸发工艺（超声乳化）制备了番茄红素纳米分散体,用分光光度法研究了番茄红素纳米分散体体外清除活性氧自由基（过氧化氢、羟自由基）的作用,并与溶于四氢呋喃的番茄红素进行比较。结果表明,番茄红素纳米分散体对这两种自由基均有不同程度的清除作用,且均呈现一定的量效关系;在相同浓度下,其清除能力均强于番茄红素四氢呋喃溶液,其中对过氧化氢清除能力远远高于番茄红素四氢呋喃溶液。      

宰后成熟过程中活性氧介导的氧化应激对肉品质的影响

宰后成熟是提高肉品质的有效途径之一,为了改善肉品质,活性氧(reactive oxygen species,ROS)介导的氧化应激已成为国内外学者的研究热点,但ROS介导的氧化应激的研究主要集中在医学领域。通过综述ROS介导的细胞死亡方式(凋亡、自噬与坏死)可能与肉品质之间的关系,以及ROS对肉色泽、风味、嫩度的影响,来阐述ROS介导的氧化应激可能在宰后成熟过程中对肉品质的影响。     

Ligand-enhanced reactive oxidant generation by nanoparticulate zero-valent iron and oxygen

The reaction of zero-valent iron or ferrous iron with oxygen produces reactive oxidants capable of oxidizing organic compounds. However, the oxidant yield in the absence of ligands is too low for practical applications. The addition of oxalate, nitrilotriacetic acid (NTA), or ethylenediaminetetraacetic acid (EDTA) to oxygen-containing solutions of nanoparticulate zero-valent iron (nZVI) significantly increases oxidant yield, with yields approaching their theoretical maxima near neutral pH. These ligands improve oxidant production by limiting iron precipitation and by accelerating the rates of key reactions, including ferrous iron oxidation by oxygen and hydrogen peroxide. Product yields indicate that the oxic nZVI system produces hydroxyl radical (OH*) over the entire pH range in the presence of oxalate and NTA. In the presence of EDTA, probe compound oxidation is attributed to OH under acidic conditions and a mixture of OH* and ferryl ion (Fe[IV]) at circumneutral pH.     

油茶总皂甙抑菌活性及清除活性氧能力初步研究

用溶剂法提取油茶饼中油茶总皂甙,经柱层析制备标准品标定有效成分含量为82.5%。以油茶皂甙为材料对枯草杆菌、大肠杆菌、金黄色葡萄球菌进行抑菌试验,结果表明均有抑制作用;其中对枯草杆菌和金黄色葡萄球菌抑制作用较强,20%浓度对三种细菌均具有致死作用,抑菌圈直径大小与油茶总皂甙浓度呈正相关;且初步测定油茶总皂甙清除活性氧能力强于维生素C。     

Chemistry and reactions of reactive oxygen species in foods

Reactive oxygen species (ROS) are formed enzymatically, chemically, photochemically, and by irradiation of food. They are also formed by the decomposition and the inter-reactions of ROS. Hydroxy radical is the most reactive ROS, followed by singlet oxygen. Reactions of ROS with food components produce undesirable volatile compounds and carcinogens, destroy essential nutrients, and change the functionalities of proteins, lipids, and carbohydrates. Lipid oxidation by ROS produces low molecular volatile aldehydes, alcohols, and hydrocarbons. ROS causes crosslink or cleavage of proteins and produces low molecular carbonyls from carbohydrates. Vitamins are easily oxidized by ROS, especially singlet oxygen. The singlet oxygen reaction rate was the highest in ss-carotene, followed by tocopherol, riboflavin, vitamin D, and ascorbic acid.     

The inhibition effects of reactive oxygen species by Maillard reaction products in Caco‐2 cells

Maillard reaction products (MRPs) were prepared by heating a mixture of rice starch with different dextrose equivalents (DE 10, 30, 50 and 70) and glycine. The glycine was added to the sample pastes at the same molar concentration as the sugar contained in each sample. As the dextrose equivalent of rice starch increased, the browning intensity and fluorescence of the MRPs increased. The antioxidant properties of the MRPs were investigated by DPPH radical scavenging activity, reducing power, and phenolic content in a chemical system, and were evaluated by measuring reactive oxygen species levels and antioxidant enzymes activities in Caco‐2 cells. The darkest MRPs, MRPs‐4, showed the highest antioxidant activity and phenolic content among the samples; in addition, it inhibited the cellular oxidative stress. The decrease of cell viability and antioxidant enzymes activities caused by reactive oxygen species and apoptosis were recovered by MRPs‐4. Actually, the addition of the MRPs suppressed apoptosis by decreasing the proportion of cells in the sub‐G1 phase. Therefore, these MRPs, as compounds formed by the Maillard reaction, are considered to possess an effective antioxidant activity against oxidizable substrates.     

Size dependent and reactive oxygen species related nanosilver toxicity to nitrifying bacteria

The intrinsic slow growth of nitrifying bacteria and their high sensitivity to environmental perturbations often result in cell growth inhibition by toxicants. Nanoparticles are of great concern to the environment because of their small size and high catalytic properties. This work sought to determine size-dependent inhibition by Ag nanoparticles and evaluate the relationship between the inhibition and reactive oxygen species (ROS). Nanoparticles with an average size range of 9-21 nm were synthesized by varying the molar ratios of BH4-/Ag+ in the solution. The resulting ROS generation was quantified in the presence and absence of the bacteria while the degree of inhibition was inferred from specific oxygen uptake rate measurements, determined by extant respirometry. By examining the correlation between nanoparticle size distribution, photocatalytic ROS generation, intracellular ROS accumulation, and nitrification inhibition, we observed that inhibition to nitrifying organisms correlated with the fraction of Ag nanoparticles less than 5 nm in the suspension. It appeared that these size nanoparticles could be more toxic to bacteria than any other fractions of nanoparticles or their counterpart bulk species. Furthermore, inhibition by Ag nanoparticles as well as other forms of silver (AgCl colloid and Ag+ ion) correlated well with the intracellular ROS concentrations, but not with the photocatalytic ROS fractions. The ROS correlations were different for the different forms of silver, indicating that factors other than ROS are also important in determining nanosilver toxicity.     

Soy isoflavone genistein induces cell death in breast cancer cells through mobilization of endogenous copper ions and generation of reactive oxygen species

Scope: Worldwide geographical variation in cancer incidence indicates a correlation between dietary habits and cancer risk. Epidemiological studies have suggested that populations with high isoflavone intake through soy consumption have lower rates of breast, prostate, and colon cancer. Isoflavone genistein in soybean is considered a potent chemopreventive agent against cancer. Although several mechanisms have been proposed, a clear anticancer action mechanism of genistein is still not known. Methods and results: Here, we show that the cytotoxic action of genistein against breast cancer cells involves mobilization of endogenous copper. Further, whereas the copper specific chelator neocuproine is able to inhibit the apoptotic potential of genistein, the molecules which specifically bind iron (desferroxamine mesylate) and zinc (histidine) are relatively ineffective in causing such inhibition. Also, genistein‐induced apoptosis in these cells is inhibited by scavengers of reactive oxygen species (ROS) implicating ROS as effector elements leading to cell death. Conclusions: As copper levels are known to be considerably elevated in almost all types of cancers, in this proof‐of‐concept study we show that genistein is able to target endogenous copper leading to prooxidant signaling and consequent cell death. We believe that such a mechanism explains the anticancer effect of genistein as also its preferential cytotoxicity towards cancer cells.     

Enantioselectivity tuning of chiral herbicide dichlorprop by copper: roles of reactive oxygen species

Reactive oxygen species (ROS) are considered to be the key players in cell toxicity. However, cross talk between the enantioselective toxicity of pesticides, heavy metals, and ROS is poorly understood. To decipher the puzzle, the effects of copper (Cu) on the enantioselective ecotoxicity of the chiral pesticide dichlorprop (DCPP) to Scenedesmus obliquus were investigated. The results showed that the presence of DCPP and Cu, both individually and in combination, caused a sudden increase of ROS. This in turn stimulated the response of antioxidant defenses, impaired subcellular structure and physiological function, and finally resulted in cell growth inhibition. In the absence of Cu, ROS production after exposure to the herbicidally active (R)-enantiomer was higher than that of the (S)-enantiomer, suggesting a preference for an (R)-enantiomer-induced production of ROS. When DCPP and Cu were both added to algae simultaneously, (R)-DCPP preferentially induced production of ROS was observed. However, the enantioselective induced production of ROS was reversed when DCPP was mixed with Cu for 24 h prior to addition to the algae solution. It was also found that the generation of ROS, antioxidant response, and growth inhibition rate in Scenedesmus obliquus were all (R)-enantiomer preferentially induced. These findings implied that ROS play a primary role in chemical contaminant toxicity, and interactions between contaminants can tune the enantioselectivity of chiral herbicides, which should be considered in future risk assessment.     

Low abundance of mitophagy markers is associated with reactive oxygen species overproduction in cows with fatty liver and causes reactive oxygen species overproduction and lipid accumulation in calf hepatocytes

Mitochondria are the main site of fatty acid oxidation and reactive oxygen species (ROS) formation. Damaged or dysfunctional mitochondria induce oxidative stress and increase the risk of lipid accumulation. During the process of mitophagy, PTEN induced kinase 1 (PINK1) accumulates on damaged mitochondria and recruits cytoplasmic Parkin to mitochondria. As an autophagy receptor protein, sequestosome-1 (p62) binds Parkin-ubiquitinated outer mitochondrial membrane proteins and microtubule-associated protein 1 light chain 3 (LC3) to facilitate degradation of damaged mitochondria. In nonruminants, clearance of dysfunctional mitochondria through the PINK1/Parkin-mediated mitophagy pathway contributes to reducing ROS production and maintaining metabolic homeostasis. Whether PINK1/Parkin-mediated mitophagy plays a similar role in dairy cow liver is not well known. Thus, the objective of this study was to investigate mitophagy status in dairy cows with fatty liver and its role in free fatty acid (FFA)-induced oxidative stress and lipid accumulation. Liver and blood samples were collected from healthy dairy cows (n = 10) and cows with fatty liver (n = 10) that had a similar number of lactations (median = 3, range = 2 to 4) and days in milk (median = 6 d, range = 3 to 9 d). Calf hepatocytes were isolated from 5 healthy newborn female Holstein calves (1 d of age, 30–40 kg). Hepatocytes were transfected with small interfering RNA targeted against PRKN for 48 h or transfected with PRKN overexpression plasmid for 36 h, followed by treatment with FFA (0.3 or 1.2 mM) for 12 h. Mitochondria were isolated from fresh liver tissue or calf hepatocytes. Serum concentrations of β-hydroxybutyrate were higher in dairy cows with fatty liver. Hepatic malondialdehyde (MDA) and hydrogen peroxide (H₂O₂) were greater in cows with fatty liver. The lower protein abundance of PINK1, Parkin, p62, and LC3-II in hepatic mitochondrial fraction of dairy cows with fatty liver indicated the mitophagy was impaired. In hepatocytes, knockdown of PRKN decreased protein abundance of p62 and LC3-II in the mitochondrial fraction, and increased contents of triacylglycerol (TG), MDA, and H₂O₂. In addition, protein abundances of PINK1, Parkin, p62, and LC3-II were lower in the mitochondrial fraction from hepatocytes treated with 1.2 mM FFA than the hepatocytes treated with 0.3 mM FFA, whereas the content of TG, MDA, and H₂O₂ increased. In 1.2 mM FFA-treated hepatocytes, PRKN overexpression increased protein abundance of p62 and LC3-II in the mitochondrial fraction and decreased contents of TG, MDA, and H₂O₂. Together, our data demonstrate that low abundance of mitophagy markers is associated with ROS overproduction in dairy cows with fatty liver and impaired mitophagy induced by a high concentration of FFA promotes ROS production and lipid accumulation in female calf hepatocytes.     

Chemiluminescent screening of quenching effects of natural colorants against reactive oxygen species: Evaluation of grape seed,monascus, gardenia and red radish extracts as multi-functional food additives

The quenching effects of commercial available natural colorants against reactive oxygen species (ROS) were investigated. The effectiveness was evaluated by measuring the quenching ratio of chemiluminescence (CL) intensity with each colorant on luminol or methyl-6-(p-methoxyphenyl)-3,7-dihydroimidaz[1,2-a]pyrazin-3-one CL induced by ROS. As a result, the grape seed extracts and polyphenolics dose-dependently quenched CL. The quenching effects of grape seed extract A at 1 mg/ml for superoxide, singlet oxygen, hydroxyl radical, hypochlorite ion and linolenic acid peroxide were 81.1 ± 0.5%, 95.7 ± 0.4%, 99.3 ± 0.3%, 27.7 ± 4.2% and 88.3 ± 1.0%, respectively (n = 3). Except for chalcone, the polyphenolics, such as pelargonidin, cyanidin, delphinidin and trans-resveratrol, also showed the high quenching effects against ROS. These results suggested that the grape seed extract might be useful as a multi-functional food additive.     

Induction of apoptosis by Myagropsis myagroides extracts is associated with a caspase dependent pathway and reactive oxygen species generation in human cancer cells

The purpose of this study was to elucidate the mechanisms underlying apoptosis induced by an ethanol extracts from Myagropsis myagroides (ME) in HeLa, U937, and PC-3 cells. ME treatment for 24 h significantly inhibited cell viability in a dose-dependent manner, and induced apoptosis. Moreover, ME treatment triggered the cleavage of caspase-8, ?9, ?3, and poly(ADP-ribose) polymerase (PARP). A general caspase inhibitor (z-VAD-fmk) inhibited ME-induced activation of caspase-3, PARP cleavage, and cell death. ME treatment also triggered the release of cytochrome c from the mitochondria to the cytosol and stimulated the cleavage of Bid, up-regulation of Bax, and down-regulation of Bcl-2. Furthermore, ME treatment caused reactive oxygen species (ROS) generation. An antioxidant N-acetylcysteine (NAC) blocked MEinduced activation of caspase-3, PARP cleavage, and cell death. Overall, these results suggest that ME-induced apoptosis is mediated by a caspase dependent pathway and ROS generation in HeLa, U937, and PC-3 cells.