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1.
Cho ES  Lee KW  Lee HJ 《Mutation research》2008,640(1-2):123-130
Oxidative stress induced by reactive oxygen species has been strongly associated with the pathogenesis of neurodegenerative disorders, including Alzheimer's disease. In this study, we investigated the possible protective effects of a cocoa procyanidin fraction (CPF) and procyanidin B2 (epicatechin-(4β-8)-epicatechin) – a major polyphenol in cocoa – against apoptosis of PC12 rat pheochromocytoma (PC12) cells induced by hydrogen peroxide (H2O2). CPF (1 and 5 μg/ml) and procyanidin B2 (1 and 5 μM) reduced PC12 cell death caused by H2O2, as determined by MTT and trypan blue exclusion assays. CPF and procyanidin B2 attenuated the H2O2-induced fragmentation of nucleus and DNA in PC12 cells. Western blot data demonstrated that H2O2 induced cleavage of poly(ADP-ribose)polymerase (PARP), downregulated Bcl-XL and Bcl-2 in PC12 cells. Pretreatment with CPF or procyanidin B2 before H2O2 treatment diminished PARP cleavage and increased Bcl-XL and Bcl-2 expression compared with those only treated with H2O2. Activation of caspase-3 by H2O2 was inhibited by pretreatment with CPF or procyanidin B2. Furthermore, H2O2-induced rapid and significant phosphorylation of c-Jun N-terminal protein kinase (JNK) and p38 mitogen-activated protein kinase (MAPK), and both of these effects were attenuated by CPF or procyanidin B2 treatment. These results suggest that the protective effects of CPF and procyanidin B2 against H2O2-induced apoptosis involve inhibiting the downregulation of Bcl-XL and Bcl-2 expression through blocking the activation of JNK and p38 MAPK.  相似文献   

2.
Introduction Excess of intracellular reactive oxygen species in relation to antioxidative systems results in an oxidative environment which may modulate gene expression or damage cellular molecules. These events are expected to greatly contribute to processes of carcinogenesis. Only few studies are available on the oxidative/reductive conditions in the colon, an important tumour target tissue. It was the objective of this work to further develop methods to assess intracellular oxidative stress within human colon cells as a tool to study such associations in nutritional toxicology.

Methods We have measured H2O2-induced oxidative stress in different colon cell lines, in freshly isolated human colon crypts, and, for comparative purposes, in NIH3T3 mouse embryo fibroblasts. Detection was performed by loading the cells with the fluorigenic peroxide-sensitive dye 6-carboxy-2',7'-dichlorodihydrofluorescein diacetate (diacetoxymethyl ester), followed by in vitro treatment with H2O2 and fluorescence detection with confocal laser scanning microscopy (CLSM). Using the microgel electrophoresis (“Comet”) Assay, we also examined HT29 stem and clone 19A cells and freshly isolated primary colon cells for their relative sensitivity toward H2O2-induced DNA damage and for steady-state levels of endogenous oxidative DNA damage.

Results A dose-response relationship was found for the H2O2-induced dye decomposition in NIH3T3 cells (7.8-125 μM H2O2) whereas no effect occurred in the human colon tumour cell lines HT29 stem and HT29 clone 19A (62-1000 μM H2O2). Fluorescence was significantly increased at 62 μM H2O2 in the human colon adenocarcinoma cell line Caco-2. In isolated human colon crypts, the lower crypt cells (targets of colon cancer) were more sensitive towards H2O2 than the more differentiated upper crypt cells. In contrast to the CLSM results, oxidative DNA damage was detected in both cell lines using the Comet Assay. Endogenous oxidative DNA damage was highest in HT29 clone 19A, followed by the primary colon cells and HT29 stem cells.

Conclusions Oxidative stress in colon cells leads to damage of macromolecules which is sensitively detected in the Comet Assay. The lacking response of the CLSM-approach in colon tumour cells is probably due to intrinsic modes of protective activities of these cells. In general, however, the CLSM method is a sensitive technique to detect very low concentrations of H2O2-induced oxidative stress in NIH3T3 cells. Moreover, by using colon crypts it provides the unique possibility of assessing cell specific levels of oxidative stress in explanted human tissues. Our results demonstrate that the actual target cells of colon cancer induction are indeed susceptible to the oxidative activity of H2O2.  相似文献   

3.
The toxicity of H2O2 in Escherichia coli wild type and superoxide dismutase mutants was investigated under different experimental conditions. Cells were either grown aerobically, and then treated in M9 salts or K medium, or grown anoxically, and then treated in K medium. Results have demonstrated that the wild type and superoxide dismutase mutants display a markedly different sensitivity to both modes of lethality produced by H2O2 (i.e. mode one killing, which is produced by concentrations of H2O2 lower than 5 mM, and mode two killing which results from the insult generated by concentrations of H2O2 higher than 10 mM). Although the data obtained do not clarify the molecular basis of H2O2 toxicity and/or do not explain the specific function of superoxide ions in H2O2-induced bacterial inactivation, they certainly demonstrate that the latter species plays a key role in both modes of H2O2 lethality. A mechanism of H2O2 toxicity in E. coli is proposed, involving the action of a hypothetical enzyme which should work as an O2-• generating system. This enzyme should be active at low concentrations of H2O2 (<5 mM) and high concentrations of the oxidant (>5 mM) should inactivate the same enzyme. Superoxide ions would then be produced and result in mode one lethality. The resistance at intermediate H2O2 concentrations may be dependent on the inactivation of such enzyme with no superoxide ions being produced at levels of H2O2 in the range 5–10 mM. Mode two killing could be produced by the hydroxyl radical in concert with superoxide ions, chemically produced via the reaction of high concentrations of H2O2 (>10 mM) with hydroxyl radicals. The rate of hydroxyl radical production may be increased by the higher availability of Fe2+ since superoxide ions may also reduce trivalent iron to the divalent form.  相似文献   

4.
The turning point between apoptosis and necrosis induced by hydrogen peroxide (H2O2) have been investigated using human T-lymphoma Jurkat cells. Cells treated with 50 μM H2O2 exhibited caspase-9 and caspase-3 activation, finally leading to apoptotic cell death. Treatment with 500 μM H2O2 did not exhibit caspase activation and changed the mode of death to necrosis. On the other hand, the release of cytochrome c from the mitochondria was observed under both conditions. Treatment with 500 μM H2O2, but not with 50 μM H2O2, caused a marked decrease in the intracellular ATP level; this is essential for apoptosome formation. H2O2-reducing enzymes such as cellular glutathione peroxidase (cGPx) and catalase, which are important for the activation of caspases, were active under the 500 μM H2O2 condition. Prevention of intracellular ATP loss, which did not influence cytochrome c release, significantly activated caspases, changing the mode of cell death from necrosis to apoptosis. These results suggest that ATP-dependent apoptosome formation determines whether H2O2-induced cell death is due to apoptosis or necrosis.  相似文献   

5.
In the present study, triphlorethol-A, a phlorotannin, was isolated from Ecklonia cava and its antioxidant properties were investigated. Triphlorethol-A was found to scavenge intracellular reactive oxygen species (ROS) and 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical, and thus prevented lipid peroxidation. The radical scavenging activity of triphlorethol-A protected the Chinese hamster lung fibroblast (V79-4) cells exposed to hydrogen peroxide (H2O2) against cell death, via the activation of ERK protein. Furthermore, triphlorethol-A reduced the apoptotic cells formation induced by H2O2. Triphlorethol-A increased the activities of cellular antioxidant enzymes like, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). Hence, from the present study, it is suggestive that triphlorethol-A protects V79-4 cells against H2O2 damage by enhancing the cellular antioxidative activity.  相似文献   

6.
Lin HJ  Wang X  Shaffer KM  Sasaki CY  Ma W 《FEBS letters》2004,570(1-3):102-106
In the present study, we characterized hydrogen peroxide (H2O2)-induced cell apoptosis and related cell signaling pathways in cultured embryonic neural stem/progenitor cells (NS/PCs). Our data indicated that H2O2 induced acute cell apoptosis in NS/PC in concentration- and time-dependent manners and selectively, it transiently increased PI3K-Akt and Mek-Erk1/2 in a dose-dependent manner. Inhibition of PI3K-Akt with wortmannin, a PI3-K inhibitor, was found to significantly increase H2O2-induced acute apoptosis and dramatically decrease basal pGSK3β levels. The level of pGSK3β remained unchanged with H2O2 exposure. We conclude that the transient activation of PI3K-Akt signaling delays the H2O2-induced acute apoptosis in cultured NS/PCs in part through maintaining the basal pGSK3β level and activating other downstream effectors.  相似文献   

7.
Three-dimensionally (3D) ordered macroporous active carbon has been fabricated and used as electrode substrate for the direct electrochemistry of horse heart cytochrome c (Cyt c). The Cyt c immobilized on the surface of the ordered macroporous active carbon shows a pair of well-defined and nearly reversible redox waves at the formal potential of −0.033 V in pH 6.8 phosphate buffer solution. The interaction between Cyt c and the 3D macroporous active carbon makes the formal potential shift negatively compared to that of Cyt c in solution. Spectrophotometric and electrochemical methods have been used to investigate the interaction between Cyt c and the porous active carbon. The immobilized Cyt c maintains its biological activity, and shows a surface controlled electrode process with the electron-transfer rate constant (ks) of 17.6 s−1 and the charge-transfer coefficient (a) of 0.52, and displays the features of a peroxidase in the electrocatalytic reduction of hydrogen peroxide (H2O2). A potential application of the Cyt c-immobilized porous carbon electrode as a biosensor to monitor H2O2 has been investigated. The steady-state current response increases linearly with H2O2 concentration from 2.0 × 10−5 to 2.4 × 10−4 mol l−1. The detection limit (3σ) for determination of H2O2 has been found to be 1.46 × 10−5 mol l−1.  相似文献   

8.
The effect of oxygen transfer rate (OTR) on β-carotene production by Blakelsea trispora in shake flask culture was investigated. The results indicated that the concentration of β-carotene (704.1 mg/l) was the highest in culture grown at maximum OTR of 20.5 mmol/(l h). In this case, the percentage of zygospores was over 50.0% of the biomass dry weight. On the other hand, OTR level higher than 20.5 mmol/(l h) was found to be detrimental to cell growth and pigment formation. To elucidate the effect of oxidative stress on β-carotene synthesis, the accumulation of hydrogen peroxide during fermentation under different OTRs was determined. A linear response of β-carotene synthesis to the level of H2O2 was observed, indicating that β-carotene synthesis is stimulated by H2O2. However, there was an optimal concentration of H2O2 (2400 μM) in enhancing β-carotene synthesis. At a higher concentration of H2O2, β-carotene decreased significantly due to its toxicity.  相似文献   

9.
Tea (Camellia sinensis) catechins have been studied for disease prevention. These compounds undergo oxidation and produce H2O2. We have previously shown that holding tea solution or chewing tea leaves generates high salivary catechin levels. Herein, we examined the generation of H2O2 in the oral cavity by green tea solution or leaves. Human volunteers holding green tea solution (0.1-0.6%) developed salivary H2O2 with Cmax = 2.9-9.6 μM and AUC0 → ∞ = 8.5-285.3 μM min. Chewing 2 g green tea leaves produced higher levels of H2O2 (Cmax = 31.2 μM, AUC0 → ∞ = 1290.9 μM min). Salivary H2O2 correlated with catechin levels and with predicted levels of H2O2 (Cmax(expected) = 36 μM vs Cmax(determined) = 31.2 μM). Salivary H2O2 and catechin concentrations were similar to those that are biologically active in vitro. Catechin-generated H2O2 may, therefore, have a role in disease prevention by green tea.  相似文献   

10.
Wu LT  Chu CC  Chung JG  Chen CH  Hsu LS  Liu JK  Chen SC 《Mutation research》2004,556(1-2):75-82
The effect of tannic acid (TA), gallic acid (GA), propyl gallate (PA) and ellagic acid (EA) on DNA damage in human lymphocytes induced by food mutagens [3-amino-1-methyl-5H-pyrido (4,3-b) indole (Trp-P-2) and 2-amino-1-methyl-6-phenylimadazo (4,5-b) pyridine (PhIP) or H2O2 was evaluated by using single-cell electrophoresis (comet assay). The toxicity of these tested compounds (0.1–100 μg/ml) on lymphocytes was not found. These compounds did not cause DNA strand breaks at lower concentrations of 0.1–10 μg/ml. At a concentration of 100 μg/ml, TA and GA exhibited slight DNA damage, whereas PA and EA showed no DNA strand breaks. TA and its related compounds decreased the DNA strand breaks induced by Trp-P-2, PhIP or H2O2 at concentrations of 0.1–10 μg/ml. DNA repair enzymes endonuclease III (Endo III) and formamidopyrimidine-DNA glycoslase (FPG)] were used to examine the levels of oxidised pyrimidines and purines in human lymphocytes induced by H2O2. All the compounds at 10 μg/ml can reduce the level of FPG sensitive sites. However, only EA inhibited the formation of EndoIII sensitive sites. The results indicated that these compounds can enhance lymphocytes resistance towards DNA strand breaks induced by food mutagens or H2O2 in vitro.  相似文献   

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