ascorbic-acid and 2--7--dichlorodihydrofluorescein

How single-walled carbon nanotubes (SWCNT) function in redox reactions may be related to their behaviors in the induction of oxidative stress. Herein, oxidation of several biologically relevant reducing agents in the presence of SWCNT was studied in aqueous solutions. The selected reductants included a common indicator for intracellular reactive oxygen species (ROS) (2,7-dichlorodihydrofluorescein), small antioxidants (vitamin C, Trolox, and cysteine), and a high-molecular-weight ROS scavenger (bovine serum albumin). The unmodified or carboxylated SWCNT acted as both the oxidants and the catalysts in reactions. Moreover, they accelerated the oxidation reactions mediated by horseradish peroxidase, a representative member of the enzyme family actively involved in balancing oxidative stress. These diverse roles in redox reactions may serve the chemistry basis for SWCNT to induce oxidative stress in biological systems as potential environmental pollutants.

Topics: Animals; Argon; Ascorbic Acid; Cattle; Fluoresceins; Horseradish Peroxidase; Kinetics; Nanotubes, Carbon; Oxidation-Reduction; Serum Albumin, Bovine; Solutions; Spectrophotometry, Ultraviolet; Water

Oxygen enhancement of tumor radiosensitivity is attributed to DNA damage by reactive oxygen species. The mechanism remains unclear but may involve mitochondria as major sources of oxygen and nitrogen radicals as well as central effectors of energy homeostasis and apoptosis. Here we used dihydrorhodamine and 2',7'-dichlorodihydrofluorescein to compare mitochondrial and total cell generation, respectively, of reactive oxygen or nitrogen species in cells irradiated at 5 Gy. Irradiation in the presence of oxygen selectively stimulated mitochondrial radical production in HeLa and MeWo cells, but in MCF7 cells radical production was more generalized. In all three cell lines oxygen impaired cell proliferation as measured by resazurin reduction 7 days after irradiation. Antioxidants N-acetylcysteine, ascorbic acid, and melatonin largely prevented dye oxidation during normoxic irradiation yet had no effect on oxygen-dependent irradiation injury. However, NO synthase inhibitor N(G)-monomethyl-L-arginine protected HeLa and MCF7 though not MeWo cells, consistent with their different levels of constitutive NO generation. SB203580 inhibition of p38 MAPK appreciably protected HeLa and marginally protected MCF7 cells against oxygen-dependent irradiation injury, while the less specific JNK/SAPK inhibitor SP600125 and ERK inhibitor U0126 had no effect. None of the inhibitors affected MeWo radiosensitivity. Therefore oxygen-enhanced radiosensitivity in these tumor cell lines does not depend on extensive production of oxygen radicals and is cell-type dependent. NO mediates oxygen-dependent injury in HeLa and MCF7 cells, by p38-dependent and MAPK-independent mechanisms, respectively. In MeWo cells this oxygen-enhanced radiosensitivity is independent of both NO and MAPK signaling.

Topics: Acetylcysteine; Anthracenes; Antioxidants; Apoptosis; Ascorbic Acid; Butadienes; Cell Line, Tumor; Cell Proliferation; DNA Damage; Enzyme Inhibitors; Fluoresceins; HeLa Cells; Humans; Imidazoles; MAP Kinase Signaling System; Melatonin; Mitochondria; Nitriles; Nitrites; Oxazines; Oxygen; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Pyridines; Radiation Tolerance; Rhodamines; Spectrometry, Fluorescence; Xanthenes

In this study, we investigated the structure-activity relationship of four flavonoids, i.e. eriodictyol, luteolin, quercetin, and taxifolin, in cultured retinal cells after ascorbate/Fe(2+)-induced oxidative stress. The relative order of antioxidant efficacy, determined by the thiobarbituric acid method, was the following: eriodictyol > quercetin > luteolin > taxifolin. Upon preincubation, the flavonoids were also effective in reducing the extent of lipid peroxidation. Oxidative stress, determined by the changes in fluorescence of 2',7'-dichlorodihydrofluorescein, was also decreased in the presence of the flavonoids, showing the following order of antioxidant efficacy: eriodictyol > taxifolin approximately quercetin > luteolin. Ascorbate/Fe(2+)-induced oxidative stress or incubation in the presence of the flavonoids did not significantly affect the viability of retinal cells. We also evaluated the degree of membrane partition of the flavonoids. In this system, the results strongly suggest that the higher antioxidant activity of the flavonoids is not correlated with the presence of a double bond at C(2)-C(3) and/or a hydroxyl group at C(3) on the C ring, but rather may depend on the capacity to inhibit the production of reactive oxygen species to interact hydrophobically with membranes. Eriodictyol was shown to be the most efficient antioxidant in protecting against oxidative stress induced by ascorbate/Fe(2+) in the retinal cells.

Topics: Animals; Antioxidants; Ascorbic Acid; Cell Survival; Cells, Cultured; Chick Embryo; Drug Interactions; Ferrous Compounds; Flavanones; Flavonoids; Fluoresceins; Lipid Peroxidation; Oxidative Stress; Reactive Oxygen Species; Retina; Structure-Activity Relationship

The purpose of this study was to examine distribution and time history of oxidative stress during the hyperacute period of reperfusion in the liver grafts undergoing cold ischemia and to investigate roles of Kupffer cells as a potential oxidant source. Rat livers were harvested at 4 degrees C in University of Wisconsin solution and followed by reperfusion with Krebs-Henseleit buffer under monitoring bile excretion. To investigate oxidative changes, laser-confocal microfluorography was performed in reperfused livers preloaded with dichlorodihydrofluorescein diacetate succinimidyl ester, a fluorescence precursor sensing intracellular hydroperoxide generation. Livers undergoing the 16-hour cold storage displayed an impaired recovery of bile acid-dependent bile output concurrent with a marked increase in hydroperoxide generation in hepatocytes, which occurred as early as 5 minutes after the onset of reperfusion, whereas the status of lobular perfusion was well maintained. Pretreatment with liposome-encapsulated dichloromethylene diphosphonate, a Kupffer cell-depleting reagent, did neither alter the reperfusion-induced periportal oxidative changes nor improve the recovery of bile output in the graft. On the other hand, EPCK, a hepatotropic antioxidant composed of vitamin E phosphate ester bound to vitamin C, not only diminished the oxidative changes but also improved the reduction of bile acid-dependent bile output. Furthermore, the reagent was capable of inhibiting H(2)O(2)-induced oxidative stress in cultured hepatocytes. These results suggest that hepatocytes constitute a major site of the oxidative insult triggered through Kupffer cell-independent mechanisms and serve as an important cellular component to be protected by antioxidant therapeutics.

Topics: Animals; Antioxidants; Ascorbic Acid; Bile; Clodronic Acid; Cold Temperature; Deferoxamine; Fluoresceins; Hydrogen Peroxide; Kupffer Cells; Liver; Liver Transplantation; Male; Mitochondria, Liver; Oxidative Stress; Rats; Rats, Wistar; Reperfusion Injury; Taurocholic Acid; Vitamin E