dihydrorhodamine 123 oxidation test

1. Oxidative stress induced by reactive oxygen species (ROS) is a key mediator of haemorrhagic shock (HS)-induced vascular hyperpermeability. In the present study, curcumin, a natural anti-oxidant obtained from turmeric (Curcuma longa), was tested against HS-induced hyperpermeability and associated ROS formation in rat mesenteric post-capillary venules in vivo and in rat lung microvascular endothelial cells (RLMEC) in vitro. 2. In rats, HS was induced by withdrawing blood to reduce mean arterial pressure to 40 mmHg for 60 min, followed by resuscitation for 60 min...

Mycoplasma bovis (M. bovis) contributes to a number of clinical syndromes in cattle; in particular, chronic pneumonia that is poorly responsive to therapy has been increasingly recognized as an important cause of morbidity, mortality, and financial loss. M. bovis impairs host immune function, but little is known about whether field isolates vary significantly in their effect on immune function. This research tested the hypothesis that different field isolates vary in their ability to suppress cellular metabolism and cellular production of radical oxygen species (ROS) by bovine leukocytes...

PURPOSE: The production of reactive oxygen species (ROS) can lead to oxidative stress, which is a strong contributory factor to many ocular diseases. In this study, the removal of trophic factors is used as a model system to investigate the effects of stress in the retina. The aims were to determine if both rod and cone photoreceptor cells produce ROS when they are deprived of trophic factor support and to demonstrate if the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (Nox) enzymes are responsible for this ROS production...

Sepsis-induced acute kidney injury (AKI) is a complex disease characterized by generation of inducible nitric oxide synthase (iNOS)-derived reactive nitrogen species (RNS) by the renal tubular epithelium. While most in vitro models of sepsis use combinations of lipopolysaccharide and cytokines to simulate exposure to inflammatory mediators thought to play a role in sepsis, the relevance of these models is limited. To address the need for a model that more closely mimics the tubular microenvironment during sepsis, we developed an in vitro model where mIMCD-3 (murine tubular epithelial) cells are treated with media containing 5% serum collected from mice at 4 h after cecal ligation and puncture (CLP) or sham surgery (no sepsis)...

The evaluation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) production by neutrophils is currently a matter of extensive research, with scientific reports showing an enormous variability on the detection methods as well as the concentration of the detecting probes. Also the incubation media used to test neutrophils and the respective ionic concentration, as well as the glucose concentration, varies enormously from study to study. This variability often results in different sensibility and/or response of neutrophils to stimulating agents, which can be a focus of confounding and sometime contradictory results among reports...

Growth factors are important in the development, maintenance and repair of cartilage. The principal aim of this study was to test the capacity of three growth factors with established roles in cartilage, namely insulin-like growth factor (IGF)-1, fibroblast growth factor (FGF) and transforming growth factor (TGF)-beta 1, to alter intracellular reactive oxygen species (ROS) levels. Explants of articular cartilage from young, mature, and aged rats were pretreated with IGF-1, FGF, or TGF-beta 1 and intracellular ROS levels were quantified using the free radical sensing probe dihydrorhodamine 123 (DHR 123), confocal microscopy, and densitometric image analysis...

Polyurethane cardiovascular implants are subject to oxidation initiated surface degradation, which is mediated by monocyte-derived macrophages (MDM); this often leads to surface cracking and device failure. The present studies examined the hypothesis that covalently attaching antioxidant, di-tert-butylphenol (DBP), to the urethane nitrogens of a polyether polyurethane (PU) via bromo-alkylation reactions could prevent this problem. PU was configured with two dosages of DBP, 0.14 mM DBP/g PU of DBP (PU-DBP) and a more highly modified (HM) 0...

Neutrophil dysfunction can result from oxidative burst defect or from glucose-6-phosphate dehydrogenase (G6PD) deficiency; we noted both in the same patient. A 4-month-old male infant with G6PD deficiency presented with swelling of the left middle finger, left leg, and right big toe. At 5 weeks of age he was hospitalized for fever for 2 days. A maternal uncle died at 5 years of age and a male maternal cousin died at the age of 21 months, both reportedly diagnosed with chronic granulomatous disease (CGD). On physical examination, he had a swollen erythematous left third finger, left distal leg swelling, and right big toe abscess...

The effect of oxidative stress induced by the ascorbate/menadione-redox association was examined in K562 cells, a human erythromyeloid leukaemia cell line. Our results show that ascorbate enhances menadione redox cycling, leading to the formation of intracellular reactive oxygen species (as shown by dihydrorhodamine 123 oxidation). The incubation of cells in the presence of both ascorbate/menadione and aminotriazole, a catalase inhibitor, resulted in a strong decrease of cell survival, reinforcing the role of H(2)O(2) as the main oxidizing agent killing K562 cells...

Although increasing evidence shows the nutritional benefits of calcium fructoborate (CF) on animals and humans, its action mechanism has not been clearly identified. The present study aims to investigate the possible antioxidant function of CF. Based on its efficiency in skin wound healing, the authors tested whether CF possesses antioxidant properties on human keratinocytes cultures, in a complete serum-free medium (KMK-2; Sigma). The cells treated with CF (0-450 nmol/culture medium) were exposed to exogenous 100 micromol of hydrogen peroxide to mimic the oxidative stress...

The yeast Candida intermedia, as a model organism, was used to examine the links between the metal ions exposure, reactive oxygen species generation and oxidative stress response. To estimate intracellular peroxide and superoxide levels, the fluorescence indicators dihydrorhodamine 123 and dihydroethidium were used, respectively. Antioxidant defence systems were investigated by measuring the activity of catalase, glutathione peroxidase and superoxide dismutase and the content of reduced glutathione. Altered superoxide, peroxide, glutathione levels, and the catalase activity were perceived after the treatment with copper...

Diphenyleneiodonium (DPI) inhibits activity of flavoenzymes like NADPH oxidase, the major source of superoxide anion in cardiovascular system, but affects also other oxidoreductases. Contradictory data have been published concerning the effect of diphenyleneiodonium on the production of reactive oxygen species in cells, both inhibitory and stimulatory action of DPI being reported. We have examined the effect of DPI on the cellular production of reactive oxygen and nitrogen species (ROS/RNS) and on the proliferation and apoptosis of human vascular endothelial cells...

A microchip electrophoresis method coupled with laser-induced fluorescence (LIF) detection was established for simultaneous determination of two kinds of intracellular signaling molecules (reactive oxygen species, ROS, and reduced glutathione, GSH) related to apoptosis and oxidative stress. As the probe dihydrorhodamine-123 (DHR-123) can be converted intracellularly by ROS to the fluorescent rhodamine-123 (Rh-123), and the probe naphthalene-2,3-dicarboxaldehyde (NDA) can react quickly with GSH to produce a fluorescent adduct, rapid determination of Rh-123 and GSH was achieved on a glass microchip within 27 s using a 20 mM borate buffer (pH 9...

Three hydroquinone glucosides and four caffeoylquinic esters were examined for their effect on tyrosine nitration, as well as on the oxidation of dihydrorhodamine (DHR) 123 and cytochrome c(2+) induced by peroxynitrite. All these phenolics, which had previously been characterized as the active principles of the plant Phagnalon rupestre, were fairly active in preventing the oxidation of DHR 123, though inefficient in the cytochrome c test. While their antioxidant potency is associated with the presence of a caffeoyl moiety, not so an obvious chemical character was correlated to a greater activity against nitration of tyrosine...

BACKGROUND: Hemin, the oxidized form of heme, accumulates in intracranial hematomas and is a potent oxidant. Growing evidence suggests that it contributes to delayed injury to surrounding tissue, and that this process is affected by the heme oxygenase enzymes. In a prior study, heme oxygenase-2 gene deletion increased the vulnerability of cultured cortical astrocytes to hemin. The present study tested the effect of HO-2 gene deletion on protein oxidation, reactive oxygen species formation, and cell viability after mixed cortical neuron/astrocyte cultures were incubated with neurotoxic concentrations of hemin...

Two monohydroxamates of l-aspartic acid beta-hydroxamate (AAH) and l-glutamic acid gamma-hydroxamate (GAH) were used for testing antioxidant and angiotensin converting enzyme (ACE) inhibitory activities in comparison with those of asparagine and glutamine, respectively. The half-inhibition concentrations, IC(50), of scavenging activity against 1,1-diphenyl-2-picrylhydrazyl (DPPH) were 36 and 48 microM and against superoxide radicals were 18.99 and 6.33 mM, respectively, for AAH and GAH. However, no activities of asparagine and glutamine were found...

Ultra-high molecular weight polyethylene (UHMWPE) is a biopolymer widely used in orthopaedic implants and its oxidation is considered as major responsible for inflammation and the prosthesis failure. We have studied the effect on the activation of resting human granulocytes of the addition of Vitamin E (Vit.E, alpha-tocopherol), a natural biological antioxidant and antiinflammatory agent, to UHMWPE. We have measured changes in granulocytes morphology and respiratory burst by flow cytometry using Dihydrorhodamine 123 and matrix metalloproteinase 9 (MMP-9, gelatinase B) release and activity in the growth medium using substrate zymography following contact (60 min at 37 degrees C) with cell grade polystyrene (PS), normal UHMWPE (PE) and Vit...

When cortical neurons are exposed to hemoglobin, they undergo oxidative stress that ultimately results in iron-dependent cell death. Heme oxygenase (HO)-2 is constitutively expressed in neurons and catalyzes heme breakdown. Its role in the cellular response to hemoglobin is unclear. We tested the hypothesis that HO-2 attenuates hemoglobin neurotoxicity by comparing reactive oxygen species (ROS) formation and cell death in wild-type and HO-2 knockout cortical cultures. Consistent with prior observations, hemoglobin increased ROS generation, detected by fluorescence intensity after dihydrorhodamine 123 or dichlorofluorescin-diacetate loading, in wild-type neurons...

Leukocytosis in tobacco smokers has been well recognized; however, the exact cause has not been elucidated. To test the hypothesis that tobacco nicotine stimulates neutrophils in the respiratory tract to produce IL-8, which causes neutrophilia in vivo, we examined whether nicotine induces neutrophil-IL-8 production in vitro; the causative role of NF-kappaB in its production, in association with the possible production of reactive oxygen intermediates that activate NF-kappaB; and the nicotinic acetylcholine receptors (nAChRs) involved in IL-8 production...

The potato (Solanum tuberosum L.) tuber storage protein, patatin, was purified to homogeneity with a molecular mass of 45 kDa. The purified patatin showed antioxidant or antiradical activity by a series of in vitro tests, including 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical (half-inhibition concentration, IC(50), was 0.582 mg/mL) scavenging activity assays, anti-human low-density lipoprotein peroxidation tests, and protections against hydroxyl radical-mediated DNA damages and peroxynitrite-mediated dihydrorhodamine 123 oxidations...