Activation Of Nicotinamide Adenine Dinucleotide Phosphate Research Articles

Downregulation of p22phox in Retinal Pigment Epithelial Cells Inhibits Choroidal Neovascularization in Mice

Choroidal neovascularization (CNV) occurs in a variety of chorioretinal diseases including age-related macular degeneration (AMD), and is the major cause of severe visual loss in patients with AMD. Oxidative stress has been thought to play an important role in the development of CNV. Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase is one of the major intracellular sources of reactive oxygen species (ROS) in the vascular system. In this study, we examined the expression of p22phox, an integral subunit in the NADPH oxidase complex, in the mouse eye. We determined that p22phox is expressed in the retinal pigment epithelial (RPE) cells and inner retinal neurons. A small-interfering RNA (siRNA) designed against p22phox efficiently reduced the expression of the protein in the eye when delivered by means of recombinant adeno-associated virus (AAV) vector. Vector treatment inhibited CNV in the mouse when delivered into the subretinal space where RPE cells were transduced. These results suggest that NADPH oxidase-mediated ROS production in RPE cells may play an important role in the pathogenesis of neovascular AMD, and that this pathway may represent a new target for therapeutic intervention in AMD.

Arthritis suppression by NADPH activation operates through an interferon-β pathway

BackgroundA polymorphism in the activating component of the nicotinamide adenine dinucleotide phosphate (NADPH) oxidase complex, neutrophil cytosolic factor 1 (NCF1), has previously been identified as a regulator of arthritis severity in mice and rats. This discovery resulted in a search for NADPH oxidase-activating substances as a potential new approach to treat autoimmune disorders such as rheumatoid arthritis (RA). We have recently shown that compounds inducing NCF1-dependent oxidative burst, e.g. phytol, have a strong ameliorating effect on arthritis in rats. However, the underlying molecular mechanism is still not clearly understood. The aim of this study was to use gene-expression profiling to understand the protective effect against arthritis of activation of NADPH oxidase in the immune system.ResultsSubcutaneous administration of phytol leads to an accumulation of the compound in the inguinal lymph nodes, with peak levels being reached approximately 10 days after administration. Hence, global gene-expression profiling on inguinal lymph nodes was performed 10 days after the induction of pristane-induced arthritis (PIA) and phytol administration. The differentially expressed genes could be divided into two pathways, consisting of genes regulated by different interferons. IFN-γ regulated the pathway associated with arthritis development, whereas IFN-β regulated the pathway associated with disease protection through phytol. Importantly, these two molecular pathways were also confirmed to differentiate between the arthritis-susceptible dark agouti (DA) rat, (with an Ncf-1DA allele that allows only low oxidative burst), and the arthritis-protected DA.Ncf-1E3 rat (with an Ncf1E3 allele that allows a stronger oxidative burst).ConclusionNaturally occurring genetic polymorphisms in the Ncf-1 gene modulate the activity of the NADPH oxidase complex, which strongly regulates the severity of arthritis. We now show that the Ncf-1 allele that enhances oxidative burst and protects against arthritis is operating through an IFN-β-associated pathway, whereas the arthritis-driving allele operates through an IFN-γ-associated pathway. Treatment of arthritis-susceptible rats with an NADPH oxidase-activating substance, phytol, protects against arthritis. Interestingly, the treatment led to a restoration of the oxidative-burst effect and induction of a strikingly similar IFN-β-dependent pathway, as seen with the disease-protective Ncf1 polymorphism.

Role of Oxidative Stress in Cardiac Hypertrophy and Remodeling

Cardiac adaptation in response to intrinsic or external stress involves a complex process of chamber remodeling and myocyte molecular modifications. A fundamental response to increased biomechanical stress is cardiomyocyte and chamber hypertrophy. Although this may provide initial salutary compensation to the stress, sustained hypertrophic stimulation becomes maladaptive, worsening morbidity and mortality risks because of congestive heart failure and sudden death.1 Growing evidence highlights oxidative and nitrosative stresses as important mechanisms for this maladaptation.2–9 Oxidative stress occurs when excess reactive oxygen species (ROS) are generated that cannot be adequately countered by intrinsic antioxidant systems. Superoxide anion (O2−) can further combine with NO, forming reactive compounds such as peroxynitrite, generating nitroso-redox imbalance.4 ROS generation is a normal component of oxidative phosphorylation and plays a role in normal redox control of physiological signaling pathways.5,8,9 However, excessive ROS generation triggers cell dysfunction, lipid peroxidation, and DNA mutagenesis and can lead to irreversible cell damage or death.5,8,9 In this review, we discuss recent experimental evidence for the role of oxidant stress on cardiac remodeling, focusing on pressure- overload–induced hypertrophy and dilation. ROS include free radicals such as superoxide (O2−) and hydroxyl radical and compounds such as hydrogen peroxide (H2O2) that can be converted to radicals, and they participate in both normal and pathologic biochemical reactions.9 O2− is formed intracellularly (Figure 1) by activation of nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase or xanthine oxidase (XO), uncoupling of NO synthase (NOS), and electron transport and “leakage” during oxidative phosphorylation in the mitochondria.5,8,9 H2O2 can generate the highly reactive hydroxyl radical via Fenton chemistry under pathological conditions.9 Figure 1. General schematic of generation pathways for ROS and antioxidant systems in the heart. Low levels of ROS are thought to …

In vivo evidence for antioxidant potential of estrogen in microvessels of female spontaneously hypertensive rats.

In studies conducted in vitro, it has been demonstrated that estrogen has an antioxidant potential that may contribute to its protective effects on the cardiovascular system. However, the antioxidant effect of estrogen in vivo has not been demonstrated. To address this issue, in this study the effects of estrogen on oxidative stress were evaluated in microvessels studied in vivo. Oxidative stress was evaluated by using intravital microscopy in mesenteric arterioles from female spontaneously hypertensive rats (SHR) in physiological estrous (OE), ovariectomized (OVX), OVX treated with estradiol (E(2)), or estradiol + progesterone (E/P). The mesenteries were superfused with hydroethidine, a reduced and nonfluorescent precursor of ethidium bromide (EB). In the presence of reactive oxygen species, hydroethidine is transformed intracellularly in EB, which binds to DNA and can be detected by its red fluorescence. The percentage of EB-positive nuclei along the arteriolar wall in OVX (28.4 +/- 4.3) was significantly increased compared with OE (14.2 +/- 3.9; P<0.05). The OVX overproduction of oxyradicals was attenuated by E(2) (15.7 +/- 2.2) and E/P (14.8 +/- 0.8). Treatment with the superoxide dismutase mimetic MnTMPyP attenuated by 75% the oxidation of hydroethidine in both OE and OVX. Conversely, mannitol, that decomposes hydroxyl radical, and L-NAME, a nitric oxide synthase inhibitor, had no significant effects on hydroethidine oxidation. No differences on hydrogen peroxide plasma concentration were observed among the groups, suggesting that superoxide anion is the most likely oxyradical involved in the increased oxidative stress observed in OVX. The treatment of mesenteries with diphenyleneiodonium (DPI), an nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase inhibitor, but not with oxypurinol, a xanthine-oxidase inhibitor, produced a significant reduction of oxyradical generation in OVX microvessels and a slight decrease in those from OE. Chronic treatment of female SHR with losartan caused similar decreases in oxyradicals in both OE and OVX, whereas diclofenac and verapamil had no effects. Together these data suggest that estrogen reduces superoxide anion bioavailability in vivo. The antioxidant effect of estrogen, which can contribute to a less pronounced endothelial dysfunction in female SHR, may be dependent on a direct modulatory action of estrogen on NADPH activity.

Zinc inhibition of respiratory burst in zymosan-stimulated neutrophils: a possible membrane action of zinc.

The effect of Zn2+ on the respiratory burst of rat pleural neutrophils was studiesd. Serum treated zymosan (STZ)-stimulated O2 consumption was inhibited by Zn2+ depending on hte Zn2+ concentration and time of cell incubation. Addition of Zn2+ after the stimulation of cells with STZ did not inhibit the O2 consumption Zn2+ failed to affect the cell viability or the opsonizing process of STZ, indicating that the inhibition of O2 consumption was not due to te direct cytotoxic action of Zn2+ or to the interaction of Zn2+ with STZ. In addition, the activity of reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, an enzyme responsile for the respiratory burst of neutrophils, was not inhibited by Zn2+. These results suggested that Zn2+ may inhibit some process of the activation of NADPH oxidase.Equimolar Zn2+ 8-hydroxyquinoline complex (Zn-8HQ), which is known to be unable to penetrate the cell membrane, inhibited the O2 consumption more drastically than did Zn2+ alone. 8-Hydroxyquinoline alone did not cause significant inhibition of O2 consumption, indicating that the inhibitory effect of Zn2+ is potentiated by complexing with 8-hydrozyquinoline. The inhibition of O2 consumption Zn2+ was almost completely restored by washing and reincubating the Zn2+-treated cells in Zn2+-free medium. On the other hand, in the cells treated with Zn-8HQ, the same treatment of cell washing and reincubation only partially restored the O2 consumption. These results suggested that Zn2+ may be acting on the cell membrane of neutrophils.