Carbonate Anion Radical Generated by the Peroxidase Activity of Copper-Zinc Superoxide Dismutase:Scavenging of Radical and Protection of Enzyme by Hypotaurine and Cysteine Sulfinic Acid

Copper-zinc superoxide dismutase (SOD) is considered one of the most important mammalian antioxidant defenses and plays a relevant role due to its main function in catalyzing the dismutation of superoxide anion to oxygen and hydrogen peroxide. However, interaction between SOD and H2O2 produced a strong copper-bound oxidant (Cu(II)●OH) that seems able to contrast the self-inactivation of the enzyme or oxidize other molecules through its peroxidase activity. The bicarbonate presence enhances the peroxidase activity and produces the carbonate anion radical (CO3●–). CO3●– is a freely diffusible reactive species capable of oxidizing several molecules that are unwieldy to access into the reactive site of the enzyme. Cu(II)●OH oxidizes bicarbonate to the CO3●–, which spreads out of the binding site and oxidizes hypotaurine and cysteine sulfinic acid to the respective sulfonates through an efficient reaction. These findings suggest a defense role for sulfinates against the damage caused by CO3●–. The effect of hypotaurine and cysteine sulfinic acid on the CO3●–-mediated oxidation of the peroxidase probe ABTS to ABTS cation radical (ABTS●+) has been studied. Both sulfinates are able to inhibit the oxidation of ABTS mediated by CO3●–. The effect of hypotaurine and cysteine sulfinic acid against SOD inactivation by H2O2 (~42% protection of enzyme activity) has also been investigated. Interestingly, hypotaurine and cysteine sulfinic acid partially avoid the H2O2-mediated SOD inactivation, suggesting that the two sulfinates may have access to the SOD reactive site and preserve it by reacting with the copper-bound oxidant. In this way hypotaurine and cysteine sulfinic acid not only intercept CO3●–which could move out from the reactive site and cause oxidative damage, but also prevents the inactivation of SOD

Comparative study of the antioxidant and reactive oxygen species scavenging properties in the extracts of the fruits of Terminalia chebula, Terminalia belerica and Emblica officinalis

<p>Abstract</p> <p>Background</p> <p>Cellular damage caused by reactive oxygen species (ROS) has been implicated in several diseases, and hence natural antioxidants have significant importance in human health. The present study was carried out to evaluate the <it>in vitro </it>antioxidant and reactive oxygen species scavenging activities of <it>Terminalia chebula</it>, <it>Terminalia belerica </it>and <it>Emblica officinalis </it>fruit extracts.</p> <p>Methods</p> <p>The 70% methanol extracts were studied for <it>in vitro </it>total antioxidant activity along with phenolic and flavonoid contents and reducing power. Scavenging ability of the extracts for radicals like DPPH, hydroxyl, superoxide, nitric oxide, hydrogen peroxide, peroxynitrite, singlet oxygen, hypochlorous acid were also performed to determine the potential of the extracts.</p> <p>Results</p> <p>The ability of the extracts of the fruits in exhibiting their antioxative properties follow the order <it>T. chebula </it>><it>E. officinalis </it>><it>T. belerica</it>. The same order is followed in their flavonoid content, whereas in case of phenolic content it becomes <it>E. officinalis </it>><it>T. belerica </it>><it>T. chebula</it>. In the studies of free radicals' scavenging, where the activities of the plant extracts were inversely proportional to their IC₅₀values, <it>T. chebula </it>and <it>E. officinalis </it>were found to be taking leading role with the orders of <it>T. chebula </it>><it>E. officinalis </it>><it>T. belerica </it>for superoxide and nitric oxide, and <it>E. officinalis </it>><it>T. belerica </it>><it>T. chebula </it>for DPPH and peroxynitrite radicals. Miscellaneous results were observed in the scavenging of other radicals by the plant extracts, viz., <it>T. chebula </it>><it>T. belerica </it>><it>E. officinalis </it>for hydroxyl, <it>T. belerica </it>><it>T. chebula </it>><it>E. officinalis </it>for singlet oxygen and <it>T. belerica </it>><it>E. officinalis </it>><it>T. chebula </it>for hypochlorous acid. In a whole, the studied fruit extracts showed quite good efficacy in their antioxidant and radical scavenging abilities, compared to the standards.</p> <p>Conclusions</p> <p>The evidences as can be concluded from the study of the 70% methanol extract of the fruits of <it>Terminalia chebula</it>, <it>Terminalia belerica </it>and <it>Emblica officinalis</it>, imposes the fact that they might be useful as potent sources of natural antioxidant.</p

The Interaction of Hypotaurine and Other Sulfinates with Reactive Oxygen and Nitrogen Species:A Survey of Reaction Mechanisms

Considerable strides have been made in understanding the oxidative mechanisms involved in the final steps of the cysteine pathway leading to taurine. The oxidation of sulfinates, hypotaurine and cysteine sulfinic acid, to the respective sulfonates, taurine and cysteic acid, has never been associated with any specific enzyme. Conversely, there is strong evidence that in vivo formation of taurine and cysteic acid is the result of sulfinate interaction with a variety of biologically relevant oxidants. In the last decade, many experiments have been performed to understand whether peroxynitrite, nitrogen dioxide and carbonate radical anion could be included in the biologically relevant reactive species capable of oxidizing sulfinates. Thanks to this work, it has been possible to highlight two possible reaction mechanisms (direct and indirect reaction) of sulfinates with reactive oxygen and nitrogen species.The sulfinates oxidation, mediated by peroxynitrite, is an example of both reaction mechanisms: through a two-electron-direct-reaction with peroxynitrite or through a one-electron-indirect-transfer reaction. In the indirect mechanism, the peroxynitrite homolysis releases hydroxyl and nitrogen dioxide radical and in addition the degradation of short-lived adduct formed by peroxynitrite and CO2 can generate carbonate radical anion. The reaction of hypotaurine and cysteine sulfinic acid with peroxynitrite-derived radicals is accompanied by extensive oxygen uptake with the generation of transient intermediates, which can begin a reaction by an oxygen-dependent mechanism with the sulfonates, taurine, and cysteic acid as final products. Due to pulse radiolysis studies, it has been shown that transient sulfonyl radicals (RSO2(•)) have been produced during the oxidation of both sulfinates by one-electron transfer reaction.The purpose is to analyze all the aspects of the reactive mechanism in the sulfinic group oxidation of hypotaurine and cysteine sulfinic acid through the results obtained from our laboratory in recent years

Photovoltaic restoration of sight with high visual acuity

Patients with retinal degeneration lose sight due to the gradual demise of photoreceptors. Electrical stimulation of surviving retinal neurons provides an alternative route for the delivery of visual information. We demonstrate that subretinal implants with 70-μm-wide photovoltaic pixels provide highly localized stimulation of retinal neurons in rats. The electrical receptive fields recorded in retinal ganglion cells were similar in size to the natural visual receptive fields. Similarly to normal vision, the retinal response to prosthetic stimulation exhibited flicker fusion at high frequencies, adaptation to static images and nonlinear spatial summation. In rats with retinal degeneration, these photovoltaic arrays elicited retinal responses with a spatial resolution of 64 ± 11 μm, corresponding to half of the normal visual acuity in healthy rats. The ease of implantation of these wireless and modular arrays, combined with their high resolution, opens the door to the functional restoration of sight in patients blinded by retinal degeneration

eNOS Protects from Atherosclerosis Despite Relevant Superoxide Production by the Enzyme in apoE−/− Mice

All three nitric oxide synthase (NOS) isoforms are expressed in atherosclerotic plaques. NOS enzymes in general catalyse NO production. However, under conditions of substrate and cofactor deficiency, the enzyme directly catalyse superoxide formation. Considering this alternative chemistry, the effects of NOS on key events in spontaneous hyperlipidemia driven atherosclerosis have not been investigated yet. Here, we evaluate how endothelial nitric oxide synthase (eNOS) modulates leukocyte/endothelial- (L/E) and platelet/endothelial- (P/E) interactions in atherosclerosis and the production of nitric oxide (NO) and superoxide by the enzyme. Intravital microscopy (IVM) of carotid arteries revealed significantly increased L/E-interactions in apolipoproteinE/eNOS double knockout mice (apoE(-/-)/eNOS(-/-)), while P/E-interactions did not differ, compared to apoE(-/-). eNOS deficiency increased macrophage infiltration in carotid arteries and vascular cell adhesion molecule-1 (VCAM-1) expression, both in endothelial and smooth muscle cells. Despite the expression of other NOS isoforms (inducible NOS, iNOS and neuronal NOS, nNOS) in plaques, Electron Spin Resonance (ESR) measurements of NO showed significant contribution of eNOS to total circulating and vascular wall NO production. Pharmacological inhibition and genetic deletion of eNOS reduced vascular superoxide production, indicating uncoupling of the enzyme in apoE(-/-) vessels. Overt plaque formation, increased vascular inflammation and L/E- interactions are associated with significant reduction of superoxide production in apoE(-/-)/eNOS(-/-) vessels. Therefore, lack of eNOS does not cause an automatic increase in oxidative stress. Uncoupling of eNOS occurs in apoE(-/-) atherosclerosis but does not negate the enzyme's strong protective effects

Attenuation of Skeletal Muscle and Renal Injury to the Lower Limb following Ischemia-Reperfusion Using mPTP Inhibitor NIM-811.

INTRODUCTION: Operation on the infrarenal aorta and large arteries of the lower extremities may cause rhabdomyolysis of the skeletal muscle, which in turn may induce remote kidney injury. NIM-811 (N-metyl-4-isoleucine-cyclosporine) is a mitochondria specific drug, which can prevent ischemic-reperfusion (IR) injury, by inhibiting mitochondrial permeability transition pores (mPTP). OBJECTIVES: Our aim was to reduce damages in the skeletal muscle and the kidney after IR of the lower limb with NIM-811. MATERIALS AND METHODS: Wistar rats underwent 180 minutes of bilateral lower limb ischemia and 240 minutes of reperfusion. Four animal groups were formed called Sham (receiving vehicle and sham surgery), NIM-Sham (receiving NIM-811 and sham surgery), IR (receiving vehicle and surgery), and NIM-IR (receiving NIM-811 and surgery). Serum, urine and histological samples were taken at the end of reperfusion. NADH-tetrazolium staining, muscle Wet/Dry (W/D) ratio calculations, laser Doppler-flowmetry (LDF) and mean arterial pressure (MAP) monitoring were performed. Renal peroxynitrite concentration, serum TNF-alpha and IL-6 levels were measured. RESULTS: Less significant histopathological changes were observable in the NIM-IR group as compared with the IR group. Serum K+ and necroenzyme levels were significantly lower in the NIM-IR group than in the IR group (LDH: p<0.001; CK: p<0.001; K+: p = 0.017). Muscle mitochondrial viability proved to be significantly higher (p = 0.001) and renal function parameters were significantly better (creatinine: p = 0.016; FENa: p<0.001) in the NIM-IR group in comparison to the IR group. Serum TNF-alpha and IL-6 levels were significantly lower (TNF-alpha: p = 0.003, IL-6: p = 0.040) as well as W/D ratio and peroxynitrite concentration were significantly lower (p = 0.014; p<0.001) in the NIM-IR group than in the IR group. CONCLUSION: NIM-811 could have the potential of reducing rhabdomyolysis and impairment of the kidney after lower limb IR injury

Computational Insights on the Competing Effects of Nitric Oxide in Regulating Apoptosis

Despite the establishment of the important role of nitric oxide (NO) on apoptosis, a molecular- level understanding of the origin of its dichotomous pro- and anti-apoptotic effects has been elusive. We propose a new mathematical model for simulating the effects of nitric oxide (NO) on apoptosis. The new model integrates mitochondria-dependent apoptotic pathways with NO-related reactions, to gain insights into the regulatory effect of the reactive NO species N2O3, non-heme iron nitrosyl species (FeLnNO), and peroxynitrite (ONOO−). The biochemical pathways of apoptosis coupled with NO-related reactions are described by ordinary differential equations using mass-action kinetics. In the absence of NO, the model predicts either cell survival or apoptosis (a bistable behavior) with shifts in the onset time of apoptotic response depending on the strength of extracellular stimuli. Computations demonstrate that the relative concentrations of anti- and pro-apoptotic reactive NO species, and their interplay with glutathione, determine the net anti- or pro-apoptotic effects at long time points. Interestingly, transient effects on apoptosis are also observed in these simulations, the duration of which may reach up to hours, despite the eventual convergence to an anti-apoptotic state. Our computations point to the importance of precise timing of NO production and external stimulation in determining the eventual pro- or anti-apoptotic role of NO