The SOD1 Inhibitor, LCS-1, Oxidizes H2S to Reactive Sulfur Species, Directly and Indirectly, through Conversion of SOD1 to an Oxidase.

LCS-1, a putative selective inhibitor of SOD1, is a substituted pyridazinone with rudimentary similarity to quinones and naphthoquinones. As quinones catalytically oxidize HS to biologically active reactive sulfur species (RSS), we hypothesized LCS-1 might have similar attributes. Here, we examine LCS-1 reactions with HS and SOD1 using thiol-specific fluorophores, liquid chromatography-mass spectrometry, electron paramagnetic resonance (EPR), UV-vis spectrometry, and oxygen consumption.

Reaction Mechanisms of HS Oxidation by Naphthoquinones.

1,4-naphthoquinones (NQs) catalytically oxidize HS to per- and polysufides and sulfoxides, reduce oxygen to superoxide and hydrogen peroxide, and can form NQ-SH adducts through Michael addition. Here, we measured oxygen consumption and used sulfur-specific fluorophores, liquid chromatography tandem mass spectrometry (LC-MS/MS), and UV-Vis spectrometry to examine HS oxidation by NQs with various substituent groups. In general, the order of HS oxidization was DCNQ ~ juglone > 1,4-NQ > plumbagin >DMNQ ~ 2-MNQ > menadione, although this order varied somewhat depending on the experimental conditions.

Redox and Nucleophilic Reactions of Naphthoquinones with Small Thiols and Their Effects on Oxidization of HS to Inorganic and Organic Hydropolysulfides and Thiosulfate.

Naphthoquinone (1,4-NQ) and its derivatives (NQs, juglone, plumbagin, 2-methoxy-1,4-NQ, and menadione) have a variety of therapeutic applications, many of which are attributed to redox cycling and the production of reactive oxygen species (ROS). We previously demonstrated that NQs also oxidize hydrogen sulfide (HS) to reactive sulfur species (RSS), potentially conveying identical benefits. Here we use RSS-specific fluorophores, mass spectroscopy, EPR and UV-Vis spectrometry, and oxygen-sensitive optodes to examine the effects of thiols and thiol-NQ adducts on HS-NQ reactions.

Naphthoquinones Oxidize HS to Polysulfides and Thiosulfate, Implications for Therapeutic Applications.

1,4-Napththoquinones (NQs) are clinically relevant therapeutics that affect cell function through production of reactive oxygen species (ROS) and formation of adducts with regulatory protein thiols. Reactive sulfur species (RSS) are chemically and biologically similar to ROS and here we examine RSS production by NQ oxidation of hydrogen sulfide (H2S) using RSS-specific fluorophores, liquid chromatography-mass spectrometry, UV-Vis absorption spectrometry, oxygen-sensitive optodes, thiosulfate-specific nanoparticles, HPLC-monobromobimane derivatization, and ion chromatographic assays. We show that NQs, catalytically oxidize H2S to per- and polysulfides (H2Sn, n = 2−6), thiosulfate, sulfite and sulfate in reactions that consume oxygen and are accelerated by superoxide dismutase (SOD) and inhibited by catalase.

The Effects of Antioxidant Nutraceuticals on Cellular Sulfur Metabolism and Signaling.

Nutraceuticals are ingested for health benefits, in addition to their general nutritional value. These dietary supplements have become increasingly popular since the late 20th century and they are a rapidly expanding global industry approaching a half-trillion U.S.

Coenzyme Q and related quinones oxidize HS to polysulfides and thiosulfate.

In the canonical pathway for mitochondrial HS oxidation electrons are transferred from sulfide:quinone oxidoreductase (SQR) to complex III via ubiquinone (CoQ). We previously observed that a number of quinones directly oxidize HS and we hypothesize that CoQ may have similar properties. Here we examine HS oxidation by CoQ and more hydrophilic, truncated forms, CoQ and CoQ, in buffer using HS and polysulfide fluorophores (AzMC and SSP4), silver nanoparticles to measure thiosulfate (HSO), mass spectrometry to identify polysulfides and O-sensitive optodes to measure O consumption.

'Antioxidant' berries, anthocyanins, resveratrol and rosmarinic acid oxidize hydrogen sulfide to polysulfides and thiosulfate: A novel mechanism underlying their biological actions.

Nutraceutical polyphenol catechins in green tea oxidize HS to polysulfides (PS) in buffer and in cells thereby conveying their cytoprotective effects. Here we measure HS oxidation in buffer and HEK293 cells by over-the-counter nutraceuticals, blueberry, bilberry and cranberry, and by polyphenols, cyanadin (Cya), quercetin (Que), rosmarinic acid (RA) and resveratrol (Res). HS and PS were measured with specific fluorophores, AzMc and SSP4 respectively, and thiosulfate (TS) production was measured in buffer using silver nanoparticles (AgNPs).

Oxidation of Hydrogen Sulfide by Quinones: How Polyphenols Initiate Their Cytoprotective Effects.

We have shown that autoxidized polyphenolic nutraceuticals oxidize HS to polysulfides and thiosulfate and this may convey their cytoprotective effects. Polyphenol reactivity is largely attributed to the B ring, which is usually a form of hydroxyquinone (HQ). Here, we examine the effects of HQs on sulfur metabolism using HS- and polysulfide-specific fluorophores (AzMC and SSP4, respectively) and thiosulfate sensitive silver nanoparticles (AgNP).

Manganese Porphyrin-Based SOD Mimetics Produce Polysulfides from Hydrogen Sulfide.

Manganese-centered porphyrins (MnPs), MnTE-2-PyP (MnTE), MnTnHex-2-PyP (MnTnHex), and MnTnBuOE-2-PyP (MnTnBuOE) have received considerable attention because of their ability to serve as superoxide dismutase (SOD) mimetics thereby producing hydrogen peroxide (HO), and oxidants of ascorbate and simple aminothiols or protein thiols. MnTE-2-PyP and MnTnBuOE-2-PyP are now in five Phase II clinical trials warranting further exploration of their rich redox-based biology. Previously, we reported that SOD is also a sulfide oxidase catalyzing the oxidation of hydrogen sulfide (HS) to hydrogen persulfide (HS) and longer-chain polysulfides (HS, = 3-7).

Metabolism of hydrogen sulfide (HS) and Production of Reactive Sulfur Species (RSS) by superoxide dismutase.

Reactive sulfur species (RSS) such as HS, HS, HS, (n = 2-7) and HS are chemically similar to HO and the reactive oxygen species (ROS) HO, HO, O and act on common biological effectors. RSS were present in evolution long before ROS, and because both are metabolized by catalase it has been suggested that "antioxidant" enzymes originally evolved to regulate RSS and may continue to do so today. Here we examined RSS metabolism by Cu/Zn superoxide dismutase (SOD) using amperometric electrodes for dissolved HS, a polysulfide-specific fluorescent probe (SSP4), and mass spectrometry to identify specific polysulfides (HS-HS).

Fluorescence quenching by metal centered porphyrins and poryphyrin enzymes.

Fluorescence spectroscopy and microscopy have been used extensively to monitor biomolecules, especially reactive oxygen species (ROS) and, more recently, reactive sulfide (RSS) species. Nearly all fluorophores are either excited by or emit light between 450 and 550 nm, which is similar to the absorbance of heme proteins and metal-centered porphyrins. Here we examined the effects of catalase (Cat), reduced and oxidized hemoglobin (Hb and metHb), albumin (alb), manganese (III) tetrakis (4-benzoic acid) porphyrin chloride (MnTBAP), iron protoporphyrin IX (hemin), and copper protoporphyrin IX (CuPPIX) on the fluorescence properties of fluorescein.

Catalase as a sulfide-sulfur oxido-reductase: An ancient (and modern?) regulator of reactive sulfur species (RSS).

Catalase is well-known as an antioxidant dismutating HO to O and HO. However, catalases evolved when metabolism was largely sulfur-based, long before O and reactive oxygen species (ROS) became abundant, suggesting catalase metabolizes reactive sulfide species (RSS). Here we examine catalase metabolism of HS, the sulfur analog of HO, hydrogen sulfide (HS) and other sulfur-bearing molecules using HS-specific amperometric electrodes and fluorophores to measure polysulfides (HS; SSP4) and ROS (dichlorofluorescein, DCF).

The Role of Hydrogen Sulfide in Evolution and the Evolution of Hydrogen Sulfide in Metabolism and Signaling.

The chemical versatility of sulfur and its abundance in the prebiotic Earth as reduced sulfide (H2S) implicate this molecule in the origin of life 3.8 billion years ago and also as a major source of energy in the first seven-eighths of evolution. The tremendous increase in ambient oxygen ∼ 600 million years ago brought an end to H2S as an energy source, and H2S-dependent animals either became extinct, retreated to isolated sulfide niches, or adapted.

AAV2/8-humanFOXP3 gene therapy shows robust anti-atherosclerosis efficacy in LDLR-KO mice on high cholesterol diet.

Inflammation is a key etiologic component in atherogenesis. Previously we demonstrated that adeno-associated virus (AAV) 2/8 gene delivery of Netrin1 inhibited atherosclerosis in the low density lipoprotein receptor knockout mice on high-cholesterol diet (LDLR-KO/HCD). One important finding from this study was that FOXP3 was strongly up-regulated in these Netrin1-treated animals, as FOXP3 is an anti-inflammatory gene, being the master transcription factor of regulatory T cells.

Acute thyrotoxic bulbar myopathy with encephalopathic behaviour: an uncommon complication of hyperthyroidism.

Objective. Acute thyrotoxic bulbar palsy is rare, severe, and rapidly progressive. We describe a case of thyrotoxicosis with bulbar palsy, encephalopathy, and pyramidal tract dysfunction.

Brain ablation in the rat cerebral cortex using a tunable-free electron laser.

Background And Objectives: We used the MARK III free electron laser (FEL) tuned to molecular vibrational absorbance maxima in the infrared (IR) wavelength range of 3.0-6.45 microm to study the effect of these various wavelengths and a power level of 5 mJ/2 microseconds macropulse on photoablation of CNS tissue.

Observation of a cw dark-field signal in an absorption spectroscopic experiment using a phase locked free-electron laser.

We introduce a technique for ultrasensitive absorption spectroscopy using the GHz-rate pulse train from a phase-locked free-electron laser (FEL), in which the fractional power absorbed from one or more laser lines reappears as a signal on the dark background between the pulses emerging from the sample. Preliminary absorption experiments in 15 Torr cm of methane at 3.25 &mgr;m, using phase-locked pulses from the Mark III FEL, clearly reveal an interpulse beat signal due to absorption by adjacent molecular rotational lines which is generated only in the presence of interpulse phase coherence.

Laser energy reaching the posterior pole during transscleral cyclophotocoagulation.

Objective: To measure scattered laser energy reaching the posterior pole during transscleral cyclophotocoagulation.

Methods: Transscleral cyclophotocoagulation was performed on 4 cadaver eyes with Nd:YAG noncontact, Nd:YAG contact, and diode contact lasers. Energy was measured with a photodiode through a 7-mm trephined hole in the posterior pole.

Reperfusion injury in ischemic myocardium: effects of nifedipine and verapamil.

Coronary reperfusion following myocardial ischemia may result in further damage to injured myocytes, as judged by their ultrastructural appearance. Calcium entry into myocytes has been implicated in this effect, and calcium channel-blocking agents have been used in attempts to prevent or limit such damage. In this study, we produced myocardial ischemia in pigs by means of reversible coronary artery occlusion.

Microvasculature sparing with controlled reperfusion of ischemic myocardium.

Reperfusion of ischemic myocardium may result in further ultrastructural damage to cardiac fibers, a phenomenon known as reperfusion injury. We have recently shown that controlled reperfusion, with maintenance of reperfusion flow rates near preischemia levels, prevents much of this reperfusion damage. This observation suggests that mechanical damage to the myocardial microvasculature is important in the pathogenesis of reperfusion injury.

Reperfusion injury in ischemic myocardium: protective effect of controlled reperfusion.

Restoration of coronary artery flow following a period of ischemia often results in further ultrastructural damage to cardiac fibers, a phenomenon known as reperfusion injury. We have compared the ultrastructural effects of uncontrolled reperfusion in vivo of ischemic pig myocardium with the ultrastructural effects of reperfusion controlled at flow rates comparable to preischemia levels. Myocardial ischemia was produced for 60 minutes in 9 pigs by means of a reversible coronary artery occlusion, after which coronary artery flow was restored for 120 minutes.

Controlled versus hyperemic flow during reperfusion of jeopardized ischemic myocardium.

Controlled versus uncontrolled reperfusion of ischemic myocardium after experimental coronary artery occlusion was studied to determine the effect on regional ventricular wall motion and associated biochemical alterations. Fourteen pigs underwent coronary artery occlusion for 1 hour followed by 2 hours of reperfusion. In seven animals uncontrolled reperfusion was achieved by complete release of the arterial occlusion resulting in hyperemic flow.

Reperfusion injury in ischemic myocardium: protective effects of ruthenium red and of nitroprusside.

Coronary reperfusion following myocardial ischemia may result in further damage to injured myocytes, as judged by their ultrastructural appearance. Ruthenium red is an inorganic dye with calcium flux-inhibiting properties which protects ischemic myocardium against reperfusion damage, as judged by biochemical indices of mitochondrial function. In this study, we produced myocardial ischemia in pigs by means of reversible coronary artery occlusion.

Evaluation of biochemical functions and ventricular performance in regional ischemic-reperfused myocardium by afterload reduction: differential effects of calcium blocking and non-calcium blocking vasodilators.

The effects of afterload reduction with and without calcium blockade on reperfusion injury were studied in the pig. Reversible occlusion of the left anterior descending coronary artery was performed for 60 minutes followed by 120 minutes of reperfusion. For 15 minutes prior to and throughout reperfusion, treatment was administered with a calcium blocker (nifedipine or verapamil), a metallic organic dye and Ca2+ antagonist (ruthenium red), a vasodilator (nitroprusside), or saline.

The relationship between hypertrophy and dilatation in the postmortem heart.

Confusion may exist at the time of postmortem examination as to whether the diseased heart is dilated, hypertrophied, or both. Ventricular dilatation and ventricular hypertrophy were therefore evaluated by cardiac partition techniques in 441 subjects at autopsy to determine their relationship. Specific weight and surface area of each ventricle were obtained and patients were divided into categories of disease.