Rapid HPLC method reveals dynamic shifts in coenzyme Q redox state.

Ubiquinol or coenzyme Q (CoQ) is a lipid-soluble electron carrier in the respiratory chain and an electron acceptor for various enzymes in metabolic pathways that intersect at this cofactor hub in the mitochondrial inner membrane. The reduced form of CoQ is an antioxidant, which protects against lipid peroxidation. In this study, we have optimized a UV-detected HPLC method for CoQ analysis from biological materials, which involves a rapid single-step extraction into n-propanol followed by direct sample injection onto a column.

Sulfide oxidation promotes hypoxic angiogenesis and neovascularization.

Angiogenic programming in the vascular endothelium is a tightly regulated process for maintaining tissue homeostasis and is activated in tissue injury and the tumor microenvironment. The metabolic basis of how gas signaling molecules regulate angiogenesis is elusive. Here, we report that hypoxic upregulation of ·NO in endothelial cells reprograms the transsulfuration pathway to increase biogenesis of hydrogen sulfide (HS), a proangiogenic metabolite.

Prediction of Oscillations in Glycolysis in Ethanol-Consuming Erythrocyte-Bioreactors.

A mathematical model of energy metabolism in erythrocyte-bioreactors loaded with alcohol dehydrogenase and acetaldehyde dehydrogenase was constructed and analyzed. Such erythrocytes can convert ethanol to acetate using intracellular NAD and can therefore be used to treat alcohol intoxication. Analysis of the model revealed that the rate of ethanol consumption by the erythrocyte-bioreactors increases proportionally to the activity of incorporated ethanol-consuming enzymes until their activity reaches a specific threshold level.

A growth chamber for chronic exposure of mammalian cells to HS.

HS is a redox-active signaling molecule that exerts an array of cellular and physiological effects. While intracellular HS concentrations are estimated to be in the low nanomolar range, intestinal luminal concentrations can be significantly higher due to microbial metabolism. Studies assessing HS effects are typically conducted with a bolus treatment with sulfide salts or slow releasing sulfide donors, which are limited by the volatility of HS, and by potential off-target effects of the donor molecules.

Sulfide oxidation promotes hypoxic angiogenesis and neovascularization.

Unlabelled: Angiogenic programming in the vascular endothelium is a tightly regulated process to maintain tissue homeostasis and is activated in tissue injury and the tumor microenvironment. The metabolic basis of how gas signaling molecules regulate angiogenesis is elusive. Herein, we report that hypoxic upregulation of NO synthesis in endothelial cells reprograms the transsulfuration pathway and increases H S biogenesis.

Redox integration of signaling and metabolism in a head and neck cancer model of radiation resistance using COSM.

Redox metabolism is increasingly investigated in cancer as driving regulator of tumor progression, response to therapies and long-term patients' quality of life. Well-established cancer therapies, such as radiotherapy, either directly impact redox metabolism or have redox-dependent mechanisms of action defining their clinical efficacy. However, the ability to integrate redox information across signaling and metabolic networks to facilitate discovery and broader investigation of redox-regulated pathways in cancer remains a key unmet need limiting the advancement of new cancer therapies.

A redox cycle with complex II prioritizes sulfide quinone oxidoreductase-dependent HS oxidation.

The dual roles of HS as an endogenously synthesized respiratory substrate and as a toxin raise questions as to how it is cleared when the electron transport chain is inhibited. Sulfide quinone oxidoreductase (SQOR) catalyzes the first step in the mitochondrial HS oxidation pathway, using CoQ as an electron acceptor, and connects to the electron transport chain at the level of complex III. We have discovered that at high HS concentrations, which are known to inhibit complex IV, a new redox cycle is established between SQOR and complex II, operating in reverse.

The hepatic compensatory response to elevated systemic sulfide promotes diabetes.

Impaired hepatic glucose and lipid metabolism are hallmarks of type 2 diabetes. Increased sulfide production or sulfide donor compounds may beneficially regulate hepatic metabolism. Disposal of sulfide through the sulfide oxidation pathway (SOP) is critical for maintaining sulfide within a safe physiological range.

Hydrogen sulfide stimulates lipid biogenesis from glutamine that is dependent on the mitochondrial NAD(P)H pool.

Mammalian cells synthesize HS from sulfur-containing amino acids and are also exposed to exogenous sources of this signaling molecule, notably from gut microbes. As an inhibitor of complex IV in the electron transport chain, HS can have a profound impact on metabolism, suggesting the hypothesis that metabolic reprogramming is a primary mechanism by which HS signals. In this study, we report that HS increases lipogenesis in many cell types, using carbon derived from glutamine rather than from glucose.

The mitochondrial NADH pool is involved in hydrogen sulfide signaling and stimulation of aerobic glycolysis.

Hydrogen sulfide is synthesized by enzymes involved in sulfur metabolism and oxidized via a dedicated mitochondrial pathway that intersects with the electron transport chain at the level of complex III. Studies with HS are challenging since it is volatile and also reacts with oxidized thiols in the culture medium, forming sulfane sulfur species. The half-life of exogenously added HS to cultured cells is unknown.

Thioredoxin regulates human mercaptopyruvate sulfurtransferase at physiologically-relevant concentrations.

3-Mercaptopyruvate sulfur transferase (MPST) catalyzes the desulfuration of 3-mercaptopyruvate (3-MP) and transfers sulfane sulfur from an enzyme-bound persulfide intermediate to thiophilic acceptors such as thioredoxin and cysteine. Hydrogen sulfide (HS), a signaling molecule implicated in many physiological processes, can be released from the persulfide product of the MPST reaction. Two splice variants of MPST, differing by 20 amino acids at the N terminus, give rise to the cytosolic MPST1 and mitochondrial MPST2 isoforms.

Hydrogen sulfide perturbs mitochondrial bioenergetics and triggers metabolic reprogramming in colon cells.

Unlike most other tissues, the colon epithelium is exposed to high levels of HS derived from gut microbial metabolism. HS is a signaling molecule that modulates various physiological effects. It is also a respiratory toxin that inhibits complex IV in the electron transfer chain (ETC).

-3-Carboxypropyl-l-cysteine specifically inhibits cystathionine γ-lyase-dependent hydrogen sulfide synthesis.

Hydrogen sulfide (HS) is a gaseous signaling molecule, which modulates a wide range of mammalian physiological processes. Cystathionine γ-lyase (CSE) catalyzes HS synthesis and is a potential target for modulating HS levels under pathophysiological conditions. CSE is inhibited by propargylglycine (PPG), a widely used mechanism-based inhibitor.

Rat liver folate metabolism can provide an independent functioning of associated metabolic pathways.

Folate metabolism in mammalian cells is essential for multiple vital processes, including purine and pyrimidine synthesis, histidine catabolism, methionine recycling, and utilization of formic acid. It remains unknown, however, whether these processes affect each other via folate metabolism or can function independently based on cellular needs. We addressed this question using a quantitative mathematical model of folate metabolism in rat liver cytoplasm.

Cytochrome c Reduction by HS Potentiates Sulfide Signaling.

Hydrogen sulfide (HS) is an endogenously produced gas that is toxic at high concentrations. It is eliminated by a dedicated mitochondrial sulfide oxidation pathway, which connects to the electron transfer chain at the level of complex III. Direct reduction of cytochrome c (Cyt C) by HS has been reported previously but not characterized.

Structural and Mechanistic Insights into Hemoglobin-catalyzed Hydrogen Sulfide Oxidation and the Fate of Polysulfide Products.

Hydrogen sulfide is a cardioprotective signaling molecule but is toxic at elevated concentrations. Red blood cells can synthesize HS but, lacking organelles, cannot dispose of HS via the mitochondrial sulfide oxidation pathway. We have recently shown that at high sulfide concentrations, ferric hemoglobin oxidizes HS to a mixture of thiosulfate and iron-bound polysulfides in which the latter species predominates.

Macrophage-Mediated Clofazimine Sequestration Is Accompanied by a Shift in Host Energy Metabolism.

Prolonged (8 weeks) oral administration of clofazimine results in a profound pharmacodynamic response-bioaccumulation in macrophages (including Kupffer cells) as intracellular crystal-like drug inclusions (CLDIs) with an associated increase in interleukin-1 receptor antagonist production. Notably, CLDI formation in Kupffer cells concomitantly occurs with the formation of macrophage-centric granulomas. Accordingly, we sought to understand the impact of these events on host metabolism using H-nuclear magnetic resonance metabolomics.

Hydrogen Sulfide Oxidation by Myoglobin.

Enzymes in the sulfur network generate the signaling molecule, hydrogen sulfide (H2S), from the amino acids cysteine and homocysteine. Since it is toxic at elevated concentrations, cells are equipped to clear H2S. A canonical sulfide oxidation pathway operates in mitochondria, converting H2S to thiosulfate and sulfate.

Biosynthesis and Reactivity of Cysteine Persulfides in Signaling.

Hydrogen sulfide (H2S) elicits pleiotropic physiological effects ranging from modulation of cardiovascular to CNS functions. A dominant method for transmission of sulfide-based signals is via posttranslational modification of reactive cysteine thiols to persulfides. However, the source of the persulfide donor and whether its relationship to H2S is as a product or precursor is controversial.

H2S analysis in biological samples using gas chromatography with sulfur chemiluminescence detection.

Hydrogen sulfide (H2S) is a metabolite and signaling molecule in biological tissues that regulates many physiological processes. Reliable and sensitive methods for H2S analysis are necessary for a better understanding of H2S biology and for the pharmacological modulation of H2S levels in vivo. In this chapter, we describe the use of gas chromatography coupled to sulfur chemiluminescence detection to measure the rates of H2S production and degradation by tissue homogenates at physiologically relevant concentrations of substrates.

Sulfide oxidation by a noncanonical pathway in red blood cells generates thiosulfate and polysulfides.

A cardioprotectant at low concentrations, H2S is a toxin at high concentrations and inhibits cytochrome c oxidase. A conundrum in H2S homeostasis is its fate in red blood cells (RBCs), which produce H2S but lack the canonical mitochondrial sulfide oxidation pathway for its clearance. The sheer abundance of RBCs in circulation enhances the metabolic significance of their clearance strategy for H2S, necessary to avoid systemic toxicity.

Organization of the human mitochondrial hydrogen sulfide oxidation pathway.

Sulfide oxidation is expected to play an important role in cellular switching between low steady-state intracellular hydrogen sulfide levels and the higher concentrations where the physiological effects are elicited. Yet despite its significance, fundamental questions regarding how the sulfide oxidation pathway is wired remain unanswered, and competing proposals exist that diverge at the very first step catalyzed by sulfide quinone oxidoreductase (SQR). We demonstrate that, in addition to sulfite, glutathione functions as a persulfide acceptor for human SQR and that rhodanese preferentially synthesizes rather than utilizes thiosulfate.

Sulfur as a signaling nutrient through hydrogen sulfide.

Hydrogen sulfide (H₂S) has emerged as an important signaling molecule with beneficial effects on various cellular processes affecting, for example, cardiovascular and neurological functions. The physiological importance of H₂S is motivating efforts to develop strategies for modulating its levels. However, advancement in the field of H₂S-based therapeutics is hampered by fundamental gaps in our knowledge of how H₂S is regulated, its mechanism of action, and its molecular targets.

Bile salts act as effective protein-unfolding agents and instigators of disulfide stress in vivo.

Commensal and pathogenic bacteria must deal with many different stress conditions to survive in and colonize the human gastrointestinal tract. One major challenge that bacteria encounter in the gut is the high concentration of bile salts, which not only aid in food absorption but also act as effective physiological antimicrobials. The mechanism by which bile salts limit bacterial growth is still largely unknown.

Sulfur-based redox alterations in long-lived Snell dwarf mice.

Changes in sulfur-based redox metabolite profiles in multiple tissues of long-lived Snell dwarf mice were compared with age- and sex-matched controls. Plasma methionine and its oxidation products, hypotaurine and taurine, were increased in Snell dwarfs while cystine and glutathione levels were decreased, leading to an oxidative shift in the redox potential. Sexual dimorphism in renal cystathionine β-synthase (CBS) activity was observed in control mice but not in Snell dwarfs.