The Interaction of Myeloperoxidase with the Industrial Contaminant 6-PPD: A Potential Pathway for Reactive Metabolites.

6-PPD (N-[1,3-dimethylbutyl]-N'-phenyl-p-phenylenediamine) is an industrial antioxidant reported to be an environmental contaminant. It was found to be highly toxic to coho salmon and potentially other aquatic organisms. The toxicity of 6-PPD in humans, however, remains unknown. The neutrophil enzyme myeloperoxidase (MPO) is known to catalyze xenobiotic metabolism; therefore, its role in 6-PPD cytotoxicity was investigated using the MPO-rich HL-60 cell line. UV-visible spectroscopy and liquid chromatography-mass spectrometry (LC/MS) were performed to investigate the MPO-mediated oxidation of 6-PPD and identify possible metabolites in the absence and presence of glutathione (GSH). 6-PPD's cytotoxicity, effect on mitochondrial membrane potential (MMP), and GSH-depleting ability in HL-60 cells were assessed. Electron paramagnetic resonance (EPR) was used to determine GSH radical formation using DMPO, and mitochondrial-derived superoxide was assessed with the mito-TEMPO-H probe. Evaluation of the 6-PPD-induced cellular injury pathways was performed by preincubating an antioxidant and an MPO inhibitor with HL-60 cells. UV-vis analysis of MPO-catalyzed oxidation of 6-PPD demonstrated changes in the 6-PPD spectrum, whereas the addition of GSH altered the spectrum, indicating possible GSH conjugate formation. LC/MS showed the formation of multiple products, including GSH-6-PPD conjugates and a GSH conjugate to a 4-hydroxydiphenylamine (a known 6-PPD degradant), which could potentially induce cytotoxicity. 6-PPD demonstrated concentration-dependent cytotoxicity, and cellular GSH levels were decreased by 6-PPD. Similarly, the level of MMP decreased, suggesting mitochondrial depolarization. Furthermore, the EPR spin probe for mitochondrial superoxide showed a positive relationship with 6-PPD concentration, and EPR spin-trapping demonstrated 6-PPD concentration-dependent GSH radical signal intensity using MPO/H2O2. The GSH precursor, NAC, demonstrated partial cytoprotection against 6-PPD; however, the MPO inhibitor PF-1355 surprisingly showed no significant cytoprotective effect. Our results suggest that MPO could be a potential catalyst for 6-PPD toxicity in humans. However, MPO inhibition did not significantly affect cellular viability, suggesting an MPO-independent toxicity pathway. These findings warrant a deeper investigation to determine 6-PPD mammalian toxicity pathways.

Global change impacts on cacti (Cactaceae): current threats, challenges and conservation solutions.

BACKGROUND: The plant family Cactaceae provides some of the most striking examples of adaptive evolution, expressing undeniably the most spectacular New World radiation of succulent plants distributed across arid and semi-arid regions of the Americas. Cacti are widely regarded for their cultural, economic and ecological value, yet they are also recognized as one of the most threatened and endangered taxonomic groups on the planet. SCOPE: This paper reviews current threats to species of cacti that have distributions in arid to semi-arid subtropical regions. Our review focuses primarily on four global change forces: (1) increases in atmospheric CO2 concentrations; (2) increases in mean annual temperatures and heat waves; (3) increases in the duration, frequency and intensity of droughts; and (4) and increases in competition and wildfire frequency from invasion by non-native species. We provide a broad range of potential priorities and solutions for stemming the extinction risk of cacti species and populations. CONCLUSIONS: Mitigating ongoing and emerging threats to cacti will require not only strong policy initiatives and international cooperation, but also new and creative approaches to conservation. These approaches include determining species at risk from climate extremes, enhancing habitat quality after disturbance, approaches and opportunities for ex situ conservation and restoration, and the potential use of forensic tools for identifying plants that have been removed illegally from the wild and sold on open markets.

Neutrophil Myeloperoxidase-Mediated N-Demethylation of Quetiapine Leads to N-Desalkylquetiapine, a Pharmacologically Active Cytochrome P450 Metabolite.

The atypical antipsychotic drugs, quetiapine and clozapine, are associated with idiosyncratic drug reactions (such as agranulocytosis or neutropenia) that are thought to involve reactive metabolites. Neutrophil myeloperoxidase (MPO) metabolism of quetiapine is not well-studied, but is metabolized by cytochrome P450. Based on structural similarity to clozapine, we hypothesized that quetiapine can be metabolized by MPO and that there is overlap between cytochrome P450 and MPO metabolism of quetiapine. The interaction of quetiapine and clozapine with MPO and MPO chlorination activity was studied using UV-vis spectrophotometry. The metabolites were characterized using liquid chromatography-mass spectrometry (LC-MS), and electron paramagnetic resonance (EPR) spectroscopy was used for detecting drug-catalyzed glutathione oxidation. In the presence of quetiapine, MPO compound II accumulated for about 7.5 min, whereas in the presence of clozapine, MPO compound II was not observed as it was rapidly reduced back to the resting state. Increasing quetiapine concentrations resulted in a decrease in MPO chlorination activity, while the opposite result was found in the case of clozapine. UV-vis spectral studies showed no change when quetiapine was oxidized in the absence and presence of chloride anion (Cl-, to catalyze chlorination reactions). Significant changes, however, were observed in the same assay with clozapine, where Cl- appeared to hinder the rate of clozapine metabolism. The MPO-catalyzed hydroxylated and dealkylated metabolites of quetiapine and hydroxylated metabolites of clozapine were observed from the LC-MS analyses, particularly when Cl- was included in the reaction. In addition, hydroxylated, dealkylated, and a proposed sulfoxide metabolite of quetiapine were also observed in the reaction catalyzed by human microsomes/NADPH. Lastly, compared to quetiapine, clozapine metabolism by MPO/H2O2 and glutathione produced more glutathionyl radicals using EPR spin trapping. In conclusion, MPO/H2O2/Cl- was shown to metabolize quetiapine to S-oxidation and P450-like dealkylation products, and quetiapine metabolites were generally less reactive than clozapine.

Impacts of hydraulic redistribution on grass-tree competition vs facilitation in a semi-arid savanna.

A long-standing ambition in ecosystem science has been to understand the relationship between ecosystem community composition, structure and function. Differential water use and hydraulic redistribution have been proposed as one mechanism that might allow for the coexistence of overstory woody plants and understory grasses. Here, we investigated how patterns of hydraulic redistribution influence overstory and understory ecophysiological function and how patterns vary across timescales of an individual precipitation event to an entire growing season. To this end, we linked measures of sap flux within lateral and tap roots, leaf-level photosynthesis, ecosystem-level carbon exchange and soil carbon dioxide efflux with local meteorology data. The hydraulic redistribution regime was characterized predominantly by hydraulic descent relative to hydraulic lift. We found only a competitive interaction between the overstory and understory, regardless of temporal time scale. Overstory trees used nearly all water lifted by the taproot to meet their own transpirational needs. Our work suggests that alleviating water stress is not the reason we find grasses growing in the understory of woody plants; rather, other stresses, such as excessive light and temperature, are being ameliorated. As such, both the two-layer model and stress gradient hypothesis need to be refined to account for this coexistence in drylands.

Comparing the oxidative functions of neutrophil myeloperoxidase and cytochrome P450 enzymes in drug metabolism.

Drug metabolism is an essential process that chemically alters xenobiotic substrates to activate or terminate drug activity. Myeloperoxidase (MPO) is a neutrophil-derived haem-containing enzyme that is involved in killing invading pathogens, although consequentially, this same oxidative activity can produce metabolites that damage host tissue and play a role in various human pathologies. Cytochrome P450s (CYPs) are a superfamily of haem-containing enzymes that are significantly involved in the metabolism of drugs by functioning as monooxygenases and can be induced or inhibited, resulting in significant drug-drug interactions that lead to unanticipated adverse drug reactions. In this review, the functions of drug metabolism of MPO and CYPs are explored, along with their involvement and association for common enzymatic pathways by certain xenobiotics. MPO and CYPs metabolize numerous xenobiotics, although few reported studies have made a direct comparison between both enzymes. Additionally, we employed molecular docking to compare the active site and haem prosthetic group of MPO and CYPs, supporting their similar catalytic activities. Furthermore, we performed LCMS analysis and observed a shared hydroxylated mefenamic acid metabolite produced in both enzymatic systems. A proper understanding of the enzymology and mechanisms of action of MPO and CYPs is of significant importance when enhancing the beneficial functions of drugs in health and diminishing their damaging effects on diseases. Therefore, awareness of drugs and xenobiotic substrates involved in MPO and CYPs metabolism pathways will add to the knowledge base to foresee and prevent potential drug interactions and adverse events.