Electrochemical Investigation of a Microbial Solar Cell Reveals a Nonphotosynthetic Biocathode Catalyst

Microbial solar cells (MSCs) are microbial fuel cells (MFCs) that generate their own oxidant and/or fuel through photosynthetic reactions. Here, we present electrochemical analyses and biofilm 16S rRNA gene profiling of biocathodes of sediment/seawater-based MSCs inoculated from the biocathode of a previously described sediment/seawater-based MSC. Electrochemical analyses indicate that for these second-generation MSC biocathodes, catalytic activity diminishes over time if illumination is provided during growth, whereas it remains relatively stable if growth occurs in the dark. For both illuminated and dark MSC biocathodes, cyclic voltammetry reveals a catalytic-current–potential dependency consistent with heterogeneous electron transfer mediated by an insoluble microbial redox cofactor, which was conserved following enrichment of the dark MSC biocathode using a three-electrode configuration. 16S rRNA gene profiling showed Gammaproteobacteria, most closely related to Marinobacter spp., predominated in the enriched biocathode. The enriched biocathode biofilm is easily cultured on graphite cathodes, forms a multimicrobe-thick biofilm (up to 8.2 μm), and does not lose catalytic activity after exchanges of the reactor medium. Moreover, the consortium can be grown on cathodes with only inorganic carbon provided as the carbon source, which may be exploited for proposed bioelectrochemical systems for electrosynthesis of organic carbon from carbon dioxide. These results support a scheme where two distinct communities of organisms develop within MSC biocathodes: one that is photosynthetically active and one that catalyzes reduction of O₂ by the cathode, where the former partially inhibits the latter. The relationship between the two communities must be further explored to fully realize the potential for MSC applications.     

Application of exogenous substances reduces tobacco-specific nitrosamines content by regulating biosynthesis of nicotine and nitrite in burley tobacco

Tobacco-specific nitrosamines (TSNAs) are carcinogenic chemicals found in tobacco plants. The increasing health consciousness of individuals had led to an increased interest in research on reducing TSNAs content. The aim of this study was to use a pot experiment in which exogenous substances were applied to burley tobacco to dissect the mechanism of TSNAs production. The results indicated that spraying the exogenous substances IAA, NAA, SA and combination thereof on burley tobacco after topping decreased TSNAs content by 2.69–29.4 % in upper leaves and 0.23–39.3 % in middle leaves without affecting total sugar, total nitrogen, potassium and chlorine contents. The application of exogenous substances could down-regulate expression of the NR gene and the activity of the NR enzyme, resulting in less accumulation of the TSNAs precursor nitrite. The exogenous substances significantly reduced nicotine accumulation, which was consistent with low enzyme activities and the down-regulated expressions of genes involved in nicotine biosynthesis, especially significant in the case of quinolinate phosphoribosyltransferase. These results suggested that the application of exogenous substances on burley tobacco after topping could reduce TSNAs content which may be attributed to the regulation of exogenous substances on nitrite and nicotine. This also implies one potential improvement to agronomic practices aimed at controlling the accumulation of TSNAs in burley tobacco.     

Characterization and constitutive expression of a novel endo-1,4-β-d-xylanohydrolase from Aspergillus niger in Pichia pastoris

A putative endo-1,4-β-d-xylanohydrolase gene xyl10 from Aspergillus niger, encoding a 308-residue mature xylanase belonging to glycosyl hydrolase family 10, was constitutively expressed in Pichia pastoris. The recombinant Xyl10 exhibited optimal activity at pH 5.0 and 60 °C with more than 50 % of the maximum activity from 40 to 70 °C. It retained more than 90 % of the original activity after incubation at 60 °C (pH 5.0) for 30 min and more than 74 % after incubation at pH 3.0–13.0 for 2 h (25 °C). The specific activity, K _m and V _max values for purified Xyl10 were, respectively, 3.2 × 10³ U mg^?1, 3.6 mg ml^?1 and 5.4 × 10³ μmol min^?1 mg^?1 towards beechwood xylan. The enzyme degraded xylan to a series of xylooligosaccharides and xylose. The recombinant enzyme with these properties has the potential for various industrial applications.     

Mechanisms for L-channel-mediated increase in [Ca]i and its reduction by anti-bipolar drugs in cultured astrocytes combined with its mRNA expression in freshly isolated cells support the importance of astrocytic L-channels

The importance of Ca²⁺ signaling in astrocytes is undisputed but a potential role of Ca²⁺ influx via L-channels in the brain in vivo is disputed, although expression of these channels in cultured astrocytes is recognized. This study shows that an increase in free cytosolic Ca²⁺ concentration ([Ca²⁺]_i) in astrocytes in primary cultures in response to an increased extracellular K⁺ concentration (45 mM) is inhibited not only by nifedipine (confirming previous observations) but also to a very large extent by ryanodine, inhibiting ryanodine receptor-mediated release of Ca²⁺, known to occur in response to an elevation in [Ca²⁺]_i. This means that the actual influx of Ca²⁺ is modest, which may contribute to the difficulty in demonstrating L-channel-mediated Ca²⁺ currents in astrocytes in intact brain tissue. Chronic treatment with any of the 3 conventional anti-bipolar drugs lithium, carbamazepine or valproic acid similarly causes a pronounced inhibition of K⁺-mediated increase in [Ca²⁺]_i. This is shown to be due to an inhibition of capacitative Ca²⁺ influx, reflected by decreased mRNA and protein expression of the ‘transient receptor potential channel’ (TRPC1), a constituent of store-operated channels (SOCEs). Literature data are cited (i) showing that depolarization-mediated Ca²⁺ influx in response to an elevated extracellular K⁺ concentration is important for generation of Ca²⁺ oscillations and for the stimulatory effect of elevated K⁺ concentrations in intact, non-cultured brain tissue, and (ii) that Ca²⁺ channel activity is dependent upon availability of metabolic substrates, including glycogen. Finally, expression of mRNA for Ca_v1.3 is demonstrated in freshly separated astrocytes from normal brain.     

Zinc Transporter 7 Induced by High Glucose Attenuates Epithelial-to-Mesenchymal Transition of Peritoneal Mesothelial Cells

Zinc (Zn) is an essential micronutrient and cytoprotectant involved in preventing many types of epithelial-to-mesenchymal transition (EMT)-driven fibrosis in vivo. The zinc-transporter family SLC30A (ZnT) is a pivotal factor in the regulation of Zn homeostasis. However, its function in EMT in peritoneal mesothelial cells (PMCs) remains unknown. This study explored the regulation of zinc transporters and the role they play in cell EMT, particularly in rat peritoneal mesothelial cells (RPMCs), surrounding glucose concentrations and the molecular mechanism involved. The effects of high glucose (HG) on zinc transporter gene expression were measured in RPMCs by real-time PCR. We explored ZnT7 (Slc30A7): the effect of ZnT7 over-expression and siRNA-mediated knock-down on HG-induced EMT was investigated as well as the underlying molecular mechanisms. Over-expression of ZnT7 resulted in significantly inhibited HG-induced EMT in RPMCs, while inhibition of ZnT7 expression using a considerable siRNA-mediated knock-down of RPMCs increased the levels of EMT. Furthermore, over-expression of ZnT7 is accompanied by down-regulation of TGF-β/Smad pathway, phospho-Smad3,4 expression levels. The finding suggests that the zinc-transporting system in RPMCs is influenced by the exposure to HG. The ZnT7 may account for the inhibition of HG-induced EMT in RPMCs, likely through targeting TGF-β/Smad signaling.     

An N-terminal Nuclear Export Signal Regulates Trafficking and Aggregation of Huntingtin (Htt) Protein Exon 1

Huntington disease is a dominantly inherited neurodegenerative condition caused by polyglutamine expansion in the N terminus of the huntingtin protein (Htt). The first 17 amino acids (N17) of Htt play a key role in regulating its toxicity and aggregation. Both nuclear export and cytoplasm retention functions have been ascribed to N17. We have determined that N17 acts as a nuclear export sequence (NES) within Htt exon and when fused to yellow fluorescent protein. We have defined amino acids within N17 that constitute the nuclear export sequence (NES). Mutation of any of the conserved residues increases nuclear accumulation of Htt exon 1. Nuclear export of Htt is sensitive to leptomycin B and is reduced by knockdown of exportin 1. In HEK293 cells, NES mutations decrease overall Htt aggregation but increase the fraction of cells with nuclear inclusions. In primary cultured neurons, NES mutations increase nuclear accumulation and increase overall aggregation. This work defines a bona fide nuclear export sequence within N17 and links it to effects on protein aggregation. This may help explain the important role of N17 in controlling Htt toxicity.     

Intratumoral,not circulating,endothelial progenitor cells share genetic aberrations with glial tumor cells

Literature data indicate that glioma stem cells may give rise to both tumor cells and endothelial progenitor cells (EPCs). Malignant glioma patients usually have increased levels of circulating (EPCs) and these cells are known to contribute to the glioma neovasculature. In this study we compared the intratumoral and circulating EPCs of glioma patients for a set of common glioma genotypical aberrations (amplification of EGFR; deletion of PTEN and aneusomy of chromosomes 7 and 10). We found that the EPCs present in the tumor tissues, not the circulating EPCs, share genetic aberrations with the tumor cells. EPCs with EGFR amplification were found in 46% and with PTEN deletion in 36% of the cases. EPCs with polysomy 7 and monosomy 10 were detected in 56% and 38% of the cases while centrosomal abnormalities in EPCs were found in 68% of the cases. The presence of genetic aberrations of glioma cells in intratumoral EPCs may point to transdifferentiation of glioma stem cells into EPCs. However, the tissue specific CD133 splice variant of blood EPCs was detected in the glioma tissues but not in control brains, suggestive of a blood origin of at least part of the intratumoral EPCs. The findings highlight the complexity of the cellular constituents of glioma neovascularization which should be taken into account when developing anti‐angiogenic strategies for gliomas. J. Cell. Physiol. 228: 1383–1390, 2013. © 2012 Wiley Periodicals, Inc.     

Biosynthesis,isolation, and NMR analysis of leukotriene A epoxides: substrate chirality as a determinant of the cis or trans epoxide configuration

Leukotriene (LT)A₄ and closely related allylic epoxides are pivotal intermediates in lipoxygenase (LOX) pathways to bioactive lipid mediators that include the leukotrienes, lipoxins, eoxins, resolvins, and protectins. Although the structure and stereochemistry of the 5-LOX product LTA₄ is established through comparison to synthetic standards, this is the exception, and none of these highly unstable epoxides has been analyzed in detail from enzymatic synthesis. Understanding of the mechanistic basis of the cis or trans epoxide configuration is also limited. To address these issues, we developed methods involving biphasic reaction conditions for the LOX-catalyzed synthesis of LTA epoxides in quantities sufficient for NMR analysis. As proof of concept, human 15-LOX-1 was shown to convert 15S-hydroperoxy-eicosatetraenoic acid (15S-HPETE) to the LTA analog 14S,15S-trans-epoxy-eicosa-5Z,8Z,10E,12E-tetraenoate, confirming the proposed structure of eoxin A₄. Using this methodology we then showed that recombinant Arabidopsis AtLOX1, an arachidonate 5-LOX, converts 5S-HPETE to the trans epoxide LTA₄ and converts 5R-HPETE to the cis epoxide 5-epi-LTA₄, establishing substrate chirality as a determinant of the cis or trans epoxide configuration. The results are reconciled with a mechanism based on a dual role of the LOX nonheme iron in LTA epoxide biosynthesis, providing a rational basis for understanding the stereochemistry of LTA epoxide intermediates in LOX-catalyzed transformations.     

IL‐1β and TNF‐α induce neurotoxicity through glutamate production: a potential role for neuronal glutaminase

Glutaminase 1 is the main enzyme responsible for glutamate production in mammalian cells. The roles of macrophage and microglia glutaminases in brain injury, infection, and inflammation are well documented. However, little is known about the regulation of neuronal glutaminase, despite neurons being a predominant cell type of glutaminase expression. Using primary rat and human neuronal cultures, we confirmed that interleukin‐1β (IL‐1β) and tumor necrosis factor‐α (TNF‐α), two pro‐inflammatory cytokines that are typically elevated in neurodegenerative disease states, induced neuronal death and apoptosis in vitro. Furthermore, both intracellular and extracellular glutamate levels were significantly elevated following IL‐1β and/or TNF‐α treatment. Pre‐treatment with N‐Methyl‐d ‐aspartate (NMDA) receptor antagonist MK‐801 blocked cytokine‐induced glutamate production and alleviated the neurotoxicity, indicating that IL‐1β and/or TNF‐α induce neurotoxicity through glutamate. To determine the potential source of excess glutamate production in the culture during inflammation, we investigated the neuronal glutaminase and found that treatment with IL‐1β or TNF‐α significantly upregulated the kidney‐type glutaminase (KGA), a glutaminase 1 isoform, in primary human neurons. The up‐regulation of neuronal glutaminase was also demonstrated in situ in a murine model of HIV‐1 encephalitis. In addition, IL‐1β or TNF‐α treatment increased the levels of KGA in cytosol and TNF‐α specifically increased KGA levels in the extracellular fluid, away from its main residence in mitochondria. Together, these findings support neuronal glutaminase as a potential component of neurotoxicity during inflammation and that modulation of glutaminase may provide therapeutic avenues for neurodegenerative diseases.