Coenzyme Q biosynthesis inhibition induces HIF-1α stabilization and metabolic switch toward glycolysis.

Coenzyme Q10 (CoQ, ubiquinone) is a redox-active lipid endogenously synthesized by the cells. The final stage of CoQ biosynthesis is performed at the mitochondrial level by the 'complex Q', where coq2 is responsible for the prenylation of the benzoquinone ring of the molecule. We report that the competitive coq2 inhibitor 4-nitrobenzoate (4-NB) decreased the cellular CoQ content and caused severe impairment of mitochondrial function in the T67 human glioma cell line. In parallel with the reduction in CoQ biosynthesis, the cholesterol level increased, leading to significant perturbation of the plasma membrane physicochemical properties. We show that 4-NB treatment did not significantly affect the cell viability, because of an adaptive metabolic rewiring toward glycolysis. Hypoxia-inducible factor 1α (HIF-1α) stabilization was detected in 4-NB-treated cells, possibly due to the contribution of both reduction in intracellular oxygen tension and ROS overproduction. Exogenous CoQ supplementation partially recovered cholesterol content, HIF-1α degradation, and ROS production, whereas only weakly improved the bioenergetic impairment induced by the CoQ depletion. Our data provide new insights on the effect of CoQ depletion and contribute to shed light on the pathogenic mechanisms of ubiquinone deficiency syndrome.

Characterization of a ß2 adrenergic receptor protein precursor in the European eel (Anguilla anguilla) and its tissue distribution across silvering.

This study provides the characterization and tissue distribution of a ß2-AR in the female European eel during silvering, aiming to better understand the adrenergic system involvement in this critical maturation event. A putative ß2-AR (ADRB2) mRNA was cloned and sequenced. Amino acid residues and motifs important for ligand binding are generally conserved across fish and between fish and mammals, although the occurrence of some sequence variabilities may explain the noted peculiarities of eel AR interaction with pharmacological ligands. The tissue distribution of the ADRB2 gene product was analyzed in five tissues of the eel at different silvering stages and compared with that of the ADRA1 mRNA encoding an α1-AR subtype. On the whole, data suggested that relative ADRA1/ADRB2 tissue expression across silvering is part of the preparatory (molecular) adjustments required to face changes in habitats and migration efforts.

Renewable Alkenes from the Hydrothermal Treatment of Polyhydroxyalkanoates-Containing Sludge.

Polyhydroxyalkanoates (PHA) are a key constituent of excess sludge produced by Aerobic Sewage Sludge Treatment plants. The accumulation of significant amount of PHA inside aerobic microbial cells occurs when a surplus of an easily degradable carbon source (e.g., volatile fatty acids, VFA) is found in combination with other nutrients limitation. Herein, hydrothermal treatment (HT) of PHA-containing sludge at 300 and 375 °C was demonstrated to be effective in converting most (>70% w/w) of the bacterial PHA stored inside microbial cells into alkene/CO2 gas mixtures. Simultaneously, most of non-PHA biomass was converted into water-soluble compounds (50% carbon yield) that were acidogenic fermented to produce volatile fatty acids, ideal substrate to feed aerobic bacteria and produce more PHA. According to results here presented, HT of excess sludge with moderate (13%) PHA content can produce about 50 kg of alkenes per tonne of suspended solids treated, with a significant reduction of sludge mass (80% reduction of wet sludge volume) and consequent disposal cost.

A comprehensive evaluation of the environmental quality of a coastal lagoon (Ravenna, Italy): Integrating chemical and physiological analyses in mussels as a biomonitoring strategy.

This study aimed at evaluating the environmental quality of a coastal lagoon (Pialassa Piomboni, NW-Adriatic, Italy) by combining analyses of biomarkers of environmental stress and bioaccumulation of contaminants in marine mussels (Mytilus galloprovincialis) transplanted for 28days to six selected sites. Assessed biomarkers encompassed lysosomal endpoints, oxidative stress and detoxification parameters, specific responses to metals, neuro- and genotoxic substances; chemical analyses focused on PAHs, metals, pesticide and pharmaceuticals. Results showed up to a 67-fold bioaccumulation of 4- to 6-ring PAHs, including pyrene, fluoranthene, chrysene and benzo(ghi)perylene in transplanted mussels compared to reference conditions (T0). A 10-fold increase of Fe, Cr and Mn was observed, while pesticides and pharmaceuticals were not or slightly detected. The onset of a significant (p<0.05) general stress syndrome occurred in exposed mussels, as outlined by a 50-57.7% decrease in haemocytes lysosomal membrane stability and an increased lysosomal volume (22.6-26.9%) and neutral lipid storage (18.9-48.8%) observed in digestive gland. Data also revealed a diffuse lipofuscin accumulation (86.5-139.3%; p<0.05) in digestive gland, occasionally associated to a catalase activity inhibition in gill, indicating an increased vulnerability toward pro-oxidant factors. Higher levels of primary DNA damage (258%; p<0.05) and PAH accumulation were found in mussels exposed along the eastern shoreline, hosting a petrochemical settlement. Bioaccumulated metals showed a positive correlation with increased metallothionein content (85-208%; p<0.05) observed in mussels from most sites. Overall, the use of physiological and chemical analyses detected chronic alterations of the mussel health status induced by specific toxicological pathways, proving a suitable approach in the framework of biomonitoring programs of coastal lagoons.

Chemical and ecotoxicological properties of three bio-oils from pyrolysis of biomasses.

In view of the potential use of pyrolysis-based technologies, it is crucial to understand the environmental hazards of pyrolysis-derived products, in particular bio-oils. Here, three bio-oils were produced from fast pyrolysis of pine wood and intermediate pyrolysis of corn stalk and poultry litter. They were fully characterized by chemical analysis and tested for their biodegradability and their ecotoxicity on the crustacean Daphnia magna and the green alga Raphidocelis subcapitata. These tests were chosen as required by the European REACH regulation. These three bio-oils were biodegradable, with 40-60% of biodegradation after 28 days, and had EC50 values above 100mgL(-1) for the crustacean and above 10mgL(-1) for the alga, showing low toxicity to the aquatic life. The toxic unit approach was applied to verify whether the observed toxicity could be predicted from the data available for the substances detected in the bio-oils. The predicted values largely underestimated the experimental values.

Use of an integrated biomarker-based strategy to evaluate physiological stress responses induced by environmental concentrations of caffeine in the Mediterranean mussel Mytilus galloprovincialis.

The occurrence of caffeine (CF), a biologically active drug, has widely been documented in coastal waters, and whether its environmental concentrations do represent a threat for marine organisms is unclear. The present study aimed at assessing sub-lethal effects induced by a 7-day exposure to environmentally relevant concentrations of CF (5, 50 and 500ng/L) in the Mediterranean mussel, Mytilus galloprovincialis. CF in water and mussel tissues, and a battery of biomarkers, including lysosomal parameters of general stress, oxidative stress responses and endpoints of neurological and genetic damages, were evaluated and tested for significance vs controls (p<0.05). CF exposure triggered a significant decrease of lysosomal membrane stability in both haemocytes and digestive gland (at 50 and 500ng/L CF) and a significant increase of lysosomal content of neutral lipids (at 500ng/L CF), indicating the onset of a stress syndrome. No effects were noted on lipid peroxidation parameters, such as malondialdehyde and lipofuscin content. The activity of the antioxidant enzymes glutathione S-transferase (GST) and catalase was unmodified in gills, while a significant increase of GST activity was observed in digestive gland (at 5 and 500ng/L CF), suggesting the occurrence of GST-mediated phase II detoxifying processes. CF did not induce geno/neurotoxicity, as shown by the lack of effects on primary DNA damages and acetylcholinesterase activity. In line with its high hydrophilicity, CF did not bioaccumulate in mussel tissues. Data were integrated using the Mussel Expert System, which assigned a low stress level to mussels exposed to 500ng/L CF, whereas no alterations of animal health status were highlighted at lower dosages. This study revealed a low profile of toxicity for environmental concentrations of CF, and confirmed the suitability of an integrated biomarker-based approach to provide a comprehensive picture of the degree of stress induced by emerging contaminants in marine invertebrates.

Selection of best-performing reference gene products for investigating transcriptional regulation across silvering in the European eel (Anguilla anguilla).

The focus of the present study was to set a methodological approach for evaluating molecular mechanisms underlying silvering transformation in the European eel, Anguilla anguilla. Silvering is a tightly controlled process during which eels undergo significant morphological, physiological and behavioral changes, pre-adapting for the oceanic spawning migration. Female eels showing different silver indexes were caught in different seasons in the Comacchio Lagoon (North Adriatic Sea, Italy). Isolated hepatocytes from these eels were selected as the experimental model given the relevant role of these cells in metabolic functions potentially altered during silvering. Expression profiles of 7 candidate reference transcripts were analyzed seeking the most viable and robust strategies for accurate qPCR data normalization during silvering. Stability analysis and further statistical validation identified transcripts encoding the ribosomal proteins L13 and ARP as the appropriate reference genes in studies on A. anguilla through silvering. The identified reference transcripts were further used to evaluate expression profiles of target transcripts encoding the thyroid hormone receptor ß (THRß) and vitellogenin (vtg), known to be involved in silvering processes. To the best of our knowledge, this is the first study comparing THRß expression in European eels across silvering.