1,727 research outputs found

    Determination of propofol by GC/MS and fast GC/MS-TOF in two cases of poisoning

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    Two cases of suspected acute and lethal intoxication caused by propofol were delivered by the judicial authority to the Department of Sciences for Health Promotion and Mother-Child Care in Palermo, Sicily. In the first case a female nurse was found in a hotel room, where she lived with her mother; four 10 mg/mL vials and two 20 mg/mL vials of propofol were found near the decedent along with syringes and needles. In the second case a male nurse was found in the operating room of a hospital, along with a used syringe. In both cases a preliminary systematic and toxicological analysis indicated the presence of propofol in the blood and urine. As a result, a method for the quantitative determination of propofol in biological fluids was optimized and validated using a liquid-liquid extraction protocol followed by GC/MS and fast GC/MS-TOF. In the first case, the concentration of propofol in blood was determined to be 8.1 \u3bcg/mL while the concentration of propofol in the second case was calculated at 1.2 \u3bcg/mL. Additionally, the tissue distribution of propofol was determined for both cases. Brain and liver concentrations of propofol were, respectively, 31.1 and 52.2 \u3bcg/g in Case 1 and 4.7 and 49.1 \u3bcg/g in Case 2. Data emerging from the autopsy findings, histopathological exams as well as the toxicological results aided in establishing that the deaths were due to poisoning, however, the manner of death in each were different: homicide in Case 1 and suicide in Case 2

    The kinetics of folding of the NSH2 domain from p85

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    SH2 domains are protein domains that mediate protein-protein interaction through the recognition and binding of specific sequences containing phosphorylated tyrosines. The p85 protein is the regulatory subunit of the heterodimeric enzyme PI3K, an important enzyme involved in several molecular pathways. In this work we characterize the folding kinetics of the NSH2 domain of p85. Our data clearly reveal peculiar folding kinetics, characterized by an apparent mismatch between the observed folding and unfolding kinetics. Taking advantage of double mixing stopped flow experiments and site directed mutagenesis we demonstrate that such behavior is due to the cis/trans isomerization of the peptide bond between D73 and P74, being in a cis conformation in the native protein. Our data are discussed in comparison with previous works on the folding of other SH2 domains

    Temperature modulates the response of the thermophilous sea urchin Arbacia lixula early life stages to CO2-driven acidification

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    The increasing abundances of the thermophilous black sea urchin Arbacia lixula in the Mediterranean Sea are attributed to the Western Mediterranean warming. However, few data are available on the potential impact of this warming on A. lixula in combination with other global stressors such as ocean acidification. The aim of this study is to investigate the interactive effects of increased temperature and of decreased pH on fertilization and early development of A. lixula. This was tested using a fully crossed design with four temperatures (20, 24, 26 and 27 C) and two pH levels (pHNBS 8.2 and 7.9). Temperature and pH had no significant effect on fertilization and larval survival (2d) for temperature <27 C. At 27 C, the fertilization success was very low (<1%) and all larvae died within 2d. Both temperature and pH had effects on the developmental dynamics. Temperature appeared to modulate the impact of decreasing pH on the % of larvae reaching the pluteus stage leading to a positive effect (faster growth compared to pH 8.2) of low pH at 20 C, a neutral effect at 24 C and a negative effect (slower growth) at 26 C. These results highlight the importance of considering a range of temperatures covering today and the future environmental variability in any experiment aiming at studying the impact of ocean acidificatio

    The Salt Tolerance Related Protein (STRP) Mediates Cold Stress Responses and Abscisic Acid Signalling in Arabidopsis thaliana

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    Low temperature stress is one of the major causes of crop yield reduction in agriculture. The alteration of gene expression pattern and the accumulation of stress-related proteins are the main strategies activated by plants under this unfavourable condition. Here we characterize the Arabidopsis thaliana Salt Tolerance Related Protein (STRP). The protein rapidly accumulates under cold treatment, and this effect is not dependent on transcriptional activation of the STRP gene, but on the inhibition of proteasome-mediated degradation. Subcellular localization of STRP was determined by the transient expression of STRP-YFP in A. thaliana protoplasts. STRP is localized into the cytosol, nucleus, and associated to the plasma membrane. Under cold stress, the membrane-associated fraction decreases, while in the cytosol and in the nucleus STRP levels strongly increase. STRP has high similarity with WCI16, a wheat Late Embryogenesis Abundant (LEA)-like protein. Despite no canonical LEA motifs in the STRP sequence are present, physicochemical characterization demonstrated that STRP shares common features with LEA proteins, being a high hydrophilic unstructured protein, highly soluble after boiling and with cryoprotectant activity. To clarify the physiological function of STRP, we characterized the phenotype and the response to low temperature stress of the strp knockout mutant. The mutation causes an equal impairment of plant growth and development both in physiological and cold stress conditions. The strp mutant is more susceptible to oxidative damage respect to the wild type, showing increased lipid peroxidation and altered membrane integrity. Furthermore, the analysis of Abscisic acid (ABA) effects on protein levels demonstrated that the hormone induces the increase of STRP levels, an effect in part ascribable to its ability to activate STRP expression. ABA treatments showed that the strp mutant displays an ABA hyposensitive phenotype in terms of seed germination, root development, stomata closure and in the expression of ABA-responsive genes. In conclusion, our results demonstrate that STRP acts as a multifunctional protein in the response mechanisms to low temperature, suggesting a crucial role for this protein in stress perception and in the translation of extracellular stimuli in an intracellular response

    The Surprising Story of Fusicoccin: A Wilt-Inducing Phytotoxin, a Tool in Plant Physiology and a 14-3-3-Targeted Drug

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    Fusicoccin is the alpha glucoside of a carbotricyclic diterpene, produced by the fungus Phomopsis amygdali (previously classified as Fusicoccum amygdali), the causal agent of almond and peach canker disease. A great interest in this molecule started when it was discovered that it brought about an irreversible stomata opening of higher plants, thereby inducing the wilting of their leaves. Since then, several studies were carried out to elucidate its biological activity, biosynthesis, structure, structure-activity relationships and mode of action. After sixty years of research and more than 1800 published articles, FC is still the most studied phytotoxin and one of the few whose mechanism of action has been elucidated in detail. The ability of FC to stimulate several fundamental plant processes depends on its ability to activate the plasma membrane H+-ATPase, induced by eliciting the association of 14-3-3 proteins, a class of regulatory molecules widespread in eukaryotes. This discovery renewed interest in FC and prompted more recent studies aimed to ascertain the ability of the toxin to influence the interaction between 14-3-3 proteins and their numerous client proteins in animals, involved in the regulation of basic cellular processes and in the etiology of different diseases, including cancer. This review covers the different aspects of FC research partially treated in different previous reviews, starting from its discovery in 1964, with the aim to outline the extraordinary pathway which led this very uncommon diterpenoid to evolve from a phytotoxin into a tool in plant physiology and eventually into a 14-3-3-targeted drug

    Aircraft based four-channel thermal dissociation laser induced fluorescence instrument for simultaneous measurements of NO2, total peroxy nitrate, total alkyl nitrate, and HNO3

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    A four-channel thermal dissociation laser induced fluorescence (TD-LIF) instrument has been developed for simultaneous measurements of nitrogen dioxide (NO2), total peroxy nitrate (∑PNs), total alkyl nitrate (∑ANs) and nitric acid (HNO3). NO2 is measured directly by LIF at 532 nm, whereas organic nitrates and nitric acid are thermally dissociated at distinct temperatures in the inlet to form NO2, which is then measured by LIF. The concentrations of each dissociated species are derived by the differences in measured NO2 relative to the reference colder inlet channel. The TD-LIF was adapted to fly on board the UK Facility for Airborne Atmospheric Measurements (FAAM) BAe 146-301 atmospheric research aircraft in summer 2010, and to date has successfully flown in five field campaigns. This paper reports novel improvements in the TD-LIF instrumentations, including (1) the use of a single wavelength laser, which makes the system compact and relatively cheap; (2) the use of a single beam laser that allows easy alignment and optical stability against the vibrational aircraft environment; and (3) the optical assembly of four detection cells that allow simultaneous and fast (time resolution up to 0.1 s) measurements of NO2, ∑PNs, ∑ANs and HNO3. Laboratory-generated mixtures of PNs, ANs and HNO3 in zero air are converted into NO2 and used to fix the dissociation temperatures of each heated inlet to test the selectivity of the instrument and potential interferences due to recombination reactions of the dissociated products. The effectiveness of the TD-LIF was demonstrated during the RONOCO aircraft campaign (summer 2010). A chemiluminescence system that was measuring NO2 and a broadband cavity enhanced absorption spectrometer (BBCEAS) that was measuring one of the PNs (N2O5) were installed on the same aircraft during the campaign. The in-flight intercomparison of the new TD-LIF with the chemiluminescence system for NO2 measurements and the intercomparison between ∑PNs measured by the TD-LIF and N2O5 by the BBCEAS are used to assess the performance of the TD-LIF

    ASTRI SST-2M prototype and mini-array simulation chain, data reduction software, and archive in the framework of the Cherenkov Telescope Array

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    The Cherenkov Telescope Array (CTA) is a worldwide project aimed at building the next-generation ground-based gamma-ray observatory. Within the CTA project, the Italian National Institute for Astrophysics (INAF) is developing an end-to-end prototype of the CTA Small-Size Telescopes with a dual-mirror (SST-2M) Schwarzschild-Couder configuration. The prototype, named ASTRI SST-2M, is located at the INAF "M.C. Fracastoro" observing station in Serra La Nave (Mt. Etna, Sicily) and is currently in the scientific and performance validation phase. A mini-array of (at least) nine ASTRI telescopes has been then proposed to be deployed at the Southern CTA site, by means of a collaborative effort carried out by institutes from Italy, Brazil, and South-Africa. The CTA/ASTRI team is developing an end-to-end software package for the reduction of the raw data acquired with both ASTRI SST-2M prototype and mini-array, with the aim of actively contributing to the global ongoing activities for the official data handling system of the CTA observatory. The group is also undertaking a massive Monte Carlo simulation data production using the detector Monte Carlo software adopted by the CTA consortium. Simulated data are being used to validate the simulation chain and evaluate the ASTRI SST-2M prototype and mini-array performance. Both activities are also carried out in the framework of the European H2020-ASTERICS (Astronomy ESFRI and Research Infrastructure Cluster) project. A data archiving system, for both ASTRI SST-2M prototype and mini-array, has been also developed by the CTA/ASTRI team, as a testbed for the scientific archive of CTA. In this contribution, we present the main components of the ASTRI data handling systems and report the status of their development.Comment: Proceedings of the 35th International Cosmic Ray Conference (ICRC 2017), Bexco, Busan, Korea. All CTA contributions at arXiv:1709.0348

    Specificity of Δ and non-Δ isoforms of Arabidopsis 14-3-3 proteins towards the H+-ATPase and other targets

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    14-3-3 proteins are a family of ubiquitous dimeric proteins that modulate many cellular functions in all eukaryotes by interacting with target proteins. 14-3-3s exist as a number of isoforms that in Arabidopsis identifies two major groups named Δ and non-Δ. Although isoform specificity has been demonstrated in many systems, the molecular basis for the selection of specific sequence contexts has not been fully clarified. In this study we have investigated isoform specificity by measuring the ability of different Arabidopsis 14-3-3 isoforms to activate the H+-ATPase. We observed that GF14 isoforms of the non-Δgroup were more effective than Δ group isoforms in the interaction with the H+-ATPase and in the stimulation of its activity. Kinetic and thermodynamic parameters of the binding of GF14Δ and GF14ω isoforms, representative of Δ and non-Δ groups respectively, with the H+-ATPase, have been determined by Surface Plasmon Resonance analysis demonstrating that the higher affinity of GF14ω is mainly due to slower dissociation. The role of the C-terminal region and of a Gly residue located in the loop 8 and conserved in all non-Δ isoforms has also been studied by deletion and site-specific mutagenesis. The C-erminal domains, despite their high divergence, play an auto-inhibitory role in both isoforms and they, in addition to a specific residue located in the loop 8, contribute to isoform specificity. To investigate the generality of these findings, we have used the SPOT-synthesis technology to array a number of phosphopeptides matching known or predicted 14-3-3 binding sites present in a number of clients. The results of this approach confirmed isoform specificity in the recognition of several target peptides, suggesting that the isoform specificity may have an impact on the modulation of a variety of additional protein activities, as suggested by probing of a phosphopeptide array with members of the two 14-3-3 groups. © 2014 Pallucca et al
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