Gross and Microscopic Morphological Anatomical Study of the Guinea Pig (Cavia porcellus) and the Capybara (Hydrochoerus hydrochaeris), Aimed at the Preparation of a Comparative Anatomical Atlas of the Different Systems

In recent years, there has been an exponential increase in the so-called “new pets”, including the domestic guinea pig (Cavia porcellus) and the capybara (Hydrochoerus hydrochaeris), two closely related Caviid rodents native to South America. Both historically bred for food purposes, they have more recently become increasingly popular as pets in the European and American continents, respectively. This led to an increasing veterinary interest in deepening the knowledge regarding their normal anatomy, as a basic contribution to other fields of veterinary medicine, including diagnostic imaging, surgery, and pathological anatomy. Being part of a bilateral framework co-tutelage agreement leading to a joint Doctoral Degree between the Alma Mater Studiorum of Bologna, Italy and the Universidad Nacional del Litoral of Santa Fe, Argentina, this research project was partly carried out in Italy (study of guinea pigs) and partly in Argentina (study of capybaras). It consisted in the macroscopic study, through anatomical dissections of carcasses of both species as well as the use of anatomical casts, and in the histological study of the various systems in the two species, and was aimed at creating a gross and microscopic comparative anatomical atlas. From the gross and microscopic morphological and morphometrical anatomical study of the different system of the guinea pig and capybara, several analogies and differences emerged. The creation of a comparative anatomical atlas of gross and microscopic anatomy of the capybara and the guinea pig might prove useful for clinical, zootechnical and research purposes.

The novel hydroxylamine derivative NG-094 suppresses polyglutamine protein toxicity in Caenorhabditis elegans.

Aggregation-prone polyglutamine (polyQ) expansion proteins cause several neurodegenerative disorders, including Huntington disease. The pharmacological activation of cellular stress responses could be a new strategy to combat protein conformational diseases. Hydroxylamine derivatives act as co-inducers of heat-shock proteins (HSPs) and can enhance HSP expression in diseased cells, without significant adverse effects. Here, we used Caenorhabditis elegans expressing polyQ expansions with 35 glutamines fused to the yellow fluorescent protein (Q35-YFP) in body wall muscle cells as a model system to investigate the effects of treatment with a novel hydroxylamine derivative, NG-094, on the progression of polyQ diseases. NG-094 significantly ameliorated polyQ-mediated animal paralysis, reduced the number of Q35-YFP aggregates and delayed polyQ-dependent acceleration of aging. Micromolar concentrations of NG-094 in animal tissues with only marginal effects on the nematode fitness sufficed to confer protection against polyQ proteotoxicity, even when the drug was administered after disease onset. NG-094 did not reduce insulin/insulin-like growth factor 1-like signaling, but conferred cytoprotection by a mechanism involving the heat-shock transcription factor HSF-1 that potentiated the expression of stress-inducible HSPs. NG-094 is thus a promising candidate for tests on mammalian models of polyQ and other protein conformational diseases.

The novel ss469415590 variant predicts virological response to therapy in patients with chronic hepatitis C virus type 1 infection.

BACKGROUND: A novel dinucleotide variant TT/∆G (ss469415590) has been associated with hepatitis C virus clearance. AIM: To assess the role of the ss469415590 variant, compared with the known IL28B polymorphisms (rs8099917, rs12979860 and rs12980275) for predicting virological response to therapy in chronic hepatitis C, and its association with the CXCL10 chemokine serum levels - a surrogate marker of interferon-stimulated genes activation. METHODS: Multivariate analysis of factors predicting rapid and sustained virological response in 280 consecutive, treatment-naïve, nondiabetic, Caucasian patients with chronic hepatitis C treated with peginterferon alpha and ribavirin. RESULTS: In hepatitis C virus genotype 1, the OR (95% CI) for rapid and sustained virological response for the wild-type ss469415590 TT was 9.88 (1.99-48.99) and 7.25 (1.91-27.51), respectively, similar to those found for rs12979860 CC [9.55 (1.93-47.37) and 6.30 (1.71-23.13)] and for rs12980275 AA [9.62 (1.94-47.77] and 7.83 (2.02-30.34)], but higher than for rs8099917 TT [4.8 (1.73-13.33) and 4.75 (2.05-10.98)]. In hepatitis C virus genotype 1, mean (SD) CXCL10 levels in patients with the TT/TT, TT/∆G and ∆G/∆G variants were, respectively, 355.1 (240.6), 434.4 (247.4) and 569.9 (333.3) (P = 0.04). In patients with genotypes 2 and 3 no significant association was found for TT/∆G with viral response. The predictive value of ss469415590 was stronger in patients with advanced fibrosis. CONCLUSIONS: The novel IL28B variants at marker ss469415590 predict response to IFN alpha in chronic hepatitis C patients, especially in those with advanced fibrosis. Their determination may be superior to that of known IL28B variants for patient management using IFN-based regimens.

Saccamoeba lacustris, sp. nov. (Amoebozoa: Lobosea: Hartmannellidae), a new lobose amoeba, parasitized by the novel chlamydia 'Candidatus Metachlamydia lacustris' (Chlamydiae: Parachlamydiaceae).

An amoeba isolated from an aquatic biotope, identified morphologically as Saccamoeba limax, was found harbouring mutualistic rod-shaped gram-negative bacteria. During their cultivation on agar plates, a coinfection also by lysis-inducing chlamydia-like organisms was found in some subpopulations of that amoeba. .Here we provide a molecular-based identification of both the amoeba host and the two bacterial endosymbionts. Analysis of the 18S rRNA gene revealed that this strain is the sister-group to Glaeseria, for which we proposed the name Saccamoeba lacustris. The rod-shaped endosymbiont was identified as a member of Variovorax paradoxus group (Comamonadaceae, Beta-Proteobacteria). No growth on bacteriological agars was recorded, hence this symbiont might be strictly intracellular. The chlamydia-like parasite was unable to infect Acanthamoeba and other amoebae in coculture, showing high host specificity. Phylogenetic analysis based on the 16S rDNA indicated that it is a new member of the family Parachlamydiaceae (order Chlamydiales), for which we proposed the name 'Candidatus Metachlamydia lacustris'.

Probing the interaction between feline immunodeficiency virus and CD134 by using the novel monoclonal antibody 7D6 and the CD134 (Ox40) ligand.

The feline immunodeficiency virus (FIV) targets activated CD4-positive helper T cells preferentially, inducing an AIDS-like immunodeficiency in its natural host species, the domestic cat. The primary receptor for FIV is CD134, a member of the tumor necrosis factor receptor superfamily, and all primary viral strains tested to date use CD134 for infection. We examined the expression of CD134 in the cat using a novel anti-feline CD134 monoclonal antibody (MAb), 7D6, and showed that as in rats and humans, CD134 expression is restricted tightly to CD4+, and not CD8+, T cells, consistent with the selective targeting of these cells by FIV. However, FIV is also macrophage tropic, and in chronic infection the viral tropism broadens to include B cells and CD8+ T cells. Using 7D6, we revealed CD134 expression on a B220-positive (B-cell) population and on cultured macrophages but not peripheral blood monocytes. Moreover, macrophage CD134 expression and FIV infection were enhanced by activation in response to bacterial lipopolysaccharide. Consistent with CD134 expression on human and murine T cells, feline CD134 was abundant on mitogen-stimulated CD4+ T cells, with weaker expression on CD8+ T cells, concordant with the expansion of FIV into CD8+ T cells with progression of the infection. The interaction between FIV and CD134 was probed using MAb 7D6 and soluble CD134 ligand (CD134L), revealing strain-specific differences in sensitivity to both 7D6 and CD134L. Infection with isolates such as PPR and B2542 was inhibited well by both 7D6 and CD134L, suggesting a lower affinity of interaction. In contrast, GL8, CPG, and NCSU were relatively refractory to inhibition by both 7D6 and CD134L and, accordingly, may have a higher-affinity interaction with CD134, permitting infection of cells where CD134 levels are limiting.

The novel proteins Rng8 and Rng9 regulate the myosin-V Myo51 during fission yeast cytokinesis.

The myosin-V family of molecular motors is known to be under sophisticated regulation, but our knowledge of the roles and regulation of myosin-Vs in cytokinesis is limited. Here, we report that the myosin-V Myo51 affects contractile ring assembly and stability during fission yeast cytokinesis, and is regulated by two novel coiled-coil proteins, Rng8 and Rng9. Both rng8Δ and rng9Δ cells display similar defects as myo51Δ in cytokinesis. Rng8 and Rng9 are required for Myo51's localizations to cytoplasmic puncta, actin cables, and the contractile ring. Myo51 puncta contain multiple Myo51 molecules and walk continuously on actin filaments in rng8(+) cells, whereas Myo51 forms speckles containing only one dimer and does not move efficiently on actin tracks in rng8Δ. Consistently, Myo51 transports artificial cargos efficiently in vivo, and this activity is regulated by Rng8. Purified Rng8 and Rng9 form stable higher-order complexes. Collectively, we propose that Rng8 and Rng9 form oligomers and cluster multiple Myo51 dimers to regulate Myo51 localization and functions.

The novel ruthenium-gamma-linolenic complex [Ru(2)(aGLA)(4)Cl] inhibits C6 rat glioma cell proliferation and induces changes in mitochondrial membrane potential, increased reactive oxygen species generation and apoptosis in vitro

The present study reports the synthesis of a novel compound with the formula [Ru(2)(aGLA)(4)Cl] according to elemental analyses data, referred to as Ru(2)GLA. The electronic spectra of Ru(2)GLA is typical of a mixed valent diruthenium(II,III) carboxylate. Ru(2)GLA was synthesized with the aim of combining and possibly improving the anti-tumour properties of the two active components ruthenium and gamma-linolenic acid (GLA). The properties of Ru(2)GLA were tested in C6 rat glioma cells by analysing cell number, viability, lipid droplet formation, apoptosis, cell cycle distribution, mitochondrial membrane potential and reactive oxygen species. Ru(2)GLA inhibited cell proliferation in a time and concentration dependent manner. Nile Red staining suggested that Ru(2)GLA enters the cells and ICP-AES elemental analysis found all increase in ruthenium from <0.02 to 425 mg/Kg in treated cells. The sub-G1 apoptotic cell population was increased by Ru(2)GLA (22 +/- 5.2%) when analysed by FACS and this was confirmed by Hoechst staining of nuclei. Mitochondrial membrane potential was decreased in the presence of Ru(2)GLA (44 +/- 2.3%). In contrast, the cells which maintained a high mitochondrial membrane potential had an increase (18 +/- 1.5%) in reactive oxygen species generation. Both decreased mitochondrial membrane potential and increased reactive oxygen species generation may be involved in triggering apoptosis in Ru(2)GLA exposed cells. The EC(50) for Ru(2)GLA decreased with increasing time of exposure from 285 mu M at 24h, 211 mu M at 48 h to 81 mu M at 72 h. In conclusion, Ru(2)GLA is a novel drug with anti proliferative properties in C6 glioma cells and is a potential candidate for novel therapies in gliomas. Copyright (C) 2009 John Wiley & Sons, Ltd.

Development of a novel microextraction by packed sorbent-based approach followed by ultrahigh pressure liquid chromatography as a powerful technique for quantification phenolic constituents of biological interest in wines

A novel analytical approach, based on a miniaturized extraction technique, the microextraction by packed sorbent (MEPS), followed by ultrahigh pressure liquid chromatography (UHPLC) separation combined with a photodiode array (PDA) detection, has been developed and validated for the quantitative determination of sixteen biologically active phenolic constituents of wine. In addition to performing routine experiments to establish the validity of the assay to internationally accepted criteria (linearity, sensitivity, selectivity, precision, accuracy), experiments are included to assess the effect of the important experimental parameters on the MEPS performance such as the type of sorbent material (C2, C8, C18, SIL, and M1), number of extraction cycles (extract-discard), elution volume, sample volume, and ethanol content, were studied. The optimal conditions of MEPS extraction were obtained using C8 sorbent and small sample volumes (250 μL) in five extraction cycle and in a short time period (about 5 min for the entire sample preparation step). The wine bioactive phenolics were eluted by 250 μL of the mixture containing 95% methanol and 5% water, and the separation was carried out on a HSS T3 analytical column (100 mm × 2.1 mm, 1.8 μm particle size) using a binary mobile phase composed of aqueous 0.1% formic acid (eluent A) and methanol (eluent B) in the gradient elution mode (10 min of total analysis). The method gave satisfactory results in terms of linearity with r2-values > 0.9986 within the established concentration range. The LOD varied from 85 ng mL−1 (ferulic acid) to 0.32 μg mL−1 ((+)-catechin), whereas the LOQ values from 0.028 μg mL−1 (ferulic acid) to 1.08 μg mL−1 ((+)-catechin). Typical recoveries ranged between 81.1 and 99.6% for red wines and between 77.1 and 99.3% for white wines, with relative standard deviations (RSD) no larger than 10%. The extraction yields of the MEPSC8/UHPLC–PDA methodology were found between 78.1 (syringic acid) and 99.6% (o-coumaric acid) for red wines and between 76.2 and 99.1% for white wines. The inter-day precision, expressed as the relative standard deviation (RSD%), varied between 0.2% (p-coumaric and o-coumaric acids) and 7.5% (gentisic acid) while the intra-day precision between 0.2% (o-coumaric and cinnamic acids) and 4.7% (gallic acid and (−)-epicatechin). On the basis of analytical validation, it is shown that the MEPSC8/UHPLC–PDA methodology proves to be an improved, reliable, and ultra-fast approach for wine bioactive phenolics analysis, because of its capability for determining simultaneously in a single chromatographic run several bioactive metabolites with high sensitivity, selectivity and resolving power within only 10 min. Preliminary studies have been carried out on 34 real whole wine samples, in order to assess the performance of the described procedure. The new approach offers decreased sample preparation and analysis time, and moreover is cheaper, more environmentally friendly and easier to perform as compared to traditional methodologies.

MetaPPase by in silico analyses predicting the novel gene operate as H+ pumps.

The lastyears declined the discovery of compounds to use in industrial and naturaldiversity has been the best supplier for novel genes, enzymes and compounds inhigh demand by the biotechnology industry. We know immense diversity of microorganisms,yet most remains unexplored. For these reason we use the metagenômica approach toinvestigate the potential of uncultured microorganisms. With this purpose weused the metagenomic library of from Eucalyptus spp. arboretum (EAA), wedid screening to found positive clone and them was submitted to the process of shotgun,the data obtained was submitted a bioinformatics analyses. Our results showsthe hypothesis of high unexplored microbial diversity of soil are able to foundnovel genes and metagenomic approach is and allowed to isolate novel genes and insilico analyses are essential part to identify a novel Inorganicpyrophosphatase (PPase) prediction indicated the novel gene operate as H+ pumps. Thissuggests that a special feature, our work in situ will be cloning thegene expression vector for subsequent kinetic characterization and crystallization.

Hyperverzweigte Lacton-Copolymere: enzymatische Synthese, Charakterisierung und Darstellung strukturierter Polymernetzwerke = (Hyperbranched lactone copolymers by enzyme catalysis: synthesis, characterization and preparation of structured polymer networks)

Hyperverzweigte Polymere erfuhren in den letzten Jahren immer mehr Beachtung, da sie im Vergleich zu ihren linearen Analoga besondere Eigenschaften besitzen. Im Jahre 2002 wurde die erste enzymkatalysierte Darstellung hyperverzweigter Poly(epsilon-caprolacton)e (hb-PCL) beschrieben. Hier ermöglichte das Konzept der konkurrierenden ringöffnenden Polymerisation und Polykondensation die Kontrolle der Eigenschaften des dargestellten Polymers. Detaillierte Untersuchungen in Hinblick auf Grenzen und Möglichkeiten, aber auch die Synthese im Technikumsmaßstab sind wesentliche Aspekte dieser Arbeit. Außerdem wird ein neues Konzept eingeführt, das Reknitting genannt wurde. Ziel desselben ist das Recycling kommerziellen, linearen PCLs mittels Umesterung zu hb-PCL durch Enzymkatalyse. Diese hb-PCLs zeigen vergleichbare Eigenschaften zu den aus den Comonomeren dargestellten. Ausgehend von hb-PCL sollte eine geeignete Route zu methacrylierten Vernetzerverbindungen entwickelt werden. Aus Mischungen derselben mit 2-Hydroxyethylmethacrylat wurden komplexe Netzwerkarchitekturen durch Copolymerisation erhalten. Diese Netzwerke wurden in Hinblick auf ihre mechanisch physikalischen Eigenschaften untersucht. Zuletzt wurden Screeningexperimente an anderen zyklischen Estern durchgeführt, da ein Transfer des oben vorgestellten Konzepts angestrebt wurde. Zwei neue hyperverzweigte Polymerklassen, hb-Poly(delta-valerolacton) und hb-Polytrimethylencarbonat wurden detaillierter untersucht und in Ihren Eigenschaften mit hb-PCL verglichen.

Preparation of silicon-metal nanocomposites based on electrophoretic deposition

Silicon has long been considered as one of the most promising anode material for lithium-ion batteries. However, the poor cycle life due to stress during charge/discharge cycling has been a major concern for its practical applications. In this report, novel Si-metal nanocomposites have been explored to accommodate the stress generated in the intercalation process. Several approaches have been studied with the aim of getting uniform mixing, good mechanical stability and high Si content. Among the three approaches being investigated, Si- Galinstan nanocomposite based on electrophoretic deposition showed the best promise by achieving at least 32.3% Si theoretical weight percentage, and our in current experiments we’ve already get 13% Silicon weight percentage, which gave us an anode material 46% more capacity than the current commercial product.

Evidence that the novel receptor FGFRL1 signals indirectly via FGFR1

Fibroblast growth factor (FGF) receptor-like protein 1 (FGFRL1) is a recently discovered member of the FGF receptor (FGFR) family. Similar to the classical FGFRs, it contains three extracellular immunoglobulin-like domains and interacts with FGF ligands. However, in contrast to the classical receptors, it does not contain any intracellular tyrosine kinase domain and consequently cannot signal by transphosphorylation. In mouse kidneys, FgfrL1 is expressed primarily at embryonic stages E14-E15 in regions where nascent nephrons develop. In this study, we used whole-mount in situ hybridization to show the spatial pattern of five different Fgfrs in the developing mouse kidney. We compared the expression pattern of FgfrL1 with that of other Fgfrs. The expression pattern of FgfrL1 closely resembled that of Fgfr1, but clearly differed from that of Fgfr2‑Fgfr4. It is therefore conceivable that FgfrL1 signals indirectly via Fgfr1. The mechanisms by which FgfrL1 affects the activity of Fgfr1 remain to be elucidated.

Rapid and simple LC-MS/MS-Screening of 64 novel psychoactive substances using dried blood spots

The range of novel psychoactive substances (NPS) including phenethylamines, cathinones, piperazines, tryptamines, etc. is continuously growing. Therefore, fast and reliable screening methods for these compounds are essential and needed. The use of dried blood spots (DBS) for a fast straightforward approach helps to simplify and shorten sample preparation significantly. DBS were produced from 10 µl of whole blood and extracted offline with 500 µl methanol followed by evaporation and reconstitution in mobile phase. Reversed-phase chromatographic separation and mass spectrometric detection (RP-LC-MS/MS) was achieved within a run time of 10 min. The screening method was validated by evaluating the following parameters: limit of detection (LOD), matrix effect, selectivity and specificity, extraction efficiency, and short-term and long-term stability. Furthermore, the method was applied to authentic samples and results were compared with those obtained with a validated whole blood method used for Routine analysis of NPS. LOD was between 1 and 10 ng/ml. No interference from Matrix compounds was observed. The method was proven to be specific and selective for the analytes, although with limitations for 3-FMC/flephedrone and MDDMA/MDEA. Mean extraction efficiency was 84.6 %. All substances were stable in DBS for at least a week when cooled. Cooling was essential for the stability of cathinones. Prepared samples were stable for at least 3 days. Comparison to the validated whole blood method yielded similar results. DBS were shown to be useful in developing a rapid screening method for NPS with simplified sample preparation. Copyright © 2013 John Wiley & Sons, Ltd