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Hypothalamic gonadotropin-releasing hormone (GnRH) neurons regulate fertility and integrate hormonal status with environmental cues to ensure reproductive success. Here we show that GnRH neurons in the olfactory bulb (GnRHOB) of adult mice can mediate social recognition. Specifically, we show that GnRHOBneurons extend neurites into the vomeronasal organ and olfactory epithelium and project to the median eminence. GnRHOBneurons in males express vomeronasal and olfactory receptors, are activated by female odors and mediate gonadotropin release in response to female urine. Male preference for female odors required the presence and activation of GnRHOBneurons, was impaired after genetic inhibition or ablation of these cells and relied on GnRH signaling in the posterodorsal medial amygdala. GnRH receptor expression in amygdala kisspeptin neurons appear to be required for GnRHOBneurons’ actions on male mounting behavior. Taken together, these results establish GnRHOBneurons as regulating fertility, sex recognition and mating in male mice.

We report here the first fully automated capillary electrophoresis (CE) system that can be operated underwater. The system performs sample acquisition and analysis by coupling CE to contactless conductivity detection. Using 5 M acetic acid as the background electrolyte (BGE), inorganic cations and amino acids at concentrations as low as 5.2 μM can be separated and identified. This technology could be augmented to include a variety of other detection modes. This system serves as an early prototype for potential future underwater explorers on ocean worlds of the outer solar system such as Europa or Enceladus. This work documents the first step in the development of this general-purpose technology platform.

The present study investigates the effect of different reaction times on the crystallinity, surface morphology and size of iron oxide nanoparticles. In this synthetic system, aqueous iron (III) nitrate (Fe (NO3)3·9H2O) nonahydrate, provided the iron source and triethylamine was the precipitant and alkaline agent. The as-synthesised iron oxide nanoparticles were characterised by X-ray diffraction (XRD), Rietveld analysis, Scanning Electron Microscopy (SEM) and Fourier transform infrared spectroscopy (FTIR). Prolonged reaction times indicated the change on nanoparticle shape from elongated nanorods to finally distorted nanocubes. Analysis on the crystallinity of the iron oxide nanoparticles suggest that the samples mainly consist of two phases, which are Goethite (α-FeOOH) and Hematite (α-Fe2O3) respectively.

The in situ search for chemical signatures of life on extraterrestrial worlds requires automated hardware capable of performing detailed compositional analysis during robotic missions of exploration. The use of electrophoretic separations in this search is particularly powerful, enabling analysis of a wide range of soluble organic compounds potentially indicative of life, as well as inorganic compounds that can serve as indicators of habitability. However, to detect this broad range of compounds with a single electrophoresis instrument, a combination of different detection modes is required. For detection of any ionizable species, including organic compounds that do not participate in terrestrial biology (i.e., “unknown unknowns”), mass spectrometry (MS) is essential. Inorganic ions, or any dissolved charged species present, can be analyzed using capacitively coupled contactless conductivity detection (C4D). Additionally, for the trace analysis of compounds of key interest to astrobiology (particularly, amino acids), laser-induced fluorescence (LIF) detection holds unique promise, due to the fact that it has the highest demonstrated sensitivity of any form of detection. Here, we demonstrate a fully automated, portable capillary electrophoresis analyzer that is capable of all these modes of detection. The prototype system developed here addresses the three most significant challenges for doing electrophoretic separations: precise sample injection, HV isolation, and automation of all operational steps. These key challenges were successfully addressed with the use of custom-designed rotor-stator valves with optimized operational sequences incorporating gas purging steps, rinses, and HV application.

Objective: Atorvastatin (ATV) is effective in reducing total cholesterol and low-density lipoprotein levels. Furthermore, it produces pleiotropic effects in neurodegenerative conditions such as Parkinson's, Alzheimer's, and epilepsy. However, due to the effective defense system of the central nervous system (CNS), the development of new medicines for clinical conditions has proven difficult. In this context, nanotechnology was applied as a promising solution to promote drug vectorization to the brain. Methods: The solvent emulsification-diffusion method was used to develop nanoparticles (NPs) based on polylactic acid and coated with polysorbate 80 containing ATV. Quality-by-Design (QbD) was used in the optimization of nanoparticles production through the application of the experimental design Box-Behnken Design. Results: After optimizing the independent factors including sonication time, surfactant concentration and surfactant volume, the NPs presented physicochemical characteristics such as entrapment efficiency of 86.4 ± 2.4%, mean size of 225.2 ± 4.8 nm, and zeta potential of -14.4 ± 0.36 mV. In the in vitro release study, approximately 20% of the encapsulated ATV was released. Conclusion: The application of QbD was very useful in demonstrating its applicability in the nanotechnological pharmaceutical area for controlling and predicting the influence of the variables in the production of NPs. The NPs developed in this study presented adequate physicochemical characteristics, which is promising for future in vivo studies. The physicochemical characteristics included entrapment efficiency of 86.4 ± 2.4%, mean size of 225.2 ± 4.8 nm, and zeta potential of -14.4 ± 0.36 mV. In the in vitro release study, approximately 20% of the encapsulated ATV was released. The application of QbD was very useful in demonstrating its applicability in the nanotechnological pharmaceutical area for controlling and predicting the influence of the variables in the production of NPs. The NPs developed in this study presented adequate physicochemical characteristics, which is promising for future in vivo studies.

The discovery of isozyme-selective histone deacetylase (HDAC) inhibitors is critical for understanding the biological functions of individual HDACs and for validating HDACs as drug targets. The isozyme HDAC10 contributes to chemotherapy resistance and has recently been described to be a polyamine deacetylase, but no studies toward selective HDAC10 inhibitors have been published. Using two complementary assays, we found Tubastatin A, an HDAC6 inhibitor, to potently bind HDAC10. We synthesized Tubastatin A derivatives and found that a basic amine in the cap group was required for strong HDAC10 binding. HDAC10 inhibitors mimicked knockdown by causing dose-dependent accumulation of acidic vesicles in a neuroblastoma cell line. Furthermore, docking into human HDAC10 homology models indicated that a hydrogen bond between a cap group nitrogen and the gatekeeper residue Glu272 was responsible for potent HDAC10 binding. Taken together, our data provide an optimal platform for the development of HDAC10-selective inhibitors, as exemplified with the Tubastatin A scaffold.

The identification of possible targets for a known bioactive compound is of the utmost importance for drug design and development. Molecular docking is one possible approach for in-silicoprotein target prediction, whereas a molecule is docked into several different protein structures to identify potential targets. This reverse docking approach is hampered by the limitation of current scoring functions to correctly discriminate between targets and nontargets. In this work, a development of target-specific scoring functions is described that showed improved prediction performances for the correct target prediction of both actives and decoys on three validation data sets. In contrast to pure ligand-based approaches, that are in general faster and include a greater target space, docking-based approaches can cover also unknown chemical space that lies outside the known bioactivity data. These target-specific scoring functions are based on known bioactivity data retrieved from ChEMBL and supervised machine learning approaches. Neural Networks and Support Vector Machines (SVMs) models were trained for 20 different protein targets. Our protein–ligand interaction fingerprint PADIF (Protein Atom Score Contributions Derived Interaction Fingerprint) represents the input for training, whereas the PADIFs are calculated based on docking poses of active and inactive compounds. Different data sets of previously unseen molecules were used for the final evaluation and analysis of the prediction performance of the created models. For a single-target selectivity data set, the correct target model returns in most of the cases the highest probabilities scores for their active molecules and with statistically significant differences from the other targets. These probability scores were also predicted and successfully used to rank the targets for molecules of a multitarget data set with activity data described simultaneously for two, three, and four to seven protein targets.

The present study deals with a comparative evaluation of a single-zone (SZ) thermodynamic model and a 3D computational fluid dynamics (CFD) model for heat release calculation in internal combustion engines. The first law, SZ, model is based on the first law of thermodynamics. This model is characterized by a very simplified modeling of the combustion phenomenon allowing for a great simplicity in the mathematical formulation and very low computational time. The CFD 3D models, instead, are able to solve the chemistry of the combustion process, the interaction between turbulence and flame propagation, the heat exchange with walls and the dissociation and re-association of chemical species. They provide a high spatial resolution of the combustion chamber as well. Nevertheless, the computation requirements of CFD models are enormously larger than the SZ techniques. However, the SZ model needs accurate experimental in-cylinder pressure data for initializing the heat release calculation. Therefore, the main objective of an SZ model is to evaluate the heat release, which is very difficult to measure in experiments, starting from the knowledge of the in-cylinder pressure data. Nevertheless, the great simplicity of the SZ numerical formulation has a margin of uncertainty which cannot be known a priori. The objective of this paper was, therefore, to evaluate the level of accuracy and reliability of the SZ model comparing the results with those obtained with a CFD 3D model. The CFD model was developed and validated using cooperative fuel research (CFR) engine experimental in-cylinder pressure data. The CFR engine was fueled with 2,2,4-trimethylpentane, at a rotational speed of 600 r/min, an equivalence ratio equal to 1 and a volumetric compression ratio of 5.8. The analysis demonstrates that, considering the simplicity and speed of the SZ model, the heat release calculation is sufficiently accurate and thus can be used for a first investigation of the combustion process.

The aerodynamic characterization of airfoils is of crucial importance for the design and optimization of wind turbines. The present paper tries to provide an engineering methodology for the improvement of the accuracy and reliability of 2D airfoil computational fluid dynamics models, by coupling the ANSYS Fluent solver and a Micro-Genetic Algorithm. The modeling strategy provided includes meshing optimization, solver settings, comparison between different turbulence models and, mainly, the calibration of the local correlation parameters of the transition turbulence model by Menter, which was found to be the most accurate model for the simulation of transitional flows. Specifically, the Micro-Genetic Algorithm works by generating populations of the missing local correlation parameters. In doing so, it is possible to search for the minimization of the error in lift calculations. For each specific Reynolds number, the calibration was carried out only at the Angle of Attack where the lift drop occurs and the airfoil completely stalls. This new idea allowed for a relatively rapid and good calibration as demonstrated by the experimental–numerical comparisons presented in this paper. Only the experimental stall angle and the relative lift coefficient were, therefore, necessary for obtaining a good calibration. The calibration was made using the widely known S809 profile data. The correlation parameters, obtained as so, were subsequently used for testing on the NACA 0018 airfoil with satisfactory results. Therefore, the calibration obtained using the S809 airfoil data appeared to be reliable and may be used for the simulation of other airfoils. This can be done without the need for further wind tunnel experimental data or recalibrations. The proposed methodology will, therefore, be of essential help in obtaining accurate aerodynamic coefficients data. This will drastically improve the capabilities of the 1D design codes at low Reynolds numbers thanks to the possibility of generating accurate databases of 2D airfoil aerodynamic coefficients. The advantages of the proposed calibration will be helpful in the generation of more accurate 3D wind turbine models as well. The final objective of the paper was thus to obtain a fine and reliable calibration of the transition turbulence model by Menter. This was specifically made for an accurate prediction of the aerodynamic coefficients of any airfoil at low Reynolds numbers and for the improvements of 3D rotor models.

Two-dimensional (2D) crystals with C2N stoichiometry have recently been synthesized. This novel material, dubbed nitrogenated holey graphene (NHG), is a semiconductor unlike pristine graphene. For any novel material, it is fundamental to understand the behaviors of different adatoms on its surface, a process responsible for a rich phenomenology. In this work, we employed first-principles calculations and a hybrid quantum mechanics/molecular mechanics method to investigate the adsorption of H, B, and O on NHG sheets. The adsorption of H atoms could prove important for applications in hydrogen storage and gas sensors, whereas the adsorption of O in any new material is important to understand its oxidation process. Both N and B are common dopants in carbon-based systems, such as in BNC structures. We found that H and B prefer to adsorb on top of a nitrogen atom, whereas O prefers to adsorb on top of a carbon–carbon bond. The electronic structure of NHG also changes as a result of the presence of adatoms, with the appearance of midgap states close to the Fermi level. In the case of NHG + H and NHG + B, we observed the appearance of a finite magnetic moment, related to the midgap states, which could give rise to a magnetoresistance effect. Our results provide insight into the adsorption of impurities on this novel 2D carbon-based material, with potential for applications in novel electronic devices.

In this paper the authors describe a design and optimization process of micro HAWTs using a numerical and experimental approach. An in-house 1D BEM model was used to obtain a first geometrical draft. It allowed to quickly optimize blade geometry to maximize energy production as well. As these models are quite sensitive to airfoil coefficients, above all at low Reynolds numbers, an accurate 3D CFD model was developed to support and validate the 1D BEM design, analyzing and fixing the discrepancies between model output. The 3D CFD model was developed and optimized using ANSYS Fluent solver and a RANS transition turbulence model. This allowed to correctly reproduce the transition and stall phenomena that characterize the aerodynamic behavior of micro wind turbines, solving the issues related to low Reynolds flows. The procedure was completed, thus building two micro HAWTs with different scales, testing them in the subsonic wind tunnel of the University of Catania. Wind tunnel features, experimental set-up and testing procedures are presented in the paper. Through the comparison of numerical CFD and experimental test results, a good compatibility was found. This allowed the authors to analyze and compare numerical calculation results and verify blockage effects on the prototypes as well.

Networks of silver nanowires appear set to replace expensive indium tin oxide as the transparent conducting electrode material in next generation devices. The success of this approach depends on optimizing the material conductivity, which until now has largely focused on minimizing the junction resistance between wires. However, there have been no detailed reports on what the junction resistance is, nor is there a known benchmark for the minimum attainable sheet resistance of an optimized network. In this paper, we present junction resistance measurements of individual silver nanowire junctions, producing for the first time a distribution of junction resistance values and conclusively demonstrating that the junction contribution to the overall resistance can be reduced beyond that of the wires through standard processing techniques. We find that this distribution shows the presence of a small percentage (6%) of high-resistance junctions, and we show how these may impact the performance of network-based materials. Finally, through combining experiment with a rigorous model, we demonstrate the important role played by the network skeleton and the specific connectivity of the network in determining network performance.

Networks comprised of randomly oriented overlapping nanowires offer the possibility of simple fabrication on a variety of substrates, in contrast with the precise placement required for devices with single or aligned nanowires. Metal nanowires typically have a coating of surfactant or oxide that prevents aggregation, but also prevents electrical connection. Prohibitively high voltages can be required to electrically activate nanowire networks, and even after activation many nanowire junctions remain nonconducting. Nonelectrical activation methods can enhance conductivity but destroy the memristive behavior of the junctions that comprise the network. We show through both simulation and experiment that electrical stimulation, microstructured electrode geometry, and feature scaling can all be used to manipulate the connectivity and thus electrical conductivity of networks of silver nanowires with a nonconducting polymer coating. More generally, these results describe a strategy to integrate nanomaterials into controllable, adaptive macroscale materials.

The aim of the study was to test the clinical efficacy and effect on colonic transit time (CTT) of a dietary fiber mixture given to children with controlled chronic constipation (CC) after the withdrawal of stool softeners and enemas.

The aim of the study was to test the clinical efficacy and effect on colonic transit time (CTT) of a dietary fiber mixture given to children with controlled chronic constipation (CC) after the withdrawal of stool softeners and enemas. This randomized, placebo-controlled, double-blind clinical trial involved 54 patients aged 4 to 12 years and had CC that was controlled by the use of low-dose stool softeners. The use of these softeners was discontinued when the patients were admitted to the clinical trial. The patients were randomized into 2 groups for the 4-week study period. One group received a dietary fiber mixture and the other group received a placebo (maltodextrin). The primary outcome was therapeutic failure (oral stool softeners or enemas was required to prescribe during the trial). Secondary outcomes included defecation frequency, stool consistency (measured using the Bristol Stool Form Scale), and CTT. Therapeutic failure was observed in 34.6% (9/26) of the patients in the dietary fiber mixture group and in 35.7% (10/28) in the control group (P= 0.933). The mean increase in daily bowel movements was 0.53 in the dietary fiber mixture group and 0.23 in the control group (P= 0.014). The patients in the dietary fiber mixture group (60.0%) passed nonhardened stools more frequently than did those in the control group (16.7%, P= 0.003). The CTT was similar for both groups. The fiber mixture did not prevent the suspension of stool softeners or lead to reduced CTT; however, the mixture promoted an increased frequency of defecation and an improvement in the stool consistency.

ABSTRACTSequestration of Plasmodium falciparum-infected erythrocytes (Pf-iEs) in the microvasculature of vital organs plays a key role in the pathogenesis of life-threatening malaria complications, such as cerebral malaria and malaria in pregnancy. This phenomenon is marked by the cytoadhesion of Pf-iEs to host receptors on the surfaces of endothelial cells, on noninfected erythrocytes, and in the placental trophoblast; therefore, these sites are potential targets for antiadhesion therapies. In this context, glycosaminoglycans (GAGs), including heparin, have shown the ability to inhibit Pf-iE cytoadherence and growth. Nevertheless, the use of heparin was discontinued due to serious side effects, such as bleeding. Other GAG-based therapies were hampered due to the potential risk of contamination with prions and viruses, as some GAGs are isolated from mammals. In this context, we investigated the effects and mechanism of action of fucosylated chondroitin sulfate (FucCS), a unique and highly sulfated GAG isolated from the sea cucumber, with respect to P. falciparumcytoadhesion and development. FucCS was effective in inhibiting the cytoadherence of Pf-iEs to human lung endothelial cells and placenta cryosections under static and flow conditions. Removal of the sulfated fucose branches of the FucCS structure virtually abolished the inhibitory effects of FucCS. Importantly, FucCS rapidly disrupted rosettes at high levels, and it was also able to block parasite development by interfering with merozoite invasion. Collectively, these findings highlight the potential of FucCS as a candidate for adjunct therapy against severe malaria.

Copyright of American Indian Rock Art is the property of American Rock Art Research Association and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

Copyright of American Indian Rock Art is the property of American Rock Art Research Association and its content may not be copied or emailed to multiple sites or posted to a listserv without the copyright holder's express written permission. However, users may print, download, or email articles for individual use. This abstract may be abridged. No warranty is given about the accuracy of the copy. Users should refer to the original published version of the material for the full abstract. (Copyright applies to all Abstracts.)

AbstractWe measured the 41K thermal neutron absorption and resonance integral cross sections after the irradiation of KNO3samples near the core of the IEA-R1 IPEN pool-type research reactor. Bare and cadmium-covered targets were irradiated in pairs with Au-Al alloy flux-monitors. The residual activities were measured by gamma-ray spectroscopy with a HPGe detector, with special care to avoid the 42K decay ß-emission effects on the spectra. The gamma-ray self-absorption was corrected with the help of MCNP simulations. We applied the Westcott formalism in the average neutron flux determination and calculated the depression coefficients for thermal and epithermal neutrons due to the sample thickness with analytical approximations. We obtained 1.57(4) b and 1.02(4) b, for thermal and resonance integral cross sections, respectively, with correlation coefficient equal to 0.39.

Propulsion system can experience large power absorption fluctuations during tight maneuvers. In the case of a turning circle maneuver for a twin-screw ship, the power required by the two shaft lines can be completely different; in case of non conventional propulsion system, like cross-connect configurations, a compromise must be met in order to design a safe control system without affecting dramatically the vessel's maneuvering performance. In this work, a series of free running model tests have been carried out in order to investigate the influence of different propulsion system operation settings on the vessel's maneuvering characteristics.

Blockade of the inducible costimulator (ICOS) pathway has been shown to prolong allograft survival; however, its utility in xenotransplantation is unknown. We hypothesize that local expression of ICOS-Ig by the xenograft will suppress the T-cell response resulting in significant prolonged graft survival.

Long-term acceptance of transplanted organs without requirement for indefinite immunosuppression remains the ultimate goal of transplant clinicians and scientists. This clinical state of allograft acceptance termed “operational tolerance” has been elusive in routine practice. However, there are published reports of recipients where immunosuppression has been discontinued, by intention or patient noncompliance, in which the outcome is a nondestructive immune response and normal function. The question now arises how clinical operational tolerance might be achieved in the majority of recipients. This review provides an overview of current approaches to achieve operational tolerance, including the use of donor bone marrow and depletion of recipient T cells and the resistance of liver transplants to rejection. It also describes the key role of clinical immune monitoring and future approaches to tolerance induction including inhibition of T-cell signaling, manipulation of costimulatory pathways, and expansion of regulatory T cells. The principles of these experimental approaches may ultimately be extended to provide safe and effective control of transplant rejection and induction of clinical operational tolerance.

Introduction:The existence of specific carbohydrates on the surface of a wide range of cells provides the opportunity for the development of highly targeted therapeutic agents. The potential applications of such agents are diverse, and include vaccines against pathogenic microorganisms, cancer and HIV, and anti-rejection agents for organ transplantation. However, the use of carbohydrates as either therapeutic agents or immunogens is frequently problematic, as they are often rapidly metabolized and poorly immunogenic. Therefore, the search for carbohydrate-mimetic agents is of considerable therapeutic value, for the potential of such agents to both interfere with carbohydrate–protein interactions and to generate carbohydrate-specific immune responses.Areas covered:The review discusses recent examples of carbohydrate-mimetic peptides with regard to the structural and functional aspects of mimicry and the implications of peptide mimicry for application in therapeutics. The reader will gain knowledge of the various mechanisms of peptide carbohydrate mimicry, and the potential importance of these mechanisms in targeted therapeutic design.Expert opinion:Peptide carbohydrate mimicry is manifested by distinct mechanisms, any one of which may be relevant to specific protein targets. As structural information becomes available for a wider variety of systems, the questions about mimicry will be more effectively addressed.

Earthquakes have been found to be associated with increased prevalence of psychiatric disorders: Post-traumatic Stress Disorder (PTSD) with a diagnosis range of 1.5%-74%, depression, anxiety and sleep disorders and substance abuse. Risk factors are varied: exposure to the earthquake, closeness to the epicenter, disruption of social network, financial loss, female sex, low educational level, etc. PTSD diagnosis is difficult because people unconsciously neglect traumatic history and ignore consequences. Between April 6th2009 and September 2009, 323 young survivors aged between 18 and 30 years were screened for PTSD symptoms at the S.M.I.L.E., a psychiatric service for young people at the L'Aquila Camp Hospital. The screening assessment consisted of: a socio-demographic schedule with questions about earthquake experience, the General Health Questionnaire-12 items (GHQ-12), Impact Event Scale-Revised (IES-R) and Semi-structured Clinical Interview Diagnosis II (SCID-II). Regarding psychiatric morbidity, 44.2% and 37.4% had respectively moderate and high stress levels. Female gender and unemployment were significantly correlated (p<.001) with stress level and Obsessive-Compulsive (OC) trait. After screening assessment, the 66.7% (N=215) of total sample showed a positive post-traumatic symptomatology with 13.8% of PTSD diagnosis. Obsessive-Compulsive trait, female gender, destruction of housing and high level of stress (GHQ = 20) were significant predictors for a PTSD diagnosis when compared with subjects positive for PTSD symptomatology but without a PTSD diagnosis according to DSM-IV criteria. Personality features must be considered important risks for post-traumatic consequences: OC trait was significantly associated with a high score on the GHQ-12 (=20) and is a significant predictor of PTSD symptoms (“re-experiencing” dimension). Houses destroyed, gender and unemployed were also important risk factors. Our study confirms that a natural disaster produces high levels of mental disorders with significant long-term risk of chronic impairment, such as the development of a PTSD diagnosis (13.8%) with enormously costly consequences for the mental health care system and society. This should be a high priority in a public health program.

Carbohydrate–antibody interactions mediate many cellular processes and immune responses. Carbohydrates expressed on the surface of cells serve as recognition elements for particular cell types, for example, in the ABO(H) blood group system. Antibodies that recognize host-incompatible ABO(H) system antigens exist in the bloodstream of all individuals (except AB individuals), preventing bloodtransfusion and organ transplantation between incompatible donors and recipients. A similar barrier exists for cross-species transplantation (xenotransplantation), in particular for pig-to-human transplantation. All humans express antibodies against the major carbohydrate xenoantigen, Gala(1,3)Gal (aGal), preventing successful xenotransplantation. Although antibody binding sites are precisely organized so as to selectively bind a specific antigen, many antibodies recognize molecules other than their native antigen. A range of peptides have been identified that can mimic carbohydrates and inhibit anti-aGal antibodies. However, the structural basis of how the peptides achieved this was not known. Previously, we developed an in silico method which we used to investigate carbohydrate recognition by a panel of anti-aGal antibodies. The method involves molecular docking of carbohydrates to antibodies and uses the docked carbohydrate poses to generate maps of the antibody binding sites in terms of prevalent hydrogen bonding and van der Waals interactions. We have applied this method to investigate peptide recognition by the anti-aGal antibodies. It was found that the site maps of the peptides and the carbohydrates were similar, indicating that the peptides interact with the same residues as those involved in carbohydrate recognition. This study demonstrates the potential for “design by mapping” of anti-carbohydrate antibody inhibitors. © 2010 Wiley Periodicals, Inc. Biopolymers (Pept Sci) 96: 177–180, 2011.

Fibrosis is the end point of most renal diseases, and several glycosaminoglycans have been shown to attenuate this process. Marine invertebrate glycosaminoglycans with unique structures have opened the possibility to test these new compounds on renal fibrosis. The effect of a fucosylated chondroitin sulfate from an echinoderm marine species is reported with the use of a model of renal fibrosis in rats, termed unilateral ureteral obstruction. Animals were given 4 mg/kg body wt of fucosylated chondroitin sulfate intraperitoneally, once a day. After 14 days, their kidneys were examined by histological, immunohistochemical, and biochemical methods. Compared with control mice, collagen deposition decreased in the course of renal fibrosis in the animals receiving fucosylated chondroitin sulfate, as revealed by Sirius red staining and hydroxyproline content. The cellularity related to myofibroblasts and macrophages was also reduced, as was the production of transforming growth factor (TGF)-β. The glycosaminoglycan content increased in the renal interstitium of animals submitted to unilateral ureteral obstruction compared with the control contralateral kidney, mostly due to an increase of chondroitin sulfate content. Interestingly, no change in the pattern of glycosaminoglycan deposition was observed after administration of fucosylated chondroitin sulfate. Fibrosis induced by unilateral ureteral obstruction is attenuated in P-selectin-deficient mice, which also do not respond to the invertebrate glycosaminoglycan. In conclusion, fucosylated chondroitin sulfate attenuates renal fibrosis on a ureteral obstruction model in mice preponderantly through a P-selectin-mediated mechanism.

Single-walled carbon nanotubes substitutionally doped with the light-element phosphorus are synthesized and are investigated by electrical and nuclear magnetic resonance measurements. Decreased spin−lattice relaxation times compared to undoped tubes point toward enhanced spin-sensitive scattering. Temperature dependence of the zero-bias conductance shows step-like features, a signature of scattering from a very low density (few sites per nanotube) of localized spin moments at oxidized phosphorus sites, consistent with density functional calculations. This supports recent predictions that localized magnetic moments must be indirectly magnetically coupled through the nanotube conduction electrons.

Nephropathy complicates the course and adversely impacts on the prognosis of HIV-infected patients. We examined trends and correlates of all-cause nephropathy (ACN).

This paper presents an analysis of the performance of a Pd-MOS structure with respect to their sensitivity to hydrogenated gases. Assays using an experimental arrangement of the scanning light pulse technique (SLPT) were carried out. This arrangement allowed the chemical images patterns generation that show the sensitivity of the device to different gas composition, which the sensor was exposed.

Food products are susceptible to mycotoxin contamination. The intoxication produces distinct signs, depending on mycotoxin, duration, dose, and species susceptibility. Behavioral impairments after mycotoxin exposure critically affect animal health and welfare. Nevertheless, this topic has been barely explored in the specific literature.

Transplantation of organs across species (xenotransplantation) is being considered to overcome the shortage of human donor organs. However, unmodified pig organs undergo an antibody-mediated hyperacute rejection that is brought about by the presence of natural antibodies to Galα(1,3)Gal, which is the major carbohydrate xenoantigen. Genetic modification of pig organs to remove most of the Galα(1,3)Gal epitopes has been achieved, but the human immune system may still recognize residual lipid-linked Galα(1,3)Gal carbohydrates, new (cryptic) carbohydrates or additional non-Galα(1,3)Gal carbohydrate xenoantigens. The structural basis for lectin and antibody recognition of Galα(1,3)Gal carbohydrates is starting to be understood and is discussed in this review. Antibody binding to Galα(1,3)Gal carbohydrates is predicted to primarily involve end-on insertion of the terminal αGal residue, but it is possible that groove-type binding can occur, as for some lectins. It is likely that similar antibody and lectin recognition will occur with other non-Galα(1,3)Gal xenoantigens, which potentially represent new barriers for pig-to-human xenotransplantation.

ABSTRACTHuman herpesvirus 6 (HHV-6) is a ubiquitous virus with which infections have been associated with pathologies ranging from delayed bone marrow engraftment to a variety of neurological diseases. The lack of a standardized assay that can be used to detect and estimate HHV-6 DNA contents in various clinical specimens can lead and has led to discordant results among investigators and on the potential association of HHV-6 to diseases. To identify the most reliable and sensitive assays, an identical set of 11 coded serum samples spiked with various quantities of the HHV-6A variant (range, 4 to 400,000 genome copies/ml) was sent to eight independent laboratories around the world. Each laboratory was asked to estimate the HHV-6 DNA content by use of its own protocols and assays. Among the various assays, three TaqMan-based real-time PCR assays yielded quantities that were closest to the quantity of HHV-6 that had been spiked. To provide better homogeneity between the results from the different laboratories working on HHV-6, we propose that investigators interested in quantifying HHV-6 in clinical samples adopt one of these assays.

The generation of knockout pigs was a major achievement for the field of xenotransplantation. This review will look at key findings reported within the past 12 months relevant to the role of carbohydrates in the α1,3-galactosyltransferase knockout pig lines.

In end‐stage renal disease patients treated by hemodialysis with HBeAg‐negative chronic hepatitis B virus (HBV) infection, the evaluation of the presence of viral replication is essential in the assessment for renal transplantation. Data on HBV viral load, prevalence of precore mutations, as well as the influence of HCV coinfection on HBV‐DNA levels in this group of patients is scarce. The aim of this study was to determine the HBV viral load in HBsAg‐positive/HBeAg‐negative hemodialysis patients; to compare HBV‐DNA levels between isolated HBV infection carriers and HBV‐HCV coinfected patients, and to evaluate the prevalence of precore mutations in these patients. Fifty hemodialysis patients with chronic HBeAg‐negative HBV infection were studied. Viral load was determined by PCR (Amplicor HBV Monitor‐Roche). The detection of precore mutations was made by sequencing. Of a total of 50 patients, 76% were male, with a mean age of 44 ± 11 years. Anti‐HCV was positive in 56% of patients. HBV‐DNA was undetectable in 58% of patients; 24% had HBV‐DNA <10,000 copies/ml, 12% between 10,000–100,000 copies/ml, and only 6% had HBV‐DNA >100,000 copies/ml. There was no difference in the viral load of patients infected only by HBV and HBV‐HCV co‐infected patients (P= 0.96). Precore mutations were detected in only 8% of cases. In conclusion, hemodialysis patients with HBeAg‐negative HBV infection had a low viral load. Precore mutations were infrequent and the presence of anti‐HCV has not influenced the levels of HBV‐DNA. J. Med. Virol. 78:1284–1288, 2006. © 2006 Wiley‐Liss, Inc.

Heparin exerts beneficial effects in different experimental models of nephropathy, as observed by the preservation of the structural morphology of the kidney after heparin therapy. Here we investigate molecular and cellular events involved in the protective effects of heparin in the progression of renal disease after unilateral ureteral obstruction.

Heparin cofactor II (HCII) is a plasma protein that inhibits thrombin rapidly in the presence of dermatan sulfate or heparin. We previously reported that the time to thrombotic occlusion of the carotid artery after photochemical injury was shorter in HCII-deficient mice than in wild-type control animals. In this paper, we describe the antithrombotic activity of dermatan sulfate in wild-type and HCII-deficient mice. Intravenous administration of porcine skin dermatan sulfate induced a dose-dependent prolongation of the carotid artery occlusion time in HCII+/+ mice that was not observed in HCII-/- animals. Pharmacokinetic studies suggested that porcine skin dermatan sulfate expresses antithrombotic activity after being transferred from the plasma to sites in the vessel wall. Using invertebrate dermatan sulfate preparations, we showed that N-acetylgalactosamine-4-O-sulfate residues are required for the HCII-dependent antithrombotic effect. Furthermore, the invertebrate dermatan sulfates, which have higher charge densities than mammalian dermatan sulfate, slightly prolonged the thrombotic occlusion time of HCII-/- mice. These results indicate that HCII mediates the antithrombotic effect of porcine skin dermatan sulfate after injury to the carotid arterial endothelium in mice, whereas more highly charged dermatan sulfates possess weak antithrombotic activity independent of HCII. (Blood. 2004;104:3965-3970)

ABSTRACTSensitive and specific techniques to detect and identify Mycobacterium tuberculosisdirectly in clinical specimens are important for the diagnosis and management of patients with tuberculosis (TB). We developed two real-time PCR assays, based on the IS6110multicopy element and on the senX3-regX3intergenic region, which provide a rapid method for the diagnosis of mycobacterial infections. The sensitivity and specificity of both assays were established by using purified DNA from 71 clinical isolates and 121 clinical samples collected from 83 patients, 20 of whom were affected by TB. Both assays are accurate, sensitive, and specific, showing a complementary pattern of Mycobacteriumrecognition: broader for the IS6110-based assay and restricted to the M. tuberculosiscomplex for the senX3-regX3-based assay. Moreover, the addition of a synthetic DNA calibrator prior to DNA extraction allowed us to measure the efficiency of DNA recovery and to control for the presence of PCR inhibitors. The mycobacterial burden of the clinical samples, as assessed by direct microscopy, correlates with the M. tuberculosisDNA load measured by the senX3-regX3-based assay. In addition, reduced levels of M. tuberculosisDNA load are present in those patients subjected to successful therapy, suggesting a potential use of this assay for monitoring treatment efficacy. Therefore, these assays represent a fully controlled high-throughput system for the evaluation of mycobacterial burden in clinical specimens.

The distribution of hepatitis C virus (HCV) genotypes in patients on hemodialysis and in kidney transplant recipients was compared with that observed in a control group composed of HCV‐infected individuals from the general population. A total of 340 patients were included in the study: 46 with end‐stage renal disease on regular hemodialysis treatment, 22 kidney transplant recipients and 272 controls matched for sex and age at a 4:1 ratio (controls to patient). HCV genotype was determined by sequencing of the 5′ untranslated region of the HCV genome. No difference was observed in the distribution of HCV genotypes in hemodialysis patients and renal transplant patients (P= 0.47). However, when each of these groups was compared with the control group, a significant difference was detected in the genotype distribution (P< 0.001). In hemodialysis and renal transplant patients the most prevalent subtype was 1a, followed by 1b, 3, and other less prevalent genotypes (2, 4, and 5), whereas in the control group the most prevalent subtype was 1b, followed by 3, 1a, and others. That observation may reflect differences in the epidemiology of HCV infection, viral characteristics and host factors in renal patients in comparison to the control group. J. Med. Virol. 69:489–494, 2003. © 2003 Wiley‐Liss, Inc.

ABSTRACTAccurate laboratory tests for the diagnosis of active human herpesvirus 8 (HHV-8) infection are becoming essential to study the pathogenesis of HHV-8-associated tumors and for the clinical management of HHV-8-infected individuals. We have developed a highly sensitive, calibrated quantitative real-time PCR assay for the measurement of cell-free HHV-8 DNA in body fluids, based on the addition of a synthetic DNA calibrator prior to DNA extraction. The calibrator controls each sample for the presence of PCR inhibitors, determines a cutoff value of sensitivity for negative samples, and normalizes positive samples for the efficiency of DNA recovery. The assay shows a wide dynamic range of detection (between 1 and 106viral genome equivalents/reaction) and a high degree of accuracy even in the presence of high amounts (up to 1 µg) of human genomic DNA. Moreover, the assay has a very high sensitivity (lower detection limit, 10 genome equivalents/ml) and a high degree of reproducibility and repeatability with a coefficient of variation (CV) of <15 and 23%, respectively. Furthermore, the use of the calibrator improves the accuracy of quantitation and decreases the intersample variability (CV, 9 and 6%, respectively). The sensitivity and specificity of the assay were tested with a series of clinical specimens obtained from patients affected by various HHV-8-related diseases, as well as from a wide number of controls. In conclusion, our calibrated real-time PCR assay provides a reliable high-throughput method for quantitation of HHV-8 DNA in clinical and laboratory specimens.

Alpha(1,3)Galactosyltransferase (GT) is a Golgi-localized enzyme that catalyzes the transfer of a terminal galactose to N-acetyllactosamine to create Galalpha(1,3)Gal. This glycosyltransferase has been studied extensively because the Galalpha(1,3)Gal epitope is involved in hyperacute rejection of pig-to-human xenotransplants. The original crystal structure of bovine GT defines the amino acids forming the catalytic pocket; however, those directly involved in the interaction with the donor nucleotide sugars were not characterized. Comparison of amino acid sequences of GT from several species with the human A and B transferases suggest that His271 of pig GT may be critical for recognition of the donor substrate, UDP-Gal. Using pig GT as the representative member of the GT family, we show that replacement of His271 with Ala, Leu, or Gly caused complete loss of function, in contrast to replacement with Arg, another basic charged residue, which did not alter the ability of GT to produce Galalpha(1,3)Gal. Molecular modeling showed that His271 may interact directly with the Gal moiety of UDP-Gal, an interaction possibly retained by replacing His with Arg. However, replacing His271 with amino acids found in alpha(1,3)GalNAc transferases did not change the donor nucleotide specificity. Thus His271 is critical for enzymatic function of pig GT.

Historically, the most effective means of modifying cell surface carbohydrates has required the intracellular overexpression of glycosyltransferases or glycosidases and is dependent on the enzymes occupying a cellular localization close to the carbohydrate structures they modify. We report on relocalizing the lysosomal resident glycosidase human α-galactosidase to other regions of the cell, Golgi and cell surface, where it is in closer proximity for cleaving the carbohydrate structure Galα(1,3)Gal. Relocalization of α-galactosidase was achieved by using the transmembrane and cytoplasmic domains from the human protein furin, which is known to localize in the trans-Golgi network (TGN) and cell surface. Two chimeric forms of α-galactosidase were generated, one directing it to the TGN of the cell and the other to the cell surface, as shown by confocal microscopy. The relocalized enzymes have the ability to cleave terminal α-galactose as detected by expression on the cell surface. Furthermore, when expressed as a transgene in mice, the TGN form of α-galactosidase was more effective at decreasing cell surface terminal α-galactose than was the native lysosomal form. When expressed in conjunction with the α1,2fucosyltransferase that also decreases Galα(1,3)Gal, the reduction was additive. The ability to relocalize enzymes that modify cell surface carbohydrate structures has far-reaching implications in biology and may be useful in such fields as xenotransplantation and treatment of glycosidase disorders.

Historically, the most effective means of modifying cell surface carbohydrates has required the intracellular overexpression of glycosyltransferases or glycosidases and is dependent on the enzymes occupying a cellular localization close to the carbohydrate structures they modify. We report on relocalizing the lysosomal resident glycosidase human alpha-galactosidase to other regions of the cell, Golgi and cell surface, where it is in closer proximity for cleaving the carbohydrate structure Galalpha(1,3)Gal. Relocalization of alpha-galactosidase was achieved by using the transmembrane and cytoplasmic domains from the human protein furin, which is known to localize in the trans-Golgi network (TGN) and cell surface. Two chimeric forms of alpha-galactosidase were generated, one directing it to the TGN of the cell and the other to the cell surface, as shown by confocal microscopy. The relocalized enzymes have the ability to cleave terminal alpha-galactose as detected by expression on the cell surface. Furthermore, when expressed as a transgene in mice, the TGN form of alpha-galactosidase was more effective at decreasing cell surface terminal alpha-galactose than was the native lysosomal form. When expressed in conjunction with the alpha1,2fucosyltransferase that also decreases Galalpha(1,3)Gal, the reduction was additive. The ability to relocalize enzymes that modify cell surface carbohydrate structures has far-reaching implications in biology and may be useful in such fields as xenotransplantation and treatment of glycosidase disorders.

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The variance matrix between the emission probabilities of the strong gamma-rays following the βdecay of 152Eu were determined in a specially designed experiment. The emission probabilities are deduced from the decay scheme, whose branching-ratios and beta feeding fractions were fitted to the observed peak areas.

Nineteen monoclonal antibodies (MAbs) against tissue-nonspecific (liver/bone/kidney) alkaline phosphatase (TNALP) were investigated in the ISOBM TD-9 Workshop. These MAbs were generated with antigens obtained from human bone tissue (n = 9), human osteosarcoma cell lines (SaOS-2 and TPX; n = 7) and human liver tissue (n = 3). The evaluation included the following antigen forms: (a) commercially available preparations of human bone ALP (BALP) and liver ALP (LALP); (b) human BALP isoforms, B/I, B1 and B2; and (c) soluble secreted epitope-tagged recombinant human TNALP (setTNALP) expressed in COS-1, osteosarcoma (SaOS-2) and hepatoma (Huh2) cell lines. In addition, 16 TNALP mutant cDNAs corresponding to a wide spectrum of reported hypophosphatasia mutations were used in an attempt to map specific immunoreactive epitopes on the surface of the TNALP molecule. The TD-9 MAbs were evaluated by immunoradiometric (IRMA) assays, cross-inhibition and different enzyme immunoassay designs. No indications of explicit tissue discriminatory immunoreactivities of the investigated MAbs against TNALP were found. However, certain IRMA combinations of MAbs increased the specificity of BALP measurements. All MAbs bound to the three BALP isoforms B/I, B1 and B2, but none of the investigated MAbs were specific for any of the isoforms. Significant differences were, however, found in immunoreactivity between these isoforms, with cross-reactivities ranging from 21 to 109% between the two major BALP isoforms B1 and B2. Desialylation with neuraminidase significantly increased the MAb affinity for the BALP isoforms B/I, B1 and B2, and also decreased the observed differences in cross-reactivity between these isoforms. We suggest, therefore, that the MAb affinity is dependent on the amount/number of terminal sialic acid residues located at the five putative N-glycosylation sites. Based on the overall results, we present a putative three-dimensional model of the TNALP molecule with positioning of the four major antigenic domains (designated A–D) of the investigated MAbs. The TNALP molecule is depicted as a homodimer, hence most, but not necessarily all, epitopes are displayed twice. The antigenic domains were positioned with the following assumptions: domain A was positioned close to the active site since most of these MAbs interfered with the catalytic activity. Interestingly, both MAbs included in the commercial BALP kits were grouped with domain A. Moreover, 4 of the 5 putative N-glycosylation sites (with terminal sialic acid residues) are located within, or with close proximity to, domain A. Domain B was localized at the top flexible loop (crown domain) of the TNALP molecule. Domain C was clearly defined by the IRMA assay combinations and by site-directed mutants of TNALP to be close to residue E281, which is located near the fourth metal binding site, likely to be occupied by a calcium ion. Domain D was positioned close to residues A115, A162 and E174, but this domain was also close to the GPI anchor site. In conclusion, none of the 19 investigated TD-9 MAbs were entirely specific for BALP or LALP, thus indicating that all MAbs bind mainly to epitopes on the common protein core of BALP and LALP and/or common glycosylated epitopes. However, some MAbs (either single or in combination with other MAbs) work sufficiently well to measure BALP when the assayed samples do not contain elevated levels of LALP.

It is currently under debate whether the mechanism of Golgi retention of different glycosyltransferases is determined by sequences in the transmembrane, luminal, or cytoplasmic domains or a combination of these domains. We have shown that the cytoplasmic domains of α1,3-galactosyltransferase (GT) and α1,2-fucosyltransferase (FT) are involved in Golgi localization. Here we show that the cytoplasmic tails of GT and FT are sufficient to confer specific Golgi localization. Further, we show that the expression of only the cytoplasmic tail of GT can lead to displacement or inhibition of binding of the whole transferase and that cells expressing the cytoplasmic tail of GT were not able to express full-length GT or its product, Galα1,3Gal. Thus, the presence of the cytoplasmic tail prevented the localization and function of full-length GT, suggesting a possible specific Golgi binding site for GT. The effect was not altered by the inclusion of the transmembrane domain. Although the transmembrane domain may act as an anchor, these data show that, for GT, only the cytoplasmic tail is involved in specific localization to the Golgi.

The cross section for the production of ωmesons in proton–proton collisions has been measured in a previously unexplored region of incident energies. Cross sections of σ=(7.5±1.9) μb and σ=(30.8±3.4) μb (with 20% systematic uncertainties) were extracted at ϵ=92 MeV and 173 MeV excess energy above the ωthreshold, respectively. The angular distribution of the ωat ϵ=173 MeV is strongly anisotropic, demonstrating the importance of partial waves beyond pure s-wave production at this energy.

Two brothers with myopathic coenzyme Q10(CoQ10) deficiency responded dramatically to CoQ10supplementation. Muscle biopsies before therapy showed ragged-red fibers, lipid storage, and complex I III and II III deficiency. Approximately 30 of myofibers had multiple features of apoptosis. After 8 months of treatment, excessive lipid storage resolved, CoQ10level normalized, mitochondrial enzymes increased, and proportion of fibers with TUNEL-positive nuclei decreased to 10. The authors conclude that muscle CoQ10deficiency can be corrected by supplementation of CoQ10, which appears to stimulate mitochondrial proliferation and to prevent apoptosis.

Natural antibodies that react with galactose-(1,3)galactose gal(1,3)gal carbohydrate epitopes exist in humans and Old World primates because of the inactivation of the 1,3-galactosyltransferase (1,3GT) gene in these species and the subsequent production of antibodies to environmental microbes that express the gal(1,3)gal antigen. The Gal knockout (Gal o/o) mouse, produced by homologous disruption of the 1,3GT gene, spontaneously makes anti-gal(1,3)gal antibodies and can be used to study the genetic control of humoral immune responses to this carbohydrate epitope.