Graph-based description of tertiary lymphoid organs at single-cell level.
Our aim is to complement observer-dependent approaches of immune cell evaluation in microscopy images with reproducible measures for spatial composition of lymphocytic infiltrates. Analyzing such patterns of inflammation is becoming increasingly important for therapeutic decisions, for example in transplantation medicine or cancer immunology. We developed a graph-based assessment of lymphocyte clustering in full whole slide images. Based on cell coordinates detected in the full image, a Delaunay triangulation and distance criteria are used to build neighborhood graphs. The composition of nodes and edges are used for classification, e.g. using a support vector machine. We describe the variability of these infiltrates on CD3/CD20 duplex staining in renal biopsies of long-term functioning allografts, in breast cancer cases, and in lung tissue of cystic fibrosis patients. The assessment includes automated cell detection, identification of regions of interest, and classification of lymphocytic clusters according to their degree of organization. We propose a neighborhood feature which considers the occurrence of edges with a certain type in the graph to distinguish between phenotypically different immune infiltrates. Our work addresses a medical need and provides a scalable framework that can be easily adjusted to the requirements of different research questions.


Numerical correction of anti-symmetric aberrations in single HRTEM images of weakly scattering 2D-objects.
Here, we present a numerical post-processing method for removing the effect of anti-symmetric residual aberrations in high-resolution transmission electron microscopy (HRTEM) images of weakly scattering 2D-objects. The method is based on applying the same aberrations with the opposite phase to the Fourier transform of the recorded image intensity and subsequently inverting the Fourier transform. We present the theoretical justification of the method, and its verification based on simulated images in the case of low-order anti-symmetric aberrations. Ultimately the method is applied to experimental hardware aberration-corrected HRTEM images of single-layer graphene and MoSe₂ resulting in images with strongly reduced residual low-order aberrations, and consequently improved interpretability. Alternatively, this method can be used to estimate by trial and error the residual anti-symmetric aberrations in HRTEM images of weakly scattering objects.


The effect of isotretinoin on ovarian reserve based on hormonal parameters, ovarian volume, and antral follicle count in women with acne.
BACKGROUD/AIMS: Widely prescribed in routine practice, isotretinoin has an unknown impact on ovarian reserve. With a long history in acne treatment and numerous potential side effects, it is surprising that very few prospective studies have investigated its effect on ovarian reserve. Therefore, we aimed to evaluate the impact of oral isotretinoin on ovarian reserve based on hormonal parameters, anti-Müllerian hormone (AMH), ovarian volume (OV), and antral follicle count (AFC) in women of reproductive age with acne. Our study group consisted of 82 women of reproductive age with acne who were treated with oral isotretinoin. The patients were evaluated for ovarian reserve prior to therapy and reevaluated 6 months after isotretinoin treatment with regard to hormonal parameters, AMH, OV, and AFC. Significant differences were found between the pre- and posttreatment period for AMH [2.20 ng/ml (25th-75th percentile 1.14-4.07) vs. 1.31 ng/ml (0.32-2.28)], total AFC [16 (14-18.25) vs. 12.5 (10-15)], and total OV [23 ml (18-29) vs. 15 ml (13-18); p < 0.001]. Our study is the first to analyze the levels of serum AMH, AFC, and OV together in patients treated with oral isotretinoin for acne. The results of our study demonstrated that oral isotretinoin had a significant negative effect on ovarian reserve.


Theoretical and experimental analysis of tunable Sagnac high-birefringence loop filter for dual-wavelength laser application.
We present detailed investigations of the spectral dependencies of the transmission of a fiber optical loop mirror (FOLM) consisting of a coupler with output ports spliced at arbitrary angles to a high-birefringence (Hi-Bi) fiber. The application for dual-wavelength lasers is discussed. For this aim, the spectral dependence of the reflection is tuned by the temperature of the Hi-Bi fiber that allows a fine adjustment of the cavity loss for generated wavelengths. The ratio between maximum and minimum reflection can be adjusted by the twist angle of the fiber at the splices, which also provides useful possibilities for the adjustment of cavity losses. We used the twist and temperature variation of the Hi-Bi fiber to change the operation from single wavelength to stable dual-wavelength generation with either equal or unequal powers of wavelengths.


Reliability of diabetic patients' gait parameters in a challenging environment.
Activities of daily life require us to move about in challenging environments and to walk on varied surfaces. Irregular terrain has been shown to influence gait parameters, especially in a population at risk for falling. A precise portable measurement system would permit objective gait analysis under such conditions. The aims of this study are to (a) investigate the reliability of gait parameters measured with the Physilog in diabetic patients walking on different surfaces (tar, grass, and stones); (b) identify the measurement error (precision); (c) identify the minimal clinical detectable change. 16 patients with Type 2 diabetes were measured twice within 8 days. After clinical examination patients walked, equipped with a Physilog, on the three aforementioned surfaces. ICC for each surface was excellent for within-visit analyses (>0.938). Inter-visit ICC's (0.753) were excellent except for the knee range parameter (>0.503). The coefficient of variation (CV) was lower than 5% for most of the parameters. Bland and Altman Plots, SEM and SDC showed precise values, distributed around zero for all surfaces. Good reliability of Physilog measurements on different surfaces suggests that Physilog could facilitate the study of diabetic patients' gait in conditions close to real-life situations. Gait parameters during complex locomotor activities (e.g. stair-climbing, curbs, slopes) have not yet been extensively investigated. Good reliability, small measurement error and values of minimal clinical detectable change recommend the utilization of Physilog for the evaluation of gait parameters in diabetic patients.


Student help seeking from pastoral care in UK high schools: a qualitative study.
Little is known about high school students' perceptions of school-based pastoral support. This study aimed to explore this in the context of a high school-based emotional support project. Qualitative interviews explored perspectives on help seeking of students (N = 23) and staff (N = 27) in three UK high schools where a pastoral project was introduced. Data were analysed thematically. Student peer groups perceived help seeking as a sign of weakness. However, students valued learning skills in managing emotions and friendships. Staff expressed concerns about students' ability to help seek proactively, and highlighted organisational influences on pastoral support. Increasing student control over the process, and involving trusted staff, could encourage help seeking in high school. It is possible to access the views of students who do not help seek, to improve understanding of help-seeking behaviour.


Phase IB randomized, double-blinded, placebo-controlled, dose escalation study of polyphenon E in women with hormone receptor-negative breast cancer.
Epidemiologic data support an inverse association between green tea intake and breast cancer risk, and numerous experimental studies have shown the antitumor effects of its main component, epigallocatechin gallate (EGCG). We conducted a phase IB dose escalation trial in women with a history of stage I to III hormone receptor-negative breast cancer of an oral green tea extract, polyphenon E (Poly E) 400, 600, 800 twice daily or matching placebo for 6 months. The primary endpoint was to determine the maximum tolerated dose (MTD), defined as the dose that causes 25% dose-limiting toxicity (DLT, grade ≥II). Assignment to dose level was based upon an adaptive design, the continual reassessment method. A mammogram and random core biopsy of the contralateral breast were obtained at baseline and 6 months and serial blood/urine collections every 2 months for biomarker analyses. Forty women were randomized: 10 to placebo, 30 to Poly E (16 at 400 mg, 11 at 600 mg, 3 at 800 mg). There was one DLT at 400 mg (grade III rectal bleeding), three DLTs at 600 mg (grade II weight gain, grade III indigestion and insomnia), and one DLT at 800 mg (grade III liver function abnormality). The DLT rate at 600 mg was 27% (3 of 11). Pharmacologic levels of total urinary tea polyphenols were achieved with all three dose levels of Poly E. Using a novel phase I trial design, we determined the MTD for Poly E to be 600 mg twice daily. This study highlights the importance of assessing toxicity for any chemopreventive agent being developed for chronic use in healthy individuals.


Selegiline Recovers Synaptic Plasticity in the Medial Prefrontal Cortex and Improves Corresponding Depression-Like Behavior in a Mouse Model of Parkinson's Disease.
In patients with Parkinson's disease (PD), non-motor symptoms (NMS) including depression and anxiety are often recognized before motor symptoms develop. Monoamine oxidase (MAO)-B inhibitors are therapeutically effective for motor symptoms; however, their effects on NMS in PD are yet to be fully assessed. Here, we aimed to explore the antidepressant-like effects of propargyl MAO-B inhibitors, selegiline and rasagiline, in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) as a PD model, and to elucidate the mechanisms underlying these effects. Four repeated intraperitoneal injections of MPTP at 17.5 mg/kg to C57BL/6 mice led to a partial reduction in the number of nigrostriatal tyrosine hydroxylase-positive neurons and to the extension of immobility time during the tail suspension test (TST), without any obvious induction of motor deficits. A single subcutaneous administration of selegiline at 10 mg/kg shortened the extended immobility time of MPTP mice in the TST, without any increase in motor activities, suggesting that selegiline exerts antidepressant-like effects. In this test, rasagiline did not produce antidepressant-like effects, although the inhibitory effect of 3 mg/kg rasagiline on brain MAO activity was comparable to that of 10 mg/kg selegiline. The shortened immobility time in the TST correlated with reduced cortical dopamine (DA) turnover rates in MPTP mice treated with selegiline, but not in MPTP mice treated with rasagiline. These results suggest that MAO inhibition does not entirely account for the antidepressant-like effects of selegiline. Administration of selegiline (10 mg/kg), but not rasagiline (1 mg/kg), to MPTP mice restored the impaired long-term potentiation induced by high-frequency stimulation in the medial prefrontal cortex (mPFC), and normalized the reduced phosphorylation of Ca2+/calmodulin-dependent protein kinase IIα, which is known to be involved in neuroplasticity, in the frontal cortex. In MPTP mice, the antiparkinsonian drug pramipexole (0.3 mg/kg), a DA D2 and D3 receptor agonist, that has been shown to be effective in treating depression in PD, ameliorated depression-like behavior and synaptic dysfunction in the mPFC. Taken together, the antidepressant-like effects of selegiline in MPTP mice are attributable to the restoration of impaired synaptic plasticity in the mPFC, suggesting its potential for treating depression in early PD.


Toward high-throughput screening of NAD(P)-dependent oxidoreductases using boron-doped diamond microelectrodes and microfluidic devices.
Although oxidoreductases are widely used in many applications, such as biosensors and biofuel cells, improvements in the function of existing oxidoreductases or the discovery of novel oxidoreductases with greater activities is desired. To increase the activity of oxidoreductases by directed evolution, a powerful screening technique for oxidoreductases is required. In this study, we demonstrate the utility of boron-doped diamond (BDD) microelectrodes for quantitative and potentially high-throughput measurement of the activity of NAD(P)-dependent oxidoreductases. We first confirmed that BDD microelectrodes can quantify the activity of low concentrations (10-100 pM) of glucose-6-phosphate dehydrogenase and alcohol dehydrogenase with a measuring time of 1 ms per sample. In addition, we found that poisoning of BDD microelectrodes can be repressed by optimizing the pH and by adding l-arginine to the enzyme solution as an antiaggregation agent. Finally, we fabricated a microfluidic device containing a BDD electrode for the first time and observed the elevation of the oxidation current of NADH with increasing flow rate. These results imply that the combination of a BDD microelectrode and microfluidics can be used for high-throughput screening of an oxidoreductase library containing a large number (>10(6)) of samples, each with a small (nanoliter) sample volume.


Comparison of Detection Methods for a Novel Tobamovirus Isolated from Florida Hibiscus.
A novel tobamovirus recently was isolated from hibiscus in Florida. Serological and molecular methods, including enzyme-linked immunosorbent assay (ELISA), dot-blot immunoassay (DBIA), tissue-blot immunoassay (TBIA), and immunocapture reverse-transcription poly-merase chain reaction (IC-RT-PCR) were compared to evaluate their usefulness for diagnosis of this virus. Each method was tested with partially purified virus preparations and tissue samples from infected hibiscus and Chenopodium quinoa plants. Indirect ELISA was more sensitive than double-antibody sandwich (DAS)-ELISA with all samples tested. The Florida hibiscus virus was detectable in hibiscus leaves and bark up to 1:12,800 and 1:6,400 dilutions, respectively, by indirect ELISA and up to 1:3,200 and 1:400 dilutions by DAS-ELISA. End-point dilutions of partially purified virus preparations from indirect and DAS-ELISA were 4 and 31 ng/ml, respectively. Florida hibiscus virus was detected by DBIA in sap from hibiscus bark and leaves at dilutions up to 1:400 and 1:800, respectively, showing that DBIA was less sensitive than either ELISA method. The virus also was detected reliably by TBIA from leaves and bark of hibiscus plants. The most sensitive method was IC-RT-PCR, which could detect as little as 500 pg/ml of virus in partially purified preparations and was 16- and 32-fold more sensitive than DAS-ELISA with hibiscus bark and leaf extracts, respectively. Over 600 hibiscus samples were tested by various combinations of these methods to validate their usefulness.


How does the brain discriminate familiar and unfamiliar faces?: a PET study of face categorical perception.
Where and how does the brain discriminate familiar and unfamiliar faces? This question has not been answered yet by neuroimaging studies partly because different tasks were performed on familiar and unfamiliar faces, or because familiar faces were associated with semantic and lexical information. Here eight subjects were trained during 3 days with a set of 30 faces. The familiarized faces were morphed with unfamiliar faces. Presented with continua of unfamiliar and familiar faces in a pilot experiment, a group of eight subjects presented a categorical perception of face familiarity: there was a sharp boundary in percentage of familiarity decisions between 40% and 60% faces. In the main experiment, subjects were scanned (PET) on the fourth day (after 3 days of training) in six conditions, all requiring a sex classification task. Completely novel faces (0%) were presented in Condition 1 and familiar faces (100%) in Condition 6, while faces of steps of 20% in the continuum of familiarity were presented in Conditions 2 to 5 (20% to 80%). A principal component analysis (PCA) indicated that most variations in neural responses were related to the dissociation between faces perceived as familiar (60% to 100%) and faces perceived as unfamiliar (0 to 40%). Subtraction analyses did not disclose any increase of activation for faces perceived as familiar while there were large relative increases for faces perceived as unfamiliar in several regions of the right occipito-temporal visual pathway. These changes were all categorical and were observed mainly in the right middle occipital gyrus, the right posterior fusiform gyrus, and the right inferotemporal cortex. These results show that (1) the discrimination between familiar and unfamiliar faces is related to relative increases in the right ventral pathway to unfamiliar/novel faces; (2) familiar and unfamiliar faces are discriminated in an all-or-none fashion rather than proportionally to their resemblance to stored representations; and (3) categorical perception of faces is associated with abrupt changes of brain activity in the regions that discriminate the two extremes of the multidimensional continuum.


Phylum-wide comparative genomics unravel the diversity of secondary metabolism in Cyanobacteria.
Cyanobacteria are an ancient lineage of photosynthetic bacteria from which hundreds of natural products have been described, including many notorious toxins but also potent natural products of interest to the pharmaceutical and biotechnological industries. Many of these compounds are the products of non-ribosomal peptide synthetase (NRPS) or polyketide synthase (PKS) pathways. However, current understanding of the diversification of these pathways is largely based on the chemical structure of the bioactive compounds, while the evolutionary forces driving their remarkable chemical diversity are poorly understood. We carried out a phylum-wide investigation of genetic diversification of the cyanobacterial NRPS and PKS pathways for the production of bioactive compounds. 452 NRPS and PKS gene clusters were identified from 89 cyanobacterial genomes, revealing a clear burst in late-branching lineages. Our genomic analysis further grouped the clusters into 286 highly diversified cluster families (CF) of pathways. Some CFs appeared vertically inherited, while others presented a more complex evolutionary history. Only a few horizontal gene transfers were evidenced amongst strongly conserved CFs in the phylum, while several others have undergone drastic gene shuffling events, which could result in the observed diversification of the pathways. Therefore, in addition to toxin production, several NRPS and PKS gene clusters are devoted to important cellular processes of these bacteria such as nitrogen fixation and iron uptake. The majority of the biosynthetic clusters identified here have unknown end products, highlighting the power of genome mining for the discovery of new natural products.


Development of pulmonary endovascular metastases following vertebroplasty: case report.
A 69-year-old man developed pulmonary metastases following vertebroplasties for pathological fractures of vertebrae T12-L4. The fractures developed due to spinal metastases from castrate-resistant prostate cancer. A CT scan performed 1 month prior indicated no evidence of pulmonary malignancy. However, CT scans performed 2 months after the vertebroplasties demonstrated intravascular pulmonary metastases distributed similarly to embolized polymethylmethacrylate. Vertebroplasty is a well-established procedure for symptomatic management of vertebral compression fractures. However, studies have demonstrated an increase in circulating tumor cells following vertebroplasties, theoretically increasing the risk of distant metastases. In this case, the chronicity and radiological findings suggest that the pulmonary intravascular metastases may have resulted from the vertebroplasties.


Integrated computational prediction and experimental validation identifies promiscuous T cell epitopes in the proteome of Mycobacterium bovis.
The discovery of novel antigens is an essential requirement in devising new diagnostics or vaccines for use in control programmes against human tuberculosis (TB) and bovine tuberculosis (bTB). Identification of potential epitopes recognised by CD4+ T cells requires prediction of peptide binding to MHC class-II, an obligatory prerequisite for T cell recognition. To comprehensively prioritise potential MHC-II-binding epitopes from Mycobacterium bovis, the agent of bTB and zoonotic TB in humans, we integrated three binding prediction methods with the M. bovisproteome using a subset of human HLA alleles to approximate the binding of epitope-containing peptides to the bovine MHC class II molecule BoLA-DRB3. Two parallel strategies were then applied to filter the resulting set of binders: identification of the top-scoring binders or clusters of binders. Our approach was tested experimentally by assessing the capacity of predicted promiscuous peptides to drive interferon-γ secretion from T cells of M. bovis infected cattle. Thus, 376 20-mer peptides, were synthesised (270 predicted epitopes, 94 random peptides with low predictive scores and 12 positive controls of known epitopes). The results of this validation demonstrated significant enrichment (>24 %) of promiscuously recognised peptides predicted in our selection strategies, compared with randomly selected peptides with low prediction scores. Our strategy offers a general approach to the identification of promiscuous epitopes tailored to target populations where there is limited knowledge of MHC allelic diversity.


Ezrin-radixin-moesin-binding phosphoprotein 50 is expressed at the apical membrane of rat liver epithelia.
Ezrin-radixin-moesin (ERM)-binding phosphoprotein 50 (EBP50) and NHE3 Kinase A regulatory protein (E3KARP) are membrane-cytoskeleton linking proteins that utilize 2 PSD-95/DIg/ZO-1 (PDZ) domains and an ERM binding site to coordinate cyclic adenosine monophosphate (cAMP)-regulated ion transport in a number of distinct epithelia. ERM family members serve to anchor EBP50 and E3KARP to the actin cytoskeleton and sequester protein kinase A (PKA) to these protein complexes. In hepatocytes and cholangiocytes, the epithelial cells of the bile secretory unit, cAMP-activated PKA stimulates secretion and bile formation, but the molecular mechanisms, including the potential contribution of EBP50 and E3KARP, remain undetermined. The present studies evaluated the comparative expression and localization of EBP50 and E3KARP in rat hepatocytes and cholangiocytes. Complementary DNAs encoding rat EBP50 and E3KARP were identified by reverse transcription-polymerase chain reaction in both epithelial cell types and subsequently sequenced. Northern and Western analysis showed the presence of EBP50 messenger RNA and protein in both hepatocytes and cholangiocytes. Confocal immunofluorescence revealed EBP50 was concentrated at the apical domain of both cell types. E3KARP was also expressed in cholangiocytes but had a distinct cytoplasmic/nuclear distribution. In dominant-negative transfection studies, patch clamp analysis of Mz-ChA1 cholangiocarcinoma cells showed that expression of the PDZ1 domain of EBP50 selectively decreased the endogenous cAMP-mediated Cl secretory response. The apical expression of EBP50, presence of specific ERM proteins, and functional effects of PDZ1 expression on cholangiocyte secretion suggest EBP50 is positioned to contribute to the organization and regulation of bile secretory proteins in both hepatocytes and cholangiocytes.


Proteomics show antigen presentation processes in human immune cells after AS03-H5N1 vaccination.
Adjuvants enhance immunity elicited by vaccines through mechanisms that are poorly understood. Using a systems biology approach, we investigated temporal protein expression changes in five primary human immune cell populations: neutrophils, monocytes, natural killer cells, T cells, and B cells after administration of either an Adjuvant System 03 adjuvanted or unadjuvanted split-virus H5N1 influenza vaccine. Monocytes demonstrated the strongest differential signal between vaccine groups. On day 3 post-vaccination, several antigen presentation-related pathways, including MHC class I-mediated antigen processing and presentation, were enriched in monocytes and neutrophils and expression of HLA class I proteins was increased in the Adjuvant System 03 group. We identified several protein families whose proteomic responses predicted seroprotective antibody responses (>1:40 hemagglutination inhibition titer), including inflammation and oxidative stress proteins at day 1 as well as immunoproteasome subunit (PSME1 and PSME2) and HLA class I proteins at day 3 in monocytes. While comparison between temporal proteomic and transcriptomic results showed little overlap overall, enrichment of the MHC class I antigen processing and presentation pathway in monocytes and neutrophils was confirmed by both approaches.


Muscle glucose transport and phosphorylation in type 2 diabetic, obese nondiabetic, and genetically predisposed individuals.
Our objectives were to quantitate insulin-stimulated inward glucose transport and glucose phosphorylation in forearm muscle in lean and obese nondiabetic subjects, in lean and obese type 2 diabetic (T2DM) subjects, and in normal glucose-tolerant, insulin-resistant offspring of two T2DM parents. Subjects received a euglycemic insulin (40 mU.m(-2).min(-1)) clamp with brachial artery/deep forearm vein catheterization. After 120 min of hyperinsulinemia, a bolus of d-mannitol/3-O-methyl-d-[(14)C]glucose/d-[3-(3)H]glucose (triple-tracer technique) was given into brachial artery and deep vein samples obtained every 12-30 s for 15 min. Insulin-stimulated forearm glucose uptake (FGU) and whole body glucose metabolism (M) were reduced by 40-50% in obese nondiabetic, lean T2DM, and obese T2DM subjects (all P < 0.01); in offspring, the reduction in FGU and M was approximately 30% (P < 0.05). Inward glucose transport and glucose phosphorylation were decreased by approximately 40-50% (P < 0.01) in obese nondiabetic and T2DM groups and closely paralleled the decrease in FGU. The intracellular glucose concentration in the space accessible to glucose was significantly greater in obese nondiabetic, lean T2DM, obese T2DM, and offspring compared with lean controls. We conclude that 1) obese nondiabetic, lean T2DM, and offspring manifest moderate-to-severe muscle insulin resistance (FGU and M) and decreased insulin-stimulated glucose transport and glucose phosphorylation in forearm muscle; these defects in insulin action are not further reduced by the combination of obesity plus T2DM; and 2) the increase in intracelullar glucose concentration under hyperinsulinemic euglycemic conditions in obese and T2DM groups suggests that the defect in glucose phosphorylation exceeds the defect in glucose transport.


Translocation to a fragmented landscape: survival, movement, and site fidelity of Northern Bobwhites.
Habitat fragmentation, degradation, and loss have taxed early-successional species including the Northern Bobwhite (Colinus virginianus) and numerous grassland obligate birds. Translocation is often applied to counteract the consequences of habitat fragmentation through the creation, reestablishment, or augmentation of wild populations for the purposes of conservation, biodiversity maintenance. However, the implementation of these techniques is often conducted without valid experimental designs and therefore lacks robust, empirical data needed to evaluate and advance the knowledge and application of translocation. Despite the increasing amount of habitat management applied to patches among fragmented landscapes, a paucity of source populations often limits natural (re)colonization. As such, translocation may serve as a surrogate to natural dispersal, but its efficacy among fragmented landscapes is uncertain. Few studies exist that have assessed site fidelity, movement, and survival of individuals following translocation among fragmented landscapes. Thus, we experimentally evaluated the efficacy of translocation using known-fate and multi-strata models to evaluate hypotheses of temporal, biological, and group effects on survival and movement of translocated and resident bobwhites. We did not detect differences in survival or movement between translocated and resident bobwhites, suggesting that movement of individuals to a fragmented habitat does not negatively influence these demographic attributes. Based on these data, we suggest that two site-specific criteria should be met prior to instituting translocation: habitat management should be conducted to ensure that quality habitat exists and the patch size should be a minimum of 600 ha of quality habitat (poorer sites may warrant even larger patches). Translocation is a viable conservation method for increasing abundance in patches when habitat quality is high but source populations are limited.


Cooling-mediated protection from chemotherapy drug-induced cytotoxicity in human keratinocytes by inhibition of cellular drug uptake.
Chemotherapy-induced alopecia (CIA) represents the most distressing side-effect for cancer patients. Scalp cooling is currently the only treatment to combat CIA, yet little is known about its cytoprotective effects in human hair follicles (HF). We have previously established in vitro human keratinocyte models to study the effects of taxanes and anthracyclines routinely-used clinically and reported that cooling markedly-reduced or even completely-prevented cytotoxicity in a temperature dependent manner. Using these models (including HF-derived primary keratinocytes), we now demonstrate that cooling markedly attenuates cellular uptake of the anthracyclines doxorubicin and epirubicin to reduce or prevent drug-mediated human keratinocyte cytotoxicity. We show marked reduction in drug uptake and nuclear localization qualitatively by fluorescence microscopy. We have also devised a flow cytometry-based methodology that permitted semi-quantitative analysis of differences in drug uptake, which demonstrated that cooling can reduce drug uptake by up to ~8-fold in comparison to normal/physiological temperature, an effect that was temperature-dependent. Our results provide evidence that attenuation of cellular drug uptake represents at least one of the mechanisms underpinning the ability of cooling to rescue human keratinocytes from chemotherapy drug-cytotoxicity, thus supporting the clinical efficacy of scalp cooling.


Protein interaction networks--more than mere modules.
It is widely believed that the modular organization of cellular function is reflected in a modular structure of molecular networks. A common view is that a "module" in a network is a cohesively linked group of nodes, densely connected internally and sparsely interacting with the rest of the network. Many algorithms try to identify functional modules in protein-interaction networks (PIN) by searching for such cohesive groups of proteins. Here, we present an alternative approach independent of any prior definition of what actually constitutes a "module". In a self-consistent manner, proteins are grouped into "functional roles" if they interact in similar ways with other proteins according to their functional roles. Such grouping may well result in cohesive modules again, but only if the network structure actually supports this. We applied our method to the PIN from the Human Protein Reference Database (HPRD) and found that a representation of the network in terms of cohesive modules, at least on a global scale, does not optimally represent the network's structure because it focuses on finding independent groups of proteins. In contrast, a decomposition into functional roles is able to depict the structure much better as it also takes into account the interdependencies between roles and even allows groupings based on the absence of interactions between proteins in the same functional role. This, for example, is the case for transmembrane proteins, which could never be recognized as a cohesive group of nodes in a PIN. When mapping experimental methods onto the groups, we identified profound differences in the coverage suggesting that our method is able to capture experimental bias in the data, too. For example yeast-two-hybrid data were highly overrepresented in one particular group. Thus, there is more structure in protein-interaction networks than cohesive modules alone and we believe this finding can significantly improve automated function prediction algorithms.


A "novel" association to treat pain: tramadol/dexketoprofen. The first drug of a "new pharmacological class".
Acute and chronic pain have an important socio-economical impact. In order to help physicians to choose the appropriate drug, especially for cancer pain, in 1986 WHO has developed a three-step analgesic "ladder" for cancer pain relief in adults. Later it has also been used for acute pain and chronic non-cancer pain. In step I nonsteroidal anti-inflammatory drugs (NSAIDs) are considered with or without adjuvants, in step II the use of weak opioids for mild-moderate pain, with or without NSAIDs and adjuvant, is suggested, while the step III is reserved to strong opioids for moderate-severe pain with or without non-opioids or adjuvants. In the last two decades, a better pathophysiology knowledge has improved pain management shifting our view from the pain ladder to a modern pain pyramid, in which drugs are selected not only on the basis of pain intensity, but mainly according to mechanisms underlying pain, including peripheral and spinal sensitization which is the main trigger of chronic pain. The best pharmacological approach has become multimodal, in which drugs belonging to different steps should be combined, matching the mechanisms of action with the type of pain. An important corollary of combining analgesic drugs with different mechanism of action is that proper matching achieves the same effect with lower doses, better outcome and fewer adverse effects. In this new perspective, fixed-dose pharmaceutical combinations of different drugs are very useful to fulfil pharmacodynamics, pharmacokinetics and adherence criteria, enriching the pain pyramid of half-steps between the first and second step and between the second and third step. Hence, a new fixed combination of a NSAID with peripheral and central anti-infilammatory activities, such as dexketoprofen, and a weak opioid, such as tramadol, with double analgesic activity in the spinal cord as an opioid and, at the same time, on the descending modulatory pathways, is expected to cover a wide range of acute and recurrent painful conditions, ranging from nociceptive inflammatory pain to neuropathic pain of moderate/severe intensity. In this review we evaluate the rationale that justifies its use as new class of pharmacological modality to treat pain accordingly also to a more update view of WHO pain ladder.


De novo transcriptomes of 14 gammarid individuals for proteogenomic analysis of seven taxonomic groups.
Gammarids are amphipods found worldwide distributed in fresh and marine waters. They play an important role in aquatic ecosystems and are well established sentinel species in ecotoxicology. In this study, we sequenced the transcriptomes of a male individual and a female individual for seven different taxonomic groups belonging to the two genera Gammarus and Echinogammarus: Gammarus fossarum A, G. fossarum B, G. fossarum C, Gammarus wautieri, Gammarus pulex, Echinogammarus berilloni, and Echinogammarus marinus. These taxa were chosen to explore the molecular diversity of transcribed genes of genotyped individuals from these groups. Transcriptomes were de novo assembled and annotated. High-quality assembly was confirmed by BUSCO comparison against the Arthropod dataset. The 14 RNA-Seq-derived protein sequence databases proposed here will be a significant resource for proteogenomics studies of these ecotoxicologically relevant non-model organisms. These transcriptomes represent reliable reference sequences for whole-transcriptome and proteome studies on other gammarids, for primer design to clone specific genes or monitor their specific expression, and for analyses of molecular differences between gammarid species.


Comprehensive dietary evaluation of Italian primary school children: food consumption and intake of energy, nutrients and phenolic compounds.
Information on children's diet including bioactive compounds is quite scarce. This observational study investigated the composition of the diet of children living in Parma (Italy; n = 172, 8-10 years) using 3-day food records completed in winter and spring. Mean daily intakes of food groups, energy and nutrients were obtained using the national food database, while (poly)phenol contents were estimated from Phenol-Explorer or by specific literature searches. Food consumption, energy and nutrient intakes decreased in spring and were partially in line with national data. Adherence to the nutritional recommendations was not satisfied for the majority of nutrients. Main contributors to the phenolic intake were flavonoids (flavan-3-ols) and phenolic acids (hydroxycinnamic acids), while main dietary sources were fruit, chocolate-based products, vegetables, and tea & coffee (decaffeinated). This study provided the first comprehensive analysis of the nutritional composition of children's diet. Future research should look at the health implications of dietary choices in children.


Empiric potassium supplementation and increased survival in users of loop diuretics.
The effectiveness of the clinical strategy of empiric potassium supplementation in reducing the frequency of adverse clinical outcomes in patients receiving loop diuretics is unknown. We sought to examine the association between empiric potassium supplementation and 1) all-cause death and 2) outpatient-originating sudden cardiac death (SD) and ventricular arrhythmia (VA) among new starters of loop diuretics, stratified on initial loop diuretic dose. We conducted a one-to-one propensity score-matched cohort study using 1999-2007 US Medicaid claims from five states. Empiric potassium supplementation was defined as a potassium prescription on the day of or the day after the initial loop diuretic prescription. Death, the primary outcome, was ascertained from the Social Security Administration Death Master File; SD/VA, the secondary outcome, from incident, first-listed emergency department or principal inpatient SD/VA discharge diagnoses (positive predictive value = 85%). We identified 654,060 persons who met eligibility criteria and initiated therapy with a loop diuretic, 27% of whom received empiric potassium supplementation (N = 179,436) and 73% of whom did not (N = 474,624). The matched hazard ratio for empiric potassium supplementation was 0.93 (95% confidence interval, 0.89-0.98, p = 0.003) for all-cause death. Stratifying on initial furosemide dose, hazard ratios for empiric potassium supplementation with furosemide < 40 and ≥ 40 milligrams/day were 0.93 (0.86-1.00, p = 0.050) and 0.84 (0.79-0.89, p < 0.0001). The matched hazard ratio for empiric potassium supplementation was 1.02 (0.83-1.24, p = 0.879) for SD/VA. Empiric potassium supplementation upon initiation of a loop diuretic appears to be associated with improved survival, with a greater apparent benefit seen with higher diuretic dose. If confirmed, these findings support the use of empiric potassium supplementation upon initiation of a loop diuretic.


Prevalence of antibody to poliovirus in England and Wales 1984-6.
Serum from 995 subjects aged 6 months to 99 years was screened at a dilution of 1/8 for neutralising antibodies to poliovirus. Of these samples, 975 (98%) contained antibody to at least one serotype, and 763 (77%) contained antibody to all three, an improvement since previous studies. Children aged 8 to 15, however, had a low prevalence of antibody to poliovirus type 3, with only four (40%) of those aged 12 protected. This finding is possibly due to the waning of antibodies induced by the type 3 component of oral poliovirus vaccine and emphasises the continued importance of a booster dose of vaccine for those leaving school.


First Report of Ash Anthracnose Caused by Discula fraxinea in Oregon.
Anthracnose on ash trees has been observed on landscape trees in western Oregon, yet there has been no formal report of the disease or its causal fungus. Anthracnose symptoms are observed annually in May and become severe by July when defoliation starts to occur. From 1989 to now, samples have been received from Benton, Josephine, and Marion counties, suggesting that ash anthracnose has been present throughout western Oregon for some time. To identify the causal agent, a fungus was isolated from acervuli on necrotic lesions on leaves of cultivated white ash (Fraxinus americana L.) trees in Benton County in July 2003. The acervuli produced hyaline, nonseptate, ellipsoid conidia 5 to 11 × 3.5 to 6 µm in diameter. The fungus was identified as Discula fraxinea (Peck) Redlin & Stack (teleomorph Gnomoniella fraxini Redlin & Stack) (2). The sequences of the internal transcribed spacer (ITS) region (GenBank Accession No. AY455814) and large subunit (GenBank Accession No. AY455818) nrDNA agreed with those of D. fraxinea from Maryland, except for three single-base substitutions and three insertions/deletions in ITS1. Ash anthracnose has been reported from the central and eastern United States and California, the prairie provinces in Canada, and recently, from British Columbia (1). A specimen (U.S. National Fungus Collections BPI 843391) and culture (Centraal Bureau voor Schimmelcultures CBS 114053) of D. fraxinea from Oregon were deposited. References: (1) D. F. Farr et al. Fungal Databases. Systematic Botany and Mycology Laboratory. On-line publication. ARS USDA, 2003. (2) S. C. Redlin and R. W. Stack. Mycotaxon 32:175, 1988.


Adenoviral transgene delivery provides an approach to identifying important molecular processes in inflammation: evidence for heterogenecity in the requirement for NFkappaB in tumour necrosis factor production.
The success of anti-tumour necrosis factor (TNF) treatment, either using antibodies or soluble receptors, has defined TNF as a major factor of the inflammatory response in rheumatoid arthritis (RA). As a result of this success, much attention has been devoted to understanding the molecular mechanisms by which TNF expression and activity is elicited and controlled. By understanding these pathways, it is hoped that key elements of the molecular pathology associated with RA will be uncovered and, as a result, new targets for therapeutic intervention will be identified. However, studying the cell and molecular biology of model systems for RA, such as primary human macrophages, or tissue from rheumatoid joints may present technical problems. In an attempt to overcome this, we have investigated the use of adenovirus as a means of delivering transgenes by which different intracellular pathways can be modulated and examined. Our data show that adenovirus can be successfully used to efficiently deliver transgenes to primary human macrophages and RA joint tissue. Using a virus encoding IkappaBalpha, the natural inhibitor of NFkappaB, we show that the requirement for the transcription factor is not universal, but is dependent on the nature of the stimulus. Furthermore, while NFkappaB is of importance for the expression of TNF and other pro-inflammatory cytokines (for example, interleukin 6) and the destructive matrix metalloproteinases, this factor is not required for the expression of anti-inflammatory cytokines interleukin 10 and interleukin 1 receptor antagonist.


One-sample determination of glomerular filtration rate (GFR) in children. An evaluation based on 75 consecutive patients.
From a clinical point of view determination of glomerular filtration rate (clearance) is important. The aim of the present study was to compare the one-sample clearance to reference multiple-sample (51)Cr-EDTA clearance in consecutively referred children suspected of or with established nephro-urological disorders. A total of 75 children, age ½-13 years, received a quantitative intravenous injection of (51)Cr-EDTA followed by six plasma samples 10-120 min after injection. The multiple-sample clearance was measured as injected dose relative to the area under the plasma radioactivity curve. The one-sample clearance was determined from a single plasma sample collected at 60, 90 or 120 min after injection according to the one-pool method. The overall accuracy of one-sample clearance was excellent with mean numeric difference to the reference value of 0.7-1.7 mL/min. In 64 children, the one-sample clearance was within ± 4 mL/min of the multiple-sample value. However, in 11 children the numeric difference exceeded 4 mL/min (4.4-19.5). Analysis of age, body size, distribution volume, indicator retention time, clearance level, curve fitting, and sampling time could not explain the observed large difference between one-sample and multiple-sample clearance in these 15% of the children. In the majority of children there is an excellent agreement between one-sample clearance and multiple-sample clearance. However, unexpectedly in a small fraction (15%) larger discrepancies are found. If an accurate clearance value is essential a multiple-sample determination should be performed.


Effectiveness of decision aids for smoking cessation in adults: a quantitative systematic review.
The objective of the review was to determine the effectiveness of the use of decision aids to facilitate the shared decision-making process between a person and a healthcare provider on smoking cessation rates, knowledge of smoking cessation methods and decisional quality. The tobacco epidemic is one of the biggest health threats in the world, and it is the single most important preventable cause of disease. In the smoking cessation process, people move through different stages in their readiness to quit. Healthcare providers are well positioned to impact smoking cessation, however, challenges continue to exist. Using decision aids in the smoking cessation decision-making process may be one means in moving people toward the decision to quit smoking. The review considered randomized controlled trials and quasi-experimental studies which included adults, 18 years and over, who smoke tobacco products. The intervention of interest was implementation of decision aids to promote shared decision-making related to smoking cessation. The comparator was usual care without the use of decision aids. Outcomes of interest were smoking abstinence, smoking quit attempts, knowledge of smoking cessation methods and decisional quality. A comprehensive search was undertaken to locate relevant published and unpublished quantitative English language studies from the inception of each database searched through July 30, 2017. Databases searched included: CINAHL, Cochrane Central Register of Controlled Trials, Embase, PsycINFO and PubMed. A search of gray literature was also performed. Titles and abstracts were screened by two independent reviewers for assessment against the inclusion criteria. Studies meeting the inclusion criteria were retrieved in full and assessed in detail against the inclusion criteria by two independent reviewers. Two reviewers independently evaluated the included studies for methodological quality using critical appraisal tools from the Joanna Briggs Institute (JBI). Data related to the methods, participants, interventions and findings were extracted by two independent reviewers using a standardized data extraction tool from JBI. Due to clinical and methodological heterogeneity in the interventions and outcome measures of the included studies, statistical meta-analysis was not possible. Results have been presented in narrative form. Eight papers representing seven unique studies evaluating smoking cessation decision aids were included. Of the six studies that measured abstinence, only two had positive findings, with one reaching statistical significance. The fours studies that measured quit attempts all demonstrated a trend toward an increase in the number of quit attempts, with one study reaching statistical significance. Four of the five studies that measured knowledge reported an initial increase in the knowledge of smoking cessation methods, with two studies reaching statistical significance. Two of the three studies that measured decisional quality showed an improvement in decisional quality after use of the decision aid, with one study reaching statistical significance. The evidence suggests that decision aids may be effective in increasing smoking cessation knowledge and decisional quality while increasing quit attempts. There is, however, inconclusive evidence to support the definitive effectiveness of decision aids for long term and sustained smoking cessation.


Detection of Ebola viral antigen by enzyme-linked immunosorbent assay using a novel monoclonal antibody to nucleoprotein.
With the increase in international traffic, the risk of introducing rare but severe infectious diseases like Ebola hemorrhagic fever is increasing all over the world. However, the system for the diagnosis of Ebola virus infection is available in a limited number of countries. In the present study, we developed an Ebola virus antigen-detection enzyme-linked immunosorbent assay (ELISA) system using a novel monoclonal antibody (MAb) to the nucleoprotein (NP). This antibody recognized an epitope defined by a 26-amino-acid stretch near the C terminus of NP. In a sandwich ELISA system with the MAb, as little as 30 ng of purified recombinant NP (rNP) was detected. Although this MAb was prepared by immunization with rNP of subtype Zaire, it also reacted to the corresponding region of NP derived from the Reston and Sudan subtypes. These results suggest that our ELISA system should work with three of four Ebola subtypes. Furthermore, our ELISA system detected the NP in subtype Reston-infected monkey specimens, while the background level in noninfected specimens was very low, suggesting the usefulness of the ELISA for laboratory diagnosis with clinical specimens.


Clinical outcome after hypofractionated stereotactic radiotherapy (HSRT) for benign skull base tumors.
Surgical resection of skull base tumors can be associated with significant morbidity. In cases where the risks outweigh the benefits, radiation therapy can offer an alternative means to effectively control tumor growth. However, the optimal dose regime for radiation therapy remains controversial. The objective of this study was to assess the neurological outcome, local control rate and morbidity associated with a 5-fraction regime of hypofractionated stereotactic radiotherapy (HSRT) for benign skull base tumors. Twenty-six patients presenting with two of the most prevalent benign skull base tumors were included in the study. The tumors comprised 16 meningiomas and 10 acoustic neuromas. All patients exhibited preserved cranial nerve function prior to treatment, and a detailed audiological assessment was performed pre- and post-treatment for those patients with acoustic neuroma. Stereotactic radiosurgery was administered with the frameless CyberKnife Robotic Radiosurgery System. In each case, a 5-fraction HSRT regime was used: a dose of 5 Gy × 5 = 25 Gy to 6 Gy × 5 = 30 Gy was prescribed for skull base meningiomas, and 5 Gy × 5 = 25 Gy was prescribed for acoustic neuromas. The clinical and radiographic median follow-up was 22 months (range: 6-54 months). Radiological assessment showed local control in all 26 tumors (100%), and in 5/26 patients (20%) the tumor showed a decrease in size. Cranial nerve function was preserved in all cases thus far studied; however, 28% of patients had transient Grade II side effects, including fatigue, headaches, unsteadiness and transient subjective worsening of hearing. In two of these patients, the period of transient worsening of hearing was associated with a temporary increase in the size of the tumor on control T2 MR images, consistent with radiation-induced edema. One patient had transient decrease in visual acuity from treatment-related edema. At the last follow-up, 3/16 patients with meningiomas (19%) and 2/10 with acoustic neuromas (20%) showed a decrease in tumor volume and improvement in hearing. A 5-fraction stereotactic radiotherapy regime, as used in this study, seems to be effective for local control of benign skull base tumors in this early follow-up evaluation. Neurological function preservation is excellent with this short regime in the early post-treatment period, but long-term follow-up is crucial for validation.


Inference and quantification of peptidoforms in large sample cohorts by SWATH-MS.
Consistent detection and quantification of protein post-translational modifications (PTMs) across sample cohorts is a prerequisite for functional analysis of biological processes. Data-independent acquisition (DIA) is a bottom-up mass spectrometry approach that provides complete information on precursor and fragment ions. However, owing to the convoluted structure of DIA data sets, confident, systematic identification and quantification of peptidoforms has remained challenging. Here, we present inference of peptidoforms (IPF), a fully automated algorithm that uses spectral libraries to query, validate and quantify peptidoforms in DIA data sets. The method was developed on data acquired by the DIA method SWATH-MS and benchmarked using a synthetic phosphopeptide reference data set and phosphopeptide-enriched samples. IPF reduced false site-localization by more than sevenfold compared with previous approaches, while recovering 85.4% of the true signals. Using IPF, we quantified peptidoforms in DIA data acquired from >200 samples of blood plasma of a human twin cohort and assessed the contribution of heritable, environmental and longitudinal effects on their PTMs.


Multifunctional metal-organic frameworks: from academia to industrial applications.
After three decades of intense and fundamental research on metal-organic frameworks (MOFs), is there anything left to say or to explain? The synthesis and properties of MOFs have already been comprehensively described elsewhere. It is time, however, to prove the nature of their true usability: technological applications based on these extended materials require development and implementation as a natural consequence of the up-to-known intensive research focused on their design and preparation. The current large number of reviews on MOFs emphasizes practical strategies to develop novel networks with varied crystal size, shape and topology, being mainly devoted to academic concerns. The present survey intends to push the boundaries and summarise the state-of-the-art on the preparation of promising (multi)functional MOFs in worldwide laboratories and their use as materials for industrial implementation. This review starts, on the one hand, to describe several tools and striking examples of remarkable and recent (multi)functional MOFs exhibiting outstanding properties (e.g., in gas adsorption and separation, selective sorption of harmful compounds, heterogeneous catalysis, luminescent and corrosion protectants). On the other hand, and in a second part, it intends to use these examples of MOFs to incite scientists to move towards the transference of knowledge from the laboratories to the industry. Within this context, we exhaustively review the many efforts of several worldwide commercial companies to bring functional MOFs towards the daily use, analysing the various patents and applications reported to date. Overall, this review goes from the very basic concepts of functional MOF engineering and preparation ending up in their industrial production on a large scale and direct applications in society.


Antioxidants and viral infections: host immune response and viral pathogenicity.
Malnutrition has long been associated with increased susceptibility to infectious disease. The increase in severity from and susceptibility to infectious disease in malnourished hosts is thought to be the result of an impaired immune response. For example, malnutrition could influence the immune response by inducing a less effective ability to manage the challenge of an infectious disease. Work in our laboratory has demonstrated that not only is the host affected by the nutritional deficiency, but the invading pathogen is as well. Using a deficiency in selenium (Se) as a model system, mice deficient in Se were more susceptible to infection with coxsackievirus, as well as with influenza virus. Se-deficient mice develop myocarditis when infected with a normally benign strain of coxsackievirus. They also develop severe pneumonitis when infected with a mild strain of influenza virus. The immune system was altered in the Se-deficient animals, as was the viral pathogen itself. Sequencing of viral isolates recovered from Se-deficient mice demonstrated mutations in the viral genome of both coxsackievirus and influenza virus. These changes in the viral genome are associated with the increased pathogenesis of the virus. The antioxidant selenoenzyme, glutathione peroxidase-1, was found to be critically important, as glutathione peroxidase knockout mice developed myocarditis, similar to the Se-deficient mice, when infected with the benign strain of myocarditis. This work points to the importance of host nutrition in not only optimizing the host immune response, but also in preventing viral mutations which could increase the viral pathogenicity.


U S. Medical Eligibility Criteria for Contraceptive Use, 2010.
CDC created U.S. Medical Eligibility Criteria for Contraceptive Use, 2010, from guidance developed by the World Health Organization (WHO) and finalized the recommendations after consultation with a group of health professionals who met in Atlanta, Georgia, during February 2009. This guidance comprises recommendations for the use of specific contraceptive methods by women and men who have certain characteristics or medical conditions. The majority of the U.S. guidance does not differ from the WHO guidance and covers >60 characteristics or medical conditions. However, some WHO recommendations were modified for use in the United States, including recommendations about contraceptive use for women with venous thromboembolism, valvular heart disease, ovarian cancer, and uterine fibroids and for postpartum and breastfeeding women. Recommendations were added to the U.S. guidance for women with rheumatoid arthritis, history of bariatric surgery, peripartum cardiomyopathy, endometrial hyperplasia, inflammatory bowel disease, and solid organ transplantation. The recommendations in this document are intended to assist health-care providers when they counsel women, men, and couples about contraceptive method choice. Although these recommendations are meant to serve as a source of clinical guidance, health-care providers should always consider the individual clinical circumstances of each person seeking family planning services.


Rabies exposures and pre-exposure vaccination practices among individuals with an increased risk of rabies exposure in the United States.
OBJECTIVE To identify knowledge and practices related to rabies vaccination and serologic monitoring among animal care workers in the United States. DESIGN Cross-sectional survey. SAMPLE 2,334 animal care workers (ie, veterinarians, veterinary technicians, animal control workers, and wildlife rehabilitators). PROCEDURES Participants were contacted through relevant professional organizations to participate in an anonymous web-based survey. The survey collected demographic and occupational information, animal handling and potential rabies exposure information, and individual rabies vaccination and serologic monitoring practices. Comparisons of animal bite and rabies exposure rates were made between occupational groups. Multiple logistic regression was used to evaluate factors associated with rabies vaccination status and adherence to serologic monitoring recommendations. RESULTS Respondents reported 0.77 animal bites/person-year or 0.10 bites/1,000 animals handled. The overall rate of postexposure prophylaxis due to an occupational rabies exposure was 1.07/100 person-years. Veterinarians reported the highest rabies vaccination rate (98.7% [367/372]), followed by animal control workers (78.5% [344/438]), wildlife rehabilitators (78.2% [122/156]), and veterinary technicians (69.3% [937/1,352]). Respondents working for employers requiring rabies vaccination and serologic monitoring were 32.16 and 6.14 times, respectively, as likely to be vaccinated or have a current serologic monitoring status as were respondents working for employers without such policies. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested that, given the high reported rates of animal bites and potential rabies exposures among animal care workers, improvements in rabies vaccination and serologic monitoring practices are needed.


Urokinase type plasminogen activator receptor (uPAR) as a new therapeutic target in cancer.
The urokinase (uPA)-type plasminogen activator receptor (uPAR) is a GPI-anchored receptor that focuses urokinase (uPA) proteolytic activity on the cell surface. uPAR also regulates cell adhesion, migration and proliferation, protects from apoptosis and contributes to epithelial mesenchymal transition (EMT), independently of uPA enzymatic activity. Indeed, uPAR interacts with beta1, beta2 and beta3 integrins, thus regulating their activities. uPAR cross-talks with receptor tyrosine kinases through integrins and regulates cancer cell dormancy, proliferation and angiogenesis. Moreover, uPAR mediates uPA-dependent cell migration and chemotaxis induced by fMet-Leu-Phe (fMLF), through its association with fMLF-receptors (fMLF-Rs). Further, uPAR is an adhesion receptor because it binds vitronectin (VN), a component of provisional extracellular matrix. High uPAR expression predicts for more aggressive disease in several cancer types for its ability to increase invasion and metastasis. In fact, uPAR has been hypothesized to be the link between tumor cell dormancy and proliferation that usually precedes the onset of metastasis. Thus, inhibiting uPAR could be a feasible approach to affect tumor growth and metastasis. Here, we review the more recent advances in the development of uPAR-targeted anti-cancer therapeutic agents suitable for further optimization or ready for the evaluation in early clinical trials.


Evolutionary history, structural features and biochemical diversity of the NlpC/P60 superfamily of enzymes.
Peptidoglycan is hydrolyzed by a diverse set of enzymes during bacterial growth, development and cell division. The N1pC/P60 proteins define a family of cell-wall peptidases that are widely represented in various bacterial lineages. Currently characterized members are known to hydrolyze D-gamma-glutamyl-meso-diaminopimelate or N-acetylmuramate-L-alanine linkages. Detailed analysis of the N1pC/P60 peptidases showed that these proteins define a large superfamily encompassing several diverse groups of proteins. In addition to the well characterized P60-like proteins, this superfamily includes the AcmB/LytN and YaeF/YiiX families of bacterial proteins, the amidase domain of bacterial and kinetoplastid glutathionylspermidine synthases (GSPSs), and several proteins from eukaryotes, phages, poxviruses, positive-strand RNA viruses, and certain archaea. The eukaryotic members include lecithin retinol acyltransferase (LRAT), nematode developmental regulator Egl-26, and candidate tumor suppressor H-rev107. These eukaryotic proteins, along with the bacterial YaeF/poxviral G6R family, show a circular permutation of the catalytic domain. We identified three conserved residues, namely a cysteine, a histidine and a polar residue, that are involved in the catalytic activities of this superfamily. Evolutionary analysis of this superfamily shows that it comprises four major families, with diverse domain architectures in each of them. Several related, but distinct, catalytic activities, such as murein degradation, acyl transfer and amide hydrolysis, have emerged in the N1pC/P60 superfamily. The three conserved catalytic residues of this superfamily are shown to be equivalent to the catalytic triad of the papain-like thiol peptidases. The predicted structural features indicate that the N1pC/P60 enzymes contain a fold similar to the papain-like peptidases, transglutaminases and arylamine acetyltransferases.


Does activation of the FcgammaRIIa play a role in the pathogenesis of the acute lung injury/acute respiratory distress syndrome?
ALI (acute lung injury) and its more severe form ARDS (acute respiratory distress syndrome) are inflammatory diseases of the lung characterized by hypoxaemia and diffuse bilateral infiltrates. Disruption of epithelial integrity and injury to endothelium are contributing factors of the development of ALI/ARDS, and alveolar damage is the most pronounced feature of ALI/ARDS. The resulting increase in lung microvascular permeability promotes influx of inflammatory cells to the alveolar spaces. Oedema fluid contains pro-nflammatory mediators and plasma proteins, including Igs (immunoglobulins). Moreover, several reports describe the presence of autoantibodies and immune complexes [anti-IL-8 (interleukin-8) autoantibody/IL-8 complexes] in lung fluids (oedema and bronchoalveolar lavage fluids) from patients with ALI/ARDS. These immune complexes associate with FcgammaRIIa (Fcgamma IIa receptor) in lungs of patients with ARDS. Furthermore, the expression of FcgammaRIIa is substantially elevated in lungs of these patients. FcgammaRIIa appears on virtually all myeloid cells, platelets and endothelial cells. It is a low-affinity receptor for IgG that preferentially binds aggregated immunoglobulins and immune complexes. FcgammaRs regulate phagocytosis and cell-mediated cytotoxicity, and initiate the release of inflammatory mediators. It should be noted that immune complexes formed between either anti-neutrophil autoantibodies and their specific antigens or anti-HLA (human leucocyte antigen) antibodies and target antigens are implicated in the pathogenesis of TRALI (transfusion-related acute lung injury), and importantly, animal studies indicate that FcgammaRs are essential for these complexes to cause damage to the lungs. Therefore, we hypothesize that FcgammaRs such as FcgammaRIIa could contribute to the pathogenesis of ALI/ARDS.


Target Displacements during Eye Blinks Trigger Automatic Recalibration of Gaze Direction.
Eye blinks cause disruptions to visual input and are accompanied by rotations of the eyeball [1]. Like every motor action, these eye movements are subject to noise and introduce instabilities in gaze direction across blinks [2]. Accumulating errors across repeated blinks would be debilitating for visual performance. Here, we show that the oculomotor system constantly recalibrates gaze direction during blinks to counteract gaze instability. Observers were instructed to fixate a visual target while gaze direction was recorded and blinks were detected in real time. With every spontaneous blink-while eyelids were closed-the target was displaced laterally by 0.5° (or 1.0°). Most observers reported being unaware of displacements during blinks. After adapting for ∼35 blinks, gaze positions after blinks showed significant biases toward the new target position. Automatic eye movements accompanied each blink, and an aftereffect persisted for a few blinks after target displacements were eliminated. No adaptive gaze shift occurred when blinks were simulated with shutter glasses at random time points or actively triggered by observers, or when target displacements were masked by a distracting stimulus. Visual signals during blinks are suppressed by inhibitory mechanisms [3-6], so that small changes across blinks are generally not noticed [7, 8]. Additionally, target displacements during blinks can trigger automatic gaze recalibration, similar to the well-known saccadic adaptation effect [9-11]. This novel mechanism might be specific to the maintenance of gaze direction across blinks or might depend on a more general oculomotor recalibration mechanism adapting gaze position during intrinsically generated disruptions to visual input.


Global burden of childhood diarrhea and interventions.
Diarrhea is a leading cause of morbidity and mortality among children under 5 years in low-income and middle-income countries. Over the past 2 decades under-five mortality has decreased substantially, but reductions have been uneven and unsatisfactory in resource-poor regions. There are known interventions which can prevent diarrhea or manage children who suffer from it. Interventions with proven effectiveness at the prevention level include water, sanitation, and hygiene interventions, breastfeeding, complementary feeding, vitamin A and zinc supplementation, and vaccines for diarrhea (rotavirus and cholera). Oral rehydration solution, zinc treatment, continued feeding, and antibiotic treatment for certain strains of diarrhea (cholera, Shigella, and cryptosporidiosis) are effective strategies for treatment of diarrhea. The recent Lancet series using the 'Lives Saved' tool suggested that if these identified interventions were scaled up to a global coverage to at least 80%, and immunizations to at least 90%; almost all deaths due to diarrhea could be averted. The current childhood mortality burden highlights the need of a focused global diarrhea action plan. The findings suggest that with proper packaging of interventions and delivery platforms, the burden of childhood diarrhea can be reduced to a greater extent. All that is required is greater attention and steps toward right direction.


Hydration status and physiological workload of UAE construction workers: A prospective longitudinal observational study.
The objective of the study was to investigate the physiological responses of construction workers labouring in thermally stressful environments in the UAE using Thermal Work Limit (TWL) as a method of environmental risk assessment. The study was undertaken in May 2006. Aural temperature, fluid intake, and urine specific gravity were recorded and continuous heart rate monitoring was used to assess fatigue. Subjects were monitored over 3 consecutive shifts. TWL and WBGT were used to assess the thermal stress. Most subjects commenced work euhydrated and maintained this status over a 12-hour shift. The average fluid intake was 5.44 L. There were no changes in core temperature or average heart rate between day 1 and day 3, nor between shift start and finish, despite substantial changes in thermal stress. The results obtained indicated that the workers were not physiologically challenged despite fluctuating harsh environmental conditions. Core body temperatures were not elevated suggesting satisfactory thermoregulation. The data demonstrate that people can work, without adverse physiological effects, in hot conditions if they are provided with the appropriate fluids and are allowed to self-pace. The findings suggested that workers will self-pace according to the conditions. The data also demonstrated that the use of WBGT (a widely used risk assessment tool) as a thermal index is inappropriate for use in Gulf conditions, however TWL was found to be a valuable tool in assessing thermal stress.


Role of Sirt3 in mitochondrial biogenesis and developmental competence of human in vitro matured oocytes.
Does Sirt3 dysfunction result in poor developmental outcomes for human oocytes after in vitro maturation (IVM)? Inefficient Sirt3 expression induced decreased mitochondrial DNA copy number and biogenesis, and therefore impaired the developmental competence of human IVM oocytes. Cytoplasmic immaturity in IVM oocytes may lead to reduced developmental competence. Mitochondrial dysfunction results in the accumulation of free radicals and leads to DNA mutations, protein damage, telomere shortening and apoptosis. SIRT3 (in the Sirtuin protein family) has emerged as a mitochondrial fidelity protein that directs energy generation and regulates reactive oxygen species scavenging proteins. In vivo matured metaphase II (IVO-MII) oocytes and IVM-MII oocytes were donated by 324 infertile patients undergoing assisted reproductive technology cycles (12 patients for 60 IVO oocytes, and 312 patients for 403 IVM oocytes). Five oocytes each in the germinal vesicle (GV), IVM and IVO groups were compared with respect to mRNA levels for Sirt1-7 mRNA, and five samples at each developmental stage were analysed for Sirt3 mRNA. IVM-MII oocytes were injected with in vitro transcribed mRNA (n = 59) or small interfering RNA (siRNA) (n = 78). In human and mouse, IVM, mRNA-injection IVM, and siRNA-injection IVM groups (n = 5 each) were analysed for mitochondrial DNA copy number and abundance of Sirt3 and Pgc1α (an inducer of mitochondrial biogenesis) mRNAs. Human blastocysts in the IVO (n = 12), IVM (n = 9), mRNA-injection IVM (n = 13) and siRNA-injection IVM (n = 6) groups were used to generate embryonic stem cells (ESCs). In addition, 587 IVO-MII and 1737 IVM-MII oocytes from 83 mice were collected to compare the preliminary human oocyte data with another species. mRNA abundance was analysed by single-cell real-time PCR. Karyotyping of human embryos was performed with an array comparative genomic hybridization method, and that of ESCs by cytogenetic analysis. The function of the Sirt3 gene was investigated using siRNA and in vitro transcribed mRNA injection. Markers of ESCs were identified using immunofluorescence. A retrospective analysis revealed a higher spontaneous abortion rate (P < 0.01) and decrease in high quality embryo rate (P < 0.01) in patients with IVM versus controlled ovarian stimulation (COS) cycles. A decrease in abundance of Sirt3 mRNA (P < 0.01) and mitochondrial biogenesis (P < 0.05) were identified in human IVM compared with IVO oocytes. The developmental potential of human IVM-MII oocytes to the blastocyst stage was significantly reduced when Sirt3 mRNA was inhibited by siRNA (P < 0.05 versus IVM-MII group) but could be up-regulated by injection of Sirt3 mRNAs. Compared with IVO-MII group, comparable generation efficiency of human ESCs can be obtained using blastocysts from IVM-MII oocytes with Sirt3 mRNA injection. Sirt3 mRNA was significantly increased in mouse zygotes after IVF (P < 0.001 versus MII oocytes) but gradually declined until the blastocyst stage. In mice, lower Sirt3 mRNA levels were observed IVM-MII oocytes and preimplantation embryos compared with in vivo controls, and mitochondrial biogenesis and the developmental efficiency from oocytes to blastocyst were affected by the abundance of Sirt3 mRNA in accordance with human. Therefore a similar role for Sirt3 mRNA in IVM-MII oocytes was observed in mouse and human. The couples in the study had a variety of different simple and complex factors causing infertility. Additional studies with a larger number of oocytes are required to confirm the present results owing to the limited number of human oocytes in the present study. To our knowledge, this is the first study investigating a role of the Sirt3 gene in mitochondrial biogenesis and the developmental competence of human IVM-MII oocytes. The observation may help to improve clinical application of the IVM procedure. This work was supported in part by the National Natural Science Foundation of Key Program (31230047), Ministry of Science and Technology of China Grants (973 program; 2014CB943203), the National Natural Science Foundation of General Program (31371521 and 81571400), Beijing Nova Program (xxjh2015011), and Specialized Research Fund for the Doctoral Program of Higher Education (20120001130008) and the National Natural Science Foundation of Young Scholar (31501201). The authors have declared that no conflict of interest exists.


Consumption of long-chain n-3 PUFA, α-linolenic acid and fish is associated with the prevalence of chronic kidney disease.
Due to the anti-inflammatory properties of PUFA, it has been suggested that they may protect against kidney damage in adults. However, relatively few epidemiological studies have examined this hypothesis in human subjects. We investigated the association between dietary intakes of PUFA (n-3, n-6 and α-linolenic acid), fish and the prevalence of chronic kidney disease (CKD). A total of 2600 Blue Mountains Eye Study (1997-9) participants aged ≥ 50 years were analysed. Dietary data were collected using a semi-quantitative FFQ, and PUFA and fish intakes were calculated. Baseline biochemistry including serum creatinine was measured. Moderate CKD was defined as an estimated glomerular filtration rate of < 60 ml/min per 1·73 m2. Participants in the highest quartile of long-chain n-3 PUFA intake had a significantly reduced likelihood of having CKD compared with those in the lowest quartile of intake (multivariable-adjusted OR 0·69, 95 % CI 0·49, 0·99). α-Linolenic acid intake was positively associated with CKD (OR, per standard deviation increase in α-linolenic acid, 1·18, 95 % CI 1·05, 1·32). Total n-3 PUFA or total n-6 PUFA were not significantly associated with CKD. The highest compared with the lowest quartile of fish consumption was associated with a reduced likelihood of CKD (OR 0·68, 95 % CI 0·48, 0·97; P for trend = 0·02). The present study shows that an increased dietary intake of long-chain n-3 PUFA and fish reduces the prevalence of CKD. Hence, a diet rich in n-3 PUFA and fish could have a role in maintaining healthy kidney function, in addition to roles of these nutrients in the prevention and modulation of other diseases.


Determining liver stage parasite burden by real time quantitative PCR as a method for evaluating pre-erythrocytic malaria vaccine efficacy.
The detection and quantitation of blood stage parasitaemia is typically used as a surrogate endpoint for estimating the efficacy of vaccines targeted against the hepatic stage, as well as the erythrocytic stage, of the parasite. However, this does not provide an adequate means of evaluating the efficacy of vaccines, which may be only partially effective at the liver-stage. This is a particular concern for effective evaluation of immune enhancement strategies for candidate pre-erythrocytic stage vaccines. Here, we have developed and validated a method for detecting and quantitating liver stage parasites, using the TaqMan fluorescent real-time quantitative PCR system (PE Applied Biosystems). This method uses TaqMan primers designed to the Plasmodium yoelii 18S rRNA gene and rodent GAPDH to amplify products from infected mouse liver cDNA. The technique is highly reproducible as demonstrated with plasmid controls and capable of efficiently quantitating liver-stage parasite burden following a range of sporozoite challenge doses in strains of mice, which differ in their susceptibility to sporozoite infection. We have further demonstrated the capacity of this technique to evaluate the efficacy of a range of pre-erythrocytic stage vaccines. Our data establish this quantitative real-time PCR assay to be a fast and reproducible way of accurately assessing liver stage parasite burden and vaccine efficacy in rodent malaria models.


Effects of inducer continuity on auditory stream segregation: comparison of physical and perceived continuity in different contexts.
The factors influencing the stream segregation of discrete tones and the perceived continuity of discrete tones as continuing through an interrupting masker are well understood as separate phenomena. Two experiments tested whether perceived continuity can influence the build-up of stream segregation by manipulating the perception of continuity during an induction sequence and measuring streaming in a subsequent test sequence comprising three triplets of low and high frequency tones (LHL-[ellipsis (horizontal)]). For experiment 1, a 1.2-s standard induction sequence comprising six 100-ms L-tones strongly promoted segregation, whereas a single extended L-inducer (1.1 s plus 100-ms silence) did not. Segregation was similar to that following the single extended inducer when perceived continuity was evoked by inserting noise bursts between the individual tones. Reported segregation increased when the noise level was reduced such that perceived continuity no longer occurred. Experiment 2 presented a 1.3-s continuous inducer created by bridging the 100-ms silence between an extended L-inducer and the first test-sequence tone. This configuration strongly promoted segregation. Segregation was also increased by filling the silence after the extended inducer with noise, such that it was perceived like a bridging inducer. Like physical continuity, perceived continuity can promote or reduce test-sequence streaming, depending on stimulus context.


Genetic regulation of nitrogen metabolism in the fungi.
In the fungi, nitrogen metabolism is controlled by a complex genetic regulatory circuit which ensures the preferential use of primary nitrogen sources and also confers the ability to use many different secondary nitrogen sources when appropriate. Most structural genes encoding nitrogen catabolic enzymes are subject to nitrogen catabolite repression, mediated by positive-acting transcription factors of the GATA family of proteins. However, certain GATA family members, such as the yeast DAL80 factor, act negatively to repress gene expression. Selective expression of the genes which encode enzymes for the metabolism of secondary nitrogen sources is often achieved by induction, mediated by pathway-specific factors, many of which have a GAL4-like C6/Zn2 DNA binding domain. Regulation within the nitrogen circuit also involves specific protein-protein interactions, as exemplified by the specific binding of the negative-acting NMR protein with the positive-acting NIT2 protein of Neurospora crassa. Nitrogen metabolic regulation appears to play a significant role in the pathogenicity of certain animal and plant fungal pathogens.


Transcending scale dependence in identifying habitat with resource selection functions.
Multi-scale resource selection modeling is used to identify factors that limit species distributions across scales of space and time. This multi-scale nature of habitat suitability complicates the translation of inferences to single, spatial depictions of habitat required for conservation of species. We estimated resource selection functions (RSFs) across three scales for a threatened ungulate, woodland caribou (Rangifer tarandus caribou), with two objectives: (1) to infer the relative effects of two forms of anthropogenic disturbance (forestry and linear features) on woodland caribou distributions at multiple scales and (2) to estimate scale-integrated resource selection functions (SRSFs) that synthesize results across scales for management-oriented habitat suitability mapping. We found a previously undocumented scale-specific switch in woodland caribou response to two forms of anthropogenic disturbance. Caribou avoided forestry cut-blocks at broad scales according to first- and second-order RSFs and avoided linear features at fine scales according to third-order RSFs, corroborating predictions developed according to predator-mediated effects of each disturbance type. Additionally, a single SRSF validated as well as each of three single-scale RSFs when estimating habitat suitability across three different spatial scales of prediction. We demonstrate that a single SRSF can be applied to predict relative habitat suitability at both local and landscape scales in support of critical habitat identification and species recovery.


The epithelial barrier-protecting properties of a soy hydrolysate.
Enhancing the epithelial barrier function could be a possible strategy to prevent food allergy or reduce its symptoms. Soy hydrolysates containing bioactive peptides could be instrumental in this. In this study, the protective effects of pretreatment with 6 soy hydrolysates on calcium ionophore A23187-induced TEER reduction were studied in T84 cells. The effects of the most potent soy hydrolysate on tight junction gene expression were studied. In order to identify the underlying pathways involved, the barrier disruptor specificity of the effect was studied by comparing the protective effects on TEER and Lucifer Yellow flux after the exposure to barrier disruptors that work via different intracellular pathways, i.e. the disruptors A23187, mellitin, and deoxynivalenol (DON). Preincubation with one of the six hydrolysates protected the epithelial cells from a decrease in TEER induced by A23187 (restored to 105% of the starting point, while A23187 alone decreased to 53% of the starting value) and mellitin (restored to 11% of the starting point, while mellitin alone decreased to 3.8% of the starting value). This soy hydrolysate was found to increase claudin-1 and decrease claudin-2 expression. The protective effect of the hydrolysate on TEER was specific for the barrier disruptors A23187 and mellitin, but was not observed for DON. This observation suggests that the soy hydrolysate may act via PKC isoforms, which are known to lead to changes in the expression of claudin-1 and 2. Our data suggest that specific soy hydrolysates may be designed to strengthen the epithelial barrier which might be instrumental in the management of the barrier function in individuals at risk of developing food allergy.


Chlorpyrifos and chlorpyrifos oxon impair the transport of membrane bound organelles in rat cortical axons.
Chlorpyrifos (CPF) is an extensively used organophosphorus pesticide that has recently come under increasing scrutiny due to environmental health concerns particularly its association with neurodevelopmental defects. While the insecticidal actions and acute toxicity of CPF are attributed to its oxon metabolite (CPO) which potently inhibits the cholinergic enzyme acetylcholinesterase (AChE), there is significant evidence that CPF, CPO, and other organophosphates may affect a variety of neuronal targets and processes that are not directly related to AChE. Previously, in adult rat sciatic nerves ex vivo and postnatal neurons from rats in vitro we observed that CPF and CPO impaired the movements of vesicles and mitochondria in axons. Here, in embryonic neurons from rats in culture, we evaluated 24h exposures to CPF and CPO across picomolar to micromolar concentrations for effects on fast axonal transport of membrane bound organelles (MBOs) that contained the amyloid precursor protein (APP) tagged with the fluorescent marker, Dendra2 (APPDendra2). The most notable observations of this study were concentration-dependent decreases in the velocity and percentage of MBOs moving in the anterograde direction, an increase in the number of stationary MBOs, and an increased frequency of pauses associated with both CPF and CPO. These effects occurred at concentrations that did not significantly inhibit AChE activity, they were not blocked by cholinergic receptor antagonists, and they were not associated with compromised cell viability. These effects of CPF and CPO may be significant given the importance of axonal transport to neuronal development as well the function of fully developed neurons.


Activation of transforming growth factor-β/Smad signaling reduces aggregate formation of mislocalized TAR DNA-binding protein-43.
TAR DNA-binding protein of 43 kDa (TDP-43) is naturally located in the nucleus and has been identified as the major component of cytoplasmic ubiquitinated inclusions in patients with amyotrophic lateral sclerosis (ALS). We have reported that TDP-43 and phosphorylated Smad2 (pSmad2), an intracellular mediator protein of transforming growth factor-β (TGFβ) signaling, are co-localized within cytoplasmic inclusions in the anterior horn cells of sporadic ALS patients. To investigate the possible pathophysiological linkage between pathologic cytoplasmic inclusions containing TDP-43 and TGFβ/Smad signaling. We replicated cytoplasmic aggregates of TDP-43 in HEK293T cells by transfecting the cells with a nuclear localization signal deletion mutant of TDP-43 and inhibiting proteasome activity, and assessed the effect of TGFβ/Smad signaling on the cytoplasmic aggregate formation. The aggregates contained ubiquitinated, phosphorylated, and fragmented TDP-43, consistent with the essential features of the human pathology. Moreover, the aggregates were co-localized with pSmad2 under continuous TGFβ stimulation. Overexpression of Smad2 reduced the amount of cytoplasmic aggregates in HEK293T cells, and TGFβ stimulation augmented this reduction effect in a dose-dependent manner. Activation of the TGFβ/Smad signaling system is protective against aggregate formation of cytoplasmically mislocalized TDP-43 and may be a potential therapeutic approach to delay progression of ALS.


An alphabeta T-cell-independent immunoprotective response towards gut coccidia is supported by gammadelta cells.
Although gammadelta cells are commonly hypothesized to provide a 'first line of defence', gammadelta-cell-deficient mice are generally only marginally more susceptible to pathogens. Because gammadelta cells are enriched within epithelia, it is important to resolve whether immunoprotective capacity towards epithelial-tropic pathogens is absent from the gammadelta-cell compartment, or whether such activity is present but simply redundant with that of alphabeta T cells. In this work, following infection of the intestinal epithelium of alphabeta T-cell-deficient mice with the coccidian parasite, Eimeria vermiformis, gammadelta cells were shown to support the rapid activation of other lymphoid cells and to confer a transferable antipathogen effect that could be eradicated by neutralization of interferon-gamma. However, unlike alphabeta T cells, these effects of gammadelta cells showed no evidence of functional immunological memory. These results are directly relevant to coccidiosis, an economically significant disease of livestock, and should have general relevance to infections involving alphabeta T-cell deficiencies, e.g. cryptosporidiosis in patients with acquired immune deficiency syndrome (AIDS).


Molecular Basis for the Potent Inhibition of the Emerging Carbapenemase VCC-1 by Avibactam.
In 2016, we identified a new class A carbapenemase, VCC-1, in a nontoxigenic Vibrio cholerae strain that had been isolated from retail shrimp imported into Canada for human consumption. Shortly thereafter, seven additional VCC-1-producing V. cholerae isolates were recovered along the German coastline. These isolates appear to have acquired the VCC-1 gene (blaVCC-1) independently from the Canadian isolate, suggesting that blaVCC-1 is mobile and widely distributed. VCC-1 hydrolyzes penicillins, cephalothin, aztreonam, and carbapenems and, like the broadly disseminated class A carbapenemase KPC-2, is only weakly inhibited by clavulanic acid or tazobactam. Although VCC-1 has yet to be observed in the clinic, its encroachment into aquaculture and other areas with human activity suggests that the enzyme may be emerging as a public health threat. To preemptively address this threat, we examined the structural and functional biology of VCC-1 against the FDA-approved non-β-lactam-based inhibitor avibactam. We found that avibactam restored the in vitro sensitivity of V. cholerae to meropenem, imipenem, and ertapenem. The acylation efficiency was lower for VCC-1 than for KPC-2 and akin to that of Pseudomonas aeruginosa PAO1 AmpC (k2/Ki = 3.0 × 103 M-1 s-1). The tertiary structure of VCC-1 is similar to that of KPC-2, and they bind avibactam similarly; however, our analyses suggest that VCC-1 may be unable to degrade avibactam, as has been found for KPC-2. Based on our prior genomics-based surveillance, we were able to target VCC-1 for detailed molecular studies to gain early insights that could be used to combat this carbapenemase in the future.


Relationship between price paid for off-trade alcohol, alcohol consumption and income in England: a cross-sectional survey.
In order to examine the potential impact of an increase in the minimum price per unit of alcohol to 50 pence ($0.78), we examined drinking patterns and household incomes of people who purchase alcohol in England at above and below this price. Cross-sectional survey of 515 members of the public in seven towns and cities in the south of England. The primary outcome was whether the participant had purchased alcohol at <50 p/unit. The main exposures were annual household income and alcohol consumption, measured using the Alcohol Use Disorders Identification Test-Consumption (AUDIT-C). The median price paid per unit of alcohol was 53.1 pence (range 16.4-297.0 pence). Those buying alcohol at <50 p/unit had a mean AUDIT-C score of 6.2 compared with 5.5 among those buying alcohol at above this price. The odds ratio (OR) of a person on low income with high-risk drinking purchasing alcohol at <50 p/unit was 1.29 [95% confidence interval (CI) = 0.82-1.79] compared with all other study participants. The OR of a person on low income with low-risk drinking purchasing alcohol below this price was 0.51 (95% CI = 0.30-0.87) compared with all other participants. These data suggest that an increase in the minimum price of alcohol to 50 pence price per unit is only likely to disproportionately affect people on low incomes if their alcohol consumption is excessive.


The spectrum of ocular alterations in patients with β-thalassemia syndromes suggests a pathology similar to pseudoxanthoma elasticum.
To determine the prevalence and spectrum of ocular fundus abnormalities in patients with β-thalassemia and to investigate risk factors for their development. Cross-sectional, observational study. A total of 255 patients with β-thalassemia major (TM) and β-thalassemia intermedia (TI) were consecutively recruited and investigated. Patients underwent best correct visual acuity, indirect ophthalmoscopy, and fundus photography, including fundus autofluorescence (FAF) and near-infrared reflectance imaging using a confocal scanning laser ophthalmoscope (cSLO). Hematologic parameters were determined, including mean ferritin levels, aspartate amino transferase, alanine amino transferase, calcium, pre-transfusion hemoglobin, history of splenectomy, and liver iron concentration. Factors associated with the ocular phenotype were assessed using logistic regression. Ocular phenotype as determined by clinical examination and used multimodal imaging. A total of 153 patients (60.0%) affected by TM and 102 patients (40.0%) affected by TI participated, of whom 216 (84.7%) were receiving iron-chelating therapy. Ocular fundus abnormalities characteristic of pseudoxanthoma elasticum (PXE) were detected by cSLO in 70 of 255 patients (27.8%) and included peau d'orange (19.6%), angioid streaks (12.9%), pattern dystrophy-like changes (7.5%), and optic disc drusen (2.0%). Pseudoxanthoma elasticum-like changes were more frequent in patients with TI (P<0.001). Patients with PXE-like fundus changes were older than patients without these fundus changes (P<0.001). In both patients with TI and TM, age (P = 0.001) and splenectomy (P = 0.001) had the strongest association with presence of PXE-like fundus changes in multivariate analyses. A total of 43 of 255 patients (16.9%) showed increased retinal vascular tortuosity independently of the PXE-like fundus changes, which was associated with aspartate amino transferase (P = 0.036), hemoglobin (P = 0.008), and ferritin levels (P = 0.005). Pseudoxanthoma elasticum-like fundus changes are a frequent finding in patients with β-thalassemia. In TI, these changes increase with duration or severity of the disease. This particular ocular phenotype suggests an ocular pathology similar to PXE. Retinal vascular tortuosity may be an additional disease manifestation independent of the PXE-like syndrome. Patients with long-standing disease requiring iron-chelating treatment and a history of splenectomy need regular ophthalmic checkups because they are at risk of developing PXE-like fundus changes and potentially of subsequent choroidal neovascularization.


Telephone counselling by nurses in Norwegian primary care out-of-hours services: a cross-sectional study.
The primary care out-of-hours (OOH) services in Norway are characterized by high contact rates by telephone. The telephone contacts are handled by local emergency medical communication centres (LEMCs), mainly staffed by registered nurses. When assessment by a medical doctor is not required, the nurse often handles the contact solely by nurse telephone counselling. Little is known about this group of contacts. Thus, the aim of this study was to investigate characteristics of encounters with the OOH services that are handled solely by nurse telephone counselling. Nurses recorded ICPC-2 reason for encounter (RFE) codes and patient characteristics of all patients who contacted six primary care OOH services in Norway during 2014. Descriptive statistics and frequency analyses were applied. Of all telephone contacts (n = 61,441), 23% were handled solely by nurse counselling. Fever was the RFE most frequently handled (7.3% of all nurse advice), followed by abdominal pain, cough, ear pain and general symptoms. Among the youngest patients, 32% of the total telephone contacts were resolved by nurse advice compared with 17% in the oldest age group. At night, 31% of the total telephone contacts were resolved solely by nurse advice compared with 21% during the day shift and 23% in the evening. The share of nurse advice was higher on weekdays compared to weekends (mean share 25% versus 20% respectively). This study shows that nurses make a significant contribution to patient management in the Norwegian OOH services. The findings indicate which conditions nurses should be able to handle by telephone, which has implications for training and routines in the LEMCs. There is the potential for more nurse involvement in several of the RFEs with a currently low share of nurse counselling.