Subjective health of undocumented migrants in Germany - a mixed methods approach.
Health of migrants is known to be above-average in the beginning of the migration trajectory. At the same time reports from non-government organisations (NGOs) suggest that undocumented migrants in Germany tend to present late and in poor health at healthcare facilities. In this paper, we explore the health status of undocumented migrants with a mixed method approach including complementary qualitative and quantitative datasets. Undocumented migrants attending a NGO based in Hamburg, Germany, were asked to fill in the SF-12v2, a standardized questionnaire measuring health-related quality of life (HRQOL). The SF-12v2 was analyzed in comparison to the U.S. American norm sample and a representative German sample. Differences in mean scores for HRQOL were evaluated with a t-test and with a generalized linear model analyzing the impact of living without legal status on HRQOL. The quantitative research was complemented by a qualitative ethnographic study on undocumented migration and health in Berlin, Germany. The study included semi-structured interviews, informal conversations and participant observation with Latin American migrants over the course of three years. The study focused on subjective experiences of illness and health and the impact of illegality on migrants' health and access to health care. HRQOL was significantly worse in the sample of undocumented migrants (n = 96) as compared to the U.S. American sample (p < 0.005). Living without legal status displayed a significant negative effect on subjective mental and physical health (p ≤ 0.003) in the generalized linear model when adjusted for age and gender compared to the representative German population sample. The ethnographic study, which included 35 migrants, identified socio-economic conditions, the subjective experiences of criminalization, and late presentation at healthcare-facilities as the three main factors impacting on health from migrant perspective. The present research suggests a high morbidity and mortality in this comparatively young population. The ethnographic research confirms negative impacts on health of social determinants in general and stress associated with living without legal status in particular, both are further aggravated by exclusion from health care services. In addition to the provision of health care it appears to be important to structurally tackle the underlying social conditions which affect undocumented migrants' health. Living without legal status has a negative impact on health and well-being. Limited access to care may further exacerbate physical and mental illness. Possibilities to claim basic rights and protection as well as access to care without legal status appear to be important measures to improve health and well-being.


A Bayesian Noisy Logic Model for Inference of Transcription Factor Activity from Single Cell and Bulk Transcriptomic Data.
The advent of high-throughput sequencing has made it possible to measure the expression of genes at relatively low cost. However, direct measurement of regulatory mechanisms, such as Transcription Factor (TF) activity is still not readily feasible in a high-throughput manner. Consequently, there is a need for computational approaches that can reliably estimate regulator activity from observable gene expression data. In this work, we present a noisy Boolean logic Bayesian model for TF activity inference from differential gene expression data and causal graphs. Our approach provides a flexible framework to incorporate biologically motivated TF-gene regulation logic models. Using simulations and controlled over-expression experiments in cell cultures, we demonstrate that our method can accurately identify TF activity. Moreover, we apply our method to bulk and single cell transcriptomics measurements to investigate transcriptional regulation of fibroblast phenotypic plasticity. Finally, to facilitate usage, we provide user-friendly software packages and a web-interface to query TF activity from user input differential gene expression data: https://umbibio.math.umb.edu/nlbayes/.


Changes in signaling pathways induced by vandetanib in a human medullary thyroid carcinoma model, as analyzed by reverse phase protein array.
Medullary thyroid carcinoma (MTC) is a rare tumor that is caused by activating mutations in the proto-oncogene RET. Vandetanib, a tyrosine-kinase inhibitor, has been recently approved to treat adult patients with metastatic MTC. The aim of this study was to investigate changes in signaling pathways induced by vandetanib treatment in preclinical MTC models, using the reverse-phase protein array method (RPPA). The human TT cell line was used to assess in vitro and in vivo activity of vandetanib. Protein extracts from TT cells or TT xenografted mice, treated by increasing concentrations of vandetanib for different periods of time, were probed with a set of 12 antibodies representing major signaling pathways, using RPPA. Results were validated using two distinct protein detection methods: Western immunoblotting and immunohistochemistry. Vandetanib displays antiproliferative and antiangiogenic activities and inhibits RET autophosphorylation. The MAPK and AKT pathways were the two major signaling pathways inhibited by vandetanib. Interestingly, phosphorylated levels of NFκB-p65 were significantly increased by vandetanib. Comparable results were obtained in both the in vitro and in vivo approaches, as well as for the protein detection methods. However, some discrepancies were observed between RPPA and Western immunoblotting, possibly due to lack of specificity of the primary antibodies used. Overall, our results confirmed the interest of RPPA for screening global changes induced in signaling pathways by kinase inhibitors. MAPK and AKT were identified as the main pathways involved in vandetanib response in MTC models. Our results also suggest alternative routes for controlling the disease, and provide a rationale for the development of therapeutic combinations based on the comprehensive identification of molecular events induced by inhibitors.


Temporal dynamics of remapping captured by peri-saccadic continuous motion.
Different attention and saccade control areas contribute to space constancy by remapping target activity onto their expected post-saccadic locations. To visualize this dynamic remapping, we used a technique developed by Honda (2006) where a probe moved vertically while participants made a saccade across the motion path. Observers do not report any large excursions of the trace at the time of the saccade that would correspond to the classical peri-saccadic mislocalization effect. Instead, they reported that the motion trace appeared to be broken into two separate segments with a shift of approximately one-fifth of the saccade amplitude representing an overcompensation of the expected retinal displacement caused by the saccade. To measure the timing of this break in the trace, we introduced a second, physical shift that was the same size but opposite in direction to the saccade-induced shift. The trace appeared continuous most frequently when the physical shift was introduced at the midpoint of the saccade, suggesting that the compensation is in place when the saccade lands. Moreover, this simple linear shift made the combined traces appear continuous and linear, with no curvature. In contrast, Honda (2006) had reported that the pre- and post-saccadic portion of the trace appeared aligned and that there was often a small, visible excursion of the trace at the time of the saccade. To compare our results more directly, we increased the contrast of our moving probe in a third experiment. Now some observers reported seeing a deviation in the motion path but the misalignment remained present. We conclude that the large deviations at the time of saccade are generally masked for a continuously moving target but that there is nevertheless a residual misalignment between pre- and post-saccadic coordinates of approximately 20% of the saccade amplitude that normally goes unnoticed.


Modelling of ciprofloxacin killing enhanced by hyperbaric oxygen treatment in Pseudomonas aeruginosa PAO1 biofilms.
In chronic lung infections by Pseudomonas aeruginosa (PA) the bacteria thrive in biofilm structures protected from the immune system of the host and from antibiotic treatment. Increasing evidence suggests that the susceptibility of the bacteria to antibiotic treatment can be significantly enhanced by hyperbaric oxygen treatment. The aim of this study is to simulate the effect of ciprofloxacin treatment in a PAO1 biofilm model with aggregates in agarose when combined with hyperbaric oxygen treatment. This is achieved in a reaction-diffusion model that describes the combined effect of ciprofloxacin diffusion, oxygen diffusion and depletion, bacterial growth and killing, and adaptation of the bacteria to ciprofloxacin. In the model, the oxygen diffusion and depletion use a set of parameters derived from experimental results presented in this work. The part of the model describing ciprofloxacin killing uses parameter values from the literature in combination with our estimates (Jacobs, et al., 2016; Grillon, et al., 2016). Micro-respirometry experiments were conducted to determine the oxygen consumption in the P. aeruginosa strain PAO1. The parameters were validated against existing data from an HBOT experiment by Kolpen et al. (2017). The complete oxygen model comprises a reaction-diffusion equation describing the oxygen consumption by using a Michaelis-Menten reaction term. The oxygen model performed well in predicting oxygen concentrations in both time and depth into the biofilm. At 2.8 bar pure oxygen pressure, HBOT increases the penetration depth of oxygen into the biofilm almost by a of factor 4 in agreement with the scaling that follows from the stationary balance between the consumption term and diffusion term. In the full reaction-diffusion model we see that hyperbaric oxygen treatment significantly increases the killing by ciprofloxacin in a PAO1 biofilm in alignment with the experimental results from Kolpen et al. (Kolpen, et al., 2017; Kolpen, et al. 2016). The enhanced killing, in turn, lowers the oxygen consumption in the outer layers of the biofilm, and leads to even deeper penetration of oxygen into the biofilm.


Recent advances of bioinspired functional materials with specific wettability: from nature and beyond nature.
Through 3.7 billion years of evolution and natural selection, plants and animals in nature have ingeniously fulfilled a broad range of fascinating functions to achieve optimized performance in responding and adapting to changes in the process of interacting with complex natural environments. It is clear that the hierarchically organized micro/nanostructures of the surfaces of living organisms decisively manage fascinating and amazing functions, regardless of the chemical components of their building blocks. This conclusion now allows us to elucidate the underlying mechanisms whereby these hierarchical structures have a great impact on the properties of the bulk material. In this review, we mainly focus on advances over the last three years in bioinspired multiscale functional materials with specific wettability. Starting from selected naturally occurring surfaces, manmade bioinspired surfaces with specific wettability are introduced, with an emphasis on the cooperation between structural characteristics and macroscopic properties, including lotus leaf-inspired superhydrophobic surfaces, fish scale-inspired superhydrophilic/underwater superoleophobic surfaces, springtail-inspired superoleophobic surfaces, and Nepenthes (pitcher plant)-inspired slippery liquid-infused porous surfaces (SLIPSs), as well as other multifunctional surfaces that combine specific wettability with mechanical properties, optical properties and the unidirectional transport of liquid droplets. Afterwards, various top-down and bottom-up fabrication techniques are presented, as well as emerging cutting-edge applications. Finally, our personal perspectives and conclusions with regard to the transfer of micro- and nanostructures to engineered materials are provided.


Evaluation of the orientation of the steepest meridian of regular astigmatism among highly myopic Egyptian patients seeking non-ablative surgical correction of the refractive error.
LASIK surgery is currently the preferred procedure to correct low to moderate myopia. The aim of this study was to determine the orientation of the steepest meridian of regular astigmatism in order to determine the relative incidence of vertical, horizontal, and oblique regular astigmatism among highly myopic Egyptian patients seeking non-ablative surgical correction of the refractive error. One hundred and one eyes of 68 highly myopic patients who were seeking refractive surgery were included in this consecutive case series study. The refractive errors were measured using an autorefractometer and confirmed by trial. We measured the uncorrected and best corrected visual acuity in Snellen lines. Keratometry, central corneal thickness, and anterior chamber depth also were measured. The cylinder power in diopters and the axis in degrees were reported. Astigmatism was graded as with the rule (i.e., vertical meridian steeper), against the rule (i.e., horizontal meridian steeper), and oblique astigmatism. The number and the percentage of eyes with the rule, against the rule, and oblique astigmatism were calculated, and the chi-squared test was performed to analyze the data. The spherical refractive error ranged from -6.5 to -24.5 diopters (-13.45 ± 4.60). The cylinder power (Cyl) ranged from -0.25 to -7.5 diopters (-2.23 ± 1.28). The uncorrected visual acuity (UCVA) in Snellen lines ranged from 0.01 - 0.1 (0.03 ± 0.02). The mean for best corrected visual acuity (BCVA) in Snellen lines was 0.40 (± 0.23). The steepest meridian was vertical (i.e., with-the-rule astigmatism) in 44 eyes (43.56%), horizontal (i.e., against-the-rule astigmatism) in 27 eyes (26.73%), and oblique (i.e., oblique astigmatism) in 30 eyes (29.70%). The incidence of with-the-rule astigmatism in patients with high myopia was found to be much lower than in previous studies for non-myopic patients, with a higher incidence for against-the-rule astigmatism and oblique astigmatism.


Safety evaluation for a cell-based immune support system in an ex vivo rat model of gram-negative sepsis.
Granulocyte dysfunction is a central component of immunodeficiency in septic patients. Granulocyte transfusions appear to be pathophysiologically useful; however, they cause unwanted side-effects in the lungs and other organs. This study evaluates the safety of an extracorporeal immune support system with granulocytic cells in a rat model of Gram-negative sepsis. Three groups of male CD rats received either saline (control group, I), a dose of Escherichia coli O7:K1 lethal to 90% of the animals (LD90) (septic group, II), or an LD90 dose of E. coli that was incubated with the human promyelocytic leukemia cell line (HL-60) (differentiated into the granulocytic direction) for 20 min prior to infusion (second septic group, III). The animals were observed for seven days. Pre-treatment with HL-60 cells resulted in no adverse effects in the group III animals. Significantly lower bacterial counts and endotoxin levels in the plasma were detected after 24 h as compared to group II (P < 0.05). Group III animals had better weight gain and more stable hemodynamics than group II animals (P < 0.01). Seven day survival was 0/8 in group II, 6/8 in group III, and 8/9 in group I (log-rank test: II-III: P < 0.001). The data suggest that extracorporeal use of granulocytes allows the therapeutic use of these cells while avoiding unwanted effects resulting from direct contact to internal organs.


Inhibition of serotonin synthesis: A novel therapeutic paradigm.
The rate-limiting enzyme in serotonin synthesis is tryptophan hydroxylase (TPH). There are two independent serotonin systems in the body characterized by two isoforms of TPH, TPH1 and TPH2. While TPH2 synthesizes serotonin in the brain, TPH1 is expressed in the gut and in other peripheral tissues and supplies platelets in the circulation with serotonin. This duality of the serotonin system is enforced by the blood-brain barrier which is impermeable for serotonin. In the brain serotonin acts as neurotransmitter and is a main target for the treatment of psychiatric disorders. In the periphery it is released by platelets at the site of activation and elicits numerous physiological effects. TPH1 deficient mice were shown to be protected from diverse diseases including hemostatic, inflammatory, fibrotic, gastrointestinal, and metabolic disorders and therefore serotonin synthesis inhibition emerged as a reasonable therapeutic paradigm. Recently the first TPH inhibitor, telotristat ethyl, came on the market for the treatment of carcinoid syndrome. This review summarizes the state of development and the therapeutic opportunities of such compounds.


Quantitative mechanical analysis of indentations on layered, soft elastic materials.
Atomic force microscopy (AFM) is becoming an increasingly popular method for studying cell mechanics, however the existing analysis tools for determining the elastic modulus from indentation experiments are unable to quantitatively account for mechanical heterogeneity commonly found in biological samples. In this work, we numerically calculated force-indentation curves onto two-layered elastic materials using an analytic model. We found that the effect of the underlying substrate can be quantitatively predicted by the mismatch in elastic moduli and the homogeneous-case contact radius relative to the layer height for all tested probe geometries. The effect is analogous to one-dimensional Hookean springs in series and was phenomenologically modeled to obtain an approximate closed-form equation for the indentation force onto a two-layered elastic material which is accurate for up to two orders of magnitude mismatch in Young's modulus when the contact radius is less than the layer height. We performed finite element analysis simulations to verify the model and AFM microindentation experiments and macroindentation experiments to demonstrate its ability to deconvolute the Young's modulus of each layer. The model can be broadly used to quantify and serve as a guideline for designing and interpreting indentation experiments into mechanically heterogeneous samples.


Vpx rescues HIV-1 transduction of dendritic cells from the antiviral state established by type 1 interferon.
Vpx is a virion-associated protein encoded by SIVSM, a lentivirus endemic to the West African sooty mangabey (Cercocebus atys). HIV-2 and SIVMAC, zoonoses resulting from SIVSM transmission to humans or Asian rhesus macaques (Macaca mulatta), also encode Vpx. In myeloid cells, Vpx promotes reverse transcription and transduction by these viruses. This activity correlates with Vpx binding to DCAF1 (VPRBP) and association with the DDB1/RBX1/CUL4A E3 ubiquitin ligase complex. When delivered experimentally to myeloid cells using VSV G-pseudotyped virus-like particles (VLPs), Vpx promotes reverse transcription of retroviruses that do not normally encode Vpx. Here we show that Vpx has the extraordinary ability to completely rescue HIV-1 transduction of human monocyte-derived dendritic cells (MDDCs) from the potent antiviral state established by prior treatment with exogenous type 1 interferon (IFN). The magnitude of rescue was up to 1,000-fold, depending on the blood donor, and was also observed after induction of endogenous IFN and IFN-stimulated genes (ISGs) by LPS, poly(I:C), or poly(dA:dT). The effect was relatively specific in that Vpx-associated suppression of soluble IFN-β production, of mRNA levels for ISGs, or of cell surface markers for MDDC differentiation, was not detected. Vpx did not rescue HIV-2 or SIVMAC transduction from the antiviral state, even in the presence of SIVMAC or HIV-2 VLPs bearing additional Vpx, or in the presence of HIV-1 VLPs bearing all accessory genes. In contrast to the effect of Vpx on transduction of untreated MDDCs, HIV-1 rescue from the antiviral state was not dependent upon Vpx interaction with DCAF1 or on the presence of DCAF1 within the MDDC target cells. Additionally, although Vpx increased the level of HIV-1 reverse transcripts in MDDCs to the same extent whether or not MDDCs were treated with IFN or LPS, Vpx rescued a block specific to the antiviral state that occurred after HIV-1 cDNA penetrated the nucleus. Vpx provides a tool for the characterization of a potent, new HIV-1 restriction activity, which acts in the nucleus of type 1 IFN-treated dendritic cells.


Minimizing risks and monitoring safety of an antenatal care intervention to mitigate domestic violence among young Indian women: The Dil Mil trial.
Domestic violence - physical, psychological, or sexual abuse perpetrated against women by one or more family members - is highly prevalent in India. However, relatively little research has been conducted on interventions with the potential to mitigate domestic violence and its adverse health consequences, and few resources exist to guide safety planning and monitoring in the context of intervention research. Dil Mil is a promising women's empowerment-based intervention developed in India that engages with young women (daughters-in-law) and their mothers-in-law to mitigate domestic violence and related adverse health outcomes. This paper describes the design of a randomized controlled trial of Dil Mil in Bengaluru, India, with a focus on strategies used to minimize study-related risks and monitor safety. A phase 2 randomized controlled trial using a parallel comparison of the Dil Mil intervention versus standard care will be implemented in three public primary health centers in Bengaluru. Young pregnant women in the first or second trimester of pregnancy will be recruited from antenatal services at study health centers and through community outreach. If eligible and willing, their mother-in-law will also be recruited. Once enrolled, dyads will participate in a baseline interview and then randomized either to the control arm and receive standard care or to the intervention arm and receive standard care plus the Dil Mil intervention. Additional evaluations will be conducted at 3 months and 6 months postpartum. Data will be analyzed to examine the feasibility and safety of the intervention and the effect of the intervention on intermediary outcomes (the empowerment of daughters-in-law and mothers-in-law), incidence of domestic violence among daughters-in-law, and health outcomes including perceived quality of life, psychosocial status and maternal and infant health outcomes. This study offers approaches that may help guide safety planning and monitoring in other domestic violence intervention trials in similar settings. Moreover, given the staggeringly high prevalence of domestic violence against young women in India (and indeed globally) and the dearth of data on effective interventions, this study is poised to make an important contribution to the evidence-base for domestic violence prevention. ClinicalTrials.gov Identifier: NCT01337778.


Locating macromolecular assemblies in cells by 2D template matching with cisTEM.
For a more complete understanding of molecular mechanisms, it is important to study macromolecules and their assemblies in the broader context of the cell. This context can be visualized at nanometer resolution in three dimensions (3D) using electron cryo-tomography, which requires tilt series to be recorded and computationally aligned, currently limiting throughput. Additionally, the high-resolution signal preserved in the raw tomograms is currently limited by a number of technical difficulties, leading to an increased false-positive detection rate when using 3D template matching to find molecular complexes in tomograms. We have recently described a 2D template matching approach that addresses these issues by including high-resolution signal preserved in single-tilt images. A current limitation of this approach is the high computational cost that limits throughput. We describe here a GPU-accelerated implementation of 2D template matching in the image processing software cisTEM that allows for easy scaling and improves the accessibility of this approach. We apply 2D template matching to identify ribosomes in images of frozen-hydrated Mycoplasma pneumoniae cells with high precision and sensitivity, demonstrating that this is a versatile tool for in situ visual proteomics and in situ structure determination. We benchmark the results with 3D template matching of tomograms acquired on identical sample locations and identify strengths and weaknesses of both techniques, which offer complementary information about target localization and identity.


Isolation and Investigation of Presumptive Murine Lacrimal Gland Stem Cells.
Aqueous tear deficiency due to lacrimal gland insufficiency is one of the major causes of dry eye disease. In severe cases, such as Sjogren's syndrome, Stevens-Johnson syndrome, or ocular cicatricial pemphigoid, therapy with artificial tears is often insufficient to relieve severe discomfort, prevent progressive ocular surface disease, or enable visual rehabilitation by corneal transplantation. Cell or organ generation from stem cells, resulting in tear-like secretion, presents an option as a suitable alternative treatment. To obtain deeper insights into lacrimal gland stem cells we analyzed murine lacrimal glands for markers of pluripotency, self-renewal, and differentiation. A special, patented technique with mechanical and enzymatic digestion was used to generate high numbers of cells in vitro from murine lacrimal glands. These presumptive "murine lacrimal gland stem cells" ("mLGSCs") can be propagated as monolayer cultures over multiple passages. By means of RT-PCR, Western blot, and immunohistochemistry, markers of pluripotency and differentiation were demonstrated. Hanging drop culture was used to build organoid bodies from mLGSCs to investigate their spontaneous differentiation in three-dimensional culture with histology, immunohistochemistry, and transmission electron microscopy methods. Isolated mLGSCs were cultured over more than 65 passages. Murine lacrimal gland stem cells expressed markers of pluripotency such as Nanog, Sox2, Kruppel-like factor 4 (Klf4), as well as early-lineage markers of all three germ layers. Three-dimensional culture of these cells revealed their ability to differentiate into various cell types. Our results suggest that mLGSCs were isolated and cultured successfully. These cells have the ability to differentiate into all three germ layers. The results provide further insights into lacrimal gland stem cell physiology for engineering of a lacrimal gland construct to treat severe cases of tear deficiency in the future.


Redisplacement after closed reduction of forearm fractures in children.
Reangulation and displacement after closed reduction of pediatric forearm fractures were reviewed in 90 patients treated with 1978-1984. All fractures were remanipulated and followed to union. No complications occurred and all had a functionally satisfactory result. Both intrinsic and extrinsic factors were identified as contributing to the complication of reangulation/displacement. Nonepiphyseal fractures were safely remanipulated up to 24 days postfracture, with the majority at 1-2 weeks. We conclude that 7% of pediatric forearm fractures treated by closed reduction are subject to reangulation and/or displacement following routine acceptable primary treatment, and that remanipulation provides a safe, effective means to obtain and maintain reduction.


Aptamer-based fluorescence polarization assay for separation-free exosome quantification.
Tumor-derived exosomes have emerged as promising cancer biomarkers and attracted increasing interest in non-invasive cancer diagnosis and treatment monitoring. However, the identification and quantification of exosomes in clinical samples such as blood remains challenging due to the difficulty in trade-off between recognition specificity and isolation efficiency. Here we have developed an aptamer-based fluorescence polarization assay for exosome quantification, which is a separation-free, amplification-free and sensitive approach enabling direct quantification of exosomes in human plasma. While the key specificity of this assay is based on the aptamer's inherent affinity to membrane proteins on exosomes, exosomes' inherent huge mass/volume acts as mass-based fluorescence polarization amplifier. Our assay allows quantitative analysis of exosomes in the range of 5 × 102 to 5 × 105 particles per μL with a detect limitation of 500 particles per μL for the cell line deprived exosomes. The total assay time is about 30 min with just one mix-and-read step to achieve high sensitivity. We have also demonstrated quantification of exosomes from lung cancer patients and healthy donors in clinical samples. This work describes a new and simple liquid biopsy assay to directly detect exosomes in the biological matrix, which facilitates cancer diagnosis and therapy monitoring.


Alveolar gas composition before and after maximal breath-holds in competitive divers.
The urge to breathe, as stimulated by hypercapnia, is generally considered to cause a breath-hold diver to end the breath-hold, and pre-breath hold hyperventilation has been suggested to cause hypoxic loss of consciousness (LOC) due to the reduced urge to breathe. Competitors hyperventilate before "Static Apnea", yet only 10% surface with symptoms of hypoxia such as loss of motor control (LMC) or LOC. We hypothesized that the extensive hyperventilation would prevent hypercapnia even during prolonged breath-holding and we also recorded breaking-point end-tidal PO2 in humans. Nine breath-hold divers performed breath-holds of maximal duration according to their chosen "Static Apnea" procedure. They floated face down in a swimming pool (28 degrees C). The only non-standard procedure was that they exhaled into a sampling tube for end-expiratory air, before starting the breath-hold and before resuming breathing. Breath-hold duration was 284 +/- 25 (SD) seconds. End-tidal PCO2 was 18.9 +/- 2.0 mmHg before apnea and 38.3 +/- 4.7 mmHg at apnea termination. End-tidal PO2 was 131.7 +/- 2.7 mmHg before apnea and 26.9 +/- 7.5 mmHg at apnea termination. Two of the subjects showed LMC after exhaling into the sampling tube; their end-tidal PAO2 values were 19.6 and 21.0 mmHg, respectively. End-tidal CO2 was normocapnic or hypocapnic at the termination of breath-holds. These data suggest that the athletes rely primarily on the hypoxic stimuli, probably in interaction with CO2 stimuli to determine when to end breath-holds. The severity of hypoxia close to LOC was similar to that reported for acute hypobaric hypoxia in humans.


A comparison of (18)F-dopa PET and inversion recovery MRI in the diagnosis of Parkinson's disease.
To quantify structural changes in the substantia nigra of patients with PD with inversion recovery MRI and to compare these with striatal dopaminergic function measured with (18)F-dopa PET. The authors studied 10 patients with PD and eight age-matched control subjects with a combination of MR sequences previously reported to be sensitive to nigral cell loss. Striatal regions of interest were defined on T1-weighted MRI coregistered to (18)F-dopa PET in all subjects. Discriminant function analysis of the quantified MR nigral signal correctly classified 83% of the combined PD patient/control group; three of 10 PD cases were incorrectly classified as "normal" (Wilks' lambda = 0.724, p > 0.05). Discriminant function analysis correctly classified 100% of PD patients and control subjects with (18)F-dopa PET based on mean caudate and putamen K(i) values (Wilks' lambda = 0.065, p < 0.001). Correlations between mean putamen K(i) and rostral and caudal nigral MR signal changes and mean caudate K(i) and caudal nigral MR signal changes were found (r = -0.76, -0.69, -0.80, p < 0.05). (18)F-dopa PET is more reliable than inversion recovery MRI in discriminating patients with moderately severe PD from normal subjects. However, the structural changes detected within the substantia nigra of patients with PD found using inversion recovery MRI correlate with measures of striatal dopaminergic function using (18)F-dopa PET.


Formation of F-ring isoprostane-like compounds (F3-isoprostanes) in vivo from eicosapentaenoic acid.
Eicosapentaenoic acid (EPA, C20:5, omega-3) is the most abundant polyunsaturated fatty acid (PUFA) in fish oil. Recent studies suggest that the beneficial effects of fish oil are due, in part, to the generation of various free radical-generated non-enzymatic bioactive oxidation products from omega-3 PUFAs, although the specific molecular species responsible for these effects have not been identified. Our research group has previously reported that pro-inflammatory prostaglandin F2-like compounds, termed F2-isoprostanes (IsoPs), are produced in vivo by the free radical-catalyzed peroxidation of arachidonic acid and represent one of the major products resulting from the oxidation of this PUFA. Based on these observations, we questioned whether F2-IsoP-like compounds (F3-IsoPs) are formed from the oxidation of EPA in vivo. Oxidation of EPA in vitro yielded a series of compounds that were structurally established to be F3-IsoPs using a number of chemical and mass spectrometric approaches. The amounts formed were extremely large (up to 8.7 + 1.0 microg/mg EPA) and greater than levels of F2-IsoPs generated from arachidonic acid. We then examined the formation of F3-IsoPs in vivo in mice. Levels of F3-IsoPs in tissues such as heart are virtually undetectable at baseline, but supplementation of animals with EPA markedly increases quantities up to 27.4 + 5.6 ng/g of heart. Interestingly, EPA supplementation also markedly reduced levels of pro-inflammatory arachidonate-derived F2-IsoPs by up to 64% (p < 0.05). Our studies provide the first evidence that identify F3-IsoPs as novel oxidation products of EPA that are generated in vivo. Further understanding of the biological consequences of F3-IsoP formation may provide valuable insights into the cardioprotective mechanism of EPA.


A phylogenomic and molecular signature based approach for characterization of the phylum Spirochaetes and its major clades: proposal for a taxonomic revision of the phylum.
The Spirochaetes species cause many important diseases including syphilis and Lyme disease. Except for their containing a distinctive endoflagella, no other molecular or biochemical characteristics are presently known that are specific for either all Spirochaetes or its different families. We report detailed comparative and phylogenomic analyses of protein sequences from Spirochaetes genomes to understand their evolutionary relationships and to identify molecular signatures for this group. These studies have identified 38 conserved signature indels (CSIs) that are specific for either all members of the phylum Spirochaetes or its different main clades. Of these CSIs, a 3 aa insert in the FlgC protein is uniquely shared by all sequenced Spirochaetes providing a molecular marker for this phylum. Seven, six, and five CSIs in different proteins are specific for members of the families Spirochaetaceae, Brachyspiraceae, and Leptospiraceae, respectively. Of the 19 other identified CSIs, 3 are uniquely shared by members of the genera Sphaerochaeta, Spirochaeta, and Treponema, whereas 16 others are specific for the genus Borrelia. A monophyletic grouping of the genera Sphaerochaeta, Spirochaeta, and Treponema distinct from the genus Borrelia is also strongly supported by phylogenetic trees based upon concatenated sequences of 22 conserved proteins. The molecular markers described here provide novel and more definitive means for identification and demarcation of different main groups of Spirochaetes. To accommodate the extensive genetic diversity of the Spirochaetes as revealed by different CSIs and phylogenetic analyses, it is proposed that the four families of this phylum should be elevated to the order level taxonomic ranks (viz. Spirochaetales, Brevinematales ord. nov., Brachyspiriales ord. nov., and Leptospiriales ord. nov.). It is further proposed that the genera Borrelia and Cristispira be transferred to a new family Borreliaceae fam. nov. within the order Spirochaetales.


The Tonoplastic Inositol Transporter INT1 From Arabidopsis thaliana Impacts Cell Elongation in a Sucrose-Dependent Way.
The tonoplastic inositol transporter INT1 is the only known transport protein in Arabidopsis that facilitates myo-inositol import from the vacuole into the cytoplasm. Impairment of the release of vacuolar inositol by knockout of INT1 results in a severe inhibition of cell elongation in roots as well as in etiolated hypocotyls. Importantly, a more strongly reduced cell elongation was observed when sucrose was supplied in the growth medium, and this sucrose-dependent effect can be complemented by the addition of exogenous myo-inositol. Comparing int1 mutants (defective in transport) with mutants defective in myo-inositol biosynthesis (mips1 mutants) revealed that the sucrose-induced inhibition in cell elongation does not just depend on inositol depletion. Secondary effects as observed for altered availability of inositol in biosynthesis mutants, as disturbed membrane turnover, alterations in PIN protein localization or alterations in inositol-derived signaling molecules could be ruled out to be responsible for impairing the cell elongation in int1 mutants. Although the molecular mechanism remains to be elucidated, our data implicate a crucial role of INT1-transported myo-inositol in regulating cell elongation in a sucrose-dependent manner and underline recent reports of regulatory roles for sucrose and other carbohydrate intermediates as metabolic semaphores.


(Homo)polymer-mediated colloidal stability of micellar solutions.
Despite their wide range of applications, there is a remarkable lack of fundamental understanding about how micelles respond to other components in solution. The colloidal stability of micellar solutions in presence of (homo)polymers is investigated here following a theoretical bottom-up approach. A polymer-mediated micelle-micelle interaction is extracted from changes in the micelle-unimer equilibrium as a function of the inter-micelle distance. The homopolymer-mediated diblock copolymer micelle-micelle interaction is studied both for depletion and adsorption of the homopolymer. The fluffy nature of the solvophilic domain (corona) of the micelle weakens the depletion-induced destabilization. Accumulation of polymers into the corona induces bridging attraction between micelles. In fact, both depletion and adsorption phenomena are regulated by the coronal thickness relative to the size of the added polymer. Penetration of guest compounds into the coronal domain of crew-cut micelles, with a narrower yet denser corona, is less pronounced as for starlike micelles (with a more diffuse corona). Therefore, crew-cut micelles are less sensitive to the effect of added compounds, and hence more suitable for applications in multicomponent systems, such as industrial formulations or biological fluids. The trends observed for the colloidal stability of crew-cut micelles qualitatively match with our experimental observations on aqueous dispersions of polycaprolactone-polyethylene glycol (PCL-PEO) micellar suspensions with added PEO chains.


Effectiveness of pandemic influenza A/H1N1 vaccine for prevention of otitis media in children.
Our aim was to evaluate effectiveness of pandemic influenza A/H1N1 vaccine in preventing acute otitis media (AOM) and/or otitis media with effusion (OME), in a randomized, prospective and single-blind study conducted in the children aged of 6-60 months. This study was done between December 1, 2009 and April 30, 2010 during the pandemia between June 2009 and May 2010. On the healthy children, vaccinated against pandemic influenza A/H1N1 and age-matched unvaccinated controls, the rate of AOM, OME, and any otitis media (OM) attack (sum of AOM and OME attacks) confirmed by otoscopic and tympanometric examination, and their associations with risk factors were looked for. Otoscopic and tympanometric evaluation was done twice within the follow-up period of 4-8 weeks. Totally 46 vaccinated and 46 unvaccinated healthy children were enrolled. No difference in rates of AOM, OME, or OM was found between vaccinated and unvaccinated children. But logistic regression analysis revealed that unvaccinated children had 2.9-folds more risk for OME and OM, but not for AOM. Further, male gender and bottle feeding and/or using pacifier revealed significant relationships with AOM. We conclude that pandemic influenza A/H1N1 vaccine prevented OME rather than AOM attacks in children with 6-60 months of age.


Intraocular Lens Implantation in the Absence of Zonular Support: An Outcomes and Safety Update: A Report by the American Academy of Ophthalmology.
To review the published literature on the visual acuity results and complications of different surgical techniques for intraocular lens (IOL) implantation in the absence of zonular support. Peer-reviewed literature searches were conducted last in PubMed and the Cochrane Library in July 2019. The searches yielded 734 citations of articles published in English. The panel reviewed the abstracts of these mostly retrospective case series studies, and 45 were determined to be relevant to the assessment objectives. Three articles were rated as level II evidence, and 42 articles were rated as level III evidence. Eight different types of IOL fixation techniques with at least 6-month follow-up were evaluated: anterior chamber IOL (ACIOL), iris-claw IOL, retropupillary iris-claw IOL, 10-0 polypropylene iris-sutured posterior chamber IOL (PCIOL), 10-0 polypropylene scleral-sutured PCIOL, 8-0 polypropylene scleral-sutured PCIOL, CV-8 polytetrafluoroethylene, and intrascleral haptic fixation (ISHF). Eight articles reported data comparing 2 techniques. The 45 studies had insufficient statistical power to compare the techniques conclusively. A qualitative analysis of similar types showed that trends in visual acuity outcomes were not inferior to those of ACIOL implantation, but the severity of preoperative pathologic features was not controlled for. Compared with ACIOL, complications of cystoid macular edema were higher in 10-0 polypropylene iris-sutured PCIOL and 8-0 polypropylene scleral-sutured PCIOL. Non-anterior chamber IOL techniques were less likely to report chronic uveitis. Chronic glaucoma was highest in the 8-0 polypropylene scleral-sutured PCIOL group. Although retinal detachment was infrequent overall, it was twice as common in both iris- and scleral-sutured PCIOLs (except CV-8 polytetrafluoroethylene suture) compared with nonsutured methods: ACIOL, iris-clipped IOL, and ISHF PCIOL. The evidence reviewed shows no superiority of any single IOL implantation technique in the absence of zonular support. The various techniques seem to have equivalent visual acuity outcomes and safety profiles. Each technique has its own profile of inherent risk of postoperative complications. Surgeons must educate patients on the importance of close, long-term follow-up as a result of the uncertain nature of these techniques. Large prospective studies are needed to confirm the long-term complication profiles of these various IOL implantation techniques.


Chick metabolic rate and growth in three species of albatross: a comparative study.
The relative importance of genetic vs. environmental factors in determining the pattern of avian post-embryonic development is much debated. Previous cross-fostering of albatrosses suggested that although inter-specific variation in growth rate was determined primarily by differences in dietary energy content, species-specific constraints might have evolved that could limit maximal growth, even in chicks fed at similar rates and on similar diets. This study aimed to determine whether intrinsic differences in resting metabolic rate were apparent during the linear phase of growth in chicks of three species (black-browed, grey-headed and light-mantled sooty albatrosses). There was a gradual increase in absolute, and a reduction in mass-specific metabolic rate from 5.0 W kg(-1) during the earliest part of linear growth, to 3.5 W kg(-1) by the time chicks reached peak mass. These values are considerably higher than in resting adults of comparable or lower mass, presumably reflecting the large size and high metabolic demand of organs involved in rapid nutrient processing and tissue synthesis by chicks. The lack of any detectable inter-specific variation in the pattern of metabolic rate changes casts some doubt on the existence of fundamental differences in growth rate that cannot be attributed simply to differences in dietary energy or nutrient delivery rate.


Influence of phosphate anion adsorption on the kinetics of oxygen electroreduction on low index Pt(hkl) single crystals.
The detrimental effects of phosphate anion adsorption on the oxygen reduction reactions (ORR) on low index Pt single crystal electrodes were studied in 0.1 M perchloric acid by using a hanging meniscus rotating disk electrode in the presence of varied concentrations of H(3)PO(4). The kinetic current for ORR decreased dramatically on Pt(100), Pt(110), Pt(111), and PtSn(111) even with the addition of a small amount (1 mM) of H(3)PO(4) into the perchloric acid solution, most probably due to the adsorption of phosphate anions onto the Pt active sites that impeded the electroreduction of O(2). Remarkably, the extent of decline was found to vary with the specific single crystal surface, following the order of Pt(111) > PtSn(111) > Pt(110) ∼ Pt(100). Consistent behaviors were also observed in Tafel analysis and in electrochemical impedance spectroscopic measurements. Within the present experimental context, Pt(110) was found to be the optimal crystal surface for ORR in phosphoric acid fuel cells with the smallest charge transfer resistance, whereas the poisoning effects of phosphate anion adsorption were the most pronounced on Pt(111), most likely because the phosphate anions primarily adsorbed on the 3-fold sites on the Pt(111) faces, as manifested in in situ X-ray absorption spectroscopic measurements.


High residual platelet reactivity on clopidogrel: its significance and therapeutic challenges overcoming clopidogrel resistance.
Over the last decade, dual antiplatelet therapy has been the mainstay of the management of Acute Coronary Syndrome, with clopidogrel therapy providing clear benefits over aspirin monotherapy and becoming the agent of choice for the prevention of stent thrombosis. While newer antiplatelet agents have now become available, clopidogrel is still widely used due to its low cost and efficacy. However, many patients still experience recurrent ischemic events. A poor response of the platelets to clopidogrel, called High Residual Platelet Reactivity (HRPR), has been incriminated to account for this dilemma. Despite the absence of a universal definition of HRPR or the gold standard test to quantify it, persistent high platelet reactivity has consistently been associated with recurrence of ischemic events. Clopidogrel metabolism is highly variable, and genetics, comorbidities and drug interactions can affect it. In this article we review all definitions of HRPR, explore the available tests to quantify it, the clinical outcomes associated with it, as well as strategies that have shown success in overcoming it.


Social and physical contexts of schools and neighborhoods: associations with physical activity among young people in New Zealand.
We sought to determine the association between school- and neighborhood-level characteristics and physical activity among young people. We collected the data as part of Youth'07, a nationally representative survey of the health and well-being of high school students in New Zealand. In total, 9107 students from 96 schools participated (63% response rate). Students answered questions about their schools (e.g., support for physical activity) and neighborhoods (e.g., community cohesion, disintegration, safety, and recreational facilities). We created school-level measures by aggregating the students' reports within their schools and we created neighborhood-level measures by aggregating the students' reports of their neighborhoods to the census area unit of their residential address. We conducted analyses by using cross-classified random-effects models controlling for individual variables, with school and neighborhoods treated as random effects. Schools characterized by high sports team participation and neighborhoods characterized by high social connections were positively associated with student physical activity. We observed few other significant characteristics of school and neighborhood environments. Our findings highlight that opportunity for sports participation and strong social connections in neighborhoods are particularly important for youths' physical activity.


Pathogen-mediated manipulation of arthropod microbiota to promote infection.
Arthropods transmit diverse infectious agents; however, the ways microbes influence their vector to enhance colonization are poorly understood. Ixodes scapularis ticks harbor numerous human pathogens, including Anaplasma phagocytophilum, the agent of human granulocytic anaplasmosis. We now demonstrate that A. phagocytophilum modifies the I. scapularis microbiota to more efficiently infect the tick. A. phagocytophilum induces ticks to express Ixodes scapularis antifreeze glycoprotein (iafgp), which encodes a protein with several properties, including the ability to alter bacterial biofilm formation. IAFGP thereby perturbs the tick gut microbiota, which influences the integrity of the peritrophic matrix and gut barrier-critical obstacles for Anaplasma colonization. Mechanistically, IAFGP binds the terminal d-alanine residue of the pentapeptide chain of bacterial peptidoglycan, resulting in altered permeability and the capacity of bacteria to form biofilms. These data elucidate the molecular mechanisms by which a human pathogen appropriates an arthropod antibacterial protein to alter the gut microbiota and more effectively colonize the vector.


Subject and authorship of records related to the Organization for Tropical Studies (OTS) in BINABITROP, a comprehensive database about Costa Rican biology.
BINABITROP is a bibliographical database of more than 38000 records about the ecosystems and organisms of Costa Rica. In contrast with commercial databases, such as Web of Knowledge and Scopus, which exclude most of the scientific journals published in tropical countries, BINABITROP is a comprehensive record of knowledge on the tropical ecosystems and organisms of Costa Rica. We analyzed its contents in three sites (La Selva, Palo Verde and Las Cruces) and recorded scientific field, taxonomic group and authorship. We found that most records dealt with ecology and systematics, and that most authors published only one article in the study period (1963-2011). Most research was published in four journals: Biotropica, Revista de Biología Tropical/ International Journal of Tropical Biology and Conservation, Zootaxa and Brenesia. This may be the first study of a such a comprehensive database for any case of tropical biology literature.


Nonviable Staphylococcus aureus and its peptidoglycan stimulate macrophage recruitment, angiogenesis, fibroplasia, and collagen accumulation in wounded rats.
We have previously shown that local application at the time of operation of Staphylococcus aureus, nonviable S. aureus, its cell wall, or S. aureus peptidoglycan accelerates wound healing. We hypothesized that this effect is due to both direct and indirect mechanisms, among which is an increase in the inflammatory response to wounding, resulting in an increase in macrophages, angiogenesis, and fibroblasts. Twenty-seven Sprague-Dawley male rats were anesthetized, and two 7-cm paravertebral skin incisions were made. Four polyvinyl alcohol sponges, two on each side, containing either 100 microliter of isotonic saline or 0.5 mg of nonviable S. aureus or S. aureus peptidoglycan in 100-microliter saline were implanted subcutaneously. Nonviable S. aureus or S. aureus peptidoglycan (860 microgram/cm incision) in 200-microliter saline were inoculated into the incisions at closure. The rats ate a commercial rat chow and drank tap water ad libitum throughout. After days 3 and 7 postwounding, rats were euthanized, and tissues were examined for immunohistochemical features of reparative tissue using ED-1, Factor VIII, and vimentin antibodies, markers for monocyte/macrophages, endothelial cells, and mesenchymal cells (including fibroblasts), respectively. Incisions treated with nonviable S. aureus or S. aureus peptidoglycan showed more macrophages along and deep in the wound tract 7 days postoperatively. Nonviable S. aureus or S. aureus peptidoglycan-treated sponges were surrounded and penetrated by much larger capsules of reparative tissue than saline-treated sponges at both 3 and 7 days. Neutrophil influx was much greater in nonviable S. aureus or S. aureus peptidoglycan-treated sponges, especially in central regions, and there were many more ED-1-stained macrophages in distinct geographic locations, specifically, the more peripheral-cortical areas. Some clustering of macrophages occurred around areas of invasion by reparative tissue into the surrounding subcutaneous fat and within the interstices of the sponges at the interface between reparative tissue and acute inflammatory cells. In contrast, saline-treated sponge reparative tissue had significantly fewer macrophages, much thinner and flimsy reparative tissue, with proportionately fewer macrophages clustering centrally. There were many more mesenchymal cells (notably fibroblasts) and new blood vessels and much more reparative collagen in the nonviable S. aureus or S. aureus peptidoglycan-treated sponges. We conclude that local application of nonviable S. aureus or S. aureus peptidoglycan at wounding induces an increased number and alteration in location of macrophages, increased influx (or proliferation) of mesenchymal cells (notably fibroblasts), and increased angiogenesis and reparative collagen accumulation, as well as increasing the overall acute inflammatory response to wounding.


Evaluation of Different Primers for Detection of Brucella by Using PCR Method.
Brucellosis is a worldwide zoonosis and a significant cause of loss of health in humans and animals. Traditionally, classic diagnosis is carried out by isolation of Brucella, which is time-consuming, technically challenging and potentially dangerous. The aim of this study was to expand a molecular test that would be used for the develop detection of Brucella in a single reaction with high sensitivity and specificity, by targeting IS711element. This study was carried out from 2015 to 2016 at the Ayatolla Rohani hospital in Babol, Iran. The present study was designed to develop PCR assay, based on IS711 gene for rapid diagnosis of Brucella spp. and immediate detection of Brucella, with high sensitivity and specificity. Four pairs of oligo-nucleotide primers with sizes of 547, 403, 291 and 127bp respectively, were planned to exclusively amplify the targeted genes of Brucella species. Our results show that, five PCR primers set up, would be helpful in amplifying the DNAs from the genus Brucella with high specificity and sensitivity so it can be 12 fg, for Brucella species to provide a valuable tool for diagnosis. This method can be more useful than serological and biochemical tests and in addition, this reduces the number of required tests more rapidly and economically.


Identification of the pattern of copper as an etiological factor in oral submucous fibrosis: a cytological study.
Oral Submucous Fibrosis (OSF) is a well-recognized, potentially malignant disorder causing generalized fibrosis of the submucosal oral soft tissues. Though this disease is believed to be multi-factorial, areca nut chewing has emerged as the most important causative factor for OSF. Areca nut is known to have high levels of copper, which is believed to cause lysyl oxidase associated fibrosis. To evaluate the pattern of copper in buccal mucosal cells of OSF patients, areca nut chewers and normal healthy individuals and to elicit the etiology of copper in OSF. Patients were divided into three groups each comprising of 20 individuals- Healthy individuals (Group I); areca nut chewers without OSF (Group II); histopathologically confirmed OSF (Group Ill). The cytological smears made from each patient were stained with rhodanine stain for copper and evaluated for the qualitative and quantitative parameters of copper by using specific grading criteria. Quantitative estimation of copper content showed a marked variation in the mean values. Mean value of group I was 0.11 ± 0.39; group II was 1.09 ± 0.81 and group III was 2.34 ± 0.74 (p<0.001). Mean values for qualitative estimation of copper were - 0.01 ± 0.36 for group I, 1.08 ± 0.82 for group II and 2.39 ± 0.72 for group III (p<0.001). Chi square analysis was used to assess the percentage distribution of copper granules. This revealed that the colour intensity and the number of granules were seen to maximum in OSF patients, areca nut chewers without OSF having intermediated values and normal healthy individuals having the least values. An evident increase in the copper staining in group III individuals as compared to group I and group II was well appreciated. Increased copper levels in the local environment of the oral cavity indicates its role in lysyl oxidase associated submucosal fibrosis.


Unique T cell effector functions elicited by Plasmodium falciparum epitopes in malaria-exposed Africans tested by three T cell assays.
Natural immunity to malaria is characterized by low level CD4 T cell reactivity detected by either lymphoproliferation or IFN-gamma secretion. Here we show a doubling in the detection rate of responders to the carboxyl terminus of circumsporozoite protein (CS) of Plasmodium falciparum by employing three T cell assays simultaneously: rapid IFN-gamma secretion (ex vivo ELISPOT), IFN-gamma secretion after reactivation of memory T cells and expansion in vitro (cultured ELISPOT), and lymphoproliferation. Remarkably, for no individual peptide did a positive response for one T cell effector function correlate with any other. Thus these CS epitopes elicited unique T cell response patterns in malaria-exposed donors. Novel or important epitope responses may therefore be missed if only one T cell assay is employed. A borderline correlation was found between anti-CS Ab levels and proliferative responses, but no correlation was found with ex vivo or cultured IFN-gamma responses. This suggested that the proliferating population, but not the IFN-gamma-secreting cells, contained cells that provide help for Ab production. The data suggest that natural immunity to malaria is a complex function of T cell subgroups with different effector functions and has important implications for future studies of natural T cell immunity.


Notch signaling distinguishes 2 waves of definitive hematopoiesis in the zebrafish embryo.
Recent studies have revealed that definitive hematopoiesis in vertebrates initiates through the formation of a non-self-renewing progenitor with limited multilineage differentiation potential termed the erythromyeloid progenitor (EMP). EMPs are specified before hematopoietic stem cells (HSCs), which self-renew and are capable of forming all mature adult blood lineages including lymphoid cells. Despite their differences, EMPs and HSCs share many phenotypic traits, making precise study of their respective functions difficult. Here, we examine whether embryonic specification of EMPs requires Notch signaling as has been shown for HSCs. In mindbomb mutants, which lack functional Notch ligands, we show that EMPs are specified normally: we detect no significant differences in cell number, gene expression, or differentiation capacity between EMPs purified from wild-type (WT) or mindbomb mutant embryos. Similarly N-[N-(3,5-difluorophenacetyl)-l-alanyl]-S-phenylglycine t-butyl ester (DAPT), a chemical inhibitor of Notch receptor activation, has no effect on EMP specification. These studies establish that HSCs are the only hematopoietic precursor that requires Notch signaling and help to clarify the signaling events underlying the specification of the 2 distinct waves of definitive hematopoiesis.


Validation of an air-puff passive-avoidance paradigm for assessment of aversive learning and memory in rat models of chronic pain.
Chronic pain is associated with cognitive deficits. Considerable overlap in brain regions involved in pain and aversion suggests that aversive learning and memory may be affected during chronic pain. Passive-avoidance paradigms traditionally use foot-shock to induce context-conditioned avoidance and may be unsuitable for use in animal models of chronic pain, which are commonly associated with hypersensitivity of the hind-paws. The aim of the present study was to develop and validate a novel passive-avoidance paradigm in rats, employing air-puff as the aversive stimulus, and to use this paradigm to assess aversive learning and memory in rat models of chronic inflammatory and neuropathic pain. Air-puff exposure produced a significant passive-avoidance and this response was attenuated following administration of scopolamine. Nerve-ligated rats and rats injected with complete Freund's adjuvant developed allodynia and hyperalgesia. Air-puff produced a significant passive-avoidance response in both chronic pain models. However, there was no difference in the response between either model and its respective control group. Thus, air-puff can be used as an alternative to foot-shock to induce a passive-avoidance response. The data generated using this model suggest that aversive learning and memory remain intact in the rat spinal nerve ligation and complete Freund's adjuvant models of chronic neuropathic and inflammatory pain, respectively.


Review of the Novel Investigational Antifungal Olorofim.
The incidence of invasive fungal infections caused by molds and endemic fungi is increasing. There is also concern regarding increased rates of reduced susceptibility or frank resistance among Aspergillus and Coccidioides species, while Scedosporium species, Lomentospora prolificans, and Fusarium species are inherently less susceptible or intrinsically resistant to clinically available antifungals. Olorofim (formerly F901318) is the first member of the orotomide class of antifungals to be evaluated clinically for the treatment of invasive mold infections. This agent inhibits dihydroorotate dehydrogenase, a key enzyme in the biosynthesis of pyrimidines. Olorofim has activity against many molds and thermally dimorphic fungi, including species that are resistant to azoles and amphotericin B, but lacks activity against yeasts and the Mucorales. It is currently being developed for both oral and intravenous administration. Although published clinical outcome data have been limited to case reports to date, the results against invasive and refractory infections are promising. This review describes the mechanism of action of olorofim, its in vitro spectrum of activity, and what is currently known about its pharmacokinetic profile and clinical efficacy.


Incorporating language phenotypes strengthens evidence of linkage to autism.
We investigated the effect of incorporating information about proband and parental structural language phenotypes into linkage analyses in the two regions for which we found the highest signals in our first-stage affected sibling pair genome screen: chromosomes 13q and 7q. We were particularly interested in following up on our chromosome 7q finding in light of two prior reports of linkage of this region to developmental language disorder, since one of the diagnostic criteria for autism is absent or abnormal language development. We hypothesized that if the language phenotype were genetically relevant to linkage at the chromosome 7q locus, then incorporating parents phenotypes would increase the signal at that locus, and most of the signal would originate from the subset of families in which both probands had severe language delay. The results support these hypotheses. The linkage signals we obtained on chromosome 7q as well as at least one signal on chromosome 13q are mainly attributable to the subgroup of families in which both probands had language delay. This became apparent only when the parents' history of language-related difficulties was also incorporated into the analyses. Although based on our data, we were not able to distinguish between epistasis or heterogeneity models, we tentatively concluded that there may be more than one autism susceptibility locus related to language development.


Design of a study to improve accuracy in reading mammograms.
This paper is concerned with the design and analysis of mammography reading studies. In particular we consider studies aimed at evaluating interventions to improve the accuracy with which mammograms are read. A simple randomized design is suggested in which a relatively large group of readers read sets of mammograms before and after an intervention phase. We propose solutions to three difficult statistical issues that arise in the context of such studies: (i) the choice of primary outcome measure; (ii) the data analysis technique to be employed; and (iii) the methodology for calculating sample sizes for readers and images to be read. First, we argue in favor of using sensitivity and specificity as the primary outcome measures rather than receiver operating characteristic (ROC) curves in mammography studies, although the latter are considered state of the art for many types of radiology reading studies. We argue that sensitivity and specificity are more clinically relevant and conceptually more straightforward than ROC curves. Second, we suggest a bivariate approach to data analysis for evaluating intervention effects on sensitivity and specificity. This accommodates the correlations inherent between these measures and allows for estimation of joint effects on them. Finally we propose a method for power calculations that uses computer simulation techniques. Simple formulas for sample size calculations are not available in part because variability in accuracy amongst readers and variation in difficulty among images introduce complexity into power calculations. The simulation method that we propose accommodates such complexity and is easy to implement. The methodology was motivated by a study funded by the Department of Defense to evaluate the potential efficacy of an educational intervention. In the context of this study we illustrate the steps involved in power calculations and apply the data analytic techniques to the sort of data expected to result from this study. Though the proposed methods were motivated by this particular study, the statistical considerations are relevant more broadly in mammography and indeed in other types of radiologic imaging studies. Standards for the conduct of radiologic reading studies are not yet well developed, as they are for randomized clinical trials and for case-control studies. We hope that the discussion in this paper will add to the dialogue necessary for development of such standards.


Alterations in oxidative stress status modulate terminal differentiation in Atlantic salmon adipocytes cultivated in media rich in n-3 fatty acids.
Regulation of oxidative stress (OS) in adipocytes is an important mediator of their development and dysfunction. Highly unsaturated fatty acids (HUFAs) play essential roles in marine fish, where they have anti-lipogenic effects, but they are prone to peroxidation. The aim of this study was to investigate how the effects of HUFAs in fish adipocytes are modulated by changes in their intracellular redox status. Adipocytes from Atlantic salmon were cultivated on HUFA-rich media and treated with buthionine sulfoximine (BSO), which is known to deplete stores of the antioxidant glutathione (GSH) thus increasing OS, and alpha-tocopherol (alpha-TOC), which protects from OS. Gene expression was assessed by qPCR. In addition, phospholipid composition, total fatty acid (FA) composition, TBARS, the activities of pro-apoptotic caspase 3 (CASP3) and antioxidant superoxide dismutase (SOD) were determined. BSO treatment decreased the expression of genes encoding GSH-based antioxidant enzymes glutathione peroxidase (GPX) 2 and GPX3. Consequently, depletion of GSH resulted in the highest level of peroxidation products TBARS despite the increased activity of SOD in this group. Significant reduction of TBARS was achieved by alpha-TOC. Further, in comparison to two alpha-TOC supplemented groups, GSH-depleted cells accumulated less fat and their gene expression profile of adipogenic markers was lower. The formation of large intracellular vesicles was prominent in the control and BSO groups while reduction of OS by alpha-TOC coincided with the increased gene expression of the activating transcription factor 6 (ATF6), a transducer of the endoplasmic reticulum (ER)-stress response. CASP3 assay showed no difference between groups; however, depletion of GSH resulted in the increased gene expression of several apoptotic markers. Up-regulation of the apoptosis inducible factor (AIF) implied higher probability of CASP3-independent apoptosis in cells under increased OS. In conclusion, the study provides several lines of evidence in favour of anti-adipogenic effects of OS in a cold blooded vertebrate.


Differences in frailty in older men and women.
Frailty describes an individual's vulnerability to adverse health outcomes and is a useful construct that assists health professionals to understand the heterogeneity of the ageing population. While the pathophysiological pathways that lead to frailty are not well defined, an individual's sex appears to be a key factor influencing the ageing trajectory. Compared with age-matched men, women tend to have poorer health status (ie, they are more frail) but longer life expectancy (ie, they are more resilient). It seems likely that a combination of biological, behavioural and social factors underpin this male-female health-survival paradox. Randomised controlled trial data for frailty interventions in older adults are emerging, with multicomponent programs incorporating exercise and nutrition-based strategies showing promise. Pharmaceutical and other innovative therapeutic strategies for frailty are highly anticipated. Sex differences in the effectiveness of frailty interventions have not been addressed in the research literature to date. In the future, successful interventions may target many (if not all) biopsychosocial domains, with careful consideration of issues relevant to each sex.


Effectiveness of the Engagement and Counseling for Latinos (ECLA) intervention in low-income Latinos.
Persistent disparities in access and quality of mental health care for Latinos indicate a need for evidence-based, culturally adapted, and outside-the-clinic-walls treatments. Evaluate treatment effectiveness of telephone (ECLA-T) or face-to-face (ECLA-F) delivery of a 6-8 session cognitive behavioral therapy and care management intervention for low-income Latinos, as compared to usual care for depression. Multisite randomized controlled trial. Eight community health clinics in Boston, Massachusetts and San Juan, Puerto Rico. 257 Latino patients recruited from primary care between May 2011 and September 2012. The primary outcome was severity of depression, assessed with the Patient Health Questionnaire-9 and the Hopkins Symptom Checklist-20. The secondary outcome was functioning over the previous 30 days, measured using the World Health Organization Disability Assessment Schedule (WHO-DAS 2.0). Both telephone and face-to-face versions of the Engagement and Counseling for Latinos (ECLA) were more effective than usual care. The effect sizes of both intervention conditions on Patient Health Questionnaire-9 were moderate when combined data from both sites are analyzed (0.56 and 0.64 for face-to-face and telephone, respectively). Similarly, effect sizes of ECLA-F and ECLA-T on the Hopkins Symptom Checklist were quite large in the Boston site (0.64 and 0.73. respectively) but not in Puerto Rico (0.10 and 0.03). The intervention appears to help Latino patients reduce depressive symptoms and improve functioning. Of particular importance is the higher treatment initiation for the telephone versus face-to-face intervention (89.7% vs. 78.8%), which suggests that telephone-based care may improve access and quality of care.


Social factors determining maternal and neonatal mortality in South Africa: A qualitative study.
South Africa's maternal mortality ratio has increased from 150/100 000 in 1990 to 269/100 000 live births in 2015 against the Millennium Development Goals 5 (MDG5) target of 38/100 000, indicating slow progress in improving maternal health. The neonatal mortality rate was 14/1000 live births against the MDG4 target of 7/1000. The purpose of the article was to outline the socio-economic factors that determine maternal and neonatal mortality in South African communities. To identify and describe the social determinants of maternal and neonatal mortality in South Africa. A qualitative study using audio-taped individual interviews was conducted. The interviews included 10 pregnant women who were purposefully recruited from the antenatal clinic attendees in a public hospital. The interviews were conducted in isiZulu and later translated into English by the researcher who is fluent in both. Data were analysed using the World Health Organization's (WHO) Commission on Social Determinants of Health framework. Findings revealed that poverty was an underlying factor to the vulnerability to illness and death of the mothers and their neonates. Other determinants were found to be the nutritional inadequacies, neglect and abuse by male partners, HIV or AIDS, inattention to reproductive health and violation of reproductive rights, and powerlessness of women and health system issues such as poor quality and incompetent health care. It is apparent that poverty plays a major role in determining the health of mothers and neonates. This requires more coordinated multi-sectorial interventions to address both the social determinants and direct causes of maternal and neonatal deaths.


Phosphene perceptions and safety of chronic visual cortex stimulation in a blind subject.
Stimulation of primary visual cortices has the potential to restore some degree of vision to blind individuals. Developing safe and reliable visual cortical prostheses requires assessment of the long-term stability, feasibility, and safety of generating stimulation-evoked perceptions.A NeuroPace responsive neurostimulation system was implanted in a blind individual with an 8-year history of bare light perception, and stimulation-evoked phosphenes were evaluated over 19 months (41 test sessions). Electrical stimulation was delivered via two four-contact subdural electrode strips implanted over the right medial occipital cortex. Current and charge thresholds for eliciting visual perception (phosphenes) were measured, as were the shape, size, location, and intensity of the phosphenes. Adverse events were also assessed.Stimulation of all contacts resulted in phosphene perception. Phosphenes appeared completely or partially in the left hemifield. Stimulation of the electrodes below the calcarine sulcus elicited phosphenes in the superior hemifield and vice versa. Changing the stimulation parameters of frequency, pulse width, and burst duration affected current thresholds for eliciting phosphenes, and increasing the amplitude or frequency of stimulation resulted in brighter perceptions. While stimulation thresholds decreased between an average of 5% and 12% after 19 months, spatial mapping of phosphenes remained consistent over time. Although no serious adverse events were observed, the subject experienced mild headaches and dizziness in three instances, symptoms that did not persist for more than a few hours and for which no clinical intervention was required.Using an off-the-shelf neurostimulator, the authors were able to reliably generate phosphenes in different areas of the visual field over 19 months with no serious adverse events, providing preliminary proof of feasibility and safety to proceed with visual epicortical prosthetic clinical trials. Moreover, they systematically explored the relationship between stimulation parameters and phosphene thresholds and discovered the direct relation of perception thresholds based on primary visual cortex (V1) neuronal population excitation thresholds.


Hematopoietic stem cell transplantation for patients with AML in first complete remission.
Postremission therapy in patients with acute myeloid leukemia (AML) may consist of continuing chemotherapy or transplantation using either autologous or allogeneic stem cells. Patients with favorable subtypes of AML generally receive chemotherapeutic consolidation, although recent studies have also suggested favorable outcome after hematopoietic stem cell transplantation (HSCT). Although allogeneic HSCT (alloHSCT) is considered the preferred type of postremission therapy in poor- and very-poor-risk AML, the place of alloHSCT in intermediate-risk AML is being debated, and autologous HSCT is considered a valuable alternative that may be preferred in patients without minimal residual disease after induction chemotherapy. Here, we review postremission transplantation strategies using either autologous or allogeneic stem cells. Recent developments in the field of alternative donors, including cord blood and haploidentical donors, are highlighted, and we discuss reduced-intensity alloHSCT in older AML recipients who represent the predominant category of patients with AML who have a high risk of relapse in first remission.


Synchronizing N release from organic residues: opportunities for integrated management of N.
In intensive cropping systems, mineral nitrogen (N) fertilizers represent the largest component of the N cycle because the indigenous N supply is not adequate. The requirement for mineral fertilizer may be reduced with the use of organic nutrient sources. A more realistic use of organic matter, particularly in sub-Saharan Africa due to limited amounts and availability, is the combined use of organic nutrient sources and mineral fertilizers. The beneficial effects of integrated use of inorganic fertilizers and organic manures on improved nutrient recoveries, soil moisture retention, cation exchange capacity, and erosion control have been reported. However, there are as many reports indicating negligible benefits or even disadvantages of combining nutrient sources on crop production. This is not surprising given the combination of organic residue sources, soils, climatic, crops, and management factors that influence nutrient dynamics. The most widely accepted function of organic materials is improving the nutrient availability to crops by supplying N. The key to both improving efficiency of N use and reducing N losses is synchronization of N supply from soil, biological N2 fixation, organic residues, and inorganic fertilizers with the crop N demand. Organic materials are not magic; N losses also result from their use. Controlling N release from organic sources depends on their nutrient content and quality, soil properties, and the environmental and management factors. This paper will synthesize the information generated from integrated nutrient management trials in sub-Saharan Africa and the Philippines. Management strategies based on an organic resources database and a dynamic soil-crop simulation model are used to identify organic sources as N fertilizers or soil amendments. The decision support tools are also used to attain optimum synchrony between release from organic sources and soils with crop N demand.


Chemical immobilization of adult female Weddell seals with tiletamine and zolazepam: effects of age, condition and stage of lactation.
Chemical immobilization of Weddell seals (Leptonychotes weddellii) has previously been, for the most part, problematic and this has been mainly attributed to the type of immobilizing agent used. In addition to individual sensitivity, physiological status may play an important role. We investigated the use of the intravenous administration of a 1:1 mixture of tiletamine and zolazepam (Telazol) to immobilize adult females at different points during a physiologically demanding 5-6 week lactation period. We also compared performance between IV and IM injection of the same mixture. The tiletamine:zolazepam mixture administered intravenously was an effective method for immobilization with no fatalities or pronounced apnoeas in 106 procedures; however, there was a 25 % (one animal in four) mortality rate with intramuscular administration. Induction time was slightly longer for females at the end of lactation (54.9 +/- 2.3 seconds) than at post-parturition (48.2 +/- 2.9 seconds). In addition, the number of previous captures had a positive effect on induction time. There was no evidence for effects due to age, condition (total body lipid), stage of lactation or number of captures on recovery time. We suggest that intravenous administration of tiletamine and zolazepam is an effective and safe immobilizing agent for female Weddell seals. Although individual traits could not explain variation in recovery time, we suggest careful monitoring of recovery times during longitudinal studies (> 2 captures). We show that physiological pressures do not substantially affect response to chemical immobilization with this mixture; however, consideration must be taken for differences that may exist for immobilization of adult males and juveniles. Nevertheless, we recommend a mass-specific dose of 0.50-0.65 mg/kg for future procedures with adult female Weddell seals and a starting dose of 0.50 mg/kg for other age classes and other phocid seals.


Tetrahydrobiopterin regulates superoxide and nitric oxide generation by recombinant endothelial nitric oxide synthase.
Nitric oxide (NO) produced by the endothelial isoform of nitric oxide synthase (NOS III) is a key determinant of the anti-atherosclerotic properties of the endothelium. Recent in vivo studies suggest that NOS III may also be a source of superoxide production, which would limit its role as a NO-producing enzyme. In the current study we examined both the NO and the superoxide generating potential of recombinant NOS III obtained from a baculovirus/Sf9 expression system. Using lucigenin chemiluminesence we could indeed demonstrate (superoxide dismutase inhibitable) superoxide production by NOS III. This superoxide production was not affected by administration of L-arginine, but could be inhibited dose-dependently by the co-factor tetrahydrobiopterin (BH4). BH4 also dose dependently decreased superoxide generation by hypoxanthine/xantine oxidase suggesting a direct antioxidant effect. Superoxide generation by NOS III could be completely inhibited by diphenyleneiodonium (DPI), an inhibitor of the flavin moiety of the enzyme, indicating that this group is a main source of superoxide production by the enzyme. Using measurement of [3H-L-arginine] conversion to [3H-L-citrulline], it appeared that BH4 directly increased the production of NO by NOS III. In addition, we observed that BH4 stablized the NOS III in its dimeric form, suggesting that an effect on allosteric conformation could be involved in this effect on NO production. NOS III thus appears to be a superoxide generating enzyme probably through its flavin moiety, as well as a BH4-dependent NO producing enzyme.


Introduction of complementary foods and the relationship to food allergy.
To address questions regarding breastfeeding, complementary feeding, allergy development, and current infant-feeding recommendations. This was a nested, case-control within a cohort study in which mothers of 41 infants diagnosed with food allergy by the age of 2 years (according to double-blind, placebo-controlled food challenge) and their 82 age-matched controls kept prospective food diaries of how their infants were fed in the first year of life. Infants who were diagnosed with food allergy by the time they were 2 years of age were introduced to solids earlier (≤16 weeks of age) and were less likely to be receiving breast milk when cow's milk protein was first introduced into their diet. This study supports the current American Academy of Pediatrics' allergy prevention recommendations and the European Society of Pediatric Gastroenterology, Hepatology and Nutrition recommendations on complementary feeding to not introduce solids before 4 to 6 months of age. It also supports the American Academy of Pediatrics' breastfeeding recommendations that breastfeeding should continue while solids are introduced into the diet and that breastfeeding should continue for 1 year, or longer, as mutually desired by mother and infant.


Eight-Channel AC Magnetosusceptometer of Magnetic Nanoparticles for High-Throughput and Ultra-High-Sensitivity Immunoassay.
An alternating-current magnetosusceptometer of antibody-functionalized magnetic nanoparticles (MNPs) was developed for immunomagnetic reduction (IMR). A high-sensitivity, high-critical-temperature superconducting quantum interference device was used in the magnetosusceptometer. Minute levels of biomarkers of early-stage neurodegeneration diseases were detectable in serum, but measuring each biomarker required approximately 4 h. Hence, an eight-channel platform was developed in this study to fit minimal screening requirements for Alzheimer's disease. Two consistent results were measured for three biomarkers, namely Aβ40, Aβ42, and tau protein, per human specimen. This paper presents the instrument configuration as well as critical characteristics, such as the low noise level variations among channels, a high signal-to-noise ratio, and the coefficient of variation for the biomarkers' IMR values. The instrument's ultrahigh sensitivity levels for the three biomarkers and the substantially shorter total measurement time in comparison with the previous single- and four-channels platforms were also demonstrated in this study. Thus, the eight-channel instrument may serve as a powerful tool for clinical high-throughput screening of Alzheimer's disease.


A randomized controlled trial of an Internet-based cognitive-behavioural skills package for carers of people with anorexia nervosa.
Anorexia nervosa (AN) poses a major burden on families. Carers (e.g. parents or partners) of people with AN are often highly distressed and may inadvertently respond in ways that can contribute to the maintenance of the disorder, e.g. through high levels of over-involvement and criticism [also known as expressed emotion (EE)]. This study aimed to evaluate the efficacy of a novel web-based systemic cognitive-behavioural (CBT) intervention for carers of people with AN, designed to reduce carer distress and teach skills in how to offer effective support. Carers of people with AN (n=64) were randomly allocated to either the web-intervention, overcoming anorexia online, with limited clinician supportive guidance (by email or phone), or to ad-hoc usual support from the UK patient and carer organization Beat. Carer outcomes were assessed at post-treatment (4 months) and follow-up (6 months). Compared with the control intervention, web-based treatment significantly reduced carers' anxiety and depression (primary outcome) at post-treatment, with a similar trend in carers' EE. Other secondary outcomes did not favour the online intervention. Gains were maintained at follow-up. This is the first ever study to use an online CBT program to successfully reduce carer distress and improve carers' ability to support the person with AN.


Protective effect of N-acetylcysteine against radiation induced DNA damage and hepatic toxicity in rats.
The present study was designed to evaluate the radioprotective effect of N- acetylcysteine (NAC) on gamma-radiation induced toxicity in hepatic tissue in rat. The cellular changes were estimated using malondialdehyde (MDA, an index of lipid peroxidation), superoxide dismutase (SOD), glutathione peroxidase (GSHPx), reduced glutathione (GSH), and total nitrate/nitrite (NO(x)) as markers of hepatic oxidative stress in rats following gamma-irradiation. The DNA damage was determined by agarose gel electrophoresis. To achieve the ultimate goal of this study, 40 adult rats were randomly divided into 4 groups of 10 animals each. Group I was injected intraperitoneally with saline solution for 7 consecutive days and served as control group. Group II was irradiated with a single dose of 6Gy gamma-radiation. Group III was daily injected with NAC (1g/kg, i.p.) for 7 consecutive days. Group IV received a daily i.p. injection of NAC (1g/kg, i.p.) for 7 consecutive days and 1h after the last dose, rats were irradiated with a single dose (6Gy) gamma-radiation. The animals were sacrificed after 24h. DNA damage was observed in tissue after total body irradiation with a single dose of 6Gy. Malondialdehyde and total nitrate/nitrite were increased significantly whereas the levels of GSH and antioxidant enzymes were significantly decreased in gamma-irradiated group. Pretreatment with NAC showed a significant decrease in the levels of MDA, NO(x) and DNA damage. The antioxidant enzymes increased significantly along with the levels of GSH. Moreover, histopathological examination of liver tissues confirmed the biochemical data. Thus, our results show that pretreatment with N-acetylcysteine offers protection against gamma-radiation induced cellular damage.


Aspp1: A Guardian of Hematopoietic Stem Cell Integrity.
Eliminating hematopoietic stem cells (HSCs) with DNA damage is necessary to maintain the homeostasis of HSCs, but the mechanisms underlying this apoptotic elimination are unclear. Now in Cell Stem Cell, Yamashita et al. (2015) show that Aspp1 coordinates with p53 to protect HSC pool integrity, guarding against hematological malignancies.


Risk factors for refractory anastomotic strictures after oesophageal atresia repair: a multicentre study.
To determine the incidence of refractory anastomotic strictures after oesophageal atresia (OA) repair and to identify risk factors associated with refractory strictures. Retrospective national multicentre study in patients with OA born between 1999 and 2013. Exclusion criteria were isolated fistula, inability to obtain oesophageal continuity, death prior to discharge and follow-up <6 months. A refractory oesophageal stricture was defined as an anastomotic stricture requiring ≥5 dilations at maximally 4-week intervals. Risk factors for development of refractory anastomotic strictures after OA repair were identified with multivariable logistic regression analysis. We included 454 children (61% male, 7% isolated OA (Gross type A)). End-to-end anastomosis was performed in 436 (96%) children. Anastomotic leakage occurred in 13%. Fifty-eight per cent of children with an end-to-end anastomosis developed an anastomotic stricture, requiring a median of 3 (range 1-34) dilations. Refractory strictures were found in 32/436 (7%) children and required a median of 10 (range 5-34) dilations. Isolated OA (OR 5.7; p=0.012), anastomotic leakage (OR 5.0; p=0.001) and the need for oesophageal dilation ≤28 days after anastomosis (OR 15.9; p<0.001) were risk factors for development of a refractory stricture. The incidence of refractory strictures of the end-to-end anastomosis in children treated for OA was 7%. Risk factors were isolated OA, anastomotic leakage and the need for oesophageal dilation less than 1 month after OA repair.


Helicobacter pylori infection in functional dyspepsia.
Functional dyspepsia is the most common reason for patients to experience chronic epigastric pain or discomfort. The causes of functional dyspepsia are multifactorial but Helicobacter pylori infection is one likely candidate. Infection with this bacterial pathogen clearly results in chronic mucosal inflammation in the stomach and duodenum, which, in turn, might lead to abnormalities in gastroduodenal motility and sensitivity. Chronic gastritis might also affect a variety of endocrine functions of the stomach including the production of the gastrointestinal hormones and neurotransmitters somatostatin, gastrin and ghrelin. Although these abnormalities might generate symptoms in some patients with functional dyspepsia, the clinical evidence needs to be critically evaluated before this hypothesis can be confirmed. A Cochrane review reported that eradication of H. pylori in these patients had a small but statistically significant long-term effect on symptom relief when compared with placebo, lasting at least 12 months after 1 week of eradication therapy. The efficacy of eradication therapy was seen in all symptom subtypes of functional dyspepsia, but was more marked in Asian than Western patients. This evidence has led to alterations in most of the major guidelines throughout the world, which now recommend H. pylori eradication in patients with functional dyspepsia if they test positive for this bacterium.


[Expression of OCT4 protein in bladder cancer and its clinicopathological implications].
To study the expression of OCT4 protein in bladder cancer and its correlation to the clinicopathologic features and prognosis of bladder cancer. OCT4 mRNA and protein expression was detected in 5 bladder cancer cell lines (RT-4, Tcc-Sup, KK47, T24, and 5637) and 1 normal bladder cell lines by real-time PCR and Western blotting, respectively. Immunohistochemical analysis was used to detect the expression of OCT4 protein in 46 bladder cancer samples. All the 5 bladder cancer cell lines expressed detectable levels of OCT4 mRNA and proteins, whereas the normal bladder cell line SV-HUC-1 was negative for OCT4 expression. The clinical bladder cancer tissues showed a high positivity rate of OCT4 expression (76.1%), which was not detected in normal bladder tissues. Specific OCT-4 signals were localized mainly in the nuclei of the cancer cells. The expression rate of OCT4 protein was significantly higher in bladder cancer tissue than in normal bladder epithelium (P<0.05), and showed a positive correlation to the grade of tumor differentiation and metastasis (P<0.05) but not to the patients' age, gender or TNM stage. OCT4 protein expression is associated with tumor differentiation and metastasis in bladder cancer and may play an important role in the early diagnosis and prognostic evaluation of bladder cancer.


Dual disruption of DNA repair and telomere maintenance for the treatment of head and neck cancer.
Poly(ADP-ribose) polymerases (PARP) and the Mre11, Rad50, and Nbs1 (MRN) complex are key regulators of DNA repair, and have been recently shown to independently regulate telomere length. Sensitivity of cancers to PARPi is largely dependent on the BRCAness of the cells. Unfortunately, the vast majority of cancers are BRCA-proficient. In this study, therefore, we investigated whether a targeted molecular "hit" on the MRN complex, which is upstream of BRCA, can effectively sensitize BRCA-proficient head and neck squamous cell carcinoma (HNSCC) to PARP inhibitor (PARPi). Human HNSCC cell lines and a mouse model with HNSCC xenografts were used in this study. In vitro and in vivo studies were conducted to evaluate the effects and underlying mechanisms of dual molecular disruption of PARP and the MRN complex, using a pharmacologic inhibitor and a dominant-negative Nbs1 expression vector, respectively. Our findings demonstrate that downregulation of the MRN complex disrupts homologous recombination, and, when combined with PARPi, leads to accumulation of lethal DNA double-strand breaks. Moreover, we show that PARPi and MRN complex disruption induces significantly shortening telomere length. Together, our results demonstrate that dual disruption of these pathways causes significant cell death in BRCA-proficient tumor cells both in vitro and in vivo. Our study, for the first time, elucidates a novel mechanism for MRN complex and PARP inhibition beyond DNA repair, demonstrating the feasibility of a dual disruption approach that extends the utility of PARPi to the treatment of BRCA-proficient cancers.


Investigation of association between the TRAF family genes and RA susceptibility.
The tumour necrosis factor (TNF) receptor-associated factor (TRAF) family is important in activating multiple inflammatory and immune related processes induced by cytokines such as TNFalpha and interleukin-1. These genes therefore represent strong candidate susceptibility factors for rheumatoid arthritis (RA). A study was undertaken to investigate the association between single nucleotide polymorphisms (SNPs) spanning six TRAF genes and RA in a British population. Twenty-three haplotype tagging (ht) SNPs and 26 random SNPs spanning the six TRAF genes were initially tested for association in a cohort of 351 unrelated patients with RA and 368 controls. Any SNPs demonstrating an association were genotyped in further samples. Sequenom MassARRAY technology was preferentially used for genotyping. Both single point and haplotypic analyses were performed. Forty-four SNPs were successfully genotyped and conformed to Hardy-Weinberg expectation. A single SNP, rs7514863, mapping upstream of the TRAF5 gene and affecting a putative transcription factor binding site, demonstrated a significant association across the entire cohort of 1273 cases with RA compared with 2463 healthy controls (OR for minor T allele 1.2 (95% CI 1.06 to 1.36), p = 0.005). The association was stronger in the subgroup carrying at least one copy of the shared epitope alleles (OR 1.43 (95% CI 1.18 to 1.73), p = 0.0003). These findings provide evidence for the association of an SNP upstream of a strong candidate RA susceptibility gene, TRAF5, in a large cohort of patients and controls. Further association and functional studies are required to investigate the role of this variant, or one in linkage disequilibrium with it, in RA disease causation.


SUMO and ubiquitin modifications during steroid hormone synthesis and function.
Steroid hormones control many aspects of animal physiology and behaviour. They are highly regulated, among other mechanisms, by post-translational modifications of the transcription factors involved in their synthesis and response. In the present review, we will focus on the influence of SUMO (small ubiquitin-related modifier) and ubiquitin modifications on the function of transcription factors involved in adrenal cortex formation, steroidogenesis and the hormonal response.