Nanoscale structure and microscale stiffness of DNA nanotubes.
We measure the stiffness of tiled DNA nanotubes (HX-tubes) as a function of their (defined) circumference by analyzing their micrometer-scale thermal deformations using fluorescence microscopy. We derive a model that relates nanoscale features of HX-tube architecture to the measured persistence lengths. Given the known stiffness of double-stranded DNA, we use this model to constrain the average spacing between and effective stiffness of individual DNA duplexes in the tube. A key structural feature of tiled nanotubes that can affect stiffness is their potential to form with discrete amounts of twist of the DNA duplexes about the tube axis (supertwist). We visualize the supertwist of HX-tubes using electron microscopy of gold nanoparticles, attached to specific sites along the nanotube. This method reveals that HX-tubes tend not to form with supertwist unless forced by sequence design, and, even when forced, supertwist is reduced by elastic deformations of the underlying DNA lattice. We compare the hybridization energy gained upon closing a duplex sheet into a tube with the elastic energy paid for deforming the sheet to allow closure. In estimating the elastic energy we account for bending and twisting of the individual duplexes as well as shearing between them. We find the minimum supertwist state has minimum free energy, and global untwisting of forced supertwist is energetically favorable, consistent with our experimental data. Finally, we show that attachment of Cy3 dyes or changing counterions can cause nanotubes to adopt a permanent writhe with micrometer-scale pitch and amplitude. We propose that the coupling of local twist and global counter-twist may be useful in characterizing perturbations of DNA structure.


The influence of polygenic risk scores on heritability of anti-CCP level in RA.
The objective of this study was to study genetic factors that influence quantitative anticyclic citrullinated peptide (anti-CCP) antibody levels in RA patients. We carried out a genome-wide association study (GWAS) meta-analysis using 1975 anti-CCP+ RA patients from three large cohorts, the Brigham Rheumatoid Arthritis Sequential Study (BRASS), North American Rheumatoid Arthritis Consortium (NARAC) and the Epidemiological Investigation of RA (EIRA). We also carried out a genome-wide complex trait analysis (GCTA) to estimate the heritability of anti-CCP levels. GWAS-meta-analysis showed that anti-CCP levels were most strongly associated with the human leukocyte antigen (HLA) region with a P-value of 2 × 10(-11) for rs1980493. There were 112 SNPs in this region that exceeded the genome-wide significance threshold of 5 × 10(-8), and all were in linkage disequilibrium (LD) with the HLA- DRB1*03 allele with LD r(2) in the range of 0.25-0.88. Suggestive novel associations outside of the HLA region were also observed for rs8063248 (near the GP2 gene) with a P-value of 3 × 10(-7). None of the known RA risk alleles (∼52 loci) were associated with anti-CCP level. Heritability analysis estimated that 44% of anti-CCP variation was attributable to genetic factors captured by GWAS variants. In summary, anti-CCP level is a heritable trait, and HLA-DR3 and GP2 are associated with lower anti-CCP levels.


The motor cortex shows adaptive functional changes to brain injury from multiple sclerosis.
Although multiple sclerosis (MS) is an inflammatory demyelinating disease, there can be substantial axonal injury and loss. We therefore hypothesized that adaptive cortical changes may contribute to limiting functional impairment, particularly in the early stages of the disease. To test our hypothesis, we used functional magnetic resonance imaging (MRI) to characterize the localization and volumes of activation in the motor cortex during simple flexion-extension finger movements. There were differences in the patterns of cortical activation with movement between the 12 MS patients and the 12 normal controls. All patients showed greater relative supplementary motor area activation than did the normal controls. The relative hemispheric lateralization of sensorimotor cortex (SMC) activation decreased in direct proportion to the total cerebral T2-weighted MRI hyperintense lesion load. This appeared to be due primarily to increases in ipsilateral SMC activation with increasing lesion load in white matter of the hemisphere contralateral to the limb moved. The center of activation in the contralateral SMC was shifted a mean of 8.8 mm posterior in patients relative to controls, providing additional evidence for cortical adaptive responses to injury. The magnitude of this posterior shift in the SMC activation increased with greater T2 lesion loads. These observations demonstrate that cortical recruitment for simple finger movements can change both quantitatively and qualitatively in the SMCs of MS patients, suggesting that cortical reorganization or "unmasking" of latent pathways can contribute to functional recovery. These adaptive changes are another factor potentially limiting the strength of the relationship between MRI measures of pathology and clinical measures of disability.


On Textual Analysis and Machine Learning for Cyberstalking Detection.
Cyber security has become a major concern for users and businesses alike. Cyberstalking and harassment have been identified as a growing anti-social problem. Besides detecting cyberstalking and harassment, there is the need to gather digital evidence, often by the victim. To this end, we provide an overview of and discuss relevant technological means, in particular coming from text analytics as well as machine learning, that are capable to address the above challenges. We present a framework for the detection of text-based cyberstalking and the role and challenges of some core techniques such as author identification, text classification and personalisation. We then discuss PAN, a network and evaluation initiative that focusses on digital text forensics, in particular author identification.


Structure-activity relationships of human AKR-type oxidoreductases involved in bile acid synthesis: AKR1D1 and AKR1C4.
Two members of the human aldo-keto reductase (AKR) superfamily participate in the biosynthesis of bile acids by catalyzing the NADP(H) dependent reduction of 3-keto groups (AKR1C4) and Delta4 double bonds (AKR1D1) of oxysterol precursors. Structure determination of human AKR1C4 and homology modelling of AKR1D1 followed by docking experiments were used to explore active site geometries. Substrate docking resulted in ligand poses satisfying catalytic constraints, and indicates a critical role for Trp227/230 in positioning the substrate in a catalytically competent orientation. Based on the evidence gathered from our docking experiments and experimental structures, this tryptophan residue emerges as a major determinant governing substrate specificity of a subset of enzymes belonging to the AKR1 subfamily.


Effects of psychotherapy on trauma-related cognitions in posttraumatic stress disorder: a meta-analysis.
In the fifth edition of the Diagnostic and Statistical Manual of Mental Disorders criteria for posttraumatic stress disorder (PTSD) incorporate trauma-related cognitions. This adaptation of the criteria has consequences for the treatment of PTSD. Until now, comprehensive information about the effect of psychotherapy on trauma-related cognitions has been lacking. Therefore, the goal of our meta-analysis was to determine which psychotherapy most effectively reduces trauma-related cognitions. Our literature search for randomized controlled trials resulted in 16 studies with data from 994 participants. We found significant effect sizes favoring trauma-focused cognitive-behavioral therapy as compared to nonactive or active nontrauma-focused control conditions of Hedges' g = 1.21, 95% CI [0.69, 1.72], p < .001 and g = 0.36, 95% CI [0.09, 0.63], p = .009, respectively. Treatment conditions with elements of cognitive restructuring and treatment conditions with elements of exposure, but no cognitive restructuring reduced trauma-related cognitions almost to the same degree. Treatments with cognitive restructuring had small advantages over treatments without cognitive restructuring. We concluded that trauma-focused cognitive-behavioral therapy effectively reduces trauma-related cognitions. Treatments comprising either combinations of cognitive restructuring and imaginal exposure and in vivo exposure, or imaginal exposure and in vivo exposure alone showed the largest effects.


Plasma phospholipid fatty acids and CHD in older men: Whitehall study of London civil servants.
Dietary fatty acids (FA) are the major determinants of blood lipids, and measurements of plasma phospholipid FA (PL-FA) composition that reflect the dietary intake of FA may provide insights into the relationships between diet and CHD. We assessed CHD mortality associations with PL-FA (SFA, PUFA and MUFA) levels measured in a nested case-control study of 116 cases of CHD death and 239 controls that were frequency-matched for age and employment grade. The participants had plasma levels of total cholesterol, LDL-cholesterol (LDL-C), HDL-cholesterol, apo B and apo A1, C-reactive protein (CRP) and fibrinogen recorded. SFA levels were significantly positively correlated with total cholesterol, LDL-C, apo B, CRP protein and fibrinogen. By contrast, phospholipid-PUFA were inversely associated with CRP, but not with any of the lipids. A higher SFA content (top v. bottom quarter) was associated with a 2-fold higher risk of CHD (OR and 95 % CI: OR 2.12; 95 % CI: 1.13, 3.99), and an equivalent difference in PUFA was associated with a halving in CHD risk (OR 0.49; 95 % CI: 0.26, 0.94), but MUFA was unrelated to CHD risk. These associations were substantially attenuated, after additional adjustment for lipids and inflammatory markers. Higher levels of saturated fat and lower levels of polyunsaturated fats were each associated with a higher risk of CHD in elderly men, and these associations were partly explained by their effects on blood lipids and biomarkers of inflammation.


Diffusion imaging of whole, post-mortem human brains on a clinical MRI scanner.
Diffusion imaging of post mortem brains has great potential both as a reference for brain specimens that undergo sectioning, and as a link between in vivo diffusion studies and "gold standard" histology/dissection. While there is a relatively mature literature on post mortem diffusion imaging of animals, human brains have proven more challenging due to their incompatibility with high-performance scanners. This study presents a method for post mortem diffusion imaging of whole, human brains using a clinical 3-Tesla scanner with a 3D segmented EPI spin-echo sequence. Results in eleven brains at 0.94 × 0.94 × 0.94 mm resolution are presented, and in a single brain at 0.73 × 0.73 × 0.73 mm resolution. Region-of-interest analysis of diffusion tensor parameters indicate that these properties are altered compared to in vivo (reduced diffusivity and anisotropy), with significant dependence on post mortem interval (time from death to fixation). Despite these alterations, diffusion tractography of several major tracts is successfully demonstrated at both resolutions. We also report novel findings of cortical anisotropy and partial volume effects.


ATXN2-CAG42 sequesters PABPC1 into insolubility and induces FBXW8 in cerebellum of old ataxic knock-in mice.
Spinocerebellar Ataxia Type 2 (SCA2) is caused by expansion of a polyglutamine encoding triplet repeat in the human ATXN2 gene beyond (CAG)(31). This is thought to mediate toxic gain-of-function by protein aggregation and to affect RNA processing, resulting in degenerative processes affecting preferentially cerebellar neurons. As a faithful animal model, we generated a knock-in mouse replacing the single CAG of murine Atxn2 with CAG42, a frequent patient genotype. This expansion size was inherited stably. The mice showed phenotypes with reduced weight and later motor incoordination. Although brain Atxn2 mRNA became elevated, soluble ATXN2 protein levels diminished over time, which might explain partial loss-of-function effects. Deficits in soluble ATXN2 protein correlated with the appearance of insoluble ATXN2, a progressive feature in cerebellum possibly reflecting toxic gains-of-function. Since in vitro ATXN2 overexpression was known to reduce levels of its protein interactor PABPC1, we studied expansion effects on PABPC1. In cortex, PABPC1 transcript and soluble and insoluble protein levels were increased. In the more vulnerable cerebellum, the progressive insolubility of PABPC1 was accompanied by decreased soluble protein levels, with PABPC1 mRNA showing no compensatory increase. The sequestration of PABPC1 into insolubility by ATXN2 function gains was validated in human cell culture. To understand consequences on mRNA processing, transcriptome profiles at medium and old age in three different tissues were studied and demonstrated a selective induction of Fbxw8 in the old cerebellum. Fbxw8 is encoded next to the Atxn2 locus and was shown in vitro to decrease the level of expanded insoluble ATXN2 protein. In conclusion, our data support the concept that expanded ATXN2 undergoes progressive insolubility and affects PABPC1 by a toxic gain-of-function mechanism with tissue-specific effects, which may be partially alleviated by the induction of FBXW8.


The role of mitochondria in aging.
The biological basis of human aging remains one of the greatest unanswered scientific questions. Increasing evidence, however, points to a role for alterations in mitochondrial function as a potential central regulator of the aging process. Here, we focus primarily on three aspects of mitochondrial biology that link this ancient organelle to how and why we age. In particular, we discuss the role of mitochondria in regulating the innate immune system, the mechanisms linking mitochondrial quality control to age-dependent pathology, and the possibility that mitochondrial-to-nuclear signaling might regulate the rate of aging.


Using Gaussian-process regression for meta-analytic neuroimaging inference based on sparse observations.
The purpose of neuroimaging meta-analysis is to localize the brain regions that are activated consistently in response to a certain intervention. As a commonly used technique, current coordinate-based meta-analyses (CBMA) of neuroimaging studies utilize relatively sparse information from published studies, typically only using (x,y,z) coordinates of the activation peaks. Such CBMA methods have several limitations. First, there is no way to jointly incorporate deactivation information when available, which has been shown to result in an inaccurate statistic image when assessing a difference contrast. Second, the scale of a kernel reflecting spatial uncertainty must be set without taking the effect size (e.g., Z-stat) into account. To address these problems, we employ Gaussian-process regression (GPR), explicitly estimating the unobserved statistic image given the sparse peak activation "coordinate" and "standardized effect-size estimate" data. In particular, our model allows estimation of effect size at each voxel, something existing CBMA methods cannot produce. Our results show that GPR outperforms existing CBMA techniques and is capable of more accurately reproducing the (usually unavailable) full-image analysis results.


Nitric oxide donors enhance the frequency dependence of dopamine release in nucleus accumbens.
Dopamine (DA) neurotransmission in the nucleus accumbens (NAc) is critically involved in normal as well as maladaptive motivated behaviors including drug addiction. Whether the striatal neuromodulator nitric oxide (NO) influences DA release in NAc is unknown. We investigated whether exogenous NO modulates DA transmission in NAc core and how this interaction varies depending on the frequency of presynaptic activation. We detected DA with cyclic voltammetry at carbon-fiber microelectrodes in mouse NAc in slices following stimuli spanning a full range of DA neuron firing frequencies (1-100 Hz). NO donors 3-morpholinosydnonimine hydrochloride (SIN-1) or z-1-[N-(3-ammoniopropyl)-N-(n-propyl)amino]diazen-1-ium-1,2-diolate (PAPA/NONOate) enhanced DA release with increasing stimulus frequency. This NO-mediated enhancement of frequency sensitivity of DA release was not prevented by inhibition of soluble guanylyl cyclase (sGC), DA transporters, or large conductance Ca(2+)-activated K(+) channels, and did not require glutamatergic or GABAergic input. However, experiments to identify whether frequency-dependent NO effects were mediated via changes in powerful acetylcholine-DA interactions revealed multiple components to NO modulation of DA release. In the presence of a nicotinic receptor antagonist (dihydro-β-erythroidine), NO donors increased DA release in a frequency-independent manner. These data suggest that NO in the NAc can modulate DA release through multiple GC-independent neuronal mechanisms whose net outcome varies depending on the activity in DA neurons and accumbal cholinergic interneurons. In the presence of accumbal acetylcholine, NO promotes the sensitivity of DA release to presynaptic activation, but with reduced acetylcholine input, NO will promote DA release in an activity-independent manner through a direct action on dopaminergic terminals.


Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks in 188 countries, 1990-2013: a systematic analysis for the Global Burden of Disease Study 2013.
The Global Burden of Disease, Injuries, and Risk Factor study 2013 (GBD 2013) is the first of a series of annual updates of the GBD. Risk factor quantification, particularly of modifiable risk factors, can help to identify emerging threats to population health and opportunities for prevention. The GBD 2013 provides a timely opportunity to update the comparative risk assessment with new data for exposure, relative risks, and evidence on the appropriate counterfactual risk distribution. Attributable deaths, years of life lost, years lived with disability, and disability-adjusted life-years (DALYs) have been estimated for 79 risks or clusters of risks using the GBD 2010 methods. Risk-outcome pairs meeting explicit evidence criteria were assessed for 188 countries for the period 1990-2013 by age and sex using three inputs: risk exposure, relative risks, and the theoretical minimum risk exposure level (TMREL). Risks are organised into a hierarchy with blocks of behavioural, environmental and occupational, and metabolic risks at the first level of the hierarchy. The next level in the hierarchy includes nine clusters of related risks and two individual risks, with more detail provided at levels 3 and 4 of the hierarchy. Compared with GBD 2010, six new risk factors have been added: handwashing practices, occupational exposure to trichloroethylene, childhood wasting, childhood stunting, unsafe sex, and low glomerular filtration rate. For most risks, data for exposure were synthesised with a Bayesian meta-regression method, DisMod-MR 2.0, or spatial-temporal Gaussian process regression. Relative risks were based on meta-regressions of published cohort and intervention studies. Attributable burden for clusters of risks and all risks combined took into account evidence on the mediation of some risks such as high body-mass index (BMI) through other risks such as high systolic blood pressure and high cholesterol. All risks combined account for 57·2% (95% uncertainty interval [UI] 55·8-58·5) of deaths and 41·6% (40·1-43·0) of DALYs. Risks quantified account for 87·9% (86·5-89·3) of cardiovascular disease DALYs, ranging to a low of 0% for neonatal disorders and neglected tropical diseases and malaria. In terms of global DALYs in 2013, six risks or clusters of risks each caused more than 5% of DALYs: dietary risks accounting for 11·3 million deaths and 241·4 million DALYs, high systolic blood pressure for 10·4 million deaths and 208·1 million DALYs, child and maternal malnutrition for 1·7 million deaths and 176·9 million DALYs, tobacco smoke for 6·1 million deaths and 143·5 million DALYs, air pollution for 5·5 million deaths and 141·5 million DALYs, and high BMI for 4·4 million deaths and 134·0 million DALYs. Risk factor patterns vary across regions and countries and with time. In sub-Saharan Africa, the leading risk factors are child and maternal malnutrition, unsafe sex, and unsafe water, sanitation, and handwashing. In women, in nearly all countries in the Americas, north Africa, and the Middle East, and in many other high-income countries, high BMI is the leading risk factor, with high systolic blood pressure as the leading risk in most of Central and Eastern Europe and south and east Asia. For men, high systolic blood pressure or tobacco use are the leading risks in nearly all high-income countries, in north Africa and the Middle East, Europe, and Asia. For men and women, unsafe sex is the leading risk in a corridor from Kenya to South Africa. Behavioural, environmental and occupational, and metabolic risks can explain half of global mortality and more than one-third of global DALYs providing many opportunities for prevention. Of the larger risks, the attributable burden of high BMI has increased in the past 23 years. In view of the prominence of behavioural risk factors, behavioural and social science research on interventions for these risks should be strengthened. Many prevention and primary care policy options are available now to act on key risks. Bill & Melinda Gates Foundation.


Non fitting based FRET-FLIM analysis approaches applied to quantify protein-protein interactions in live cells.
New imaging methodologies in quantitative fluorescence microscopy and nanoscopy have been developed in the last few years and are beginning to be extensively applied to biological problems, such as the localization and quantification of protein interactions. Fluorescence resonance energy transfer (FRET) detected by fluorescence lifetime imaging microscopy (FLIM) is currently employed not only in biophysics or chemistry but also in bio-medicine, thanks to new advancements in technology and also new developments in data treatment. FRET-FLIM can be a very useful tool to ascertain protein interactions occurring in single living cells. In this review, we stress the importance of increasing the acquisition speed when working in vivo employing Time-Domain FLIM. The development of the new mathematical-based non-fitting methods allows the determining of the fraction of interacting donor without the requirement of high count statistics, and thus allows the performing of high speed acquisitions in FRET-FLIM to still be quantitative.


The increase in pulmonary arterial pressure caused by hypoxia depends on iron status.
Hypoxia is a major cause of pulmonary hypertension. Gene expression activated by the transcription factor hypoxia-inducible factor (HIF) is central to this process. The oxygen-sensing iron-dependent dioxygenase enzymes that regulate HIF are highly sensitive to varying iron availability. It is unknown whether iron similarly influences the pulmonary vasculature. This human physiology study aimed to determine whether varying iron availability affects pulmonary arterial pressure and the pulmonary vascular response to hypoxia, as predicted biochemically by the role of HIF. In a controlled crossover study, 16 healthy iron-replete volunteers undertook two separate protocols. The 'Iron Protocol' studied the effects of an intravenous infusion of iron on the pulmonary vascular response to 8 h of sustained hypoxia. The 'Desferrioxamine Protocol' examined the effects of an 8 h intravenous infusion of the iron chelator desferrioxamine on the pulmonary circulation. Primary outcome measures were pulmonary artery systolic pressure (PASP) and the PASP response to acute hypoxia (DeltaPASP), assessed by Doppler echocardiography. In the Iron Protocol, infusion of iron abolished or greatly reduced both the elevation in baseline PASP (P < 0.001) and the enhanced sensitivity of the pulmonary vasculature to acute hypoxia (P = 0.002) that are induced by exposure to sustained hypoxia. In the Desferrioxamine Protocol, desferrioxamine significantly elevated both PASP (P < 0.001) and DeltaPASP (P = 0.01). We conclude that iron availability modifies pulmonary arterial pressure and pulmonary vascular responses to hypoxia. Further research should investigate the potential for therapeutic manipulation of iron status in the management of hypoxic pulmonary hypertensive disease.


Arginine Methylation Regulates MEIS2 Nuclear Localization to Promote Neuronal Differentiation of Adult SVZ Progenitors.
Adult neurogenesis is regulated by stem cell niche-derived extrinsic factors and cell-intrinsic regulators, yet the mechanisms by which niche signals impinge on the activity of intrinsic neurogenic transcription factors remain poorly defined. Here, we report that MEIS2, an essential regulator of adult SVZ neurogenesis, is subject to posttranslational regulation in the SVZ olfactory bulb neurogenic system. Nuclear accumulation of MEIS2 in adult SVZ-derived progenitor cells follows downregulation of EGFR signaling and is modulated by methylation of MEIS2 on a conserved arginine, which lies in close proximity to nested binding sites for the nuclear export receptor CRM1 and the MEIS dimerization partner PBX1. Methylation impairs interaction with CRM1 without affecting PBX1 dimerization and thereby allows MEIS2 nuclear accumulation, a prerequisite for neuronal differentiation. Our results describe a form of posttranscriptional modulation of adult SVZ neurogenesis whereby an extrinsic signal fine-tunes neurogenesis through posttranslational modification of a transcriptional regulator of cell fate.


Glutathione facilitates enterovirus assembly by binding at a druggable pocket.
Enteroviruses cause a range of human and animal diseases, some life-threatening, but there remain no licenced anti-enterovirus drugs. However, a benzene-sulfonamide derivative and related compounds have been shown recently to block infection of a range of enteroviruses by binding the capsid at a positively-charged surface depression conserved across many enteroviruses. It has also been established that glutathione is essential for the assembly of many enteroviruses, interacting with the capsid proteins to facilitate the formation of the pentameric assembly intermediate, although the mechanism is unknown. Here we show, by high resolution structure analyses of enterovirus F3, that reduced glutathione binds to the same interprotomer pocket as the benzene-sulfonamide derivative. Bound glutathione makes strong interactions with adjacent protomers, thereby explaining the underlying biological role of this druggable binding pocket and delineating the pharmacophore for potential antivirals.


Functional localization of the system for visuospatial attention using positron emission tomography.
PET was used to image the neural system underlying visuospatial attention. Analysis of data at both the group and individual-subject level provided anatomical resolution superior to that described to date. Six right-handed male subjects were selected from a pilot behavioural study in which behavioural responses and eye movements were recorded. The attention tasks involved covert shifts of attention, where peripheral cues indicated the location of subsequent target stimuli to be discriminated. One attention condition emphasized reflexive aspects of spatial orientation, while the other required controlled shifts of attention. PET activations agreed closely with the cortical regions recently proposed to form the core of a neural network for spatial attention. The two attention tasks evoked largely overlapping patterns of neural activation, supporting the existence of a general neural system for visuospatial attention with regional functional specialization. Specifically, neocortical activations were observed in the right anterior cingulate gyrus (Brodmann area 24), in the intraparietal sulcus of right posterior parietal cortex, and in the mesial and lateral premotor cortices (Brodmann area 6).


The Magpie Trial: a randomised trial comparing magnesium sulphate with placebo for pre-eclampsia. Outcome for women at 2 years.
The aim of this study was to assess long-term effects for women following the use of magnesium sulphate for pre-eclampsia. Assessment at 2-3 years after delivery for women recruited to the Magpie Trial (recruitment in 1998-2001, ISRCTN 86938761), which compared magnesium sulphate with placebo for pre-eclampsia. Follow up after discharge from hospital at 125 centres in 19 countries across five continents. A total of 7927 women were randomised at the follow-up centres. Of these women, 2544 were not included for logistic reasons and 601 excluded (109 at a centre where <20% of women were contacted, 466 discharged without a surviving child and 26 opted out). Therefore, 4782 women were selected for follow-up, of whom 3375 (71%) were traced. Questionnaire assessment was administered largely by post or in a dedicated clinic. Interview assessment of selected women was performed. Main outcome measures Death or serious morbidity potentially related to pre-eclampsia at follow up, other morbidity and use of health service resources. Median time from delivery to follow up was 26 months (interquartile range 19-36). Fifty-eight of 1650 (3.5%) women allocated magnesium sulphate died or had serious morbidity potentially related to pre-eclampsia compared with 72 of 1725 (4.2%) women allocated placebo (relative risk 0.84, 95% CI 0.60-1.18). The reduction in the risk of eclampsia following prophylaxis with magnesium sulphate was not associated with an excess of death or disability for the women after 2 years.


The role of non-resolving inflammation in atherosclerosis.
Non-resolving inflammation drives the development of clinically dangerous atherosclerotic lesions by promoting sustained plaque inflammation, large necrotic cores, thin fibrous caps, and thrombosis. Resolution of inflammation is not merely a passive return to homeostasis, but rather an active process mediated by specific molecules, including fatty acid-derived specialized pro-resolving mediators (SPMs). In advanced atherosclerosis, there is an imbalance between levels of SPMs and proinflammatory lipid mediators, which results in sustained leukocyte influx into lesions, inflammatory macrophage polarization, and impaired efferocytosis. In animal models of advanced atherosclerosis, restoration of SPMs limits plaque progression by suppressing inflammation, enhancing efferocytosis, and promoting an increase in collagen cap thickness. This Review discusses the roles of non-resolving inflammation in atherosclerosis and highlights the unique therapeutic potential of SPMs in blocking the progression of clinically dangerous plaques.


Cushing syndrome: uncovering Carney complex due to novel PRKAR1A mutation.
Carney complex (CNC) is a rare multiple neoplasia syndrome characterized by spotty pigmentation of the skin and mucosa in association with various non-endocrine and endocrine tumors, including primary pigmented nodular adrenocortical disease (PPNAD). A 20-year-old woman was referred for suspected Cushing syndrome. She had signs of cortisol excess as well as skin lentigines on physical examination. Biochemical investigation was suggestive of corticotropin (ACTH)-independent Cushing syndrome. Unenhanced computed tomography scan of the abdomen did not reveal an obvious adrenal mass. She subsequently underwent bilateral laparoscopic adrenalectomy, and histopathology was consistent with PPNAD. Genetic testing revealed a novel frameshift pathogenic variant c.488delC/p.Thr163MetfsX2 (ClinVar Variation ID: 424516) in the PRKAR1A gene, consistent with clinical suspicion for CNC. Evaluation for other clinical features of the complex was unrevealing. We present a case of PPNAD-associated Cushing syndrome leading to the diagnosis of CNC due to a novel PRKAR1A pathogenic variant. Learning points: PPNAD should be considered in the differential for ACTH-independent Cushing syndrome, especially when adrenal imaging appears normal. The diagnosis of PPNAD should prompt screening for CNC. CNC is a rare multiple neoplasia syndrome caused by inactivating pathogenic variants in the PRKAR1A gene. Timely diagnosis of CNC and careful surveillance can help prevent potentially fatal complications of the disease.


Preventing Alzheimer's disease-related gray matter atrophy by B-vitamin treatment.
Is it possible to prevent atrophy of key brain regions related to cognitive decline and Alzheimer's disease (AD)? One approach is to modify nongenetic risk factors, for instance by lowering elevated plasma homocysteine using B vitamins. In an initial, randomized controlled study on elderly subjects with increased dementia risk (mild cognitive impairment according to 2004 Petersen criteria), we showed that high-dose B-vitamin treatment (folic acid 0.8 mg, vitamin B6 20 mg, vitamin B12 0.5 mg) slowed shrinkage of the whole brain volume over 2 y. Here, we go further by demonstrating that B-vitamin treatment reduces, by as much as seven fold, the cerebral atrophy in those gray matter (GM) regions specifically vulnerable to the AD process, including the medial temporal lobe. In the placebo group, higher homocysteine levels at baseline are associated with faster GM atrophy, but this deleterious effect is largely prevented by B-vitamin treatment. We additionally show that the beneficial effect of B vitamins is confined to participants with high homocysteine (above the median, 11 µmol/L) and that, in these participants, a causal Bayesian network analysis indicates the following chain of events: B vitamins lower homocysteine, which directly leads to a decrease in GM atrophy, thereby slowing cognitive decline. Our results show that B-vitamin supplementation can slow the atrophy of specific brain regions that are a key component of the AD process and that are associated with cognitive decline. Further B-vitamin supplementation trials focusing on elderly subjets with high homocysteine levels are warranted to see if progression to dementia can be prevented.


Tightrope act: autophagy in stem cell renewal, differentiation, proliferation, and aging.
Autophagy is a constitutive lysosomal catabolic pathway that degrades damaged organelles and protein aggregates. Stem cells are characterized by self-renewal, pluripotency, and quiescence; their long life span, limited capacity to dilute cellular waste and spent organelles due to quiescence, along with their requirement for remodeling in order to differentiate, all suggest that they require autophagy more than other cell types. Here, we review the current literature on the role of autophagy in embryonic and adult stem cells, including hematopoietic, mesenchymal, and neuronal stem cells, highlighting the diverse and contrasting roles autophagy plays in their biology. Furthermore, we review the few studies on stem cells, lysosomal activity, and autophagy. Novel techniques to detect autophagy in primary cells are required to study autophagy in different stem cell types. These will help to elucidate the importance of autophagy in stem cells during transplantation, a promising therapeutic approach for many diseases.


Determination and optimization of joint torques and joint reaction forces in therapeutic exercises with elastic resistance.
A model has been developed to definitively characterize the resistance properties and the joint loading (i.e., shear and compressive components of the joint reaction force) in single-joint exercises with ideal elastic bands. The model accounts for the relevant geometric and elastic properties of the band, the band pre-stretching, and the relative positioning among the joint center of rotation and the fixation points of the band. All the possible elastic torque profiles of ascending-descending, descending, or ascending type were disclosed in relation to the different ranges of joint angles. From these results the elastic resistance setting that best reproduces the average-user's knee extensor torque in maximal isometric/isokinetic efforts was determined. In this optimized setting, the shear tibiofemoral reaction force corresponding to an anterior (posterior) tibial displacement was 65% smaller than (nearly the same as) that obtained in a cam-equipped leg-extension equipment for equal values of resistance torque peak, whereas the compressive tibiofemoral reaction force was 22% higher. Compared to a weight-stack leg-extension equipment, an elastic resistance optimized setting has the potential to give a more effective quadriceps activation across the range of motion, and greatly reduces the anterior cruciate ligament strain force, which represents the main drawback of existing open kinetic-chain knee-extension exercises.


Comparative MIC evaluation of a generic ceftriaxone by broth microdilution on clinically relevant isolates from an academic hospital complex in South Africa.
We evaluated the in vitro microbiological efficacy of a generic ceftriaxone product against several clinically significant organisms collected from sterile sites. The minimum inhibitory concentration (MIC) of each was determined simultaneously with the reference and the generic ceftriaxone product. Comparative analysis of MICs between the two products for each isolate was performed using both categorical (interpretive) agreement and essential (actual MIC value) agreement. A total of 260 isolates were tested. Overall, there was categorical agreement of 98.9% and essential agreement of 95.8%. The categorical agreement for all isolates (96.7 - 100%) accorded with international standards, as no very major errors were seen and the major error rate was less than 3%. Of the 90 isolates of E. coli (40), Klebsiella spp. (40) and Salmonella spp. (10), 87.6% had an MIC less than or equal to 0.12mg/l. The generic ceftriaxone product showed equivalent efficacy by MIC determination to the reference formulation. Ceftriaxone remains a viable and useful antimicrobial agent against a variety of clinically relevant organisms in our setting.


Mechanisms of disease: genetic insights into the etiology of type 2 diabetes and obesity.
Until recently, progress in identification of the genetic variants influencing predisposition to common forms of diabetes and obesity has been slow, a sharp contrast to the large number of genes implicated in rare monogenic forms of both conditions. Recent advances have transformed the situation, however, enabling researchers to undertake well-powered scans able to detect association signals across the entire genome. For type 2 diabetes, the six high-density genome-wide association studies so far performed have extended the number of loci harboring common variants implicated in diabetes susceptibility into double figures. One of these loci, mapping to the fat mass and obesity associated gene (FTO), influences diabetes risk through a primary effect on fat mass, making this the first common variant known to influence weight and individual risk of obesity. These findings offer two main avenues for clinical translation. First, the identification of new pathways involved in disease predisposition-for example, those influencing zinc transport and pancreatic islet regeneration in the case of type 2 diabetes-offers opportunities for development of novel therapeutic and preventative approaches. Second, with continuing efforts to identify additional genetic variants, it may become possible to use patterns of predisposition to tailor individual management of these conditions.


Atypical manifestation of parathyroid carcinoma with late-onset distant metastases.
Parathyroid carcinoma is an extremely rare endocrine malignancy that accounts for less than 1% of cases of primary hyperparathyroidism. We report a 44-year-old woman who presented with fatigue and diffuse bone pain. Laboratory findings revealed highly elevated serum calcium and parathyroid hormone (PTH) levels and a 4.5 × 3 × 2.5 cm cystic lesion in the lower pole of the right thyroid lobe that was shown histologically to be a parathyroid carcinoma. Ten years later, the patient developed brain and pulmonary metastases and recurrence of PTH-related hypercalcemia. Treatment of hypercalcemia along with localized radiotherapy and various chemotherapy regimens failed to induce a biochemical or radiological response. In conclusion, parathyroid carcinoma is a rare neoplasia that may develop metastases even after prolonged follow-up, for which there is no evidence-based treatment besides surgery. Different chemotherapeutic schemes did not prove to be of any benefit in our case highlighting the need for registering such patients to better understand tumor biology and develop specific treatment. Metastases can develop many years after parathyroid cancer diagnosis.Surgery is the only curative treatment for parathyroid carcinoma.Chemotherapy and radiotherapy prove to be ineffective in parathyroid cancer treatment.Patient registering is required in order to delineate underlining pathology and offer specific treatment.


Attention biases visual activity in visual short-term memory.
In the current study, we tested whether representations in visual STM (VSTM) can be biased via top-down attentional modulation of visual activity in retinotopically specific locations. We manipulated attention using retrospective cues presented during the retention interval of a VSTM task. Retrospective cues triggered activity in a large-scale network implicated in attentional control and led to retinotopically specific modulation of activity in early visual areas V1-V4. Importantly, shifts of attention during VSTM maintenance were associated with changes in functional connectivity between pFC and retinotopic regions within V4. Our findings provide new insights into top-down control mechanisms that modulate VSTM representations for flexible and goal-directed maintenance of the most relevant memoranda.


Chemoresistance acquisition induces a global shift of expression of aniogenesis-associated genes and increased pro-angogenic activity in neuroblastoma cells.
Chemoresistance acquisition may influence cancer cell biology. Here, bioinformatics analysis of gene expression data was used to identify chemoresistance-associated changes in neuroblastoma biology. Bioinformatics analysis of gene expression data revealed that expression of angiogenesis-associated genes significantly differs between chemosensitive and chemoresistant neuroblastoma cells. A subsequent systematic analysis of a panel of 14 chemosensitive and chemoresistant neuroblastoma cell lines in vitro and in animal experiments indicated a consistent shift to a more pro-angiogenic phenotype in chemoresistant neuroblastoma cells. The molecular mechanisms underlying increased pro-angiogenic activity of neuroblastoma cells are individual and differ between the investigated chemoresistant cell lines. Treatment of animals carrying doxorubicin-resistant neuroblastoma xenografts with doxorubicin, a cytotoxic drug known to exert anti-angiogenic activity, resulted in decreased tumour vessel formation and growth indicating chemoresistance-associated enhanced pro-angiogenic activity to be relevant for tumour progression and to represent a potential therapeutic target. A bioinformatics approach allowed to identify a relevant chemoresistance-associated shift in neuroblastoma cell biology. The chemoresistance-associated enhanced pro-angiogenic activity observed in neuroblastoma cells is relevant for tumour progression and represents a potential therapeutic target.


MRC/BHF Heart Protection Study of cholesterol-lowering with simvastatin in 5963 people with diabetes: a randomised placebo-controlled trial.
Individuals with diabetes are at increased risk of cardiovascular morbidity and mortality, although typically their plasma concentrations of LDL cholesterol are similar to those in the general population. Previous evidence about the effects of lowering cholesterol in people with diabetes has been limited, and most diabetic patients do not currently receive cholesterol-lowering therapy despite their increased risk. 5963 UK adults (aged 40-80 years) known to have diabetes, and an additional 14573 with occlusive arterial disease (but no diagnosed diabetes), were randomly allocated to receive 40 mg simvastatin daily or matching placebo. Prespecified analyses in these prior disease subcategories, and other relevant subcategories, were of first major coronary event (ie, non-fatal myocardial infarction or coronary death) and of first major vascular event (ie, major coronary event, stroke or revascularisation). Analyses were also conducted of subsequent vascular events during the scheduled treatment period. Comparisons are of all simvastatin-allocated versus all placebo-allocated participants (ie, intention to treat), which yielded an average difference in LDL cholesterol of 1.0 mmol/L (39 mg/dL) during the 5-year treatment period. Both among the participants who presented with diabetes and among those who did not, there were highly significant reductions of about a quarter in the first event rate for major coronary events, for strokes, and for revascularisations. For the first occurrence of any of these major vascular events among participants with diabetes, there was a definite 22% (95% CI 13-30) reduction in the event rate (601 [20.2%] simvastatin-allocated vs 748 [25.1%] placebo-allocated, p<0.0001), which was similar to that among the other high-risk individuals studied. There were also highly significant reductions of 33% (95% CI 17-46, p=0.0003) among the 2912 diabetic participants who did not have any diagnosed occlusive arterial disease at entry, and of 27% (95% CI 13-40, p=0.0007) among the 2426 diabetic participants whose pretreatment LDL cholesterol concentration was below 3.0 mmol/L (116 mg/dL). The proportional reduction in risk was also about a quarter among various other subcategories of diabetic patient studied, including: those with different duration, type, or control of diabetes; those aged over 65 years at entry or with hypertension; and those with total cholesterol below 5.0 mmol/L (193 mg/dL). In addition, among participants who had a first major vascular event following randomisation, allocation to simvastatin reduced the rate of subsequent events during the scheduled treatment period. The present study provides direct evidence that cholesterol-lowering therapy is beneficial for people with diabetes even if they do not already have manifest coronary disease or high cholesterol concentrations. Allocation to 40 mg simvastatin daily reduced the rate of first major vascular events by about a quarter in a wide range of diabetic patients studied. After making allowance for non-compliance, actual use of this statin regimen would probably reduce these rates by about a third. For example, among the type of diabetic patient studied without occlusive arterial disease, 5 years of treatment would be expected to prevent about 45 people per 1000 from having at least one major vascular event (and, among these 45 people, to prevent about 70 first or subsequent events during this treatment period). Statin therapy should now be considered routinely for all diabetic patients at sufficiently high risk of major vascular events, irrespective of their initial cholesterol concentrations.


Treatment trials in progressive MS--current challenges and future directions.
The introduction of immunomodulatory treatments has transformed the management of patients with relapsing-remitting multiple sclerosis (MS), but has had no consistent benefit in progressive MS. Patients with primary or secondary progressive MS, therefore, are faced with relentless functional decline that remains without treatment. Clinical trials in progressive MS are clearly needed, but their design and conduct is challenging, and different from that of trials in relapsing-remitting MS. Challenges to reliable measurement of clinical progression, uncertainties about the natural history of progressive MS, and the unclear role of imaging outcomes all impede optimal trial design. Clinical trials in progressive MS have used time to a predefined change on the Expanded Disability Status Scale as their main outcome measure, which has had important consequences for trial duration and has led to inclusion of only a highly selected minority of patients. Here, we review the current approach to clinical trial design in progressive MS, outline key ongoing challenges, and suggest strategies to overcome such hurdles.


Selenium and Prostate Cancer: Analysis of Individual Participant Data From Fifteen Prospective Studies.
Some observational studies suggest that a higher selenium status is associated with a lower risk of prostate cancer but have been generally too small to provide precise estimates of associations, particularly by disease stage and grade. Collaborating investigators from 15 prospective studies provided individual-participant records (from predominantly men of white European ancestry) on blood or toenail selenium concentrations and prostate cancer risk. Odds ratios of prostate cancer by selenium concentration were estimated using multivariable-adjusted conditional logistic regression. All statistical tests were two-sided. Blood selenium was not associated with the risk of total prostate cancer (multivariable-adjusted odds ratio [OR] per 80 percentile increase = 1.01, 95% confidence interval [CI] = 0.83 to 1.23, based on 4527 case patients and 6021 control subjects). However, there was heterogeneity by disease aggressiveness (ie, advanced stage and/or prostate cancer death, Pheterogeneity = .01), with high blood selenium associated with a lower risk of aggressive disease (OR = 0.43, 95% CI = 0.21 to 0.87) but not with nonaggressive disease. Nail selenium was inversely associated with total prostate cancer (OR = 0.29, 95% CI = 0.22 to 0.40, Ptrend < .001, based on 1970 case patients and 2086 control subjects), including both nonaggressive (OR = 0.33, 95% CI = 0.22 to 0.50) and aggressive disease (OR = 0.18, 95% CI = 0.11 to 0.31, Pheterogeneity = .08). Nail, but not blood, selenium concentration is inversely associated with risk of total prostate cancer, possibly because nails are a more reliable marker of long-term selenium exposure. Both blood and nail selenium concentrations are associated with a reduced risk of aggressive disease, which warrants further investigation.


The ontogeny of bumblebee flight trajectories: from naïve explorers to experienced foragers.
Understanding strategies used by animals to explore their landscape is essential to predict how they exploit patchy resources, and consequently how they are likely to respond to changes in resource distribution. Social bees provide a good model for this and, whilst there are published descriptions of their behaviour on initial learning flights close to the colony, it is still unclear how bees find floral resources over hundreds of metres and how these flights become directed foraging trips. We investigated the spatial ecology of exploration by radar tracking bumblebees, and comparing the flight trajectories of bees with differing experience. The bees left the colony within a day or two of eclosion and flew in complex loops of ever-increasing size around the colony, exhibiting Lévy-flight characteristics constituting an optimal searching strategy. This mathematical pattern can be used to predict how animals exploring individually might exploit a patchy landscape. The bees' groundspeed, maximum displacement from the nest and total distance travelled on a trip increased significantly with experience. More experienced bees flew direct paths, predominantly flying upwind on their outward trips although forage was available in all directions. The flights differed from those of naïve honeybees: they occurred at an earlier age, showed more complex looping, and resulted in earlier returns of pollen to the colony. In summary bumblebees learn to find home and food rapidly, though phases of orientation, learning and searching were not easily separable, suggesting some multi-tasking.


Men with prostate cancer make positive dietary changes following diagnosis and treatment.
Few studies have measured dietary changes made among men diagnosed with prostate cancer (PC) without formal dietary interventions, yet they may offer insight into the needs of PC survivors. This study examined dietary changes in men before and after treatment for PC within the prostate testing for cancer and treatment randomized trial. This was a prospective cohort study in community-based men aged 50-69 tested for PC in nine UK areas. 3,935 men completed food frequency questionnaires before diagnosis and 678 with localized PC repeated the questionnaire 1 year later (response 82.7 %). Men subsequently diagnosed with or without PC all consumed similar diets before diagnosis. Diagnosis of PC led to dietary changes, with 234 (34.7 %) men eating more fresh tomatoes (p < 0.0001) and 156 (23.5 %) more tomato products (p = 0.01). 271 (40.0 %) men consumed more protein (p < 0.0001) and 193 (28.6 %) more fruit/vegetable juice (p < 0.0001). Fewer macronutrients were obtained from dairy products (p < 0.01). Men undergoing active monitoring increased their fruit/vegetable juice intake after diagnosis (p = 0.0023) more than men who had surgery or radiotherapy. Around one-third of men spontaneously adopted a healthier diet and also consumed more 'prostate-healthy' foods following a diagnosis of PC. Dietary choices also differed by radical or monitoring treatments, indicating that men undergoing active surveillance may be more likely to pursue dietary changes as an adjunct therapy. PC survivors can adopt healthier diets, thus providing clinicians with opportunities to support PC survivorship by providing targeted advice beneficial to general and potentially prostate-specific health.


The structure of tumour necrosis factor--implications for biological function.
The three-dimensional structure of TNF has been determined at 0.29 nm using the technique of X-ray crystallography. Published data on site-directed mutagenesis and antibody binding may now be assessed in the light of the structure, thus the links between structure and function for TNF may be addressed. TNF is a compact trimer composed of three identical subunits of 157 amino acids. The main-chain topology for a single subunit is essentially a beta-sandwich structure formed by two anti-parallel beta-pleated sheets. This mainchain fold corresponds to the 'jelly roll' motif observed in viral coat proteins such as VP1, VP2 and VP3 of rhinovirus, or the hemagglutinin molecule of influenza. TNF is the first non-viral protein to contain this motif. The subunits associate tightly about a threefold axis interacting through a simple edge-to-face packing of the beta-sandwich to form the solid, conical shaped trimer. A large number of the residues conserved between the amino acid sequences of TNF and lymphotoxin lie within the beta-sandwich or at the threefold axis of the trimer. This implies the presence of the same beta-sandwich motif in the lymphotoxin monomer and preservation of the edge-to-face mode of trimeric association. The detailed three dimensional structure for TNF explains a wide range of observations, including data on antibody binding and site directed mutagenesis. The currently available evidence points to a region of biological importance situated at the interface between two subunits on the lower half of the trimer.


Cigarette smoke and HIV synergistically affect lung pathology in cynomolgus macaques.
In the era of combined antiretroviral therapy (cART), lung diseases such as chronic bronchitis (CB) and chronic obstructive pulmonary disease (COPD) are common among persons living with HIV (PLWH), particularly smokers. Although smoking is highly prevalent among PLWH, HIV may be an independent risk factor for lung diseases; however, the role of HIV and cigarette smoke (CS) and their potential interaction in the development of chronic lung diseases among PLWH has not been delineated. To investigate this interaction, cynomolgus macaques were exposed to CS and/or simian-adapted human immunodeficiency virus (SHIV) and treated with cART. The development of CB and the lung functions were evaluated following CS±SHIV treatment. The results showed that in the lung, SHIV was a strong independent risk factor for goblet cell metaplasia/hyperplasia and mucus formation, MUC5AC synthesis, loss of tight junction proteins, and increased expression of Th2 cytokines/transcription factors. In addition, SHIV and CS synergistically reduced lung function and increased extrathoracic tracheal ring thickness. Interestingly, SHIV infection generated significant numbers of HIV-gp120+ epithelial cells (HGECs) in small airways and alveoli, and their numbers doubled in CS+SHIV-infected lungs. We conclude that even with cART, SHIV independently induces CB and pro-COPD changes in the lung, and the effects are exacerbated by CS.


Antidepressant drug treatment modifies the neural processing of nonconscious threat cues.
The amygdala is believed to play a key role in processing emotionally salient, threat-relevant, events that require further online processing by cortical regions. Emotional disorders such as depression and anxiety have been associated with hyperactivity of the amygdala, but it is unknown whether antidepressant treatment directly affects amygdala responses to emotionally significant information. The current study assessed the effects of 7 days administration of the selective serotonin reuptake inhibitor (SSRI), citalopram, on amygdala responses to masked presentations of fearful and happy facial expressions in never-depressed volunteers using blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging. A double-blind, between-groups design was used with volunteers randomized to 20 mg/day citalopram versus placebo. Volunteers receiving citalopram showed decreased amygdala responses to masked presentations of threat compared with those receiving placebo. Citalopram also reduced responses within the hippocampus and medial prefrontal cortex (mPFC) specifically during the fear-relevant stimuli. These neural differences were accompanied by decreased recognition of fearful facial expressions assessed after the scan. By contrast, there was no effect of citalopram on the neural or behavioral response to the happy facial expressions. These results suggest a direct effect of serotonin potentiation on amygdala response to threat-relevant stimuli in humans. Such effects may be important in the therapeutic actions of antidepressants in depression and anxiety.


Infection control management of patients with suspected highly infectious diseases in emergency departments: data from a survey in 41 facilities in 14 European countries.
In Emergency and Medical Admission Departments (EDs and MADs), prompt recognition and appropriate infection control management of patients with Highly Infectious Diseases (HIDs, e.g. Viral Hemorrhagic Fevers and SARS) are fundamental for avoiding nosocomial outbreaks. The EuroNHID (European Network for Highly Infectious Diseases) project collected data from 41 EDs and MADs in 14 European countries, located in the same facility as a national/regional referral centre for HIDs, using specifically developed checklists, during on-site visits from February to November 2009. Isolation rooms were available in 34 facilities (82,9%): these rooms had anteroom in 19, dedicated entrance in 15, negative pressure in 17, and HEPA filtration of exhausting air in 12. Only 6 centres (14,6%) had isolation rooms with all characteristics. Personnel trained for the recognition of HIDs was available in 24 facilities; management protocols for HIDs were available in 35. Preparedness level for the safe and appropriate management of HIDs is partially adequate in the surveyed EDs and MADs.


Altered adipose tissue and adipocyte function in the pathogenesis of metabolic syndrome.
Over the past decade, great progress has been made in understanding the complexity of adipose tissue biology and its role in metabolism. This includes new insights into the multiple layers of adipose tissue heterogeneity, not only differences between white and brown adipocytes, but also differences in white adipose tissue at the depot level and even heterogeneity of white adipocytes within a single depot. These inter- and intra-depot differences in adipocytes are developmentally programmed and contribute to the wide range of effects observed in disorders with fat excess (overweight/obesity) or fat loss (lipodystrophy). Recent studies also highlight the underappreciated dynamic nature of adipose tissue, including potential to undergo rapid turnover and dedifferentiation and as a source of stem cells. Finally, we explore the rapidly expanding field of adipose tissue as an endocrine organ, and how adipose tissue communicates with other tissues to regulate systemic metabolism both centrally and peripherally through secretion of adipocyte-derived peptide hormones, inflammatory mediators, signaling lipids, and miRNAs packaged in exosomes. Together these attributes and complexities create a robust, multidimensional signaling network that is central to metabolic homeostasis.


In vitro resolution of poxvirus replicative intermediates into linear minichromosomes with hairpin termini by a virally induced Holliday junction endonuclease.
Available evidence suggests that one or more late viral gene products are involved in processing poxvirus replicative intermediates into mature progeny hairpin-terminated genomes. Cloned versions of the Shope fibroma virus (SFV) replicated telomere in the inverted repeat configuration were used as substrates to assay lysates from poxvirus-infected cells for protein fractions that participate in the resolution of the circular substrate plasmid into a linear minichromosome with viral hairpin termini. An activity in a crude protein fraction obtained from vaccinia virus-infected cells at late times during the replicative cycle was capable of accurately resolving all poxviral inverted repeat replicative intermediates tested. The resolved linear products are identical to the products of in vivo resolution and possessed symmetrical nicks which mapped at the borders of the inverted repeat sequence. Strand-specific nicks were also identified, which mapped within the telomere resolution target sequence known to be required for telomere resolution in vivo. The resolving activity that we have identified is specific to virus-infected cells at late times during replication and cleaves cloned poxviral telomeric substrates in a fashion expected of a classic Holliday junction-resolving enzyme in addition to possessing a telomere resolution target-specific nicking activity. Although a Holliday junction-resolving activity would also be expected to play a role in the recombination induced by poxvirus infection, the appearance of the activity described here only after the commencement of viral late protein synthesis suggests that it functions strictly at late times. Other non-viral Holliday junction analogs can also be cleaved by this extract, suggesting that this component of the resolution activity may also play a role in other viral processes that require cleavage of a branched DNA structure. Thus, we have identified a poxviral activity that may be a part of a protein complex which resolves concatemeric replicative intermediates of viral DNA as well as participate in general recombination late during infection.


Expression QTL-based analyses reveal candidate causal genes and loci across five tumor types.
The majority of trait-associated loci discovered through genome-wide association studies are located outside of known protein coding regions. Consequently, it is difficult to ascertain the mechanism underlying these variants and to pinpoint the causal alleles. Expression quantitative trait loci (eQTLs) provide an organizing principle to address both of these issues. eQTLs are genetic loci that correlate with RNA transcript levels. Large-scale data sets such as the Cancer Genome Atlas (TCGA) provide an ideal opportunity to systematically evaluate eQTLs as they have generated multiple data types on hundreds of samples. We evaluated the determinants of gene expression (germline variants and somatic copy number and methylation) and performed cis-eQTL analyses for mRNA expression and miRNA expression in five tumor types (breast, colon, kidney, lung and prostate). We next tested 149 known cancer risk loci for eQTL effects, and observed that 42 (28.2%) were significantly associated with at least one transcript. Lastly, we described a fine-mapping strategy for these 42 eQTL target-gene associations based on an integrated strategy that combines the eQTL level of significance and the regulatory potential as measured by DNaseI hypersensitivity. For each of the risk loci, our analyses suggested 1 to 81 candidate causal variants that may be prioritized for downstream functional analysis. In summary, our study provided a comprehensive landscape of the genetic determinants of gene expression in different tumor types and ranked the genes and loci for further functional assessment of known cancer risk loci.


Reduced global cooperativity is a common feature underlying the amyloidogenicity of pathogenic lysozyme mutations.
One of the 20 or so human amyloid diseases is associated with the deposition in vital organs of full-length mutational variants of the antibacterial protein lysozyme. Here, we report experimental data that permit a detailed comparison to be made of the behaviour of two of these amyloidogenic variants, I56T and D67H, under identical conditions. Hydrogen/deuterium exchange experiments monitored by NMR and mass spectrometry reveal that, despite their different locations and the different effects of the two mutations on the structure of the native state of lysozyme, both mutations cause a cooperative destabilisation of a remarkably similar segment of the structure, comprising in both cases the beta-domain and the adjacent C-helix. As a result, both variant proteins populate transiently a closely similar, partially unstructured intermediate in which the beta-domain and the adjacent C-helix are substantially and simultaneously unfolded, whereas the three remaining alpha-helices that form the core of the alpha-domain still have their native-like structure. We show, in addition, that the binding of a camel antibody fragment, cAb-HuL6, which was raised against wild-type lysozyme, restores to both variant proteins the stability and cooperativity characteristic of the wild-type protein; as a consequence, it inhibits the formation of amyloid fibrils by both variants. These results indicate that the reduction in global cooperativity, and the associated ability to populate transiently a specific, partly unfolded intermediate state under physiologically relevant conditions, is a common feature underlying the behaviour of these two pathogenic mutations. The formation of intermolecular interactions between lysozyme molecules that are in this partially unfolded state is therefore likely to be the fundamental trigger of the aggregation process that ultimately leads to the formation and deposition in tissue of amyloid fibrils.


Nicotinic acid adenine dinucleotide phosphate (NAADP) is a second messenger in muscarinic receptor-induced contraction of guinea pig trachea.
Nicotinic acid adenine dinucleotide phosphate (NAADP) is increasingly being demonstrated to be involved in calcium signaling in many cell types and species. Although it has been shown to play a role in smooth muscle cell contraction in several tissues, nothing is known about its possible role in tracheal smooth muscle, a muscle type that is clinically relevant to asthma. To determine whether NAADP functions as a second messenger in tracheal smooth muscle contraction, we used the criteria set out by Sutherland for a molecule to be designated a second messenger. We report that NAADP satisfies all five criteria as follows. First, the NAADP antagonist Ned-19 inhibited contractions in tracheal rings and calcium increases in isolated smooth muscle cells induced by the muscarinic agonist carbachol. Second, NAADP increased cytosolic calcium in isolated cells when microinjected and was blocked by Ned-19. Third, tracheal homogenates could synthesize NAADP by base exchange from exogenous NADP and nicotinic acid and metabolize exogenous NAADP to nicotinic acid adenine dinucleotide by a 2'-phosphatase. Fourth, carbachol induced a rapid and transient increase in endogenous NAADP levels. Fifth, tracheal homogenates contained NAADP-binding sites of high affinity. Taken together, these data demonstrate that NAADP functions as a second messenger in tracheal smooth muscle, and therefore, steps in the NAADP signaling pathway might provide possible new drug targets.


Acupuncture as an adjunctive therapy to pharmacological treatment in patients with chronic pain due to osteoarthritis of the knee: a 3-armed, randomized, placebo-controlled trial.
The efficacy of acupuncture as an adjunctive therapy to pharmacological treatment of chronic pain due to knee osteoarthritis was studied with a 3-armed, single-blind, randomized, sham-controlled trial; it compared acupuncture combined with pharmacological treatment, sham acupuncture including pharmacological treatment, and pharmacological treatment alone. A total of 120 patients with knee osteoarthritis were randomly allocated to 3 groups: group I was treated with acupuncture and etoricoxib, group II with sham acupuncture and etoricoxib, and group III with etoricoxib. The primary efficacy variable was the Western Ontario and McMaster Universities (WOMAC) index and its subscales at the end of treatment at week 8. Secondary efficacy variables included the WOMAC index at the end of weeks 4 and 12, a visual analogue scale (VAS) at the end of weeks 4, 8, and 12, and the Short Form 36 version 2 (SF-36v2) health survey at the end of week 8. An algometer was used to determine changes in a predetermined unique fixed trigger point for every patient at the end of weeks 4, 8, and 12. Group I exhibited statistically significant improvements in primary and secondary outcome measures, except for Short Form mental component, compared with the other treatment groups. We conclude that acupuncture with etoricoxib is more effective than sham acupuncture with etoricoxib, or etoricoxib alone for the treatment of knee osteoarthritis.


Antigen-specific T cell responses to BK polyomavirus antigens identify functional anti-viral immunity and may help to guide immunosuppression following renal transplantation.
Infection with the polyoma virus BK (BKV) is a major cause of morbidity following renal transplantation. Limited understanding of the anti-viral immune response has prevented the design of a strategy that balances treatment with the preservation of graft function. The proven utility of interferon-gamma enzyme-linked immunospot (ELISPOT) assays to measure T cell responses in immunocompetent hosts was the basis for trying to develop a rational approach to the management of BKV following renal transplantation. In a sample of transplant recipients and healthy controls, comparisons were made between T cell responses to the complete panel of BKV antigens, the Epstein-Barr virus (EBV) antigens, BZLF1 and EBNA1, and the mitogen phytohaemagglutinin (PHA). Correlations between responses to individual antigens and immunosuppressive regimens were also analysed. Antigen-specific T cell responses were a specific indicator of recent or ongoing recovery from BKV infection (P < 0·05), with responses to different BKV antigens being highly heterogeneous. Significant BKV immunity was undetectable in transplant patients with persistent viral replication or no history of BKV reactivation. Responses to EBV antigens and mitogen were reduced in patients with BKV reactivation, but these differences were not statistically significant. The T cell response to BKV antigens is a useful and specific guide to recovery from BKV reactivation in renal transplant recipients, provided that the full range of antigenic responses is measured.


Targeting the extrinsic apoptotic pathway in cancer: lessons learned and future directions.
Apoptosis is a metazoan process of controlled cell elimination that plays critical roles in embryonic development and adult tissue homeostasis. Apoptosis dysregulation contributes to several important diseases, including cancer. Two distinct yet interconnected signaling pathways control apoptosis by activating a core intracellular machinery of death proteases called caspases. The intrinsic apoptotic pathway engages caspases via members of the BCL-2 protein family and the mitochondria in reaction to severe cellular damage or stress. The extrinsic pathway activates caspases via cell-surface death receptors, which respond to cognate death ligands expressed on immune-effector cells. Tumor cells can acquire various apoptosis-evasion mechanisms; nevertheless, the transformed state of these cells makes them uniquely susceptible to apoptosis reactivation if resistance is circumvented. Molecular approaches to reengage the apoptotic pathways in cancer have been underway for over two decades. Gratifyingly, BCL-2 antagonists - which drive the intrinsic pathway - are beginning to bear clinical fruit. In contrast, clinical attempts to stimulate the extrinsic pathway with proapoptotic receptor agonists (PARAs) have been disappointing, despite compelling preclinical efficacy with this class of agents. Here, I discuss some of the possible reasons for this translational discrepancy and suggest strategies to overcome it with the next generation of PARAs.


Strengthening the reporting of genetic risk prediction studies: the GRIPS statement.
• The rapid and continuing progress in gene discovery for complex diseases is fuelling interest in the potential application of genetic risk models for clinical and public health practice. • The number of studies assessing the predictive ability is steadily increasing, but the quality and completeness of reporting vary. • A multidisciplinary workshop sponsored by the Human Genome Epidemiology Network developed a checklist of 25 items recommended for strengthening the reporting of Genetic RIsk Prediction Studies (GRIPS), building on the principles established by prior reporting guidelines. • These recommendations aim to enhance the transparency of study reporting and thereby to improve the synthesis and application of information from multiple studies that might differ in design, conduct or analysis. • A detailed Explanation and Elaboration document is published as an accompanying article [1].


The natural history of chronic airflow obstruction.
A prospective epidemiological study of the early stages of the development of chronic obstructive pulmonary disease was performed on London working men. The findings showed that forced expiratory volume in one second (FEV1) falls gradually over a lifetime, but in most non-smokers and many smokers clinically significant airflow obstruction never develops. In susceptible people, however, smoking causes irreversible obstructive changes. If a susceptible smoker stops smoking he will not recover his lung function, but the average further rates of loss of FEV1 will revert to normal. Therefore, severe or fatal obstructive lung disease could be prevented by screening smokers' lung function in early middle age if those with reduced function could be induced to stop smoking. Infective processes and chronic mucus hypersecretion do not cause chronic airflow obstruction to progress more rapidly. There are thus two largely unrelated disease processes, chronic airflow obstruction and the hypersecretory disorder (including infective processes).


The origin, evolution, and functional impact of short insertion-deletion variants identified in 179 human genomes.
Short insertions and deletions (indels) are the second most abundant form of human genetic variation, but our understanding of their origins and functional effects lags behind that of other types of variants. Using population-scale sequencing, we have identified a high-quality set of 1.6 million indels from 179 individuals representing three diverse human populations. We show that rates of indel mutagenesis are highly heterogeneous, with 43%-48% of indels occurring in 4.03% of the genome, whereas in the remaining 96% their prevalence is 16 times lower than SNPs. Polymerase slippage can explain upwards of three-fourths of all indels, with the remainder being mostly simple deletions in complex sequence. However, insertions do occur and are significantly associated with pseudo-palindromic sequence features compatible with the fork stalling and template switching (FoSTeS) mechanism more commonly associated with large structural variations. We introduce a quantitative model of polymerase slippage, which enables us to identify indel-hypermutagenic protein-coding genes, some of which are associated with recurrent mutations leading to disease. Accounting for mutational rate heterogeneity due to sequence context, we find that indels across functional sequence are generally subject to stronger purifying selection than SNPs. We find that indel length modulates selection strength, and that indels affecting multiple functionally constrained nucleotides undergo stronger purifying selection. We further find that indels are enriched in associations with gene expression and find evidence for a contribution of nonsense-mediated decay. Finally, we show that indels can be integrated in existing genome-wide association studies (GWAS); although we do not find direct evidence that potentially causal protein-coding indels are enriched with associations to known disease-associated SNPs, our findings suggest that the causal variant underlying some of these associations may be indels.


The absence of the paternal penis.
Girls' experiences of object loss, in conjunction with female anatomical structure, may lend themselves to a particular genital anxiety regarding openness and emptiness. The relational void in giving up the mother as love object may lead to an internal self-representation of a "hole" to be filled, much as the mouth sucks the pacifier in the absence of the nipple. This image may then be extended to the genital representation. In turning to the father, a girl may find that she lacks a relationship with him in the relational space opened up by the loss of the mother; the penis is symbolically withheld from her in the father's relational distance. This lack of sexual and relational gratification, it is proposed, may be schematized by a female as her body being empty of something. The father's absence--the absence of the paternal penis--may lead to an absence of the mental representation of the vagina and to an inhibition of the role the vagina then plays for a woman in sexual desire. Vaginal repression may serve to disguise object hunger that might otherwise be experienced as vaginal longing. An abbreviated clinical vignette, revolving around a masturbatory fantasy, is offered in partial illustration of the thesis.


Blocking type I interferon signaling enhances T cell recovery and reduces HIV-1 reservoirs.
Despite the efficient suppression of HIV-1 replication that can be achieved with combined antiretroviral therapy (cART), low levels of type I interferon (IFN-I) signaling persist in some individuals. This sustained signaling may impede immune recovery and foster viral persistence. Here we report studies using a monoclonal antibody to block IFN-α/β receptor (IFNAR) signaling in humanized mice (hu-mice) that were persistently infected with HIV-1. We discovered that effective cART restored the number of human immune cells in HIV-1-infected hu-mice but did not rescue their immune hyperactivation and dysfunction. IFNAR blockade fully reversed HIV-1-induced immune hyperactivation and rescued anti-HIV-1 immune responses in T cells from HIV-1-infected hu-mice. Finally, we found that IFNAR blockade in the presence of cART reduced the size of HIV-1 reservoirs in lymphoid tissues and delayed HIV-1 rebound after cART cessation in the HIV-1-infected hu-mice. We conclude that low levels of IFN-I signaling contribute to HIV-1-associated immune dysfunction and foster HIV-1 persistence in cART-treated hosts. Our results suggest that blocking IFNAR may provide a potential strategy to enhance immune recovery and reduce HIV-1 reservoirs in individuals with sustained elevations in IFN-I signaling during suppressive cART.


Magnetic resonance imaging for the study of ovarian follicles in the mouse.
Additional tools to analyze follicle development would be highly advantageous because current methods require sacrifice of animals at specific times and time-consuming sectioning of tissues for histologic analysis. Magnetic resonance imaging (MRI) may provide a less involved, faster and more cost-effective method to analyze follicles in whole ovaries. Fixed ovaries were collected at different stages of the estrus cycle and after stimulation with gonadotrophins (24 and 48 h post pregnant mares serum (PMSG), and 10 and 24 h post human chorionic gonadotrophin (hCG)) with or without administration of the contrast agent gadodiamide. The MR images were generated using a vertical-bore, 11.7 Tesla MR system. Analysis of the MR images revealed large antral follicles in fixed ovaries with the oocyte and cumulus mass identifiable within preovulatory follicles. The use of gadodiamide had no impact on the quality of MR images obtained. The fixed ovaries were paraffin embedded, sectioned, and hematoxylin stained. Follicles were counted using the MR images and the histology sections. Preovulatory follicle numbers determined using MR images were comparable to those using histology; however counts of smaller follicles were inconsistent. MRI of gonadotrophin-stimulated ovaries in situ did not reveal discernable ovarian structures. Therefore, MRI is a useful tool for studying whole fixed ovaries leaving the ovary intact for additional analyses or for selection of samples based on morphology. The MRI is also useful for identifying preovulatory follicles, although analysis of smaller follicles is not possible, and thus the potential exists for cyst analysis in mouse models of polycystic ovarian syndrome (PCOS).


Structural studies of the O-antigen polysaccharide from Escherichia coli TD2158 having O18 serogroup specificity and aspects of its interaction with the tailspike endoglycosidase of the infecting bacteriophage HK620.
We have analyzed the O-antigen polysaccharide of the previously uncharacterized Escherichia coli strain TD2158 which is a host of bacteriophage HK620. This bacteriophage recognizes and cleaves the polysaccharide with its tailspike protein (TSP). The polysaccharide preparation as well as oligosaccharides obtained from HK620TSP endoglycosidase digests were analyzed with NMR spectroscopy. Additionally, sugar analysis was performed on the O-antigen polysaccharide and MALDI-TOF MS was used in oligosaccharide analysis. The present study revealed a heterogeneous polysaccharide with a hexasaccharide repeating unit of the following structure: α-D-Glcp-(1→6|) →2)-α-L-Rhap-91→6)-α-D-Glcp-(1→4)-α-D-Ga|lp-(1→3)-α-D-GlcpNAc-(1→ β-D-Glcp/β-D-GlcpNAc-(1→3) A repeating unit with a D-GlcNAc substitution of D-Gal has been described earlier as characteristic for serogroup O18A1. Accordingly, we termed repeating units with D-Glc substitution at D-Gal as O18A2. NMR analyses of the polysaccharide confirmed that O18A1- and O18A2-type repeats were present in a 1:1 ratio. However, HK620TSP preferentially bound the D-GlcNAc-substituted O18A1-type repeating units in its high affinity binding pocket with a dissociation constant of 140 μM and disfavored the O18A2-type having a β-D-Glcp-(1→3)-linked group. As a result, in hexasaccharide preparations, O18A1 and O18A2 repeats were present in a 9:1 ratio stressing the clear preference of O18A1-type repeats to be cleaved by HK620TSP.


Alcohol drinking, tobacco smoking and subtypes of haematological malignancy in the UK Million Women Study.
Previous research suggests associations of lower alcohol intake and higher tobacco consumption with increased risks of haematological malignancy. The prospective Million Women Study provides sufficient power for reliable estimates of subtype-specific associations in women. Approximately 1.3 million middle-aged women were recruited in the United Kingdom during 1996-2001 and followed for death, emigration and cancer registration until 2009 (mean 10.3 years per woman); potential risk factors were assessed by questionnaire. Adjusted relative risks were estimated by Cox regression. During follow-up, 9162 incident cases of haematological malignancy were recorded, including 7047 lymphoid and 2072 myeloid cancers. Among predominantly moderate alcohol drinkers, higher intake was associated with lower risk of lymphoid malignancies, in particular diffuse large B-cell lymphoma (relative risk 0.85 per 10 g alcohol per day (95% confidence interval 0.75-0.96)), follicular lymphoma (0.86 (0.76-0.98)) and plasma cell neoplasms (0.86 (0.77-0.96)). Among never- and current smokers, higher cigarette consumption was associated with increased risk of Hodgkin lymphoma (1.45 per 10 cigarettes per day (1.22-1.72)), mature T-cell malignancies (1.38 (1.10-1.73)) and myeloproliferative/myelodysplastic disease (1.42 (1.31-1.55)). These findings confirm and extend existing evidence for associations of subtypes of haematological malignancy with two common exposures in women.


Redistribution of HIV outside the lymphoid system with onset of AIDS.
The basis for many of the symptoms and pathological changes found in patients with the acquired immunodeficiency syndrome (AIDS) remains poorly understood. We have used a quantitative polymerase chain reaction technique to investigate the extent to which direct infection with human immunodeficiency virus (HIV) produces the disease manifestations of AIDS. In five patients who died with AIDS-defining illnesses (Centers for Disease Control and Prevention class IV), we found variable, but in many cases extensive, infection by HIV at various sites, including brain, lung, colon, and liver. By contrast, in three HIV-positive subjects who died without HIV-related disease (CDC class II), we found no evidence of significant infection of any non-lymphoid organ. In both groups of patients there were high levels of infection in cells of the spleen, lymph nodes, and peripheral blood. Pathological examination of tissues from the AIDS patients revealed many abnormalities, of which some, such as giant-cell encephalitis in the brain, were specifically associated with the presence of high levels of HIV infection. These findings suggest that spread of HIV outside cells of the immune system is a late event in HIV infection and is extremely sensitive to the degree of immunosuppression in the patient.


A differential genome-wide transcriptome analysis: impact of cellular copper on complex biological processes like aging and development.
The regulation of cellular copper homeostasis is crucial in biology. Impairments lead to severe dysfunctions and are known to affect aging and development. Previously, a loss-of-function mutation in the gene encoding the copper-sensing and copper-regulated transcription factor GRISEA of the filamentous fungus Podospora anserina was reported to lead to cellular copper depletion and a pleiotropic phenotype with hypopigmentation of the mycelium and the ascospores, affected fertility and increased lifespan by approximately 60% when compared to the wild type. This phenotype is linked to a switch from a copper-dependent standard to an alternative respiration leading to both a reduced generation of reactive oxygen species (ROS) and of adenosine triphosphate (ATP). We performed a genome-wide comparative transcriptome analysis of a wild-type strain and the copper-depleted grisea mutant. We unambiguously assigned 9,700 sequences of the transcriptome in both strains to the more than 10,600 predicted and annotated open reading frames of the P. anserina genome indicating 90% coverage of the transcriptome. 4,752 of the transcripts differed significantly in abundance with 1,156 transcripts differing at least 3-fold. Selected genes were investigated by qRT-PCR analyses. Apart from this general characterization we analyzed the data with special emphasis on molecular pathways related to the grisea mutation taking advantage of the available complete genomic sequence of P. anserina. This analysis verified but also corrected conclusions from earlier data obtained by single gene analysis, identified new candidates of factors as part of the cellular copper homeostasis system including target genes of transcription factor GRISEA, and provides a rich reference source of quantitative data for further in detail investigations. Overall, the present study demonstrates the importance of systems biology approaches also in cases were mutations in single genes are analyzed to explain the underlying mechanisms controlling complex biological processes like aging and development.


Fluorescent labeling of COS-7 expressing SNAP-tag fusion proteins for live cell imaging.
SNAP-tag and CLIP-tag protein labeling systems enable the specific, covalent attachment of molecules, including fluorescent dyes, to a protein of interest in live cells. These systems offer a broad selection of fluorescent substrates optimized for a range of imaging instrumentation. Once cloned and expressed, the tagged protein can be used with a variety of substrates for numerous downstream applications without having to clone again. There are two steps to using this system: cloning and expression of the protein of interest as a SNAP-tag fusion, and labeling of the fusion with the SNAP-tag substrate of choice. The SNAP-tag is a small protein based on human O(6)-alkylguanine-DNA-alkyltransferase (hAGT), a DNA repair protein. SNAP-tag labels are dyes conjugated to guanine or chloropyrimidine leaving groups via a benzyl linker. In the labeling reaction, the substituted benzyl group of the substrate is covalently attached to the SNAP-tag. CLIP-tag is a modified version of SNAP-tag, engineered to react with benzylcytosine rather than benzylguanine derivatives. When used in conjunction with SNAP-tag, CLIP-tag enables the orthogonal and complementary labeling of two proteins simultaneously in the same cells.


Genetic analysis of 7 medieval skeletons from the Aragonese Pyrenees.
To perform a genetic characterization of 7 skeletons from medieval age found in a burial site in the Aragonese Pyrenees. Allele frequencies of autosomal short tandem repeats (STR) loci were determined by 3 different STR systems. Mitochondrial DNA (mtDNA) and Y-chromosome haplogroups were determined by sequencing of the hypervariable segment 1 of mtDNA and typing of phylogenetic Y chromosome single nucleotide polymorphisms (Y-SNP) markers, respectively. Possible familial relationships were also investigated. Complete or partial STR profiles were obtained in 3 of the 7 samples. Mitochondrial DNA haplogroup was determined in 6 samples, with 5 of them corresponding to the haplogroup H and 1 to the haplogroup U5a. Y-chromosome haplogroup was determined in 2 samples, corresponding to the haplogroup R. In one of them, the sub-branch R1b1b2 was determined. mtDNA sequences indicated that some of the individuals could be maternally related, while STR profiles indicated no direct family relationships. Despite the antiquity of the samples and great difficulty that genetic analyses entail, the combined use of autosomal STR markers, Y-chromosome informative SNPs, and mtDNA sequences allowed us to genotype a group of skeletons from the medieval age.


An unusual case of an ACTH-secreting macroadenoma with a germline variant in the aryl hydrocarbon receptor-interacting protein (AIP) gene.
A patient of Cushing's disease (CD) characterized by a large tumor and only subtle symptoms of hormonal hypersecretion was examined. The patient had a germline variant in the aryl hydrocarbon receptor-interacting protein (AIP) gene. A 50-year-old male presenting with headache was diagnosed with a large pituitary tumor by magnetic resonance imaging (MRI). His visual fields were intact and he exhibited no features of CD. Owing to an exuberant response to synacthen, an overnight dexamethasone suppression test was performed revealing inadequate suppression of plasma cortisol (419 nmol/l). Owing to tumor growth and visual field impairment, he underwent transsphenoidal surgery and developed hypocortisolemia. The pathology specimen revealed a sparsely granulated corticotrope adenoma. Postoperative MRI showed a large tumor remnant. The patient developed skin hyperpigmentation and a synacthen test demonstrated high basal and stimulated cortisol levels; an overnight dexamethasone suppression test showed no suppression (791 nmol/l) and elevated plasma ACTH levels (135 ng/l). A transcranial operation was performed followed by radiotherapy. Two months after radiotherapy, he developed secondary adrenocortical failure. Genetic testing revealed an AIP variant of unknown significance (p.R16H) without loss of the normal AIP allele in the tumor. A literature review showed ten CD patients with AIP gene variants, of whom five (including our case) were p.R16H. CD is occasionally dominated by pituitary tumor growth rather than symptoms of hypersecretion. The particular AIP gene variant identified in our patient is shared by four other reported cases of CD. Future studies are needed to assess whether the reported AIP gene variant is more than just coincidental. CD is occasionally dominated by pituitary tumor growth rather than symptoms of hypersecretion.Resolution of both tumor remnant and hormonal hypersecretion may occur within 2 months after postoperative radiotherapy.The particular AIP gene variant identified in our patient is shared by four other reported cases of CD.


Energy Transfer Between Squaraine Polymer Sections: From Helix to Zigzag and All the Way Back.
We provide a joint experimental and theoretical study of squaraine polymers in solution. The absorption spectra show evidence that two different conformations are present in the polymer: a helix and a zigzag structure. This unique situation allows investigating ultrafast energy-transfer processes between different structural segments within a single polymer chain in solution. The understanding of the underlying dynamics is of fundamental importance for the development of novel materials for light-harvesting and optoelectronic applications. Here, we combine femtosecond transient absorption spectroscopy with time-resolved 2D electronic spectroscopy in order to demonstrate that ultrafast energy transfer within the squaraine polymer chains proceeds from initially excited helix segments to zigzag segments or vice versa, depending on the solvent as well as on the excitation wavenumber. These observations contrast other conjugated polymers such as MEH-PPV where much slower intrachain energy transfer was reported. The reason for the very fast energy transfer in squaraine polymers is most likely a close matching of the density of states between donor and acceptor polymer segments because of the very small reorganization energy in these cyanine-like chromophores.


Neuropsychological functioning and jumping to conclusions in delusions.
It has been consistently demonstrated that delusions are related to jumping to conclusions (JTC), a data-gathering bias and potential candidate endophenotype of psychosis. Recent research suggests that JTC may be a marker of treatment response. However, we know little about the factors contributing to the occurrence of this reasoning bias. This study investigated the relationship between JTC and hypothesised deficits in working memory, employing standard well-validated neuropsychological tests, in people with current delusions. One hundred and twenty six people with schizophrenia spectrum psychosis and current delusions were assessed for current symptoms, and tested for JTC. We compared performance on tests of working memory in those with the reasoning bias and those without. As expected, 30-40% of this sample of people with current delusions showed the JTC bias. There were no differences in premorbid IQ between those with and without the JTC reasoning bias. However, the performance of the JTC group was significantly worse on tests of working memory. The JTC data-gathering bias is associated with impairments in working memory. New non-pharmacological interventions for people with delusions, designed to improve data gathering, may benefit from incorporating strategies to overcome deficits in working memory.


Reducing teachers' psychological distress through a mindfulness training program.
Teachers constitute one of the professional collectives most affected by psychological problems. The purpose of this quasi-experimental study is to examine the efficacy of a mindfulness training programme to reduce psychological distress in a group of teachers. The sample comprised 68 teachers of Secondary School Education, from various public schools; half of them formed the experimental group, and the another half the control group. The levels of psychological distress were measured, in both groups, by the Symptom Checklist-90-R (SCL-90-R) before and after the application of the programme. Statistical analysis shows the significant reduction of three general measures of psychological distress (Global Severity Index, Positive Symptom Distress Index, and Positive Symptom Total), as well in all its dimensions (somatization, obsessive-compulsive, interpersonal sensibility, depression, anxiety, hostility, phobic anxiety, paranoid ideation, and psychoticism), in the experimental group compared with the control group. Follow-up measures show that these results were maintained for four months after termination of the intervention in the experimental group.