Cannabidiol for the treatment of psychosis in Parkinson's disease.
The management of psychosis in Parkinson's disease (PD) has been considered a great challenge for clinicians and there is a need for new pharmacological intervention. Previously an antipsychotic and neuroprotective effect of Cannabidiol (CBD) has been suggested. Therefore, the aim of the present study was to directly evaluate for the first time, the efficacy, tolerability and safety of CBD on PD patients with psychotic symptoms. This was an open-label pilot study. Six consecutive outpatients (four men and two women) with the diagnosis of PD and who had psychosis for at least 3 months were selected for the study. All patients received CBD in flexible dose (started with an oral dose of 150 mg/day) for 4 weeks, in addition to their usual therapy. The psychotic symptoms evaluated by the Brief Psychiatric Rating Scale and the Parkinson Psychosis Questionnaire showed a significant decrease under CBD treatment. CBD did not worsen the motor function and decreased the total scores of the Unified Parkinson's Disease Rating Scale. No adverse effect was observed during the treatment. These preliminary data suggest that CBD may be effective, safe and well tolerated for the treatment of the psychosis in PD.


Manganese neurotoxicity and glutamate-GABA interaction.
Brain extracellular concentrations of amino acids (e.g. aspartate, glutamate, taurine) and divalent metals (e.g. zinc, copper, manganese) are primarily regulated by astrocytes. Adequate glutamate homeostasis is essential for the normal functioning of the central nervous system (CNS). Glutamate is of central importance for nitrogen metabolism and, along with aspartate, is the primary mediator of the excitatory pathways in the brain. Similarly, the maintenance of proper manganese levels is important for normal brain functioning. Several in vivo and in vitro studies have linked increased manganese concentrations with alterations in the content and metabolism of neurotransmitters, namely dopamine, gamma-aminobutyric acid, and glutamate. It has been reported by our laboratory and others, that cultured rat primary astrocytes exposed to manganese displayed decreased glutamate uptake, thereby increasing the excitotoxic potential of glutamate. Furthermore, decreased uptake of glutamate has been associated with decreased gene expression of glutamate:aspartate transporter (GLAST) in manganese-exposed astroctyes. Additional studies have suggested that attenuation of astrocytic glutamate uptake by manganese may be a consequence of reactive oxygen species (ROS) generation. Collectively, these data suggest that excitotoxicity may occur due to manganese-induced altered glutamate metabolism, representing a proximate mechanism for manganese-induced neurotoxicity.


Efficacy of artesunate-amodiaquine for treating uncomplicated falciparum malaria in sub-Saharan Africa: a multi-centre analysis.
Artesunate and amodiaquine (AS&AQ) is at present the world's second most widely used artemisinin-based combination therapy (ACT). It was necessary to evaluate the efficacy of ACT, recently adopted by the World Health Organization (WHO) and deployed over 80 countries, in order to make an evidence-based drug policy. An individual patient data (IPD) analysis was conducted on efficacy outcomes in 26 clinical studies in sub-Saharan Africa using the WHO protocol with similar primary and secondary endpoints. A total of 11,700 patients (75% under 5 years old), from 33 different sites in 16 countries were followed for 28 days. Loss to follow-up was 4.9% (575/11,700). AS&AQ was given to 5,897 patients. Of these, 82% (4,826/5,897) were included in randomized comparative trials with polymerase chain reaction (PCR) genotyping results and compared to 5,413 patients (half receiving an ACT). AS&AQ and other ACT comparators resulted in rapid clearance of fever and parasitaemia, superior to non-ACT. Using survival analysis on a modified intent-to-treat population, the Day 28 PCR-adjusted efficacy of AS&AQ was greater than 90% (the WHO cut-off) in 11/16 countries. In randomized comparative trials (n = 22), the crude efficacy of AS&AQ was 75.9% (95% CI 74.6-77.1) and the PCR-adjusted efficacy was 93.9% (95% CI 93.2-94.5). The risk (weighted by site) of failure PCR-adjusted of AS&AQ was significantly inferior to non-ACT, superior to dihydroartemisinin-piperaquine (DP, in one Ugandan site), and not different from AS+SP or AL (artemether-lumefantrine). The risk of gametocyte appearance and the carriage rate of AS&AQ was only greater in one Ugandan site compared to AL and DP, and lower compared to non-ACT (p = 0.001, for all comparisons). Anaemia recovery was not different than comparator groups, except in one site in Rwanda where the patients in the DP group had a slower recovery. AS&AQ compares well to other treatments and meets the WHO efficacy criteria for use against falciparum malaria in many, but not all, the sub-Saharan African countries where it was studied. Efficacy varies between and within countries. An IPD analysis can inform general and local treatment policies. Ongoing monitoring evaluation is required.


Cardiovascular regulation profile predicts developmental trajectory of BMI and pediatric obesity.
The present study examined the role of cardiovascular regulation in predicting pediatric obesity. Participants for this study included 268 children (141 girls) obtained from a larger ongoing longitudinal study. To assess cardiac vagal regulation, resting measures of respiratory sinus arrhythmia (RSA) and RSA change (vagal withdrawal) to three cognitively challenging tasks were derived when children were 5.5 years of age. Heart period (HP) and HP change (heart rate (HR) acceleration) were also examined. Height and weight measures were collected when children were 5.5, 7.5, and 10.5 years of age. Results indicated that physiological regulation at age 5.5 was predictive of both normal variations in BMI development and pediatric obesity at age 10.5. Specifically, children with a cardiovascular regulation profile characterized by lower levels of RSA suppression and HP change experienced significantly greater levels of BMI growth and were more likely to be classified as overweight/at-risk for overweight at age 10.5 compared to children with a cardiovascular regulation profile characterized by high levels of RSA suppression and HP change. However, a significant interaction with racial status was found suggesting that the association between cardiovascular regulation profile and BMI growth and pediatric obesity was only significant for African-American children. An autonomic cardiovascular regulation profile consisting of low parasympathetic activity represents a significant individual risk factor for the development of pediatric obesity, but only for African-American children. Mechanisms by which early physiological regulation difficulties may contribute to the development of pediatric obesity are discussed.


P2P proteomics -- data sharing for enhanced protein identification.
In order to tackle the important and challenging problem in proteomics of identifying known and new protein sequences using high-throughput methods, we propose a data-sharing platform that uses fully distributed P2P technologies to share specifications of peer-interaction protocols and service components. By using such a platform, information to be searched is no longer centralised in a few repositories but gathered from experiments in peer proteomics laboratories, which can subsequently be searched by fellow researchers. The system distributively runs a data-sharing protocol specified in the Lightweight Communication Calculus underlying the system through which researchers interact via message passing. For this, researchers interact with the system through particular components that link to database querying systems based on BLAST and/or OMSSA and GUI-based visualisation environments. We have tested the proposed platform with data drawn from preexisting MS/MS data reservoirs from the 2006 ABRF (Association of Biomolecular Resource Facilities) test sample, which was extensively tested during the ABRF Proteomics Standards Research Group 2006 worldwide survey. In particular we have taken the data available from a subset of proteomics laboratories of Spain's National Institute for Proteomics, ProteoRed, a network for the coordination, integration and development of the Spanish proteomics facilities. We performed queries against nine databases including seven ProteoRed proteomics laboratories, the NCBI Swiss-Prot database and the local database of the CSIC/UAB Proteomics Laboratory. A detailed analysis of the results indicated the presence of a protein that was supported by other NCBI matches and highly scored matches in several proteomics labs. The analysis clearly indicated that the protein was a relatively high concentrated contaminant that could be present in the ABRF sample. This fact is evident from the information that could be derived from the proposed P2P proteomics system, however it is not straightforward to arrive to the same conclusion by conventional means as it is difficult to discard organic contamination of samples. The actual presence of this contaminant was only stated after the ABRF study of all the identifications reported by the laboratories.


Original antigenic sin and apoptosis in the pathogenesis of dengue hemorrhagic fever.
Dengue virus presents a growing threat to public health in the developing world. Four major serotypes of dengue virus have been characterized, and epidemiological evidence shows that dengue hemorrhagic fever (DHF), the more serious manifestation of the disease, occurs more frequently upon reinfection with a second serotype. We have studied dengue virus-specific T-cell responses in Thai children. During acute infection, few dengue-responsive CD8+ T cells were recovered; most of those present showed an activated phenotype and were undergoing programmed cell death. Many dengue-specific T cells were of low affinity for the infecting virus and showed higher affinity for other, probably previously encountered strains. Profound T-cell activation and death may contribute to the systemic disturbances leading to DHF, and original antigenic sin in the T-cell responses may suppress or delay viral elimination, leading to higher viral loads and increased immunopathology.


Global and local fMRI signals driven by neurons defined optogenetically by type and wiring.
Despite a rapidly-growing scientific and clinical brain imaging literature based on functional magnetic resonance imaging (fMRI) using blood oxygenation level-dependent (BOLD) signals, it remains controversial whether BOLD signals in a particular region can be caused by activation of local excitatory neurons. This difficult question is central to the interpretation and utility of BOLD, with major significance for fMRI studies in basic research and clinical applications. Using a novel integrated technology unifying optogenetic control of inputs with high-field fMRI signal readouts, we show here that specific stimulation of local CaMKIIalpha-expressing excitatory neurons, either in the neocortex or thalamus, elicits positive BOLD signals at the stimulus location with classical kinetics. We also show that optogenetic fMRI (of MRI) allows visualization of the causal effects of specific cell types defined not only by genetic identity and cell body location, but also by axonal projection target. Finally, we show that of MRI within the living and intact mammalian brain reveals BOLD signals in downstream targets distant from the stimulus, indicating that this approach can be used to map the global effects of controlling a local cell population. In this respect, unlike both conventional fMRI studies based on correlations and fMRI with electrical stimulation that will also directly drive afferent and nearby axons, this of MRI approach provides causal information about the global circuits recruited by defined local neuronal activity patterns. Together these findings provide an empirical foundation for the widely-used fMRI BOLD signal, and the features of of MRI define a potent tool that may be suitable for functional circuit analysis as well as global phenotyping of dysfunctional circuitry.


Cytotoxicity of ketamine, xylazine and Hellabrunn mixture in liver-, heart- and kidney-derived cells from fallow deer.
Chemical restraint of wild animals is practiced to accomplish intended procedures such as capture, clinical examination, collection of diagnostic samples, treatment and/or transport. Extra-label use of animal medicinal drugs is often necessary in wildlife because most approved therapeutics do not list wild species on the labelling. Here, we used cellular in vitro models, a cutting-edge tool of biomedical research, to examine cytotoxicity of anaesthetic agents in fallow deer and extrapolate these data for anaesthetic risks in wildlife. We examined the cytotoxic effects of ketamine, xylazine, and ketamine-xylazine, i.e. the Hellabrunn mixture, on liver-, heart- and kidney-derived cell cultures prepared from a fallow deer (Dama dama) specimen. In line with preliminary studies we exposed cells to 10 µM, 50 µM, 100 µM, 1 mM, and 10 mM ketamine or xylazine. The combination of ketamine-xylazine was dosed at 0.025+0.02 mg/ml, 0.05+0.04 mg/ml, 0.75+0.06 mg/ml, 0.1+0.08 mg/ml, and 0.125+0.1 mg/ml per one well containing 10 000 cells. The quantification of cytotoxicity was based on lactate dehydrogenase activity released from damaged cells. Liver-derived cells show higher sensitivity to the cytotoxic effects of both ketamine and xylazine administered as single drugs when compared with cells cultured from the heart and kidney. The Hellabrunn mixture induced significantly higher cytotoxicity for kidney-derived cells ranging from 16.78% to 35.6%. Single and combined exposures to ketamine and xylazine resulted only in high-dose cytotoxicity in the heart-derived cells. Our results indicate that immobilization drugs significantly differ in their cytotoxic effects on cells derived from various organs of the fallow deer.


Therapy for Duchenne muscular dystrophy: renewed optimism from genetic approaches.
Duchenne muscular dystrophy (DMD) is a devastating progressive disease for which there is currently no effective treatment except palliative therapy. There are several promising genetic approaches, including viral delivery of the missing dystrophin gene, read-through of translation stop codons, exon skipping to restore the reading frame and increased expression of the compensatory utrophin gene. The lessons learned from these approaches will be applicable to many other disorders.


ELSID-Diabetes study-evaluation of a large scale implementation of disease management programmes for patients with type 2 diabetes. Rationale, design and conduct--a study protocol [ISRCTN08471887].
Diabetes model projects in different regions of Germany including interventions such as quality circles, patient education and documentation of medical findings have shown improvements of HbA1c levels, blood pressure and occurrence of hypoglycaemia in before-after studies (without control group). In 2002 the German Ministry of Health defined legal regulations for the introduction of nationwide disease management programs (DMP) to improve the quality of care in chronically ill patients. In April 2003 the first DMP for patients with type 2 diabetes was accredited. The evaluation of the DMP is essential and has been made obligatory in Germany by the Fifth Book of Social Code. The aim of the study is to assess the effectiveness of DMP by example of type 2 diabetes in the primary care setting of two German federal states (Rheinland-Pfalz and Sachsen-Anhalt). The study is three-armed: a prospective cluster-randomized comparison of two interventions (DMP 1 and DMP 2) against routine care without DMP as control group. In the DMP group 1 the patients are treated according to the current situation within the German-Diabetes-DMP. The DMP group 2 represents diabetic care within ideally implemented DMP providing additional interventions (e.g. quality circles, outreach visits). According to a sample size calculation a sample size of 200 GPs (each GP including 20 patients) will be required for the comparison of DMP 1 and DMP 2 considering possible drop-outs. For the comparison with routine care 4000 patients identified by diabetic tracer medication and age (> 50 years) will be analyzed. This study will evaluate the effectiveness of the German Diabetes-DMP compared to a Diabetes-DMP providing additional interventions and routine care in the primary care setting of two different German federal states.


Lifestyle and nutritional determinants of bioavailable androgens and related hormones in British men.
To investigate the lifestyle and nutritional determinants of serum bioavailable androgens and their related hormones in men. This study is based on a sample of 696 men with a wide range of nutrient intakes, whose diet and lifestyle characteristics were assessed with a questionnaire and serum sex hormones measured using immunoassays. Men aged 70 years or older had 12% lower testosterone and 40% lower free-testosterone (FT) and androstanediol glucuronide (A-diol-g) concentrations than men who were 20-29 years of age. Conversely, sex hormone-binding globulin (SHBG) and luteinizing hormone (LH) concentrations were 90% and 49% higher in the oldest age group compared with the lowest, respectively. Men who had a body mass index (BMI) of 30+ kg/m2 had 30% lower testosterone, 45% lower SHBG, 22% lower LH and 5% lower FT concentrations compared with men with a BMI of <20 kg/m2. Conversely, A-diol-g concentration was 15% higher in the highest BMI category compared with the lowest. A high waist circumference was further associated with a 12% lower testosterone and SHBG concentration, after adjusting for BMI. Compared with never-smokers, smoking 10+ cigarettes/day was associated with 15% higher testosterone, 22% higher SHBG and 17% higher LH concentrations; FT and A-diol-g were not associated with smoking. Compared with no exercise, vigorous exercise of 3+ hours/week was associated with 11% higher testosterone and 16% higher SHBG concentrations, whilst LH, FT, and A-diol-g were not associated with vigorous exercise. Dietary factors were not strongly associated with hormones, although saturated fat intake was negatively associated with SHBG (r = -0.10; p = 0.01) and alcohol intake was positively associated with A-diol-g (r = 0.11; p = 0.004). No dietary factors were associated with testosterone, FT, or LH. Age is the strongest determinant of serum bioavailable androgens. BMI and some lifestyle and dietary factors influence SHBG and testosterone concentrations, but have no strong association with FT, suggesting that homeostasis is effective. A-diol-g shows broadly similar associations to FT, with the exception of the effect of BMI and alcohol.


A model of motor neuron loss: selective deficits after ricin injection.
This study characterizes a model of motor neuron (MN) loss on the molecular, cellular, and behavioral levels. Injection of the toxic lectin Ricinus communis agglutinin I (RCA I or ricin) caused cellular deficit and loss of function by damaging the sciatic nerve. Since the sciatic nerve supplies movement to most of the lower limb, damaging this motor system models lower limb paralysis and the deficits that occur in diseases like amyotrophic lateral sclerosis (ALS) and infantile progressive spinal muscular atrophy (SMA). We used motor-, sensorimotor-, locomotor-, and reflex-based tests to demonstrate loss of function after ricin injection. Loss of function was also demonstrated by decreased retrograde transport, and supported by measurements of muscle wasting. Histochemical and molecular methods were used to characterize sciatic nerve damage in axons and cell bodies, including apoptotic cell death in MNs. This battery of tests documents the extent of the ricin-induced damage and provides a baseline that can be used to judge the efficacy of MN treatment strategies in preclinical studies.


Prospectively determined impact of type 1 diabetes on brain volume during development.
The impact of type 1 diabetes mellitus (T1DM) on the developing central nervous system is not well understood. Cross-sectional, retrospective studies suggest that exposure to glycemic extremes during development is harmful to brain structure in youth with T1DM. However, these studies cannot identify brain regions that change differentially over time depending on the degree of exposure to glycemic extremes. We performed a longitudinal, prospective structural neuroimaging study of youth with T1DM (n = 75; mean age = 12.5 years) and their nondiabetic siblings (n = 25; mean age = 12.5 years). Each participant was scanned twice, separated by 2 years. Blood glucose control measurements (HbA(1c), glucose meter results, and reports of severe hypoglycemia) were acquired during the 2-year follow-up. Sophisticated image registration algorithms were performed, followed by whole brain and voxel-wise statistical analyses of the change in gray and white matter volume, controlling for age, sex, and age of diabetes onset. The T1DM and nondiabetic control (NDC) sibling groups did not differ in whole brain or voxel-wise change over the 2-year follow-up. However, within the T1DM group, participants with more hyperglycemia had a greater decrease in whole brain gray matter compared with those with less hyperglycemia (P < 0.05). Participants who experienced severe hypoglycemia had greater decreases in occipital/parietal white matter volume compared with those with no severe hypoglycemia (P < 0.05) and compared with the NDC sibling group (P < 0.05). These results demonstrate that within diabetes, exposure to hyperglycemia and severe hypoglycemia may result in subtle deviation from normal developmental trajectories of the brain.


Spin-history artifact during functional MRI: potential for adaptive correction.
Functional magnetic resonance imaging (fMRI) is limited by sensitivity to millimetre-scale head motion. Adaptive correction is a strategy to adjust the imaging plane in response to measured head motion, thereby suppressing motion artifacts. This strategy should correct for motion in all six degrees of freedom and also holds promise for through-plane motion that creates "spin-history" artifact that cannot easily be removed by postprocessing methods. Improved quantitative understanding of the MRI signal behavior associated with spin-history artifact would be useful for implementing adaptive correction robustly. A numerical simulation was developed to predict MRI artifact signal amplitude in a single-slice for simple motions, implemented with and without adaptive correction, and compared with experiment by imaging a phantom at 3.0 T. Functional MRI was also performed of a human volunteer to illustrate adaptive correction in the presence of spin-history artifact. Good agreement was achieved between simulation and experimental results. Although time-averaged artifact signal amplitude was observed to correlate linearly with motion speed, artifact time-courses were nonlinearly related to motion waveforms. In addition, experimental results demonstrated effective adaptive correction of spin-history artifact when the phantom underwent complex motions. Adaptive correction during human fMRI suppressed spin-history artifacts and spurious activations associated with task-correlated motion. Overall, this work suggests that adaptive correction, especially when implemented with minimal lag between motion measurement and scan plane update, may help to expand the populations for which fMRI can be performed robustly.


Low-dose aspirin, coxibs, and other NSAIDS: a clinical mosaic emerges.
Aspirin has been a commercial drug for over a century, although for most of this history, an understanding of its mechanism of action, as an inhibitor of cyclooxygenase (COX) activity and thus of prostanoid synthesis, was lacking. Over the past fifty years, a large number of other nonsteroidal antiinflammatory drugs (NSAIDs) have been developed, and a much deeper understanding of inflammation and prostanoid action has emerged. Indeed, a new class of selective inhibitors of the cyclooxygenase-2 isozyme was introduced, about ten years ago, and these so-called coxibs quickly became regarded as preferable, in certain clinical contexts, to avoid side effects associated with the use of aspirin and previously developed NSAIDs. This regard for coxibs has been challenged, sometimes infamously, as cardiovascular events associated with coxib use have become apparent. A variety of clinical trials have led to seemingly conflicting data concerning the roles of COX-1 and COX-2, and the implications of their relative inhibition, in cardiovascular health and disease. In this Review, the authors offer an assessment of drug pharmacokinetics and enzyme physiology that reconciles cardiovascular appraisals from a wide array of clinical data.


Measles virus glycoprotein-based lentiviral targeting vectors that avoid neutralizing antibodies.
Lentiviral vectors (LVs) are potent gene transfer vehicles frequently applied in research and recently also in clinical trials. Retargeting LV entry to cell types of interest is a key issue to improve gene transfer safety and efficacy. Recently, we have developed a targeting method for LVs by incorporating engineered measles virus (MV) glycoproteins, the hemagglutinin (H), responsible for receptor recognition, and the fusion protein into their envelope. The H protein displays a single-chain antibody (scFv) specific for the target receptor and is ablated for recognition of the MV receptors CD46 and SLAM by point mutations in its ectodomain. A potential hindrance to systemic administration in humans is pre-existing MV-specific immunity due to vaccination or natural infection. We compared transduction of targeting vectors and non-targeting vectors pseudotyped with MV glycoproteins unmodified in their ectodomains (MV-LV) in presence of α-MV antibody-positive human plasma. At plasma dilution 1:160 MV-LV was almost completely neutralized, whereas targeting vectors showed relative transduction efficiencies from 60% to 90%. Furthermore, at plasma dilution 1:80 an at least 4-times higher multiplicity of infection (MOI) of MV-LV had to be applied to obtain similar transduction efficiencies as with targeting vectors. Also when the vectors were normalized to their p24 values, targeting vectors showed partial protection against α-MV antibodies in human plasma. Furthermore, the monoclonal neutralizing antibody K71 with a putative epitope close to the receptor binding sites of H, did not neutralize the targeting vectors, but did neutralize MV-LV. The observed escape from neutralization may be due to the point mutations in the H ectodomain that might have destroyed antibody binding sites. Furthermore, scFv mediated cell entry via the target receptor may proceed in presence of α-MV antibodies interfering with entry via the natural MV receptors. These results are promising for in vivo applications of targeting vectors in humans.


Noninvasive associative plasticity induction in a corticocortical pathway of the human brain.
Coincident pairing of presynaptic and postsynaptic activity selectively strengthens synaptic connections, a key mechanism underlying cortical plasticity. Using paired associative transcranial magnetic stimulation (TMS), we demonstrate selective potentiation of physiological connectivity between two human brain regions, ventral premotor cortex (PMv) and primary motor cortex (M1) after repeated paired-pulse TMS of PMv and M1. The effect was anatomically specific: paired stimulation of the presupplementary motor area and M1 did not induce changes in PMv-M1 pathway connectivity. The effect was dependent on stimulation order: repeated stimulation of PMv before M1 led to strengthening of the PMv-M1 pathway, while repeated stimulation of M1 before PMv diminished the strength of the PMv-M1 pathway. The expression of the change in the pathway depended on the cognitive state of the subject at the time of testing: when the subject was tested at rest, paired PMv-M1 stimulation led to an increased inhibitory influence of PMv over M1, but when the subject was tested while engaged in a visuomotor task, PMv-M1 stimulation led to an increased facilitatory influence of PMv over M1. Plasticity evolved rapidly, lasted for at least 1 h, and began to reverse 3 h after intervention.


Phylogenetic relationship of the populations within and around Japan using 105 short tandem repeat polymorphic loci.
We have analyzed 105 autosomal polymorphic short tandem repeat (STR) loci for nine East and South-eastern Asian populations (two Japanese, five Han Chinese, Thai, and Burmese populations) and a Caucasian population using a multiplex PCR typing system. All the STR loci are genomewide tetranucleotide repeat markers of which the total number of observed alleles and the observed heterozygosity were 756 and 0.743, respectively, for Japanese populations. Phylogenetic analysis for these allele frequency data suggested that the Japanese populations are more closely related with southern Chinese populations than central and/or northern ones. STRUCTURE program analysis revealed the almost clearly divided and accountable population structure at K=2-6, that the two Japanese populations always formed one group separated from the other populations and never belong to different groups at K> or =3. Furthermore, our new allele frequency data for 91 loci were analyzed with those for 52 worldwide populations published by previous studies. Phylogenetic and multidimensional scaling (MDS) analyses indicated that Asian populations with large population size (six Han Chinese, three Japanese, two Southeast Asia) formed one distinct cluster and are closer to each other than other ethnic minorities in east and Southeast Asia. This pattern may be the caviar of comparing populations with greatly differing population sizes when STR loci were analyzed.


The harmonic mean p-value for combining dependent tests.
Analysis of "big data" frequently involves statistical comparison of millions of competing hypotheses to discover hidden processes underlying observed patterns of data, for example, in the search for genetic determinants of disease in genome-wide association studies (GWAS). Controlling the familywise error rate (FWER) is considered the strongest protection against false positives but makes it difficult to reach the multiple testing-corrected significance threshold. Here, I introduce the harmonic mean p-value (HMP), which controls the FWER while greatly improving statistical power by combining dependent tests using generalized central limit theorem. I show that the HMP effortlessly combines information to detect statistically significant signals among groups of individually nonsignificant hypotheses in examples of a human GWAS for neuroticism and a joint human-pathogen GWAS for hepatitis C viral load. The HMP simultaneously tests all ways to group hypotheses, allowing the smallest groups of hypotheses that retain significance to be sought. The power of the HMP to detect significant hypothesis groups is greater than the power of the Benjamini-Hochberg procedure to detect significant hypotheses, although the latter only controls the weaker false discovery rate (FDR). The HMP has broad implications for the analysis of large datasets, because it enhances the potential for scientific discovery.


Managing misaligned paternity findings in research including sickle cell disease screening in Kenya: 'consulting communities' to inform policy.
The management of misaligned paternity findings raises important controversy worldwide. It has mainly, however, been discussed in the context of high-income countries. Genetic and genomics research, with the potential to show misaligned paternity, are becoming increasingly common in Africa. During a genomics study in Kenya, a dilemma arose over testing and sharing information on paternal sickle cell disease status. This dilemma may be paradigmatic of challenges in sharing misaligned paternity findings in many research and health care settings. Using a deliberative approach to community consultation to inform research practice, we explored residents' views on paternal testing and sharing misaligned paternity information. Between December 2009 and November 2010, 63 residents in Kilifi County were engaged in informed deliberative small group discussions, structured to support normative reflection within the groups, with purposive selection to explore diversity. Analysis was based on a modified framework analysis approach, drawing on relevant social science and bioethics literature. The methods generated in-depth individual and group reflection on morally important issues and uncovered wide diversity in views and values. Fundamental and conflicting values emerged around the importance of family interests and openness, underpinned by disagreement on the moral implications of marital infidelity and withholding truth. Wider consideration of ethical issues emerging in these debates supports locally-held reasoning that paternal sickle cell testing should not be undertaken in this context, in contrast to views that testing should be done with or without the disclosure of misaligned paternity information. The findings highlight the importance of facilitating wider testing of family members of affected children, contingent on the development and implementation of national policies for the management of this inherited disorder. Their richness also illustrates the potential for the approach adopted in this study to strengthen community consultation.


Quantitative Bayesian model-based analysis of amide proton transfer MRI.
Amide Proton Transfer (APT) reports on contrast derived from the exchange of protons between amide groups and water. Commonly, APT contrast is quantified by asymmetry analysis, providing an ensemble contrast of both amide proton concentration and exchange rate. An alternative is to sample the off-resonant spectrum and fit an exchange model, permitting the APT effect to be quantified, correcting automatically for confounding effects of spillover, field inhomogeneity, and magnetization transfer. Additionally, it should permit amide concentration and exchange rate to be independently quantified. Here, a Bayesian method is applied to this problem allowing pertinent prior information to be specified. A three-pool model was used incorporating water protons, amide protons, and magnetization transfer effect. The method is demonstrated in simulations, creatine phantoms with varying pH and in vivo (n = 7). The Bayesian model-based approach was able to quantify the APT effect accurately (root-mean-square error < 2%) even when subject to confounding field variation and magnetization transfer effect, unlike traditional asymmetry analysis. The in vivo results gave approximate APT concentration (relative to water) and exchange rate values of 3 × 10(-3) and 15 s(-1) . A degree of correlation was observed between these parameter making the latter difficult to quantify with absolute accuracy, suggesting that more optimal sampling strategies might be required.


Myo-inositol oxygenase expression profile modulates pathogenic ferroptosis in the renal proximal tubule.
Overexpression of myo-inositol oxygenase (MIOX), a proximal tubular enzyme, exacerbates cellular redox injury in acute kidney injury (AKI). Ferroptosis, a newly coined term associated with lipid hydroperoxidation, plays a critical role in the pathogenesis of AKI. Whether or not MIOX exacerbates tubular damage by accelerating ferroptosis in cisplatin-induced AKI remains elusive. Cisplatin-treated HK-2 cells exhibited notable cell death, which was reduced by ferroptosis inhibitors. Also, alterations in various ferroptosis metabolic sensors, including lipid hydroperoxidation, glutathione peroxidase 4 (GPX4) activity, NADPH and reduced glutathione (GSH) levels, and ferritinophagy, were observed. These perturbations were accentuated by MIOX overexpression, while ameliorated by MIOX knockdown. Likewise, cisplatin-treated CD1 mice exhibited tubular damage and derangement of renal physiological parameters, which were alleviated by ferrostatin-1, a ferroptosis inhibitor. To investigate the relevance of MIOX to ferroptosis, WT mice, MIOX-overexpressing transgenic (MIOX-Tg) mice, and MIOX-KO mice were subjected to cisplatin treatment. In comparison with cisplatin-treated WT mice, cisplatin-treated MIOX-Tg mice had more severe renal pathological changes and perturbations in ferroptosis metabolic sensors, which were minimal in cisplatin-treated MIOX-KO mice. In conclusion, these findings indicate that ferroptosis, an integral process in the pathogenesis of cisplatin-induced AKI, is modulated by the expression profile of MIOX.


Effects of aspirin on risk and severity of early recurrent stroke after transient ischaemic attack and ischaemic stroke: time-course analysis of randomised trials.
Aspirin is recommended for secondary prevention after transient ischaemic attack (TIA) or ischaemic stroke on the basis of trials showing a 13% reduction in long-term risk of recurrent stroke. However, the risk of major stroke is very high for only the first few days after TIA and minor ischaemic stroke, and observational studies show substantially greater benefits of early medical treatment in the acute phase than do longer-term trials. We hypothesised that the short-term benefits of early aspirin have been underestimated. Pooling the individual patient data from all randomised trials of aspirin versus control in secondary prevention after TIA or ischaemic stroke, we studied the effects of aspirin on the risk and severity of recurrent stroke, stratified by the following time periods: less than 6 weeks, 6-12 weeks, and more than 12 weeks after randomisation. We compared the severity of early recurrent strokes between treatment groups with shift analysis of modified Rankin Scale (mRS) score. To understand possible mechanisms of action, we also studied the time course of the interaction between effects of aspirin and dipyridamole in secondary prevention of stroke. In a further analysis we pooled data from trials of aspirin versus control in which patients were randomised less than 48 h after major acute stroke, stratified by severity of baseline neurological deficit, to establish the very early time course of the effect of aspirin on risk of recurrent ischaemic stroke and how this differs by severity at baseline. We pooled data for 15,778 participants from 12 trials of aspirin versus control in secondary prevention. Aspirin reduced the 6 week risk of recurrent ischaemic stroke by about 60% (84 of 8452 participants in the aspirin group had an ischaemic stroke vs 175 of 7326; hazard ratio [HR] 0·42, 95% CI 0·32-0·55, p<0·0001) and disabling or fatal ischaemic stroke by about 70% (36 of 8452 vs 110 of 7326; 0·29, 0·20-0·42, p<0·0001), with greatest benefit noted in patients presenting with TIA or minor stroke (at 0-2 weeks, two of 6691 participants in the aspirin group with TIA or minor stroke had a disabling or fatal ischaemic stroke vs 23 of 5726 in the control group, HR 0·07, 95% CI 0·02-0·31, p=0·0004; at 0-6 weeks, 14 vs 60 participants, 0·19, 0·11-0·34, p<0·0001). The effect of aspirin on early recurrent ischaemic stroke was due partly to a substantial reduction in severity (mRS shift analysis odds ratio [OR] 0·42, 0·26-0·70, p=0·0007). These effects were independent of dose, patient characteristics, or aetiology of TIA or stroke. Some further reduction in risk of ischaemic stroke accrued for aspirin only versus control from 6-12 weeks, but there was no benefit after 12 weeks (stroke risk OR 0·97, 0·84-1·12, p=0·67; severity mRS shift OR 1·00, 0·77-1·29, p=0·97). By contrast, dipyridamole plus aspirin versus aspirin alone had no effect on risk or severity of recurrent ischaemic stroke within 12 weeks (OR 0·90, 95% CI 0·65-1·25, p=0·53; mRS shift OR 0·90, 0·37-1·72, p=0·99), but dipyridamole did reduce risk thereafter (0·76, 0·63-0·92, p=0·005), particularly of disabling or fatal ischaemic stroke (0·64, 0·49-0·84, p=0·0010). We pooled data for 40,531 participants from three trials of aspirin versus control in major acute stroke. The reduction in risk of recurrent ischaemic stroke at 14 days was most evident in patients with less severe baseline deficits, and was substantial by the second day after starting treatment (2-3 day HR 0·37, 95% CI 0·25-0·57, p<0·0001). Our findings confirm that medical treatment substantially reduces the risk of early recurrent stroke after TIA and minor stroke and identify aspirin as the key intervention. The considerable early benefit from aspirin warrants public education about self-administration after possible TIA. The previously unrecognised effect of aspirin on severity of early recurrent stroke, the diminishing benefit with longer-term use, and the contrasting time course of effects of dipyridamole have implications for understanding mechanisms of action. Wellcome Trust, the National Institute of Health Research (NIHR) Biomedical Research Centre, Oxford.


High-density mapping of the MHC identifies a shared role for HLA-DRB1*01:03 in inflammatory bowel diseases and heterozygous advantage in ulcerative colitis.
Genome-wide association studies of the related chronic inflammatory bowel diseases (IBD) known as Crohn's disease and ulcerative colitis have shown strong evidence of association to the major histocompatibility complex (MHC). This region encodes a large number of immunological candidates, including the antigen-presenting classical human leukocyte antigen (HLA) molecules. Studies in IBD have indicated that multiple independent associations exist at HLA and non-HLA genes, but they have lacked the statistical power to define the architecture of association and causal alleles. To address this, we performed high-density SNP typing of the MHC in >32,000 individuals with IBD, implicating multiple HLA alleles, with a primary role for HLA-DRB1*01:03 in both Crohn's disease and ulcerative colitis. Noteworthy differences were observed between these diseases, including a predominant role for class II HLA variants and heterozygous advantage observed in ulcerative colitis, suggesting an important role of the adaptive immune response in the colonic environment in the pathogenesis of IBD.


From social to genetic structures in central Asia.
Pastoral and farmer populations, who have coexisted in Central Asia since the fourth millennium B.C., present not only different lifestyles and means of subsistence but also various types of social organization. Pastoral populations are organized into so-called descent groups (tribes, clans, and lineages) and practice exogamous marriages (a man chooses a bride in a different lineage or clan). In Central Asia, these descent groups are patrilineal: The children are systematically affiliated with the descent groups of the father. By contrast, farmer populations are organized into families (extended or nuclear) and often establish endogamous marriages with cousins. This study aims at better understanding the impact of these differences in lifestyle and social organization on the shaping of genetic diversity. We show that pastoral populations exhibit a substantial loss of Y chromosome diversity in comparison to farmers but that no such a difference is observed at the mitochondrial-DNA level. Our analyses indicate that the dynamics of patrilineal descent groups, which implies different male and female sociodemographic histories, is responsible for these sexually-asymmetric genetic patterns. This molecular signature of the pastoral social organization disappears over a few centuries only after conversion to an agricultural way of life.


Mef2C is a lineage-restricted target of Scl/Tal1 and regulates megakaryopoiesis and B-cell homeostasis.
The basic helix-loop-helix transcription factor stem cell leukemia gene (Scl) is a master regulator for hematopoiesis essential for hematopoietic specification and proper differentiation of the erythroid and megakaryocyte lineages. However, the critical downstream targets of Scl remain undefined. Here, we identified a novel Scl target gene, transcription factor myocyte enhancer factor 2 C (Mef2C) from Scl(fl/fl) fetal liver progenitor cell lines. Analysis of Mef2C(-/-) embryos showed that Mef2C, in contrast to Scl, is not essential for specification into primitive or definitive hematopoietic lineages. However, adult VavCre(+)Mef2C(fl/fl) mice exhibited platelet defects similar to those observed in Scl-deficient mice. The platelet counts were reduced, whereas platelet size was increased and the platelet shape and granularity were altered. Furthermore, megakaryopoiesis was severely impaired in vitro. Chromatin immunoprecipitation microarray hybridization analysis revealed that Mef2C is directly regulated by Scl in megakaryocytic cells, but not in erythroid cells. In addition, an Scl-independent requirement for Mef2C in B-lymphoid homeostasis was observed in Mef2C-deficient mice, characterized as severe age-dependent reduction of specific B-cell progenitor populations reminiscent of premature aging. In summary, this work identifies Mef2C as an integral member of hematopoietic transcription factors with distinct upstream regulatory mechanisms and functional requirements in megakaryocyte and B-lymphoid lineages.


Epilogue: Lessons from the CONVERGE study of major depressive disorder in China.
This review summarizes the first clinical reports from the CONVERGE consortium: China, Oxford and VCU Experimental Research on Genetic Epidemiology. CONVERGE sets out to investigate the nature and mode of action of the genetic and environmental risk factors for major depressive disorder (MDD). CONVERGE aims to collect 6000 cases of recurrent MDD and 6000 controls. The consortium includes hospitals in 30 cities, all with populations exceeding 5million, and has collected over 2000 cases and controls. High quality phenotype data on MDD collected in China is now available to determine the source and nature of this highly heterogeneous condition. Analyses reported in a series of papers indicate that the clinical features and risk factors of MDD are sufficiently similar to those in the West that we can confidently predict that the results of subsequent analyses will be widely applicable.


Diverse Streptococcus pneumoniae Strains Drive a Mucosal-Associated Invariant T-Cell Response Through Major Histocompatibility Complex class I-Related Molecule-Dependent and Cytokine-Driven Pathways.
Mucosal-associated invariant T (MAIT) cells represent an innate T-cell population that can recognize ligands generated by the microbial riboflavin synthesis pathway, presented via the major histocompatibility complex class I-related molecule (MR1). Streptococcus pneumoniae is a major human pathogen that is also associated with commensal carriage; thus, host control at the mucosal interface is critical. The recognition of pneumococci by MAIT cells has not been defined nor have the genomics and transcriptomics of the riboflavin operon. We observed robust recognition of pneumococci by MAIT cells, using both MR1-dependent and MR1-independent pathways. The pathway used was dependent on the antigen-presenting cell. The riboflavin operon was highly conserved across a range of 571 pneumococci from 39 countries, dating back to 1916, and different versions of the riboflavin operon were also identified in related Streptococcus species. These data indicate an important functional relationship between MAIT cells and pneumococci.


Mu rhythm desynchronization reveals motoric influences of hand action on object recognition.
We examined the effect of hand grip on object recognition by studying the modulation of the mu rhythm when participants made object decisions to objects and non-objects shown with congruent or incongruent hand-grip actions. Despite the grip responses being irrelevant to the task, mu rhythm activity on the scalp over motor and pre-motor cortex was sensitive to the congruency of the hand grip-in particular the event-related desynchronization of the mu rhythm was more pronounced for familiar objects grasped with an appropriate grip than for objects given an inappropriate grasp. Also the power of mu activity correlated with RTs to congruently gripped objects. The results suggest that familiar motor responses evoked by the appropriateness of a hand grip facilitate recognition responses to objects.


Clinical profiles and outcomes of deep brain stimulation in G2019S LRRK2 Parkinson disease.
The objective of this study was to evaluate clinical features and response to deep brain stimulation (DBS) in G2019S LRRK2-Parkinson disease (LRRK2-PD) and idiopathic PD (IPD). The authors conducted a clinic-based cohort study of PD patients recruited from the Mount Sinai Beth Israel Genetics database of PD studies. The cohort included 87 participants with LRRK2-PD (13 who underwent DBS) and 14 DBS participants with IPD enrolled between 2009 and 2017. The baseline clinical features, including motor ratings and levodopa-equivalent daily dose (LEDD), were compared among LRRK2-PD patients with and without DBS, between LRRK2-PD with DBS and IPD with DBS, and between LRRK2-PD with subthalamic nucleus (STN) and internal segment of the globus pallidus (GPi) DBS. Longitudinal motor scores (Unified Parkinson's Disease Rating Scale-part III) and medication usage were also assessed pre- and postoperatively. Compared to LRRK2-PD without DBS (n = 74), the LRRK2-PD with DBS cohort (n = 13) had a significantly younger age of onset, longer disease duration, were more likely to have dyskinesia, and were less likely to experience hand tremor at disease onset. LRRK2-PD participants were also more likely to be referred for surgery because of severe dyskinesia (11/13 [85%] vs 6/14 [43%], p = 0.04) and were less likely to be referred for medically refractory tremor (0/13 [0%] vs 6/14 [43%], p = 0.02) than were IPD patients. Among LRRK2-PD patients, both STN-DBS and GPi-DBS targets were effective, although the sample size was small for both groups. There were no revisions or adverse effects reported in the GPi-DBS group, while 2 of the LRRK2-PD participants who underwent STN-DBS required revisions and a third reported depression as a stimulation-related side effect. Medication reduction favored the STN group. The LRRK2-PD cohort referred for DBS had a slightly different profile, including earlier age of onset and dyskinesia. Both the STN and GPi DBS targets were effective in symptom suppression. Patients with G2019S LRRK2 PD were well-suited for DBS therapy and had favorable motor outcomes regardless of the DBS target. LRRK2-DBS patients had longer disease durations and tended to have more dyskinesia. Dyskinesia commonly served as the trigger for DBS surgical candidacy. Medication-refractory tremor was not a common indication for surgery in the LRRK2 cohort.


De novo sequencing, assembly and analysis of the genome of the laboratory strain Saccharomyces cerevisiae CEN.PK113-7D, a model for modern industrial biotechnology.
Saccharomyces cerevisiae CEN.PK 113-7D is widely used for metabolic engineering and systems biology research in industry and academia. We sequenced, assembled, annotated and analyzed its genome. Single-nucleotide variations (SNV), insertions/deletions (indels) and differences in genome organization compared to the reference strain S. cerevisiae S288C were analyzed. In addition to a few large deletions and duplications, nearly 3000 indels were identified in the CEN.PK113-7D genome relative to S288C. These differences were overrepresented in genes whose functions are related to transcriptional regulation and chromatin remodelling. Some of these variations were caused by unstable tandem repeats, suggesting an innate evolvability of the corresponding genes. Besides a previously characterized mutation in adenylate cyclase, the CEN.PK113-7D genome sequence revealed a significant enrichment of non-synonymous mutations in genes encoding for components of the cAMP signalling pathway. Some phenotypic characteristics of the CEN.PK113-7D strains were explained by the presence of additional specific metabolic genes relative to S288C. In particular, the presence of the BIO1 and BIO6 genes correlated with a biotin prototrophy of CEN.PK113-7D. Furthermore, the copy number, chromosomal location and sequences of the MAL loci were resolved. The assembled sequence reveals that CEN.PK113-7D has a mosaic genome that combines characteristics of laboratory strains and wild-industrial strains.


Managing differences: the central challenge of global strategy.
The main goal of any international strategy should be to manage the large differences that arise at the borders of markets. Yet executives often fail to exploit market and production discrepancies, focusing instead on the tensions between standardization and localization. In this article, Pankaj Ghemawat presents a new framework that encompasses all three effective responses to the challenges of globalization. He calls it the AAA Triangle. The A's stand for the three distinct types of international strategy. Through adaptation, companies seek to boost revenues and market share by maximizing their local relevance. Through aggregation, they attempt to deliver economies of scale by creating regional, or sometimes global, operations. And through arbitrage, they exploit disparities between national or regional markets, often by locating different parts of the supply chain in different places--for instance, call centers in India, factories in China, and retail shops in Western Europe. Ghemawat draws on several examples that illustrate how organizations use and balance these strategies and describes the trade-offs they make as they do so. Because most enterprises should draw from all three A's to some extent, the framework can be used to develop a summary scorecard indicating how well the company is globalizing. However, given the tensions among the strategies, it's not enough simply to tick off the corresponding boxes. Strategic choice requires some degree of prioritization--and the framework can help with that as well. While it is possible to make progress on all three strategies, companies usually must focus on one or two when trying to build competitive advantage.


Buffering of recovery from acidification by organic acids.
In the United Kingdom, as in other regions of Europe and North America, recent decreases in surface water sulphate concentrations, due to reduced sulphur emissions, have coincided with marked increases in dissolved organic carbon (DOC) concentrations. Since many of the compounds comprising DOC are acidic, the resulting increases in organic acidity may have the potential to offset the benefits of a decrease in mineral (sulphate) acidity. To test this, we used a triprotic model of organic acid dissociation to estimate the proportional organic acid buffering of reduced mineral acidity as measured in the 22 lakes and streams monitored by the UK Acid Waters Monitoring Network. For an average non-marine sulphate decrease of 30 mueq l(-1) over 15 years from 1988-2003, we estimate that around 28% was counterbalanced by rising strong organic acids, 20% by rising alkalinity (partly attributable to an increase in weak organic acids), 11% by falling inorganic aluminium and 41% by falling non-marine base cations. The situation is complicated by a concurrent decrease in marine ion concentrations, and the impact this may have had on both DOC and acidity, but results clearly demonstrate that organic acid increases have substantially limited the amount of recovery from acidification (in terms of rising alkalinity and falling aluminium) that have resulted from reducing sulphur emissions. The consistency and magnitude of sulphate and organic acid changes are consistent with a causal link between the two, possibly due to the effects of changing acidity, ionic strength and aluminium concentrations on organic matter solubility. If this is the case, then organic acids can be considered effective but partial buffers to acidity change in organic soils, and this mechanism needs to be considered in assessing and modelling recovery from acidification, and in defining realistic reference conditions. However, large spatial variations in the relative magnitude of organic acid and sulphate changes, notably for low-deposition sites in northwestern areas where organic acid increases apparently exceed non-marine sulphate decreases, suggest that additional factors, such as changes in sea-salt deposition and climatic factors, may be required to explain the full magnitude of DOC increases in UK surface waters.


High survivin expression as a risk factor in patients with anal carcinoma treated with concurrent chemoradiotherapy.
To investigate the prognostic value of survivin expression in pretreatment specimens from patients with anal cancer treated with concurrent 5-FU and mitomycin C-based chemoradiation (CRT). Immunohistochemical staining for survivin was performed in pretreatment biopsies of 62 patients with anal carcinoma. Survivin expression was correlated with clinical and histopathological characteristics as well as local failure free- (LFFS), distant metastases free- (DMFS), cancer specific- (CSS), and overall survival (OS). Survivin staining intensity was weak in 10%, intermediate in 48% and intense in 42% of the patients. No association between survivin expression and clinicopathologic factors (tumor stage, age and HIV status) could be shown. In univariate analysis, the level of survivin staining was significantly correlated with DMFS (low survivin vs. high survivin: 94% vs. 74%, p = 0.04). T-stage, N-stage and the tumor grading were significantly associated with OS and CSS and with DMFS and LFFS, respectively. In multivariate analysis, survivin was confirmed as independent prognostic parameter for DMFS (RR, 0.04; p = 0.02) and for OS (RR, 0.27; p = 0.04). Our results demonstrated that the level of pretreatment survivin is correlated with the clinical outcome in patients with anal carcinoma treated with concurrent CRT. Further studies are warranted to elucidate the complex role of survivin for the oncologic treatment and to exploit the protein as a therapeutic target in combined modality treatment of anal cancer.


Fibroblast growth factor-23 regulates parathyroid hormone and 1alpha-hydroxylase expression in cultured bovine parathyroid cells.
Fibroblast growth factor-23 (FGF23) is a circulating factor that decreases serum levels of inorganic phosphate (Pi) as well as 1,25-dihydroxyvitamin D(3). Recent studies also suggest a correlation between serum levels of FGF23 and parathyroid hormone (PTH) in patients with chronic kidney disease. It is, however, unknown whether FGF23 directly modulates PTH expression, or whether the correlation is secondary to abnormalities in Pi and vitamin D metabolism. The objective of the current study was therefore to elucidate possible direct effects of FGF23 on bovine parathyroid cells in vitro. Treatment of parathyroid cells with a stabilized form of recombinant FGF23 (FGF23(R176Q)) induced a rise in early response gene-1 mRNA transcripts, a marker of FGF23 signaling. FGF23(R176Q) potently and dose-dependently decreased the PTH mRNA level within 12 h. In agreement, FGF23(R176Q) also decreased PTH secretion into conditioned media. In contrast, FGF23(R176Q) dose-dependently increased 1alpha-hydroxylase expression within 3 h. FGF23 (R176Q) did not affect cell viability nor induce apoptosis, whereas a small but significant increase in cell proliferation was found. We conclude that FGF23 is a negative regulator of PTH mRNA expression and secretion in vitro. Our data suggest that FGF23 may be a physiologically relevant regulator of PTH. This defines a novel function of FGF23 in addition to the previously established roles in controlling vitamin D and Pi metabolism.


The Magpie Trial: a randomised trial comparing magnesium sulphate with placebo for pre-eclampsia. Outcome for children at 18 months.
To assess the long-term effects of in utero exposure to magnesium sulphate for children whose mothers had pre-eclampsia. Assessment at 18 months of age for children whose mothers were recruited to the Magpie Trial (recruitment 1998-2001 ISRCTN 86938761), which compared magnesium sulphate with placebo. Follow-up of children born at 125 centres in 19 countries across five continents. A total of 6922 children were born to women randomised before delivery at follow-up centres. Of these, 2271 were not included for logistic reasons and 168 were excluded (101 at a centre where <20% were contacted, 40 whose death or disability was due to a problem at conception or embryogenesis and 27 whose parent/s opted out). Therefore, 4483 children were included in follow-up, of whom 3283 (73%) were contacted. Assessment by questionnaire, with interview and neurodevelopmental testing of selected children. Death or neurosensory disability at age of 18 months. Of those allocated magnesium sulphate, 245/1635 (15.0%) were dead or had neurosensory disability at 18 months compared with 233/1648 (14.1%) allocated placebo (relative risk [RR] 1.06, 95% CI 0.90-1.25), and of survivors, 19/1409 (1.3%) had neurosensory disability at 18 months compared with 27/1442 (1.9%) (RR 0.72, 95% CI 0.40-1.29). There were no substantial differences in causes of death or in the risk of individual impairments or disabilities. The lower risk of eclampsia following prophylaxis with magnesium sulphate was not associated with a clear difference in the risk of death or disability for children at 18 months.


Reversal of disease-related pathologies in the fragile X mouse model by selective activation of GABAB receptors with arbaclofen.
Fragile X syndrome (FXS), the most common inherited cause of intellectual disability and autism, results from the transcriptional silencing of FMR1 and loss of the mRNA translational repressor protein fragile X mental retardation protein (FMRP). Patients with FXS exhibit changes in neuronal dendritic spine morphology, a pathology associated with altered synaptic function. Studies in the mouse model of fragile X have shown that loss of FMRP causes excessive synaptic protein synthesis, which results in synaptic dysfunction and altered spine morphology. We tested whether the pharmacologic activation of the γ-aminobutyric acid type B (GABA(B)) receptor could correct or reverse these phenotypes in Fmr1-knockout mice. Basal protein synthesis, which is elevated in the hippocampus of Fmr1-knockout mice, was corrected by the in vitro application of the selective GABA(B) receptor agonist STX209 (arbaclofen, R-baclofen). STX209 also reduced to wild-type values the elevated AMPA receptor internalization in Fmr1-knockout cultured neurons, a known functional consequence of increased protein synthesis. Acute administration of STX209 in vivo, at doses that modify behavior, decreased mRNA translation in the cortex of Fmr1-knockout mice. Finally, the chronic administration of STX209 in juvenile mice corrected the increased spine density in Fmr1-knockout mice without affecting spine density in wild-type mice. Thus, activation of the GABA(B) receptor with STX209 corrected synaptic abnormalities considered central to fragile X pathophysiology, a finding that suggests that STX209 may be a potentially effective therapy to treat the core symptoms of FXS.


Clinical and epidemiological features of COVID-19 family clusters in Beijing, China.
Since its discovery, SARS-CoV-2 has been spread throughout China before becoming a global pandemic. In Beijing, family clusters are the main mode of human-human transmission accounting for 57.6% of the total confirmed cases. We present the epidemiological and clinical features of the clusters of three large and one small families. Our results revealed that SARS-CoV-2 is transmitted quickly through contact with index case, and a total of 22/24 infections were observed. Among those infected, 20/22 had mild symptoms and only two had moderate to severe clinical manifestations. Children in the families generally showed milder symptoms. The incubation period varied from 2 to 13 days, and the shedding of virus from the upper respiratory tract lasted from 5 to over 30 days. A prolonged period of virus shedding (>30 days) in upper respiratory tract was observed in 6/24 cases. SARS-CoV-2 is transmitted quickly in the form of family clusters. While the infection rate is high within the cluster, the disease manifestations, latent period, and virus shedding period varied greatly. We therefore recommend rigorously testing contacts even during the no-symptom phase and consider whether viral shedding has ceased before stopping isolation measures for an individual.


Big brains, meat, tuberculosis and the nicotinamide switches: co-evolutionary relationships with modern repercussions on longevity and disease?
Meat eating has been an important trigger for human evolution however the responsible component in meat has not been clearly identified. Here we propose that the limiting factors for expanding brains and increasing longevity were the micronutrient nicotinamide (vitamin B3) and the metabolically related essential amino-acid, tryptophan. Meat offers significant sourcing challenges and lack causes a deficiency of nicotinamide and tryptophan and consequently the energy carrier nicotinamide adenine dinucleotide (NAD) that gets consumed in regulatory circuits important for survival, resulting in premature ageing, poor cognition and brain atrophy. If a trophic supply of dietary nicotinamide/tryptophan is so essential for building brains, constraining their size and connectivity, we hypothesise that back-up mechanisms to ensure the supply evolved. One strategy may be increasing the reliance on gut symbionts to break down celluloses that produces NADH and only nicotinamide indirectly, and may cause diarrhoea. We suggest that a direct supplier was the chronic mycobacterial infection tuberculosis (TB) that is a surprise candidate but it co-evolved early, does not inevitably cause disease (90-95% of those infected are healthy), and secretes (and is inhibited by) nicotinamide. We hypothesise that TB evolved first as a symbiont that enabled humans to cope with short-lived shortages of meat and only later behaved as a pathogen when the supply deteriorated chronically, for those in poverty. (TB immunology and epidemiology is riddled with paradoxes for a conventional pathogen). We test this in pilot data showing that sharp declines in TB (and diarrhoea) - `environmental enteropathy' strongly correlate with increasing meat consumption and therefore nicotinamide exposure, unlike later onset cancers and Parkinson's disease that increased in incidence, perhaps - as we propose a hypothetical hypervitaminosis B3 (to include obesity and the metabolic syndrome) - as the trade-off for increased brain power and longevity, a recently evolved human characteristic.


Alcohol intake modulates the genetic association between HDL cholesterol and the PPARgamma2 Pro12Ala polymorphism.
The peroxisome proliferator-activated receptor gamma (PPARgamma) Pro12Ala polymorphism affects plasma lipids, but to what extent alcohol intake interferes with this association remains unknown. We randomly recruited 251 nuclear families (433 parents and 493 offspring) in the framework of the European Project on Genes in Hypertension study and genotyped 926 participants in whom all serum lipid variables and information on alcohol consumption were available for PPARgamma2 Pro12Ala. Genotype-phenotype relations were assessed using generalized estimating equations (GEE) and a quantitative transmission disequilibrium test (QTDT). The Ala12 allele was more frequent in Novosibirsk (0.17) than in Cracow (0.12) and Mirano (0.11) (P < 0.01). Using GEE (P = 0.03) or QTDT (P = 0.007), Italian offspring carrying the Ala12 allele had higher serum HDL cholesterol than noncarriers. HDL cholesterol levels were on average 0.086 mmol/l (P = 0.001) higher in drinkers than in nondrinkers. Compared with Pro12 homozygotes, Ala12 allele carriers consuming alcohol had higher serum total and HDL cholesterol, with the opposite trend occurring in nondrinkers. This genotype-alcohol interaction was independent of the type of alcoholic beverage and more pronounced in moderate than in heavy drinkers. We conclude that alcohol intake modulates the relation between the PPARgamma2 Pro12Ala and HDL cholesterol level and that, therefore, the Pro12Ala polymorphism, pending confirmation of our findings, might affect cardiovascular prognosis.


Training induces changes in white-matter architecture.
Although experience-dependent structural changes have been found in adult gray matter, there is little evidence for such changes in white matter. Using diffusion imaging, we detected a localized increase in fractional anisotropy, a measure of microstructure, in white matter underlying the intraparietal sulcus following training of a complex visuo-motor skill. This provides, to the best of our knowledge, the first evidence for training-related changes in white-matter structure in the healthy human adult brain.


Sleep-like behavior and 24-h rhythm disruption in the Tc1 mouse model of Down syndrome.
Down syndrome is a common disorder associated with intellectual disability in humans. Among a variety of severe health problems, patients with Down syndrome exhibit disrupted sleep and abnormal 24-h rest/activity patterns. The transchromosomic mouse model of Down syndrome, Tc1, is a trans-species mouse model for Down syndrome, carrying most of human chromosome 21 in addition to the normal complement of mouse chromosomes and expresses many of the phenotypes characteristic of Down syndrome. To date, however, sleep and circadian rhythms have not been characterized in Tc1 mice. Using both circadian wheel-running analysis and video-based sleep scoring, we showed that these mice exhibited fragmented patterns of sleep-like behaviour during the light phase of a 12:12-h light/dark (LD) cycle with an extended period of continuous wakefulness at the beginning of the dark phase. Moreover, an acute light pulse during night-time was less effective in inducing sleep-like behaviour in Tc1 animals than in wild-type controls. In wheel-running analysis, free running in constant light (LL) or constant darkness (DD) showed no changes in the circadian period of Tc1 animals although they did express subtle behavioural differences including a reduction in total distance travelled on the wheel and differences in the acrophase of activity in LD and in DD. Our data confirm that Tc1 mice express sleep-related phenotypes that are comparable with those seen in Down syndrome patients with moderate disruptions in rest/activity patterns and hyperactive episodes, while circadian period under constant lighting conditions is essentially unaffected.


Therapeutic gene silencing in the nervous system.
Progress in the understanding of RNA biology has brought into focus the prospect of using RNA-based therapeutics as a novel approach to treat human disease. In particular, following the discovery of the RNA interference (RNAi) pathway, the emergence of technology based on small interfering RNA (siRNA) now offers a powerful and highly specific tool for therapeutic gene silencing. Many neurological diseases, including neurodegenerative disorders, tumours and retinal disease are likely candidates to benefit from such advances. The challenges ahead will be to identify appropriate disease gene targets and, crucially, to understand the biological parameters that determine safe, precise and effective delivery and function of RNA-based therapeutic molecules within the unique environment of the nervous system.


Salivary cortisol as a predictor of socioemotional adjustment during kindergarten: a prospective study.
This study, based on a sample of 172 children, examined the relation between average afternoon salivary cortisol levels measured at home at age 4.5 years and socioemotional adjustment a year and a half later, as reported by mothers, fathers, and teachers. Cortisol levels were hypothesized to be positively associated with withdrawal-type behaviors (e.g., internalizing, social wariness) and inversely related to approach-type behaviors, both negative and positive (e.g., externalizing, school engagement). Higher cortisol levels at age 4.5 predicted more internalizing behavior and social wariness as reported by teachers and mothers, although child gender moderated the relation between cortisol and mother report measures. An inverse relation was found between boys' cortisol levels and father report of externalizing behavior. A marginal inverse relation was found between child cortisol levels and teacher report of school engagement. Behavior assessed concurrently with cortisol collection did not account for the prospective relations observed,suggesting that cortisol adds uniquely to an understanding of behavioral development.


Tuning DNA binding kinetics in an optical trap by plasmonic nanoparticle heating.
We report on the tuning of specific binding of DNA attached to gold nanoparticles at the individual particle pair (dimer) level in an optical trap by means of plasmonic heating. DNA hybridization events are detected optically by the change in the plasmon resonance frequency due to plasmonic coupling of the nanoparticles. We find that at larger trapping powers (i.e., larger temperatures and stiffer traps) the hybridization rates decrease by more than an order of magnitude. This result is explained by higher temperatures preventing the formation of dimers with lower binding energies. Our results demonstrate that plasmonic heating can be used to fine tune the kinetics of biomolecular binding events.


The hazards of time.
Temporal expectations are continuously formed and updated, and interact with expectations about other relevant attributes of events, in order to optimise our interaction with unfolding sensory stimulation. In this paper, we will highlight some evidence revealing the pervasive effects of temporal expectations in modulating perception and action, and reflect on the current state of understanding about their underlying neural systems and mechanisms.


Treating rheumatoid arthritis to target: 2014 update of the recommendations of an international task force.
Reaching the therapeutic target of remission or low-disease activity has improved outcomes in patients with rheumatoid arthritis (RA) significantly. The treat-to-target recommendations, formulated in 2010, have provided a basis for implementation of a strategic approach towards this therapeutic goal in routine clinical practice, but these recommendations need to be re-evaluated for appropriateness and practicability in the light of new insights. To update the 2010 treat-to-target recommendations based on systematic literature reviews (SLR) and expert opinion. A task force of rheumatologists, patients and a nurse specialist assessed the SLR results and evaluated the individual items of the 2010 recommendations accordingly, reformulating many of the items. These were subsequently discussed, amended and voted upon by >40 experts, including 5 patients, from various regions of the world. Levels of evidence, strengths of recommendations and levels of agreement were derived. The update resulted in 4 overarching principles and 10 recommendations. The previous recommendations were partly adapted and their order changed as deemed appropriate in terms of importance in the view of the experts. The SLR had now provided also data for the effectiveness of targeting low-disease activity or remission in established rather than only early disease. The role of comorbidities, including their potential to preclude treatment intensification, was highlighted more strongly than before. The treatment aim was again defined as remission with low-disease activity being an alternative goal especially in patients with long-standing disease. Regular follow-up (every 1-3 months during active disease) with according therapeutic adaptations to reach the desired state was recommended. Follow-up examinations ought to employ composite measures of disease activity that include joint counts. Additional items provide further details for particular aspects of the disease, especially comorbidity and shared decision-making with the patient. Levels of evidence had increased for many items compared with the 2010 recommendations, and levels of agreement were very high for most of the individual recommendations (≥9/10). The 4 overarching principles and 10 recommendations are based on stronger evidence than before and are supposed to inform patients, rheumatologists and other stakeholders about strategies to reach optimal outcomes of RA.


Test-retest variability of serotonin 5-HT2A receptor binding measured with positron emission tomography and [18F]altanserin in the human brain.
The role of serotonin in CNS function and in many neuropsychiatric diseases (e.g., schizophrenia, affective disorders, degenerative dementias) support the development of a reliable measure of serotonin receptor binding in vivo in human subjects. To this end, the regional distribution and intrasubject test-retest variability of the binding of [18F]altanserin were measured as important steps in the further development of [18F]altanserin as a radiotracer for positron emission tomography (PET) studies of the serotonin 5-HT2A receptor. Two high specific activity [18F]altanserin PET studies were performed in normal control subjects (n = 8) on two separate days (2-16 days apart). Regional specific binding was assessed by distribution volume (DV), estimates that were derived using a conventional four compartment (4C) model, and the Logan graphical analysis method. For both analysis methods, levels of [18F]altanserin binding were highest in cortical areas, lower in the striatum and thalamus, and lowest in the cerebellum. Similar average differences of 13% or less were observed for the 4C model DV determined in regions with high receptor concentrations with greater variability in regions with low concentrations (16-20%). For all regions, the absolute value of the test-retest differences in the Logan DV values averaged 12% or less. The test-retest differences in the DV ratios (regional DV values normalized to the cerebellar DV) determined by both data analysis methods averaged less than 10%. The regional [18F]altanserin DV values using both of these methods were significantly correlated with literature-based values of the regional concentrations of 5-HT2A receptors determined by postmortem autoradiographic studies (r2 = 0.95, P < 0.001 for the 4C model and r2 = 0.96, P < 0.001 for the Logan method). Brain uptake studies in rats demonstrated that two different radiolabeled metabolites of [18F]altanserin (present at levels of 3-25% of the total radioactivity in human plasma 10-120 min postinjection) were able to penetrate the blood-brain barrier. However, neither of these radiolabeled metabolites bound specifically to the 5-HT2A receptor and did not interfere with the interpretation of regional [18F]altanserin-specific binding parameters obtained using either a conventional 4C model or the Logan graphical analysis method. In summary, these results demonstrate that the test-retest variability of [18F]altanserin-specific binding is comparable to that of other PET radiotracers and that the regional specific binding of [18F]altanserin in human brain was correlated with the known regional distribution of 5-HT2A receptors. These findings support the usefulness of [18F]altanserin as a radioligand for PET studies of 5-HT2A receptors.


The N-terminus of ecdysteroid receptor isoforms and ultraspiracle interacts with different ecdysteroid response elements in a sequence specific manner to modulate transcriptional activity.
The functional insect ecdysteroid receptor is comprised of two nuclear receptors, the ecdysteroid receptor (EcR) and the RXR homologue, ultraspiracle (USP), which form a heterodimer. The dimer recognizes various hormone response elements and the effect of these elements on transcriptional activity of EcR isoforms was determined in vertebrate cells transfected with EcR and USP. Only constitutive activity mediated by the core response elements was preserved after elimination of nonspecific binding sites on the DNA of the vector. The constitutive transcriptional activity was regulated in a complex manner by the N-termini of both EcR and USP, the DBD of USP and the type and number of hormone response elements (HRE). Cooperative effects at oligomeric response elements particularly DR1 depended on the type of ecdysteroid response element and the N-termini of EcR and USP. The DBD of USP abolishes or attenuates synergistic effects. The data show that in the absence of hormone, transcriptional activity is regulated in a complex manner that offers additional possibilities for ecdysteroid receptor mediated gene regulation during development.


Adolescents with ADHD: patterns of behavioral adjustment, academic functioning, and treatment utilization.
Adolescents with attention deficit hyperactivity disorder (ADHD) were compared with a control group on a comprehensive assessment battery. More ADHD teenagers had oppositional defiant disorder (68%) and conduct disorder (39%) and were rated as more impaired in social competence, behavioral and emotional adjustment, and school performance by parents and teachers than control teens. The ADHD youths, however, rated themselves as better adjusted than did their parents and teachers, differing only from controls in depressive symptoms and antisocial acts. Poorer performances in verbal learning and vigilance and greater ADHD behaviors during a math task also distinguished the ADHD from control teenagers.


The timing of neural activity during shifts of spatial attention.
We developed a new experimental task to investigate the relative timing of neural activity during shifts of spatial attention with event-related potentials. The task enabled the investigation of nonlateralized as well as lateralized neural activity associated with spatial shifts. Participants detected target stimuli within one of two peripheral streams of visual letters. Colored letters embedded within the streams indicated which stream was to be used for target detection, signaling that participants should "hold" or "shift" their current focus of spatial attention. A behavioral experiment comparing performance in these focused-attention conditions with performance in a divided-attention condition confirmed the efficacy of the spatial cues. Another behavioral experiment showed that overt shifts of spatial attention were mainly complete by around 400 msec, placing an upper boundary for isolating neural activity that was instrumental in controlling spatial shifts. Event-related potentials recorded during a covert version of the focused-attention task showed a large amount of nonlateralized neural activity associated with spatial shifts, with significant effects starting around 330 msec. The effects started over posterior scalp regions, where they remained pronounced. Transient effects were also observed over frontal scalp regions. The results are compatible with a pivotal role of posterior parietal areas in initiating shifts of spatial attention.


T-cell metagene predicts a favorable prognosis in estrogen receptor-negative and HER2-positive breast cancers.
Lymphocyte infiltration (LI) is often seen in breast cancer but its importance remains controversial. A positive correlation of human epidermal growth factor receptor 2 (HER2) amplification and LI has been described, which was associated with a more favorable outcome. However, specific lymphocytes might also promote tumor progression by shifting the cytokine milieu in the tumor. Affymetrix HG-U133A microarray data of 1,781 primary breast cancer samples from 12 datasets were included. The correlation of immune system-related metagenes with different immune cells, clinical parameters, and survival was analyzed. A large cluster of nearly 600 genes with functions in immune cells was consistently obtained in all datasets. Seven robust metagenes from this cluster can act as surrogate markers for the amount of different immune cell types in the breast cancer sample. An IgG metagene as a marker for B cells had no significant prognostic value. In contrast, a strong positive prognostic value for the T-cell surrogate marker (lymphocyte-specific kinase (LCK) metagene) was observed among all estrogen receptor (ER)-negative tumors and those ER-positive tumors with a HER2 overexpression. Moreover ER-negative tumors with high expression of both IgG and LCK metagenes seem to respond better to neoadjuvant chemotherapy. Precise definitions of the specific subtypes of immune cells in the tumor can be accomplished from microarray data. These surrogate markers define subgroups of tumors with different prognosis. Importantly, all known prognostic gene signatures uniformly assign poor prognosis to all ER-negative tumors. In contrast, the LCK metagene actually separates the ER-negative group into better or worse prognosis.


Recent genetic findings in schizophrenia and their therapeutic relevance.
Over 100 loci are now associated with schizophrenia risk as identified by single nucleotide polymorphisms (SNPs) in genome-wide association studies. These findings mean that 'genes for schizophrenia' have unquestionably been found. However, many questions remain unanswered, including several which affect their therapeutic significance. The SNPs individually have minor effects, and even cumulatively explain only a modest fraction of the genetic predisposition. The remainder likely results from many more loci, from rare variants, and from gene-gene and gene-environment interactions. The risk SNPs are almost all non-coding, meaning that their biological significance is unclear; probably their effects are mediated via an influence on gene regulation, and emerging evidence suggests that some key molecular events occur during early brain development. The loci include novel genes of unknown function as well as genes and pathways previously implicated in the pathophysiology of schizophrenia, e.g. NMDA receptor signalling. Genes in the latter category have the clearer therapeutic potential, although even this will be a challenging process because of the many complexities concerning the genetic architecture and mediating mechanisms. This review summarises recent schizophrenia genetic findings and some key issues they raise, particularly with regard to their implications for identifying and validating novel drug targets.


Do patients with fibromyalgia show abnormal neural responses to the observation of pain in others?
Chronic widespread pain is a hallmark of fibromyalgia (FM). Previous neuroimaging studies have reported that the pain neuro-matrix in patients with FM showed augmented activation in response to actual pain. However, the effect of observing pain in others among patients with FM remains poorly understood. Both healthy female control subjects (n=24) and female patients with FM (n=23) underwent functional magnetic resonance imaging while observing a series of color pictures depicting others' hands and feet being injured, and a matched set of control pictures that did not show any painful events. Compared with healthy subjects, patients with FM showed a smaller neural response to pain-related versus neutral stimuli in several neural regions, including the thalamus, anterior cingulate cortex, dorsolateral prefrontal cortex, pre- and post-central gyrus, and supplementary motor area. In contrast to augmented pain processing in response to actual experimental pain, patients with FM did not show an enhanced pain response but generally showed lesser activation in cortical regions known to play a role in processing of pain. These hemodynamic alterations observed in patients with FM suggest that patients with chronic pain may empathize less with others in pain, possibly in order to lessen arousal and aversive self-oriented emotions.


Skewed maturation of memory HIV-specific CD8 T lymphocytes.
Understanding the lineage differentiation of memory T cells is a central question in immunology. We investigated this issue by analysing the expression of the chemokine receptor CCR7, which defines distinct subsets of naive and memory T lymphocytes with different homing and effector capacities and antiviral immune responses to HIV and cytomegalovirus. Ex vivo analysis of the expression of CD45RA and CCR7 antigens, together with in vitro analysis of the cell-division capacity of different memory CD8+ T-cell populations, identified four subsets of HIV- and CMV-specific CD8+ T lymphocytes, and indicated the following lineage differentiation pattern: CD45RA+ CCR7+ --> CD45RA- CCR7+ --> CD45RA- CCR7- --> CD45RA+ CCR7-. Here we demonstrate through analysis of cell division (predominantly restricted to the CCR7+ CD8+ T-cell subsets) that the differentiation of antigen-specific CD8+ T cells is a two-step process characterized initially by a phase of proliferation largely restricted to the CCR7+ CD8+ cell subsets, followed by a phase of functional maturation encompassing the CCR7- CD8+ cell subsets. The distribution of these populations in HIV- and CMV-specific CD8+ T cells showed that the HIV-specific cell pool was predominantly (70%) composed of pre-terminally differentiated CD45RA- CCR7- cells, whereas the CMV-specific cell pool consisted mainly (50%) of the terminally differentiated CD45RA+ CCR7- cells. These results demonstrate a skewed maturation of HIV-specific memory CD8+ T cells during HIV infection.


A Latino Patient-Centered, Evidence-Based Approach to Diabetes Prevention.
Cultural tailoring of evidence-based diabetes prevention program (DPP) interventions is needed to effectively address obesity and its related chronic diseases among Latinos in primary care. This article describes the patient-centered process used to adapt the DPP and reports cultural adaptations. We used a 2-stage formative research process to culturally adapt an evidence-based DPP intervention in the context of primary care. The first stage involved 5 focus groups of Latino patients and interviews with 5 stakeholders (3 with primary care physicians and 2 with medical directors) to inform a first round of adaptations. The second stage included pretesting the stage I-adapted intervention with a Latino patient advisory board to complete a second round of adaptations. Key stakeholders involved in this 2-stage adaptation process included 34 Latino patients who participated in 5 focus groups and 5 physicians and medical directors who participated in key informant interviews during stage I and 11 patients who attended the 16 advisory board meetings and their family members who attended 1 of the meetings during stage II. Using this patient-centered stakeholder-engaged approach, we found the original intervention was largely congruent with the cultural values of the study population. To further strengthen the cultural relevance of the intervention, salient cultural values emphasized by patients and stakeholders underscored the importance of family and community support for behavior change. Accordingly, key adaptations were made to (1) invite family members to the orientation session and at 2 other key timepoints to facilitate family support, (2) provide participants support from the coach and each other via smartphone applications, and (3) provide healthy, easy, low-cost culturally appropriate meals at each group session. The 2-stage approach actively engaging patients, family members, providers, and health care system leaders reinforced the cultural congruence of the existing intervention while further strengthening it with adaptations promoting Latino family and community support.


The number needed to treat (NNT) can be adjusted for bias when the outcome is measured with error.
We consider the number needed to treat (NNT) when the event of interest is defined by dichotomizing a continuous response at a threshold level. If the response is measured with error, the resulting NNT is biased. We consider methods to reduce this bias. Bias adjustment was studied using simulations in which we varied the distributions of the underlying response and measurement error, including both normal and nonnormal distributions. We studied a maximum likelihood estimate (MLE) based on normality assumptions, and also considered a simulation-extrapolation estimate (SIMEX) without such assumptions. The treatment effect across all potential thresholds was summarized using an NNT threshold curve. Crude NNT estimation was substantially biased due to measurement error. The MLE performed well under normality, and it continued to perform well with nonnormal measurement error, but when the underlying response was nonnormal the MLE was unacceptably biased and was outperformed by the SIMEX estimate. The simulation results were also reflected in empirical data from a randomized study of cholesterol-lowering therapy. Ignoring measurement error can lead to substantial bias in NNT, which can have an important practical effect on the interpretation of analyses. Analysis methods that adjust for measurement error bias can be used to assess the sensitivity of NNT estimates to this effect.


Interpreting Viral Deep Sequencing Data with GLUE.
Using deep sequencing technologies such as Illumina's platform, it is possible to obtain reads from the viral RNA population revealing the viral genome diversity within a single host. A range of software tools and pipelines can transform raw deep sequencing reads into Sequence Alignment Mapping (SAM) files. We propose that interpretation tools should process these SAM files, directly translating individual reads to amino acids in order to extract statistics of interest such as the proportion of different amino acid residues at specific sites. This preserves per-read linkage between nucleotide variants at different positions within a codon location. The samReporter is a subsystem of the GLUE software toolkit which follows this direct read translation approach in its processing of SAM files. We test samReporter on a deep sequencing dataset obtained from a cohort of 241 UK HCV patients for whom prior treatment with direct-acting antivirals has failed; deep sequencing and resistance testing have been suggested to be of clinical use in this context. We compared the polymorphism interpretation results of the samReporter against an approach that does not preserve per-read linkage. We found that the samReporter was able to properly interpret the sequence data at resistance-associated locations in nine patients where the alternative approach was equivocal. In three cases, the samReporter confirmed that resistance or an atypical substitution was present at NS5A position 30. In three further cases, it confirmed that the sofosbuvir-resistant NS5B substitution S282T was absent. This suggests the direct read translation approach implemented is of value for interpreting viral deep sequencing data.


Tunable shrink-induced honeycomb microwell arrays for uniform embryoid bodies.
Embryoid body (EB) formation closely recapitulates early embryonic development with respect to lineage commitment. Because it is greatly affected by cell-cell and cell-substrate interactions, the ability to control the initial number of cells in the aggregates and to provide an appropriate substrate are crucial parameters for uniform EB formation. Here we report of an ultra-rapid fabrication and culture method utilizing a laser-jet printer to generate closely arrayed honeycomb microwells of tunable sizes for the induction of uniform EBs from single cell suspension. By printing various microwell patterns onto pre-stressed polystyrene sheets, and through heat induced shrinking, high aspect micromolds are generated. Notably, we achieve rounded bottom polydimethylsiloxane (PDMS) wells not easily achievable with standard microfabrication methods, but critical to achieve spherical EBs. Furthermore, by simply controlling the size of the microwells and the concentration of the cell suspension we can control the initial size of the cell aggregate, thus influencing lineage commitment. In addition, these microwells are easily adaptable and scalable to most standard well plates and easily integrated into commercial liquid handling systems to provide an inexpensive and easy high throughput compound screening platform.


Acute and delayed effects of the neonicotinoid insecticide thiacloprid on seven freshwater arthropods.
Ecotoxicological risk assessment of contaminants often is based on toxicity tests with continuous-exposure profiles. However, input of many contaminants (e.g., insecticides) to surface waters typically occurs in pulses rather than continuously. Neonicotinoids are a new group of insecticides, and little is known about their toxicity to nontarget freshwater organisms and potential effects on freshwater ecosystems. The aim of the present research was to assess effects of short-term (24-h) exposure to the neonicotinoid insecticide thiacloprid, including a postexposure observation period. A comparison of several freshwater insect and crustacean species showed an increase of sensitivity by three orders of magnitude in the following order: Daphnia magna < Asellus aquaticus = Gammarus pulex < Simpetrum striolatum < Culex pipiens = Notidobia ciliaris = Simulium latigonium, with median lethal concentrations (LC50s) of 4,400, 153, 190, 31.2, 6.78, 5.47, and 5.76 mug/L, respectively (postexposure observation 11-30 d). Thiacloprid caused delayed lethal and sublethal effects, which were observed after 4 to 12 d following exposure. Reduction in LC50s found when postexposure observation was extended from 1 d to a longer period (11-30 d) was up to >50-fold. Hence, delayed effects occurring after short-term exposure should be considered in risk assessment. The 5% hazardous concentration (HC5) of thiacloprid obtained in the present study (0.72 microg/L) is more than one order of magnitude below the currently predicted worst-case environmental concentrations in surface water. Concerning the selection of test organisms, we observed that the widely employed test organism D. magna is least sensitive among the arthropods tested and that, for neonicotinoid insecticides, an insect like the mosquito C. pipiens would be more suitable for predicting effects on sensitive species.