FIGO stage IIIC endometrial carcinoma: resection of macroscopic nodal disease and other determinants of survival.
The objective of this study was to evaluate the potential survival benefit of debulking macroscopic adenopathy and other clinical prognostic factors among patients with node-positive endometrial carcinoma. Demographic, operative, pathologic, and follow-up data were abstracted retrospectively for 41 eligible patients with FIGO stage IIIC endometrial cancer. Survival curves were generated using the Kaplan-Meier method and statistical comparisons were performed using the log rank test, logistic regression analysis, and the Cox proportional hazards regression model. All patients had positive pelvic lymph nodes and 20 patients (48.8%) had positive para-aortic lymph nodes. Postoperatively, all patients received whole pelvic radiation therapy, 17 received extended-field radiation therapy, and 15 patients received chemotherapy. The median disease-specific survival (DSS) time for all patients was 30.6 months (median follow-up 34. 0 months). Patients with completely resected macroscopic lymphadenopathy had a significantly longer median DSS time (37.5 months), compared to patients left with gross residual nodal disease (8.8 months, P = 0.006). On multivariate analysis, independent predictors of DSS were gross residual nodal disease (HR 7.96, 95% CI 2.54-24.97, P < 0. 001), age > or = 65 years (HR 6.22, 95% CI 2.05-18.87, P = 0.001), and the administration of adjuvant chemotherapy (HR 0.22, 95% CI 0.07-0.76, P = 0.016). We conclude that in patients with stage IIIC endometrial carcinoma, complete resection of macroscopic nodal disease and the administration of adjuvant chemotherapy, in addition to directed radiation therapy, are associated with improved survival.


Alteration of tight junctional permeability in the rat parotid gland after isoproterenol stimulation.
The permeability of junctional complexes to ultrastructural tracers of different molecular weight and the freeze-fracture appearance of junctional structure were investigated in the resting and stimulated rat parotid gland. Tracers were administered retrogradely via the main excretory duct, and allowed to flow by gravity (16 mmHg) into the gland for 15-60 min. Secretion was induced in some animals by intraperitoneal injection of isoproterenol. In resting glands, the tracers microperoxidase , cytochrome c, myoglobin, tyrosinase (subunits), and hemoglobin were restricted to the luminal space of the acini and ducts. In glands stimulated 1-4 h before tracer administration, reaction product for microperoxidase , cytochrome c, myoglobin, and tyrosinase was found in the intercellular and interstitial spaces, whereas hemoglobin was usually retained in the lumina. In contrast, horseradish peroxidase and lactoperoxidase appeared to penetrate the tight junctions and reaction product was localized in the extracellular spaces in both resting and stimulated glands. Diffuse cytoplasmic staining for horseradish peroxidase and lactoperoxidase was frequently observed in acinar and duct cells. The distribution of horseradish peroxidase was similar in both Sprague-Dawley and Wistar-Furth rats, and at concentrations of 0.1-10 mg/ml in the tracer solution. Freeze-fracture replicas of stimulated acinar cells revealed an increased irregularity of the tight junction meshwork, but no obvious gaps or discontinuities were observed. These findings indicate that (a) tight junctions in the resting rat parotid gland are impermeable to tracers of molecular weight greater than or equal to 1,900; (b) stimulation with isoproterenol results in a transient increase in junctional permeability allowing passage of tracers of molecular weight less than or equal to 34,500; (c) junctional permeability cannot be directly correlated with junctional structure; and (d) the behavior of horseradish peroxidase and lactoperoxidase in the rat parotid gland is inconsistent with their molecular weights. Cell membrane damage due to the enzymatic activity or binding of these two tracers may account for the observed distribution.


Heterogeneity of Treatment Effects of Long-Acting Injectable Antipsychotic Medications.
To investigate subgroup responses to long-acting injectable (LAI) medications haloperidol decanoate (HD) and paliperidone palmitate (PP) in a randomized controlled trial that found no difference between the treatments on the primary outcome of efficacy failure. A Comparison of Long-Acting Injectable Medications for Schizophrenia (ACLAIMS) enrolled 311 participants from March 2011 to July 2013 meeting DSM-IV-TR criteria for diagnoses of schizophrenia or schizoaffective disorder at risk of relapse due to medication nonadherence or substance abuse. Participants were randomly assigned to double-blinded treatment with HD or PP and followed for up to 2 years. A committee blinded to treatment assignment adjudicated efficacy failure on the basis of participants' meeting at least 1 of these criteria: psychiatric hospitalization, crisis stabilization, increased outpatient visits, could not discontinue oral antipsychotic, discontinued assigned LAI due to inadequate therapeutic benefit, or ongoing or repeated need for adjunctive oral antipsychotic medication. Survival analyses examined modification of treatment effects on efficacy failure by age, sex, race, substance abuse, baseline symptom severity, and baseline adherence. Mixed-effect linear models and analysis of covariance examined this modification on safety outcomes. An interaction between age and treatment (P = .009) revealed younger participants assigned HD had longer time to efficacy failure than those assigned PP. Interactions were not significant between treatment group and sex, race, substance use disorder, baseline symptom severity, or baseline adherence. An interaction of treatment and age on akathisia (P = .047) found an advantage for PP that was larger among younger persons. An advantage for HD on serum prolactin levels was larger among younger women (P = .033). Among younger persons, HD was associated with lower rates of efficacy failure than PP. Age effects on adverse effects were mixed. Age-related heterogeneity of antipsychotic treatment effects warrants further investigation and consideration in clinical practice. ClinicalTrials.gov identifier: NCT01136772.


Anaerobic co-digestion: A critical review of mathematical modelling for performance optimization.
Anaerobic co-digestion (AcoD) is a pragmatic approach to simultaneously manage organic wastes and produce renewable energy. This review demonstrates the need for improving AcoD modelling capacities to simulate the complex physicochemical and biochemical processes. Compared to mono-digestion, AcoD is more susceptible to process instability, as it operates at a higher organic loading and significant variation in substrate composition. Data corroborated here reveal that it is essential to model the transient variation in pH and inhibitory intermediates (e.g. ammonia and organic acids) for AcoD optimization. Mechanistic models (based on the ADM1 framework) have become the norm for AcoD modelling. However, key features in current AcoD models, especially relationships between system performance and co-substrates' properties, organic loading, and inhibition mechanisms, remain underdeveloped. It is also necessary to predict biogas quantity and composition as well as biosolids quality by considering the conversion and distribution of sulfur, phosphorus, and nitrogen during AcoD.


Affinity-based chromatography utilizing genetically engineered proteins. Interaction of Bordetella pertussis adenylate cyclase with calmodulin.
An engineered calmodulin differs from vertebrate calmodulin in its ability to activate Bordetella pertussis adenylate cyclase, and this difference has been utilized as the basis for a new purification protocol for the adenylate cyclase. VU-8 calmodulin, in which 3 glutamic acid residues (residues 82-84) have been substituted with 3 lysine residues, has a 1000-fold lower apparent affinity for the adenylate cyclase, compared to vertebrate calmodulin, and decreased maximal activity. Because of the relatively calcium-independent nature of the interaction between calmodulin and the cyclase, the use of calmodulin-Sepharose conjugates in the purification of the cyclase requires the use of chaotropic agents for elution. However, when immobilized VU-8 calmodulin was tested as a calcium-dependent, affinity-based, adsorption chromatography step in the purification of the cyclase from culture media or bacterial extracts, the enzyme bound to the column in a calcium-dependent manner, and a nearly homogeneous enzyme was obtained in high yield. These results demonstrate the feasibility of using engineered calmodulins that have selective differences in activity for the rational design of rapid purification protocols for calmodulin-binding proteins as well as indicate the importance of the conserved negative charge cluster at residues 82-84 of calmodulin for activation of this cyclase.


Development of hepatocellular carcinoma in a murine model of nonalcoholic steatohepatitis induced by use of a high-fat/fructose diet and sedentary lifestyle.
Obesity is increasingly prevalent, strongly associated with nonalcoholic liver disease, and a risk factor for numerous cancers. Here, we describe the liver-related consequences of long-term diet-induced obesity. Mice were exposed to an extended obesity model comprising a diet high in trans-fats and fructose corn syrup concurrent with a sedentary lifestyle. Livers were assessed histologically using the nonalcoholic fatty liver disease (NAFLD) activity score (Kleiner system). Mice in the American Lifestyle-Induced Obesity Syndrome (ALIOS) model developed features of early nonalcoholic steatohepatitis at 6 months (mean NAFLD activity score = 2.4) and features of more advanced nonalcoholic steatohepatitis at 12 months, including liver inflammation and bridging fibrosis (mean NAFLD activity score = 5.0). Hepatic expression of lipid metabolism and insulin signaling genes were increased in ALIOS mice compared with normal chow-fed mice. Progressive activation of the mouse hepatic stem cell niche in response to ALIOS correlated with steatosis, fibrosis, and inflammation. Hepatocellular neoplasms were observed in 6 of 10 ALIOS mice after 12 months. Tumors displayed cytological atypia, absence of biliary epithelia, loss of reticulin, alteration of normal perivenular glutamine synthetase staining (absent or diffuse), and variable α-fetoprotein expression. Notably, perivascular tumor cells expressed hepatic stem cell markers. These studies indicate an adipogenic lifestyle alone is sufficient for the development of nonalcoholic steatohepatitis, hepatic stem cell activation, and hepatocarcinogenesis in wild-type mice.


Beyond parity: primary care physicians' perspectives on access to mental health care.
About two-thirds of primary care physicians (PCPs) reported in 2004-05 that they could not get outpatient mental health services for patients-a rate that was at least twice as high as that for other services. Shortages of mental health care providers, health plan barriers, and lack of coverage or inadequate coverage were all cited by PCPs as important barriers to mental health care access. The probability of having mental health access problems for patients varied by physician practice, health system, and policy factors. The results suggest that implementing mental health parity nationally will reduce some but not all of the barriers to mental health care.


Minimal change chronic pancreatitis.
In patients with severe abdominal pain, of pancreatic origin, there are a few with minimal or equivocal findings on pancreatic investigation and in whom the aetiology of their pancreatic disease is elusive. The findings and outcome in 16 of these patients (four men and 12 women) who underwent resection are reported. Pancreatic imaging showed minimal or equivocal findings in all 16; pancreas divisum was present in five. All were managed conservatively at first but resection was required for progression of symptoms. A drainage procedure was performed initially in five patients but relief of pain was at best transitory before further surgery was required. Partial resection was needed in 12, of whom eight required subsequent completion pancreatectomy and four had a one stage total resection. Nine patients are currently pain free after resection or are very much improved, while six are no better and one patient has died from an unrelated cause. Histology of resected specimens showed chronic inflammatory changes accompanied by subtle non-inflammatory changes in all but one. These changes include duct proliferation, duct complex formation, adenomatous nodules, and acinar cell atrophy, the significance of which is unclear. These findings suggest a syndrome of minimal macroscopic and radiological change chronic pancreatitis with pain as its chief clinical feature and a distinct histology, the aetiology of which is unclear. It seems that there is a distinct syndrome of minimal change pancreatitis, among the group of patients which presents with the clinical features of chronic pancreatitis.


The interaction between the Drosophila secreted protein argos and the epidermal growth factor receptor inhibits dimerization of the receptor and binding of secreted spitz to the receptor.
Drosophila Argos (Aos), a secreted protein with an epidermal growth factor (EGF)-like domain, has been shown to inhibit the activation of the Drosophila EGF receptor (DER). However, it has not been determined whether Aos binds directly to DER or whether regulation of the DER activation occurs through some other mechanism. Using DER-expressing cells (DER/S2) and a recombinant DER extracellular domain-Fc fusion protein (DER-Fc), we have shown that Aos binds directly to the extracellular domain of DER with its carboxyl-terminal region, including the EGF-like domain. Furthermore, Aos can block the binding of secreted Spitz (sSpi), a transforming growth factor alpha-like ligand of DER, to the extracellular domain of DER. We observed that sSpi stimulates the dimerization of both the soluble DER extracellular domain (sDER) and the intact DER in the DER/S2 cells and that Aos can block the sSpi-induced dimerization of both sDER and intact DER. Moreover, we have shown that, by directly interacting with DER, Aos and SpiAos (a chimeric protein that is composed of the N-terminal region of Spi and the C-terminal region of Aos) inhibit the dimerization and phosphorylation of DER that are induced by DER's overexpression in the absence of sSpi. These results indicate that Aos exerts its inhibitory function through dual molecular mechanisms: by blocking both the receptor dimerization and the binding of activating ligand to the receptor. This is the first description of this novel inhibitory mechanism for receptor tyrosine kinases.


Carer satisfaction with end-of-life care in Powys, Wales: a cross-sectional survey.
A cross-sectional postal survey of bereaved carers was conducted in order to examine levels of satisfaction with services provided for people in their last year of life in the rural county of Powys, Wales, UK. A self-complete questionnaire, using a modified version of the Views of Informal Carers - Evaluation of Services instrument was sent to all bereaved carers of all those people dying of cancer in Powys between 1 April 1999 and 30 June 2001. Eight hundred and five (out of a possible of 815 people) were contacted and 407 agreed to receive the questionnaire. Out of these 407 individuals, 301 (74% of those who agreed to receive a questionnaire and 37% of the 815 contacted) returned a completed questionnaire. A single reminder letter was sent to non-responders. It was found that the majority of those who received help from district nurses or practice nurses (90%) said that they were excellent or good. However, nearly 40% of respondents reported needing more nursing help. More help was also needed from social care services. For 103 out of the 301 respondents, it was known that the deceased person wanted to die at home; only 44 did so. Only one-fifth of respondents had the opportunity to talk to someone from health and social services after their bereavement; a large majority (four-fifths) found this helpful. One-tenth of respondents reported untreated pain at home; however, there was evidence for an increasing proportion of those treated having received good pain relief. Although there are high levels of satisfaction with care and services received by Powys residents, deficits exist in relation to: symptom control, nursing help, assistance from social services with transport and bathing, communication, and bereavement support.


Zika in the Americas, year 2: What have we learned? What gaps remain? A report from the Global Virus Network.
In response to the outbreak of Zika virus (ZIKV) infection in the Western Hemisphere and the recognition of a causal association with fetal malformations, the Global Virus Network (GVN) assembled an international taskforce of virologists to promote basic research, recommend public health measures and encourage the rapid development of vaccines, antiviral therapies and new diagnostic tests. In this article, taskforce members and other experts review what has been learned about ZIKV-induced disease in humans, its modes of transmission and the cause and nature of associated congenital manifestations. After describing the make-up of the taskforce, we summarize the emergence of ZIKV in the Americas, Africa and Asia, its spread by mosquitoes, and current control measures. We then review the spectrum of primary ZIKV-induced disease in adults and children, sites of persistent infection and sexual transmission, then examine what has been learned about maternal-fetal transmission and the congenital Zika syndrome, including knowledge obtained from studies in laboratory animals. Subsequent sections focus on vaccine development, antiviral therapeutics and new diagnostic tests. After reviewing current understanding of the mechanisms of emergence of Zika virus, we consider the likely future of the pandemic.


FoxC1 is essential for vascular basement membrane integrity and hyaloid vessel morphogenesis.
Alterations in FOXC1 dosage lead to a spectrum of highly penetrant, ocular anterior segment dysgenesis phenotypes. The most serious outcome is the development of glaucoma, which occurs in 50% to 75% of patients. Therefore, the need to identify specific pathways and genes that interact with FOXC1 to promote glaucoma is great. In this study, the authors investigated the loss of foxC1 in the zebrafish to characterize phenotypes and gene interactions that may impact glaucoma pathogenesis. Morpholino knockdown in zebrafish, RNA and protein marker analyses, transgenic reporter lines, and angiography, along with histology and transmission electron microscopy, were used to study foxC1 function and gene interactions. Zebrafish foxC1 genes were expressed dynamically in the developing vasculature and periocular mesenchyme during development. Multiple ocular and vascular defects were found after the knockdown of foxC1. Defects in the hyaloid vasculature, arteriovenous malformations, and coarctation of the aorta were observed with maximal depletion of foxC1. Partial loss of foxC1 resulted in CNS and ocular hemorrhages, defects in intersegmental vessel patterning, and increased vascular permeability. To investigate the basis for these disruptions, the ultrastructure of foxC1-depleted hyaloid vascular cells was studied. These experiments, along with laminin-111 immunoreactivity, revealed disruptions in basement membrane integrity. Finally, codepletion of laminin alpha-1 and foxC1 uncovered a genetic interaction between these genes during development. Genetic interactions between FOXC1 and basement membrane components influence vascular stability and may impact glaucoma development and increase stroke risk in FOXC1 patients.


Intoxication with bourbon versus vodka: effects on hangover, sleep, and next-day neurocognitive performance in young adults.
This study assessed the effects of heavy drinking with high or low congener beverages on next-day neurocognitive performance, and the extent to which these effects were mediated by alcohol-related sleep disturbance or alcoholic beverage congeners, and correlated with the intensity of hangover. Healthy heavy drinkers age 21 to 33 (n = 95) participated in 2 drinking nights after an acclimatization night. They drank to a mean of 0.11 g% breath alcohol concentration on vodka or bourbon one night with matched placebo the other night, randomized for type and order. Polysomnography recordings were made overnight; self-report and neurocognitive measures were assessed the next morning. After alcohol, people had more hangover and more decrements in tests requiring both sustained attention and speed. Hangover correlated with poorer performance on these measures. Alcohol decreased sleep efficiency and rapid eye movement sleep, and increased wake time and next-day sleepiness. Alcohol effects on sleep correlated with hangover but did not mediate the effects on performance. No effect of beverage congeners was found except on hangover severity, with people feeling worse after bourbon. Virtually no sex differences appeared. As drinking to this level affects complex cognitive abilities, safety could be affected, with implications for driving and for safety-sensitive occupations. Congener content affects only how people feel the next day so does not increase risk. The sleep disrupting effects of alcohol did not account for the impaired performance so other mechanisms of effect need to be sought. As hangover symptoms correlate with impaired performance, these might be contributing to the impairment.


Mindfulness-based stress reduction for the treatment of irritable bowel syndrome symptoms: a randomized wait-list controlled trial.
Irritable bowel syndrome (IBS) is a functional disorder of the lower gastrointestinal (GI) tract affected by stress, which may benefit from a biopsychosocial treatment approach such as mindfulness-based stress reduction (MBSR). A treatment as usual (TAU) wait-list controlled trial was conducted in Calgary, Canada to investigate the impact of MBSR on IBS symptoms. It was hypothesized that MBSR patients would experience greater reduction in overall IBS symptom severity and self-reported symptoms of stress relative to control patients. Ninety patients diagnosed with IBS using the Rome III criteria were randomized to either an immediate MBSR program (n = 43) or to wait for the next available program (n = 47). Patients completed IBS symptom severity, stress, mood, quality of life (QOL), and spirituality scales pre- and post-intervention or waiting period and at 6-month follow-up. Intent-to-treat linear mixed model analyses for repeated measures were conducted, followed by completers analyses. While both groups exhibited a decrease in IBS symptom severity scores over time, the improvement in the MBSR group was greater than the controls and was clinically meaningful, with symptom severity decreasing from constantly to occasionally present. Pre- to post-intervention dropout rates of 44 and 23 % for the MBSR and control groups, respectively, were observed. At 6-month follow-up, the MBSR group maintained a clinically meaningful improvement in overall IBS symptoms compared to the wait-list group, who also improved marginally, resulting in no statistically significant differences between groups at follow-up. Improvements in overall mood, QOL, and spirituality were observed for both groups over time. The results of this trial provide preliminary evidence for the feasibility and efficacy of a mindfulness intervention for the reduction of IBS symptom severity and symptoms of stress and the maintenance of these improvements at 6 months post-intervention. Attention and self-monitoring and/or anticipation of MBSR participation may account for smaller improvements observed in TAU patients.


Sensitization to gliadin induces moderate enteropathy and insulitis in nonobese diabetic-DQ8 mice.
Celiac disease (CD) is frequently diagnosed in patients with type 1 diabetes (T1D), and T1D patients can exhibit Abs against tissue transglutaminase, the auto-antigen in CD. Thus, gliadin, the trigger in CD, has been suggested to have a role in T1D pathogenesis. The objective of this study was to investigate whether gliadin contributes to enteropathy and insulitis in NOD-DQ8 mice, an animal model that does not spontaneously develop T1D. Gliadin-sensitized NOD-DQ8 mice developed moderate enteropathy, intraepithelial lymphocytosis, and barrier dysfunction, but not insulitis. Administration of anti-CD25 mAbs before gliadin-sensitization induced partial depletion of CD25(+)Foxp3(+) T cells and led to severe insulitis, but did not exacerbate mucosal dysfunction. CD4(+) T cells isolated from pancreatic lymph nodes of mice that developed insulitis showed increased proliferation and proinflammatory cytokines after incubation with gliadin but not with BSA. CD4(+) T cells isolated from nonsensitized controls did not response to gliadin or BSA. In conclusion, gliadin sensitization induced moderate enteropathy in NOD-DQ8 mice. However, insulitis development required gliadin-sensitization and partial systemic depletion of CD25(+)Foxp3(+) T cells. This humanized murine model provides a mechanistic link to explain how the mucosal intolerance to a dietary protein can lead to insulitis in the presence of partial regulatory T cell deficiency.


Functional analysis and classification of phytoplankton based on data from an automated flow cytometer.
Analytical flow cytometry (FCM) is well suited for the analysis of phytoplankton communities in fresh and sea waters. The measurement of light scatter and autofluorescence properties of particles by FCM provides optical fingerprints, which enables different phytoplankton groups to be separated. A submersible version of the CytoSense flow cytometer (the CytoSub) has been designed for in situ autonomous sampling and analysis, making it possible to monitor phytoplankton at a short temporal scale and obtain accurate information about its dynamics. For data analysis, a manual clustering is usually performed a posteriori: data are displayed on histograms and scatterplots, and group discrimination is made by drawing and combining regions (gating). The purpose of this study is to provide greater objectivity in the data analysis by applying a nonmanual and consistent method to automatically discriminate clusters of particles. In other words, we seek for partitioning methods based on the optical fingerprints of each particle. As the CytoSense is able to record the full pulse shape for each variable, it quickly generates a large and complex dataset to analyze. The shape, length, and area of each curve were chosen as descriptors for the analysis. To test the developed method, numerical experiments were performed on simulated curves. Then, the method was applied and validated on phytoplankton cultures data. Promising results have been obtained with a mixture of various species whose optical fingerprints overlapped considerably and could not be accurately separated using manual gating.


Acoustic pharyngometry - A new method to facilitate oral appliance therapy.
There is lack of reliable and accurate methods to predict treatment outcomes of oral appliance (OA) treatment. Acoustic pharyngometry (AP) is a non-invasive technique to evaluate the volume and minimal cross-sectional area of the upper airway, which may prove useful to locate the optimal position of OAs. This retrospective study aimed to evaluate the effect of applying AP to OA treatment of patients with obstructive sleep apnoea (OSA). All patients (n = 244) treated with OAs following an AP protocol at two dental clinics between 2013 and 2018 were invited to participate. A total of 129 patients accepted the invitation, and 120 patients (75 men, 45 women) were included in the analyses. Mean baseline age, BMI and apnoea hypopnea index (AHI) were 59.1 ± 0.9 years, 27.8 ± 0.4 and 21.9 ± 1.1, respectively. Mean follow-up time was 318 ± 24 days. AHI at follow-up was 6.4 ± 0.7, resulting in a treatment success rate of 86.7% (≥50% reduction of baseline AHI). The number of failures (<50% reduction of baseline AHI) did not differ significantly among patients with mild, moderate and severe OSA. 87.6% of the patients reported OA usage every night, and 95.5% reported > 5 hours usage per night, when worn. The AP protocol applied seems to contribute to the excellent effect of OA treatment in this study. Further research on the application of AP in OA treatment is necessary in order to clarify its possible beneficial contribution to improving OA therapy.


A local area analysis of racial, ethnic, and neighborhood disparities in breast cancer staging.
Few studies have examined the role of neighborhood socioeconomic condition in shaping breast cancer disparities in defined local areas. We tested associations between three measures of neighborhood socioeconomic condition (poverty, median income, and a composite neighborhood score) on breast cancer staging in two urban counties of the state of New Jersey. Data for these counties were obtained from the New Jersey Surveillance, Epidemiology, and End Results tumor registry and were selected because of their large racial/ethnic and socioeconomic diversity and pilot prevention efforts taking place in these areas. Our study population included Black, Latina, and White women (N = 4,589) diagnosed with breast cancer from 1999 to 2004. Each cancer case was geocoded and linked to socioeconomic data obtained from the 2000 U.S. census. Census tracts served as proxies for neighborhoods. Logistic regression models accounting for clustering of individuals within neighborhoods were fitted with Generalized Estimating Equations. Women living in neighborhoods with lower versus higher neighborhood scores were significantly more likely to have advanced-stage disease (odds ratio, 1.6; confidence intervals, 1.1-2.3), after adjusting for age at diagnosis and race/ethnicity. In analyses stratified by race/ethnicity, results remained significant for all neighborhood measures for White and Black women, but not for Latinas. Moreover, neighborhood poverty showed a weaker socioeconomic gradient in breast cancer staging among White women. Our study findings support the use of a multidimensional neighborhood index to better capture differences in cancer staging risk across racial/ethnic groups and provides evidence that population-based cancer data could be used to identify local needs specific to local populations.


Phase 1 Study of a Sulforaphane-Containing Broccoli Sprout Homogenate for Sickle Cell Disease.
Sickle cell disease (SCD) is the most common inherited hemoglobinopathy worldwide. Our previous results indicate that the reduced oxidative stress capacity of sickle erythrocytes may be caused by decreased expression of NRF2 (Nuclear factor (erythroid-derived 2)-like 2), an oxidative stress regulator. We found that activation of NRF2 with sulforaphane (SFN) in erythroid progenitors significantly increased the expression of NRF2 targets HMOX1, NQO1, and HBG1 (subunit of fetal hemoglobin) in a dose-dependent manner. Therefore, we hypothesized that NRF2 activation with SFN may offer therapeutic benefits for SCD patients by restoring oxidative capacity and increasing fetal hemoglobin concentration. To test this hypothesis, we performed a Phase 1, open-label, dose-escalation study of SFN, contained in a broccoli sprout homogenate (BSH) that naturally contains SFN, in adults with SCD. The primary and secondary study endpoints were safety and physiological response to NRF2 activation, respectively. We found that BSH was well tolerated, and the few adverse events that occurred during the trial were not likely related to BSH consumption. We observed an increase in the mean relative whole blood mRNA levels for the NRF2 target HMOX1 (p = 0.02) on the last day of BSH treatment, compared to pre-treatment. We also observed a trend toward increased mean relative mRNA levels of the NRF2 target HBG1 (p = 0.10) from baseline to end of treatment, but without significant changes in HbF protein. We conclude that BSH, in the provided doses, is safe in stable SCD patients and may induce changes in gene expression levels. We therefore propose investigation of more potent NRF2 inducers, which may elicit more robust physiological changes and offer clinical benefits to SCD patients. Trial registration: ClinicalTrials.gov NCT01715480.


Volunteer, lay tutors' experiences of the Chronic Disease Self-Management Course: being valued and adding value.
Chronic disease is a public health issue that could be addressed, in part, by increasing the ability of individuals to better manage their condition and its consequences on a day-to-day basis. One intervention designed to facilitate this is the Chronic Disease Self Management Course (CDSMC) that is delivered by volunteer, lay tutors who themselves have a chronic disease. Although there is growing evidence of course effectiveness for participants, the experiences of tutors have been neglected. This study aims to address this omission. Telephone interviews were conducted with 11 (six male) tutors: all interviews were transcribed and thematically analysed. Being a volunteer lay-tutor was perceived to be an enjoyable and valuable experience despite the challenges associated with course delivery, such as organizational demands and managing the diverse needs of mixed groups of chronic disease participants that led to a tension between disease-specific needs and the generic approach of the course. Being valued and adding value to the lives of others were key benefits of being a volunteer tutor, along with increased confidence that they were doing something positive for others. Course delivery prompted the initiation and maintenance of tutors' own self-management behaviours.


A RESEARCH UPON ANAESTHESIA.
THE CONCLUSIONS WHICH HAVE BEEN REACHED BY THE SERIES OF EXPERIMENTS RECORDED IN THE PRESENT MEMOIR ARE: First. Lowered arterial pressure has a comparatively feeble effect upon the respiration, but when the pressure falls sufficiently, respiratory depression does occur. Second. Even excessive lowering of blood pressure primarily stimulates the vaso-motor centre, the sensibility of the centre being evidently necessary to the automatic regulation of the circulation. Third. The circulation recovers itself more slowly after profound etherization than after a like chloroform narcosis. Fourth. It is possible for ether as well as chloroform to produce death some hours after the cessation of its administration, at a time when the cerebrum has long freed itself from distinct evidences of the narcotic, so that consciousness and intellectual action have been restored. In applying these conclusions to the subject of practical anaesthesia it is evident that the depression of the circulation produced by chloroform has effect upon the respiratory centres only when the pressure has fallen very low, and whilst it may be a factor in the production of respiratory failure during chloroformization, the failure must be chiefly due to the direct influence exercised by the drug upon the respiratory centres. Clinical experience shows that nausea and general depression are more pronounced after the use of ether than after the use of chloroform, a difference which is strongly insisted upon by the advocates of chloroform as an important agent in favor of that anaesthetic. Our research confirms clinical observation, and experimentally shows that the depression of the circulation produced by ether is more permanent than that caused by chloroform; the reason probably being the large amount of ether which is necessary to produce profound narcosis, with lowering of the arterial pressure; an amount so large that it can neither be burned up in the system nor yet eliminated in the time which would be necessary for the much smaller amount of chloroform to be gotten rid of after chloroformization.


Most neutralizing human monoclonal antibodies target novel epitopes requiring both Lassa virus glycoprotein subunits.
Lassa fever is a severe multisystem disease that often has haemorrhagic manifestations. The epitopes of the Lassa virus (LASV) surface glycoproteins recognized by naturally infected human hosts have not been identified or characterized. Here we have cloned 113 human monoclonal antibodies (mAbs) specific for LASV glycoproteins from memory B cells of Lassa fever survivors from West Africa. One-half bind the GP2 fusion subunit, one-fourth recognize the GP1 receptor-binding subunit and the remaining fourth are specific for the assembled glycoprotein complex, requiring both GP1 and GP2 subunits for recognition. Notably, of the 16 mAbs that neutralize LASV, 13 require the assembled glycoprotein complex for binding, while the remaining 3 require GP1 only. Compared with non-neutralizing mAbs, neutralizing mAbs have higher binding affinities and greater divergence from germline progenitors. Some mAbs potently neutralize all four LASV lineages. These insights from LASV human mAb characterization will guide strategies for immunotherapeutic development and vaccine design.


[The Road towards the Responsible and Safe Legalization of Cannabis Use in Portugal].
Recently, the world has seen examples of the legalization of cannabis for recreational purposes. Due to the diversity of experiences in progress, it is urgent to analyze the impacts of this legalization, from a public health perspective. Therefore, this article aims to review the accumulated knowledge in the states and countries where the use of cannabis is legal and to ponder over the relevance of starting a similar path towards legalization in Portugal, thus supporting political decisions to be properly informed and evidence-based. An extensive literature review was performed using databases and scientific journals, such as PubMed, as well as the search of institutional documentation, including the EMCDDA and SICAD. The gathered information provided insights and enabled assessment of (1) the acute and chronic effects of cannabis use on health, (2) the Portuguese situation related to cannabis and (3) the processes and lessons learned after the legalization of cannabis in other countries or states. Given the above, and according to the data presented, the authors argue for a safe and responsible strategy towards the legalization of cannabis use in Portugal. In accordance, a set of concrete proposals are presented. From a public health perspective, it is assumed that the interest of this proposal is to reduce the problematic use of cannabis, to effectively fight against illicit drug trafficking and drug-related crime, as well as health promotion and prevention of addictions and other adverse health impacts. This article reveals that the effects of legalization might, contrary to general beliefs, generate positive results with respect to these aims, given that there will be greater control on the market, price, quality, and information - to name a few - if implementation occurs with proper consideration and definition. The debate on the responsible and safe legalization of cannabis use in Portugal should be open and promoted, based on a public health perspective.


Harmonizing DTI measurements across scanners to examine the development of white matter microstructure in 803 adolescents of the NCANDA study.
Neurodevelopment continues through adolescence, with notable maturation of white matter tracts comprising regional fiber systems progressing at different rates. To identify factors that could contribute to regional differences in white matter microstructure development, large samples of youth spanning adolescence to young adulthood are essential to parse these factors. Recruitment of adequate samples generally relies on multi-site consortia but comes with the challenge of merging data acquired on different platforms. In the current study, diffusion tensor imaging (DTI) data were acquired on GE and Siemens systems through the National Consortium on Alcohol and NeuroDevelopment in Adolescence (NCANDA), a multi-site study designed to track the trajectories of regional brain development during a time of high risk for initiating alcohol consumption. This cross-sectional analysis reports baseline Tract-Based Spatial Statistic (TBSS) of regional fractional anisotropy (FA), mean diffusivity (MD), axial diffusivity (L1), and radial diffusivity (LT) from the five consortium sites on 671 adolescents who met no/low alcohol or drug consumption criteria and 132 adolescents with a history of exceeding consumption criteria. Harmonization of DTI metrics across manufacturers entailed the use of human-phantom data, acquired multiple times on each of three non-NCANDA participants at each site's MR system, to determine a manufacturer-specific correction factor. Application of the correction factor derived from human phantom data measured on MR systems from different manufacturers reduced the standard deviation of the DTI metrics for FA by almost a half, enabling harmonization of data that would have otherwise carried systematic error. Permutation testing supported the hypothesis of higher FA and lower diffusivity measures in older adolescents and indicated that, overall, the FA, MD, and L1 of the boys were higher than those of the girls, suggesting continued microstructural development notable in the boys. The contribution of demographic and clinical differences to DTI metrics was assessed with General Additive Models (GAM) testing for age, sex, and ethnicity differences in regional skeleton mean values. The results supported the primary study hypothesis that FA skeleton mean values in the no/low-drinking group were highest at different ages. When differences in intracranial volume were covaried, FA skeleton mean reached a maximum at younger ages in girls than boys and varied in magnitude with ethnicity. Our results, however, did not support the hypothesis that youth who exceeded exposure criteria would have lower FA or higher diffusivity measures than the no/low-drinking group; detecting the effects of excessive alcohol consumption during adolescence on DTI metrics may require longitudinal study.


The Abundance of Toxic Genotypes Is a Key Contributor to Anatoxin Variability in Phormidium-Dominated Benthic Mats.
The prevalence of benthic proliferations of the anatoxin-producing cyanobacterium Phormidium are increasing in cobble-bed rivers worldwide. Studies to date have shown high spatial and temporal variability in anatoxin concentrations among mats. In this study we determined anatoxin quotas (toxins per cell) in field samples and compared these results to the conventionally-used concentrations (assessed per dry weight of mat). Three mats were selected at sites in two rivers and were sampled every 2-3 h for 24-26 h. The samples were lyophilized and ground to a fine homogenous powder. Two aliquots of known weights were analyzed for anatoxin congeners using liquid chromatography-mass spectrometry, or digital droplet PCR with Phormidium-specific anaC primers to measure absolute quantities of gene copies. Anatoxin concentrations in the mats varied 59- and 303-fold in the two rivers over the study periods. A similar pattern was observed among gene copies (53- and 2828-fold). When converted to anatoxin quotas there was markedly less variability (42- and 16-fold), but significantly higher anatoxin quotas were observed in mats from the second river (p < 0.001, Student's t-test). There were no obvious temporal patterns with high and low anatoxin concentrations or quotas measured at each sampling time and across the study period. These results demonstrate that variability in anatoxin concentrations among mats is primarily due to the abundance of toxic genotypes. No consistent modulation in anatoxin production was observed during the study, although significant differences in anatoxin quotas among rivers suggest that site-specific physiochemical or biological factors may influence anatoxin production.


Quantification of Leptospira interrogans Survival in Soil and Water Microcosms.
Leptospira interrogans is the etiological agent of leptospirosis, a globally distributed zoonotic disease. Human infection usually occurs through skin exposure with water and soil contaminated with the urine of chronically infected animals. In this study, we aimed to quantitatively characterize the survival of Leptospira interrogans serovar Copenhageni in environmental matrices. We constructed laboratory microcosms to simulate natural conditions and determined the persistence of DNA markers in soil, mud, spring water and sewage using a quantitative PCR (qPCR) and a propidium monoazide (PMA)-qPCR assay. We found that L. interrogans does not survive at high concentrations in the tested matrices. No net growth was detected in any of the experimental conditions and in all cases the concentration of the DNA markers targeted decreased from the beginning of the experiment following an exponential decay with a decreasing decay rate over time. After 12 and 21 days of incubation the spiked concentration of 106L. interrogans cells/ml or g decreased to approximately 100 cells/ml or g in soil and spring water microcosms, respectively. Furthermore, culturable L. interrogans persisted at concentrations under the limit of detection by PMA-qPCR or qPCR for at least 16 days in soil and 28 days in spring water. Altogether, our findings suggest that the environment is not a multiplication reservoir but a temporary carrier of L. interrogans Copenhageni, although the observed prolonged persistence at low concentrations may still enable the transmission of the disease.IMPORTANCE Leptospirosis is a zoonotic disease caused by spirochetes of the genus Leptospira that primarily affects impoverished populations worldwide. Although leptospirosis is transmitted by contact with water and soil, little is known about the ability of the pathogen to survive in the environment. In this study, we quantitatively characterized the survival of L. interrogans in environmental microcosms and found that although it cannot multiply in water, soil or sewage, it survives for extended time periods (days to weeks depending on the matrix). The survival parameters obtained here may help to better understand the distribution of pathogenic Leptospira in the environment and improve the predictions of human infection risks in areas where such infections are endemic.


Intra-articular decorin influences the fibrosis genetic expression profile in a rabbit model of joint contracture.
The goal of this study was to determine whether intra-articular administration of the potentially anti-fibrotic agent decorin influences the expression of genes involved in the fibrotic cascade, and ultimately leads to less contracture, in an animal model. A total of 18 rabbits underwent an operation on their right knees to form contractures. Six limbs in group 1 received four intra-articular injections of decorin; six limbs in group 2 received four intra-articular injections of bovine serum albumin (BSA) over eight days; six limbs in group 3 received no injections. The contracted limbs of rabbits in group 1 were biomechanically and genetically compared with the contracted limbs of rabbits in groups 2 and 3, with the use of a calibrated joint measuring device and custom microarray, respectively. There was no statistical difference in the flexion contracture angles between those limbs that received intra-articular decorin versus those that received intra-articular BSA (66° vs 69°; p = 0.41). Likewise, there was no statistical difference between those limbs that received intra-articular decorin versus those who had no injection (66° vs 72°; p = 0.27). When compared with BSA, decorin led to a statistically significant increase in the mRNA expression of 12 genes (p < 0.01). In addition, there was a statistical change in the mRNA expression of three genes, when compared with those without injection. In this model, when administered intra-articularly at eight weeks, 2 mg of decorin had no significant effect on joint contractures. However, our genetic analysis revealed a significant alteration in several fibrotic genes. Cite this article: Bone Joint Res 2014;3:82-8.


Vav proteins regulate the plasma cell program and secretory Ig production.
Plasma cell (PC) development is initiated following B cell activation and controlled by a B lymphocyte-induced maturation protein (Blimp)-1-dependent program involving the concerted action of several proplasma transcriptional regulators. However, the factors that control Blimp-1 expression remain largely unknown. In this context, mice deficient for all three of the Vav family of proteins (Vav(null)) develop substantial B cell populations, including marginal zone B cells, yet have a virtual absence of serum Igs, indicating that Vav may be specifically required in PC development and Ig production. We show in this study that mature marginal zone B cells from Vav(null) mice proliferate following stimulation with TLR ligands but exhibit severe defects in PC differentiation and Ig secretion. Under conditions inducing PC differentiation, Vav(null) B cells fail to efficiently induce Blimp-1, X box-binding protein-1, J chain, or secretory Ig mu transcripts but express IFN-regulatory factor-4 at levels similar to wild-type cells. These data indicate a previously unknown role for Vav as an upstream regulator of Blimp-1.


Mental Health Services Use Trends in Canadian Veterans: A Population-Based Retrospective Cohort Study in Ontario.
A substantial evidence base in the peer-reviewed literature exists investigating mental illness in the military, but relatively less is documented about mental illness in veterans. This study uses provincial, administrative data to study the use of mental health services by Canadian veterans in Ontario. This was a retrospective cohort study of Canadian Armed Forces and Royal Canadian Mounted Police veterans who were released between 1990 and 2013 and resided in Ontario. Mental health-related primary care physician, psychiatrist, emergency department (ED) visits, and psychiatric hospitalisations were counted. Repeated measures were presented in 5-year intervals, stratified by age at release. The cohort included 23,818 veterans. In the first 5 years following entry into the health care system, 28.9% of veterans had ≥1 mental health-related primary care physician visit, 5.8% visited a psychiatrist at least once, and 2.4% received acute mental health services at an ED. The use of mental health services was consistent over time. Almost 8% of veterans aged 30 to 39 years saw a psychiatrist in the first 5 years after release, compared to 3.5% of veterans aged ≥50 years at release. The youngest veterans at release (<30 years) were the most frequent users of ED services for a mental health-related reason (5.1% had at least 1 ED visit). Understanding how veterans use the health care system for mental health problems is an important step to ensuring needs are met during the transition to civilian life.


Orthogonally protected artificial amino acid as tripod ligand for automated peptide synthesis and labeling with [(99m)Tc(OH(2))(3)(CO)(3)](+).
1,2-Diamino-propionic acid (Dap) is a very strong chelator for the [(99m)Tc(CO)(3)](+) core, yielding small and hydrophilic complexes. We prepared the lysine based Dap derivative l-Lys(Dap) in which the ε-NH(2) group was replaced by the tripod through conjugation to its α-carbon. The synthetic strategy produced an orthogonally protected bifunctional chelator (BFC). The -NH(2) group of the α-amino acid portion is Fmoc- and the -NH(2) of Dap are Boc-protected. Fmoc-l-Lys(Dap(Boc)) was either conjugated to the N- and C-terminus of bombesin BBN(7-14) or integrated into the sequence using solid-phase peptide synthesis (SPPS). We also replaced the native lysine in a cyclic RGD peptide with l-Lys(Dap). For all peptides, quantitative labeling with the [(99m)Tc(CO)(3)](+) core at a 10 μM concentration in PBS buffer (pH = 7.4) was achieved. For comparison, the rhenium homologues were prepared from [Re(OH(2))(3)(CO)(3)](+) and Lys(Dap)-BBN(7-14) or cyclo-(RGDyK(Dap)), respectively. Determination of integrin receptor binding showed low to medium nanomolar affinities for various receptor subtypes. The IC(50) of cyclo-(RGDyK(Dap[Re(CO)(3)])) for α(v)β(3) is 7.1 nM as compared to 3.1 nM for nonligated RGD derivative. Biodistribution studies in M21 melanoma bearing nude mice showed reasonable α(v)β(3)-integrin specific tumor uptake. Altogether, orthogonally protected l-Lys(Dap) represents a highly versatile building block for integration in any peptide sequence. Lys(Dap)-precursors allow high-yield (99m)Tc-labeling with [(99m)Tc(OH(2))(3)(CO)(3)](+), forming small and hydrophilic complexes, which in turn leads to peptide radiopharmaceuticals with excellent in vivo characteristics.


A new interpretation of chlorine leaving group kinetic isotope effects; a theoretical approach.
The chlorine leaving group kinetic isotope effects (KIEs) for the S(N)2 reactions between methyl chloride and a wide range of anionic, neutral, and radical anion nucleophiles were calculated in the gas phase and, in several cases, using a continuum solvent model. In contrast to the expected linear dependence of the chlorine KIEs on the C(alpha)-Cl bond order in the transition state, the KIEs fell in a very small range (1.0056-1.0091), even though the C(alpha)-Cl transition state bond orders varied widely from approximately 0.32 to 0.78, a range from reactant-like to very product-like. This renders chlorine KIEs, and possibly other leaving-group KIEs, less useful for studies of reaction mechanisms than commonly assumed. A partial explanation for this unexpected relationship between the C(alpha)-Cl transition state bond order and the magnitude of the chlorine KIE is presented.


Targeting modifiable risk factors in age-related macular degeneration in optometric practice in Sweden.
The purpose of this study was to investigate the extent to which ophthalmologists and optometrists in Sweden recommend the use of nutritional supplements, changes in diet, or smoking cessation to patients who are at risk of or with signs of age-related macular degeneration (AMD). In addition, this study also examined how these practitioners rate the strength of evidence for nutritional supplements in AMD management and which sources of information they consult to determine supplement recommendations for the prevention or treatment of AMD. This study implemented a cross-sectional design using data from a questionnaire. All Swedish optometrists and ophthalmologists who were registered in the membership databases of their respective professional organizations were invited to participate. The questionnaire contained 18 forced choice questions and one free text question and was organized into the following four sections: use of nutritional supplements, dietary advice, smoking and eye diseases, and strength of evidence and the sources of information regarding nutritional supplement interventions. The response rate was 40.3% for optometrists and 5% for ophthalmologists. Optometrists were more likely than ophthalmologists to recommend nutritional supplements in AMD and provided significantly more advice about diet than did the ophthalmologists for both patients at risk for AMD and those with established disease. The ophthalmologists were more likely than the optometrists to rely on the findings from the age-related eye disease studies of AMD regarding treatment with and selection of supplements and to recommend smoking cessation. Common evidence-based strategies for AMD management among eye care professionals would presumably be beneficial for AMD patients. Targeted education and implementation strategies may be needed.


Meta-analysis of epidemiologic dose-response data.
We discuss methods for summarizing epidemiologic studies of dose-response. The data from such a study typically appear as a series of dose-specific relative risks, with one category serving as the common reference group. We present methods for estimating the dose-response parameters from single and multiple study reports, for assigning levels to exposure categories when modeling relative risks, and for investigating the effects of study design and subject characteristics on dose-response curves. Finally, we discuss the choice of fixed vs random effects models. We illustrate our points with data from case-control studies of the relation between duration of oral contraceptive use and risk of breast cancer.


Education/community collaborations for undergraduate nursing gerontological clinical experiences.
In 2000, the American Association of Colleges of Nursing and the John A. Hartford Foundation Institute for Geriatric Nursing developed guidelines to help nurse educators incorporate gerontological nursing content into baccalaureate curricula. In 2001, the Hartford Foundation also provided grant monies to nursing programs to support gerontology curricular innovations and new clinical experiences. The funding allowed faculty to focus time, energy, and resources on gerontological nursing education. We, the authors, representing two funded schools of nursing, collaborated with community agencies to develop undergraduate gerontological clinical learning experiences and are encouraged by the results. This article describes the development of these collaborations and serves as a model for other schools of nursing. The education/community collaborations described here focus on clinical learning strategies, implementation activities, and outcomes/benefits of the experiences. Both educational programs had supportive administration, faculty willing to participate in curriculum change, organized plans to implement geriatric curricular enhancement, and long-standing community partnerships.


Computational insight into the mechanism and origins of high selectivities in the acylation of polyamines with 5-benzoyl-5-phenyl-1,5-dihydro-4H-pyrazol-4-one.
Amide bonds have gained much attention from numerous scientists and acyl transfer is a good way to form amide bonds. The acylation mechanism of polyamines and their high selectivity in dichloromethane were investigated by the use of the density functional theory (DFT), M06-2X/6-311+G (d, p)//M06-2X/6-31G (d, p) method combined with the solvation SMD model. The calculated results suggest that the reaction process involved two steps: an acylation step and a proton-transfer step, with the former being the rate-limiting step. Meanwhile, with the substituent group effects of amines and 5-acyl-5-phenyl-1,5-dihydro-4H-pyrazol-4-one (BCPP) on the acylation step, different substituent groups of amines have little influence on the kinetic properties of the acylation step, and the para-substituent groups of the phenyl group in BCPP lead to a linear relationship according to the electronegativity of the substituents. Furthermore, regarding the rate-selectivity of amines, the rate-selectivity of primary amines is higher than that of secondary amines, and polyamines very easily take part in acylation owing to the intramolecular hydrogen bond interaction. Moreover, it is harder for the amine group which has an α-position substituent group in polyamines to take part in an acylation reaction compared to the one without an α-position substituent group. The site-selectivity of the acylation process in polyamines is determined by steric hindrance. What's more, the auxiliary analysis of the distortion/interaction analysis and the frontier molecular orbital (FMO) analysis is used to investigate the origins of the rate- and site-selectivities.


Whole-genome sequencing of esophageal adenocarcinoma in Chinese patients reveals distinct mutational signatures and genomic alterations.
While the incidence of esophageal adenocarcinoma (EAC) has risen drastically in Western countries over the last 40 years, a similar trend has not been observed for EAC in China. Here, we analyzed mutational spectrum, copy number alterations, and structural variants from whole-genome sequencing of 10 Chinese EAC tumor samples and their matched normal samples, and compared them to previously reported EAC tumor specimens from Western countries. The mutational burden in Chinese EAC was significantly lower than that found in EAC from Western countries. The hallmark A>C mutational signature observed at high frequency in EAC from Western countries, which has been linked to acid reflux, is completely absent in Chinese samples. Furthermore, none of the Chinese samples showed evidence of chromothripsis and genome doubling that are often found in EAC from Western countries. In summary, Chinese EAC tumor samples had distinct genomic profiles and signatures, suggesting that EAC in Chinese individuals may arise from a different etiological pathway.


How malaria has affected the human genome and what human genetics can teach us about malaria.
Malaria is a major killer of children worldwide and the strongest known force for evolutionary selection in the recent history of the human genome. The past decade has seen growing evidence of ethnic differences in susceptibility to malaria and of the diverse genetic adaptations to malaria that have arisen in different populations: epidemiological confirmation of the hypotheses that G6PD deficiency, alpha+ thalassemia, and hemoglobin C protect against malaria mortality; the application of novel haplotype-based techniques demonstrating that malaria-protective genes have been subject to recent positive selection; the first genetic linkage maps of resistance to malaria in experimental murine models; and a growing number of reported associations with resistance and susceptibility to human malaria, particularly in genes involved in immunity, inflammation, and cell adhesion. The challenge for the next decade is to build the global epidemiological infrastructure required for statistically robust genomewide association analysis, as a way of discovering novel mechanisms of protective immunity that can be used in the development of an effective malaria vaccine.


Establishment of rostrocaudal polarity in tectal primordium: engrailed expression and subsequent tectal polarity.
In the E4 (embryonic day 4) chick tectal primordium, engrailed expression is strong at the caudal end and gradually weakens toward the rostral end. We used quail-chick chimeric tecta to investigate how the caudorostral gradient of engrailed expression is established and whether it is correlated with the subsequent rostrocaudal polarity of tectal development. To examine the positional value of the tectal primordium, we produced ectopic tecta in the diencephalon by transplanting a part of the mesencephalic alar plate heterotopically. In the ectopic tectum, the gradient of the engrailed expression reversed and the strength of the expression was dependent on the distance from the mes-diencephalon junction; the nearer the ectopic tectum was to the junction, the weaker the expression was. Consequently, the pattern of the engrailed expression in the host and ectopic tecta was nearly a mirror image, suggesting the existence of a repressive influence around the mes-diencephalon junction on the engrailed expression. We examined cytoarchitectonic development in the ectopic tecta, which normally proceeds in a gradient along the rostrocaudal axis; the rostral shows more advanced lamination than the caudal. In contrast, the caudal part of the ectopic tecta (near to the mes-diencephalon junction) showed more advanced lamination than the rostral. In both the host and ectopic tecta, advanced lamination was observed where the engrailed expression was repressed, and vice versa. Next we studied the correlation between engrailed expression and retinotectal projection from a view of plasticity and rigidity of rostrocaudal polarity in the tectum. We produced ectopic tecta by anisochronal transplantations between E3 host and E2 donor, and showed that there is little repressive influence at E3 around the mes-diencephalon junction. We then made chimeric double-rostral tectum (caudal half of it was replaced by rostral half of the donor tectum) or double-caudal tectum at E3. The transplants kept their original staining pattern in hosts. Consequently, the chimeric tecta showed wholly negative or positive staining of engrailed protein on the grafted side. In such tecta retinotectal projection pattern was disturbed as if the transplants retained their original position-specific characters. We propose from these heterotopic and anisochronal experiments that the engrailed expression can be a marker for subsequent rostrocaudal polarity in the tectum, both as regards cytoarchitectonic development and retinotectal projection.


Patient experiences of participation in a radical thoracic surgical trial: findings from the Mesothelioma and Radical Surgery Trial 2 (MARS 2).
The Mesothelioma and Radical Surgery Trial (MARS 2) aims to evaluate a surgical procedure by comparing chemotherapy and surgery against chemotherapy alone. The pilot study for MARS 2 evaluated the viability of recruitment. Challenges have been reported in conducting clinical research into thoracic surgical treatments and evidence is required to improve our understanding of patient experiences of trial procedures, trial treatments and the factors that influence participation. This longitudinal qualitative study was nested within the MARS 2 pilot. Semi-structured telephone interviews were conducted with 15 participants in the MARS 2 trial. Interviews were conducted post-randomisation, post-surgery (surgery arm) and at 6 and 12 months. Altogether, 41 interviews were carried out. The data were analysed using framework techniques. Challenges were identified regarding the volume and complexity of information given to participants, and their understanding of clinical equipoise and randomisation. Factors influencing participation included having an opportunity to undergo surgery, a self-assessment of their ability to cope with trial treatments, maintaining a positive approach and altruism. Obstacles included the logistics of traveling for treatment in an unfamiliar setting. Negative consequences of trial participation included increased uncertainty amplified by multiple care providers and unclear transition arrangements after the trial. Participants' descriptions provided insights that have implications for care for mesothelioma trial patients. The need for healthcare staff to be alert to the potential for misunderstanding, particularly when presenting treatment options, was identified. Patients perceived and derived benefits from taking part in the trial but experienced some negative consequences. These should be anticipated and managed proactively. ClinicalTrials.gov, NCT02040272 . Registered on 20 January 2014.


International Harmonization of Nomenclature and Diagnostic Criteria (INHAND): Non-proliferative and Proliferative Lesions of the Non-human Primate (M. fascicularis).
The INHAND (International Harmonization of Nomenclature and Diagnostic Criteria for Lesions Project (www.toxpath.org/inhand.asp) is a joint initiative of the Societies of Toxicologic Pathology from Europe (ESTP), Great Britain (BSTP), Japan (JSTP) and North America (STP) to develop an internationally accepted nomenclature for proliferative and nonproliferative lesions in laboratory animals. The purpose of this publication is to provide a standardized nomenclature for classifying microscopic lesions observed in most tissues and organs from the nonhuman primate used in nonclinical safety studies. Some of the lesions are illustrated by color photomicrographs. The standardized nomenclature presented in this document is also available electronically on the internet (http://www.goreni.org/). Sources of material included histopathology databases from government, academia, and industrial laboratories throughout the world. Content includes spontaneous lesions as well as lesions induced by exposure to test materials. Relevant infectious and parasitic lesions are included as well. A widely accepted and utilized international harmonization of nomenclature for lesions in laboratory animals will provide a common language among regulatory and scientific research organizations in different countries and increase and enrich international exchanges of information among toxicologists and pathologists.


Spectrotemporal contrast kernels for neurons in primary auditory cortex.
Auditory neurons are often described in terms of their spectrotemporal receptive fields (STRFs). These map the relationship between features of the sound spectrogram and firing rates of neurons. Recently, we showed that neurons in the primary fields of the ferret auditory cortex are also subject to gain control: when sounds undergo smaller fluctuations in their level over time, the neurons become more sensitive to small-level changes (Rabinowitz et al., 2011). Just as STRFs measure the spectrotemporal features of a sound that lead to changes in the firing rates of neurons, in this study, we sought to estimate the spectrotemporal regions in which sound statistics lead to changes in the gain of neurons. We designed a set of stimuli with complex contrast profiles to characterize these regions. This allowed us to estimate the STRFs of cortical neurons alongside a set of spectrotemporal contrast kernels. We find that these two sets of integration windows match up: the extent to which a stimulus feature causes the firing rate of a neuron to change is strongly correlated with the extent to which the contrast of that feature modulates the gain of the neuron. Adding contrast kernels to STRF models also yields considerable improvements in the ability to capture and predict how auditory cortical neurons respond to statistically complex sounds.


In Silico Fragment-Based Design Identifies Subfamily B1 Metallo-β-lactamase Inhibitors.
Zinc ion-dependent β-lactamases (MBLs) catalyze the hydrolysis of almost all β-lactam antibiotics and resist the action of clinically available β-lactamase inhibitors. We report how application of in silico fragment-based molecular design employing thiol-mediated metal anchorage leads to potent MBL inhibitors. The new inhibitors manifest potent inhibition of clinically important B1 subfamily MBLs, including the widespread NDM-1, IMP-1, and VIM-2 enzymes; with lower potency, some of them also inhibit clinically relevant Class A and D serine-β-lactamases. The inhibitors show selectivity for bacterial MBL enzymes compared to that for human MBL fold nucleases. Cocrystallization of one inhibitor, which shows potentiation of Meropenem activity against MBL-expressing Enterobacteriaceae, with VIM-2 reveals an unexpected binding mode, involving interactions with residues from conserved active site bordering loops.


Supramolecular Modification of a Sequence-Controlled Collagen-Mimicking Polymer.
Structurally and functionally well-defined recombinant proteins are an interesting class of sequence-controlled macromolecules to which different crosslinking chemistries can be applied to tune their biological properties. Herein, we take advantage of a 571-residue recombinant peptide based on human collagen type I (RCPhC1), which we functionalized with supramolecular 4-fold hydrogen bonding ureido-pyrimidinone (UPy) moieties. By grafting supramolecular UPy moieties onto the backbone of RCPhC1 (UPy-RCPhC1), increased control over the polymer structure, assembly, gelation, and mechanical properties was achieved. In addition, by increasing the degree of UPy functionalization on RCPhC1, cardiomyocyte progenitor cells were cultured on "soft" (∼26 kPa) versus "stiff" (∼68-190 kPa) UPy-RCPhC1 hydrogels. Interestingly, increased stress fiber formation, focal adhesions, and proliferation were observed on stiffer compared to softer substrates, owing to the formation of stronger cell-material interactions. In conclusion, a bioinspired hydrogel material was designed by a combination of two well-known natural components, i.e., a protein as sequence-controlled polymer and UPy units inspired on nucleobases.


Serotonin (5HT), fluoxetine, imipramine and dopamine target distinct 5HT receptor signaling to modulate Caenorhabditis elegans egg-laying behavior.
Drugs that target the serotonergic system are the most commonly prescribed therapeutic agents and are used for treatment of a wide range of behavioral and neurological disorders. However, the mechanism of the drug action remain a conjecture. Here, we dissect the genetic targets of serotonin (5HT), the selective 5HT reuptake inhibitor (SSRI) fluoxetine (Prozac), the tricyclic antidepressant imipramine, and dopamine. Using the well-established serotonergic response in C. elegans egg-laying behavior as a paradigm, we show that action of fluoxetine and imipramine at the 5HT reuptake transporter (SERT) and at 5HT receptors are separable mechanisms. Even mutants completely lacking 5HT or SERT can partially respond to fluoxetine and imipramine. Furthermore, distinct mechanisms for each drug can be recognized to mediate these responses. Deletion of SER-1, a 5HT1 receptor, abolishes the response to 5HT but has only a minor effect on the response to imipramine and no effect on the response to fluoxetine. In contrast, deletion of SER-4, a 5HT2 receptor, confers significant resistance to imipramine while leaving the responses to 5HT or fluoxetine intact. Further, fluoxetine can stimulate egg laying via the Gq protein EGL-30, independent of SER-1, SER-4, or 5HT. We also show that dopamine antagonizes the 5HT action via the 5HT-gated ion channel MOD-1 signaling, suggesting that this channel activity couples 5HT and dopamine signaling. These results suggest that the actions of these drugs at specific receptor subtypes could determine their therapeutic efficacy. SSRIs and tricyclic antidepressants may regulate 5HT outputs independently of synaptic levels of 5HT.


Costs and benefits of group living with disease: a case study of pneumonia in bighorn lambs (Ovis canadensis).
Group living facilitates pathogen transmission among social hosts, yet temporally stable host social organizations can actually limit transmission of some pathogens. When there are few between-subpopulation contacts for the duration of a disease event, transmission becomes localized to subpopulations. The number of per capita infectious contacts approaches the subpopulation size as pathogen infectiousness increases. Here, we illustrate that this is the case during epidemics of highly infectious pneumonia in bighorn lambs (Ovis canadensis). We classified individually marked bighorn ewes into disjoint seasonal subpopulations, and decomposed the variance in lamb survival to weaning into components associated with individual ewes, subpopulations, populations and years. During epidemics, lamb survival varied substantially more between ewe-subpopulations than across populations or years, suggesting localized pathogen transmission. This pattern of lamb survival was not observed during years when disease was absent. Additionally, group sizes in ewe-subpopulations were independent of population size, but the number of ewe-subpopulations increased with population size. Consequently, although one might reasonably assume that force of infection for this highly communicable disease scales with population size, in fact, host social behaviour modulates transmission such that disease is frequency-dependent within populations, and some groups remain protected during epidemic events.


Characterising hyperinsulinemia-induced insulin resistance in human skeletal muscle cells.
Hyperinsulinaemia potentially contributes to insulin resistance in metabolic tissues, such as skeletal muscle. The purpose of these experiments was to characterise glucose uptake, insulin signalling and relevant gene expression in primary human skeletal muscle-derived cells (HMDCs), in response to prolonged insulin exposure (PIE) as a model of hyperinsulinaemia-induced insulin resistance. Differentiated HMDCs from healthy human donors were cultured with or without insulin (100 nM) for 3 days followed by an acute insulin stimulation. HMDCs exposed to PIE were characterised by impaired insulin-stimulated glucose uptake, blunted IRS-1 phosphorylation (Tyr612) and Akt (Ser473) phosphorylation in response to an acute insulin stimulation. Glucose transporter 1 (GLUT1), but not GLUT4, mRNA and protein increased following PIE. The mRNA expression of metabolic (PDK4) and inflammatory markers (TNF-α) was reduced by PIE but did not change lipid (SREBP1 and CD36) or mitochondrial (UCP3) markers. These experiments provide further characterisation of the effects of PIE as a model of hyperinsulinaemia-induced insulin resistance in HMDCs.


Association between metabolic syndrome and bone fractures: a meta-analysis of observational studies.
Emerging epidemiological evidence suggest an association between metabolic syndrome and fractures. However, whether metabolic syndrome is an independent risk or protective factor of fractures remains controversial. Our goal is to provide a quantitative assessment of the association between metabolic syndrome and bone fractures by conducting a meta-analysis of observational studies. The PubMed and Embase database were searched through to March 2013 to identify studies that met pre-established inclusion criteria. Reference lists of retrieved articles were also reviewed. Summary effect estimates with 95% confidence intervals (CI) were derived using a fixed or random effects model, depending on the heterogeneity of the included studies. Eight epidemiologic studies involving 39,938 participants were included in the meta-analysis. In overall analysis, metabolic syndrome was not associated with prevalent fractures [pooled odds ratio (OR) 0.93, 95% CI 0.84 - 1.03] in cross-sectional studies or incident fractures [pooled relative risk (RR) 0.88, 95% CI 0.37 - 2.12] in prospective cohort studies. No evidence of heterogeneity was found in cross-sectional studies (p = 0.786, I2 = 0.0%). A substantial heterogeneity was detected in cohort studies (p = 0.001, I2 = 85.7%). No indication of significant publication bias was found either from Begg's test or Egger's test. Estimates of total effects were substantially consistent in the sensitivity and stratification analyses. The present meta-analysis of observational studies suggests that the metabolic syndrome has no explicit effect on bone fractures.


Evaluation of a very brief pedometer-based physical activity intervention delivered in NHS Health Checks in England: The VBI randomised controlled trial.
The majority of people do not achieve recommended levels of physical activity. There is a need for effective, scalable interventions to promote activity. Self-monitoring by pedometer is a potentially suitable strategy. We assessed the effectiveness and cost-effectiveness of a very brief (5-minute) pedometer-based intervention ('Step It Up') delivered as part of National Health Service (NHS) Health Checks in primary care. The Very Brief Intervention (VBI) Trial was a two parallel-group, randomised controlled trial (RCT) with 3-month follow-up, conducted in 23 primary care practices in the East of England. Participants were 1,007 healthy adults aged 40 to 74 years eligible for an NHS Health Check. They were randomly allocated (1:1) using a web-based tool between October 1, 2014, and December 31, 2015, to either intervention (505) or control group (502), stratified by primary care practice. Participants were aware of study group allocation. Control participants received the NHS Health Check only. Intervention participants additionally received Step It Up: a 5-minute face-to-face discussion, written materials, pedometer, and step chart. The primary outcome was accelerometer-based physical activity volume at 3-month follow-up adjusted for sex, 5-year age group, and general practice. Secondary outcomes included time spent in different intensities of physical activity, self-reported physical activity, and economic measures. We conducted an in-depth fidelity assessment on a subsample of Health Check consultations. Participants' mean age was 56 years, two-thirds were female, they were predominantly white, and two-thirds were in paid employment. The primary outcome was available in 859 (85.3%) participants. There was no significant between-group difference in activity volume at 3 months (adjusted intervention effect 8.8 counts per minute [cpm]; 95% CI -18.7 to 36.3; p = 0.53). We found no significant between-group differences in the secondary outcomes of step counts per day, time spent in moderate or vigorous activity, time spent in vigorous activity, and time spent in moderate-intensity activity (accelerometer-derived variables); as well as in total physical activity, home-based activity, work-based activity, leisure-based activity, commuting physical activity, and screen or TV time (self-reported physical activity variables). Of the 505 intervention participants, 491 (97%) received the Step it Up intervention. Analysis of 37 intervention consultations showed that 60% of Step it Up components were delivered faithfully. The intervention cost £18.04 per participant. Incremental cost to the NHS per 1,000-step increase per day was £96 and to society was £239. Adverse events were reported by 5 intervention participants (of which 2 were serious) and 5 control participants (of which 2 were serious). The study's limitations include a participation rate of 16% and low return of audiotapes by practices for fidelity assessment. In this large well-conducted trial, we found no evidence of effect of a plausible very brief pedometer intervention embedded in NHS Health Checks on objectively measured activity at 3-month follow-up. Current Controlled Trials (ISRCTN72691150).


The requirement for the highly conserved G-1 residue of Saccharomyces cerevisiae tRNAHis can be circumvented by overexpression of tRNAHis and its synthetase.
Nearly all tRNA(His) species have an additional 5' guanine nucleotide (G(-1)). G(-1) is encoded opposite C(73) in nearly all prokaryotes and in some archaea, and is added post-transcriptionally by tRNA(His) guanylyltransferase (Thg1) opposite A(73) in eukaryotes, and opposite C(73) in other archaea. These divergent mechanisms of G(-1) conservation suggest that G(-1) might have an important cellular role, distinct from its role in tRNA(His) charging. Thg1 is also highly conserved and is essential in the yeast Saccharomyces cerevisiae. However, the essential roles of Thg1 are unclear since Thg1 also interacts with Orc2 of the origin recognition complex, is implicated in the cell cycle, and catalyzes an unusual template-dependent 3'-5' (reverse) polymerization in vitro at the 5' end of activated tRNAs. Here we show that thg1-Delta strains are viable, but only if histidyl-tRNA synthetase and tRNA(His) are overproduced, demonstrating that the only essential role of Thg1 is its G(-1) addition activity. Since these thg1-Delta strains have severe growth defects if cytoplasmic tRNA(His) A(73) is overexpressed, and distinct, but milder growth defects, if tRNA(His) C(73) is overexpressed, these results show that the tRNA(His) G(-1) residue is important, but not absolutely essential, despite its widespread conservation. We also show that Thg1 catalyzes 3'-5' polymerization in vivo on tRNA(His) C(73), but not on tRNA(His) A(73), demonstrating that the 3'-5' polymerase activity is pronounced enough to have a biological role, and suggesting that eukaryotes may have evolved to have cytoplasmic tRNA(His) with A(73), rather than C(73), to prevent the possibility of 3'-5' polymerization.


Fiber-based polarization-sensitive Fourier domain optical coherence tomography using B-scan-oriented polarization modulation method.
Fiber-based high-speed polarization-sensitive Fourier domain optical coherence tomography (PS-FD-OCT) is developed at 840 nm wavelength using polarization modulation method. The incident state of polarization is modulated along B-scan. The spectrometer has a polarizing beamsplitter and two line-CCD cameras operated at a line rate of 27.7 kHz. From the 0th and 1st orders of the spatial frequencies along the B-scanning, a depth-resolved Jones matrix can be derived. Since continuous polarization modulation along B-scan causes fringe washout, equivalent discrete polarization modulation is applied to biological measurements. For the demonstration, an in vitro chicken breast muscle, an in vivo finger pad, and an in vivo caries lesion of a human tooth are measured. Three dimensional phase retardation images show the potentials for applying the system to biological and medical studies.


Efficacy of Liposomal Bupivacaine and Bupivacaine Hydrochloride vs Bupivacaine Hydrochloride Alone as a Periarticular Anesthetic for Patients Undergoing Knee Replacement: A Randomized Clinical Trial.
More than half of patients who undergo knee replacement surgery report substantial acute postoperative pain. To evaluate the efficacy and cost-effectiveness of periarticular liposomal bupivacaine for recovery and pain management after knee replacement. This multicenter, patient-blinded, pragmatic, randomized clinical superiority trial involved 533 participants at 11 institutions within the National Health Service in England. Adults undergoing primary unilateral knee replacement for symptomatic end-stage osteoarthritis were enrolled between March 29, 2018, and February 29, 2020, and followed up for 1 year after surgery. Follow-up was completed March 1, 2021. A per-protocol analysis for each coprimary outcome was performed in addition to the main intention-to-treat analysis. Two hundred sixty-six milligrams of liposomal bupivacaine admixed with 100 mg of bupivacaine hydrochloride compared with 100 mg of bupivacaine hydrochloride alone (control) administered by periarticular injection at the time of surgery. The coprimary outcomes were Quality of Recovery 40 (QoR-40) score at 72 hours and pain visual analog scale (VAS) score area under the curve (AUC) from 6 to 72 hours. Secondary outcomes included QoR-40 and mean pain VAS at days 0 (evening of surgery), 1, 2, and 3; cumulative opioid consumption for 72 hours; functional outcomes and quality of life at 6 weeks, 6 months, and 1 year; and cost-effectiveness for 1 year. Adverse events and serious adverse events up to 12 months after randomization were also assessed. Among the 533 participants included in the analysis, the mean (SD) age was 69.0 (9.7) years; 287 patients were women (53.8%) and 246 were men (46.2%). Baseline characteristics were balanced between study groups. There was no difference between the liposomal bupivacaine and control groups in QoR-40 score at 72 hours (adjusted mean difference, 0.54 [97.5% CI, -2.05 to 3.13]; P = .64) or the pain VAS score AUC at 6 to 72 hours (-21.5 [97.5% CI, -46.8 to 3.8]; P = .06). Analyses of pain VAS and QoR-40 scores demonstrated only 1 statistically significant difference, with the liposomal bupivacaine arm having lower pain scores the evening of surgery (adjusted difference -0.54 [97.5% CI, -1.07 to -0.02]; P = .02). No difference in cumulative opioid consumption and functional outcomes was detected. Liposomal bupivacaine was not cost-effective compared with the control treatment. No difference in adverse or serious adverse events was found between the liposomal bupivacaine and control groups. This study found no difference in postoperative recovery or pain associated with the use of periarticular liposomal bupivacaine compared with bupivacaine hydrochloride alone in patients who underwent knee replacement surgery. isrctn.com Identifier: ISRCTN54191675.


Experiences with global trigger tool reviews in five Danish hospitals: an implementation study.
To describe experiences with the implementation of global trigger tool (GTT) reviews in five Danish hospitals and to suggest ways to improve the performance of GTT review teams. Retrospective observational study. The measurement and monitoring of harms are crucial to campaigns to improve the safety of patients. Increasingly, teams use the GTT to review patient records and measure harms in English and non-English-speaking countries. Meanwhile, it is not clear as to how the method performs in such diverse settings. Review teams from five Danish pilot hospitals of the national Danish Safer Hospital Programme. We collected harm rates, background and anecdotal information and reported patient safety incidents (PSIs) from five pilot hospitals currently participating in the Danish Safer Hospital Programme. Experienced reviewers categorised harms by type. We plotted harm rates as run-charts and applied rules for the detection of patterns of non-random variation. The hospitals differed in size but had similar patient populations and activity. PSIs varied between 3 and 12 per 1000 patient-days. The average harm rate for all hospitals was 60 per 1000 patient-days ranging from 34 to 84. The percentage of harmed patients was 25 and ranged from 18 to 33. Overall, 96% of harms were temporary. Infections, pressure ulcers procedure-related and gastrointestinal problems were common. Teams reported differences in training and review procedures such as the role of the secondary reviewer. We found substantial variation in harm rates. Differences in training, review procedures and documentation in patient records probably contributed to these variations. Training reviewers as teams, specifying the roles of the different reviewers, training records and a database for findings of reviews may improve the application of the GTT.


Perceiving your hand moving: BOLD suppression in sensory cortices and the role of the cerebellum in the detection of feedback delays.
Sensory consequences of self-generated as opposed to externally generated movements are perceived as less intense and lead to less neural activity in corresponding sensory cortices, presumably due to predictive mechanisms. Self-generated sensory inputs have been mostly studied in a single modality, using abstract feedback, with control conditions not differentiating efferent from reafferent feedback. Here we investigated the neural processing of (a) naturalistic action-feedback associations of (b) self-generated versus externally generated movements, and (c) how an additional (auditory) modality influences neural processing and detection of delays. Participants executed wrist movements using a passive movement device (PMD) as they watched their movements in real time or with variable delays (0-417 ms). The task was to judge whether there was a delay between the movement and its visual feedback. In the externally generated condition, movements were induced by the PMD to disentangle efferent from reafferent feedback. Half of the trials involved auditory beeps coupled to the onset of the visual feedback. We found reduced BOLD activity in visual, auditory, and somatosensory areas during self-generated compared with externally generated movements in unimodal and bimodal conditions. Anterior and posterior cerebellar areas were engaged for trials in which action-feedback delays were detected for self-generated movements. Specifically, the left cerebellar lobule IX was functionally connected with the right superior occipital gyrus. The results indicate efference copy-based predictive mechanisms specific to self-generated movements, leading to BOLD suppression in sensory areas. In addition, our results support the cerebellum's role in the detection of temporal prediction errors during our actions and their consequences.


Effectiveness and Safety of Dabigatran, Rivaroxaban, and Apixaban Versus Warfarin in Nonvalvular Atrial Fibrillation.
The introduction of non-vitamin K antagonist oral anticoagulants has been a major advance for stroke prevention in atrial fibrillation; however, outcomes achieved in clinical trials may not translate to routine practice. We aimed to evaluate the effectiveness and safety of dabigatran, rivaroxaban, and apixaban by comparing each agent with warfarin. Using a large US insurance database, we identified privately insured and Medicare Advantage patients with nonvalvular atrial fibrillation who were users of apixaban, dabigatran, rivaroxaban, or warfarin between October 1, 2010, and June 30, 2015. We created 3 matched cohorts using 1:1 propensity score matching: apixaban versus warfarin (n=15 390), dabigatran versus warfarin (n=28 614), and rivaroxaban versus warfarin (n=32 350). Using Cox proportional hazards regression, we found that for stroke or systemic embolism, apixaban was associated with lower risk (hazard ratio [HR] 0.67, 95% CI 0.46-0.98, P=0.04), but dabigatran and rivaroxaban were associated with a similar risk (dabigatran: HR 0.98, 95% CI 0.76-1.26, P=0.98; rivaroxaban: HR 0.93, 95% CI 0.72-1.19, P=0.56). For major bleeding, apixaban and dabigatran were associated with lower risk (apixaban: HR 0.45, 95% CI 0.34-0.59, P<0.001; dabigatran: HR 0.79, 95% CI 0.67-0.94, P<0.01), and rivaroxaban was associated with a similar risk (HR 1.04, 95% CI 0.90-1.20], P=0.60). All non-vitamin K antagonist oral anticoagulants were associated with a lower risk of intracranial bleeding. In patients with nonvalvular atrial fibrillation, apixaban was associated with lower risks of both stroke and major bleeding, dabigatran was associated with similar risk of stroke but lower risk of major bleeding, and rivaroxaban was associated with similar risks of both stroke and major bleeding in comparison to warfarin.


On distinguishing between canonical tRNA genes and tRNA gene fragments in prokaryotes.
Automated genome annotation is essential for extracting biological information from sequence data. The identification and annotation of tRNA genes is frequently performed by the software package tRNAscan-SE, the output of which is listed for selected genomes in the Genomic tRNA database (GtRNAdb). Here, we highlight a pervasive error in prokaryotic tRNA gene sets on GtRNAdb: the mis-categorization of partial, non-canonical tRNA genes as standard, canonical tRNA genes. Firstly, we demonstrate the issue using the tRNA gene sets of 20 organisms from the archaeal taxon Thermococcaceae. According to GtRNAdb, these organisms collectively deviate from the expected set of tRNA genes in 15 instances, including the listing of eleven putative canonical tRNA genes. However, after detailed manual annotation, only one of these eleven remains; the others are either partial, non-canonical tRNA genes resulting from the integration of genetic elements or CRISPR-Cas activity (seven instances), or attributable to ambiguities in input sequences (three instances). Secondly, we show that similar examples of the mis-categorization of predicted tRNA sequences occur throughout the prokaryotic sections of GtRNAdb. While both canonical and non-canonical prokaryotic tRNA gene sequences identified by tRNAscan-SE are biologically interesting, the challenge of reliably distinguishing between them remains. We recommend employing a combination of (i) screening input sequences for the genetic elements typically associated with non-canonical tRNA genes, and ambiguities, (ii) activating the tRNAscan-SE automated pseudogene detection function, and (iii) scrutinizing predicted tRNA genes with low isotype scores. These measures greatly reduce manual annotation efforts, and lead to improved prokaryotic tRNA gene set predictions.


Biochemical characterization of highly purified leucine-rich repeat kinases 1 and 2 demonstrates formation of homodimers.
Leucine-rich repeat kinase 1 and 2 (LRRK1 and LRRK2) are large multidomain proteins containing kinase, GTPase and multiple protein-protein interaction domains, but only mutations in LRRK2 are linked to familial Parkinson's disease (PD). Independent studies suggest that LRRK2 exists in the cell as a complex compatible with the size of a dimer. However, whether this complex is truly a homodimer or a heterologous complex formed by monomeric LRRK2 with other proteins has not been definitively proven due to the limitations in obtaining highly pure proteins suitable for structural characterization. Here, we used stable expression of LRRK1 and LRRK2 in HEK293T cell lines to produce recombinant LRRK1 and LRRK2 proteins of greater than 90% purity. Both purified LRRKs are folded, with a predominantly alpha-helical secondary structure and are capable of binding GTP with similar affinity. Furthermore, recombinant LRRK2 exhibits robust autophosphorylation activity, phosphorylation of model peptides in vitro and ATP binding. In contrast, LRRK1 does not display significant autophosphorylation activity and fails to phosphorylate LRRK2 model substrates, although it does bind ATP. Using these biochemically validated proteins, we show that LRRK1 and LRRK2 are capable of forming homodimers as shown by single-particle transmission electron microscopy and immunogold labeling. These LRRK dimers display an elongated conformation with a mean particle size of 145 Å and 175 Å respectively, which is disrupted by addition of 6M guanidinium chloride. Immunogold staining revealed double-labeled particles also in the pathological LRRK2 mutant G2019S and artificial mutants disrupting GTPase and kinase activities, suggesting that point mutations do not hinder the dimeric conformation. Overall, our findings indicate for the first time that purified and active LRRK1 and LRRK2 can form dimers in their full-length conformation.