Spread from the Sink to the Patient: In Situ Study Using Green Fluorescent Protein (GFP)-Expressing Escherichia coli To Model Bacterial Dispersion from Hand-Washing Sink-Trap Reservoirs.
There have been an increasing number of reports implicating Gammaproteobacteria as often carrying genes of drug resistance from colonized sink traps to vulnerable hospitalized patients. However, the mechanism of transmission from the wastewater of the sink P-trap to patients remains poorly understood. Herein we report the use of a designated hand-washing sink lab gallery to model dispersion of green fluorescent protein (GFP)-expressing Escherichia coli from sink wastewater to the surrounding environment. We found no dispersion of GFP-expressing E. coli directly from the P-trap to the sink basin or surrounding countertop with coincident water flow from a faucet. However, when the GFP-expressing E. coli cells were allowed to mature in the P-trap under conditions similar to those in a hospital environment, a GFP-expressing E. coli-containing putative biofilm extended upward over 7 days to reach the strainer. This subsequently resulted in droplet dispersion to the surrounding areas (<30 in.) during faucet operation. We also demonstrated that P-trap colonization could occur by retrograde transmission along a common pipe. We postulate that the organisms mobilize up to the strainer from the P-trap, resulting in droplet dispersion rather than dispersion directly from the P-trap. This work helps to further define the mode of transmission of bacteria from a P-trap reservoir to a vulnerable hospitalized patient.IMPORTANCE Many recent reports demonstrate that sink drain pipes become colonized with highly consequential multidrug-resistant bacteria, which then results in hospital-acquired infections. However, the mechanism of dispersal of bacteria from the sink to patients has not been fully elucidated. Through establishment of a unique sink gallery, this work found that a staged mode of transmission involving biofilm growth from the lower pipe to the sink strainer and subsequent splatter to the bowl and surrounding area occurs rather than splatter directly from the water in the lower pipe. We have also demonstrated that bacterial transmission can occur via connections in wastewater plumbing to neighboring sinks. This work helps to more clearly define the mechanism and risk of transmission from a wastewater source to hospitalized patients in a world with increasingly antibiotic-resistant bacteria that can thrive in wastewater environments and cause infections in vulnerable patients.


Recent progress in the science of complex coacervation.
Complex coacervation is an associative, liquid-liquid phase separation that can occur in solutions of oppositely-charged macromolecular species, such as proteins, polymers, and colloids. This process results in a coacervate phase, which is a dense mix of the oppositely-charged components, and a supernatant phase, which is primarily devoid of these same species. First observed almost a century ago, coacervates have since found relevance in a wide range of applications; they are used in personal care and food products, cutting edge biotechnology, and as a motif for materials design and self-assembly. There has recently been a renaissance in our understanding of this important class of material phenomena, bringing the science of coacervation to the forefront of polymer and colloid science, biophysics, and industrial materials design. In this review, we describe the emergence of a number of these new research directions, specifically in the context of polymer-polymer complex coacervates, which are inspired by a number of key physical and chemical insights and driven by a diverse range of experimental, theoretical, and computational approaches.


Switchable telescopic contact lens.
We present design and first demonstration of optics for a telescopic contact lens with independent optical paths for switching between normal and magnified vision. The magnified optical path incorporates a telescopic arrangement of positive and negative annular concentric reflectors to achieve 2.8 x magnification on the eye, while light passing through a central clear aperture provides unmagnified vision. We present an experimental demonstration of the contact lens mounted on a life-sized optomechanical model eye and, using a pair of modified commercial 3D television glasses, demonstrate electrically operated polarization switching between normal and magnified vision.


Translocation t(2;7)(p12;q21-22) with dysregulation of the CDK6 gene mapping to 7q21-22 in a non-Hodgkin's lymphoma with leukemia.
A female patient presented with splenomegaly and lymphocytosis with atypical lymphoid cell morphology. We identified t(2;7)(p12;q21) prompting studies of the translocation breakpoint and its consequences on protein expression to confirm or otherwise the recently reported involvement of CDK6 and IG k genes in the t(2;7) leading to over-expression of CDK6 protein. A variety of clinical and laboratory techniques including cell marker, cytogenetic and histologic studies were applied in order to establish the diagnosis. Fluorescence in situ hybridization (FISH) and Southern blotting were used for mapping the translocation breakpoint and Western blotting for assessing protein expression. Immunophenotyping showed the presence of a B-cell population with strong expression of FMC7, CD22, CD79b, CD5 and k restricted surface immunoglobulins. Based on morphology and immunophenotypic markers the diagnosis of B-cell non-Hodgkin's lymphoma was made. Karyotyping revealed a clone with t(2;7)(p12;q21-22). Evidence for clonal evolution with additional abnormalities including a deletion of the TP53 was present. We established by FISH and Southern blotting that the breakpoint on 7q21-22 fell in a region 66kb telomeric to the previously reported breakpoint for the t(2;7) and was the same as that observed in a t(7;21). CDK6 protein was over-expressed. The patient received alkylating agents and splenectomy and is alive but the lymphocytosis persists with evidence of disease progression. We have demonstrated that CDK6 expression is dysregulated even when the breakpoint on 7q21-22 is located 66kb upstream from the coding region. Interestingly, the precise assignment of the lymphoma type in our case was not possible even when the splenic histology was analyzed.


Being prepared: essential to self-care and quality of life for the person with cancer.
Being adequately prepared for an experience such as cancer empowers patients, lowers distress, improves coping, supports self-management, promotes recovery, and improves quality of life. However, patients with cancer report unmet informational and support needs across the cancer trajectory. The purpose of this article is to describe the relationship of information preparation and patient outcomes, identify information and support needs across the cancer trajectory, and describe the role of oncology nurses in the delivery of high-quality patient-centered cancer care. The middle range theory of "Carrying On" was used to identify information and support needs during different phases of the cancer trajectory from treatment to survivorship. The authors concluded that nurses should engage the patient in a relational exchange of information; provide concrete, understandable information across specific times in the cancer experience; and use creative approaches to minimize barriers in meeting patient needs to achieve high-quality patient-centered cancer care.


Structural changes and thermal stability of charged LiNixMnyCozO₂ cathode materials studied by combined in situ time-resolved XRD and mass spectroscopy.
Thermal stability of charged LiNixMnyCozO2 (NMC, with x + y + z = 1, x:y:z = 4:3:3 (NMC433), 5:3:2 (NMC532), 6:2:2 (NMC622), and 8:1:1 (NMC811)) cathode materials is systematically studied using combined in situ time-resolved X-ray diffraction and mass spectroscopy (TR-XRD/MS) techniques upon heating up to 600 °C. The TR-XRD/MS results indicate that the content of Ni, Co, and Mn significantly affects both the structural changes and the oxygen release features during heating: the more Ni and less Co and Mn, the lower the onset temperature of the phase transition (i.e., thermal decomposition) and the larger amount of oxygen release. Interestingly, the NMC532 seems to be the optimized composition to maintain a reasonably good thermal stability, comparable to the low-nickel-content materials (e.g., NMC333 and NMC433), while having a high capacity close to the high-nickel-content materials (e.g., NMC811 and NMC622). The origin of the thermal decomposition of NMC cathode materials was elucidated by the changes in the oxidation states of each transition metal (TM) cations (i.e., Ni, Co, and Mn) and their site preferences during thermal decomposition. It is revealed that Mn ions mainly occupy the 3a octahedral sites of a layered structure (R3̅m) but Co ions prefer to migrate to the 8a tetrahedral sites of a spinel structure (Fd3̅m) during the thermal decomposition. Such element-dependent cation migration plays a very important role in the thermal stability of NMC cathode materials. The reasonably good thermal stability and high capacity characteristics of the NMC532 composition is originated from the well-balanced ratio of nickel content to manganese and cobalt contents. This systematic study provides insight into the rational design of NMC-based cathode materials with a desired balance between thermal stability and high energy density.


The level and distribution of the GABA(B)R1 and GABA(B)R2 receptor subunits in the rat's inferior colliculus.
The type B γ-aminobutyric acid receptor (GABA(B) receptor) is an important neurotransmitter receptor in the midbrain auditory structure, the inferior colliculus (IC). A functional GABA(B) receptor is a heterodimer consisting of two subunits, GABA(B)R1 and GABA(B)R2. Western blotting and immunohistochemical experiments were conducted to examine the expression of the two subunits over the IC including its central nucleus, dorsal cortex, and external cortex (ICc, ICd, and ICx). Results revealed that the two subunits existed in both cell bodies and the neuropil throughout the IC. The two subunits had similar regional distributions over the IC. The combined level of cell body and neuropil labeling was higher in the ICd than the other two subdivisions. Labeling in the ICc and ICx was stronger in the dorsal than the ventral regions. In spite of regional differences, no defined boundaries were formed between different areas. For both subunits, the regional distribution of immunoreactivity in the neuropil was parallel to that of combined immunoreactivity in the neuropil and cell bodies. The density of labeled cell bodies tended to be higher but sizes of cell bodies tended to be smaller in the ICd than in the other subdivisions. No systematic regional changes were found in the level of cell body immunoreactivity, except that GABA(B)R2-immunoreactive cell bodies in the ICd had slightly higher optic density (OD) than in other regions. Elongated cell bodies existed throughout the IC. Many labeled cell bodies along the outline of the IC were oriented in parallel to the outline. No strong tendency of orientation was found in labeled cell bodies in ICc. Regional distributions of the subunits in ICc correlated well with inputs to this subdivision. Our finding regarding the contrast in the level of neuropil immunoreactivity among different subdivisions is consistent with the fact that the GABA(B) receptor has different pre- and postsynaptic functions in different IC regions.


Toward a Predict and Prevent Approach in Obstructive Airway Diseases.
Asthma and chronic obstructive pulmonary disease are currently diagnosed and treated after the demonstration of variable airflow limitation and symptoms. Under this framework, undiagnosed and unchecked airway inflammation is associated with recurrent acute attacks, airway remodeling, airflow limitation, adverse effects of corticosteroids, and impaired quality of life, ultimately leading to the collection of side effects termed "people remodeling." This one-size-fits-all damage control approach aims to control symptoms and treat exacerbations rather than modify the underlying disease process. The advent of highly effective therapies targeting proximal drivers of airway inflammation calls for a paradigm shift; upstream-acting therapies offer potential to alter the disease course and achieve clinical remission. We propose moving away from downstream firefighting and toward a "predict and prevent" model, measuring inflammation and providing anti-inflammatory therapy early, without waiting for further clinical deterioration. Much in the same way that high blood pressure and cholesterol are used to predict and prevent heart attacks, in asthma, elevated blood eosinophils and/or exhaled nitric oxide can be used to predict and prevent asthma attacks. We also advocate moving research further upstream by identifying patients with subclinical airway inflammation or disease who may be at risk of progressing to airflow limitation and associated morbidities and intervening early to prevent them. In summary, we call for a predict and prevent approach in obstructive airway disease.


Barriers to Hepatitis C Virus Care and How Federally Qualified Health Centers Can Improve Patient Access to Treatment.
Despite the availability of direct-acting antiviral agents (DAAs) for hepatitis C virus (HCV) treatment, disparities in HCV care and treatment persist for underserved populations due to demographic-based and insurance-based barriers. We aim to examine the effect of barriers on HCV treatment access for a federally qualified health center (FQHC) population. We retrospectively evaluated medical records of adults diagnosed with chronic HCV at an FQHC clinic from 2016 to 2020 with follow-up through 2021. Univariate and bivariate analyses were used to describe the patient population and significant associations between predictors of linkage to HCV care and treatment access. Adjusted multivariate logistic regression analyses were used to identify predictors of starting HCV treatment. Of 279 total patients with chronic HCV, 162 patients started treatment (58%), 138 patients (50%) completed treatment, and 99 patients (35%) achieved sustained virological response (SVR). Of the total patients, 145 (52%) were seen by their primary care physician (PCP) for their HCV care and treatment, and 134 (48%) were seen by a provider that specializes in management and treatment of HCV (HCV provider). Patients seen by an HCV provider in addition to their PCP were more likely to have had their prior authorization requests for HCV treatment denied by their insurance providers than patients seen only by their PCP for HCV care (30% vs. 14%, P = 0.001). We believe that this discrepancy stems from two issues. One, prior authorizations are reviewed by insurance providers who are not specially trained in HCV management, so the verbiage used perplexes these reviewers, possibly causing them to issue denials. Two, insurance providers often require HCV genotype testing for DAA medication eligibility, and HCV providers order genotype tests for patients only when HCV treatments have failed to cure patients, so this requirement becomes another barrier to DAA medications. Patients who spoke a non-English language, lived in the USA for less than 10 years, and showed inability to pay for treatment had received treatment despite these characteristics being common barriers to HCV treatment. On multivariate regression, factors independently associated with patients starting treatment included prior denial for DAA medication (odds ratio (OR), 8.88; 95% confidence interval (CI), 3.22 - 24.6; P < 0.001) and being seen by an HCV provider (OR, 24.8; 95% CI, 11.7 - 52.5; P < 0.001). However, the most significant barrier to HCV treatment access for the FQHC population was eligibility restrictions from insurance providers. Demographic-based barriers (e.g., age, race, and income) often impede HCV care and treatment, but insurance-based barriers are the greatest challenge currently that affects treatment outcomes in our study population. Removing these restrictions would, in our opinion, help to increase treatment levels to underserved populations.


A year-long extended release nanoformulated cabotegravir prodrug.
Long-acting cabotegravir (CAB) extends antiretroviral drug administration from daily to monthly. However, dosing volumes, injection site reactions and health-care oversight are obstacles towards a broad usage. The creation of poloxamer-coated hydrophobic and lipophilic CAB prodrugs with controlled hydrolysis and tissue penetrance can overcome these obstacles. To such ends, fatty acid ester CAB nanocrystal prodrugs with 14, 18 and 22 added carbon chains were encased in biocompatible surfactants named NMCAB, NM2CAB and NM3CAB and tested for drug release, activation, cytotoxicity, antiretroviral activities, pharmacokinetics and biodistribution. Pharmacokinetics studies, performed in mice and rhesus macaques, with the lead 18-carbon ester chain NM2CAB, showed plasma CAB levels above the protein-adjusted 90% inhibitory concentration for up to a year. NM2CAB, compared with NMCAB and NM3CAB, demonstrated a prolonged drug release, plasma circulation time and tissue drug concentrations after a single 45 mg per kg body weight intramuscular injection. These prodrug modifications could substantially improve CAB's effectiveness.


Anaerobic conversion of lactic acid to acetic acid and 1, 2-propanediol by Lactobacillus buchneri.
The degradation of lactic acid under anoxic conditions was studied in several strains of Lactobacillus buchneri and in close relatives such as Lactobacillus parabuchneri, Lactobacillus kefir, and Lactobacillus hilgardii. Of these lactobacilli, L. buchneri and L. parabuchneri were able to degrade lactic acid under anoxic conditions, without requiring an external electron acceptor. Each mole of lactic acid was converted into approximately 0.5 mol of acetic acid, 0.5 mol of 1,2-propanediol, and traces of ethanol. Based on stoichiometry studies and the high levels of NAD-linked 1, 2-propanediol-dependent oxidoreductase (530 to 790 nmol min(-1) mg of protein(-1)), a novel pathway for anaerobic lactic acid degradation is proposed. The anaerobic degradation of lactic acid by L. buchneri does not support cell growth and is pH dependent. Acidic conditions are needed to induce the lactic-acid-degrading capacity of the cells and to maintain the lactic-acid-degrading activity. At a pH above 5.8 hardly any lactic acid degradation was observed. The exact function of anaerobic lactic acid degradation by L. buchneri is not certain, but some results indicate that it plays a role in maintaining cell viability.


An epoch-making application of discharge plasma phenomenon to gene-transfer.
We discovered an epoch-making gene transfer method utilizing discharge plasma. Although an electroporation method is commonly used in present gene transfer experiments, it cannot transfer genes into primary cells sufficiently. The atmospheric pressure discharge plasma employed in this study was originally used for surface treatment of non-biological materials. We hypothesized that it could provide a suitable effect on the surface of target cells and applied it to gene transfer into various types of cells. The plasma technology succeeded in the efficient transfer of green fluorescence protein (GFP) plasmid into post-mitotic neuronal cells obtained from cerebral cortices of rats, into which an electroporation with conventional equipment cannot transfer genes sufficiently, as the cells were attached. After the transfection of rat pheochromocytoma PC12 cells with the GFP gene by plasma treatment, the cells retained their function, that is, nerve growth factor-induced differentiation. Furthermore, gene transfer with the plasma technology was also applicable to other types of cell lines such as HeLa cells and Chinese hamster lung (CHL) cells as adherent cell lines, and Jurkat cells as a suspended cell line, and another type of primary cell, human umbilical vein endothelial cells (HUVEC). In conclusion, the plasma method is an epoch-making gene transfer technology which efficiently transfers genes into primary cells into which electroporation cannot transfer genes. Moreover, the method is able to universally transfer genes into various types of cells as the function of the cells was maintained.


Asymmetric division and cosegregation of template DNA strands in adult muscle satellite cells.
Satellite cells assure postnatal skeletal muscle growth and repair. Despite extensive studies, their stem cell character remains largely undefined. Using pulse-chase labelling with BrdU to mark the putative stem cell niche, we identify a subpopulation of label-retaining satellite cells during growth and after injury. Strikingly, some of these cells display selective template-DNA strand segregation during mitosis in the muscle fibre in vivo, as well as in culture independent of their niche, indicating that genomic DNA strands are nonequivalent. Furthermore, we demonstrate that the asymmetric cell-fate determinant Numb segregates selectively to one daughter cell during mitosis and before differentiation, suggesting that Numb is associated with self-renewal. Finally, we show that template DNA cosegregates with Numb in label-retaining cells that express the self-renewal marker Pax7. The cosegregation of 'immortal' template DNA strands and their link with the asymmetry apparatus has important implications for stem cell biology and cancer.


The fate of the compressed deformed spinal cord after decompressive surgery: MR imaging and histopathological findings in experimental studies.
The authors conducted a study in which they applied the spinal cord compression-decompression model in rabbits to investigate the morphological changes and histopathological findings in the deformed spinal cord over a long-term period after performing decompressive surgery. Throughout the experimental period, mangnetic resonance (MR) images were obtained frequently; after obtaining a final MR image, the spinal cord was dissected and underwent histological examination. Immediately after decompressive surgery, axial T1-wieighted MR imaging revealed an increase in the cross-sectional area of the spinal cord during the 1st and 2nd weeks. The spinal cord area achieved a peak at an average of 5.9 weeks after decompressive surgery, when it displayed isointensity on T1- and high-intensity on T2-weighted images. The main histological findings were spongy changes in the white matter, which persisted for 4 months postsurgery. There was a significant correlation between the presurgical spinal cord area and the postsurgical decreased number of motor neuron cells. Based on the MR imaging and histopathological studies, although the deformed spinal cord that underwent compression for 3 months was immediately restored morphologically after decompressive surgery, the change in quality in the spinal cord persisted at least 4 months.


Income differences in COVID-19 incidence and severity in Finland among people with foreign and native background: A population-based cohort study of individuals nested within households.
Although intrahousehold transmission is a key source of Coronavirus Disease 2019 (COVID-19) infections, studies to date have not analysed socioeconomic risk factors on the household level or household clustering of severe COVID-19. We quantify household income differences and household clustering of COVID-19 incidence and severity. We used register-based cohort data with individual-level linkage across various administrative registers for the total Finnish population living in working-age private households (N = 4,315,342). Incident COVID-19 cases (N = 38,467) were identified from the National Infectious Diseases Register from 1 July 2020 to 22 February 2021. Severe cases (N = 625) were defined as having at least 3 consecutive days of inpatient care with a COVID-19 diagnosis and identified from the Care Register for Health Care between 1 July 2020 and 31 December 2020. We used 2-level logistic regression with individuals nested within households to estimate COVID-19 incidence and case severity among those infected. Adjusted for age, sex, and regional characteristics, the incidence of COVID-19 was higher (odds ratio [OR] 1.67, 95% CI 1.58 to 1.77, p < 0.001, 28.4% of infections) among individuals in the lowest household income quintile than among those in the highest quintile (18.9%). The difference attenuated (OR 1.23, 1.16 to 1.30, p < 0.001) when controlling for foreign background but not when controlling for other household-level risk factors. In fact, we found a clear income gradient in incidence only among people with foreign background but none among those with native background. The odds of severe illness among those infected were also higher in the lowest income quintile (OR 1.97, 1.52 to 2.56, p < 0.001, 28.0% versus 21.6% in the highest quintile), but this difference was fully attenuated (OR 1.08, 0.77 to 1.52, p = 0.64) when controlling for other individual-level risk factors-comorbidities, occupational status, and foreign background. Both incidence and severity were strongly clustered within households: Around 77% of the variation in incidence and 20% in severity were attributable to differences between households. The main limitation of our study was that the test uptake for COVID-19 may have differed between population subgroups. Low household income appears to be a strong risk factor for both COVID-19 incidence and case severity, but the income differences are largely driven by having foreign background. The strong household clustering of incidence and severity highlights the importance of household context in the prevention and mitigation of COVID-19 outcomes.


Assessing the calorific significance of episodes of human cannibalism in the Palaeolithic.
Episodes of Palaeolithic cannibalism have frequently been defined as 'nutritional' in nature, but with little empirical evidence to assess their dietary significance. This paper presents a nutritional template that offers a proxy calorie value for the human body. When applied to the Palaeolithic record, the template provides a framework for assessing the dietary value of prehistoric cannibalistic episodes compared to the faunal record. Results show that humans have a comparable nutritional value to those faunal species that match our typical body weight, but significantly lower than a range of fauna often found in association with anthropogenically modified hominin remains. This could suggest that the motivations behind hominin anthropophagy may not have been purely nutritionally motivated. It is proposed here that the comparatively low nutritional value of hominin cannibalism episodes support more socially or culturally driven narratives in the interpretation of Palaeolithic cannibalism.


A reexamination of krill oil bioavailability studies.
It has proven difficult to compare the bioavailability of krill oil (KO) vs. fish oil (FO) due to several of the characteristics of KO. These include the lower concentration of the active ingredients, eicosapentaenoic acid (EPA, 20:5n-3) and docosahexaenoic acid (DHA, 22:6n3), in KO as well as differences in their ratio relative to FO as well as the red color due to astaxanthin. In addition, the lipid classes in which EPA and DHA are found are quite different with KO containing phospholipid, di- and tri-glycerides as well as non-esterified fatty acid forms and with FO being primarily triglycerides. No human study has yet been performed that matches the dose of EPA and DHA in a randomized, controlled trial with measures of bloodstream EPA and DHA content. However, several claims have been made suggesting greater bioavailability of KO vs. FO. These have largely been based on a statistical argument where a somewhat lower dose of KO has been used to result in a similar bloodstream level of EPA and/or DHA or their total. However, the magnitude of the dosage differential is shown to be too small to be expected to result in differing blood levels of the long chain n-3 PUFAs. Some studies which have claimed to provide equal doses of KO and FO have actually used differing amounts of the two major n-3 fatty acid constituents. It is concluded that there is at present no evidence for greater bioavailability of KO vs. FO and that more carefully controlled human trials must be performed to establish their relative efficacies after chronic administration.


Oat and Barley in the Food Supply and Use of Beta Glucan Health Claims.
Beta glucan is a type of soluble dietary fibre found in oats and barley with known cholesterol-lowering benefits. Many countries globally have an approved beta glucan health claim related to lowering blood cholesterol, an important biomarker for cardiovascular disease. However, the use of these claims has not been examined. The aim of this study was to explore the range and variety of oat and barley products in the Australian and global market within a defined range of grain food and beverage categories and examine the frequency of beta glucan health claims. Australian data were collected via a recognised nutrition audit process from the four major Australian supermarkets in metropolitan Sydney (January 2018 and September 2020) and Mintel Global New Product Database was used for global markets where a claim is permitted. Categories included breakfast cereals, bread, savoury biscuits, grain-based muesli bars, flour, noodles/pasta and plant-based milk alternatives and information collected included ingredients lists and nutrition and health claims. Products from Australia (n = 2462) and globally (n = 44,894) were examined. In Australia, 37 products (1.5%) made use of the beta glucan claim (84% related to oat beta glucan and 16% related to barley beta glucan, specifically BARLEYmax®). Of products launched globally, 0.9% (n = 403) displayed beta glucan cholesterol-lowering claims. Despite the number of products potentially eligible to make beta glucan claims, their use in Australia and globally is limited. The value of dietary modification in cardiovascular disease treatment and disease progression deserves greater focus, and health claims are an opportunity to assist in communicating the role of food in the management of health and disease. Further assessment of consumer understanding of the available claims would be of value.


Transcription of the human ferredoxin gene through a single promoter which contains the 3',5'-cyclic adenosine monophosphate-responsive sequence and Sp 1-binding site.
We have investigated the functional elements involved in cAMP-stimulated transcription of the human ferredoxin gene. Unlike the bovine gene, the human gene lacked a second upstream RNA initiation site as demonstrated by sequence analysis of the exon boundary, lack of upstream RNA, and analysis of the promoter. The presence of a single promoter was determined by testing the ability of various gene segments to drive the expression of the chloramphenicol acetyltransferase gene after transfection into a mouse adrenal cell line Y1. Full promoter activity was conferred by a DNA fragment spanning -209 to +55, although the -94 to +55 fragment already provided some promoter activity. Transcription from the -94 to +55 segment was stimulated by 2-fold when 8-bromo-cAMP was added to the cell. Footprinting analyses showed two GC boxes at -50 to -70 and -87 to -108 were protected by proteins from both Y1 and HeLa cells. Competition experiments showed that a protein with a recognition sequence indistinguishable from Sp1 bound to these sites. When connected to a heterologous TATA box, the sequence at -76 to -42, which contained the proximal GC box, was able to confer a high level of basal transcription and cAMP stimulation. This sequence does not show sequence homology with the known cAMP-responsive element. Mutations or deletion of the Sp 1-binding site showed diminished basal transcription and defined the cAMP responsive sequence to be from -76 to -62. Therefore the cAMP-responsive sequence of the human ferredoxin gene was located at -76 to -62, which was adjacent to the Sp 1-binding site.


A combined cryo-EM and molecular dynamics approach reveals the mechanism of ErmBL-mediated translation arrest.
Nascent polypeptides can induce ribosome stalling, regulating downstream genes. Stalling of ErmBL peptide translation in the presence of the macrolide antibiotic erythromycin leads to resistance in Streptococcus sanguis. To reveal this stalling mechanism we obtained 3.6-Å-resolution cryo-EM structures of ErmBL-stalled ribosomes with erythromycin. The nascent peptide adopts an unusual conformation with the C-terminal Asp10 side chain in a previously unseen rotated position. Together with molecular dynamics simulations, the structures indicate that peptide-bond formation is inhibited by displacement of the peptidyl-tRNA A76 ribose from its canonical position, and by non-productive interactions of the A-tRNA Lys11 side chain with the A-site crevice. These two effects combine to perturb peptide-bond formation by increasing the distance between the attacking Lys11 amine and the Asp10 carbonyl carbon. The interplay between drug, peptide and ribosome uncovered here also provides insight into the fundamental mechanism of peptide-bond formation.


[Surveillance of foodborne diseases in the province of Rio Negro, Argentina, 1993-2001].
A total of 39 outbreaks of foodborne diseases affecting 958 people in the province of Rio Negro, Argentina between 1993 and 2001 are described and evaluated. The main causal agents were identified involving food, sites of occurrence, risk factors and notification system used. Salmonella spp (38%), Trichinella spiralis (15%), Escherichia coli (13%) and Staphylococcus aureus (15%) were the most frequent agents present in outbreaks. Salmonella spp produced the largest number of cases (52%). Food involved were cooked meat (36%), cheese (10%), sandwiches (10%), deserts (10%) and ice cream (8%). Indeed, ice creams were involved in the largest number of cases and of people affected. In relation to the source of food, 41% of outbreaks were caused by homemade meals, 23% by catering or ice cream parlor, 13% in family parties, 8% in county fairs and 8% in hotel restaurants. In 28% of the outbreaks the etiological agent was identified exclusively by epidemiological analysis, in 64% isolation of the agent was carried out, and in 8% of the cases, a final diagnosis could not be obtained. Validity of epidemiological studies in foodborne disease, the necessity of strengthening the notification system of outbreaks, and the importance of good practices in food handling are analyzed.


National priority setting partnership using a Delphi consensus process to develop neonatal research questions suitable for practice-changing randomised trials in the United Kingdom.
Methodologically robust clinical trials are required to improve neonatal care and reduce unwanted variations in practice. Previous neonatal research prioritisation processes have identified important research themes rather than specific research questions amenable to clinical trials. Practice-changing trials require well-defined research questions, commonly organised using the Population, Intervention, Comparison, Outcome (PICO) structure. By narrowing the scope of research priorities to those which can be answered in clinical trials and by involving a wide range of different stakeholders, we aim to provide a robust and transparent process to identify and prioritise research questions answerable within the National Healthcare System to inform future practice-changing clinical trials. A steering group comprising parents, doctors, nurses, allied health professionals, researchers and representatives from key organisations (Neonatal Society, British Association of Perinatal Medicine, Neonatal Nurses Association and Royal College of Paediatrics and Child Health) was identified to oversee this project. We will invite submissions of research questions formatted using the PICO structure from the following stakeholder groups using an online questionnaire: parents, patients, healthcare professionals and academic researchers. Unanswered, non-duplicate research questions will be entered into a three-round eDelphi survey of all stakeholder groups. Research questions will be ranked by mean aggregate scores. The final list of prioritised research questions will be disseminated through traditional academic channels, directly to key stakeholder groups through representative organisations and on social media. The outcome of the project will be shared with key research organisations such as the National Institute for Health Research. Research ethics committee approval is not required.


Environmental harshness is positively correlated with intraspecific divergence in mammals and birds.
Life on Earth is conspicuously more diverse in the tropics. Although this intriguing geographical pattern has been linked to many biotic and abiotic factors, their relative importance and potential interactions are still poorly understood. The way in which latitudinal changes in ecological conditions influence evolutionary processes is particularly controversial, as there is evidence for both a positive and a negative latitudinal gradient in speciation rates. Here, we identify and address some methodological issues (how patterns are analysed and how latitude is quantified) that could lead to such conflicting results. To address these issues, we assemble a comprehensive data set of the environmental correlates of latitude (including climate, net primary productivity and habitat heterogeneity) and combine it with biological, historical and molecular data to explore global patterns in recent divergence events (subspeciation). Surprisingly, we find that the harsher conditions that typify temperate habitats (lower primary productivity, decreased rainfall and more variable and unpredictable temperatures) are positively correlated with greater subspecies richness in terrestrial mammals and birds. Thus, our findings indicate that intraspecific divergence is greater in regions with lower biodiversity, a pattern that is robust to both sampling variation and latitudinal biases in taxonomic knowledge. We discuss possible causal mechanisms for the link between environmental harshness and subspecies richness (faster rates of evolution, greater likelihood of range discontinuities and more opportunities for divergence) and conclude that this pattern supports recent indications that latitudinal gradients of diversity are maintained by simultaneously higher potentials for both speciation and extinction in temperate than tropical regions.


A transposable element insertion is associated with an alternative life history strategy.
Tradeoffs affect resource allocation during development and result in fitness consequences that drive the evolution of life history strategies. Yet despite their importance, we know little about the mechanisms underlying life history tradeoffs. Many species of Colias butterflies exhibit an alternative life history strategy (ALHS) where females divert resources from wing pigment synthesis to reproductive and somatic development. Due to this reallocation, a wing color polymorphism is associated with the ALHS: either yellow/orange or white. Here we map the locus associated with this ALHS in Colias crocea to a transposable element insertion located downstream of the Colias homolog of BarH-1, a homeobox transcription factor. Using CRISPR/Cas9 gene editing, antibody staining, and electron microscopy we find white-specific expression of BarH-1 suppresses the formation of pigment granules in wing scales and gives rise to white wing color. Lipid and transcriptome analyses reveal physiological differences associated with the ALHS. Together, these findings characterize a mechanism for a female-limited ALHS.


Validation study of artificial neural network models for prediction of methicillin-resistant Staphylococcus aureus carriage.
Use of active surveillance cultures for methicillin-resistant Staphylococcus aureus (MRSA) for all patients admitted to the intensive care unit has been shown to reduce nosocomial transmission. However, the cost-effectiveness and the utility of implementing use of active surveillance cultures nationwide remain controversial. We sought to develop an artificial neural network (ANN) model that would predict the likelihood of MRSA colonization. Two acute care hospitals, one in Pittsburgh (hospital A) and one in Kaohsiung, Taiwan (hospital B). Nasal cultures were performed for all patients admitted to the hospitals. A total of 46 potential risk factors in hospital A and 86 potential risk factors in hospital B associated with MRSA colonization were assessed. Culture results were obtained; 75% of the data were used for training our ANN model, and the remaining 25% were used for validating our ANN model. The culture results were the "gold standard" for determining the accuracy of the model predictions. The ANN model predictions were accurate 95.2% of the time for hospital A (sensitivity, 94.3%; specificity, 96.0%) and 94.2% of the time for hospital B (sensitivity, 96.6%; specificity, 91.8%), integrating all potential risk factors into the model. Only 17 potential risk factors were needed for the hospital A ANN model (accuracy, 90.9%; sensitivity, 98.5%; specificity, 83.4%), and only 20 potential risk factors were needed for the hospital B ANN model (accuracy, 90.5%; sensitivity, 96.6%; specificity, 84.3%), if the minimal risk factor method was used. Cross-validation analysis showed an average accuracy of 85.6% (sensitivity, 91.3%; specificity, 80.0%). Our ANN model can be used to predict with an accuracy of more than 90% which patients carry MRSA. The false-negative rates were significantly lower than the false-positive rates in the ANN predictions, which can serve as a safety buffer in case of patient misclassification.


Mutant prevention concentrations of ciprofloxacin and enrofloxacin for Salmonella enterica.
To determine the mutant prevention concentrations (MPCs) of ciprofloxacin and enrofloxacin against four strains of Salmonella enterica serovar Enteritidis and four strains of S. Typhimurium including one fully susceptible, one multiply resistant (MAR), one GyrA mutant and one GyrA/MAR mutant. Further, to examine mutants arising after exposure to sub-MPC concentrations of the antibiotics for susceptibility to ciprofloxacin and enrofloxacin, and cyclohexane tolerance. MICs were determined using the agar dilution method of the BSAC. The MPC was recorded as the lowest concentration of antibiotic to inhibit growth from an inoculum of 10(10) cfu. The MPCs and resulting MPC/MIC ratios of enrofloxacin were generally two- to four-fold higher than for ciprofloxacin. At 24 h for both antibiotics, MPCs were lowest for the fully susceptible strains (0.25-0.5 mg/L), similar for the MAR (1-4 mg/L) and GyrA (2-4 mg/L) mutants and highest for the GyrA/MAR mutants (1-8 mg/L). MPC/MIC ratios at 24 h were 2-16 for all strains except those for the MAR strains without mutation in gyrA where the ratios were 8-64. The ability to eradicate Salmonella in vivo depends on many factors such as antibiotic susceptibility of the strain, dose and route of administration. It is suggested that these MPC values will be useful when considering dosing strategies. In view of the high MPC/MIC ratio, MAR strains with wild-type gyrA, although susceptible to ciprofloxacin (MICs 0.06-0.13 mg/L), may give rise to treatment failures.


Smooth muscle calponin: an unconventional CArG-dependent gene that antagonizes neointimal formation.
Smooth muscle calponin (CNN1) contains multiple conserved intronic CArG elements that bind serum response factor and display enhancer activity in vitro. The objectives here were to evaluate these CArG elements for activity in transgenic mice and determine the effect of human CNN1 on injury-induced vascular remodeling. Mice carrying a lacZ reporter under control of intronic CArG elements in the human CNN1 gene failed to show smooth muscle cell (SMC)-restricted activity. However, deletion of the orthologous sequences in mice abolished endogenous Cnn1 promoter activity, suggesting their necessity for in vivo Cnn1 expression. Mice carrying a 38-kb bacterial artificial chromosome (BAC) harboring the human CNN1 gene displayed SMC- restricted expression of the corresponding CNN1 protein, as measured by immunohistochemistry and Western blotting. Extensive BAC recombineering studies revealed the absolute necessity of a single intronic CArG element for correct SMC-restricted expression of human CNN1. Overexpressing human CNN1 suppressed neointimal formation following arterial injury. Mice with an identical BAC carrying mutations in CArG elements that inhibit human CNN1 expression showed outward remodeling and neointimal formation. A single intronic CArG element is necessary but insufficient for proper CNN1 expression in vivo. CNN1 overexpression antagonizes arterial injury-induced neointimal formation.


Characterisation and prognosis of undiagnosed chronic obstructive pulmonary disease patients at their first hospitalisation.
Under-diagnosis of COPD is an important unmet medical need. We investigated the characteristics and prognosis of hospitalised patients with undiagnosed COPD. The PAC-COPD cohort included 342 COPD patients hospitalised for the first time for an exacerbation of COPD (2004-2006). Patients were extensively characterised using sociodemographic, clinical and functional variables, and the cohort was followed-up through 2008. We defined "undiagnosed COPD" by the absence of any self-reported respiratory disease and regular use of any pharmacological respiratory treatment. Undiagnosed COPD was present in 34% of patients. They were younger (mean age 66 vs. 68 years, p = 0.03), reported fewer symptoms (mMRC dyspnoea score, 2.1 vs. 2.6, p < 0.01), and had a better health status (SGRQ total score, 29 vs. 40, p < 0.01), milder airflow limitation (FEV1% ref., 59% vs. 49%, p < 0.01), and fewer comorbidities (two or more, 40% vs. 56%, p < 0.01) when compared with patients with an established COPD diagnosis. Three months after hospital discharge, 16% of the undiagnosed COPD patients had stopped smoking (vs. 5%, p = 0.019). During follow-up, annual hospitalisation rates were lower in undiagnosed COPD patients (0.14 vs. 0.25, p < 0.01); however, this difference disappeared after adjustment for severity. Mortality was similar in both groups. Undiagnosed COPD patients have less severe disease and lower risk of re-hospitalisation when compared with hospitalised patients with known COPD.


Solid fuel use and cooking practices as a major risk factor for ALRI mortality among African children.
Almost half of global child deaths due to acute lower respiratory infections (ALRIs) occur in sub-Saharan Africa, where three-quarters of the population cook with solid fuels. This study aims to quantify the impact of fuel type and cooking practices on childhood ALRI mortality in Africa, and to explore implications for public health interventions. Early-release World Health Survey data for the year 2003 were pooled for 16 African countries. Among 32,620 children born during the last 10 years, 1455 (4.46%) were reported to have died prior to their fifth birthday. Survival analysis was used to examine the impact of different cooking-related parameters on ALRI mortality, defined as cough accompanied by rapid breathing or chest indrawing based on maternal recall of symptoms prior to death. Solid fuel use increases the risk of ALRI mortality with an adjusted hazard ratio of 2.35 (95% CI 1.22 to 4.52); this association grows stronger with increasing outcome specificity. Differences between households burning solid fuels on a well-ventilated stove and households relying on cleaner fuels are limited. In contrast, cooking with solid fuels in the absence of a chimney or hood is associated with an adjusted hazard ratio of 2.68 (1.38 to 5.23). Outdoor cooking is less harmful than indoor cooking but, overall, stove ventilation emerges as a more significant determinant of ALRI mortality. This study shows substantial differences in ALRI mortality risk among African children in relation to cooking practices, and suggests that stove ventilation may be an important means of reducing indoor air pollution.


Binary Switching of Calendar Cells in the Pituitary Defines the Phase of the Circannual Cycle in Mammals.
Persistent free-running circannual (approximately year-long) rhythms have evolved in animals to regulate hormone cycles, drive metabolic rhythms (including hibernation), and time annual reproduction. Recent studies have defined the photoperiodic input to this rhythm, wherein melatonin acts on thyrotroph cells of the pituitary pars tuberalis (PT), leading to seasonal changes in the control of thyroid hormone metabolism in the hypothalamus. However, seasonal rhythms persist in constant conditions in many species in the absence of a changing photoperiod signal, leading to the generation of circannual cycles. It is not known which cells, tissues, and pathways generate these remarkable long-term rhythmic processes. We show that individual PT thyrotrophs can be in one of two binary states reflecting either a long (EYA3(+)) or short (CHGA(+)) photoperiod, with the relative proportion in each state defining the phase of the circannual cycle. We also show that a morphogenic cycle driven by the PT leads to extensive re-modeling of the PT and hypothalamus over the circannual cycle. We propose that the PT may employ a recapitulated developmental pathway to drive changes in morphology of tissues and cells. Our data are consistent with the hypothesis that the circannual timer may reside within the PT thyrotroph and is encoded by a binary switch timing mechanism, which may regulate the generation of circannual neuroendocrine rhythms, leading to dynamic re-modeling of the hypothalamic interface. In summary, the PT-ventral hypothalamus now appears to be a prime structure involved in long-term rhythm generation.


Small mammal responses to long-term large-scale woodland creation: the influence of local and landscape-level attributes.
Habitat loss and fragmentation greatly affect biological diversity. Actions to counteract their negative effects include increasing the quality, amount and connectivity of seminatural habitats at the landscape scale. However, much of the scientific evidence underpinning landscape restoration comes from studies of habitat loss and fragmentation, and it is unclear whether the ecological principles derived from habitat removal investigations are applicable to habitat creation. In addition, the relative importance of local- (e.g., improving habitat quality) vs. landscape-level (e.g., increasing habitat connectivity) actions to restore species is largely unknown, partly because studying species responses over sufficiently large spatial and temporal scales is challenging. We studied small mammal responses to large-scale woodland creation spanning 150 yr, and assessed the influence of local- and landscape-level characteristics on three small mammal species of varying woodland affinity. Woodland specialists, generalists, and grassland specialists were present in woodlands across a range of ages from 10 to 160 yr, demonstrating that these species can quickly colonize newly created woodlands. However, we found evidence that woodlands become gradually better over time for some species. The responses of individual species corresponded to their habitat specificity. A grassland specialist (Microtus agrestis) was influenced only by landscape attributes; a woodland generalist (Apodemus sylvaticus) and specialist (Myodes glareolus) were primarily influenced by local habitat attributes, and partially by landscape characteristics. At the local scale, high structural heterogeneity, large amounts of deadwood, and a relatively open understory positively influenced woodland species (both generalists and specialists); livestock grazing had strong negative effects on woodland species abundance. Actions to enhance habitat quality at the patch scale focusing on these attributes would benefit these species. Woodland creation in agricultural landscapes is also likely to benefit larger mammals and birds of prey feeding on small mammals and increase ecosystem processes such as seed dispersal.


Cross-situational learning in a Zipfian environment.
Both adults and children have shown impressive cross-situational word learning in which they leverage the statistics of word usage across many different scenes in order to isolate specific word meanings (e.g., Yu & Smith, 2007). However, relatively little is known about how this learning scales to real language. Some theoretical analyses suggest that when words follow a Zipfian distribution, as they do in natural language, it should be more difficult to learn a lexicon because of the many low-frequency words that are only observed a few times (Blythe, Smith, & Smith, 2010; Vogt, 2012). Although this effect can be mitigated somewhat by assuming mutual exclusivity (Reisenauer, Smith, & Blythe, 2013), no mathematical analyses suggest that learning in a Zipfian environment should be easier. In this work, we show the opposite of the predicted effect using cross-situational learning experiments with adults: when the distribution of words and meanings is Zipfian, learning is not impaired and is usually improved. Over a series of experiments, we provide evidence that this is because Zipfian distributions help people to disambiguate the meanings of the other words in the situation.


Epidemiology of angioedema without wheals in an allergy and immunology center.
We describe the diagnostic epidemiology, the clinical course, the family history and the response to treatment of patients with angioedema without wheals (AWW) at an Allergy and Immunology Clinical Center. We reviewed the case records of all patients at our office from January 1997 to April 2013. We recorded sex, age, age at onset of symptoms, family history of angioedema, number of visits to the office, type of angioedema, and response to treatment from those patients with angioedema without wheals. We classified angioedema according to its pathophysiology. We also describe those patients with angioedema mimics. From a total of 17,823 new patients, 303 had a presumptive diagnosis of angioedema without wheals. Twenty-three patients had an angioedema mimic. Forty percent were male and 60% were female. Average age at first visit was 40.6. Average number of visits was 2.4. Fifty-seven patients referred a family history. We attributed idiopathic angioedema to 55.7% of patients, 24.3% were drug related, 15.7% were due to C1 inhibitor deficiency, 2.1% were drug related+idiopathic angioedema, 1.4% were type III and 0.7% had exercise-induced angioedema. Ninety six percent of 53 evaluable idiopathic angioedema patients referred a benefit with anti-histamine therapy. AWW was a rare cause of consultation. Most of our patients had anti H1 responsive idiopathic angioedema and none had allergic angioedema. Women cases prevailed over men's. Family history and average age of onset of symptoms were different among the different types of angioedema.


Tbx1: identification of a 22q11.2 gene as a risk factor for autism spectrum disorder in a mouse model.
Although twin studies indicate clear genetic bases of autism spectrum disorder (ASD), the precise mechanisms through which genetic variations causally result in ASD are poorly understood. Individuals with 3 Mb and nested 1.5 Mb hemizygosity of the chromosome 22q11.2 represent genetically identifiable cases of ASD. However, because more than 30 genes are deleted even in the minimal deletion cases of 22q11.2 deficiency, the individual 22q11.2 gene(s) responsible for ASD remain elusive. Here, we examined the impact of constitutive heterozygosity of Tbx1, a 22q11.2 gene, on the behavioral phenotypes of ASD and characterized the regional and cellular expression of its mRNA and protein in mice. Congenic Tbx1 heterozygous (HT) mice were impaired in social interaction, ultrasonic vocalization, memory-based behavioral alternation, working memory and thigmotaxis, compared with wild-type (WT) mice. These phenotypes were not due to non-specific alterations in olfactory function, exploratory behavior, motor movement or anxiety-related behavior. Tbx1 mRNA and protein were ubiquitously expressed throughout the brains of C57BL/6J mice, but protein expression was enriched in regions that postnatally retain the capacity of neurogenesis, and in fact, postnatally proliferating cells expressed Tbx1. In postnatally derived hippocampal culture cells of C57BL/6J mice, Tbx1 levels were higher during proliferation than during differentiation, and expressed in neural progenitor cells, immature and matured neurons and glial cells. Taken together, our data suggest that Tbx1 is a gene responsible for the phenotypes of 22q11.2 hemizygosity-associated ASD possibly through its role in diverse cell types, including postnatally and prenatally generated neurons.


Unstable laser resonator modes.
In this paper, mode patterns and losses are determined for unstable laser resonators with finite, rectangular reflectors of spherical curvature. An analysis of the uniform intensity mode, based upon the Cornu spiral is given. For more general calculations, gaussian quadrature integration is used to convertthe integral equation for the modes into a matrix equation. The latter is solved using ALLMIAT on a digital computer, thereby simultaneously determining many modes. The mode competition effectreported by Siegman and Arrathoon is shown to occur because the unstable modes do not generally retain their ordering, according to relative loss, as the reflector size changes. We also discuss a perturbation calculation in which the infinite mirror solutions of Bergstein are used as expansion functions.


Hepatitis-C virus antibodies (anti HCV) in haemodialyzed vs non-dialyzed patients.
Anti HCV was checked using Enzyme Immuno assay (EIA) (C100-3-Abbott) in 68 patients with chronic renal failure (CRF) who were on maintenance haemodialysis and 48 patients on conservative management. Mean age of the patients was 50 years. The duration of illness ranged from 3 months to 18 years (mean 3 years). All patients were Hepatitis B surface antigen (HBsAg) negative. In haemodialyzed group 31 (46%) and in conservative group only 3 (6%) were Anti HCV positive. High Frequency of Anti HCV positivity in haemodialyzed group was related to period on dialysis (1 year) and number of blood transfusions (>4 units). Patients in whom dialyzer was re-used showed 60% positivity as compared to only 17% in those with single use. Anti HCV positivity predominated in females as compared to males, 42.6% vs 20.3%. Nine out of 10 patients with a history of jaundice and six out of 8 patients with raised transaminase levels were Anti HCV positive. Fifteen of 37 Anti HBc (IgG) positive cases were Anti HCV +ve.


Committee opinion No. 532: compounded bioidentical menopausal hormone therapy.
Although improvement in long-term health is no longer an indication for menopausal hormone therapy, evidence supporting fewer adverse events in younger women, combined with its high overall effectiveness, has reinforced its usefulness for short-term treatment of menopausal symptoms. Menopausal therapy has been provided not only by commercially available products but also by compounding, or creation of an individualized preparation in response to a health care provider's prescription to create a medication tailored to the specialized needs of an individual patient. The Women's Health Initiative findings, coupled with an increase in the direct-to-consumer marketing and media promotion of compounded bioidentical hormonal preparations as safe and effective alternatives to conventional menopausal hormone therapy, have led to a recent increase in the popularity of compounded bioidentical hormones as well as an increase in questions about the use of these preparations. Not only is evidence lacking to support superiority claims of compounded bioidentical hormones over conventional menopausal hormone therapy, but these claims also pose the additional risks of variable purity and potency and lack efficacy and safety data. The Committee on Gynecologic Practice of the American College of Obstetricians and Gynecologists and the Practice Committee of the American Society for Reproductive Medicine provide an overview of the major issues of concern surrounding compounded bioidentical menopausal hormone therapy and provide recommendations for patient counseling.


Multifaceted actions of Zeb2 in postnatal neurogenesis from the ventricular-subventricular zone to the olfactory bulb.
The transcription factor Zeb2 controls fate specification and subsequent differentiation and maturation of multiple cell types in various embryonic tissues. It binds many protein partners, including activated Smad proteins and the NuRD co-repressor complex. How Zeb2 subdomains support cell differentiation in various contexts has remained elusive. Here, we studied the role of Zeb2 and its domains in neurogenesis and neural differentiation in the young postnatal ventricular-subventricular zone (V-SVZ), in which neural stem cells generate olfactory bulb-destined interneurons. Conditional Zeb2 knockouts and separate acute loss- and gain-of-function approaches indicated that Zeb2 is essential for controlling apoptosis and neuronal differentiation of V-SVZ progenitors before and after birth, and we identified Sox6 as a potential downstream target gene of Zeb2. Zeb2 genetic inactivation impaired the differentiation potential of the V-SVZ niche in a cell-autonomous fashion. We also provide evidence that its normal function in the V-SVZ also involves non-autonomous mechanisms. Additionally, we demonstrate distinct roles for Zeb2 protein-binding domains, suggesting that Zeb2 partners co-determine neuronal output from the mouse V-SVZ in both quantitative and qualitative ways in early postnatal life.


The use and role of open source software applications in public and not-for-profit hospitals in the United States.
The potential cost savings and customizability of open source software (OSS) may be particularly attractive for hospitals. However, numerous health-care-specific OSS applications exist, the adoption of OSS health information technology (HIT) applications is not widespread in the United States. This disconnect between the availability of promising software and low adoption raises the basic question: If OSS HIT is so advantageous, why are more health care organizations not using it? We interviewed the chief information officer, or equivalent position, at 17 not-for-profit and public hospitals across the United States. Through targeted recruitment, our sample included nine hospitals using OSS HIT and eight hospitals not using OSS HIT. The open-ended interview questions were guided by domains included in the fit-viability theory, an organizational-level innovation adoption framework, and those suggested by a review of the literature. Transcripts were analyzed using an inductive and comparative approach, which involved an open coding for relevant themes. Interviews described the state of OSS use in hospitals. Specifically, general OSS applications were widely used by IT professionals. In addition, hospitals using OSS HIT still relied heavily on vendor support. In terms of why decisions arose to use OSS HIT, several hospitals using OSS HIT noted the cost advantages. In contrast, hospitals avoiding OSS HIT were clear, OSS as a class did not fit with clinical work and posed too much risk. Perceptions of OSS HIT ranged from enthusiastic embracement to resigned adoption, to refusal, to abandonment. Some organizations were achieving success with their OSS HIT choices, but they still relied on vendors for significant support. The decision to adopt OSS HIT was not uniform but contingent upon views of the risk posed by the technology, economic factors, and the hospital's existing capabilities.


Cellular apoptosis is associated with increased caveolin-1 expression in macrophages.
Macrophage apoptosis is an important factor in determining the efficiency of the immune response, atherosclerotic lesion stability, and clearance of aged cells by phagocytosis. The involvement of caveolin-1 in the regulation of apoptosis has been previously suggested in fibroblasts and epithelial cells. Here we show that treatment of thioglycollate-elicited mouse peritoneal macrophages with various unrelated apoptotic agents, including simvastatin, camptothecin, or glucose deprivation, is associated with a specific and large increase in caveolin-1 expression. In contrast, caveolin-2 levels remain unaffected. Induction of apoptosis was measured by changes in cell morphology, annexin V-labeling, and DNA fragmentation. We demonstrate that caveolin-1 in macrophages is present in lipid rafts and colocalizes with phosphatidylserine (PS) at the cell surface of apoptotic macrophages. Our data suggest that caveolin-1 increase is an early event, closely accompanied by PS externalization and independent of caspase activation and nuclear DNA fragmentation. The increase in caveolin-1 levels does not require new protein synthesis, as cycloheximide does not prevent the apoptosis-mediated increase in caveolin-1 levels. We propose that increased levels of caveolin-1 characterize the apoptotic phenotype of macrophages. Caveolin-1 may be involved in the efficient externalization of PS at the surface of the apoptotic cells.


Magnesium Status and Stress: The Vicious Circle Concept Revisited.
Magnesium deficiency and stress are both common conditions among the general population, which, over time, can increase the risk of health consequences. Numerous studies, both in pre-clinical and clinical settings, have investigated the interaction of magnesium with key mediators of the physiological stress response, and demonstrated that magnesium plays an inhibitory key role in the regulation and neurotransmission of the normal stress response. Furthermore, low magnesium status has been reported in several studies assessing nutritional aspects in subjects suffering from psychological stress or associated symptoms. This overlap in the results suggests that stress could increase magnesium loss, causing a deficiency; and in turn, magnesium deficiency could enhance the body's susceptibility to stress, resulting in a magnesium and stress vicious circle. This review revisits the magnesium and stress vicious circle concept, first introduced in the early 1990s, in light of recent available data.


Stem cell transplantation for spinal cord injury: a meta-analysis of treatment effectiveness and safety.
The aim of this study was to evaluate the effectiveness and safety of stem cell transplantation for spinal cord injury (SCI). PubMed, EMBASE, Cochrane, China National Knowledge Infrastructure, China Science and Technology Journal, Wanfang, and SinoMed databases were systematically searched by computer to select clinical randomized controlled trials using stem cell transplantation to treat SCI, published between each database initiation and July 2016. Randomized controlled trials comparing stem cell transplantation with rehabilitation treatment for patients with SCI. Inclusion criteria: (1) Patients with SCI diagnosed according to the American Spinal Injury Association (ASIA) International standards for neurological classification of SCI; (2) patients with SCI who received only stem cell transplantation therapy or stem cell transplantation combined with rehabilitation therapy; (3) one or more of the following outcomes reported: outcomes concerning neurological function including sensory function and locomotor function, activities of daily living, urination functions, and severity of SCI or adverse effects. Studies comprising patients with complications, without full-text, and preclinical animal models were excluded. Quality of the included studies was evaluated using the Cochrane risk of bias assessment tool and RevMan V5.3 software, provided by the Cochrane Collaboration, was used to perform statistical analysis. ASIA motor score, ASIA light touch score, ASIA pinprick score, ASIA impairment scale grading improvement rate, activities of daily living score, residual urine volume, and adverse events. Ten studies comprising 377 patients were included in the analysis and the overall risk of bias was relatively low level. Four studies did not detail how random sequences were generated, two studies did not clearly state the blinding outcome assessment, two studies lacked blinding outcome assessment, one study lacked follow-up information, and four studies carried out selective reporting. Compared with rehabilitation therapy, stem cell transplantation significantly increased the lower limb light touch score (odds ratio (OR) = 3.43, 95% confidence interval (CI): 0.01 - 6.86, P = 0.05), lower limb pinprick score (OR = 3.93, 95%CI: 0.74 - 7.12, P = 0.02), ASI grading rate (relative risk (RR) = 2.95, 95%CI: 1.64 - 5.29, P = 0.0003), and notably reduced residual urine volume (OR = -8.10, 95%CI: -15.09 to -1.10, P = 0.02). However, stem cell transplantation did not significantly improve motor score (OR = 1.89, 95%CI: -0.25 to 4.03, P = 0.08) or activities of daily living score (OR = 1.12, 95%CI: -1.17 to 4.04, P = 0.45). Furthermore, stem cell transplantation caused a high rate of mild adverse effects (RR = 14.49, 95%CI: 5.34 - 34.08, P < 0.00001); however, these were alleviated in a short time. Stem cell transplantation was determined to be an efficient and safe treatment for SCI and simultaneously improved sensory and bladder functions. Although associated minor and temporary adverse effects were observed with transplanted stem cells, spinal cord repair and axon remyelination were apparent. More randomized controlled trials with larger sample sizes and longer follow-up times are needed to further validate the effectiveness of stem cell transplantation in the treatment of SCI.


Inverse agonist and neutral antagonist actions of synthetic compounds at an insect 5-HT1 receptor.
5-Hydroxytryptamine (5-HT) has been shown to control and modulate many physiological and behavioural functions in insects. In this study, we report the cloning and pharmacological properties of a 5-HT(1) receptor of an insect model for neurobiology, physiology and pharmacology. A cDNA encoding for the Periplaneta americana 5-HT(1) receptor was amplified from brain cDNA. The receptor was stably expressed in HEK 293 cells, and the functional and pharmacological properties were determined in cAMP assays. Receptor distribution was investigated by RT-PCR and by immunocytochemistry using an affinity-purified polyclonal antiserum. The P. americana 5-HT(1) receptor (Pea5-HT(1)) shares pronounced sequence and functional similarity with mammalian 5-HT(1) receptors. Activation with 5-HT reduced adenylyl cyclase activity in a dose-dependent manner. Pea5-HT(1) was expressed as a constitutively active receptor with methiothepin acting as a neutral antagonist, and WAY 100635 as an inverse agonist. Receptor mRNA was present in various tissues including brain, salivary glands and midgut. Receptor-specific antibodies showed that the native protein was expressed in a glycosylated form in membrane samples of brain and salivary glands. This study marks the first pharmacological identification of an inverse agonist and a neutral antagonist at an insect 5-HT(1) receptor. The results presented here should facilitate further analyses of 5-HT(1) receptors in mediating central and peripheral effects of 5-HT in insects.


Circulating levels of cell-derived microparticles are reduced by mild hypobaric hypoxia: data from a randomised controlled trial.
Hypoxia is known to induce the release of microparticles in vitro. However, few publications have addressed the role of hypoxia in vivo on circulating levels of microparticles. This randomised, controlled, crossover trial aimed to determine the effect of mild hypoxia on in vivo levels of circulating microparticles in healthy individuals. Blood was obtained from 51 healthy male volunteers (mean age of 26.9 years) at baseline altitude (490 m) and after 24 and 48 h at moderate altitude (2,590 m). The order of altitude exposure was randomised. Flow cytometry was used to assess platelet-poor plasma for levels of circulating microparticles derived from platelets, endothelial cells, leucocytes, granulocytes, monocytes, red blood cells and procoagulant microparticles. Mean (standard deviation) oxygen saturation was significantly lower on the first and second day after arrival at 2,590 m, 91.0 (2.0) and 92.0 (2.0) %, respectively, compared to 490 m, 96 (1.0) %, p < 0.001 for both comparisons. A significant decrease in the levels of procoagulant microparticles (annexin V+ -221/μl 95 % CI -370.8/-119.0, lactadherin+ -202/μl 95 % CI -372.2/-93.1), platelet-derived microparticles (-114/μl 95 % CI -189.9/-51.0) and red blood cell-derived microparticles (-81.4 μl 95 % CI -109.9/-57.7) after 48 h at moderate altitude was found. Microparticles derived from endothelial cells, granulocytes, monocytes and leucocytes were not significantly altered by exposure to moderate altitude. In healthy male individuals, mild hypobaric hypoxia, induced by a short-term stay at moderate altitude, is associated with lower levels of procoagulant microparticles, platelet-derived microparticles and red blood cell-derived microparticles, suggesting a reduction in thrombotic potential.


Distinct contribution of IL-6, TNF-alpha, IL-1, and IL-10 to T cell-mediated spontaneous autoimmune arthritis in mice.
Cytokines play key roles in spontaneous CD4(+) T cell-mediated chronic autoimmune arthritis in SKG mice, a new model of rheumatoid arthritis. Genetic deficiency in IL-6 completely suppressed the development of arthritis in SKG mice, irrespective of the persistence of circulating rheumatoid factor. Either IL-1 or TNF-alpha deficiency retarded the onset of arthritis and substantially reduced its incidence and severity. IL-10 deficiency, on the other hand, exacerbated disease, whereas IL-4 or IFN-gamma deficiency did not alter the disease course. Synovial fluid of arthritic SKG mice contained high amounts of IL-6, TNF-alpha, and IL-1, in accord with active transcription of these cytokine genes in the afflicted joints. Notably, immunohistochemistry revealed that distinct subsets of synovial cells produced different cytokines in the inflamed synovium: the superficial synovial lining cells mainly produced IL-1 and TNF-alpha, whereas scattered subsynovial cells produced IL-6. Thus, IL-6, IL-1, TNF-alpha, and IL-10 play distinct roles in the development of SKG arthritis; arthritogenic CD4(+) T cells are not required to skew to either Th1 or Th2; and the appearance of rheumatoid factor is independent of joint inflammation. The results also indicate that targeting not only each cytokine but also each cell population secreting distinct cytokines could be an effective treatment of rheumatoid arthritis.


99th Percentile Upper-Reference Limit of Cardiac Troponin and the Diagnosis of Acute Myocardial Infarction.
Concerns exist regarding how the 99th percentile upper reference limit (URL) of cardiac troponin (cTn) is determined and whether it should be derived from normal healthy individuals. The 99th percentile URL of cTn is an important criterion to standardize the diagnosis of myocardial infarction (MI) for clinical, research, and regulatory purposes. Statistical heterogeneity in its calculation exists but recommendations have been proposed. Some negativity has resulted from the fact that with some high-sensitivity (hs) cTn assays, a greater number of increases above the 99th percentile are observed when transitioning from a contemporary assay. Increases reflect acute or chronic myocardial injury and provide valuable diagnostic and prognostic information. The etiology of increases can sometimes be difficult to determine, making a specific treatment approach challenging. For those reasons, some advocate higher cutoff concentrations. This approach can contribute to missed diagnoses. Contrary to claims, neither clinical or laboratory guidelines have shifted away from the 99th percentile. To support the diagnosis of acute MI, the 99th percentile URL remains the best-established approach given the absence of cTn assay standardization. Importantly, risk stratification algorithms using hs-cTn assays predict the possibility of MI diagnoses established using the 99th percentile. The 99th percentile of cTn remains the best-established criterion for the diagnosis of acute MI. While not perfect, it is analytically and clinically evidence-based. Until there are robust data to suggest some other approach, staying with the 99th percentile, a threshold that has served the field well for the past 20 years, appears prudent.


Placenta-derived exosomes continuously increase in maternal circulation over the first trimester of pregnancy.
Human placenta releases specific nanovesicles (i.e. exosomes) into the maternal circulation during pregnancy, however, the presence of placenta-derived exosomes in maternal blood during early pregnancy remains to be established. The aim of this study was to characterise gestational age related changes in the concentration of placenta-derived exosomes during the first trimester of pregnancy (i.e. from 6 to 12 weeks) in plasma from women with normal pregnancies. A time-series experimental design was used to establish pregnancy-associated changes in maternal plasma exosome concentrations during the first trimester. A series of plasma were collected from normal healthy women (10 patients) at 6, 7, 8, 9, 10, 11 and 12 weeks of gestation (n = 70). We measured the stability of these vesicles by quantifying and observing their protein and miRNA contents after the freeze/thawing processes. Exosomes were isolated by differential and buoyant density centrifugation using a sucrose continuous gradient and characterised by their size distribution and morphology using the nanoparticles tracking analysis (NTA; Nanosight™) and electron microscopy (EM), respectively. The total number of exosomes and placenta-derived exosomes were determined by quantifying the immunoreactive exosomal marker, CD63 and a placenta-specific marker (Placental Alkaline Phosphatase PLAP). These nanoparticles are extraordinarily stable. There is no significant decline in their yield with the freeze/thawing processes or change in their EM morphology. NTA identified the presence of 50-150 nm spherical vesicles in maternal plasma as early as 6 weeks of pregnancy. The number of exosomes in maternal circulation increased significantly (ANOVA, p = 0.002) with the progression of pregnancy (from 6 to 12 weeks). The concentration of placenta-derived exosomes in maternal plasma (i.e. PLAP+) increased progressively with gestational age, from 6 weeks 70.6 ± 5.7 pg/ml to 12 weeks 117.5 ± 13.4 pg/ml. Regression analysis showed that weeks is a factor that explains for >70% of the observed variation in plasma exosomal PLAP concentration while the total exosome number only explains 20%. During normal healthy pregnancy, the number of exosomes present in the maternal plasma increased significantly with gestational age across the first trimester of pregnancy. This study is a baseline that provides an ideal starting point for developing early detection method for women who subsequently develop pregnancy complications, clinically detected during the second trimester. Early detection of women at risk of pregnancy complications would provide an opportunity to develop and evaluate appropriate intervention strategies to limit acute adverse sequel.


The heterochromatin-associated protein HP-1 is an essential protein in Drosophila with dosage-dependent effects on position-effect variegation.
Chromosome rearrangements which place euchromatic genes adjacent to a heterochromatic breakpoint frequently result in gene repression (position-effect variegation). This repression is thought to reflect the spreading of a heterochromatic structure into neighboring euchromatin. Two allelic dominant suppressors of position-effect variegation were found to contain mutations within the gene encoding the heterochromatin-specific chromosomal protein HP-1. The site of mutation for each allele is given: one converts Lys169 into a nonsense (ochre) codon, while the other is a frameshift after Ser10. In flies heterozygous for one of the mutant alleles (Su(var)2-504), a truncated HP-1 protein was detectable by Western blot analysis. An HP-1 minigene, consisting of HP-1 cDNA under the control of an Hsp70 heat-inducible promoter, was transduced into flies by P element-mediated germ line transformation. Heat-shock driven expression of this minigene results in elevated HP-1 protein level and enhancement of position-effect variegation. Levels of variegating gene expression thus appear to depend upon the level of expression of a heterochromatin-specific protein. The implications of these observations for mechanism of heterochromatic position effects and heterochromatin function are discussed.


The Effects of Environmental Sustainability Labels on Selection, Purchase, and Consumption of Food and Drink Products: A Systematic Review.
This review assessed the effects of environmental labels on consumers' demand for more sustainable food products. Six electronic databases were searched for experimental studies of ecolabels and food choices. We followed standard Cochrane methods and results were synthesized using vote counting. Fifty-six studies (N = 42,768 participants, 76 interventions) were included. Outcomes comprised selection (n = 14), purchase (n = 40) and consumption (n = 2). The ecolabel was presented as text (n = 36), logo (n = 13) or combination (n = 27). Message types included: organic (n = 25), environmentally sustainable (n = 27), greenhouse gas emissions (n = 17), and assorted "other" message types (n = 7). Ecolabels were tested in actual (n = 15) and hypothetical (n = 41) environments. Thirty-nine studies received an unclear or high RoB rating. Sixty comparisons favored the intervention and 16 favored control. Ecolabeling with a variety of messages and formats was associated with the selection and purchase of more sustainable food products.


GFR estimation using standardized serum cystatin C in Japan.
Recently, the Chronic Kidney Disease Epidemiology Collaboration (CKD-EPI) developed glomerular filtration rate (GFR)-estimating equations based on standardized serum cystatin C (CKD-EPI(cys)) and standardized serum creatinine plus standardized serum cystatin C (CKD-EPI(cr-cys)). We developed new GFR-estimating equations based on standardized cystatin C for a Japanese population and compared their accuracy with the CKD-EPI equations. Accuracy of diagnostic test study. 413 (development data set) and 350 individuals (validation data set). CKD-EPI(cys); CKD-EPI(cr-cys); modifications to CKD-EPI(cys) and CKD-EPI(cr-cys) using Japanese coefficients; and newly developed Japanese eGFR equations based on standardized serum cystatin C (Eq(cys)), cystatin C with a nonrenal factor reflecting hypothesized extrarenal elimination (Eq(cys+nonrenal)), and creatinine in combination with cystatin C (Eq(cr-cys)). Standardized cystatin C values were determined by a colloidal gold immunoassay traceable to the international certified reference material ERM-DA471/IFCC. Measured GFR by inulin renal clearance. In a development data set, we calculated Japanese coefficients for CKD-EPI(cys) and CKD-EPI(cr-cys) of 0.977 (95% CI, 0.853-1.002) and 0.908 (95% CI, 0.889-0.928), respectively. In a validation data set, we compared CKD-EPI(cys), Eq(cys), and Eq(cys+nonrenal) with each other. Bias and accuracy were not significantly different among the 3 equations. The precision of CKD-EPI(cys) was significantly better than for Eq(cys) (P = 0.007) and not significantly different from Eq(cys+nonrenal) (P = 0.6). We then compared 0.908 × CKD-EPI(cr-cys), Eq(cr-cys), and Eq(average) (the average value of Eq(cr) [previous Japanese equation based on standardized serum creatinine] and Eq(cys+nonrenal)) with each other in the validation data set. Bias and accuracy were not significantly different among the 3 equations. The precision of 0.908 × CKD-EPI(cr-cys) was significantly better than for Eq(cr-cys) (P = 0.004) and not significantly different from Eq(average) (P = 0.06). Limited number of participants with measured GFR >90 mL/min/1.73 m(2). Extrarenal elimination of cystatin C was not measured. CKD-EPI(cys) performed well in Japanese individuals, suggesting that equations based on serum cystatin C could be used in patients with different races without modification. Accounting for extrarenal elimination of cystatin C may improve the performance of estimating equations.


An armrest is effective for reducing hand tremble in neurosurgeons.
Experienced neurosurgeons reduce hand tremble by placing their hand beside the operative field when performing microneurosurgery conventionally. Another solution to reduce hand tremble is an armrest. However, the reduction of hand tremble by using an armrest or finger-placing technique has not been rigorously measured in microneurosurgery. This study was performed to provide a quantitative assessment of the efficacy of an armrest to reduce hand tremble in comparison with the finger-placing technique. Hand tremble was evaluated in 11 board-certified neurosurgeons in a simulated microneurosurgery. The loci of surgical forceps handled by neurosurgeons were measured by a three-dimensional optical coordinate measuring machine. A static task was performed under four conditions: with/without the finger-placing technique and/or an armrest. The radius of an imaginative sphere including 95% of each locus was calculated and reviewed according to the four conditions. Hand tremble was significantly larger when the finger-placing technique was not implemented compared to when the technique was used (P<0.05). The armrest also reduced hand tremble (P<0.05) similar to the finger-placing technique. Non-inferiority was retained between the finger-placing technique and the armrest. Concomitant use of the armrest and the finger-placing technique did not interfere with the efficacy of the technique to reduce neurosurgeon's hand tremble. The finger-placing technique was confirmed to reduce hand tremble. Resting the neurosurgeon's forearm on an armrest also reduced the hand tremble. An armrest is a device that reduces hand tremble in neurosurgeons like the finger-placing technique.


Neurochemical individuality: genetic diversity among human dopamine and serotonin receptors and transporters.
Behavioral variation in human beings encompasses wide differences in personality and susceptibility to psychiatric illness arising from both genotype and experience. Long-lasting behavioral differences generally have heritabilities of 30% or more, and such inheritance is ultimately attributable to functional variants of genes programming brain development and function. The sequencing of the human genome is revealing a pattern of gene sequence variation. The ability of sequence variants to affect neural function either alone or in concert may reveal effects of behavioral selection on the human genome over evolutionary time frames. Dopamine and serotonin are phylogenetically ancient neurotransmitters intrinsic to brain function and behavior. Dopamine and serotonin receptor and transporter genes have been an early focus for efforts to identify and functionally characterize sequence variation. The purpose of this article is to present a preview of a developing new perspective in human behavior: the genetic variation of the brain or neurochemical individuality. Arch Gen Psychiatry. 2000;57:1105-1114.


Argovit™ silver nanoparticles to fight Huanglongbing disease in Mexican limes (Citrus aurantifolia Swingle).
Nowadays, Huanglongbing (HLB) disease, commonly known as "yellow dragon disease", affects citrus crops worldwide and has a devastating effect in the agro-industrial sector. Significant efforts have been made to fight the illness, but still, there is no effective treatment to eradicate the disease. This work is the first approach to evaluate the capacity of silver nanoparticles (AgNPs) to directly eradicate the bacteria responsible for Huanglongbing disease, Candidatus Liberibacter asiaticus (CLas), in the field. The AgNPs were administered by foliar sprinkling and trunk injection of 93 sick trees with remarkable results. Both methods produce an 80-90% decrease of bacterial titre, quantified by qRT-PCR in collected foliar tissue, compared with the control group. Scanning electron microscopy images show an essential reduction of starch accumulation in phloem vessels after AgNP treatments without evidence of bacteria in the analyzed samples. Compared with other effective methods that involve β-lactam antibiotics, the potency of AgNPs is 3 to 60-times higher when it is administered by foliar sprinkling and from 75 to 750-fold higher when the administration was by trunk-injection. All these results allow us to propose this AgNP formulation as a promising alternative for the treatment of infected trees in the field.


Disruption of basal JNK activity differentially affects key fibroblast functions important for wound healing.
We used both a gene knockout approach and pharmacologic modulation to study the implication of the JNK pathway in regulating fibroblast motility, capacity to contract mechanically unloaded collagen gels, and type I collagen gene expression in vitro. These parameters, which are important for tissue repair, are positively regulated by transforming growth factor (TGF)-beta, a cytokine viewed as playing a master role during wound healing. We demonstrate that basal JNK activity is critical for fibroblast motility because (a) mouse embryo jnk-/- fibroblasts exhibit significantly lower ability to close mechanically induced cell layer wounds than their wild-type (wt) counterparts, and (b) wound closure by human dermal fibroblasts is dramatically impaired by the specific JNK inhibitor SP600125. junAA fibroblasts, in which amino acids Ser63 and Ser73 of c-Jun are replaced by two Ala residues so that c-Jun cannot be phosphorylated by JNK, also exhibited impaired motility, suggesting that c-Jun phosphorylation by JNK is critical for fibroblast migration. In sharp contrast to their lesser motility on plastic, jnk-/- and junAA fibroblasts contracted free-floating, mechanically unloaded, collagen lattices markedly faster than wt fibroblasts. Furthermore, basal mRNA steady-state levels for types I and III collagen genes were similar in jnk-/- and wt fibroblasts. Likewise, overexpression of a dominant-negative mutant form of MKK4 in dermal fibroblasts did not affect collagen expression. We also demonstrate that basal JNK activity does not affect either TGF-beta-induced collagen gene expression or lattice contraction, whereas on the other hand, the blockage of motility initiated by JNK inhibition cannot be overcome by TGF-beta. Together these results demonstrate discrete, yet significant and highly specific, regulation of fibroblast functions important for wound healing by basal JNK activity.


Influence of head positioning on the assessment of Chiari-like malformation in Cavalier King Charles spaniels.
Chiari-like malformation (CM) is almost omnipresent in the Cavalier King Charles spaniels (CKCS), often leading to syringomyelia (SM). Morphometric studies have produced variable results concerning relationship between the brain parenchyma within the caudal cranial fossa (CCF) and SM. The present study assesses the effect of head position, one potential confounder. Magnetic resonance images of CKCS with CM were reviewed in extended and flexed head positions. Volumes were calculated from transverse T2-weighted brain images. Mid-sagittal images were used for measurement of cerebellar herniation and CSF space between cerebellum and brainstem. Fourteen CKCS were included into the study, seven dogs with CM and seven with CM/SM. There was no difference between the relative brain parenchyma within the CCF in extended position and flexed position, or the brain parenchyma within the rostral and middle cranial fossae proportion. Cerebellar herniation and CSF space between cerebellum and brainstem were significantly increased in the flexed position. Cerebellar herniation and CSF space differed significantly between CM and CM/SM in a flexed head position. Volumetric measurements did not vary with head position. Cerebellar herniation and CSF space between the cerebellum and the brainstem were larger in a flexed head position.


Congenital malformations in infants born after in vitro fertilization in Sweden.
The risk for congenital malformations is increased in infants born after in vitro fertilization (IVF). Some specific malformations appear to be more affected than others. The presence of congenital malformations in 15,570 infants born after IVF with an embryo transfer between April 1, 2001, and the end of 2006 were compared with all infants born in Sweden during 2001 to 2007 (n = 689,157). Risk estimates were made after adjusting for year of birth, maternal age, parity, smoking, and body mass index. The risks of specific malformations were compared with data from a previous study (1982 to March 31, 2001) of 16,280 infants born after IVF. Different IVF methods were compared to respect to malformation risk. Increased risks of a similar magnitude were found for most cardiovascular malformations and limb reduction defects for both study periods. For neural tube defects, cardiac septal defects, and esophageal atresia, there was still an increased risk, but it was lower during the second than during the first period. For small bowel atresia, anal atresia, and hypospadias, the risk increase observed during the first study period had disappeared during the second period. An increased risk was seen for some syndromes that have been associated with imprinting errors. No difference in malformation risk according to IVF method was apparent. A slightly increased risk for congenital malformations after IVF persists. A decreasing risk is seen for some specific malformations, either true or the result of multiple testing.


Challenges and priorities for modelling livestock health and pathogens in the context of climate change.
Climate change has the potential to impair livestock health, with consequences for animal welfare, productivity, greenhouse gas emissions, and human livelihoods and health. Modelling has an important role in assessing the impacts of climate change on livestock systems and the efficacy of potential adaptation strategies, to support decision making for more efficient, resilient and sustainable production. However, a coherent set of challenges and research priorities for modelling livestock health and pathogens under climate change has not previously been available. To identify such challenges and priorities, researchers from across Europe were engaged in a horizon-scanning study, involving workshop and questionnaire based exercises and focussed literature reviews. Eighteen key challenges were identified and grouped into six categories based on subject-specific and capacity building requirements. Across a number of challenges, the need for inventories relating model types to different applications (e.g. the pathogen species, region, scale of focus and purpose to which they can be applied) was identified, in order to identify gaps in capability in relation to the impacts of climate change on animal health. The need for collaboration and learning across disciplines was highlighted in several challenges, e.g. to better understand and model complex ecological interactions between pathogens, vectors, wildlife hosts and livestock in the context of climate change. Collaboration between socio-economic and biophysical disciplines was seen as important for better engagement with stakeholders and for improved modelling of the costs and benefits of poor livestock health. The need for more comprehensive validation of empirical relationships, for harmonising terminology and measurements, and for building capacity for under-researched nations, systems and health problems indicated the importance of joined up approaches across nations. The challenges and priorities identified can help focus the development of modelling capacity and future research structures in this vital field. Well-funded networks capable of managing the long-term development of shared resources are required in order to create a cohesive modelling community equipped to tackle the complex challenges of climate change.


Herpesvirus gB-induced fusion between the virion envelope and outer nuclear membrane during virus egress is regulated by the viral US3 kinase.
Herpesvirus capsids collect along the inner surface of the nuclear envelope and bud into the perinuclear space. Enveloped virions then fuse with the outer nuclear membrane (NM). We previously showed that herpes simplex virus (HSV) glycoproteins gB and gH act in a redundant fashion to promote fusion between the virion envelope and the outer NM. HSV mutants lacking both gB and gH accumulate enveloped virions in herniations, vesicles that bulge into the nucleoplasm. Earlier studies had shown that HSV mutants lacking the viral serine/threonine kinase US3 also accumulate herniations. Here, we demonstrate that HSV gB is phosphorylated in a US3-dependent manner in HSV-infected cells, especially in a crude nuclear fraction. Moreover, US3 directly phosphorylated the gB cytoplasmic (CT) domain in in vitro assays. Deletion of gB in the context of a US3-null virus did not add substantially to defects in nuclear egress. The majority of the US3-dependent phosphorylation of gB involved the CT domain and amino acid T887, a residue present in a motif similar to that recognized by US3 in other proteins. HSV recombinants lacking gH and expressing either gB substitution mutation T887A or a gB truncated at residue 886 displayed substantial defects in nuclear egress. We concluded that phosphorylation of the gB CT domain is important for gB-mediated fusion with the outer NM. This suggested a model in which the US3 kinase is incorporated into the tegument layer (between the capsid and envelope) in HSV virions present in the perinuclear space. By this packaging, US3 might be brought close to the gB CT tail, leading to phosphorylation and triggering fusion between the virion envelope and the outer NM.