Circling behavior after narcotic drugs and during naloxone-precipitated abstinence in rats with unilateral nigral lesions.
Unilateral lesions of the substantia nigra zona compacta (SNC) in rats were produced by electrolytic coagulation or by an injection of 6-hydroxydopamine. Two to 5 weeks later, after being preselected for amphetamine-induced ipsilateral circling behavior, the animals were administered narcotic agonists or antagonists and their circling behavior was observed. Morphine, methadone, levorphanol, nalorphine or pentazocine induced ipsilateral circling movements; both naloxone and dextrorphan were without effect. Ipsilateral circling was also observed in rats with unilateral electrolytic lesions after administration of agents that are thought to enhance central dopaminergic activities: d-amphetamine, l-dopa and apomorphine. In rats with unilateral electrolytic or 6-hydroxydopamine SNC lesions that were rendered highly morphine-dependent by multiple-morphine pellet implantation, contralateral (C) circling behavior was observed within 1 to 2 minutes after a naloxone challenge; the onset and duration of C circling behavior coincided with the initial appearance and duration of precipitated-morphine withdrawal signs. C circling was also observed after administration of putative dopamine receptor blockers, haloperidol and pimozide in rats with either unilateral electrolytic or 6-hydroxydopamine SNC lesions. Morphine pretreatment diminished both the C circling intensity and the appearance of withdrawal signs observed after a naloxone challenge in morphine-dependent, SNC-lesioned rats. The naloxone-precipitated withdrawal in unilaterally lesioned morphine-dependent rats was accompanied by a 20% elevation of neostriatal dopamine in the intact side. In contrast to the effects of a chronic SNC lesion in decreasing neostriatal dopamine, a 77% increase was observed in the lesioned side 30 minutes after electrolytic coagulation. Thus, narcotic agonists and partial agonists may enhance central dopaminergic activities and naloxone-precipitated withdrawal may involve a diminution in central dopaminergic activities of the nigroneostriatal pathway.


Diabetes and endometrial cancer in the Iowa women's health study.
Diabetes has been associated with increased risk of endometrial cancer in some epidemiological studies. Body mass index (BMI) and other measures of obesity have been associated positively with both diabetes and endometrial cancer. It is not clear whether or not the association of diabetes with endometrial cancer is explained entirely by obesity. Thus, we sought to test the hypothesis that diabetes is not associated with endometrial cancer independent of obesity. We examined the association between self-reported diabetes (onset at >30 years of age) and incident endometrial cancer in a prospective cohort study of 24,664 postmenopausal women in Iowa. Over 12 years of follow-up, 346 cases occurred among the cohort at risk. Data were analyzed using proportional hazards regression models. Diabetes was analyzed as reported at baseline and as a time-dependent variable using information obtained during follow-up. After adjustment for BMI, waist:hip ratio, and other covariates, the relative risk (RR) for women with diabetes versus women without diabetes was 1.43 [95% confidence interval (CI), 0.98-2.1]. The diabetes association was confined to women in the upper two BMI quintiles (RR, 1.47; 95% CI, 0.98-2.20), but a formal test of interaction was not statistically significant. Analyses that included diabetes ascertained at baseline and at follow-up gave similar results; the diabetes-associated RR in the higher BMI strata was 1.64 (95% CI, 1.16-2.31). We conclude that after adjustment for other risk factors, diabetes is associated with a modestly increased risk for endometrial cancer among women in this cohort.


Glucosylsphingosine evokes pruritus via activation of 5-HT2A receptor and TRPV4 in sensory neurons.
Glucosylsphingosine (GS), an endogenous sphingolipid, is highly accumulated in the epidermis of patients with atopic dermatitis (AD) due to abnormal ceramide metabolism. More importantly, GS can evoke scratching behaviours. However, the precise molecular mechanism by which GS induces pruritus has been elusive. Thus, the present study aimed to elucidate the molecular signalling pathway of GS, especially at the peripheral sensory neuronal levels. Calcium imaging was used to investigate the responses of HEK293T cells or mouse dorsal root ganglion (DRG) neurons to application of GS. Scratching behaviour tests were also performed with wild-type and Trpv4 knockout mice. GS activated DRG neurons in a manner involving both the 5-HT2A receptor and TRPV4. Furthermore, GS-induced responses were significantly suppressed by various inhibitors, including ketanserin (5-HT2A receptor antagonist), YM254890 (Gαq/11 inhibitor), gallein (Gβγ complex inhibitor), U73122 (phospholipase C inhibitor), bisindolylmaleimide I (PKC inhibitor) and HC067047 (TRPV4 antagonist). Moreover, DRG neurons from Trpv4 knockout mice exhibited significantly reduced responses to GS. Additionally, GS-evoked scratching behaviours were greatly decreased by pretreatment with inhibitors of either 5-HT2A receptor or TRPV4. As expected, GS-evoked scratching behaviour was also significantly decreased in Trpv4 knockout mice. Overall, the present study provides evidence for a novel molecular signalling pathway for GS-evoked pruritus, which utilizes both 5-HT2A receptor and TRPV4 in mouse sensory neurons. Considering the high accumulation of GS in the epidermis of patients with AD, GS could be another pruritogen in patients with AD.


Nondrug interventions for treatment of hypertension.
The treatment of hypertension is no longer limited to the simple prescription of pharmaceuticals. For many patients, maximal medical therapy is insufficient to adequately treat refractory hypertension. In addition, some patients may prefer to explore therapies that do not involve drugs as an initial step. Utilizing our broadening understanding of the physiology of hypertension, new technology and interventions have been developed that allow for treatments that do not rely on medications. In addition, dietary supplements and modification, as well as herbal supplements, may be useful under the right circumstances. Lifestyle modification remains a necessary part of treatment for all patients with hypertension. This article will review the evidence behind some available nondrug interventions for the treatment of hypertension.


Epigenetic transgenerational inheritance of somatic transcriptomes and epigenetic control regions.
Environmentally induced epigenetic transgenerational inheritance of adult onset disease involves a variety of phenotypic changes, suggesting a general alteration in genome activity. Investigation of different tissue transcriptomes in male and female F3 generation vinclozolin versus control lineage rats demonstrated all tissues examined had transgenerational transcriptomes. The microarrays from 11 different tissues were compared with a gene bionetwork analysis. Although each tissue transgenerational transcriptome was unique, common cellular pathways and processes were identified between the tissues. A cluster analysis identified gene modules with coordinated gene expression and each had unique gene networks regulating tissue-specific gene expression and function. A large number of statistically significant over-represented clusters of genes were identified in the genome for both males and females. These gene clusters ranged from 2-5 megabases in size, and a number of them corresponded to the epimutations previously identified in sperm that transmit the epigenetic transgenerational inheritance of disease phenotypes. Combined observations demonstrate that all tissues derived from the epigenetically altered germ line develop transgenerational transcriptomes unique to the tissue, but common epigenetic control regions in the genome may coordinately regulate these tissue-specific transcriptomes. This systems biology approach provides insight into the molecular mechanisms involved in the epigenetic transgenerational inheritance of a variety of adult onset disease phenotypes.


Iodine deficiency in Europe and its consequences: an update.
This paper updates the information on the prevalence of the disorders induced by iodine deficiency (IDD) in Europe. Thirty-two European countries were still affected by mild to severe iodine deficiency in the late 1990s. The most severely affected countries were in Eastern Europe, including Central Asia, but Western Europe was also still affected. National surveys recently conducted in 11 of these countries show that, with the exception of the Netherlands, none has yet reached a state of iodine sufficiency, though very significant improvement in the situation has been evidenced in many of them, e.g. Poland, Bulgaria and Macedonia. The consequences of persisting iodine deficiency are goitre, hyperavidity of the thyroid for iodide (which increases the risk of thyroid irradiation in the event of a nuclear accident) and subclinical hypothyroidism during pregnancy and early infant (with a concomitant risk of minor brain damage and irreversible impairment of the neuropsychointellectual development of offspring). Access to iodised salt at the household level in European countries affected by IDD increased from 5%-10% in 1990 to 28% in 1999. This constitutes encouraging progress. However, in terms of access of iodine-deficient countries to iodised salt, Europe remains the worst region in the world, as shown by the fact that the mean figure worldwide in 1999 was 68%. In Latin America it even reached 90%. Salt iodisation has to be further implemented in Europe. Until that goal is achieved, iodine supplementation in those groups most sensitive to the effects of iodine deficiency (pregnant and lactating women and young infants) will have to be considered in the most severely affected areas.


Interaction between the C. elegans cell-death regulators CED-9 and CED-4.
Programmed cell death (apoptosis) is an evolutionarily conserved process used by multicellular organisms to eliminate cells that are not needed or are potentially detrimental to the organism. Members of the Bcl-2 family of mammalian proteins are intimately involved in the regulation of apoptosis, but, their precise mechanism of action remains unresolved. In Caenorhabditis elegans, the Bcl-2 homologue CED-9 prevents cell death by antagonizing the death-promoting activities of CED-3, a member of the Caspase family of death proteases, and of CED-4, a protein with no known mammalian homologue. Here we show that CED-9 interacts physically with CED-4. Mutations that reduce or eliminate CED-9 activity also disrupt its ability to bind CED-4, suggesting that this interaction is important for CED-9 function. Thus, CED-9 might control C. elegans cell death by binding to and regulating CED-4 activity. We propose that mammalian Bcl-2 family members might control apoptosis in a similar way through interaction and regulation of CED-4 homologues or analogues.


Formulation strategies to improve oral peptide delivery.
Delivery of peptides by the oral route greatly appeals due to commercial, patient convenience and scientific arguments. While there are over 60 injectable peptides marketed worldwide, and many more in development, most delivery strategies do not yet adequately overcome the barriers to oral delivery. Peptides are sensitive to chemical and enzymatic degradation in the intestine, and are poorly permeable across the intestinal epithelium due to sub-optimal physicochemical properties. A successful oral peptide delivery technology should protect potent peptides from presystemic degradation and improve epithelial permeation to achieve a target oral bioavailability with acceptable intra-subject variability. This review provides a comprehensive up-to-date overview of the current status of oral peptide delivery with an emphasis on patented formulations that are yielding promising clinical data.


The role of optical coherence tomography in the setting of acute myocardial infarction.
In recent years, intravascular imaging-guided percutaneous coronary intervention (PCI) has been increasing in patients with acute myocardial infarction (AMI). However, the role of optical coherence tomography (OCT) has not been established in the setting of AMI despite OCT providing superior resolution (10μm axial resolution) and facilitating assessment of baseline lesion characteristics and post-intervention evaluation of the acute result of stent implantation, including visualization of procedural dissections, malapposition, tissue prolapse, and thrombus. We provide an overview of the potential benefits of OCT-guidance in various situations of AMI.


Extracting fluorescent reporter time courses of cell lineages from high-throughput microscopy at low temporal resolution.
The extraction of fluorescence time course data is a major bottleneck in high-throughput live-cell microscopy. Here we present an extendible framework based on the open-source image analysis software ImageJ, which aims in particular at analyzing the expression of fluorescent reporters through cell divisions. The ability to track individual cell lineages is essential for the analysis of gene regulatory factors involved in the control of cell fate and identity decisions. In our approach, cell nuclei are identified using Hoechst, and a characteristic drop in Hoechst fluorescence helps to detect dividing cells. We first compare the efficiency and accuracy of different segmentation methods and then present a statistical scoring algorithm for cell tracking, which draws on the combination of various features, such as nuclear intensity, area or shape, and importantly, dynamic changes thereof. Principal component analysis is used to determine the most significant features, and a global parameter search is performed to determine the weighting of individual features. Our algorithm has been optimized to cope with large cell movements, and we were able to semi-automatically extract cell trajectories across three cell generations. Based on the MTrackJ plugin for ImageJ, we have developed tools to efficiently validate tracks and manually correct them by connecting broken trajectories and reassigning falsely connected cell positions. A gold standard consisting of two time-series with 15,000 validated positions will be released as a valuable resource for benchmarking. We demonstrate how our method can be applied to analyze fluorescence distributions generated from mouse stem cells transfected with reporter constructs containing transcriptional control elements of the Msx1 gene, a regulator of pluripotency, in mother and daughter cells. Furthermore, we show by tracking zebrafish PAC2 cells expressing FUCCI cell cycle markers, our framework can be easily adapted to different cell types and fluorescent markers.


A nano-junction of self-assembled mixed-metal-centre molecular wires on transparent conductive oxides.
The fabrication of stable, highly conductive molecular nano-junctions is one of the main research goals in the field of molecular electronics. In this paper we report on the self-assembly and functional characterisation of highly conductive molecular wires, based on mixed-metal polynuclear complexes, at the surface of a transparent conductive oxide. The adopted synthetic approach involves metal-coordination reactions on oxide surfaces, pre-functionalised with a monolayer of terpyridine moieties that are used as anchoring sites for the integration of ditopic, redox-active ruthenium-bisterpyridine molecules through iron(ii) centres. By the stepwise iteration of the iron-coordination reaction, molecular wires of the desired length can be prepared, which alternate iron and ruthenium centres in the wire backbone. The stepwise assembly of the wires at the transparent conductive oxide surface was characterised by means of UV-Vis spectroscopy and, at the nanoscale, by means of ToF-SIMS measurements. The electrical characteristics of the wires were obtained by the liquid-metal eutectic-gain nano-junction technique, with results that show good electron transport capabilities along the wires. The demonstrated feasibility of the integration of these metal-polypyridinic, redox-active, conductive wires at the surface of a transparent and conductive oxide, and the evidence for good electrical conduction indicates prospective applications in the field of nanoscale molecular optoelectronics.


Measurement properties of translated versions of neck-specific questionnaires: a systematic review.
Several disease-specific questionnaires to measure pain and disability in patients with neck pain have been translated. However, a simple translation of the original version doesn't guarantee similar measurement properties. The objective of this study is to critically appraise the quality of the translation process, cross-cultural validation and the measurement properties of translated versions of neck-specific questionnaires. Bibliographic databases were searched for articles concerning the translation or evaluation of the measurement properties of a translated version of a neck-specific questionnaire. The methodological quality of the selected studies and the results of the measurement properties were critically appraised and rated using the COSMIN checklist and criteria for measurement properties. The search strategy resulted in a total of 3641 unique hits, of which 27 articles, evaluating 6 different questionnaires in 15 different languages, were included in this study. Generally the methodological quality of the translation process is poor and none of the included studies performed a cross-cultural adaptation. A substantial amount of information regarding the measurement properties of translated versions of the different neck-specific questionnaires is lacking. Moreover, the evidence for the quality of measurement properties of the translated versions is mostly limited or assessed in studies of poor methodological quality. Until results from high quality studies are available, we advise to use the Catalan, Dutch, English, Iranian, Korean, Spanish and Turkish version of the NDI, the Chinese version of the NPQ, and the Finnish, German and Italian version of the NPDS. The Greek NDI needs cross-cultural validation and there is no methodologically sound information for the Swedish NDI. For all other languages we advise to translate the original version of the NDI.


Cytogenetic and microsatellite alterations in tumors from patients with the syndrome of myxomas, spotty skin pigmentation, and endocrine overactivity (Carney complex).
Carney complex (CC) is a familial multiple neoplasia and lentiginosis syndrome, transmitted in an autosomal dominant manner. It is the only familial form of cardiac and skin myxomas known and includes endocrine neoplasms causing Cushing's syndrome [primary pigmented nodular adrenocortical disease (PPNAD)] and acromegaly (GH-producing adenoma). The molecular defect leading to CC remains unknown, but was recently mapped to chromosome 2p16 by linkage analysis. This region has exhibited cytogenetic aberrations in atrial myxomas from patients with CC and harbors the hMSH2 and hMSH6 genes, which are involved in the preservation of microsatellite length stability of replicating human cells. In the present study, we examined 15 tumor and normal tissue specimens from 13 patients with CC [GH-producing adenoma (n = 1), adrenal tumors (PPNAD, n = 8), thyroid cancer (n = 1), normal adrenal gland (n = 1)] and 4 cultured cell lines [heart myxoma (n = 3) and eyelid myxoma (n = 1)]. Chromosome analysis was obtained by standard cytogenetic techniques. One of the myxoma cell lines and 3 PPNAD specimens contained multiple telomeric associations (tas). The normal adrenocortical tissue from a patient with PPNAD contained no apparent chromosomal anomalies, whereas the neighboring PPNAD tissue demonstrated tas. DNA was extracted from peripheral blood, tumor cell lines, and frozen or paraffin-embedded tissues and subjected to PCR amplification with primers from 64 microsatellite locations covering chromosomes 1 and 3-22 and 14 loci covering chromosome 2. The alterations detected were loss and gain of heterozygosity (LOH and GOH; 49% and 26%, respectively), deletions of both alleles (DEL; 10%), and microsatellite length instability (15%). GOH and LOH were the most frequent changes, with telomeric markers significantly over-represented (P < 0.05). Chromosomes 6, 11, 22, 10, and 19 demonstrated mostly LOH, GOH, or DEL in over 40% of the informative loci tested (73%, 59%, 47%, 46%, and 44%, respectively), whereas markers on chromosome 2 showed only microsatellite length instability (10%). The degree of genomic instability and its type were independent of tumor type (P > 0.1). We conclude that tumors and tumor cell lines from patients with CC demonstrate significant genomic, but not microsatellite length, instability. Thus, the CC gene(s) on chromosome 2p16 is different from the hMSH2 and hMSH6 genes and has dominant, rather than recessive, tumorigenic function. This gene(s) appears to be involved in the regulation of genomic stability of dividing cells, in particular the structure of telomeres in replicating chromosomes and/or the function of the mitotic apparatus.


Array compression for MRI with large coil arrays.
Arrays with large numbers of independent coil elements are becoming increasingly available as they provide increased signal-to-noise ratios (SNRs) and improved parallel imaging performance. Processing of data from a large set of independent receive channels is, however, associated with an increased memory and computational load in reconstruction. This work addresses this problem by introducing coil array compression. The method allows one to reduce the number of datasets from independent channels by combining all or partial sets in the time domain prior to image reconstruction. It is demonstrated that array compression can be very effective depending on the size of the region of interest (ROI). Based on 2D in vivo data obtained with a 32-element phased-array coil in the heart, it is shown that the number of channels can be compressed to as few as four with only 0.3% SNR loss in an ROI encompassing the heart. With twofold parallel imaging, only a 2% loss in SNR occurred using the same compression factor.


Intraflagellar transport drives flagellar surface motility.
The assembly and maintenance of all cilia and flagella require intraflagellar transport (IFT) along the axoneme. IFT has been implicated in sensory and motile ciliary functions, but the mechanisms of this relationship remain unclear. Here, we used Chlamydomonas flagellar surface motility (FSM) as a model to test whether IFT provides force for gliding of cells across solid surfaces. We show that IFT trains are coupled to flagellar membrane glycoproteins (FMGs) in a Ca(2+)-dependent manner. IFT trains transiently pause through surface adhesion of their FMG cargos, and dynein-1b motors pull the cell towards the distal tip of the axoneme. Each train is transported by at least four motors, with only one type of motor active at a time. Our results demonstrate the mechanism of Chlamydomonas gliding motility and suggest that IFT plays a major role in adhesion-induced ciliary signaling pathways. DOI:http://dx.doi.org/10.7554/eLife.00744.001.


Concerns of Family Caregivers during COVID-19: The Concerns of Caregivers and the Surprising Silver Linings.
COVID-19 has had an enormous impact on older people around the world. As family caregivers provide a good portion of the care for older people, their lives have been drastically altered by COVID-19 too. Our study is an in-depth exploration of the greatest concerns of family caregivers in the United States during COVID-19, as well as their perspectives on the benefits of caregiving during this global pandemic. We conducted in-depth interviews with a diverse sample of 52 family caregivers in the United States between May and September of 2020 over video conferencing using a semi-structured interview guide. Thematic analysis was conducted to ascertain our participants' perceptions. Caregiver's concerns were organized into six main themes, including social isolation, decline in mental health, decline in physical and cognitive functioning, keeping their family members safe from COVID-19, lack of caregiving support, and caregiving stress. The themes related to the benefits of caregiving during COVID-19 included: enjoyed the slower pace, increased time to spend together, deepened relationships, recognizing the resilience of family members, and caregiving innovations. Our in-depth study helps social workers understand the nature of caregiving stress during COVID-19, as well as the positive aspects of caregiving, even during a global pandemic.


Integrating palliative care in long-term care facilities across Europe (PACE): protocol of a cluster randomized controlled trial of the 'PACE Steps to Success' intervention in seven countries.
Several studies have highlighted the need for improvement in palliative care delivered to older people long-term care facilities. However, the available evidence on how to improve palliative care in these settings is weak, especially in Europe. We describe the protocol of the PACE trial aimed to 1) evaluate the effectiveness and cost-effectiveness of the 'PACE Steps to Success' palliative care intervention for older people in long-term care facilities, and 2) assess the implementation process and identify facilitators and barriers for implementation in different countries. We will conduct a multi-facility cluster randomised controlled trial in Belgium, Finland, Italy, the Netherlands, Poland, Switzerland and England. In total, 72 facilities will be randomized to receive the 'Pace Steps to Success intervention' or to 'care as usual'. Primary outcome at resident level: quality of dying (CAD-EOLD); and at staff level: staff knowledge of palliative care (Palliative Care Survey). resident's quality of end-of-life care, staff self-efficacy, self-perceived educational needs, and opinions on palliative care. Economic outcomes: direct costs and quality-adjusted life years (QALYs). Measurements are performed at baseline and after the intervention. For the resident-level outcomes, facilities report all deaths of residents in and outside the facilities over a previous four-month period and structured questionnaires are sent to (1) the administrator, (2) staff member most involved in care (3) treating general practitioner, and (4) a relative. For the staff-level outcomes, all staff who are working in the facilities are asked to complete a structured questionnaire. A process evaluation will run alongside the effectiveness evaluation in the intervention group using the RE-AIM framework. The lack of high quality trials in palliative care has been recognized throughout the field of palliative care research. This cross-national cluster RCT designed to evaluate the impact of the palliative care intervention for long-term care facilities 'PACE Steps to Success' in seven countries, will provide important evidence concerning the effectiveness as well as the preconditions for optimal implementation of palliative care in nursing homes, and this within different health care systems. The study is registered at www.isrctn.com - ISRCTN14741671 (FP7-HEALTH-2013-INNOVATION-1 603111) Registration date: July 30, 2015.


Mastoparan increases the intracellular free calcium concentration in two insulin-secreting cell lines by inhibition of ATP-sensitive potassium channels.
The mechanisms underlying mastoparan-induced elevation of the intracellular free calcium concentration ([Ca2+]i) were investigated in the insulin-secreting cell lines RINm5F and HIT. In both cell types, micromolar concentrations of mastoparan induced a prompt increase of [Ca2+]i, measured as an increase in fura-2 fluorescence. This response was dependent on extracellular calcium entry and was suppressed by organic calcium channel blockers; the increase of [Ca2+]i caused by high glucose concentrations or tolbutamide was not enhanced by mastoparan. These data indicate the involvement of voltage-dependent calcium channels and suggest that depolarization, rather than a direct effect on the channels, mediates the response to mastoparan. This proposition was supported by the observation that whole-cell calcium currents measured using the nystatin-permeabilized patch technique were not affected by mastoparan. Mastoparan-induced depolarization was observed using the potentiometric indicator bis-oxonol, and it was shown not to be additive with the depolarization induced by high glucose concentrations or tolbutamide. The mechanism underlying mastoparan-induced depolarization was identified in single-channel patch-clamp experiments, where it was shown that mastoparan caused closure of ATP-sensitive potassium channels [K(ATP) channels] in cell-attached and excised membrane patches. Responsiveness to mastoparan in excised patches demonstrated the membrane-delimited character of K(ATP) channel inhibition. The observation that the response persisted in the absence of exogenous GTP and in the presence of 250 microM GDP or guanosine-5'-O-(2-thio)diphosphate suggested that this effect is not mediated via enhancement of G protein activity. Partial suppression of channel activity by mastoparan did not prevent the action of tolbutamide, which fully suppressed the remaining activity in excised patches. In summary, the increase of [Ca2+]i in the insulin-secreting tumor cell lines RINm5F and HIT in response to mastoparan is mediated via G protein-independent suppression of K(ATP) channel activity, cell depolarization, and activation of voltage-dependent calcium channels.


Sequence-variant repeats of MUC1 show higher conformational flexibility, are less densely O-glycosylated and induce differential B lymphocyte responses.
The human epithelial cancer mucin MUC1 is able to break tolerance and to induce humoral immune responses in healthy subjects and in cancer patients. We recently showed that clusters of sequence-variant repeats are interspersed in the repeat domain of MUC1 at high frequency, which should contribute to the structural and immunological features of the mucin. Here we elucidated the potential effects exerted by sequence-variant repeats on their O-glycosylation. Evidence from in vitro glycosylation with polypeptide N-acetylgalactosaminyltransferases GalNAc-T1 and GalNAc-T2 in concert with mass spectrometric analyses of in vivo glycosylated MUC1 probes from transiently transfected HEK293 cells indicated reduced glycosylation densities of repeats with three concerted replacements: AHGVTSAPESRPAPGSTAPA. The Pro to Ala replacement in STAPA exerts not only proximal effects on the ppGalNAc-T2 preferred site at -3 and -4, but also more distant effects on the ppGalNAc-T1 preferred site at -15 (TSAPESRPAPGSTAPA). We also examined the conformational changes of MUC1 glycopeptides induced by the concerted DT to ES replacements and revealed a higher conformational flexibility of ES/P peptides compared to DT/P peptides. Differences in conformational flexibilities and in O-glycosylation densities could underlie the observed differential humoral responses in humans. We were able to show that the natural immunoglobulin G (IgG) responses to the repeat domain of MUC1 in sera from nonmalignant control subjects are preferentially directed to variant repeat clusters. In contrast, the IgG response in patients with adenocarcinoma shifted to higher frequencies of preferential DTR peptide binding.


IUPHAR-DB: the IUPHAR database of G protein-coupled receptors and ion channels.
The IUPHAR database (IUPHAR-DB) integrates peer-reviewed pharmacological, chemical, genetic, functional and anatomical information on the 354 nonsensory G protein-coupled receptors (GPCRs), 71 ligand-gated ion channel subunits and 141 voltage-gated-like ion channel subunits encoded by the human, rat and mouse genomes. These genes represent the targets of approximately one-third of currently approved drugs and are a major focus of drug discovery and development programs in the pharmaceutical industry. IUPHAR-DB provides a comprehensive description of the genes and their functions, with information on protein structure and interactions, ligands, expression patterns, signaling mechanisms, functional assays and biologically important receptor variants (e.g. single nucleotide polymorphisms and splice variants). In addition, the phenotypes resulting from altered gene expression (e.g. in genetically altered animals or in human genetic disorders) are described. The content of the database is peer reviewed by members of the International Union of Basic and Clinical Pharmacology Committee on Receptor Nomenclature and Drug Classification (NC-IUPHAR); the data are provided through manual curation of the primary literature by a network of over 60 subcommittees of NC-IUPHAR. Links to other bioinformatics resources, such as NCBI, Uniprot, HGNC and the rat and mouse genome databases are provided. IUPHAR-DB is freely available at http://www.iuphar-db.org.


Combining photoredox and silver catalysis for azidotrifluoromethoxylation of styrenes.
The first example of an azidotrifluoromethoxylation of styrenes has been achieved by synergistic visible-light-mediated photoredox and silver catalysis. Trifluoromethyl arylsulfonate (TFMS) and the Zhdankin reagent were used as the trifluoromethoxylation reagent and the azide source, respectively. A good functional group tolerance and mild reaction conditions of this method are applicable to late-stage azidotrifluoromethoxylation of complex small molecules. Furthermore, the mechanistic investigations indicate the single-electron transfer involved in the reaction.


The Italian Society for Pediatric Nephrology (SINePe) consensus document on the management of nephrotic syndrome in children: Part I - Diagnosis and treatment of the first episode and the first relapse.
This consensus document is aimed at providing an updated, multidisciplinary overview on the diagnosis and treatment of pediatric nephrotic syndrome (NS) at first presentation. It is the first consensus document of its kind to be produced by all the pediatric nephrology centres in Italy, in line with what is already present in other countries such as France, Germany and the USA. It is based on the current knowledge surrounding the symptomatic and steroid treatment of NS, with a view to providing the basis for a separate consensus document on the treatment of relapses. NS is one of the most common pediatric glomerular diseases, with an incidence of around 2-7 cases per 100000 children per year. Corticosteroids are the mainstay of treatment, but the optimal therapeutic regimen for managing childhood idiopathic NS is still under debate. In Italy, shared treatment guidelines were lacking and, consequently, the choice of steroid regimen was based on the clinical expertise of each individual unit. On the basis of the 2015 Cochrane systematic review, KDIGO Guidelines and more recent data from the literature, this working group, with the contribution of all the pediatric nephrology centres in Italy and on the behalf of the Italian Society of Pediatric Nephrology, has produced a shared steroid protocol that will be useful for National Health System hospitals and pediatricians. Investigations at initial presentation and the principal causes of NS to be screened are suggested. In the early phase of the disease, symptomatic treatment is also important as many severe complications can occur which are either directly related to the pathophysiology of the underlying NS or to the steroid treatment itself. To date, very few studies have been published on the prophylaxis and treatment of these early complications, while recommendations are either lacking or conflicting. This consensus provides indications for the prevention, early recognition and treatment of these complications (management of edema and hypovolemia, therapy and prophylaxis of infections and thromboembolic events). Finally, recommendations about the clinical definition of steroid resistance and its initial diagnostic management, as well as indications for renal biopsy are provided.


Junctional sequences of fetal T cell receptor beta chains have few N regions.
T cell receptors (TCRs) and immunoglobulins (Igs) derive a large fraction of their repertoire from diversity generated at the junctions of the V, D, and J coding segments. This diversity is derived both from the random deletion of nucleotides from the ends of coding regions and from the subsequent addition of nontemplated N region nucleotides. While the vast majority of TCRs and Igs from adult mice have N regions, less than 5% of both TCR-gamma/delta and Ig from fetal and neonatal mice have N regions. This study analyzed the ontogeny of junctional diversity of TCR-alpha/beta. Genomic DNA or C beta-primed cDNA was prepared from thymocytes of mice at varying stages in ontogeny, and the rearranged V beta 8 or V beta 5 sequences were amplified by polymerase chain reactions. Sequencing of the V beta-D beta-J beta junctions showed few N regions early in ontogeny, although the fraction of sequences with N regions exceeded that previously reported for Ig and for TCR-gamma/delta. N regions were found in 13% of V beta junctional sequences from day 18-19 fetal thymocytes, 33% of sequences from newborn thymocytes, 76% of sequences from day 4 postnatal thymocytes, and 88% of sequences from 5-wk-old thymocytes. In addition, nonrandom usage of the D beta and J beta segments was observed in both fetal and adult TCR sequences. While the usage of each of the six J beta 2 segments was different, the same pattern of usage was seen regardless of whether D beta 1 or D beta 2 was used, suggesting that a factor controlling the rate of usage of each J segment is intrinsic to the J gene itself. Since TCRs derive so much of their diversity from N regions, the relative paucity of N regions in fetal alpha/beta T cells would create a fetal TCR-alpha/beta repertoire that would be quite different from, and smaller than, the adult repertoire. The lack of N regions might be predicted to limit the range of affinities of TCR-MHC + peptide interactions, which may have important consequences for positive and negative selection of fetal and newborn T cells.


Modulation of THP-1 macrophage and cholesterol-loaded foam cell apolipoprotein E levels by glycosphingolipids.
Macrophages synthesize and secrete apolipoprotein E (apoE) constitutively. This process is upregulated under conditions of cholesterol loading. The response to cholesterol is antiatherogenic as it is believed to promote cholesterol efflux from the artery wall. The concentration of lactosyl ceramide (LacCer), a glycosphingolipid recently discovered to regulate cellular signaling, proliferation, and expression of adhesion molecules, is also increased in atherosclerotic tissues. Here we have investigated the effect of exogenous LacCer on macrophage apoE levels. We show that increasing macrophage LacCer levels sevenfold led to reductions in cellular and secreted apoE (15 and 30%, respectively, over a 24-h period) as determined by enzyme-linked immunosorbent assay. A similar effect was also induced by glucosyl ceramide (GlcCer) but not by ganglioside species. When macrophages were converted to cholesterol-loaded foam cells by incubation with acetylated LDL, the resulting increase in cellular apoE levels was inhibited by 26% when the cells were subsequently enriched with LacCer. After metabolic labeling of cellular glycosphingolipids with [14C]palmitate, we also discovered that high-density lipoprotein (HDL) stimulates the efflux of glycosphingolipids from foam cells. These data imply that LacCer and GlcCer may be proatherogenic due to the suppression of macrophage apoE production. Furthermore, the efflux of glycosphingolipids from macrophage foam cells to HDL could indicate a potential pathway for their removal from the artery wall and subsequent delivery to the liver.


Visual exploration pattern in hemineglect.
The analysis of eye movement parameters in visual neglect such as cumulative fixation duration, saccade amplitude, or the numbers of saccades has been used to probe attention deficits in neglect patients, since the pattern of exploratory eye movements has been taken as a strong index of attention distribution. The current overview of the literature of visual neglect has its emphasis on studies dealing with eye movement and exploration analysis. We present our own results in 15 neglect patients. The free exploration behavior was analyzed in these patients presenting 32 naturalistic color photographs of everyday scenes. Cumulative fixation duration, spatial distribution of fixations in the horizontal and vertical plane, the number and amplitude of exploratory saccades was analyzed and compared with the results of an age-matched control group. A main result of our study was that in neglect patients, fixation distribution of free exploration of natural scenes is not only influenced by the left-right bias in the horizontal direction but also by the vertical direction.


Amyloid Cardiomyopathy in Hereditary Transthyretin V30M Amyloidosis - Impact of Sex and Amyloid Fibril Composition.
Transthyretin V30M (ATTR V30M) amyloidosis is a phenotypically diverse disease with symptoms ranging from predominant neuropathy to exclusive cardiac manifestations. The aims of this study were to determine the dispersion of the two types of fibrils found in Swedish ATTR V30M patients -Type A consisting of a mixture of truncated and full length ATTR fibrils and type B fibrils consisting of full length fibrils, and to estimate the severity of cardiac dysfunction in relation to fibril composition and sex. Echocardiographic data were analysed in 107 Swedish ATTR V30M patients with their fibril composition determined as either type A or type B. Measurements of left ventricular (LV) dimensions and evaluation of systolic and diastolic function including speckle tracking derived strain were performed. Patients were grouped according to fibril type and sex. Multivariate linear regression was utilised to determine factors of significant impact on LV thickness. There was no significant difference in proportions of the two types of fibrils between men and women. In patients with type A fibrils, women had significantly lower median septal (p = 0.007) and posterior wall thicknesses (p = 0.010), lower median LV mass indexed to height (p = 0.008), and higher septal strain (p = 0.037), as compared to males. These differences were not apparent in patients with type B fibrils. Multiple linear regression analysis revealed that fibril type, sex and age all had significant impact on LV septal thickness. This study demonstrates a clear difference between sexes in the severity of amyloid heart disease in ATTR V30M amyloidosis patients. Even though type A fibrils were associated with more advanced amyloid heart disease compared to type B, women with type A fibrils generally developed less cardiac infiltration than men. The differences may explain the better outcome for liver transplanted late-onset female patients compared to males.


High performance thin-film composite forward osmosis hollow fiber membranes with macrovoid-free and highly porous structure for sustainable water production.
The development of high-performance and well-constructed thin-film composite (TFC) hollow fiber membranes for forward osmosis (FO) applications is presented in this study. The newly developed membranes consist of a functional selective polyamide layer formed by highly reproducible interfacial polymerization on a polyethersulfone (PES) hollow fiber support. Using dual-layer coextrusion technology to design and effectively control the phase inversion during membrane formation, the support was designed to possess desirable macrovoid-free and fully sponge-like morphology. Such morphology not only provides excellent membrane strength, but it has been proven to minimize internal concentration polarization in a FO process, thus leading to the water flux enhancement. The fabricated membranes exhibited relatively high water fluxes of 32-34 LMH and up to 57-65 LMH against a pure water feed using 2 M NaCl as the draw solution tested under the FO and pressure retarded osmosis (PRO) modes, respectively, while consistently maintaining relatively low salt leakages below 13 gMH for all cases. With model seawater solution as the feed, the membranes could display a high water flux up to 15-18 LMH, which is comparable to the best value reported for seawater desalination applications.


π-Extended hexadeca-substituted cobalt phthalocyanine as an active layer for organic field-effect transistors.
Due to their flexibility and solution applicability, organic field-effect transistors (OFETs) are considered prominent candidates for application in flexible and low-cost devices. A soluble phthalocyaninato cobalt(ii) complex was designed and synthesized based on a hexadeca-substitution pattern by introducing peripheral phenylethynyl groups and non-peripheral n-butoxy groups. The cobalt phthalocyanine derivative was characterized using a wide range of spectroscopic and electrochemical methods, as well as single-crystal X-ray diffraction analysis. An OFET device was fabricated using a spin-coated film of soluble 1,4,8,11,15,18,22,25-octakisbutoxy-2,3,9,10,16,17,23,24-octakis-ethynyl phenyl phthalocyaninato cobalt(ii) with a bottom-gate top-contact device configuration. The transfer and output characteristics were investigated to evaluate the charge carrier mobility. The mechanisms of the leakage current through the gate dielectric were also investigated, which revealed that the dominant leakage current mechanism is Fowler-Nordheim tunneling.


The genomic sequence of the accidental pathogen Legionella pneumophila.
We present the genomic sequence of Legionella pneumophila, the bacterial agent of Legionnaires' disease, a potentially fatal pneumonia acquired from aerosolized contaminated fresh water. The genome includes a 45-kilobase pair element that can exist in chromosomal and episomal forms, selective expansions of important gene families, genes for unexpected metabolic pathways, and previously unknown candidate virulence determinants. We highlight the genes that may account for Legionella's ability to survive in protozoa, mammalian macrophages, and inhospitable environmental niches and that may define new therapeutic targets.


Laryngeal histologic findings in infants with palatal defects with or without craniofacial malformations.
The objective of this study was to determine whether specimens from infants with palatal defects (PDs) with or without craniofacial malformations (CFMs) exhibit aberrant laryngeal histologic findings compared with specimens from normal infants. Ten laryngeal specimens from infants with PDs with or without CFMs were histologically compared with 7 laryngeal specimens defined as normal from the same collection. Both groups were similar in terms of demographics and airway manipulation. All infants were prelingual. Comparisons were made at 3 levels: supraglottic, glottic, and subglottic. Histologically, no significant differences in primary laryngeal structures were found between the PD with or without CFM group and the group defined as normal. Acquired and intubation-type injuries, such as inflammation, ulceration, capillary congestion, and scar tissue, were more prevalent and severe in the study group. The primary laryngeal histologic findings of specimens from individuals with PDs with or without CFMs do not differ substantially from those from normal individuals; however, individuals with PDs do appear to be somewhat more susceptible to intubation injury and other acquired laryngeal injury. Meticulous airway management is essential.


Equine Amniotic Microvesicles and Their Anti-Inflammatory Potential in a Tenocyte Model In Vitro.
Administration of horse amniotic mesenchymal cells (AMCs) and their conditioned medium (AMC-CM) improves the in vivo recovery of spontaneous equine tendon lesions and inhibits in vitro proliferation of peripheral blood mononuclear cells (PBMC). This process may involve microvesicles (MVs) as an integral component of cell-to-cell communication during tissue regeneration. In this study, the presence and type of MVs secreted by AMCs were investigated and the response of equine tendon cells to MVs was studied using a dose-response curve at different concentrations and times. Moreover, the ability of MVs to counteract in vitro inflammation of tendon cells induced by lipopolysaccharide was studied through the expression of some proinflammatory genes such as metallopeptidase (MPP) 1, 9, and 13 and tumor necrosis factor-α (TNFα), and expression of transforming growth factor-β (TGF-β). Lastly, the immunomodulatory potential of MVs was investigated. Results show that AMCs secrete MVs ranging in size from 100 to 200 nm. An inverse relationship between concentration and time was found in their uptake by tendon cells: the maximal uptake occurred after 72 h at a concentration of 40 × 10(6) MVs/mL. MVs induced a downregulation of MMP1, MMP9, MMP13, and TNFα expression without affecting PBMC proliferation, contrary to CM and supernatant. Our data suggest that MVs contribute to in vivo healing of tendon lesions, alongside soluble factors in AMC-CM.


Natural history of diverticular disease of the colon. A review of 521 cases.
In a survey of the natural history of 521 patients with diverticular disease of the colon half of the patients had had symptoms for less than one month on presentation at hospital, and these carried the highest morbidity and mortality. Progression of the disease was usually within segments initially involved, and extension to other regions of the colon rarely occurred. The overall prognosis of patients with total colonic involvement was similar to those with localized disease, while the morbidity and mortality associated with a recurrent attack were higher than in the initial acute episode.


Characterization of two genes for the biosynthesis of the labdane diterpene Z-abienol in tobacco (Nicotiana tabacum) glandular trichomes.
Leaves of tobacco (Nicotiana tabacum) are covered with glandular trichomes that produce sucrose esters and diterpenoids in varying quantities, depending on cultivar type. The bicyclic diterpene Z-abienol is the major labdanoid present in some oriental tobacco cultivars, where it constitutes a precursor of important flavours and aromas. We describe here the identification and characterization of two genes governing the biosynthesis of Z-abienol in N. tabacum. As for other angiosperm labdanoid diterpenes, the biosynthesis of Z-abienol proceeds in two steps. NtCPS2 encodes a class-II terpene synthase that synthesizes 8-hydroxy-copalyl diphosphate, and NtABS encodes a kaurene synthase-like (KSL) protein that uses 8-hydroxy-copalyl diphosphate to produce Z-abienol. Phylogenetic analysis indicates that NtABS belongs to a distinct clade of KSL proteins that comprises the recently identified tomato (Solanum habrochaites) santalene and bergamotene synthase. RT-PCR results show that both genes are preferentially expressed in trichomes. Moreover, microscopy of NtCPS2 promoter-GUS fusion transgenics demonstrated a high specificity of expression to trichome glandular cells. Ectopic expression of both genes, but not of either one alone, driven by a trichome-specific promoter in transgenic Nicotiana sylvestris conferred Z-abienol formation to this species, which does not normally produce it. Furthermore, sequence analysis of over 100 tobacco cultivars revealed polymorphisms in NtCPS2 that lead to a prematurely truncated protein in cultivars lacking Z-abienol, thus establishing NtCPS2 as a major gene controlling Z-abienol biosynthesis in tobacco. These results offer new perspectives for tobacco breeding and the metabolic engineering of labdanoid diterpenes, as well as for structure-function relationship studies of terpene synthases.


PATZ1 fusions define a novel molecularly distinct neuroepithelial tumor entity with a broad histological spectrum.
Large-scale molecular profiling studies in recent years have shown that central nervous system (CNS) tumors display a much greater heterogeneity in terms of molecularly distinct entities, cellular origins and genetic drivers than anticipated from histological assessment. DNA methylation profiling has emerged as a useful tool for robust tumor classification, providing new insights into these heterogeneous molecular classes. This is particularly true for rare CNS tumors with a broad morphological spectrum, which are not possible to assign as separate entities based on histological similarity alone. Here, we describe a molecularly distinct subset of predominantly pediatric CNS neoplasms (n = 60) that harbor PATZ1 fusions. The original histological diagnoses of these tumors covered a wide spectrum of tumor types and malignancy grades. While the single most common diagnosis was glioblastoma (GBM), clinical data of the PATZ1-fused tumors showed a better prognosis than typical GBM, despite frequent relapses. RNA sequencing revealed recurrent MN1:PATZ1 or EWSR1:PATZ1 fusions related to (often extensive) copy number variations on chromosome 22, where PATZ1 and the two fusion partners are located. These fusions have individually been reported in a number of glial/glioneuronal tumors, as well as extracranial sarcomas. We show here that they are more common than previously acknowledged, and together define a biologically distinct CNS tumor type with high expression of neural development markers such as PAX2, GATA2 and IGF2. Drug screening performed on the MN1:PATZ1 fusion-bearing KS-1 brain tumor cell line revealed preliminary candidates for further study. In summary, PATZ1 fusions define a molecular class of histologically polyphenotypic neuroepithelial tumors, which show an intermediate prognosis under current treatment regimens.


A requirement for thioredoxin in redox-sensitive modulation of T-cadherin expression in endothelial cells.
T-cad (T-cadherin), a glycosylphosphatidylinositol-anchored cadherin superfamily member, is expressed widely in the brain and cardiovascular system, and absent, decreased, or even increased, in cancers. Mechanisms controlling T-cad expression are poorly understood. The present study investigated transcriptional regulation of T-cad in ECs (endothelial cells). Conditions of oxidative stress (serum-deprivation or presence of H(2)O(2)) elevate T-cad mRNA and protein levels in ECs. Reporter gene analysis, using serially deleted T-cad promoter stretches ranging from -99 to -2304 bp, located the minimal promoter region of T-cad within -285 bp from the translation start site. Reporter activity in ECs transfected with the -285 bp construct increased under conditions of oxidative stress, and this was normalized by antioxidant N-acetylcysteine. An electrophoretic-mobility-shift assay revealed a specific nucleoprotein complex unique to -156 to -203 bp, which increased when nuclear extracts from oxidatively stressed ECs were used, suggesting the presence of redox-sensitive binding element(s). MS analysis of the nucleoprotein complex unique to -156 to -203 bp after streptavidin-agarose pull-down detected the presence of the redox-active protein thioredoxin. The presence of thioredoxin-1 in a nuclear extract from oxidatively stressed ECs was demonstrated after immunoprecipitation and immunoblotting. Transfection of ECs with thioredoxin-1 small interfering RNA abrogated oxidative-stress-induced up-regulation of T-cad transcripts and protein. We conclude that thioredoxin-1 is an important determinant of redox-sensitive transcriptional up-regulation of T-cad in ECs.


Genome-wide analysis of central corneal thickness in primary open-angle glaucoma cases in the NEIGHBOR and GLAUGEN consortia.
To investigate the effects of central corneal thickness (CCT)-associated variants on primary open-angle glaucoma (POAG) risk using single nucleotide polymorphisms (SNP) data from the Glaucoma Genes and Environment (GLAUGEN) and National Eye Institute (NEI) Glaucoma Human Genetics Collaboration (NEIGHBOR) consortia. A replication analysis of previously reported CCT SNPs was performed in a CCT dataset (n = 1117) and these SNPs were then tested for association with POAG using a larger POAG dataset (n = 6470). Then a CCT genome-wide association study (GWAS) was performed. Top SNPs from this analysis were selected and tested for association with POAG. cDNA libraries from fetal and adult brain and ocular tissue samples were generated and used for candidate gene expression analysis. Association with one of 20 previously published CCT SNPs was replicated: rs12447690, near the ZNF469 gene (P = 0.001; β = -5.08 μm/allele). None of these SNPs were significantly associated with POAG. In the CCT GWAS, no SNPs reached genome-wide significance. After testing 50 candidate SNPs for association with POAG, one SNP was identified, rs7481514 within the neurotrimin (NTM) gene, that was significantly associated with POAG in a low-tension subset (P = 0.00099; Odds Ratio [OR] = 1.28). Additionally, SNPs in the CNTNAP4 gene showed suggestive association with POAG (top SNP = rs1428758; P = 0.018; OR = 0.84). NTM and CNTNAP4 were shown to be expressed in ocular tissues. The results suggest previously reported CCT loci are not significantly associated with POAG susceptibility. By performing a quantitative analysis of CCT and a subsequent analysis of POAG, SNPs in two cell adhesion molecules, NTM and CNTNAP4, were identified and may increase POAG susceptibility in a subset of cases.


Social and health outcomes following upgrades to a national housing standard: a multilevel analysis of a five-wave repeated cross-sectional survey.
While existing research indicates that housing improvements are associated with health improvements, less is known about the wider social and health benefits of meeting national housing standards, as well as those of their specific constituent measures. This study evaluates the impacts of a managed housing upgrade programme through a repeated cross-sectional survey design. A five-wave repeated cross-sectional survey was conducted over a seven-year period from 2009 to 2016 (n = 2075; n = 2219; n = 2015; n = 1991; and n = 1709, respectively). The study followed a managed upgrade programme designed to meet a national social housing standard over an extended period. The data were analysed from a multilevel perspective to take account of the time-dependent nature of the observations and differences in socio-demographic composition. The installation of the majority of individual housing measures (new windows and doors; boilers; kitchens; bathrooms; electrics; loft insulation; and cavity/external wall insulation) were associated with improvements in several social (housing suitability, satisfaction, and quality; thermal comfort and household finances) and health (mental, respiratory and general health) outcomes; and analyses showed relationships between the number of measures installed and the total amount invested on the one hand and the social and health outcomes on the other. There were however a few exceptions. Most notably, the installation of cavity wall insulation was associated with poorer health outcomes, and did not lead to better social outcomes. Also, no association was found between the number of measures installed and respiratory health. The study suggests that substantial housing investments through a managed upgrade programme may result in better social and health outcomes, and that the size of the improvements are proportionate to the number of measures installed and amount invested. However, there may be risks associated with specific measures; and more attention is needed for mechanical ventilation when upgrading energy efficiency of houses through fabric work. In addition to providing new evidence regarding the wider social and health outcomes, the study provides an analytical approach to evaluate upgrade programmes that are delivered over multiple years.


Sphingosine 1-phosphate stimulates Na+/H+ exchange in thyroid FRTL-5 cells.
Sphingosine derivatives are potent mitogens in several cell types. Many mitogens activate the Na+/H+ exchange, although the interrelationships between Na+/H+ exchange and mitogenesis are unclear. The present investigation in thyroid FRTL-5 cells shows that sphingosine 1-phosphate (SPP) activates Na+/H+ exchange in a dose-dependent manner in acid-loaded cells. The effect of SPP was abolished in a Na+-free buffer and by pretreatment of the cells with ethylisopropylamiloride. SPP did not affect basal intracellular pH (pHi). SPP stimulated the release of sequestered Ca2+ and a substantial entry of Ca2+. The effect of SPP on pH(i) was abolished in cells incubated in a Ca2+-free buffer, and in cells loaded with the intracellular Ca2+ chelator 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid. Furthermore, the effect of SPP was abolished in pertussis toxin (PTX)-treated cells. PTX decreased Ca2+ entry only, without affecting the release from intracellular stores. Phosphatidic acid (PA) did not activate Na+/H+ exchange, suggesting that the effect of SPP was not mediated via activation of phospholipase D and the production of PA. Thus one mechanism of action of SPP in FRTL-5 cells appears to be to activate Na+/H+ exchange. This action is mediated via a G protein-dependent mechanism and requires an increase in intracellular free Ca2+.


Type 2 diabetes in general practice in Norway 2005-2014: moderate improvements in risk factor control but still major gaps in complication screening.
To assess the status of type 2 diabetes care in general practice and changes in the quality of care between 2005 and 2014, and to identify areas of diabetes care requiring improvement. Two cross-sectional surveys were performed that included patients with type 2 diabetes in selected areas (n=9464 in 2014, n=5463 in 2005). Quality of care was assessed based on key recommendations in national guidelines. Differences in clinical performance between 2005 and 2014 were assessed in regression models adjusting for age, sex, counties and clustering within general practices. Treatment targets were achieved in a higher proportion of patients in 2014 compared with 2005: hemoglobin A1c ≤7.0% (≤53 mmol/mol) in 62.8% vs 54.3%, blood pressure ≤135/80 mm Hg in 44.9% vs 36.6%, and total cholesterol ≤4.5 mmol/L in 49.9% vs 33.5% (all adjusted P≤0.001). Regarding screening procedures for microvascular complications, fewer patients had recorded an eye examination (61.0% vs 71.5%, adjusted P<0.001), whereas more patients underwent monofilament test (25.9% vs 18.7%, adjusted P<0.001). Testing for albuminuria remained low (30.3%) in 2014. A still high percentage were current smokers (22.7%). We found moderate improvements in risk factor control for patients with type 2 diabetes in general practice during the last decade, which are similar to improvements reported in other countries. We report major gaps in the performance of recommended screening procedures to detect microvascular complications. The proportion of daily smokers remains high. We suggest incentives to promote further improvements in diabetes care in Norway.


Reaction kinetics in living systems.
In this report we treat reaction rates, equilibrium theory, and irreversible thermodynamics as different aspects of a single discipline. In biological reactions the rate is ultimately controlled by enzymes and other proteins of complex structure and high molecular weight. The needed formalism can be placed in one-to-one correspondence with appropriate electrical and mechanical networks. An enzyme molecule has zwitter ions anchored in the polypeptide chain, which enable it to distort the substrate by electrostatic polarization. Water weakens the induced or existing polar bonds and so speeds reaction. Several biological processes, such as luminescence, catalysis, nerve excitation, and anesthesia, in which enzymatic reactions play a major part are discussed from this point of view. We also have discussed the energy consumption and coupling effect in living systems. It is likely that a small fraction of bonds can become energy rich through the process of quenching and that unsymmetrical barriers in biological systems act like transistors in making the driving forces more efficient by a valve effect.


The changing limits and incidence of malaria in Africa: 1939-2009.
Understanding the historical, temporal changes of malaria risk following control efforts in Africa provides a unique insight into what has been and might be archived towards a long-term ambition of elimination on the continent. Here, we use archived published and unpublished material combined with biological constraints on transmission accompanied by a narrative on malaria control to document the changing incidence of malaria in Africa since earliest reports pre-second World War. One result is a more informed mapped definition of the changing margins of transmission in 1939, 1959, 1979, 1999 and 2009.


Induction of Peripheral Effector CD8 T-cell Proliferation by Combination of Paclitaxel, Carboplatin, and Bevacizumab in Non-small Cell Lung Cancer Patients.
Chemotherapy has long been the standard treatment for advanced stage non-small cell lung cancer (NSCLC), but checkpoint inhibitors are now approved for use in several patient groups and combinations. To design optimal combination strategies, a better understanding of the immune-modulatory capacities of conventional treatments is needed. Therefore, we investigated the immune-modulatory effects of paclitaxel/carboplatin/bevacizumab (PCB), focusing on the immune populations associated with the response to checkpoint inhibitors in peripheral blood. A total of 223 patients with stage IV NSCLC, enrolled in the NVALT12 study, received PCB, with or without nitroglycerin patch. Peripheral blood was collected at baseline and after the first and second treatment cycle, proportions of T cells, B cells, and monocytes were determined by flow cytometry. Furthermore, several subsets of T cells and the expression of Ki67 and coinhibitory receptors on these subsets were determined. Although proliferation of CD4 T cells remained stable following treatment, proliferation of peripheral blood CD8 T cells was significantly increased, particularly in the effector memory and CD45RA+ effector subsets. The proliferating CD8 T cells more highly expressed programmed death receptor (PD)-1 and cytotoxic T-lymphocyte-associated antigen-4 (CTLA-4) compared with nonproliferating CD8 T cells. Immunologic responders (iR; >2 fold increased proliferation after treatment) did not show an improved progression-free (PFS) or overall survival (OS). Paclitaxel/carboplatin/bevacizumab induces proliferation of CD8 T cells, consisting of effector cells expressing coinhibitory checkpoint molecules. Induction of proliferation was not correlated to clinical outcome in the current clinical setting. Our findings provide a rationale for combining PCB with checkpoint inhibition in lung cancer.


Systematic review of topic treatment for venous ulcers.
Venous ulcer patients can experience this situation for several years without achieving healing if treatment is inadequate. Evidence-based professional practice generates effective results for patients and services. This research aimed to carry out a systematic review to assess the most effective method to improve venous return and the best topic treatment for these ulcers. Studies were collected in eight databases, using the following descriptors: leg ulcer, venous ulcer and similar terms. The sample consisted of 33 primary studies and two Meta-analyses. A wide range of treatments was identified, grouped in compression therapy (54.3%) and topical treatment (45.7%). It was evidenced that compression therapy increases ulcer healing rates and should be used in patients with intact arteries. There is no consensus about the best topical treatment, although different options should be associated with compression therapy.


Human Urine Extract Cell Differentiation Agent 2 Protects PC12 Cells from Serum Deprivation-Induced Apoptosis Accompanied with Priming of Extracellular Signal-Regulated Kinase Activation and Differentiation Induction.
To investigate the potential neuroprotective effect of human urine extract cell differentiation agent 2 (CDA-2) by the model of serum deprivation-induced apoptosis of PC12 cells and study the underlying molecular mechanisms. Apoptosis of PC12 cells was induced by serum deprivation. CDA-2 at doses of 0.5-4 mg/mL was used to treat the serum-deprived PC12 cells. The cellular viability was measured by sulforhodamine B binding assay and the cell apoptosis was determined by flow cytometer. Western blot was used to analyze the expression of differentiation markers and activity of extracellular signal-regulated kinase (ERK). The cellular morphology was examined under an inverted microscope. CDA-2 inhibited apoptotic cell death of serum-deprived PC12 cells in a dose-dependent manner. Expression of low- and mid-sized neurofilaments was observed in serum-deprived PC12 cells treated with CDA-2 or nerve growth factor (NGF). However, CDA-2 did not induce proliferation of these cells like NGF. The morphology of CDA-2 treated cells was changed from rounded to neuron-like flat polygonal shape in contrast to the extensive neurite outgrowth induced by NGF. CDA-2 transiently induced the phosphorylation of ERK in serum deprived-PC12 cells and the expression of neurofilaments induced by CDA-2 was attenuated by mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) inhibitor PD98059. Human urine extract CDA-2 showed a potential neuroprotective activity which may correlate with ERK activation and differentiation induction.


"They don't have to suffer for me": why dialysis patients refuse offers of living donor kidneys.
In the present climate of organ shortage, high demand for kidney transplants, and better clinical outcomes from living donors, health care professionals expect and encourage patients to accept offers of living related donor (LRD) kidneys. When patients decide not to adhere to this course of treatment, scholars and policy makers may question why, given that it delays their chances of receiving a transplant. This article reports on patients' decisions to refuse LRD kidney offers. An 18-month study was conducted of treatment decisions by hemodialysis patients (N = 79). Fewer than half of those offered an LRD kidney (N = 64) were willing to accept it, suggesting that sociocultural factors inform decisions. Patients expressed concerns about the potential donors' well-being over their own and about compromising their relationships with the donors. This article concludes that social relations, emotions, and ethnomedical beliefs play an important role in patients' treatment decisions and thus contributes to the anthropology of decision making.


NOS inhibition enhances myogenic tone by increasing rho-kinase mediated Ca2+ sensitivity in the male but not the female gerbil spiral modiolar artery.
Cochlear blood flow regulation is important to prevent hearing loss caused by ischemia and oxidative stress. Cochlear blood supply is provided by the spiral modiolar artery (SMA). The myogenic tone of the SMA is enhanced by the nitric oxide synthase (NOS) blocker L-N(G)-nitro-arginine (LNNA) in males, but not in females. Here, we investigated whether this gender difference is based on differences in the cytosolic Ca(2+) concentration and/or the Ca(2+) sensitivity of the myofilaments. Vascular diameter, myogenic tone, cytosolic Ca(2+), and Ca(2+) sensitivity were evaluated in pressurized SMA segments isolated from male and female gerbils using laser-scanning microscopy and microfluorometry. The gender difference of the LNNA-induced tone was compared, in the same vessel segments, to tone induced by 150 mM K(+) and endothelin-1, neither of which showed an apparent gender-difference. Interestingly, LNNA-induced tone in male SMAs was observed in protocols that included changes in intramural pressure, but not when the intramural pressure was held constant. LNNA in male SMAs did not increase the global Ca(2+) concentration in smooth muscle cells but increased the Ca(2+) sensitivity. This increase in the Ca(2+) sensitivity was abolished in the presence of the guanylyl cyclase inhibitor ODQ or by extrinsic application of either the nitric oxide (NO)-donor DEA-NONOate or the cGMP analog 8-pCPT-cGMP. The rho-kinase blocker Y27632 decreased the basal Ca(2+) sensitivity and abolished the LNNA-induced increase in Ca(2+) sensitivity in male SMAs. Neither LNNA nor Y27632 changed the Ca(2+) sensitivity in female SMAs. The data suggest that the gender difference in LNNA-induced tone is based on a gender difference in the regulation of rho-kinase mediated Ca(2+) sensitivity. Rho-kinase and NO thus emerge as critical factors in the regulation of cochlear blood flow. The larger role of NO-dependent mechanisms in male SMAs predicts greater restrictions on cochlear blood flow under conditions of impaired endothelial cell function.


Dynamical view of membrane binding and complex formation of human factor VIIa and tissue factor.
The molecular mechanism of enhancement of the enzymatic activity of factor VIIa by tissue factor (TF) is not fully understood, primarily because of the lack of atomic models for the membrane-bound form of the TF-FVIIa complex. To construct the first membrane-bound model of the TF-FVIIa complex, and to investigate the dynamics of the complex in solution and on the surface of anionic membranes by using large-scale molecular dynamics (MD) simulations in full atomic detail. Membrane-bound models of the TF-FVIIa complex and the individual factors were constructed and subjected to MD simulations, in order to characterize protein-protein and protein-lipid interactions, and to investigate the dynamics of TF and FVIIa. The MD trajectories reveal that isolated FVIIa undergoes large structural fluctuation, primarily due to the hinge motions between its domains, whereas soluble TF (sTF) is structurally stable. Upon complex formation, sTF restricts the motion of FVIIa significantly. The results also show that, in the membrane-bound form, sTF directly interacts with the lipid headgroups, even in the absence of FVIIa. The first atomic models of membrane-bound sTF-FVIIa, FVIIa and sTF are presented, revealing that sTF forms direct contacts with the lipids, both in the isolated form and in complex with FVIIa. The main effect of sTF binding to FVIIa is spatial stabilization of the catalytic site of FVIIa, which ensures optimal interaction with the substrate, FX.


Desmoplastic melanoma: a pathologically and clinically distinct form of cutaneous melanoma.
Desmoplastic melanoma (DM) is a rare variant characterized by the presence of fusiform melanocytes in a sclerotic stroma. Pathologic heterogeneity within DM may account for the controversy regarding the clinical presentation and prognosis of DM compared with conventional melanoma (CM). We identified 131 patients with a diagnosis of DM seen between 1979 and 2002. Tumors were categorized as either pure DM (pDM; n = 92), if desmoplasia was prominent throughout the entire invasive tumor, or mixed DM (mDM; n = 39), if fibrosis was well developed in only parts of an otherwise non-DM. Differences in clinical behavior among pDM, mDM, and CM (n = 3976) were examined. Seventy-three percent of patients with DM had tumors >2 mm in depth, compared with 31% of patients with CM (P < .001). Regional nodal metastasis was uncommon in patients who presented with clinically localized pDM (1%) compared with those with mDM (10%) or CM (6%) (P < .05, pDM vs. CM). Five-year melanoma-specific mortality was lower for patients who presented with pDM compared with mDM (11% vs. 31%; P < .01). Patients with pDM and CM had a similar melanoma-specific mortality despite a 3-fold difference in median tumor depth (3.6 vs. 1.2 mm, respectively). DMs can be divided into two subtypes based on a histological quantification of desmoplasia. Tumors with prominent fibrosis (pure subtype) are unlikely to disseminate to regional lymph nodes and are associated with a favorable outcome when compared with those with mixed desmoplasia or CM.


The three-dimensional genome organization of Drosophila melanogaster through data integration.
Genome structures are dynamic and non-randomly organized in the nucleus of higher eukaryotes. To maximize the accuracy and coverage of three-dimensional genome structural models, it is important to integrate all available sources of experimental information about a genome's organization. It remains a major challenge to integrate such data from various complementary experimental methods. Here, we present an approach for data integration to determine a population of complete three-dimensional genome structures that are statistically consistent with data from both genome-wide chromosome conformation capture (Hi-C) and lamina-DamID experiments. Our structures resolve the genome at the resolution of topological domains, and reproduce simultaneously both sets of experimental data. Importantly, this data deconvolution framework allows for structural heterogeneity between cells, and hence accounts for the expected plasticity of genome structures. As a case study we choose Drosophila melanogaster embryonic cells, for which both data types are available. Our three-dimensional genome structures have strong predictive power for structural features not directly visible in the initial data sets, and reproduce experimental hallmarks of the D. melanogaster genome organization from independent and our own imaging experiments. Also they reveal a number of new insights about genome organization and its functional relevance, including the preferred locations of heterochromatic satellites of different chromosomes, and observations about homologous pairing that cannot be directly observed in the original Hi-C or lamina-DamID data. Our approach allows systematic integration of Hi-C and lamina-DamID data for complete three-dimensional genome structure calculation, while also explicitly considering genome structural variability.


Effect of Cytomegalovirus (CMV) and Ageing on T-Bet and Eomes Expression on T-Cell Subsets.
The differential impact of ageing and cytomegalovirus (CMV) latent infection on human T-cell subsets remains to some extent controversial. The purpose of this study was to analyse the expression of the transcription factors T-bet and Eomes and CD57 on CD4+, CD4hiCD8lo and CD8+ T-cell subsets in healthy individuals, stratified by age and CMV serostatus. The percentage of CD4+ T-cells expressing T-bet or Eomes was very low, in particular in CD4+ T-cells from young CMV-seronegative individuals, and were higher in CMV-seropositive older individuals, in both CD57- and CD57+ CD4+ T-cells. The study of the minor peripheral blood double-positive CD4hiCD8lo T-cells showed that the percentage of these T-cells expressing both Eomes and T-bet was higher compared to CD4+ T-cells. The percentage of CD4hiCD8lo T-cells expressing T-bet was also associated with CMV seropositivity and the coexpression of Eomes, T-bet and CD57 on CD4hiCD8lo T-cells was only observed in CMV-seropositive donors, supporting the hypothesis that these cells are mature effector memory cells. The percentage of T-cells expressing Eomes and T-bet was higher in CD8+ T-cells than in CD4+ T-cells. The percentages of CD8+ T-cells expressing Eomes and T-bet increased with age in CMV-seronegative and -seropositive individuals and the percentages of CD57- CD8+ and CD57+ CD8+ T-cells coexpressing both transcription factors were similar in the different groups studied. These results support that CMV chronic infection and/or ageing are associated to the expansion of highly differentiated CD4+, CD4hiCD8lo and CD8+ T-cells that differentially express T-bet and Eomes suggesting that the expression of these transcription factors is essential for the generation and development of an effector-memory and effector T lymphocytes involved in conferring protection against chronic CMV infection.


Welfare reform and the changing landscape of substance abuse services for low-income women.
Recent welfare reform policies could fundamentally change the nature of public-sector substance abuse services available to women. This review summarizes what is presently known about substance abuse services and women on welfare, and identifies limitations in our current knowledge about the potential effects of welfare reform. Five crucial areas are examined in which research on services has fallen short: (1) assessing the need for substance abuse services across a broad spectrum of welfare populations, (2) exploring the role that alcohol and drug problems play in welfare dependency, (3) examining how welfare programs can serve as pathways to alcohol and drug treatment, (4) evaluating the effectiveness and costs of innovative welfare-treatment programs, and (5) understanding systems-level adaptations in substance abuse services for women that result from changing welfare policies. We conclude that researchers who study services should take a broad view of these issues-one that considers the unique situation of poor women and single mothers, that views substance abuse within a work impairment or disabilities framework, and one that is attuned to future changes in the effects of welfare reform as the economy and labor markets undergo change.


Oral ganciclovir systemic exposure is enhanced in HIV-infected patients with diarrhea and weight loss.
To determine whether diarrhea and intestinal malabsorption during HIV infection alter oral ganciclovir systemic exposure. We studied the oral disposition of ganciclovir in 42 HIV-infected patients stratified into three groups: A (n = 15), HIV (stage A and B); B (n = 13), AIDS (stage C); and C (n = 14), AIDS with chronic diarrhea and wasting syndrome (10% or more weight loss). Each patient was evaluated for nutritional (body mass index, serum albumin and transferrin), immunologic (CD4 count, plasma viral load) and intestinal status (D-xylose test, fecal fat and nitrogen excretion, and intestinal permeability). Following an overnight fast, 1 g oral ganciclovir was given to patients. Six blood samples were collected over 24 hours. Serum was analyzed for ganciclovir by high performance liquid chromatography. Drug disposition was characterized using a population pharmacokinetic approach. Mean intestinal permeability increased as HIV disease progressed (0. 05, 0.1, and 0.2 for groups A, B, and C, respectively). Average weight-adjusted maximum concentration (Cmax) in group C was twofold more than that in group A and B patients (12.5 versus 6 and 6.4 ng/ml/kg), and average area under the curve (AUC0-infinity) was threefold greater in group C patients (193 versus 59 and 65 ng. hour/ml/kg in groups A and B, respectively). Mean oral clearance was threefold lower in group C (96 versus 258 and 212 L/hour in groups A and B, respectively). Because systemic exposure of oral ganciclovir is enhanced in AIDS patients with diarrhea and wasting syndrome, oral ganciclovir therapy may benefit these patients.


Long-term study of high-comorbidity ESRD patients converted from conventional to short daily hemodialysis.
Conventional hemodialysis (CHD) is associated with suboptimal clinical outcomes and high mortality rates. Daily hemodialysis (DHD) has been reported to improve outcomes and quality of life (QOL), predominantly in self-care or home dialysis populations. The effect of short DHD (sDHD) on patients with end-stage renal disease (ESRD) with high comorbidities has not been established. This prospective study compared clinical outcomes and QOL in high-comorbidity patients with ESRD converted from CHD to sDHD while maintaining the same total weekly dialysis time. Study patients had 4.0 +/- 1.7 major comorbid conditions in addition to ESRD. Standard dialysis parameters, antihypertensive and erythropoietin (EPO) requirements, Kidney Disease Quality of Life (KDQOL) measurements, vascular access problems, and hospitalization rates were compared while on sDHD therapy versus the previous 12 months on CHD therapy. Forty-two patients were studied on sDHD therapy for 793 patient-months during a 72-month period. During sDHD, standard Kt/V increased 31%, hospitalization days decreased significantly by 34%, and vascular access problems did not increase. Cumulative survival was 33% at 6 years. In the 20 patients who remained on sDHD therapy for 12 months, after 1 year, we found significant improvements in KDQOL scores, a 69% reduction in antihypertensive medications with stable blood pressure, and a 45% reduction in EPO requirements with stable hematocrits. We hypothesize that these improvements are the result of the less extreme solute and fluid fluctuations and greater dialysis dose provided by sDHD, even when weekly dialysis time is unchanged. High-comorbidity patients with ESRD converted to sDHD therapy had significantly improved clinical outcomes and QOL and decreased hospitalizations, with no increase in vascular access problems.


Effectiveness of St John's wort in major depression: a randomized controlled trial.
Extracts of St John's wort are widely used to treat depression. Although more than 2 dozen clinical trials have been conducted with St John's wort, most have significant flaws in design and do not enable meaningful interpretation. To compare the efficacy and safety of a standardized extract of St John's wort with placebo in outpatients with major depression. Randomized, double-blind, placebo-controlled clinical trial conducted between November 1998 and January 2000 in 11 academic medical centers in the United States. Two hundred adult outpatients (mean age, 42.4 years; 67.0% female; 85.9% white) diagnosed as having major depression and having a baseline Hamilton Rating Scale for Depression (HAM-D) score of at least 20. Participants completed a 1-week, single-blind run-in of placebo, then were randomly assigned to receive either St John's wort extract (n = 98; 900 mg/d for 4 weeks, increased to 1200 mg/d in the absence of an adequate response thereafter) or placebo (n = 102) for 8 weeks. The primary outcome measure was rate of change on the HAM-D over the treatment period. Secondary measures included the Beck Depression Inventory (BDI), Hamilton Rating Scale for Anxiety (HAM-A), the Global Assessment of Function (GAF) scale, and the Clinical Global Impression-Severity and -Improvement scales (CGI-S and CGI-I). The random coefficient analyses for the HAM-D, HAM-A, CGI-S, and CGI-I all showed significant effects for time but not for treatment or time-by-treatment interaction (for HAM-D scores, P<.001, P =.16, and P =.58, respectively). Analysis of covariance showed nonsignificant effects for BDI and GAF scores. The proportion of participants achieving an a priori definition of response did not differ between groups. The number reaching remission of illness was significantly higher with St John's wort than with placebo (P =.02), but the rates were very low in the full intention-to-treat analysis (14/98 [14.3%] vs 5/102 [4.9%], respectively). St John's wort was safe and well tolerated. Headache was the only adverse event that occurred with greater frequency with St John's wort than placebo (39/95 [41%] vs 25/100 [25%], respectively). In this study, St John's wort was not effective for treatment of major depression.


Antibody response to human immunodeficiency virus in homosexual men. Relation of antibody specificity, titer, and isotype to clinical status, severity of immunodeficiency, and disease progression.
The titers and isotypes of antibodies to specific proteins of the human immunodeficiency virus were determined by Western blot analysis of sera from 107 homosexual men. Antibody titers were generally lower in sera from patients with the acquired immunodeficiency syndrome (AIDS) and in sera from men whose condition subsequently progressed to AIDS than in sera from men who had not progressed to AIDS. We found no evidence of isotypic prominence or restriction of the antibody response. In multivariate analysis, lower levels of CD4 helper cells were most highly associated with progression to AIDS. Lower antibody titers to the envelope protein gp110, the core protein p24, and the reverse transcriptase enzyme p51/65 were also predictive of progression to AIDS independent of their association with CD4 cell levels. These data suggest that differences in antibody levels are not simply a consequence of severe immunodeficiency but may be markers for control of infection.


Analysis of factors contributing to variation in the C57BL/6J fecal microbiota across German animal facilities.
The intestinal microbiota is involved in many physiological processes and it is increasingly recognized that differences in community composition can influence the outcome of a variety of murine models used in biomedical research. In an effort to describe and account for the variation in intestinal microbiota composition across the animal facilities of participating members of the DFG Priority Program 1656 "Intestinal Microbiota", we performed a survey of C57BL/6J mice from 21 different mouse rooms/facilities located at 13 different institutions across Germany. Fresh feces was sampled from five mice per room/facility using standardized procedures, followed by extraction and 16S rRNA gene profiling (V1-V2 region, Illumina MiSeq) at both the DNA and RNA (reverse transcribed to cDNA) level. In order to determine the variables contributing to bacterial community differences, we collected detailed questionnaires of animal husbandry practices and incorporated this information into our analyses. We identified considerable variation in a number of descriptive aspects including the proportions of major phyla, alpha- and beta diversity, all of which displayed significant associations to specific aspects of husbandry. Salient findings include a reduction in alpha diversity with the use of irradiated chow, an increase in inter-individual variability (beta diversity) with respect to barrier access and open cages and an increase in bacterial community divergence with time since importing from a vendor. We further observe a high degree of facility-level individuality, which is likely due to each facility harboring its own unique combination of multiple varying attributes of animal husbandry. While it is important to account and control for such differences between facilities, the documentation of such diversity may also serve as a valuable future resource for investigating the origins of microbial-driven host phenotypes.


Maximal expiratory pressure and Valsalva manoeuvre do not produce similar cardiovascular responses in healthy men.
What is the central question of this study? This is the first study to evaluate and describe the cardiovascular responses during maximal expiratory pressure compared with the Valsalva manoeuvre, and whether those responses are similar. What is the main finding and its importance? This study showed that the duration of the manoeuvres appears to be responsible for the different physiological mechanisms involved in the cardiovascular responses to each manoeuvre and that the intensity of expiratory effort was related to the response in maximal expiratory pressure. These results are important to identify the risks to which subjects are exposed when performing these manoeuvres. The main purpose of this study was to compare the cardiovascular responses between the Valsalva manoeuvre (VM) and maximal expiratory pressure (MEP) and to evaluate the effect of age on these responses. Twenty-eight healthy men were evaluated and divided into two groups, younger (n = 15, 25 ± 5 years) and middle aged (n = 13, 50 ± 5 years), and they performed the VM and MEP measurement. The VM consisted of an expiratory effort (40 mmHg) against a manometer for 15 s, and the MEP was performed according to American Thoracic Society guidelines. The cardiovascular responses were analysed at rest, isotime (3 s), peak, nadir and recovery, and the cardiovascular variations (Δ) were calculated as peak or isotime minus resting values. For the statistical analysis, we used two-way ANOVA (P < 0.05). We observed that MEP and the VM generate similar changes in cardiac output (P > 0.05), but MEP presents higher values for mean arterial pressure (MAPPeak , MAPIsotime , ΔMAP and ΔMAPIsotime ) than those observed in the VM (P < 0.05). The execution time of the manoeuvres (VM ∼15 s and MEP ∼5 s) appears to be largely responsible for the activation of different physiological mechanisms involved in the cardiovascular control for each manoeuvre, and the intensity of expiratory effort is related to the higher response of MAP and peripheral vascular resistance (PVRIsotime and ΔPVRIsotime ) during MEP (P < 0.05). Moreover, it appears that age affects only the heart rate and PVR responses (P < 0.05), which were higher in the young and middle-aged group, respectively. Based on these findings, we can conclude that MEP and the VM do not generate similar cardiovascular responses, except for cardiac output.


A prospective study of macrophage inhibitory cytokine-1 (MIC-1/GDF15) and risk of colorectal cancer.
Chronic inflammation plays a role in the development of colorectal cancer (CRC). The novel plasma inflammatory biomarker macrophage inhibitory cytokine-1 (MIC-1, GDF15) may have a direct mechanistic role in colorectal carcinogenesis. We conducted a prospective, nested, case-control study of incident CRC among men and women who provided a prediagnostic blood specimen. We used an enzyme-linked immunosorbent assay to measure MIC-1 and examined associations between quintiles of MIC-1 and CRC using logistic regression adjusted for matching factors (age and date of blood draw), risk factors, and other plasma inflammatory markers. We also assessed the relationship between MIC-1 levels and prostaglandin-endoperoxide synthase 2 (PTGS2)/cyclooxygenase-2 (COX-2) enzyme status in tumors with available tissue for analysis. All statistical tests were two-sided. Compared with men and women within the lowest quintile of plasma MIC-1, the multivariable relative risk (RR) for CRC was 1.93 (95% confidence interval [CI] = 1.27 to 2.94) for the highest quintile (P linear trend = .004). In an exploratory analysis, we found that among individuals with high plasma MIC-1 levels (quintiles 2-5), compared with nonuse, regular use of aspirin and nonsteroidal anti-inflammatory drugs (NSAIDs) was associated with a lower risk of PTGS2-positive CRC (multivariable RR = 0.60; 95% confidence interval = 0.41 to 0.88) but not PTGS2-negative CRC (multivariable RR = 1.21; 95% CI = 0.71 to 2.07). In contrast, among individuals with low MIC-1 levels (quintile 1), aspirin and NSAID use was not associated with a lower risk of PTGS2-positive CRC (multivariable RR = 0.57; 95% CI = 0.21 to 1.54) or PTGS2-negative CRC (multivariable RR = 1.41; 95% CI = 0.47 to 4.23). Our results support an association between higher levels of circulating MIC-1 (GDF15) and CRC. Aspirin/NSAID use appeared to lower risk of PTGS2-positive cancers, particularly among individuals with high levels of circulating MIC-1.


Cell-Size-Dependent Transcription of FLC and Its Antisense Long Non-coding RNA COOLAIR Explain Cell-to-Cell Expression Variation.
Single-cell quantification of transcription kinetics and variability promotes a mechanistic understanding of gene regulation. Here, using single-molecule RNA fluorescence in situ hybridization and mathematical modeling, we dissect cellular RNA dynamics for Arabidopsis FLOWERING LOCUS C (FLC). FLC expression quantitatively determines flowering time and is regulated by antisense (COOLAIR) transcription. In cells without observable COOLAIR expression, we quantify FLC transcription initiation, elongation, intron processing, and lariat degradation, as well as mRNA release from the locus and degradation. In these heterogeneously sized cells, FLC mRNA number increases linearly with cell size, resulting in a large cell-to-cell variability in transcript level. This variation is accounted for by cell-size-dependent, Poissonian FLC mRNA production, but not by large transcriptional bursts. In COOLAIR-expressing cells, however, antisense transcription increases with cell size and contributes to FLC transcription decreasing with cell size. Our analysis therefore reveals an unexpected role for antisense transcription in modulating the scaling of transcription with cell size.


Rare occurrence of central diabetes insipidus with dermatomyositis in a young male.
Central diabetes insipidus (CDI) and several endocrine disorders previously classified as idiopathic are now considered to be of an autoimmune etiology. Dermatomyositis (DM), a rare autoimmune condition characterized by inflammatory myopathy and skin rashes, is also known to affect the gastrointestinal, pulmonary, and rarely the cardiac systems and the joints. The association of CDI and DM is extremely rare. After an extensive literature search and to the best of our knowledge this is the first reported case in literature, we report the case of a 36-year-old male with a history of CDI, who presented to the hospital's endocrine outpatient clinic for evaluation of a 3-week history of progressive facial rash accompanied by weakness and aching of the muscles. Accurate biochemical diagnosis should always be followed by etiological investigation. This clinical entity usually constitutes a therapeutic challenge, often requiring a multidisciplinary approach for optimal outcome. Dermatomyositis is an important differential diagnosis in patients presenting with proximal muscle weakness. Associated autoimmune conditions should be considered while evaluating patients with dermatomyositis. Dermatomyositis can relapse at any stage, even following a very long period of remission. Maintenance immunosuppressive therapy should be carefully considered in these patients.