Engineering of a functional human NADH-dependent cytochrome P450 system.
A functional human NADH-dependent cytochrome P450 system has been developed by altering the cofactor preference of human NADPH cytochrome P450 reductase (CPR), the redox partner for P450s. This has been achieved by a single amino acid change of the conserved aromatic amino acid Trp-676, which covers the re-side of the FAD isoalloxazine ring in the nicotinamide-binding site. Of the mutations made, the substitution of Trp-676 with alanine (W676A) resulted in a functional NADH-dependent enzyme, which catalyzed the reduction of cytochrome c and ferricyanide as well as facilitated the metabolism of 7-ethoxyresorufin by CYP1A2. Kinetic analysis measuring cytochrome c activity revealed that the NADH-dependent k(cat) of W676A is equivalent (90%) to the NADPH-dependent k(cat) of the wild-type enzyme, with W676A having an approximately 1,000-fold higher specificity for NADH. The apparent K(M)(NADPH) and K(M)(NADH) values of W676A are 80- and 150-fold decreased, respectively. In accordance with structural data, which show a bipartite binding mode of NADPH, substitution of Trp-676 does not affect 2'-AMP binding as seen by the inhibition of both wild-type CPR and the W676A mutant. Furthermore, NADPH was a potent inhibitor of the W676A NADH-dependent cytochrome c reduction and CYP1A2 activity. Overall, the results show that Trp-676 of human CPR plays a major role in cofactor discrimination, and substitution of this conserved aromatic residue with alanine results in an efficient NADH-dependent cytochrome P450 system.


Association between multilayer left ventricular rotational mechanics and the development of left ventricular remodeling after acute myocardial infarction.
The identification of patients at risk for developing left ventricular (LV) remodeling after acute myocardial infarction (AMI) has crucial prognostic implications. The aims of this study were (1) to investigate the relationship between peak subepicardial and subendocardial twist and infarct transmurality, as assessed using contrast-enhanced magnetic resonance imaging, and (2) to evaluate the association between peak subepicardial and subendocardial twist and LV remodeling 6 months after AMI. A total of 213 patients with ST-segment elevation AMIs who underwent three-dimensional echocardiography for LV volumes and functional assessment and two-dimensional speckle-tracking analysis for the evaluation of LV twist (subendocardial vs subepicardial) were retrospectively included. A subgroup of 40 patients underwent magnetic resonance imaging within 2 months for infarct size quantification. Peak subepicardial twist was strongly related to infarct size (number of segments with transmural scar: r(2) = 0.526, P < .001; total scar score: r(2) = 0.515, P < .001) compared with peak subendocardial twist (number of segments with transmural scar: r(2) = 0.379, P < .001; total scar score: r(2) = 0.331, P < .001). In the overall population, 44 patients (21%) developed significant LV remodeling at 6-month follow-up (LV end-systolic volume increase ≥ 15%). These patients showed significantly more impaired peak subepicardial and subendocardial twist at baseline compared with patients without LV remodeling (4.5 ± 1.3° vs 9.4 ± 3.5°, P < .001; 7.0 ± 3.2° vs 12.9 ± 5.8°, P < .001, respectively). Importantly, peak subepicardial twist (odds ratio, 0.241; 95% confidence interval, 0.134-0.431; P < .001) and peak troponin T (odds ratio, 1.152; 95% confidence interval, 1.006-1.320; P = .041) were independently associated with the development of LV remodeling. Peak subepicardial twist strongly reflects infarct transmurality as assessed with magnetic resonance imaging and is independently associated with LV remodeling after AMI.


Leptin levels are negatively correlated with 2-arachidonoylglycerol in the cerebrospinal fluid of patients with osteoarthritis.
There is compelling evidence in humans that peripheral endocannabinoid signaling is disrupted in obesity. However, little is known about the corresponding central signaling. Here, we have investigated the relationship between gender, leptin, body mass index (BMI) and levels of the endocannabinoids anandamide (AEA) and 2-arachidonoylglycerol (2-AG) in the serum and cerebrospinal fluid (CSF) of primarily overweight to obese patients with osteoarthritis. Patients (20 females, 15 males, age range 44-78 years, BMI range 24-42) undergoing total knee arthroplasty for end-stage osteoarthritis were recruited for the study. Endocannabinoids were quantified by liquid chromatography - mass spectrometry. AEA and 2-AG levels in the serum and CSF did not correlate with either age or BMI. However, 2-AG levels in the CSF, but not serum, correlated negatively with CSF leptin levels (Spearman's ρ -0.48, P=0.0076, n=30). No such correlations were observed for AEA and leptin. In the patient sample investigated, there is a negative association between 2-AG and leptin levels in the CSF. This is consistent with pre-clinical studies in animals, demonstrating that leptin controls the levels of hypothalamic endocannabinoids that regulate feeding behavior.


One stage pulse compression at 1554 nm through highly anomalous dispersive photonic crystal fiber.
We demonstrate the pulse compression at 1554 nm using one stage of highly anomalous dispersive photonic crystal fibers with a dispersion value of 600 ps/nm∙km. A 1.64 ps pulse is compressed down to 0.357 ps with a compression factor of 4.6, which agrees reasonably well with the simulation value of 6.1. The compressor is better suited for high energy ultra-short pulse compression than conventional low dispersive single mode fibers.


Comparison of FOLFIRI with or without cetuximab in patients with resected stage III colon cancer; NCCTG (Alliance) intergroup trial N0147.
Two arms with FOLFIRI, with or without cetuximab, were initially included in the randomized phase III intergroup clinical trial NCCTG (North Central Cancer Treatment Group) N0147. When other contemporary trials demonstrated no benefit to using irinotecan as adjuvant therapy, the FOLFIRI-containing arms were discontinued. We report the clinical outcomes for patients randomized to FOLFIRI with or without cetuximab. After resection, patients were randomized to 12 biweekly cycles of FOLFIRI, with or without cetuximab. KRAS (Kirsten rat sarcoma viral oncogene homolog) mutation status was retrospectively determined in a central lab. The primary end point was disease-free survival (DFS). Secondary end points included overall survival (OS) and toxicity. One hundred and six patients received FOLFIRI and 40 received FOLFIRI plus cetuximab. Median follow-up was 5.95 years (range, 0.1-7.0 years). The addition of cetuximab showed a trend toward improved DFS (hazard ratio [HR], 0.53; 95% CI, 0.26-1.1; P = .09) and OS (HR, 0.45; 95% CI, 0.17-1.16; P = .10) in the overall group, regardless of KRAS status, and in patients with wild type KRAS. Grade ≥ 3 nonhematologic adverse effects were significantly increased in the cetuximab versus FOLFIRI-alone arm (68% vs. 46%; P = .02). Adjuvant FOLFIRI resulted in a 3-year DFS less than that expected for FOLFOX. In this small randomized subset of patients with resected stage III colon cancer, the addition of cetuximab to FOLFIRI was associated with a nonsignificant trend toward improved DFS and OS. Nevertheless, considering the limitations of this analysis, FOLFOX without the addition of a biologic agent remains the standard of care for adjuvant therapy in resected stage III colon cancer.


Single-molecule detection: applications to ultrasensitive biochemical analysis.
Recent developments in laser-based detection of fluorescent molecules have made possible the implementation of very sensitive techniques for biochemical analysis. We present and discuss our experiments on the applications of our recently developed technique of single-molecule detection to the analysis of molecules of biological interest. These newly developed methods are capable of detecting and identifying biomolecules at the single-molecule level of sensitivity. In one case, identification is based on measuring fluorescence brightness from single molecules. In another, molecules are classified by determining their electrophoretic velocities.


Some theoretical aspects of the turbulent point-spread function.
In a previous work [J. Opt. Soc. Am. A24, 753 (2007)], we developed a theory describing the space-time statistics of the scintillation and displacement fields of the turbulent point-spread function (PSF), which we validated using imaging data. We now expand on that work by deriving the theoretical expression for the spread fields of the PSF. We also show how the displacement and spread fields are derivable from a set of potentials, and we discuss some of the properties of those potentials.


Structure of purotoxin-2 from wolf spider: modular design and membrane-assisted mode of action in arachnid toxins.
Traditionally, arachnid venoms are known to contain two particularly important groups of peptide toxins. One is disulfide-rich neurotoxins with a predominance of β-structure that specifically target protein receptors in neurons or muscle cells. The other is linear cationic cytotoxins that form amphiphilic α-helices and exhibit rather non-specific membrane-damaging activity. In the present paper, we describe the first 3D structure of a modular arachnid toxin, purotoxin-2 (PT2) from the wolf spider Alopecosa marikovskyi (Lycosidae), studied by NMR spectroscopy. PT2 is composed of an N-terminal inhibitor cystine knot (ICK, or knottin) β-structural domain and a C-terminal linear cationic domain. In aqueous solution, the C-terminal fragment is hyper-flexible, whereas the knottin domain is very rigid. In membrane-mimicking environment, the C-terminal domain assumes a stable amphipathic α-helix. This helix effectively tethers the toxin to membranes and serves as a membrane-access and membrane-anchoring device. Sequence analysis reveals that the knottin + α-helix architecture is quite widespread among arachnid toxins, and PT2 is therefore the founding member of a large family of polypeptides with similar structure motifs. Toxins from this family target different membrane receptors such as P2X in the case of PT2 and calcium channels, but their mechanism of action through membrane access may be strikingly similar.


A global-level assessment of the effectiveness of protected areas at resisting anthropogenic pressures.
One-sixth of the global terrestrial surface now falls within protected areas (PAs), making it essential to understand how far they mitigate the increasing pressures on nature which characterize the Anthropocene. In by far the largest analysis of this question to date and not restricted to forested PAs, we compiled data from 12,315 PAs across 152 countries to investigate their ability to reduce human pressure and how this varies with socioeconomic and management circumstances. While many PAs show positive outcomes, strikingly we find that compared with matched unprotected areas, PAs have on average not reduced a compound index of pressure change over the past 15 y. Moreover, in tropical regions average pressure change from cropland conversion has increased inside PAs even more than in matched unprotected areas. However, our results also confirm previous studies restricted to forest PAs, where pressures are increasing, but less than in counterfactual areas. Our results also show that countries with high national-level development scores have experienced lower rates of pressure increase over the past 15 y within their PAs compared with a matched outside area. Our results caution against the rapid establishment of new PAs without simultaneously addressing the conditions needed to enable their success.


Catalytic asymmetric synthesis of sterically hindered tertiary α-aryl ketones.
The catalytic asymmetric synthesis of a series of tertiary α-aryl cyclopentanones and cyclohexanones has been accomplished via a Pd-catalyzed decarboxylative protonation of the corresponding α-aryl-β-keto allyl esters. Enantioselectivities of up to 92% ee and 74% ee were achieved for cyclopentanone and cyclohexanone substrates, respectively. The route described gives access to these important structural motifs in moderate to high levels of enantioselectivity. In particular, this is only the second direct approach for the preparation of tertiary α-aryl cyclopentanones. The synthetic approach allows for simple modification of the aryl group. Significantly, substrates containing sterically hindered aryl groups gave the highest levels of enantioselectivity, and these aryl groups were readily installed by a Pb-mediated arylation of a β-keto allyl ester.


Selectivity and polarization in water channel membranes: lessons learned from polymeric membranes and CNTs.
Water channels are employed by nature to move pure water across cell membranes while selectively rejecting salts. At present, synthetic channels successfully mimic water permeation, yet even the best channels, such as carbon nanotubes (CNTs) and graphene oxide stacks, still fall short of the selectivity target. The present paper analyzes factors that may help to enhance and control salt rejection based on the lessons learned from conventional membranes and CNTs. First, it highlights the importance of raising the ion self-energy (dielectric mechanism), which suggests that having the channels both narrow and surrounded by a low-dielectric environment is key to high selectivity. In contrast, pore charge alone is insufficient, yet it may help to enhance and tune ion rejection, provided that non-mean-field effects enhanced in low-dielectric pores, such as ion association and sorption, especially of H+ and OH- ions, are properly understood and addressed in the channel design. Second, the role of concentration polarization (CP) is analyzed, which shows that the CP level is apparently low in isolated channels or microscopically small membranes. However, the geometry of the diffusion field should change and CP should increase drastically in macroscopic membranes incorporating densely spaced channel arrays. If not properly addressed in membrane design, the increased CP level in scaled-up channel-based membranes may significantly compromise the observed selectivity and require that target of selectivity be re-set to an even more challenging value. These points may help guide the future development of high-performance artificial water channels and their scale-up towards utilization in next-generation water purification membranes.


Low Plasma Atherogenic Index Associated with Poor Prognosis in Hospitalized Patients with Acute Myocardial Infarction.
the impact of atherogenic index of plasma (AIP), calculated as logarithmic of triglyceride:HDL ratio (log10.[TG:HDL]), on major adverse cardiovascular events (MACE) during acute myocardial infarction (AMI) has not been fully accepted. This study aims to investigate the role of AIP in predicting major adverse cardiovascular events following AMI during intensive care in the hospital. this was a prospective cohort study. We enrolled subjects with AMI hospitalized in intensive coronary care unit at Dr. Sardjito General Hospital, Yogyakarta. The AIP was measured in fasting blood within 24 hours of hospital admission. The total cholesterol, LDL, HDL, and triglyceride (TG), were measured and AIP value was determined as log10.[TG:HDL]). Based on AIP value, subjects were allocated into low AIP (<0.24) and high AIP (0.24). The outcome of the study was major adverse cardiovascular events during hospitalization, i.e. multipart of all cause mortality, acute heart failure, cardiogenic shock, reinfarction, and rescucitated VT/VF. among 277 subjects, the high AIP group comprised 213 subjects (77%) and low AIP group comprised 64 subjects (33%). During intensive hospitalisation, 66 subjects (24%) developed MACE and 20 subjects (7%) developed fatal outcome (all cause mortality). The incidence of MACE tended to be higher in low AIP group, however its difference was not significant. The incidence of all cause mortality was significantly higher in low AIP group (14%) than in high AIP group (5%). Multivariable analysis showed that low AIP predicted all cause mortality independently with a risk ratio 3.71 (95% CI 1.26-10.97, p=0.02). low AIP value (<0.24) is an independent predictor for all cause mortality in patients with acute myocardial infarction undergoing intensive hospitalisation.


Dispersal, environment, and floristic variation of western Amazonian forests.
The distribution of plant species, the species compositions of different sites, and the factors that affect them in tropical rain forests are not well understood. The main hypotheses are that species composition is either (i) uniform over large areas, (ii) random but spatially autocorrelated because of dispersal limitation, or (iii) patchy and environmentally determined. Here we test these hypotheses, using a large data set from western Amazonia. The uniformity hypothesis gains no support, but the other hypotheses do. Environmental determinism explains a larger proportion of the variation in floristic differences between sites than does dispersal limitation; together, these processes explain 70 to 75% of the variation. Consequently, it is important that management planning for conservation and resource use take into account both habitat heterogeneity and biogeographic differences.


Measuring and forecasting progress in education: what about early childhood?
A recent Nature article modelled within-country inequalities in primary, secondary, and tertiary education and forecast progress towards Sustainable Development Goal (SDG) targets related to education (SDG 4). However, their paper entirely overlooks inequalities in achieving Target 4.2, which aims to achieve universal access to quality early childhood development, care and preschool education by 2030. This is an important omission because of the substantial brain, cognitive and socioemotional developments that occur in early life and because of increasing evidence of early-life learning's large impacts on subsequent education and lifetime wellbeing. We provide an overview of this evidence and use new analyses to illustrate medium- and long-term implications of early learning, first by presenting associations between pre-primary programme participation and adolescent mathematics and science test scores in 73 countries and secondly, by estimating the costs of inaction (not making pre-primary programmes universal) in terms of forgone lifetime earnings in 134 countries. We find considerable losses, comparable to or greater than current governmental expenditures on all education (as percentages of GDP), particularly in low- and lower-middle-income countries. In addition to improving primary, secondary and tertiary schooling, we conclude that to attain SDG 4 and reduce inequalities in a post-COVID era, it is essential to prioritize quality early childhood care and education, including adopting policies that support families to promote early learning and their children's education.


Haplotype inference from unphased SNP data in heterozygous polyploids based on SAT.
Haplotype inference based on unphased SNP markers is an important task in population genetics. Although there are different approaches to the inference of haplotypes in diploid species, the existing software is not suitable for inferring haplotypes from unphased SNP data in polyploid species, such as the cultivated potato (Solanum tuberosum). Potato species are tetraploid and highly heterozygous. Here we present the software SATlotyper which is able to handle polyploid and polyallelic data. SATlotyper uses the Boolean satisfiability problem to formulate Haplotype Inference by Pure Parsimony. The software excludes existing haplotype inferences, thus allowing for calculation of alternative inferences. As it is not known which of the multiple haplotype inferences are best supported by the given unphased data set, we use a bootstrapping procedure that allows for scoring of alternative inferences. Finally, by means of the bootstrapping scores, it is possible to optimise the phased genotypes belonging to a given haplotype inference. The program is evaluated with simulated and experimental SNP data generated for heterozygous tetraploid populations of potato. We show that, instead of taking the first haplotype inference reported by the program, we can significantly improve the quality of the final result by applying additional methods that include scoring of the alternative haplotype inferences and genotype optimisation. For a sub-population of nineteen individuals, the predicted results computed by SATlotyper were directly compared with results obtained by experimental haplotype inference via sequencing of cloned amplicons. Prediction and experiment gave similar results regarding the inferred haplotypes and phased genotypes. Our results suggest that Haplotype Inference by Pure Parsimony can be solved efficiently by the SAT approach, even for data sets of unphased SNP from heterozygous polyploids. SATlotyper is freeware and is distributed as a Java JAR file. The software can be downloaded from the webpage of the GABI Primary Database at http://www.gabipd.org/projects/satlotyper/. The application of SATlotyper will provide haplotype information, which can be used in haplotype association mapping studies of polyploid plants.


Automatic extraction of ground control regions and orthorectification of remote sensing imagery.
We develop a fast and accurate method that is able to automatically select and match a large amount of ground control regions (GCRs) for orthorectifying remote sensing imagery. This new method is comprised of four modules, namely automatic extraction of GCRs, fast image-to-image matching, iterating and filtering of GCRs, and rigorous orthorectification. We assess the accuracy of this new method by processing the high-temporal- and high-spatial-resolution Formosat-2 imagery. Results show that the accurate orthoimage with a root mean square error of less than 1.5 pixels can be automatically generated from one standard Formosat-2 image (covering 12 km x 12 km) in 55 minutes. This new method has been incorporated into the Formosat-2 automatic image processing system and has been used to produce orthoimages on a daily-basis.


Antifungal innate immunity: recognition and inflammatory networks.
A large variety of fungi are present in the environment, among which a proportion colonizes the human body, usually without causing any harm. However, depending on the host immune status, commensals can become opportunistic pathogens that induce diseases ranging from superficial non-harmful infection to life-threatening systemic disease. The interplay between the host and the fungal commensal flora is being orchestrated by an efficient recognition of the microorganisms, which in turn ensures a proper balance between tolerance of the normal fungal flora and induction of immune defense mechanisms when invasion occurs. Pattern recognition receptors (PRRs) play a significant role in maintaining this balance due to their capacity to sense fungi and induce host responses such as the induction of proinflammatory cytokines involved in the activation of innate and adaptive immune responses. In the present review, we will discuss the most recent findings regarding the recognition of Candida albicans and Aspergillus fumigatus and the different types of immune cells that play a role in antifungal host defense.


Marburg virus infection in nonhuman primates: Therapeutic treatment by lipid-encapsulated siRNA.
Marburg virus (MARV) and the closely related filovirus Ebola virus cause severe and often fatal hemorrhagic fever (HF) in humans and nonhuman primates with mortality rates up to 90%. There are no vaccines or drugs approved for human use, and no postexposure treatment has completely protected nonhuman primates against MARV-Angola, the strain associated with the highest rate of mortality in naturally occurring human outbreaks. Studies performed with other MARV strains assessed candidate treatments at times shortly after virus exposure, before signs of disease are detectable. We assessed the efficacy of lipid nanoparticle (LNP) delivery of anti-MARV nucleoprotein (NP)-targeting small interfering RNA (siRNA) at several time points after virus exposure, including after the onset of detectable disease in a uniformly lethal nonhuman primate model of MARV-Angola HF. Twenty-one rhesus monkeys were challenged with a lethal dose of MARV-Angola. Sixteen of these animals were treated with LNP containing anti-MARV NP siRNA beginning at 30 to 45 min, 1 day, 2 days, or 3 days after virus challenge. All 16 macaques that received LNP-encapsulated anti-MARV NP siRNA survived infection, whereas the untreated or mock-treated control subjects succumbed to disease between days 7 and 9 after infection. These results represent the successful demonstration of therapeutic anti-MARV-Angola efficacy in nonhuman primates and highlight the substantial impact of an LNP-delivered siRNA therapeutic as a countermeasure against this highly lethal human disease.


Isolation and characterization of a chicken homolog of the c-ret proto-oncogene.
The c-ret proto-oncogene encodes a receptor tyrosine kinase that plays important roles in human disease and in normal mammalian development. Mutations in the human RET gene are associated with multiple endocrine neoplasia syndromes and Hirschsprung's disease in humans, while targeted mutagenesis of murine c-ret resulted in severe developmental abnormalities affecting the excretory and peripheral nervous systems. To examine the evolutionary conservation of the ret protein sequence and its developmental expression pattern, we isolated and sequenced cDNA clones of chicken c-ret and examined its expression in chick embryos and adult tissues. The cytoplasmic domains of chicken and human ret were relatively well conserved (91% similar), but the extracellular domains were more divergent (68% similar), although the conservation of cysteine residues in this region suggests a conserved secondary structure. As in mouse and human, chicken c-ret encodes two protein isoforms. The number and sizes of the transcripts were similar to those in human and mouse cells, and during chick embryogenesis, c-ret mRNA was observed in many of the same sites as in the mouse, including the Wolffian duct and ureteric bud, the enteric, dorsal root, sympathetic and facioacoustic ganglia, and the ventral spinal cord. Evolutionary differences in expression were observed in the trigeminal ganglion, the ventral roots of the spinal cord, the mesenchymal cells of the branchial arches and the adult testes. The results are discussed with regard to the role of the ret receptor in normal development and disease.


Receipt of medications for opioid use disorder among youth engaged in primary care: data from 6 health systems.
Little is known about prevalence and treatment of OUD among youth engaged in primary care (PC). Medications are the recommended treatment of opioid use disorder (OUD) for adolescents and young adults (youth). This study describes the prevalence of OUD, the prevalence of medication treatment for OUD, and patient characteristics associated with OUD treatment among youth engaged in PC. This cross-sectional study includes youth aged 16-25 years engaged in PC. Eligible patients had ≥ 1 PC visit during fiscal years (FY) 2014-2016 in one of 6 health systems across 6 states. Data from electronic health records and insurance claims were used to identify OUD diagnoses, office-based OUD medication treatment, and patient demographic and clinical characteristics in the FY of the first PC visit during the study period. Descriptive analyses were conducted in all youth, and stratified by age (16-17, 18-21, 22-25 years). Among 303,262 eligible youth, 2131 (0.7%) had a documented OUD diagnosis. The prevalence of OUD increased by ascending age groups. About half of youth with OUD had documented depression or anxiety and one third had co-occurring substance use disorders. Receipt of medication for OUD was lowest among youth 16-17 years old (14%) and highest among those aged 22-25 (39%). In this study of youth engaged in 6 health systems across 6 states, there was low receipt of medication treatment, and high prevalence of other substance use disorders and mental health disorders. These findings indicate an urgent need to increase medication treatment for OUD and to integrate treatment for other substance use and mental health disorders.


Advanced potential energy surfaces for condensed phase simulation.
Computational modeling at the atomistic and mesoscopic levels has undergone dramatic development in the past 10 years to meet the challenge of adequately accounting for the many-body nature of intermolecular interactions. At the heart of this challenge is the ability to identify the strengths and specific limitations of pairwise-additive interactions, to improve classical models to explicitly account for many-body effects, and consequently to enhance their ability to describe a wider range of reference data and build confidence in their predictive capacity. However, the corresponding computational cost of these advanced classical models increases significantly enough that statistical convergence of condensed phase observables becomes more difficult to achieve. Here we review a hierarchy of potential energy surface models used in molecular simulations for systems with many degrees of freedom that best meet the trade-off between accuracy and computational speed in order to define a sweet spot for a given scientific problem of interest.


Binding of HMG-I(Y) elicits structural changes in a silencer of the human beta-globin gene.
Proteins involved in repression of the human beta-globin gene may be useful in the treatment of sickle cell anemia, in conjunction with therapy to reactivate fetal globin genes. If there is a reciprocal elevation of gamma-globin expression upon repression, this approach could be useful in additional hemoglobinopathies. We previously showed that repression of the beta-globin gene appears to be mediated through two DNA sequences, silencers I and II, and identified a protein termed BP1 which binds to both silencer sequences. In this study, we cloned two cDNAs encoding proteins which bind to an oligonucleotide in silencer I containing a BP1 binding site. These cDNAs correspond to HMG-I and HMG-Y, isoforms regarded as architectural proteins. We demonstrate that binding of HMG-I(Y) to this oligonucleotide causes bending/flexure of the DNA. HMG-I(Y) also binds to a second oligonucleotide containing a BP1 binding site located in a negative control region upstream of the delta-globin gene, suggesting a role for HMG-I(Y) in repression of adult globin genes. Expression studies revealed that HMG-I(Y) is ubiquitously expressed in human tissues that do not express beta-globin, being present in 48 of 50 tissues and six hematopoietic cell lines examined. Furthermore, HMG-I(Y) expression is down-regulated during differentiation of primary erythroid cells. We present a model in which HMG-I(Y) alters DNA conformation to allow binding of repressor proteins, and in which the relative amount of HMG-I(Y) helps to determine the repressive state of the beta-globin gene.


Angiomatoid giant cellular blue nevus of vaginal wall associated with pregnancy.
Blue nevi that arise from the Müllerian tract are rare melanocytic lesions. Several histopathologic variants of cellular blue nevi have been described. The angiomatoid variant is characterized by a vascular component, and is considered to be a rare variant. Few studies have explored the influence of pregnancy on melanocytic lesions. A 29-year-old woman was presented with a pigmented vaginal lesion that increased gradually during pregnancy. A full term gynecologic examination showed a tumor mass protruding into the vaginal canal. The mass was resected during cesarean-section under the clinical impression of vaginal hemangioma. Gross examination revealed a cystic mass measuring 6.0 × 4.3 × 3.5 cm, which was filled with dark friable material. Histologically, the mass showed a subepithelial cellular proliferation of heavily pigmented dendritic melanocytes with prominent vascular stroma. Cytologic pleomorphism, junctional activity, atypical mitosis, and necrosis were not found. The proliferation was immunoreactive for HMB-45, S-100 and melan-A, and non-immunoreactive for CD34, smooth muscle actin, and AE1/AE3. The MIB-1 proliferative index was less than 1%. The patient had a postoperative course without complication. Angiomatoid giant cellular blue nevus arising from the vagina during pregnancy is extremely rare. The low proliferative index and absence of cytologic pleomorphism, or necrosis, supports a benign biological behavior. Clinical follow-up showed no evidence of recurrence at one year after the resection of the mass.


The view from elsewhere: perspectives on ALife modeling.
Many artificial life researchers stress the interdisciplinary character of the field. Against such a backdrop, this report reviews and discusses artificial life, as it is depicted in, and as it interfaces with, adjacent disciplines (in particular, philosophy, biology, and linguistics), and in the light of a specific historical example of interdisciplinary research (namely cybernetics) with which artificial life shares many features. This report grew out of a workshop held at the Sixth European Conference on Artificial Life in Prague and features individual contributions from the workshop's eight speakers, plus a section designed to reflect the debates that took place during the workshop's discussion sessions. The major theme that emerged during these sessions was the identity and status of artificial life as a scientific endeavor.


General theory for integrated analysis of growth, gene, and protein expression in biofilms.
A theory for analysis and prediction of spatial and temporal patterns of gene and protein expression within microbial biofilms is derived. The theory integrates phenomena of solute reaction and diffusion, microbial growth, mRNA or protein synthesis, biomass advection, and gene transcript or protein turnover. Case studies illustrate the capacity of the theory to simulate heterogeneous spatial patterns and predict microbial activities in biofilms that are qualitatively different from those of planktonic cells. Specific scenarios analyzed include an inducible GFP or fluorescent protein reporter, a denitrification gene repressed by oxygen, an acid stress response gene, and a quorum sensing circuit. It is shown that the patterns of activity revealed by inducible stable fluorescent proteins or reporter unstable proteins overestimate the region of activity. This is due to advective spreading and finite protein turnover rates. In the cases of a gene induced by either limitation for a metabolic substrate or accumulation of a metabolic product, maximal expression is predicted in an internal stratum of the biofilm. A quorum sensing system that includes an oxygen-responsive negative regulator exhibits behavior that is distinct from any stage of a batch planktonic culture. Though here the analyses have been limited to simultaneous interactions of up to two substrates and two genes, the framework applies to arbitrarily large networks of genes and metabolites. Extension of reaction-diffusion modeling in biofilms to the analysis of individual genes and gene networks is an important advance that dovetails with the growing toolkit of molecular and genetic experimental techniques.


Metabolism-based inactivation of penile nitric oxide synthase activity by guanabenz.
Guanabenz (Wytensin) was shown to inactivate nitric oxide synthase (NOS) activity in vitro and in vivo. In in vitro studies with the use of a cytosolic fraction from penile tissue, the inactivation was found to depend on NADPH, time, and the concentration of guanabenz. The L-, but not the D-, isomer of arginine could protect from the inactivation, suggesting an active site-directed event. The kinetics of inactivation could be described by an apparent dissociation constant for the initial reversible complex (Ki) and a pseudo first-order inactivation constant (kinact) of 38.5 microM and 0.179 min-1, respectively. In in vivo studies, guanabenz was shown to inhibit penile cytosolic NOS activity in a dose- and time-dependent manner. Treatment of rats with guanabenz (5 mg/kg/day) for 4 days caused a decrease of approximately one-half in the NOS activity of the penile cytosolic fraction with a concomitant loss in the amount of immunodetectable NOS protein. Treatment for 4 days at a dose of 0. 5 mg/kg/day showed a similar decrease in activity, whereas a dose of 0.05 mg/kg/day showed no effects. Due to the multitude of processes that are regulated by NO, the inactivation of NOS is a potential mechanism to be considered in a variety of biological effects associated with drugs.


Rehabilitation of occlusion - science or art?
The primary objective of rehabilitating occlusion is to improve stomatognathic function in patients experiencing dysfunction in mastication, speech, and swallowing as a consequence of tooth loss. The procedure of occlusal treatment involves improving the morphology and the stomatognathic function. Several practical methods and morphological endpoints have been described in occlusal rehabilitation. We made a selection of these (mandibular position, occlusal plane, occlusal guidance, occlusal contact, face-bow transfer, use of an adjustable articulator and occlusal support) and performed a literature review to verify the existence of compelling scientific evidence for each of these. A literature search was conducted using Medline/PubMed in March 2011. Over 400 abstracts were reviewed, and more than 50 manuscripts selected. An additional hand search was also conducted. Of the many studies investigating stomatognathic function in relation to specific occlusal schemes, most studies were poorly designed and of low quality, thus yielding ambiguous results. Overall, there is no scientific evidence that supports any specific occlusal scheme being superior to others in terms of improving stomatognathic function, nor that sophisticated methods are superior to simpler ones in terms of clinical outcomes. However, it is obvious that the art of occlusal rehabilitation requires accurate, reproducible, easy and quick procedures to reduce unnecessary technical failures and/or the requirement for compensatory adjustments. Therefore, despite the lack of scientific evidence for specific treatments, the acquisition of these general skills by dentists and attaining profound knowledge and skills in postgraduate training will be necessary for specialists in charge of complicated cases.


Protection by the Total Flavonoids from Rosa laevigata Michx Fruit against Lipopolysaccharide-Induced Liver Injury in Mice via Modulation of FXR Signaling.
We previously reported the effects of the total flavonoids (TFs) from Rosa laevigata Michx fruit against carbon tetrachloride-induced liver damage, non-alcoholic fatty liver disease, and liver ischemia-reperfusion injury. However, there have been no papers reporting the role of R. laevigata TFs against lipopolysaccharide (LPS)-induced liver injury. In this paper, liver injury in mice was induced by LPS, and R. Laevigata extract was intragastrically administered to the mice for 7 days. Biochemical parameters in serum and liver tissue were examined, and pathological changes were observed by transmission electron microscopy, hematoxylin and eosin (H&E) and Oil Red O staining. The results showed that the TFs markedly reduced serum ALT (alanine transferase), AST (aspartate transaminase), TG (total triglyceride), and TC (total cholesterol) levels and relative liver weights and improved liver pathological changes. In addition, the TFs markedly decreased tissue MDA (malondialdehyde) level and increased the levels of SOD (superoxide dismutase) and GSH-Px (glutathione peroxidase). A mechanistic study showed that the TFs significantly increased the expression levels of Nrf2 (nuclear erythroid factor2-related factor 2), HO-1 (heme oxygenase-1), NQO1 (NAD(P)H dehydrogenase (quinone 1), GCLC (glutamate-cysteine ligase catalytic subunit), and GCLM (glutamate-cysteine ligase regulatory subunit) and decreased Keap1 (Kelch-like ECH-associated protein 1) level by activating FXR (farnesoid X receptor) against oxidative stress. Furthermore, the TFs markedly suppressed the nuclear translocation of NF-κB (nuclear factor-kappa B) and subsequently decreased the expression levels of IL (interleukin)-1β, IL-6, HMGB-1 (high -mobility group box 1), and COX-2 (cyclooxygenase-2) by activating FXR and FOXO3a (forkhead box O3) against inflammation. Besides, the TFs obviously reduced the expression levels of SREBP-1c (sterol regulatory element-binding proteins-1c), ACC1 (acetyl-CoA carboxylase-1), FASN (fatty acid synthase), and SCD1 (stearoyl-coenzyme A desaturase 1), and improved CPT1 (carnitine palmitoyltransferase 1) level by activating FXR to regulate lipid metabolism. Our results suggest that TFs exhibited protective effect against LPS-induced liver injury by altering FXR-mediated oxidative stress, inflammation, and lipid metabolism, and should be developed as an effective food and healthcare product for the therapy of liver injury in the future.


Horizontal and Vertical Integration of Health Care Providers: A Framework for Understanding Various Provider Organizational Structures.
Current U.S. policy and payment initiatives aim to encourage health care provider accountability for population health and higher value care, resulting in efforts to integrate providers along the continuum. Providers work together through diverse organizational structures, yet evidence is limited regarding how to best organize the delivery system to achieve higher value care. In 2016, we conducted a narrative review of 10 years of literature to identify definitional components of key organizational structures in the United States. A clear accounting of common organizational structures is foundational for understanding the system attributes that are associated with higher value care. We distinguish between structures characterized by the horizontal integration of providers delivering similar services and the vertical integration of providers fulfilling different functions along the care continuum. We characterize these structures in terms of their origins, included providers and services, care management functions, and governance. Increasingly, U.S. policymakers seek to promote provider integration and coordination. Emerging evidence suggests that organizational structures, composition, and other characteristics influence cost and quality performance. Given current efforts to reform the U.S. delivery system, future research should seek to systematically examine the role of organizational structure in cost and quality outcomes.


Urinary collagen IV and πGST: potential biomarkers for detecting localized kidney injury in diabetes--a pilot study.
Urinary biomarkers can identify damage to specific parts of the nephron. We performed a cross-sectional study to characterise the pattern of diabetic nephropathy using urinary biomarkers of glomerular fibrosis (collagen IV), proximal tubular damage (α-glutathione-S-transferase, GST) and distal tubular damage (πGST). Clinical data from 457 unselected patients attending a hospital diabetes clinic were collected. Spot urine samples were analysed for albumin and creatinine. Biomarkers were measured by enzyme-linked immunosorbent assay, and corrected to urinary creatinine. All 3 biomarkers correlated weakly with albumin/creatinine ratios (Pearson correlation <0.2, p values <0.001). The most common abnormality was elevated urinary collagen IV (glomerular, 35%) compared to αGST (proximal tubule, 18%) or πGST (distal tubule, 15%). The proportion of patients with abnormal biomarker results increased across the normo-, micro- and macroalbuminuria groups, with collagen IV (26, 58, 65%) and πGST (11, 25, 35%) but not αGST. In patients with diabetes, these urinary biomarkers appear to identify renal damage that is related to, but distinct from, urine albumin/creatinine ratios. The markers of glomerular fibrosis and distal tubular damage related most closely to the degree of albuminuria. Longitudinal studies are now required to assess whether these biomarkers can detect early renal disease with greater specificity and sensitivity than the albumin/creatinine ratio.


Dissociation of 24-hour catecholamine levels from blood pressure in older men.
Increased plasma norepinephrine levels have been observed in some persons with early essential hypertension. Although both plasma norepinephrine level and mean arterial blood pressure rise with age, little is known about the state of catecholamine secretion in elderly patients with essential hypertension. We studied the 24-hour cycle levels of plasma norepinephrine, epinephrine, and dopamine in 12 elderly hypertensive subjects and 13 age-matched normotensive controls (mean ages, 63.8 +/- 1.2 yr and 64.8 +/- 1.8 yr [SEM] respectively). Blood samples were obtained at bihourly intervals from 0900 to 2100 hours and every 30 minutes from 2100 to 0900 hours, during which time sleep and breathing were continuously monitored. A circadian rhythm was displayed in both groups by plasma epinephrine levels (mesor, 49 +/- 2 pg/ml and 38 +/- 1 pg/ml; amplitude, 15 +/- 2 pg/ml and 11 +/- 1 pg/ml; acrophase, 12.20 +/- 0.40 hr and 14.41 +/- 0.34 hr in the normotensive and hypertensive groups respectively) but not by plasma norepinephrine or dopamine levels. During the 24-hour cycle plasma epinephrine, but not norepinephrine or dopamine, levels were positively related to mean arterial blood pressure (r = 0.60 for the normotensive subjects, r = 0.57 for the hypertensive subjects, p less than 0.01 for both).(ABSTRACT TRUNCATED AT 250 WORDS)


Update on the OMERACT magnetic resonance imaging task force: research and future directions.
Magnetic resonance imaging (MRI) provides an important biomarker across a range of rheumatological diseases. At the Outcome Measures in Rheumatology (OMERACT) 11 meeting, the MRI task force continued its work of developing and improving the use of MRI outcomes for use in clinical trials. The breadth of pathology in the Rheumatoid Arthritis MRI Score has been strengthened with further work on the development of a joint space narrowing score, and a series of exercises presented at OMERACT 11 demonstrated good reliability and construct validity for this assessment. Understanding the importance of residual inflammation after RA treatment remains a major focus of the group's work. Analyses were presented on defining the level of synovitis (using MRI scores of a single hand) that would predict absence of erosion progression. The development of the OMERACT Hand Osteoarthritis MRI score has continued with substantial work presented on its iterative development, including pathology definition, scaling, and subsequent reliability of the score. Optimizing the role of MRI as a robust biomarker and surrogate outcome remains a priority for this group.


Functional characterization of filamin a interacting protein 1-like, a novel candidate for antivascular cancer therapy.
Inhibiting angiogenesis has become a major therapeutic strategy for cancer treatment. To identify common intracellular mediators, we previously analyzed gene expression profiles of endothelial cells after treatment with angiogenesis inhibitors. Filamin A interacting protein 1-like (FILIP1L; previously known as down-regulated in ovarian cancer 1) was identified as one of the genes up-regulated in endothelial cells in response to these inhibitors. However, the expression and function of FILIP1L protein is uncharacterized. Here, we provide the first description of the expression and specific subcellular localization of FILIP1L protein in human tissue. Overexpression of FILIP1L resulted in inhibition of cell proliferation and migration and increased apoptosis. In addition, overexpression of FILIP1L truncation mutants showed differential antiproliferative activity. A COOH terminal truncation mutant (FILIP1LDeltaC103) was more potent than wild-type FILIP1L in mediating this activity. Targeted expression of FILIP1LDeltaC103 in tumor vasculature inhibited tumor growth in vivo. Overall, these findings suggest that the novel protein FILIP1L may be an important mediator of the effects of angiogenesis inhibitors and that FILIP1L has the potential to be an antivascular reagent for cancer therapy.


De novo DNA synthesis by human DNA polymerase lambda, DNA polymerase mu and terminal deoxyribonucleotidyl transferase.
DNA polymerases (pols) catalyse the synthesis of DNA. This reaction requires a primer-template DNA in order to grow from the 3'OH end of the primer along the template. On the other hand terminal deoxyribonucleotidyl transferase (TdT) catalyses the addition of nucleotides at the 3'OH end of a DNA strand, without the need of a template. Pol lambda and pol micro are ubiquitous enzymes, possess both DNA polymerase and terminal deoxyribonucleotidyl transferase activities and belong to pol X family, together with pol beta and TdT. Here we show that pol lambda, pol micro and TdT, all possess the ability to synthesise in vitro short fragments of DNA in the absence of a primer-template or even a primer or a template in the reaction. The DNA synthesised de novo by pol lambda, pol micro and TdT appears to have an unusual structure. Furthermore we found that the amino acid Phe506 of pol lambda is essential for the de novo synthesis. This novel catalytic activity might be related to the proposed functions of these three pol X family members in DNA repair and DNA recombination.


Ca(2+)/calmodulin-dependent protein kinase IV is expressed in spermatids and targeted to chromatin and the nuclear matrix.
Ca(2+)/calmodulin-dependent protein kinase IV and calspermin are two proteins encoded by the Camk4 gene. Both are highly expressed in the testis, where in situ hybridization studies in rat testes have demonstrated that CaMKIV mRNA is localized to pachytene spermatocytes, while calspermin mRNA is restricted to spermatids. We have examined the expression patterns of both CaMKIV and calspermin in mouse testis and unexpectedly find that CaMKIV is expressed in spermatogonia and spermatids but excluded from spermatocytes, while calspermin is found only in spermatids. CaMKIV and calspermin expression in the testis are stage-dependent and appear to be coordinately regulated. In germ cells, we find that CaMKIV is associated with the chromatin. We further demonstrate that a fraction of CaMKIV in spermatids is hyperphosphorylated and specifically localized to the nuclear matrix. These novel findings may implicate CaMKIV in chromatin remodeling during nuclear condensation of spermatids.


The significance of ultrastructural features in acute myeloid leukemia: a study of 220 patients entered into the Medical Research Council's ninth acute myeloid leukemia trial.
Samples of peripheral blood or bone marrow obtained before treatment from 220 patients entered into the Medical Research Council's ninth acute myeloid leukemia (AML) trial were examined by electron microscopy. Several ultrastructural features showed strong correlations with each other, with variants of AML defined by the Hayhoe classification scheme, and with FAB type. However, no ultrastructural feature was found either uniquely in association with any other or specifically in any variant of AML. Each ultrastructural feature was tested for association with achievement of remission, remission duration, and survival time. Only Auer rods were associated with a high remission rate (p = 0.01), but even this association was not quite conventionally significant when the analysis was not quite conventionally significant when the analysis was stratified for age (p = 0.055). Shorter duration of remission was associated with cytoplasmic projections (p = 0.04 stratified for age). Overall survival was worse when convoluted or lobed nuclei were present but only when stratified for age and Hayhoe type (p = 0.04). The possibility of testing combinations of ultrastructural features for correlation with diagnosis or prognosis is discussed but would require data from more than the present 220 patients to be meaningful.


[Development of "MADE", a French Job exposure matrix for evaluation of biomechanical exposure].
This paper presents the stages of development of an occupational biomechanical exposure matrix and preliminary reliability and validity indicators. The expertise-based job exposure matrix, called "MADE" (for "difficult physical conditions and job matrix"), was developed from the French and international classification of jobs for 17 biomechanical exposures. Three pairs of investigators independently rate the frequency and intensity of exposure of each job from 0 to 5; discordant scores within pairs (mean difference >20% for each biomechanical exposure considered) were discussed collectively. Reliability was assessed by the difference between the initial ratings, the number of revised jobs, and the correlation between the two classifications (with transcoding). Validity was studied by correlating variables similar to those from existing United States and Danish matrices. The difference between the mean scores of the pairs was considered to be "fair" (less than one point). Of the 51 paired results studied (17 exposures - 3 pairs of investigators), four coefficients were found to be less than 0.6, and 14 were between 0.6 and 0.7. Forty-nine of the 1,169 job categories were reviewed. Correlation coefficients between the initial classification and transcoding were greater than 0.7. Correlation coefficients between the French, United States and Danish MEE variables were fair to good. An expertise-based job exposure matrix with fairly reliable indicators has been developed, opening up the prospects to improve certain some fields of public health, at both national and international levels.


Chronic GvHD-associated serositis and pericarditis.
Serositis is a rare manifestation of chronic GvHD (cGvHD). No risk factors or laboratory changes associated with this syndrome have been recognized to date, and outcomes have not been described in a large series. We searched our institutional database for patients undergoing allogeneic hematopoietic cell transplant identified as having serositis or pericarditis. Laboratory studies from prior to diagnosis, at diagnosis and post diagnosis of serositis, as well as outcomes from invasive procedures were included. Twenty patients met criteria for cGvHD-associated serositis, and all but three patients had a prior diagnosis of cGvHD. Fifteen were male, and the complication occurred in the setting of immunosuppressant taper in 12 cases. Ten patients required invasive interventions, including pericardial window or stripping. A significant increase in blood monocytes and decrease in serum albumin were identified at diagnosis compared with pre-diagnosis. Out of 20 patients, 17 were treated with steroids, with 12 demonstrating a complete response. These data suggest that cGvHD-associated serositis occurs mainly in the setting of treated as opposed to de novo cGvHD and biomarkers associated with the syndrome include a decrease in albumin and an increase in absolute monocyte count. Outcome data from larger series are required to better understand the optimal management of this rare complication.


Beta bursts during continuous movements accompany the velocity decrement in Parkinson's disease patients.
Bradykinesia is reported to correlate with subthalamic beta power (13-35 Hz) recorded at rest in Parkinson's disease (PD). Pilot studies suggest adaptive deep brain stimulation triggered by amplitude threshold crossings of beta activity defined at rest is effective. This is puzzling, given that beta is suppressed during repetitive movements when bradykinesia becomes apparent. Recently, increased beta power in PD has been linked to beta bursts. Here we investigate whether beta bursts also occur during repetitive movements and relate to progressive decrement in movement velocity. Therefore, subthalamic local field potentials were recorded in 12 PD patients off medication while performing 30s blocks of rotatory movements alternating with rest periods. Bursts were defined separately for the low (13-20 Hz) and high (20-35 Hz) beta band using thresholds defined at rest. As expected, velocity significantly decreased within movement blocks. Despite the sustained suppression of both beta sub-bands, bursts could still be detected during movement. Beta bursts were reduced in amplitude, duration and rate during movement with beta rate correlating best with beta power. A mixed-effects linear model revealed that percentage time spent in beta bursts predicted velocity decreases better than averaged power. This correlation was specific for the low beta band. Our results link beta bursts during movement to bradykinesia. This helps explain how beta activity may contribute to bradykinetic movement decrement even though mean beta power is reduced during movement. Moreover, our findings help explain the effectiveness of adaptive DBS triggered off beta bursts, even though these may be defined with respect to beta levels at rest.


Yeast Trm7 interacts with distinct proteins for critical modifications of the tRNAPhe anticodon loop.
Post-transcriptional modification of the tRNA anticodon loop is critical for translation. Yeast Trm7 is required for 2'-O-methylation of C(32) and N(34) of tRNA(Phe), tRNA(Trp), and tRNA(Leu(UAA)) to form Cm(32) and Nm(34), and trm7-Δ mutants have severe growth and translation defects, but the reasons for these defects are not known. We show here that overproduction of tRNA(Phe) suppresses the growth defect of trm7-Δ mutants, suggesting that the crucial biological role of Trm7 is the modification of tRNA(Phe). We also provide in vivo and in vitro evidence that Trm7 interacts with ORF YMR259c (now named Trm732) for 2'-O-methylation of C(32), and with Rtt10 (named Trm734) for 2'-O-methylation of N(34) of substrate tRNAs and provide evidence for a complex circuitry of anticodon loop modification of tRNA(Phe), in which formation of Cm(32) and Gm(34) drives modification of m(1)G(37) (1-methylguanosine) to yW (wyebutosine). Further genetic analysis shows that the slow growth of trm7-Δ mutants is due to the lack of both Cm(32) and Nm(34), and the accompanying loss of yW, because trm732-Δ trm734-Δ mutants phenocopy trm7-Δ mutants, whereas each single mutant is healthy; nonetheless, TRM732 and TRM734 each have distinct roles, since mutations in these genes have different genetic interactions with trm1-Δ mutants, which lack m(2,2)G(26) in their tRNAs. We speculate that 2'-O-methylation of the anticodon loop may be important throughout eukaryotes because of the widespread conservation of Trm7, Trm732, and Trm734 proteins, and the corresponding modifications, and because the putative human TRM7 ortholog FTSJ1 is implicated in nonsyndromic X-linked mental retardation.


Uptake epithelia behave in a cell-centric and not systems homeostatic manner in response to zinc depletion and supplementation.
Much remains to be understood about systemic regulation of zinc uptake in vertebrates, and adequate zinc status is far from always achieved in animals or human. In addition to absorbing zinc from the diet, fish are able to take up zinc directly from the water with the gills. This provides an elegant system to study zinc uptake, how it relates to zinc status, and the expression of genes for proteins involved in zinc acquisition. A 21-day experiment was conducted in which zebrafish were acclimated to deficient, control or excess zinc concentrations in the water and feed. Deficient provision of zinc reduced whole body zinc, potassium, sodium and calcium levels whilst zinc concentrations in the uptake epithelia (gills and gut) remained unchanged. Excess levels of zinc caused accumulation of zinc in the gills, intestine and carcass, but impaired whole body iron, sodium and calcium concentrations. Fish subjected to zinc deficiency had, surprisingly, a reduced zinc influx across the gill epithelium, even when tested at a high concentration of zinc in the water. Zinc influx in the excess group was indistinct from the control. Expression of genes for metallothionein-2 (Mt2) and zinc transporters-1, -2, and -8 (Znt1, Znt2, Znt8) in uptake epithelia showed in general a direct relationship with zinc supply, while mRNA for Zip4 was inversely related to zinc supply. Transcripts for the epithelial calcium channel (Ecac/Trpv6) showed time-dependent increased expression in the gills of the deficiency group, and a transient decrease of expression during zinc excess. Transcriptome profiling by microarrays showed that in both gills and intestine, the most markedly affected biological functions were those related to cell growth, proliferation and cancer, closely followed by processes of gene transcription and protein synthesis in general. Whilst changes in zinc supply had profound effects in the intestine on genes associated with uptake and metabolism of macronutrients, many of the unique categories of genes preferentially regulated in the gill could be mapped onto signalling pathways. This included pathways for PPAR/RXR, LXR/RXR, ATM, chemokine, and BMP signalling. Overall, the responses of epithelial tissue to zinc deficiency and excess are best explained by local epithelial homeostasis with no evidence of systemic control.


Caffeine withdrawal increases cerebral blood flow velocity and alters quantitative electroencephalography (EEG) activity.
Cessation of daily caffeine consumption produces a withdrawal syndrome comprised of subjective symptoms and functional impairment. Few controlled studies have examined the physiological effects of caffeine withdrawal. The present study examined the effect of caffeine withdrawal on cerebral blood flow velocity and quantitative EEG. Ten volunteers reporting moderate caffeine intake (mean 333 mg/day) participated in this double-blind study. Subjects completed several tests when maintaining their normal diet (baseline period) and during two 1-day periods during which they consumed caffeine-free diets and received capsules containing placebo (placebo test session) or caffeine (caffeine test session) in amounts equal to their baseline daily caffeine consumption. Blood flow velocity was determined for four arteries: right and left middle (MCA), and right and left anterior (ACA) cerebral arteries using pulsed transcranial Doppler sonography. EEG was recorded for 3 min from eight scalp sites while subjects sat, with eyes closed, in a sound-attenuated electronically shielded chamber. Subjective effects were assessed with questionnaires. Results showed an effect of the placebo (21-h withdrawal) condition compared to the caffeine condition. Placebo significantly increased the mean velocity, systolic velocity and diastolic velocity (cm/s) in all four cerebral arteries. In the MCA, the pulsatility index was significantly decreased following placebo. Placebo significantly increased EEG theta power. Placebo also produces subjective effect changes, including increases in heavy feelings in arms and legs and decreases in ability to concentrate. The caffeine and baseline conditions produced similar results on both the physiological and subjective measures. Cessation of daily caffeine consumption produced changes in cerebral blood flow velocity and quantitative EEG. These changes may be related to classic caffeine withdrawal symptoms of headache, drowsiness and decreased alertness.


Implementation of an electronic surgical referral service. Collaboration, consensus and cost of the surgeon - general practitioner Delphi approach.
Poor coordination between levels of care plays a central role in determining the quality and cost of health care. To improve patient coordination, systematic structures, guidelines, and processes for creating, transferring, and recognizing information are needed to facilitate referral routines. Prospective observational survey of implementation of electronic medical record (EMR)-supported guidelines for surgical treatment. One university clinic, two local hospitals, 31 municipalities, and three EMR vendors participated in the implementation project. Surgical referral guidelines were developed using the Delphi method; 22 surgeons and seven general practitioners (GPs) needed 109 hours to reach consensus. Based on consensus guidelines, an electronic referral service supported by a clinical decision support system, fully integrated into the GPs' EMR, was developed. Fifty-five information technology personnel and 563 hours were needed (total cost 67,000 £) to implement a guideline supported system in the EMR for 139 GPs. Economical analyses from a hospital and societal perspective, showed that 504 (range 401-670) and 37 (range 29-49) referred patients, respectively, were needed to provide a cost-effective service. A considerable amount of resources were needed to reach consensus on the surgical referral guidelines. A structured approach by the Delphi method and close collaboration between IT personnel, surgeons and primary care physicians were needed to reach consensus.


Improving life expectancy: how many years behind has the USA fallen? A cross-national comparison among high-income countries from 1958 to 2007.
Many studies have documented higher mortality levels in the USA compared to other high-income nations. We add to this discussion by quantifying how many years behind comparison countries the USA has fallen and by identifying when US mortality rates began to diverge. We use full life tables, for men and women, for 17 high-income countries including the USA. We extract the life expectancy at birth and compute the mortality rates for each 5-year age group from birth up to age 80. Using the metric of how many 'years behind' a country has fallen, we compare US mortality levels with those in other high-income countries ('comparators'). We report life expectancy for 17 high-income countries, for the period 1958-2007. Up to the late 1970s, US men and especially women closely tracked comparators in life expectancy. In the late 1970s in the USA, most strikingly women began to diverge from comparators so that the US female life expectancy in 2007 corresponded to that of the comparators' average 10 years earlier. Mortality rates also began to diverge from the late 1970s, and the largest mortality gap was in the 15-49 age group, for both men and women, where the USA had fallen about 40 years behind the comparators by 2007. Some causes proposed for the relatively high US mortality today-racial differences, lack of universal health insurance, US exceptionalism-changed little while the mortality gap emerged and grew. This suggests that explanations for the growing gap lie elsewhere. Quantification of how many years behind the USA has fallen can help provide clues about where to look for potential causes and remedies.


Role of word-of-mouth for programs of voluntary vaccination: A game-theoretic approach.
We propose a model describing the synergetic feedback between word-of-mouth (WoM) and epidemic dynamics controlled by voluntary vaccination. The key feature consists in combining a game-theoretic model for the spread of WoM and a compartmental model describing VSIR disease dynamics in the presence of a program of voluntary vaccination. We evaluate and compare two scenarios for determinants of behavior, depending on what WoM disseminates: (1) vaccine advertising, which may occur whether or not an epidemic is ongoing and (2) epidemic status, notably disease prevalence. Understanding the synergy between the two strategies could be particularly important for designing voluntary vaccination campaigns. We find that, in the initial phase of an epidemic, vaccination uptake is determined more by vaccine advertising than the epidemic status. As the epidemic progresses, epidemic status becomes increasingly important for vaccination uptake, considerably accelerating vaccination uptake toward a stable vaccination coverage.


A procedure to find thermodynamic equilibrium constants for CO2 and CH4 adsorption on activated carbon.
Thermodynamic equilibrium for adsorption means that the chemical potential of gas and adsorbed phase are equal. A precise knowledge of the chemical potential is, however, often lacking, because the activity coefficient of the adsorbate is not known. Adsorption isotherms are therefore commonly fitted to ideal models such as the Langmuir, Sips or Henry models. We propose here a new procedure to find the activity coefficient and the equilibrium constant for adsorption which uses the thermodynamic factor. Instead of fitting the data to a model, we calculate the thermodynamic factor and use this to find first the activity coefficient. We show, using published molecular simulation data, how this procedure gives the thermodynamic equilibrium constant and enthalpies of adsorption for CO2(g) on graphite. We also use published experimental data to find similar thermodynamic properties of CO2(g) and of CH4(g) adsorbed on activated carbon. The procedure gives a higher accuracy in the determination of enthalpies of adsorption than ideal models do.


Construction of coffee transcriptome networks based on gene annotation semantics.
Gene annotation is a process that encompasses multiple approaches on the analysis of nucleic acids or protein sequences in order to assign structural and functional characteristics to gene models. When thousands of gene models are being described in an organism genome, construction and visualization of gene networks impose novel challenges in the understanding of complex expression patterns and the generation of new knowledge in genomics research. In order to take advantage of accumulated text data after conventional gene sequence analysis, this work applied semantics in combination with visualization tools to build transcriptome networks from a set of coffee gene annotations. A set of selected coffee transcriptome sequences, chosen by the quality of the sequence comparison reported by Basic Local Alignment Search Tool (BLAST) and Interproscan, were filtered out by coverage, identity, length of the query, and e-values. Meanwhile, term descriptors for molecular biology and biochemistry were obtained along the Wordnet dictionary in order to construct a Resource Description Framework (RDF) using Ruby scripts and Methontology to find associations between concepts. Relationships between sequence annotations and semantic concepts were graphically represented through a total of 6845 oriented vectors, which were reduced to 745 non-redundant associations. A large gene network connecting transcripts by way of relational concepts was created where detailed connections remain to be validated for biological significance based on current biochemical and genetics frameworks. Besides reusing text information in the generation of gene connections and for data mining purposes, this tool development opens the possibility to visualize complex and abundant transcriptome data, and triggers the formulation of new hypotheses in metabolic pathways analysis.


Histophagous ciliate Pseudocollinia brintoni and bacterial assemblage interaction with krill Nyctiphanes simplex. II. Host responses.
Unlike decapod crustaceans of commercial interest, the krill defense system and its response to parasites and pathogens is virtually unknown. Histophagous ciliates of the genus Pseudocollinia interact with at least 7 krill species in the northeastern Pacific. Although they can cause epizootic events, the physiology of the histophagous ciliate-host interaction and krill (host) defenses remain unknown. From 1 oceanographic survey along the southwestern coast of the Baja California Peninsula near Bahía Magdalena and 2 in the Gulf of California, we investigated parasitoid-host physiological responses (fatty acid and oxidative stress indicators) of the subtropical krill Nyctiphanes simplex infected with the ciliate P. brintoni. All life stages of P. brintoni were associated with opportunistic bacterial assemblages that have not been explicitly investigated in other Pseudocollinia species (P. beringensis, P. oregonensis, and P. similis). Parasitoid ciliates exclusively infected adult females, which showed increased lipid content during gonad development. As the infection progressed, omega-3 eicosapentaenoic and docosahexaenoic fatty acids, which may act as energy sources to produce high numbers of ciliate transmission stages, were quickly depleted. Antioxidant enzymes, components of the crustacean defense system, varied throughout infection, but without inhibiting Pseudocollinia infection, i.e. higher levels of lipid oxidative damage were detected in late stages of infection. The ineffective response of the krill antioxidant defense system against histophagous ciliates and the bacteria associated with the ciliates suggests that Pseudocollinia ciliates are functionally analogous to krill predators and may have a strong influence on the population dynamics of krill.


Copper-catalyzed oxidative cross-coupling of α-aminocarbonyl compounds with primary amines toward 2-oxo-acetamidines.
A general and mild method for the construction of a carbon-nitrogen bond via copper-catalyzed oxidative cross-coupling of amines with α-aminocarbonyl compounds was achieved. Amines, either aliphatic primary amines, aromatic primary amines or secondary amines can be used as the starting materials. When R2 was different from R3, two isomers would be observed. Therefore, this reaction system has a broad substrate scope and provides a facile pathway for the synthesis of 2-oxo-acetamidines.


Magnetic resonance imaging detects placental hypoxia and acidosis in mouse models of perturbed pregnancies.
Endothelial dysfunction as a result of dysregulation of anti-angiogenic molecules secreted by the placenta leads to the maternal hypertensive response characteristic of the pregnancy complication of preeclampsia. Structural abnormalities in the placenta have been proposed to result in altered placental perfusion, placental oxidative stress, cellular damage and inflammation and the release of anti-angiogenic compounds into the maternal circulation. The exact link between these factors is unclear. Here we show, using Magnetic Resonance Imaging as a tool to examine placental changes in mouse models of perturbed pregnancies, that T 2 contrast between distinct regions of the placenta is abolished at complete loss of blood flow. Alterations in T 2 (spin-spin or transverse) relaxation times are explained as a consequence of hypoxia and acidosis within the tissue. Similar changes are observed in perturbed pregnancies, indicating that acidosis as well as hypoxia may be a feature of pregnancy complications such as preeclampsia and may play a prominent role in the signalling pathways that lead to the increased secretion of anti-angiogenic compounds.


Maximal Respiratory Pressures and Maximum Voluntary Ventilation in Young Arabs: Association with Anthropometrics and Physical Activity.
Maximum inspiratory pressure (MIP), maximum expiratory pressure (MEP) and maximum voluntary ventilation (MVV) measurements assist in determining the respiratory muscle strength and endurance. These determinants of respiratory muscles vary significantly by age, gender, height, and ethnic origin. Normative values for maximum respiratory pressures (MRPs) and MVV would aid in evaluating respiratory muscle function in athletes, estimating performance, and assisting in rehabilitation. In addition, the reference values may aid in determining the efficacy of therapeutic interventions in young people with chronic respiratory diseases. The purpose of this study was to see how respiratory muscle strength indices correlated with anthropometric and physical activity characteristics in young Arabs. The study included 80 male volunteers and 85 female volunteers ranging in age from 18 to 30 years. MicroRPM was used to measure MIP and MEP, and pulmonary function test data, including MVV values, were recorded. All subjects completed the Global Physical Activity Questionnaire (GPAQ) and anthropometric measurements. Unpaired t-tests or Mann-Whitney U-tests were used to determine male-female differences. Using the Pearson correlation coefficient and Spearman Rho correlation coefficient tests, MIP and MEP values were correlated with body composition and physical activity. Using stepwise multiple linear regression analysis, the relationships between respiratory function (MVV, MIP, and MEP) and PFT values (FVC, FEV1, and FEV1/FVC), physical activity, and sedentary behavior were investigated. MIP, MEP, and MVV values were significantly lower in females than in males. MIP, MEP, and MVV values had a moderate correlation with forced vital capacity, forced expiratory volume in 1 second, and height, but not with weight, BMI, or GPAQ. Age, gender, and body mass index were found to be significant predictors of maximal respiratory pressures in a young Arab population. Maximum respiratory pressures and maximal voluntary ventilation were significantly lower in young Arabs than in other ethnic groups; these values were influenced by gender and height but not by levels of physical activity.


Osteopontin is up-regulated in chronic hepatitis C and is associated with cellular permissiveness for hepatitis C virus replication.
OPN (osteopontin)) is a Hh (Hedgehog)-regulated cytokine that is up-regulated during chronic liver injury and directly promotes fibrosis. We have reported that Hh signalling enhances viral permissiveness and replication in HCV (hepatitis C virus)-infected cells. Hence we hypothesized that OPN directly promotes HCV replication, and that targeting OPN could be beneficial in HCV. In the present study, we compared the expression of OPN mRNA and protein in HCV (JFH1)-infected Huh7 and Huh7.5 cells, and evaluated whether modulating OPN levels using exogenous OPN ligands (up-regulate OPN) or OPN-specific RNA-aptamers (neutralize OPN) leads to changes in HCV expression. Sera and livers from patients with chronic HCV were analysed to determine whether OPN levels were associated with disease severity or response to therapy. Compared with Huh7 cells, Huh7.5 cells support higher levels of HCV replication (15-fold) and expressed significantly more OPN mRNA (30-fold) and protein. Treating Huh7 cells with OPN ligands led to a dose-related increase in HCV (15-fold) and OPN (8-fold) mRNA. Conversely, treating Huh7.5 cells with OPN-specific RNA aptamers inhibited HCV RNA and protein by >50% and repressed OPN mRNA to basal levels. Liver OPN expression was significantly higher (3-fold) in patients with advanced fibrosis. Serum OPN positively correlated with fibrosis-stage (P=0.009), but negatively correlated with ETBCR (end-of-treatment biochemical response), ETVR (end-of-treatment virological response), SBCR (sustained biochemical response) and SVR (sustained virological response) (P=0.007). The OPN fibrosis score (serum OPN and presence of fibrosis ≥F2) may be a predictor of SVR. In conclusion, OPN is up-regulated in the liver and serum of patients with chronic hepatitis C, and supports increased viral replication. OPN neutralization may be a novel therapeutic strategy in chronic hepatitis C.


Stakeholder interaction within the ERICA Integrated Approach.
Within the ERICA project, stakeholder involvement has been addressed within three main areas: generic interactions throughout the project, specific consultation by means of attendance to events and considerations as part of the ERICA Integrated Approach and Assessment Tool. The word stakeholders meant namely any individual or group who may be affected by or have an interest in an issue, and to include experts, lay-people and the public. An End-Users-Group (EUG) was set up to facilitate the two-way dialogue between the ERICA Consortium and stakeholders. The ERICA EUG consisted of representatives of 60 organisations ranging from regulatory bodies, national advisory bodies, academia, non-governmental organisations, industry, consultants and inter-governmental organisations. Stakeholder interaction was included from the very start of the project. Inputs from the EUG were recorded and in most instances incorporated within the development of the project and thus influenced and helped to shape some of the ERICA deliverables.


MiR-34a expression has an effect for lower risk of metastasis and associates with expression patterns predicting clinical outcome in breast cancer.
MiR-34a acts as a candidate tumour suppressor gene, and its expression is reduced in several cancer types. We aimed to study miR-34a expression in breast cancer and its correlation with tumour characteristics and clinical outcome, and regulatory links with other genes. We analysed miR-34a expression in 1,172 breast tumours on TMAs. 25% of the tumours showed high, 43% medium and 32% low expression of miR-34a. High miR-34a expression associated with poor prognostic factors for breast cancer: positive nodal status (p = 0.006), high tumour grade (p<0.0001), ER-negativity (p = 0.0002), HER2-positivity (p = 0.0002), high proliferation rate (p<0.0001), p53-positivity (p<0.0001), high cyclin E (p<0.0001) and γH2AX (p<0.0001). However, multivariate analysis adjusting for conventional prognostic factors indicated that high miR-34a expression in fact associated with a lower risk of recurrence or death from breast cancer (HR = 0.63, 95% CI = 0.41-0.96, p = 0.031). Gene expression analysis by differential miR-34a expression revealed an expression signature with an effect on both the 5-year and 10-year survival of the patients (p<0.001). Functional genomic analysis highlighted a novel regulatory role of the transcription factor MAZ, apart from the known control by p53, on the expression of miR-34a and a number of miR-34a targets. Our findings suggest that while miR-34a expression activation is a marker of aggressive breast tumour phenotype it exerts an independent effect for a lower risk of recurrence or death from breast cancer. We also present an expression signature of 190 genes associated with miR-34a expression. Our analysis for regulatory loops suggest that MAZ and p53 transcription factors co-operate in modulating miR-34a, as well as miR-34a targets involved in several cellular pathways. Taken together, these results suggest that the network of genes co-regulated with and targeted by miR-34a form a group of down-stream effectors that maybe of use in predicting clinical outcome, and that highlight novel regulatory mechanisms in breast cancer.


Health care resource use and stroke outcome. Multinational comparisons within the GAIN International trial.
Outcome in patients hospitalized for acute stroke varies considerably between populations. Within the framework of the GAIN International trial, a large multicenter trial of a neuroprotective agent (gavestinel, glycine antagonist), stroke outcome in relation to health care resource use has been compared in a large number of countries, allowing for differences in case mix. This substudy includes 1,422 patients in 19 countries grouped into 10 regions. Data on prognostic variables on admission to hospital, resource use, and outcome were analyzed by regression models. All results were adjusted for differences in prognostic factors on admission (NIH Stroke Scale, age, comorbidity). There were threefold variations in the average number of days in hospital/institutional care (from 20 to 60 days). The proportion of patients who met with professional rehabilitation staff also varied greatly. Three-month case fatality ranged from 11% to 28%, and mean Barthel ADL score at three months varied between 64 and 73. There was no relationship between health care resource use and outcome in terms of survival and ADL function at three months. The proportion of patients living at home at three months did not show any relationship to ADL function across countries. There are wide variations in health care resource use between countries, unexplained by differences in case mix. Across countries, there is no obvious relationship between resource use and clinical outcome after stroke. Differences in health care traditions (treatment pathways) and social context seem to be major determinants of resource use. In making comparisons between countries, great care should be exercised in using outcome variables as indicators of quality of stroke care.


TET3 epigenetically controls feeding and stress response behaviors via AGRP neurons.
The TET family of dioxygenases promote DNA demethylation by oxidizing 5-methylcytosine to 5-hydroxymethylcytosine (5hmC). Hypothalamic agouti-related peptide-expressing (AGRP-expressing) neurons play an essential role in driving feeding, while also modulating nonfeeding behaviors. Besides AGRP, these neurons produce neuropeptide Y (NPY) and the neurotransmitter GABA, which act in concert to stimulate food intake and decrease energy expenditure. Notably, AGRP, NPY, and GABA can also elicit anxiolytic effects. Here, we report that in adult mouse AGRP neurons, CRISPR-mediated genetic ablation of Tet3, not previously known to be involved in central control of appetite and metabolism, induced hyperphagia, obesity, and diabetes, in addition to a reduction of stress-like behaviors. TET3 deficiency activated AGRP neurons, simultaneously upregulated the expression of Agrp, Npy, and the vesicular GABA transporter Slc32a1, and impeded leptin signaling. In particular, we uncovered a dynamic association of TET3 with the Agrp promoter in response to leptin signaling, which induced 5hmC modification that was associated with a chromatin-modifying complex leading to transcription inhibition, and this regulation occurred in both the mouse models and human cells. Our results unmasked TET3 as a critical central regulator of appetite and energy metabolism and revealed its unexpected dual role in the control of feeding and other complex behaviors through AGRP neurons.


Multiple deletions of small Cab-like proteins in the cyanobacterium Synechocystis sp. PCC 6803: consequences for pigment biosynthesis and accumulation.
Deletion of the genes for four or five small Cab-like proteins (SCPs) in photosystem (PS) I-less and PS I-less/PS II-less strains of Synechocystis sp. PCC 6803 caused a large decrease in the chlorophyll and carotenoid content of the cells without accumulation of early intermediates in the chlorophyll biosynthesis pathway, suggesting limited chlorophyll availability. The PS II/PS I ratio increased upon deletion of multiple SCPs in a wild type background, similar to what is observed in the presence of subsaturating concentrations of gabaculin, an inhibitor of an early step in the tetrapyrrole biosynthesis pathway. Upon deletion of multiple SCPs, neither 77 K fluorescence emission properties of phycobilisomeless thylakoids from the PS I-less/PS II-less strain nor the energy trapping efficiency of PS II were affected, indicating that under steady-state conditions SCPs do not bind much chlorophyll and do not serve as PS II antenna. Under conditions where protochlorophyllide reduction and thus chlorophyll synthesis were inhibited, chlorophyll disappeared quickly in a mutant lacking all five SCPs. This implies a role of SCPs in stabilization of chlorophyll-binding proteins and/or in reuse of chlorophylls. Under these conditions of inhibited reduction of protochlorophyllide, the accumulation kinetics of this intermediate were greatly altered in the absence of the five SCPs. This indicates an alteration of tetrapyrrole biosynthesis kinetics by SCPs. Based on this and other evidence, we propose that SCPs bind carotenoids and transiently bind chlorophyll, aiding in the supply of chlorophyll to nascent or reassembling photosynthetic complexes, and regulate the tetrapyrrole biosynthesis pathway as a function of the demand for chlorophyll.


Hypoxia and acidification have additive and synergistic negative effects on the growth, survival, and metamorphosis of early life stage bivalves.
Low oxygen zones in coastal and open ocean ecosystems have expanded in recent decades, a trend that will accelerate with climatic warming. There is growing recognition that low oxygen regions of the ocean are also acidified, a condition that will intensify with rising levels of atmospheric CO2. Presently, however, the concurrent effects of low oxygen and acidification on marine organisms are largely unknown, as most prior studies of marine hypoxia have not considered pH levels. We experimentally assessed the consequences of hypoxic and acidified water for early life stage bivalves (bay scallops, Argopecten irradians, and hard clams, Mercenaria mercenaria), marine organisms of significant economic and ecological value and sensitive to climate change. In larval scallops, experimental and naturally-occurring acidification (pH, total scale = 7.4-7.6) reduced survivorship (by >50%), low oxygen (30-50 µM) inhibited growth and metamorphosis (by >50%), and the two stressors combined produced additively negative outcomes. In early life stage clams, however, hypoxic waters led to 30% higher mortality, while acidified waters significantly reduced growth (by 60%). Later stage clams were resistant to hypoxia or acidification separately but experienced significantly (40%) reduced growth rates when exposed to both conditions simultaneously. Collectively, these findings demonstrate that the consequences of low oxygen and acidification for early life stage bivalves, and likely other marine organisms, are more severe than would be predicted by either individual stressor and thus must be considered together when assessing how ocean animals respond to these conditions both today and under future climate change scenarios.


Effects of acute exercise on long-term memory.
In this study, we tested the effect of acute exercise on long-term memory, specifically the timing of exercise relative to the memory challenge. We assessed memory via paragraph recall, in which participants listened to two paragraphs (exposure) and recounted them following a 35-min delay. Participants (n = 48) were randomly assigned to one of three groups: exercise prior to exposure, exercise after exposure, or no-exercise. Exercise consisted of 30 min on a cycle ergometer including 20 min at moderate intensity. Only the exercise prior group recalled significantly more than the control group (p < .05). Differences among the exercise groups failed to reach significance (p = .09). Results indicated that acute exercise positively influenced recall and that exercise timing relative to memory task may have an impact on this effect.


A phosphate binding pocket is a key determinant of exo- versus endo-nucleolytic activity in the SNM1 nuclease family.
The SNM1 nucleases which help maintain genome integrity are members of the metallo-β-lactamase (MBL) structural superfamily. Their conserved MBL-β-CASP-fold SNM1 core provides a molecular scaffold forming an active site which coordinates the metal ions required for catalysis. The features that determine SNM1 endo- versus exonuclease activity, and which control substrate selectivity and binding are poorly understood. We describe a structure of SNM1B/Apollo with two nucleotides bound to its active site, resembling the product state of its exonuclease reaction. The structure enables definition of key SNM1B residues that form contacts with DNA and identifies a 5' phosphate binding pocket, which we demonstrate is important in catalysis and which has a key role in determining endo- versus exonucleolytic activity across the SNM1 family. We probed the capacity of SNM1B to digest past sites of common endogenous DNA lesions and find that base modifications planar to the nucleobase can be accommodated due to the open architecture of the active site, but lesions axial to the plane of the nucleobase are not well tolerated due to constriction around the altered base. We propose that SNM1B/Apollo might employ its activity to help remove common oxidative lesions from telomeres.


Novel Immunoreactive glycoprotein orthologs of Ehrlichia spp.
We previously identified a strongly immunoreactive 43 kDa protein (p43) of Ehrlichia canis. As an immunodiagnostic antigen, the p43 had a 96% accuracy as compared with IFA and provided species-specific diagnosis of E. canis infections. Further investigation has revealed that the E. canis p43 represents the N-terminal portion of the largest immunoreactive protein described in Ehrlichia spp. with a predicted molecular mass of 153 kDa. Analysis of the recombinant N-terminal region (p43) of the p153 by protein gel electrophoresis demonstrated a larger than predicted molecular mass (approximately 30%), and presence of carbohydrate glycans, indicating that the p153 is a glycoprotein. A BLASTn search was performed on the E. chaffeensis genome sequence (95%), and the gene encoding the p153 ortholog was identified in E. chaffeensis. The E. canis p153 (4,263 bp) and E. chaffeensis p156 (4,389 bp) genes had similar chromosomal locations, downstream of the homologous (approximately 87%) deoxyguanosine-triphosphate triphosphohydrolase genes, and homologous (approximately 90%) intergenic sequences preceding the open reading frames. Nucleic acid sequence homology (52%) observed between the glycoprotein genes supported previous findings with regard to genetic divergence of the p43 gene fragment, and the p153 and p156 proteins had amino acid similarity of 32%. A native E. canis protein with a molecular mass of 200 kDa reacted with antisera produced against the N-terminal region (p43) of the p153, suggesting that the native protein was posttranslationally modified. Similarly, recombinant constructs of E. chaffeensis p156 migrated larger than predicted (approximately 200 kDa), and carbohydrate was detected on the recombinant proteins. The chromosomal location, amino acid homology, and biophysical properties support the conclusion that the p153 and p156 glycoproteins (designated gp200s) are species-specific immunoreactive orthologs.


Coupling of DNA replication and mitosis by fission yeast rad4/cut5.
The fission yeast cut5+ (identical to rad4+) gene is essential for S phase. Its temperature-sensitive (ts) mutation causes mitosis while S phase is inhibited: dependence of mitosis upon the completion of S phase is abolished. If DNA is damaged in mutant cells, however, cell division is arrested. Thus the checkpoint control system for DNA damage is functional, while that for DNA synthesis inhibition is not in the cut5 mutants. Transcription of the cut5+ gene is not under the direct control of cdc10+, which encodes a transcription factor for the START of cell cycle. The transcript level does not change during the cell cycle. The protein product has four distinct domains and is enriched in the nucleus. Its level does not alter during the cell cycle. The N-domain is important for cut5 protein function: it is essential for complementation of ts cut5 mutations and its overexpression blocks cell division. Furthermore, it resembles the N-terminal repeat domain of proto-oncoprotein Ect2, which, in the C-domain, contains a regulator-like sequence for small G proteins. We discuss a hypothesis that the cut5 protein is an essential component of the checkpoint control system for the completion of DNA synthesis. The restraint of mitosis until the completion of S phase is mediated by the cut5 protein, which can sense the state of chromosome duplication and negatively interacts with M phase regulators such as cdc25 and cdc2.


Pragmatic RAndomised controlled trial of a trauma-focused guided self-help Programme versus InDividual trauma-focused cognitive Behavioural therapy for post-traumatic stress disorder (RAPID): trial protocol.
There is good evidence that trauma-focused therapies for Post-Traumatic Stress Disorder are effective. However, they are not always feasible to deliver due a shortage of trained therapists and demands on the patient. An online trauma-focused Guided Self-Help (GSH) programme which could overcome these barriers has shown promise in a pilot study. This study will be the first to evaluate GSH against standard face-to-face therapy to assess its suitability for use in the NHS. The study is a large-scale multi-centre pragmatic randomised controlled non-inferiority trial, with assessors masked to treatment allocation. One hundred and ninety-two participants will be randomly allocated to receive either face-to-face trauma-focused cognitive behaviour therapy (TFCBT) or trauma-focused online guided self-help (GSH). The primary outcome will be the severity of symptoms of PTSD over the previous week as measured by the Clinician Administered PTSD Scale for DSM5 (CAPS-5) at 16 weeks post-randomisation. Secondary outcome measures include PTSD symptoms over the previous month as measured by the CAPS-5 at 52 weeks plus the Impact of Event Scale - revised (IES-R), Work and Social Adjustment Scale (WSAS), Patient Health Questionnaire-9 (PHQ-9), General Anxiety Disorder-7 (GAD-7), Alcohol Use Disorders Test (AUDIT-O), Multidimensional Scale for Perceived Social Support (MSPSS), short Post-Traumatic Cognitions Inventory (PTCI), Insomnia Severity Index (ISI) and General Self Efficacy Scale (GSES) measured at 16 and 52 weeks post-randomisation. Changes in health-related quality of life will be measured by the EQ-5D and the level of healthcare resource utilisation for health economic analysis will be determined by an amended version of the Client Socio-Demographic and Service Receipt Inventory European Version. The Client Satisfaction Questionnaire (CSQ) will be collected at 16 weeks post-randomisation to evaluate treatment satisfaction. This study will be the first to compare online GSH with usual face-to-face therapy for PTSD. The strengths are that it will test a rigorously developed intervention in a real world setting to inform NHS commissioning. The potential challenges of delivering such a pragmatic study may include participant recruitment, retention and adherence, therapist retention, and fidelity of intervention delivery. ISRCTN13697710 registered on 20/12/2016.