Quantifying Risk for Anxiety Disorders in Preschool Children: A Machine Learning Approach.
Early childhood anxiety disorders are common, impairing, and predictive of anxiety and mood disorders later in childhood. Epidemiological studies over the last decade find that the prevalence of impairing anxiety disorders in preschool children ranges from 0.3% to 6.5%. Yet, less than 15% of young children with an impairing anxiety disorder receive a mental health evaluation or treatment. One possible reason for the low rate of care for anxious preschoolers is the lack of affordable, timely, reliable and valid tools for identifying young children with clinically significant anxiety. Diagnostic interviews assessing psychopathology in young children require intensive training, take hours to administer and code, and are not available for use outside of research settings. The Preschool Age Psychiatric Assessment (PAPA) is a reliable and valid structured diagnostic parent-report interview for assessing psychopathology, including anxiety disorders, in 2 to 5 year old children. In this paper, we apply machine-learning tools to already collected PAPA data from two large community studies to identify sub-sets of PAPA items that could be developed into an efficient, reliable, and valid screening tool to assess a young child's risk for an anxiety disorder. Using machine learning, we were able to decrease by an order of magnitude the number of items needed to identify a child who is at risk for an anxiety disorder with an accuracy of over 96% for both generalized anxiety disorder (GAD) and separation anxiety disorder (SAD). Additionally, rather than considering GAD or SAD as discrete/binary entities, we present a continuous risk score representing the child's risk of meeting criteria for GAD or SAD. Identification of a short question-set that assesses risk for an anxiety disorder could be a first step toward development and validation of a relatively short screening tool feasible for use in pediatric clinics and daycare/preschool settings.


Survival, morbidity, growth and developmental delay for babies born preterm in low and middle income countries - a systematic review of outcomes measured.
Premature birth is the leading cause of neonatal death and second leading in children under 5. Information on outcomes of preterm babies surviving the early neonatal period is sparse although it is considered a major determinant of immediate and long-term morbidity. Systematic review of studies reporting outcomes for preterm babies in low and middle income settings was conducted using electronic databases, citation tracking, expert recommendations and "grey literature". Reviewers screened titles, abstracts and articles. Data was extracted using inclusion and exclusion criteria, study site and facilities, assessment methods and outcomes of mortality, morbidity, growth and development. The Child Health Epidemiology Reference Group criteria (CHERG) were used to assess quality. Of 197 eligible publications, few (10.7%) were high quality (CHERG). The majority (83.3%) report on the outcome of a sample of preterm babies at time of birth or admission. Only 16.0% studies report population-based data using standardised mortality definitions. In 50.5% of studies, gestational age assessment method was unclear. Only 15.8% followed-up infants for 2 years or more. Growth was reported using standardised definitions but recommended morbidity definitions were rarely used. The criteria for assessment of neurodevelopmental outcomes was variable with few standardised tools - Bayley II was used in approximately 33% of studies, few studies undertook sensory assessments. To determine the relative contribution of preterm birth to the burden of disease in children and to inform the planning of healthcare interventions to address this burden, a renewed understanding of the assessment and documentation of outcomes for babies born preterm is needed. More studies assessing outcomes for preterm babies who survive the immediate newborn period are needed. More consistent use of data is vital with clear and aligned definitions of health outcomes in newborn (preterm or term) and intervention packages aimed to save lives and improve health.


Periendoscopic management of direct oral anticoagulants: a prospective cohort study.
To assess the frequency of adverse events associated with periendoscopic management of direct oral anticoagulants (DOACs) in patients undergoing elective GI endoscopy and the efficacy and safety of the British Society of Gastroenterology (BSG) and European Society of Gastrointestinal Endoscopy (ESGE) recommendations (NCT02734316). Consecutive patients on DOACs scheduled for elective GI endoscopy were prospectively included. The timing of DOAC interruption and resumption before and after the procedures were recorded, along with clinical and procedural data. Procedures were stratified into low-risk and high-risk for GI-related bleeding, and patients into low-risk and high-risk for thromboembolic events. Patients were followed-up for 30 days for major and clinically relevant non-major bleeding events (CRNMB), arterial and venous thromboembolism and death. Of 529 patients, 38% and 62% underwent high-risk and low-risk procedures, respectively. There were 45 (8.5%; 95% CI 6.3% to 11.2%) major or CRNMB events and 2 (0.4%; 95% CI 0% to 1.4%) thromboembolic events (transient ischaemic attacks). Overall, the incidence of bleeding events was 1.8% (95% CI 0.7% to 4%) and 19.3% (95% CI 14.1% to 25.4%) in low-risk and high-risk procedures, respectively. For high-risk procedures, the incidence of intraprocedural bleeding was similar in patients who interrupted anticoagulation according to BSG/ESGE guidelines or earlier (10.3%vs10.8%, p=0.99), with a trend for a lower risk as compared with those who stopped anticoagulation later (10.3%vs25%, p=0.07). The incidence of delayed bleeding appeared similar in patients who resumed anticoagulation according to BSG/ESGE guidelines or later (6.6%vs7.7%, p=0.76), but it tended to increase when DOAC was resumed earlier (14.4%vs6.6%, p=0.27). The risk of delayed major bleeding was significantly higher in patients receiving heparin bridging than in non-bridged ones (26.6%vs5.9%, p=0.017). High-risk procedures in patients on DOACs are associated with a substantial risk of bleeding, further increased by heparin bridging. Adoption of the BSG/ESGE guidelines in periendoscopic management of DOACs seems to result in a favourable benefit/risk ratio. NCT02734316; Pre-results.


BE-ACTIV: a staff-assisted behavioral intervention for depression in nursing homes.
This article (a) describes a 10-week, behavioral, activities-based intervention for depression that can be implemented in nursing homes collaboratively with nursing home activities staff and (b) presents data related to its development, feasibility, and preliminary outcomes. We developed BE-ACTIV, which stands for Behavioral Activities Intervention, in two pilot study phases: a treatment development phase and a feasibility-outcome phase with a small, randomized trial. We first piloted the intervention with five depressed residents in a single nursing home in collaboration with the social services and activities staff. The second phase randomized 20 residents from six nursing homes to receive either the intervention or treatment as usual. The intervention was well received by residents, family, and staff members. Experience with the intervention and input from staff members resulted in modifications to streamline the intervention and improve implementation. Results suggest that BE-ACTIV reduced institutional barriers to participation in pleasant activities, increased resident control over activity participation, increased overall activity participation, and improved depressive symptoms. Despite low power, statistical and graphical comparisons suggest superiority of the intervention over treatment as usual. Because depression among nursing home residents is prevalent, heterogeneous, and often treatment resistant, there is a need for effective, low-cost interventions that are ecologically acceptable and efficient. BE-ACTIV is a promising intervention; it is brief, addresses institutional barriers, involves facility staff in treatment, and is acceptable to residents. As such, BE-ACTIV merits further evaluation to establish efficacy and effectiveness.


Monitoring and modeling wetland chloride concentrations in relationship to oil and gas development.
Extraction of oil and gas via unconventional methods is becoming an important aspect of energy production worldwide. Studying the effects of this development in countries where these technologies are being widely used may provide other countries, where development may be proposed, with some insight in terms of concerns associated with development. A fairly recent expansion of unconventional oil and gas development in North America provides such an opportunity. Rapid increases in energy development in North America have caught the attention of managers and scientists as a potential stressor for wildlife and their habitats. Of particular concern in the Northern Great Plains of the U.S. is the potential for chloride-rich produced water associated with unconventional oil and gas development to alter the water chemistry of wetlands. We describe a landscape scale modeling approach designed to examine the relationship between potential chloride contamination in wetlands and patterns of oil and gas development. We used a spatial Bayesian hierarchical modeling approach to assess multiple models explaining chloride concentrations in wetlands. These models included effects related to oil and gas wells (e.g. age of wells, number of wells) and surficial geology (e.g. glacial till, outwash). We found that the model containing the number of wells and the surficial geology surrounding a wetland best explained variation in chloride concentrations. Our spatial predictions showed regions of localized high chloride concentrations. Given the spatiotemporal variability of regional wetland water chemistry, we do not regard our results as predictions of contamination, but rather as a way to identify locations that may require more intensive sampling or further investigation. We suggest that an approach like the one outlined here could easily be extended to more of an adaptive monitoring approach to answer questions about chloride contamination risk that are of interest to managers.


A phase II study of dasatinib in patients with chemosensitive relapsed small cell lung cancer (Cancer and Leukemia Group B 30602).
SRC is an oncogene with an essential role in the invasiveness and metastasis of solid tumors including small cell lung cancer. Dasatinib is a potent inhibitor of SRC as well as other tyrosine kinases. The primary objective of this study was to determine the efficacy of second-line dasatinib in patients with chemosensitive (relapse or progression > or =90 days after completing first-line therapy) small cell lung cancer. Patients with measurable disease; performance status 0 to 1; no more than one prior platinum-based chemotherapy regimen; and adequate hematologic, hepatic, and renal function were eligible. Dasatinib was administered orally at 70 mg twice daily continuously (1 cycle = 21 days) until disease progression or unacceptable toxicity. Response was determined after every two cycles. Patients were followed until disease progression or death. The study was prospectively designed to simultaneously discriminate between complete plus partial response rates of 5% versus 20% and progression-free survival (PFS) rates at 6 weeks of 50% versus 70.7% in 53 evaluable patients with at least 92% power. The study was to be terminated early and declared negative if 1 or less objective response and 14 or fewer instances of PFS > or =6 weeks were observed among the initial 27 patients; however, patient accrual continued while the initial 27 patients were evaluated. Between April 2007 and December 2008, 45 patients were enrolled, but one patient never received any protocol therapy and one patient was ineligible: male/female, 17/26; white/black/unknown, 40/2/1; median age, 64 years (range, 35-84 years); and performance status 0/1, 12/31. No objective response was recorded among the 43 eligible and treated patients. Among the initial 27 patients, only 13 instances of PFS > or =6 weeks were observed. With a median follow-up time of 7.1 months, median estimated overall survival and PFS times for the 43 eligible and treated patients were 17.0 and 5.9 weeks, respectively. Common reasons for removal of patients from protocol treatment were progressive disease (65%) and adverse events (26%). Toxicity was generally mild to moderate: grade 3 events of >5% frequency included fatigue and pleural and pericardial effusions; and no grade 4 or 5 events were encountered. Dasatinib did not reach our specified efficacy criteria in this clinical setting, and the study was terminated.


PCR artifact in testing for homologous recombination in genomic editing in zebrafish.
We report a PCR-induced artifact in testing for homologous recombination in zebrafish. We attempted to replace the lnx2a gene with a donor cassette, mediated by a TALEN induced double stranded cut. The donor construct was flanked with homology arms of about 1 kb at the 5' and 3' ends. Injected embryos (G0) were raised and outcrossed to wild type fish. A fraction of the progeny appeared to have undergone the desired homologous recombination, as tested by PCR using primer pairs extending from genomic DNA outside the homology region to a site within the donor cassette. However, Southern blots revealed that no recombination had taken place. We conclude that recombination happened during PCR in vitro between the donor integrated elsewhere in the genome and the lnx2a locus. We conclude that PCR alone may be insufficient to verify homologous recombination in genome editing experiments in zebrafish.


Contraceptive use by women across different sexual orientation groups.
To examine contraceptive methods used across sexual orientation groups. We collected data from 118,462 female participants in two longitudinal cohorts-the Nurses' Health Study (NHS) 2 (founded in 1989, participants born 1947-1964) and NHS3 (founded in 2010, born 1965-1995). We used log-binomial models to estimate contraceptive methods ever used across sexual orientation groups and cohorts, adjusting for age and race. Lesbians were the least likely of all sexual orientation groups to use any contraceptive method. Lesbians in NHS2 were 90% less likely than heterosexuals to use long-acting reversible contraceptives (LARCs; adjusted risk ratio [aRR]; 95% confidence interval [CI]: 0.10 [0.04, 0.26]) and results were similar for other contraceptive methods and in the NHS3 cohort. Compared to the reference group of completely heterosexual participants with no same-sex partners, those who identified as completely heterosexual with same-sex partners, mostly heterosexual, or bisexual were generally more likely to use any method of contraception. Use of LARCs was especially striking across sexual minority groups, and, with the exception of lesbians, they were more likely to use LARCs; as one illustration, NHS3 bisexuals were more than twice as likely to use LARCs (aRR [95% CI]: 2.01 [1.67, 2.42]). While certain sexual minority subgroups (e.g., bisexuals) were more likely than heterosexuals to use contraceptive methods such as LARCs, lesbians were less likely to use any method. Many sexual minority patients need contraceptive counseling and providers should ensure to offer this counseling to patients in need, regardless of sexual orientation.


Macromolecular charge and reticuloendothelial function: comparison between the kinetics of administered native and cationized ferritins and the corresponding immune complexes in the mouse.
In order to evaluate the role of macromolecular charge on uptake by the reticuloendothelial system (RES), kinetic studies were carried out following the intravenous administration of 125I-labelled native ferritin (NF, pI 4.5) or cationized ferritin (CF, pI 7) to Swiss-Webster female mice subsequently killed at 2, 4, 8, 24 and 36 hr later. The same experiments were performed following the administration of radio-labelled (125I) native ferritin immune complexes (NFIC, pI 5-6.5) and cationized ferritin immune complexes (CFIC, pI 6.5-7.5). These complexes were prepared in five-times antigen excess by combination of affinity-purified anti-ferritin IgG-125I with NF or CF. A striking difference between the plasma clearance of NF and that of CF was observed in that the former was rapidly eliminated within 8 hr whereas the latter persisted in the circulation at 24 hr. This was associated with a significant increase in the uptake of NF by the liver, spleen, and kidney. No differences were observed in blood cell-associated radioactivity. Immunohistochemical studies confirmed the presence of increased amounts of NF in Kupffer cells and splenic phagocytes. Thus, the uptake of ferritin by components of the RES is highly dependent upon its pI. The present data may be explained by differences in diffusibility of CF and NF or alternatively by differential interactions with the cell surface in vivo. Contrary to the prior investigation carried out with the antigens alone, the plasma clearance and organ (liver, spleen and kidney) kinetic studies of NFIC and CFIC were similar. In addition, immunohistochemical studies demonstrated that the uptakes of NFIC (pI 5-6.5), CFIC (pI 6.5-7.5) and CFIC (pI 7-9) by Kupffer cells and splenic phagocytes were similar. As further confirmation for similarity in binding to Fc receptors of human polymorphonuclear leucocytes, Scatchard analysis failed to demonstrate any differences between NFIC and CFIC. These studies provide evidence that, within the range employed in this investigation, the charge of ferritin within the immune complex (and hence the charge of the complex itself) does not affect its uptake by receptors of phagocytic cells. In contrast, the uptake of ferritin, which is not Fc or C3 receptor dependent, is clearly conditioned by electrostatic charge.


Karyotype instability in the ponerine ant genus Diacamma.
The queenless ponerine ant Diacamma ceylonense and a population of Diacamma from the Nilgiri hills which we refer to as 'nilgiri', exhibit interesting similarities as well as dissimilarities. Molecular phylogenetic study of these morphologically almost similar taxa has shown that D. ceylonense is closely related to 'nilgiri' and indicates that 'nilgiri' is a recent diversion in the Diacamma phylogenetic tree. However, there is a striking behavioural difference in the way reproductive monopoly is maintained by the respective gamergates (mated egg laying workers), and there is evidence that they are genetically differentiated, suggesting a lack of gene flow. To develop a better understanding of the mechanism involved in speciation of Diacamma, we have analysed karyotypes of D. ceylonense and 'nilgiri'. In both, we found surprising inter-individual and intra-individual karyotypic mosaicism. The observed numerical variability, both at intra-individual and inter-individual levels, does not appear to have hampered the sustainability of the chromosomal diversity in each population under study. Since the related D. indicum displays no such intra-individual or inter-individual variability whatsoever under identical experimental conditions, these results are unlikely to be artifacts. Although no known mechanisms can account for the observed karyotypic variability of this nature, we believe that the present findings on the ants under study would provide opportunities for exciting new discoveries concerning the origin, maintenance and significance of intra-individual and inter-individual karyotypic mosaicism.


Mesenchymal stem cells enhance recovery and repair following ventilator-induced lung injury in the rat.
Bone-marrow derived mesenchymal stem cells (MSCs) reduce the severity of evolving acute lung injury (ALI), but their ability to repair the injured lung is not clear. A study was undertaken to determine the potential for MSCs to enhance repair after ventilator-induced lung injury (VILI) and elucidate the mechanisms underlying these effects. Anaesthetised rats underwent injurious ventilation which produced severe ALI. Following recovery, they were given an intravenous injection of MSCs (2×10(6) cells) or vehicle immediately and a second dose 24 h later. The extent of recovery following VILI was assessed after 48 h. Subsequent experiments examined the potential for non-stem cells and for the MSC secretome to enhance VILI repair. The contribution of specific MSC-secreted mediators was then examined in a wound healing model. MSC therapy enhanced repair following VILI. MSCs enhanced restoration of systemic oxygenation and lung compliance, reduced total lung water, decreased lung inflammation and histological lung injury and restored lung structure. They attenuated alveolar tumour necrosis factor α concentrations while increasing concentrations of interleukin 10. These effects were not seen with non-stem cells (ie, rat fibroblasts). MSC-secreted products also enhanced lung repair and attenuated the inflammatory response following VILI. The beneficial effect of the MSC secretome on repair of pulmonary epithelial wounds was attenuated by prior depletion of keratinocyte growth factor. MSC therapy enhances lung repair following VILI via a paracrine mechanism that may be keratinocyte growth factor-dependent.


Centrosome loss or amplification does not dramatically perturb global gene expression in Drosophila.
Centrosome defects are a common feature of many cancers, and they can predispose fly brain cells to form tumours. In flies, centrosome defects perturb the asymmetric division of the neural stem cells, but it is unclear how this might lead to malignant transformation. One possibility is that centrosome defects might also perturb cellular homeostasis: for example, stress pathways are often activated in response to centrosome defects in cultured cells, and stress contributes to tumourigenesis in some fly models. Here we attempt to assess whether centrosome loss or centrosome amplification perturbs cell physiology in vivo by profiling the global transcriptome of Drosophila larval brains and imaginal discs that either lack centrosomes or have too many centrosomes. Surprisingly, we find that centrosome loss or amplification leads to few changes in the transcriptional profile of these cells, indicating that centrosome defects are surprisingly well tolerated by these cells. These observations indicate that centrosome defects can predispose fly brain cells to form tumours without, at least initially, dramatically altering their physiology.


Efficacy of the Social Cognition Training Program in a Sample of Outpatients with Schizophrenia.
Social cognition is recognized to be a deficit in individuals suffering from schizophrenia. Numerous studies have explored the relationship between social cognition and social functioning in outpatients with schizophrenia through the use of different social cognition training programs. This study examines the efficacy of the Social Cognition Training Program (PECS in Spanish) in adults with a diagnosis of schizophrenia. Data were derived from a sample of 44 non-hospitalized adult patients who presented with a DSM-IV-TR Axis I diagnosis of schizophrenia and 39 healthy controls. The 44 patients were divided into an experimental group (n=20) and a control task group (n=24) that received cognitive training. Healthy controls did not receive any treatment. Sociodemographic and clinical variables correlates were computed. The 2-way ANOVA was conducted to examine differences between groups in pre- and post-treatment measures. Intragroup differences were explored using the paired-samples t-test. At the end of the training, patients in the experimental group showed a higher performance compared to patients in the control task group in the Hinting Task Test and in the emotion recognition of sadness, anger, fear, and disgust. The PECS proved to be effective in the improvement of some areas of theory of mind and emotion recognition in outpatients with schizophrenia. The PECS is one of the first programs developed in Spanish to train social cognition, and the data obtained support the importance of expanding the social cognition programs to non-English language samples.


Different neural and cognitive response to emotional faces in healthy monozygotic twins at risk of depression.
Negative cognitive bias and aberrant neural processing of emotional faces are trait-marks of depression. Yet it is unclear whether these changes constitute an endophenotype for depression and are also present in healthy individuals with hereditary risk for depression. Thirty healthy, never-depressed monozygotic (MZ) twins with a co-twin history of depression (high risk group: n = 13) or without co-twin history of depression (low-risk group: n = 17) were enrolled in a functional magnetic resonance imaging (fMRI) study. During fMRI, participants viewed fearful and happy faces while performing a gender discrimination task. After the scan, they were given a faces dot-probe task, a facial expression recognition task and questionnaires assessing mood, personality traits and coping strategies. High-risk twins showed increased neural response to happy and fearful faces in dorsal anterior cingulate cortex (ACC), dorsomedial prefrontal cortex (dmPFC), pre-supplementary motor area and occipito-parietal regions compared to low-risk twins. They also displayed stronger negative coupling between amygdala and pregenual ACC, dmPFC and temporo-parietal regions during emotional face processing. These task-related changes in neural responses in high-risk twins were accompanied by impaired gender discrimination performance during face processing. They also displayed increased attention vigilance for fearful faces and were slower at recognizing facial expressions relative to low-risk controls. These effects occurred in the absence of differences between groups in mood, subjective state or coping. Different neural response and functional connectivity within fronto-limbic and occipito-parietal regions during emotional face processing and enhanced fear vigilance may be key endophenotypes for depression.


Phase I/II trial of helical IMRT-based stereotactic body radiotherapy for hepatocellular carcinoma.
To report the results of a phase I/II study of helical IMRT-based stereotactic body radiotherapy (SBRT) for hepatocellular carcinoma (HCC). Eligibility included Child-Turcotte-Pugh class A or B, ≤3 lesions, and cumulative tumor diameter ≤6 cm. Dose was escalated from 36 Gy to 60 Gy delivered in 4 fractions. Grade ≥3 gastrointestinal toxicities (CTCAE v3.0) or radiation-induced liver disease defined dose-limiting toxicity (DLT). Thirty-two patients were enrolled: seven in dose levels 1-2 (36-44 Gy) and 25 in levels 3-4 (42-60 Gy). Failures included 1 local, 14 outfield intrahepatic, 2 distant, 1 concurrent local and outfield, 1 concurrent outfield and distant, and 1 concurrent local, outfield, and distant. Nine had grade 3 hematologic toxicities and 5 had grade 2 hepatic toxicities; no patient experienced DLT. Two-year local control (LFFS), outfield intrahepatic control (OutFFS), and overall survival (OS) rates were 80.9%, 46.7%, and 81.3%, respectively. Dose levels 3-4 and pre-radiotherapy multi-segment recurrence were independent prognostic factors for LFFS and OutFFS, respectively. Two-year LFFS, OutFFS, and OS were significantly higher for patients who were treated with dose-levels 3/4 for tumor(s) involving single segment compared with the rest of the patients. Helical IMRT-based SBRT was safe and effective, and patients with multi-segment recurrences prior to SBRT need to be closely followed.


Effects of chronic scopolamine administration on spatial working memory and hippocampal receptors related to learning.
Scopolamine has been used in neuropsychopharmacology as a standard drug that leads to symptoms mimicking cognitive deficits seen during the aging process in healthy humans and animals. Scopolamine is known to be a nonselective muscarinic receptor blocker, but its chronic effect on the expression of certain hippocampal receptors is not clear. The aim of the present study was to determine the effect of chronic scopolamine administration on hippocampal receptor expression and spatial working memory in two different learning tasks, the water maze and the eight-arm radial maze. Male rats (8-12 months) were trained in both tasks. Subsequently, different groups received physiological saline or 0.1, 0.8, or 2 mg/kg scopolamine hydrobromide, respectively, for 15 days. After drug administration, the rats were retested for both tasks, and hippocampal expressions of NR2A, NR2B, nAChRα7, and mAChRM1 receptors were assessed by western blotting analysis. In both tasks, the spatial working memory was decreased dose dependently in all groups compared with the control group. In terms of receptor expressions, 0.8 and 2 mg/kg scopolamine administration significantly decreased NR2A protein expression, which corroborates suggestions of an interaction between cholinergic and glutamatergic receptors in the hippocampus.


Characterization of phenotypic changes in Pseudomonas putida in response to surface-associated growth.
The formation of complex bacterial communities known as biofilms begins with the interaction of planktonic cells with a surface. A switch between planktonic and sessile growth is believed to result in a phenotypic change in bacteria. In this study, a global analysis of physiological changes of the plant saprophyte Pseudomonas putida following 6 h of attachment to a silicone surface was carried out by analysis of protein profiles and by mRNA expression patterns. Two-dimensional (2-D) gel electrophoresis revealed 15 proteins that were up-regulated following bacterial adhesion and 30 proteins that were down-regulated. N-terminal sequence analyses of 11 of the down-regulated proteins identified a protein with homology to the ABC transporter, PotF; an outer membrane lipoprotein, NlpD; and five proteins that were homologous to proteins involved in amino acid metabolism. cDNA subtractive hybridization revealed 40 genes that were differentially expressed following initial attachment of P. putida. Twenty-eight of these genes had known homologs. As with the 2-D gel analysis, NlpD and genes involved in amino acid metabolism were identified by subtractive hybridization and found to be down-regulated following surface-associated growth. The gene for PotB was up-regulated, suggesting differential expression of ABC transporters following attachment to this surface. Other genes that showed differential regulation were structural components of flagella and type IV pili, as well as genes involved in polysaccharide biosynthesis. Immunoblot analysis of PilA and FliC confirmed the presence of flagella in planktonic cultures but not in 12- or 24-h biofilms. In contrast, PilA was observed in 12-h biofilms but not in planktonic culture. Recent evidence suggests that quorum sensing by bacterial homoserine lactones (HSLs) may play a regulatory role in biofilm development. To determine if similar protein profiles occurred during quorum sensing and during early biofilm formation, HSLs extracted from P. putida and pure C(12)-HSL were added to 6-h planktonic cultures of P. putida, and cell extracts were analyzed by 2-D gel profiles. Differential expression of 16 proteins was observed following addition of HSLs. One protein, PotF, was found to be down-regulated by both surface-associated growth and by HSL addition. The other 15 proteins did not correspond to proteins differentially expressed by surface-associated growth. The results presented here demonstrate that P. putida undergoes a global change in gene expression following initial attachment to a surface. Quorum sensing may play a role in the initial attachment process, but other sensory processes must also be involved in these phenotypic changes.


Direct-to-consumer personal genome testing and cancer risk prediction.
The last several years has witnessed an explosion in genomics, with the advent of genome-wide association studies revealing hundreds of DNA variants significantly associated with most common diseases, including cancer. On the heels of these scientific advances came the direct-to-consumer (DTC) genetic testing industry. Genome-wide scans for disease have been marketed and sold directly to the public, without the involvement of a health care provider. Unlike genetic testing for mutations in known hereditary cancer susceptibility genes such as BRCA1/2, these genomic profiles examine DNA variants, which typically have a minimal risk impact, and account for only a fraction of the heritable component of cancer. Furthermore, risk information provided to consumers does not account for family history or other known risk factors. The clinical validity and utility of personal genome scans for disease risk prediction remain for the most part unestablished, although some argue lack of evidence of harm and the possibility that positive impacts on health behaviors or genetic awareness may result from consumer use. The DTC genetic testing industry has sparked significant controversy not only among the scientific community, but also among professional societies and government agencies.In this review, we present some of the history and methodological considerations of DTC genomic profiling, with a focus on cancer risk prediction. The literature regarding consumer awareness and utilization is explored, including understanding, expectations, and behavioral and psychological responses to DTC genomic risk prediction. Primary care provider and genetic professional knowledge and perceptions of DTC genomic profiling are also addressed. Ethical and scientific controversy surrounding the DTC genetic testing industry is presented, along with policy recommendations, regulatory actions, and the changing landscape of the DTC genetic testing market in response. Although our understanding of the human genome holds much promise in the realm of cancer prevention and treatment, DTC genomic profiling for cancer risk prediction is unlikely in its current form to have any significant impact on the health of the public. Time will tell if the next venture in genomic medicine, whole genome sequencing, will be accompanied by the translational research and emphasis on public/provider education required to ensure its successful application toward reducing the burden of cancer at a population level.


Alien species in aquaculture and biodiversity: a paradox in food production.
Aquaculture is seen as an alternative to meeting the widening gap in global rising demand and decreasing supply for aquatic food products. Asia, the epicenter of the global aquaculture industry, accounts for over 90% of the global aquaculture production quantity and about 80% of the value. Asian aquaculture, as with global aquaculture, is dependent to a significant extent on alien species, as is the case for all the major food crops and husbanded terrestrial animals. However, voluntary and or accidental introduction of exotic aquatic species (alien species) is known to negatively impact local biodiversity. In this relatively young food production industry, mitigating the dependence on alien species, and thereby minimizing potential negative impacts on biodiversity, is an imperative for a sustainable future. In this context an attempt is made in this synthesis to understand such phenomena, especially with reference to Asian inland finfish, the mainstay of global aquaculture production. It is pointed out that there is potential for aquaculture, which is becoming an increasingly important food production process, not to follow the past path of terrestrial food crops and husbanded animals in regard to their negative influences on biodiversity.


Regulator of telomere elongation helicase 1 (RTEL1) rs6010620 polymorphism contribute to increased risk of glioma.
Regulator of telomere elongation helicase 1 (RTEL1) is critical for genome stability and tumor avoidance. Many studies have reported the associations of RTEL1 rs6010620 with glioma risk, but individually published results were inconclusive. This meta-analysis was performed to quantitatively summarize the evidence for such a relationship. The PubMed, Embase, and Web of Science were systematically searched to identify relevant studies. The odds ratio (OR) and 95 % confidence interval (95 % CI) were computed to estimate the strength of the association using a fixed or random effects model. Ten studies were eligible for meta-analysis including data on glioma with 6,490 cases and 9,288 controls. Overall, there was a significant association between RTEL1 rs6010620 polymorphism and glioma risk in all four genetic models (GG vs. AA: OR=1.87, 95 % CI=1.60-2.18, P heterogeneity=0.552; GA vs. AA: OR=1.30, 95 % CI=1.16-1.46, P heterogeneity=0.495; dominant model-GG+GA vs. AA: OR=1.46, 95 % CI=1.31-1.63, P heterogeneity=0.528; recessive model-GG vs. GA+AA: OR=1.36, 95 % CI=1.27-1.46, P heterogeneity=0.093). Subgroup analyses by ethnicity showed that RTEL1 rs6010620 polymorphism resulted in a higher risk of glioma among both Asians and Caucasians. In the stratified analysis by ethnicity and source of controls, significantly increased risk was observed for Asians and Europeans in all genetic models, population-based studies in all genetic models, and hospital-based studies in three genetic models (heterozygote comparison, homozygote comparison, and dominant model). Our meta-analysis suggested that RTEL1 rs6010620 polymorphism is likely to be associated with increased glioma risk, which lends further biological plausibility to these findings.


Prognostic Relevance of Objective Response According to EASL Criteria and mRECIST Criteria in Hepatocellular Carcinoma Patients Treated with Loco-Regional Therapies: A Literature-Based Meta-Analysis.
The European Association for the Study of the Liver (EASL) criteria and the modified Response Evaluation Criteria in Solid Tumors (mRECIST) are currently adopted to evaluate radiological response in patients affected by HCC and treated with loco-regional procedures. Several studies explored the validity of these measurements in predicting survival but definitive data are still lacking. To conduct a systematic review of studies exploring mRECIST and EASL criteria usefulness in predictive radiological response in HCC undergoing loco-regional therapies and their validity in predicting survival. A comprehensive search of the literature was performed in electronic databases EMBASE, MEDLINE, COCHRANE LIBRARY, ASCO conferences and EASL conferences up to June 10, 2014. Our overall search strategy included terms for HCC, mRECIST, and EASL. Loco-regional procedures included transarterial embolization (TAE), transarterial chemoembolization (TACE) and cryoablation. Inter-method agreement between EASL and mRECIST was assessed using the k coefficient. For each criteria, overall survival was described in responders vs. non-responders patients, considering all target lesions response. Among 18 initially found publications, 7 reports including 1357 patients were considered eligible. All studies were published as full-text articles. Proportion of responders according to mRECIST and EASL criteria was 62.4% and 61.3%, respectively. In the pooled population, 1286 agreements were observed between the two methods (kappa statistics 0.928, 95% confidence interval 0.912-0.944). HR for overall survival (responders versus non responders) according to mRECIST and EASL was 0.39 (95% confidence interval 0.26-0.61, p<0.0001) and 0.38 (95% confidence interval 0.24-0.61, p<0.0001), respectively. In this literature-based meta-analysis, mRECIST and EASL criteria showed very good concordance in HCC patients undergoing loco-regional treatments. Objective response according to both criteria confirms a strong prognostic value in terms of overall survival. This prognostic value appears to be very similar between the two criteria.


Placental 11-beta hydroxysteroid dehydrogenase methylation is associated with newborn growth and a measure of neurobehavioral outcome.
There is growing evidence that the intrauterine environment can impact the neurodevelopment of the fetus through alterations in the functional epigenome of the placenta. In the placenta, the HSD11B2 gene encoding the 11-beta hydroxysteroid dehydrogenase enzyme, which is responsible for the inactivation of maternal cortisol, is regulated by DNA methylation, and has been shown to be susceptible to stressors from the maternal environment. We examined the association between DNA methylation of the HSD11B2 promoter region in the placenta of 185 healthy newborn infants and infant and maternal characteristics, as well as the association between this epigenetic variability and newborn neurobehavioral outcome assessed with the NICU Network Neurobehavioral Scales. Controlling for confounders, HSD11B2 methylation extent is greatest in infants with the lowest birthweights (P = 0.04), and this increasing methylation was associated with reduced scores of quality of movement (P = 0.04). These results suggest that factors in the intrauterine environment which contribute to birth outcome may be associated with placental methylation of the HSD11B2 gene and that this epigenetic alteration is in turn associated with a prospectively predictive early neurobehavioral outcome, suggesting in some part a mechanism for the developmental origins of infant neurological health.


The use of scaffolds in the management of articular cartilage injury.
Managing articular cartilage injury continues to be a difficult challenge for the clinician. Although the short- and intermediate-term results of autologous chondrocyte implantation appear to be favorable, resources are being directed toward research to improve the technology. One promising area of investigation is the combination of cultured chondrocytes with scaffolds. Clinicians desire techniques that may be implanted easily, reduce surgical morbidity, do not require harvesting of other tissues, exhibit enhanced cell proliferation and maturation, have easier phenotype maintenance, and allow for efficient and complete integration with surrounding articular cartilage. The characteristics that make scaffolds optimal for clinical use are that they be biocompatible, biodegradable, permeable, reproducible, mechanically stable, noncytotoxic, and capable of serving as a temporary support for the cells while allowing for eventual replacement by matrix components synthesized by the implanted cells. Clinical experience is growing with three scaffold-based cartilage repair techniques, each using a different type of scaffold material: matrix-induced autologous chondrocyte implantation, a hyaluronic acid-based scaffold, and a composite polylactic/polyglycolic acid polymer fleece. Clinical results are encouraging. Future directions in scaffold-based cartilage repair include bioactive and spatially oriented scaffolds.


Somatostatin as Inflow Modulator in Liver-transplant Recipients With Severe Portal Hypertension: A Randomized Trial.
To investigate the safety and efficacy of somatostatin as liver inflow modulator in patients with end-stage liver disease (ESLD) and clinically significant portal hypertension (CSPH) undergoing liver transplantation (LT) (ClinicalTrials.gov number,01290172). In LT, portal hyperperfusion can severely impair graft function and survival, mainly in cases of partial LT. Thirty-three patients undergoing LT for ESLD and CSPH were randomized double-blindly to receive somatostatin or placebo (2:1). The study drug was administered intraoperatively as 5-mL bolus (somatostatin: 500 μg), followed by a 2.5 mL/h infusion (somatostatin: 250 μg/h) for 5 days. Hepatic and systemic hemodynamics were measured, along with liver function tests and clinical outcomes. The ischemia-reperfusion injury (IRI) was analyzed through histological and protein expression analysis. Twenty-nine patients (18 receiving somatostatin, 11 placebo) were included in the final analysis. Ten patients responded to somatostatin bolus, with a significant decrease in hepatic venous portal gradient (HVPG) and portal flow of -28.3% and -29.1%, respectively. At graft reperfusion, HVPG was lower in patients receiving somatostatin (-81.7% vs -58.8%; P = 0.0084), whereas no difference was observed in the portal flow (P = 0.4185). Somatostatin infusion counteracted the decrease in arterial flow (-10% vs -45%; P = 0.0431). There was no difference between the groups in the severity of IRI, incidence of adverse events, long-term complications, graft, and patient survival. Somatostatin infusion during LT in patients with CSPH is safe, reduces the HVPG, and preserves the arterial inflow to the graft. This study establishes the efficacy of somatostatin as a liver inflow modulator.


Gefitinib-induced interstitial pneumonia: A case report and review of the literature.
The aim of this study was to explore the clinical characteristics of and treatment strategies for interstitial pneumonia induced by gefitinib in patients with advanced non-small cell lung cancer (NSCLC). The detailed clinical data of one patient with NSCLC and gefitinib-induced interstitial pneumonia were compiled and a review of relevant previous studies was performed. Based on this case report and the review, the clinical characteristics, mechanisms and treatment strategies of this rare disease were analyzed. The analyses showed that older, male patients with a long smoking history, high smoking index and adenocarcinoma (particularly bronchoalveolar carcinoma) were more likely to suffer from interstitial pneumonia while taking gefitinib. The onset time of interstitial pneumonia was 1-2 months subsequent to gefitinib administration. The clinical manifestations included chest tightness, shortness of breath, progressive dyspnea, severe hypoxemia and respiratory failure. Diffuse infiltration and alveolar interstitial shadows were observed on the chest tomography scan. In such circumstances, a timely judgment is required, in addition to the withdrawal of gefitinib treatment and the administration of high-dose glucocorticoids, as well as oxygen inhalation and anti-infective therapies, in order to relieve the symptoms. In conclusion, following the onset of gefitinib-induced interstitial pneumonia, the discontinuation of gefitinib is likely to alleviate the suffering of the majority of patients. Early interstitial pneumonia is not an absolute index for the permanent discontinuation of gefitinib treatment. It is necessary to comprehensively consider the benefits and hazards of gefitinib for the patients.


Response of newly established mouse myeloid leukemic cell lines to MC3T3-G2/PA6 preadipocytes and hematopoietic factors.
Some mouse myeloid leukemias induced by X-irradiation and serially transplanted into syngenic mice do not proliferate in vitro even in the presence of hematopoietic factors. To examine whether such leukemic cells can proliferate in response to stromal cells, we cocultured them with MC3T3-G2/PA6 (PA6) preadipocytes, cells that can support the growth of hematopoietic stem cells. All leukemias developed into in vitro cell lines, showing a dependence on contact with the PA6 cells. Two cell lines responded to none of the known hematopoietic factors including interleukin-3 (IL-3), IL-4, IL-5, IL-6, GM-CSF, G-CSF, M-CSF, and Epo. These results demonstrate that the mechanism of the action of PA6 cells is different from that of any of the known hematopoietic factors, and that, because these two leukemic cell lines retained the ability to grow in vivo, responsiveness to the known hematopoietic factors is not essential for the leukemic cell growth in vivo. Furthermore, all leukemic cell lines could respond also to the preadipocytes fixed with formalin, paraformaldehyde, or glutaraldehyde, suggesting that some molecule(s) associated with the surface of PA6 cells or with extracellular matrix secreted by the preadipocytes is responsible for the leukemic cell growth.


Food selection by the South Indian leaf-monkey, Presbytis johnii, in relation to leaf chemistry.
The leaf-monkey Presbytis johnii has been found to exhibit considerable selectivity in its dietary utilization of mature foliage in a rain-forest habitat. To investigate the basis of this selectivity and to examine the hypothesis that the observed selection is related to the digestibility and toxicity of the available foliage, chemical analyses have been made on 16 of the most important tree species in the monkey's habitat. It has been found that the most heavily used items, which form a staple part of the diet of P. johnii, are characterized by a low fibre content and a very low condensed tannin content. However, neither class of compound is an absolute feeding deterrent as minor, but still significant, mature leaf food items contain considerable amounts of both. It is suggested that the feeding deterrents in these minor items, which may be ingested to supply specific dietary requirements, can perhaps be tolerated because of their dilution in the gut by the dietary staples. An examination of the pepsin/cellulase digestibility of available mature foliage showed that the staple food items tended to be highly digestible. Little correlation has been found between alkaloid content and food selection and it is suggested that the colobine forestomach microflora has the ability to detoxify at least some alkaloids. Additional data on mature leaf petioles and young leaves suggest that the high ratio of cell-sap to cell-wall in these items, and their related high digestibility, explain the high relative abundance of these items in the P. johnii diet.


Temporal changes in the expression of the insulin-like growth factor II gene associated with tissue maturation in the human fetus.
The insulin-like growth factors are broadly distributed in the human conceptus and are thought to play a role in the growth and differentiation of tissues during development. Using in situ hybridization we have shown that a wide variety of specific cell types within tissues express the gene for insulin-like growth factor II at times of development from 18 days to 14 weeks of gestation. Examination of blastocysts produced by in vitro fertilization showed no expression, thus bracketing the time of first accumulation of IGF-II mRNA to between 5 and 18 days postfertilization. The pattern of IGF-II expression shows specific age-related differences in different tissues. In the kidney, for example, expression is found in the cells of the metanephric blastema which is dramatically reduced as the blastema differentiates. The reverse is also seen, and we have noted an increase in expression of IGF-II in the cytotrophoblast layer of the placenta with gestational age. The sites of expression do not correlate with areas of either high mitotic activity or specific types of differentiation, but the observed pattern of expression in the kidney, adrenal glands and liver suggests an explanation for the abnormally high IGF-II mRNA expression in developmental tumours such as Wilms' tumour.


Silver nanoparticles inhibit the function of hypoxia-inducible factor-1 and target genes: insight into the cytotoxicity and antiangiogenesis.
Hypoxia-inducible factor-1 (HIF-1) is a transcription factor that is activated upon exposure to hypoxic stress. It modulates a number of cellular responses including proliferation, apoptosis, angiogenesis, and metabolism by activating a panel of target genes in response to hypoxia. The HIF-1 level is often upregulated in the hypoxic microenvironment of solid tumors, which contributes to cancer treatment failure. Here we report that silver nanoparticles (AgNPs), which are widely used as an antimicrobial agent, are an effective inhibitor of HIF-1. AgNPs inhibited the activation of a HIF-dependent reporter construct after the cells were exposed to hypoxic conditions or treated with cobalt chloride, a hypoxia mimetic agent. The AgNPs also interfered with the accumulation of HIF-1α protein and the induction of the endogenous HIF target genes, VEGF-A and GLUT1. Since both HIF-1 and vascular endothelial growth factor-A play an important role in angiogenesis, AgNPs also inhibited angiogenesis in vitro. Our data reveal a new mechanism of how AgNPs act on cellular function, that is, they disrupt HIF signaling pathway. This finding provides a novel insight into how AgNPs can inhibit cancer cell growth and angiogenesis.


L1TD1 - a prognostic marker for colon cancer.
Prognostic markers specific to a particular cancer type can assist in the evaluation of survival probability of patients and help clinicians to assess the available treatment modalities. Gene expression data was analyzed from three independent colon cancer microarray gene expression data sets (N = 1052). Survival analysis was performed for the three data sets, stratified by the expression level of the LINE-1 type transposase domain containing 1 (L1TD1). Correlation analysis was performed to investigate the role of the interactome of L1TD1 in colon cancer patients. We found L1TD1 as a novel positive prognostic marker for colon cancer. Increased expression of L1TD1 associated with longer disease-free survival in all the three data sets. Our results were in contrast to a previous study on medulloblastoma, where high expression of L1TD1 was linked with poor prognosis. Notably, in medulloblastoma L1TD1 was co-expressed with its interaction partners, whereas our analysis revealed lack of co-expression of L1TD1 with its interaction partners in colon cancer. Our results identify increased expression of L1TD1 as a prognostic marker predicting longer disease-free survival in colon cancer patients.


Detection of Mycobacterium tuberculosis in sputum by gas chromatography-mass spectrometry of methyl mycocerosates released by thermochemolysis.
Tuberculosis requires rapid diagnosis to prevent further transmission and allow prompt administration of treatment. Current methods for diagnosing pulmonary tuberculosis lack sensitivity are expensive or are extremely slow. The identification of lipids using gas chromatography- electron impact mass spectrometry (GC-EI/MS) could provide an alternative solution. We have studied mycocerosic acid components of the phthiocerol dimycocerosate (PDIM) family of lipids using thermochemolysis GC-EI/MS. To facilitate use of the technology in a routine diagnostic laboratory a simple extraction procedure was employed where PDIMs were extracted from sputum using petroleum ether, a solvent of low polarity. We also investigated a method using methanolic tetramethylammonium hydroxide, which facilitates direct transesterification of acidic components to methyl esters in the inlet of the GC-MS system. This eliminates conventional chemical manipulations allowing rapid and convenient analysis of samples. When applied to an initial set of 40 sputum samples, interpretable results were obtained for 35 samples with a sensitivity relative to culture of 94% (95%CI: 69.2,100) and a specificity of 100% (95%CI: 78.1,100). However, blinded testing of a larger set of 395 sputum samples found the assay to have a sensitivity of 61.3% (95%CI: 54.9,67.3) and a specificity of 70.6% (95%CI: 62.3,77.8) when compared to culture. Using the results obtained we developed an improved set of classification criteria, which when applied in a blinded re-analysis increased the sensitivity and specificity of the assay to 64.9% (95%CI: 58.6,70.8) and 76.2% (95%CI: 68.2,82.8) respectively. Highly variable levels of background signal were observed from individual sputum samples that inhibited interpretation of the data. The diagnostic potential of using thermochemolytic GC-EI/MS of PDIM biomarkers for diagnosis of tuberculosis in sputum has been established; however, further refinements in sample processing are required to enhance the sensitivity and robustness of the test.


A functional role for the motor system in language understanding: evidence from theta-burst transcranial magnetic stimulation.
Does language comprehension depend, in part, on neural systems for action? In previous studies, motor areas of the brain were activated when people read or listened to action verbs, but it remains unclear whether such activation is functionally relevant for comprehension. In the experiments reported here, we used off-line theta-burst transcranial magnetic stimulation to investigate whether a causal relationship exists between activity in premotor cortex and action-language understanding. Right-handed participants completed a lexical decision task, in which they read verbs describing manual actions typically performed with the dominant hand (e.g., "to throw," "to write") and verbs describing nonmanual actions (e.g., "to earn," "to wander"). Responses to manual-action verbs (but not to nonmanual-action verbs) were faster after stimulation of the hand area in left premotor cortex than after stimulation of the hand area in right premotor cortex. These results suggest that premotor cortex has a functional role in action-language understanding.


Quantifying Guest-Host Dynamics in Supramolecular Assemblies to Analyze Their Robustness.
The most basic function of synthetic microenvironments for tissue engineering is to act as a physical substrate for cell attachment, migration, and proliferation, similar to the natural cell environment. Functionalization of supramolecular materials with guest compounds that display the same recognition moieties is a common strategy to introduce biofunctionality. However, besides a robust interaction with the material, a certain level of dynamics needs to be conserved for an adaptive interface toward the living environment. A balance between robust material functionalization and dynamic cell interaction needs to be met. The detailed analysis hereof using a ureido-pyrimidinone (UPy) poly(ethylene glycol) system in dilute and transient network regime is demonstrated. Monovalent and bivalent UPy-functionalized guest molecules are designed and their interaction with UPy-host fibers is evaluated. Analysis of guest interaction in the dilute state by microfluidics, and in the gel state, by fluorescence recovery after photobleaching and fluorescence resonance energy transfer is proven to be suitable to quantify the local and ensemble guest mobility. The results demonstrate that the interaction of bioactive moieties through supramolecular host-guest chemistry yields a dynamic system, which is stronger for divalent guests but risks unintended leakage in the case of functional monomeric units.


Different subcellular distributions of the vesicular monoamine transporter, VMAT2, in subclasses of sympathetic neurons.
A subpopulation of neurons in the rat superior cervical ganglion (SCG) was found to lack immunostaining for VMAT2, an isoform of the vesicular monoamine transporter that loads catecholamines into vesicles for release at the synapse. Double labeling with neuropeptide Y (NPY), a marker for vasomotor neurons, revealed selective cellular colocalization of NPY together with intense perinuclear staining for VMAT2. This implied that VMAT2-negative neurons were likely to have secretomotor and pilomotor phenotypes. We tested this by identifying peripheral noradrenergic axons by their expression of immunoreactivity for tyrosine hydroxylase (TH) and determining whether they also expressed NPY and VMAT2. This analysis revealed the presence of VMAT2-positive, non-vasomotor sympathetic axons in the submandibular gland and at the base of piloerector hairs. Together the results confirm earlier indications that virtually all sympathetic neurons in the rat SCG express VMAT2 and they show for the first time that functional subclasses of cells can be distinguished by different somatic levels of immunoreactivity for VMAT2.


Nutrient signalling in the regulation of human muscle protein synthesis.
The mammalian target of rapamycin (mTOR) and AMP-activated protein kinase (AMPK) are important nutrient- and energy-sensing and signalling proteins in skeletal muscle. AMPK activation decreases muscle protein synthesis by inhibiting mTOR signalling to regulatory proteins associated with translation initiation and elongation. On the other hand, essential amino acids (leucine in particular) and insulin stimulate mTOR signalling and protein synthesis. We hypothesized that anabolic nutrients would be sensed by both AMPK and mTOR, resulting in an acute and potent stimulation of human skeletal muscle protein synthesis via enhanced translation initiation and elongation. We measured muscle protein synthesis and mTOR-associated upstream and downstream signalling proteins in young male subjects (n=14) using stable isotopic and immunoblotting techniques. Following a first muscle biopsy, subjects in the 'Nutrition' group ingested a leucine-enriched essential amino acid-carbohydrate mixture (EAC). Subjects in the Control group did not consume nutrients. A second biopsy was obtained 1 h later. Ingestion of EAC significantly increased muscle protein synthesis, modestly reduced AMPK phosphorylation, and increased Akt/PKB (protein kinase B) and mTOR phosphorylation (P<0.05). mTOR signalling to its downstream effectors (S6 kinase 1 (S6K1) and 4E-binding protein 1 (4E-BP1) phosphorylation status) was also increased (P<0.05). In addition, eukaryotic elongation factor 2 (eEF2) phosphorylation was significantly reduced (P<0.05). Protein synthesis and cell signalling (phosphorylation status) was unchanged in the control group (P>0.05). We conclude that anabolic nutrients alter the phosphorylation status of both AMPK- and mTOR-associated signalling proteins in human muscle, in association with an increase in protein synthesis not only via enhanced translation initiation but also through signalling promoting translation elongation.


Reproductive aging-associated common genetic variants and the risk of breast cancer.
A younger age at menarche and an older age at menopause are well established risk factors for breast cancer. Recent genome-wide association studies have identified several novel genetic loci associated with these two traits. However, the association between these loci and breast cancer risk is unknown. In this study, we investigated 19 and 17 newly identified single nucleotide polymorphisms (SNPs) from the ReproGen Consortium that have been associated with age at menarche and age at natural menopause, respectively, and assessed their associations with breast cancer risk in 6 population-based studies among up to 3,683 breast cancer cases and 34,174 controls in white women of European ancestry. In addition, we used these SNPs to calculate genetic risk scores (GRSs) based on their associations with each trait. After adjusting for age and potential population stratification, two age at menarche associated SNPs (rs1079866 and rs7821178) and one age at natural menopause associated SNP (rs2517388) were associated with breast cancer risk (p values, 0.003, 0.009 and 0.023, respectively). The odds ratios for breast cancer corresponding to per-risk-allele were 1.14 (95% CI, 1.05 to 1.24), 1.08 (95% CI, 1.02 to 1.15) and 1.10 (95% CI, 1.01 to 1.20), respectively, and were in the direction predicted by their associations with age at menarche or age at natural menopause. These associations did not appear to be attenuated by further controlling for self-reported age at menarche, age at natural menopause, or known breast cancer susceptibility loci. Although we did not observe a statistically significant association between any GRS for reproductive aging and breast cancer risk, the 4th and 5th highest quintiles of the younger age at menarche GRS had odds ratios of 1.14 (95% CI, 1.01 to 1.28) and 1.13 (95% CI, 1.00 to 1.27), respectively, compared to the lowest quintile. Our study suggests that three genetic variants, independent of their associations with age at menarche or age at natural menopause, were associated with breast cancer risk and may contribute modestly to breast cancer risk prediction; however, the combination of the 19 age at menarche or the 17 age at natural menopause associated SNPs did not appear to be useful for identifying a high risk subgroup for breast cancer.


Biodiversity within the city: Effects of land sharing and land sparing urban development on avian diversity.
Urbanization, one of the most extreme human-induced environmental changes, is negatively affecting biodiversity worldwide, strongly suggesting that we should reconcile urban development with conservation. Urbanization can follow two extreme types of development within a continuum: land sharing (buildings mixed with dispersed green space) or land sparing (buildings interspersed with green patches that concentrate biodiversity-supporting vegetation). Recent local-scale studies indicate that biodiversity is typically favored by land sparing. We investigated which of these two types of urbanization is associated with a higher taxonomic (i.e. species richness), functional, and phylogenetic diversity of birds. To do so, we collected information on breeding and wintering bird assemblages in 45 land-sharing and 45 land-sparing areas in nine European cities, which provide the first attempt to explore this question using a large geographical scale and temporal replication. We found that land-sharing urban areas were significantly associated with a higher taxonomic and functional diversity of birds during winter, but not during the breeding season (with only a marginally significant effect for functional diversity). We found no association between the type of urban development and phylogenetic diversity. Our findings indicate that not all components of avian diversity are similarly affected by these two means of urban planning and highlight the importance of integrating the temporal perspective into this kind of studies. Our results also offer useful information to the current debate about the trade-off between biodiversity conservation and human well-being in the context of land sharing and sparing urban practices. In addition, we found that certain small-scale urban landscape characteristics (i.e. few impervious surfaces, high water or tree cover) and human practices (i.e. bird feeders or plants with berries) can help maintaining more diverse urban bird assemblages. We provide specific suggestions for both policymakers and citizens that hopefully will help to create more biodiversity-friendly cities in the future.


Synthesis and reactivity of a ruthenocene-type complex bearing an aromatic π-ligand with the heaviest group 14 element.
An anionic ruthenocene derived from a dilithioplumbole complex was prepared. In the complex, the plumbole ligand coordinates a ruthenium atom in an η5-fashion, similar to the cyclopentadienyl ligand in ferrocene. The ruthenocene that has the aromatic π-ligand with the heaviest group 14 element reacted with electrophiles to afford the plumbole complexes wherein the plumbole ligands show deviation from planarity, in contrast to the planar plumbole ring in the anionic ruthenocene. The bent angles of the plumbole ligands are dependent on the substituents on the lead atoms. Cyclic voltammetry measurements revealed that the plumbole complexes are oxidized more easily than the corresponding stannole complexes.


Modulation of hexokinase association with mitochondria analyzed with quantitative three-dimensional confocal microscopy.
Hexokinase isozyme I is proposed to be associated with mitochondria in vivo. Moreover, it has been suggested that this association is modulated in coordination with changes in cell metabolic state. To test these hypotheses, we analyzed the subcellular distribution of hexokinase relative to mitochondria in paraformaldehyde-fixed astrocytes using immunocytochemistry and quantitative three-dimensional confocal microscopy. Analysis of the extent of colocalization between hexokinase and mitochondria revealed that approximately 70% of cellular hexokinase is associated with mitochondria under basal metabolic conditions. In contrast to the immunocytochemical studies, between 15 to 40% of cellular hexokinase was found to be associated with mitochondria after fractionation of astrocyte cultures depending on the exact fractionation conditions. The discrepancy between fractionation studies and those based on imaging of distributions in fixed cells indicates the usefulness of using techniques that can evaluate the distributions of "cytosolic" enzymes in cells whose subcellular ultrastructure is not severely disrupted. To determine if hexokinase distribution is modulated in concert with changes in cell metabolism, the localization of hexokinase with mitochondria was evaluated after inhibition of glucose metabolism with 2-deoxyglucose. After incubation with 2-deoxyglucose there was an approximate 35% decrease in the amount of hexokinase associated with mitochondria. These findings support the hypothesis that hexokinase is bound to mitochondria in rat brain astrocytes in vivo, and that this association is sensitive to cell metabolic state.


Relative impact of amyloid-β, lacunes, and downstream imaging markers on cognitive trajectories.
SEE COHEN DOI101093/AWW183 FOR A SCIENTIFIC COMMENTARY ON THIS ARTICLE: Amyloid-β and cerebral small vessel disease are the two major causes of cognitive impairment in the elderly. However, the underlying mechanisms responsible for precisely how amyloid-β and cerebral small vessel disease affect cognitive impairment remain unclear. We investigated the effects of amyloid-β and lacunes on downstream imaging markers including structural network and cortical thickness, further analysing their relative impact on cognitive trajectories. We prospectively recruited a pool of 117 mild cognitive impairment patients (45 amnestic type and 72 subcortical vascular type), from which 83 patients received annual follow-up with neuropsychological tests and brain magnetic resonance imaging for 3 years, and 87 patients received a second Pittsburgh compound B positron emission tomography analysis. Structural networks based on diffusion tensor imaging and cortical thickness were analysed. We used linear mixed effect regression models to evaluate the effects of imaging markers on cognitive decline. Time-varying Pittsburgh compound B uptake was associated with temporoparietal thinning, which correlated with memory decline (verbal memory test, unstandardized β = -0.79, P < 0.001; visual memory test, unstandardized β = -2.84, P = 0.009). Time-varying lacune number was associated with the degree of frontoparietal network disruption or thinning, which further affected frontal-executive function decline (Digit span backward test, unstandardized β = -0.05, P = 0.002; Stroop colour test, unstandardized β = -0.94, P = 0.008). Of the multiple imaging markers analysed, Pittsburgh compound B uptake and the number of lacunes had the greatest association with memory decline and frontal-executive function decline, respectively: Time-varying Pittsburgh compound B uptake (standardized β = -0.25, P = 0.010) showed the strongest effect on visual memory test, followed by time-varying temporoparietal thickness (standardized β = 0.21, P = 0.010) and time-varying nodal efficiency (standardized β = 0.17, P = 0.024). Time-varying lacune number (standardized β = -0.25, P = 0.014) showed the strongest effect on time-varying digit span backward test followed by time-varying nodal efficiency (standardized β = 0.17, P = 0.021). Finally, time-varying lacune number (β = -0.22, P = 0.034) showed the strongest effect on time-varying Stroop colour test followed by time-varying frontal thickness (standardized β = 0.19, P = 0.026). Our multimodal imaging analyses suggest that cognitive trajectories related to amyloid-β and lacunes have distinct paths, and that amyloid-β or lacunes have greatest impact on cognitive decline. Our results provide rationale for the targeting of amyloid-β and lacunes in therapeutic strategies aimed at ameliorating cognitive decline.


iEEGview: an open-source multifunction GUI-based Matlab toolbox for localization and visualization of human intracranial electrodes.
The precise localization of intracranial electrodes is a fundamental step relevant to the analysis of intracranial electroencephalography (iEEG) recordings in various fields. With the increasing development of iEEG studies in human neuroscience, higher requirements have been posed on the localization process, resulting in urgent demand for more integrated, easy-operation and versatile tools for electrode localization and visualization. With the aim of addressing this need, we develop an easy-to-use and multifunction toolbox called iEEGview, which can be used for the localization and visualization of human intracranial electrodes. iEEGview is written in Matlab scripts and implemented with a GUI. From the GUI, by taking only pre-implant MRI and post-implant CT images as input, users can directly run the full localization pipeline including brain segmentation, image co-registration, electrode reconstruction, anatomical information identification, activation map generation and electrode projection from native brain space into common brain space for group analysis. Additionally, iEEGview implements methods for brain shift correction, visual location inspection on MRI slices and computation of certainty index in anatomical label assignment. All the introduced functions of iEEGview work reliably and successfully, and are tested by images from 28 human subjects implanted with depth and/or subdural electrodes. iEEGview is the first public Matlab GUI-based software for intracranial electrode localization and visualization that holds integrated capabilities together within one pipeline. iEEGview promotes convenience and efficiency for the localization process, provides rich localization information for further analysis and offers solutions for addressing raised technical challenges. Therefore, it can serve as a useful tool in facilitating iEEG studies.


Evaluation of waiting times and sonographic findings in patients with first trimester vaginal bleeding at the university hospital of the west indies. Can emergency department ultrasound make a difference?
Pregnant female patients with vaginal bleeding in the first trimester are seen commonly in the Emergency Department (ED) at the University Hospital of the West Indies (UHWI), Kingston, Jamaica. The protocol for the management of these patients requires that they have a sonographic evaluation performed for the purpose of localizing the pregnancy where possible, to assist with determining the risk for an ectopic pregnancy. The ultrasound examinations are performed in the radiology department. This retrospective study was conducted to evaluate how long patients wait for a pelvic ultrasound. We also sought to establish how many patients had ultrasound findings that would have allowed safe discharge home. The records of 150 patients seen in the six-month period from January 1 to July 30, 2008 were examined. Data were extracted pertaining to age, time to see an emergency room doctor, time taken for ultrasound examination to be obtained from the radiology department and the ultrasound findings. Fifty-four per cent presented to the Emergency Department with a complaint of vaginal bleeding and abdominal pain, 29% with bleeding only, 16% with abdominal pain only and one with syncope. One hundred and sixteen of the patients enrolled had an ultrasound performed at UHWI. The average waiting time for an ultrasound was 3.8 ± 2.5 hours. The majority (66/116) of the patients had an intrauterine pregnancy (IUP) demonstrated on ultrasound. Twenty-nine had no IUP, free fluid or adnexal mass. These 95 patients would likely have been discharged home. Ten patients had an adnexal mass with or without free fluid, and ten had free fluid only on ultrasound. One patient was found to have a definite ectopic pregnancy. These 21 patients would have been referred for evaluation by the obstetrician on call for further management. The majority of patients had sonographic findings that would have allowed safe and timely discharge from the Emergency Department had ultrasound been available at the point of care.


The association between three attitude-related indexes of oral hygiene and secondary implant failures: A retrospective longitudinal study.
This study evaluated the strength of the association between three widely used clinical indexes considered as distal behavioural indicators of attitude-related oral status (an index of oral hygiene, the plaque index [PI] and two periodontal indexes, that is the presence of bleeding on probing [BOP] and of pockets probing depth [PPD]) and secondary implant failure due to peri-implantitis in patients rehabilitated with cemented prosthesis. The study included patients who underwent implant-prosthetic rehabilitation and had joined the programme of maintenance of the same hospital. Implant failures, number of months between implant insertion and implant loading, and patients' surgical protocol were monitored and recorded. Further, PI, BOP and PPD-all attitude-related indicators of oral hygiene and periodontal inflammation-were recorded and related, in terms of odds ratios (ORs ) and corresponding risk factors, to secondary implant failures. A total of 1427 patients (2673 implants) were enrolled. The follow-up ranged from 1.5 to 9 years (mean 5.3 years±1.3). The cumulative survival rate was 98.01%. Thirty-two patients (36 implants, 1.36% of all implants) had implant failure. A statistically significant association between PI, BOP, PPD and secondary failures due to peri-implantitis was observed. Within the limitations of this study, all three attitude-related behavioural indicators-the plaque index (PI), bleeding on probing (BOP) and abnormal probing pocket depth (PPD)-proved to be significant risk indicators for secondary implant failure due to peri-implantitis, both from a clinical and from a socio-psychological attitude-related perspective.


Zooming in on cerebral small vessel function in small vessel diseases with 7T MRI: Rationale and design of the "ZOOM@SVDs" study.
Cerebral small vessel diseases (SVDs) are a major cause of stroke and dementia. Yet, specific treatment strategies are lacking in part because of a limited understanding of the underlying disease processes. There is therefore an urgent need to study SVDs at their core, the small vessels themselves. This paper presents the rationale and design of the ZOOM@SVDs study, which aims to establish measures of cerebral small vessel dysfunction on 7T MRI as novel disease markers of SVDs. ZOOM@SVDs is a prospective observational cohort study with two years follow-up. ZOOM@SVDs recruits participants with Cerebral Autosomal Dominant Arteriopathy with Subcortical Infarcts and Leukoencephalopathy (CADASIL, N = 20), sporadic SVDs (N = 60), and healthy controls (N = 40). Participants undergo 7T brain MRI to assess different aspects of small vessel function including small vessel reactivity, cerebral perforating artery flow, and pulsatility. Extensive work-up at baseline and follow-up further includes clinical and neuropsychological assessment as well as 3T brain MRI to assess conventional SVD imaging markers. Measures of small vessel dysfunction are compared between patients and controls, and related to the severity of clinical and conventional MRI manifestations of SVDs. ZOOM@SVDs will deliver novel markers of cerebral small vessel function in patients with monogenic and sporadic forms of SVDs, and establish their relation with disease burden and progression. These small vessel markers can support etiological studies in SVDs and may serve as surrogate outcome measures in future clinical trials to show target engagement of drugs directed at the small vessels.


Insecticidal Effects of Organotin(IV) Compounds on Plutella Xylostella (L.) Larvae. II. Inhibitory Potencies Against Acetylcholinesterase and Evidence for Synergism in Tests With Bacillus Thuringiensis(BER.) and Malathion.
Features of pesticide synergism and acetylcholinesterase (AChE) inhibition (in vitro) were studied using a selected range of organotin compounds against the early 4th instar larvae of a highly resistant strain of the diamondback moth (DBM), Plutella xylostella, a major universal pest of cruciferous vegetables.Fourteen triorganotin compounds were evaluated for their ability to enhance the toxicity of the microbial insecticide, Bacillus thuringiensis (BT) and of the commercial insecticide, Malathion to Plutella xylostella larvae. Supplemental synergism was observed with triphenyl- and tricyclopentyltin hydroxides in combinations with Bacillus thuringiensis. Increased synergism was observed with an increase in the number of cyclopentyl groups on tin in the mixed series, Cyp(n) Ph(3-n) SnX, where X = OH, and 1-(1,2,4-triazolyl). The combination of (p-chlorophenyl)diphenyltin N,N-dimethyldithiocarbamate at LD(10) and LD(25) concentrations with sublethal concentrations of Malathion as well as of tricyclohexyltin methanesulphonate at the 0.01% (w/v) concentration with Malathion exerted strong synergistic effects (supplemental synergism) with toxicity index (T.I) values of 7.2, 19.8 and 10.1, respectively.Studies on the in vitro inhibition of acetylcholinesterase prepared from the DBM larvae showed that while most of the triorganotin Compounds tested were without effect on the enzyme, compounds containing the thiocarbamylacetate or the dithiocarbamylacetate moieties demonstrated appreciable levels of inhibition, being comparable in efficacy to commercial grades of Malathion and Methomyl.


Short-Term Efficacy and Safety of Tolvaptan in Patients with Left Ventricular Assist Devices.
Tolvaptan is an effective therapy for heart failure patients with symptomatic congestion and hyponatremia. The efficacy of its use in patients with continuous-flow left ventricular assist devices (LVADs) is unknown. The aim of this study was to assess the clinical efficacy and safety of tolvaptan in LVAD patients. We retrospectively reviewed medical records of patients who underwent LVAD implantation between January 2014 and August 2018. Among 217 consecutive LVAD patients, tolvaptan was used in 20 patients. Mean age was 46 ± 14 years old and 14 patients were males. The duration of tolvaptan therapy was 4 (interquartile range 1-8) days. Urine volume significantly increased from 2,623 ± 1,109 ml/day before tolvaptan to 4,308 ± 1,432 ml/day during tolvaptan therapy (p < 0.001). Serum sodium increased from 127 ± 3 to 133 ± 3 mEq/L at the end of tolvaptan therapy (p < 0.001). No patients developed hypernatremia (serum sodium >150 mEq/L). The 90-day overall survival following tolvaptan therapy was 89% in both the tolvaptan group and a propensity score-matched non-tolvaptan group (p = 0.918). Survival free of heart failure readmissions was also comparable between the groups (p = 0.751). In conclusion, short-term use of tolvaptan following LVAD implantation is a safe and effective therapy to augment diuresis and improve hyponatremia.


FOXM1 contributes to treatment failure in acute myeloid leukemia.
Acute myeloid leukemia (AML) patients with NPM1 mutations demonstrate a superior response to standard chemotherapy treatment. Our previous work has shown that these favorable outcomes are linked to the cytoplasmic relocalization and inactivation of FOXM1 driven by mutated NPM1. Here, we went on to confirm the important role of FOXM1 in increased chemoresistance in AML. A multiinstitution retrospective study was conducted to link FOXM1 expression to clinical outcomes in AML. We establish nuclear FOXM1 as an independent clinical predictor of chemotherapeutic resistance in intermediate-risk AML in a multivariate analysis incorporating standard clinicopathologic risk factors. Using colony assays, we show a dramatic decrease in colony size and numbers in AML cell lines with knockdown of FOXM1, suggesting an important role for FOXM1 in the clonogenic activity of AML cells. In order to further prove a potential role for FOXM1 in AML chemoresistance, we induced an FLT3-ITD-driven myeloid neoplasm in a FOXM1-overexpressing transgenic mouse model and demonstrated significantly higher residual disease after standard chemotherapy. This suggests that constitutive overexpression of FOXM1 in this model induces chemoresistance. Finally, we performed proof-of-principle experiments using a currently approved proteasome inhibitor, ixazomib, to target FOXM1 and demonstrated a therapeutic response in AML patient samples and animal models of AML that correlates with the suppression of FOXM1 and its transcriptional targets. Addition of low doses of ixazomib increases sensitization of AML cells to chemotherapy backbone drugs cytarabine and the hypomethylator 5-azacitidine. Our results underscore the importance of FOXM1 in AML progression and treatment, and they suggest that targeting it may have therapeutic benefit in combination with standard AML therapies.


Ecosystem-based coastal defence in the face of global change.
The risk of flood disasters is increasing for many coastal societies owing to global and regional changes in climate conditions, sea-level rise, land subsidence and sediment supply. At the same time, in many locations, conventional coastal engineering solutions such as sea walls are increasingly challenged by these changes and their maintenance may become unsustainable. We argue that flood protection by ecosystem creation and restoration can provide a more sustainable, cost-effective and ecologically sound alternative to conventional coastal engineering and that, in suitable locations, it should be implemented globally and on a large scale.


Prevalence of microalbuminuria and associated electrocardiographic abnormalities in an Indo-Asian population.
Microalbuminuria (MA) is a known predictor of cardiovascular disease (CVD) in European origin populations, but such data are lacking in native Indo-Asian populations, where CVD risks are high. Major electrocardiographic (ECG) changes are predictive of cardiovascular mortality. We determined the association of MA with major ECG changes in the general population of Pakistan. A total of 3143 subjects aged >or=40 years from 12 randomly selected communities in Karachi participated. MA was defined as the urine albumin to creatinine (ACR) ratio of < 300 mg/g creatinine and >or=17 mg/g in men and >or=25 mg/g in women from a single-spot morning urine sample. Major changes on ECG were coded in duplicate using Minnesota classification. The mean age of subjects was 51.5 (10.7) years. The median (25-75 percentile) ACR was 4.2 (2.9-7.9) mg/g in men and 6.0 (3.9-10.8) mg/g in women (P < 0.001). The overall prevalence (95% CI) of MA was 12.3% (11.1-13.5%), and 20.3% in those with major ECG changes. In a multivariable model, major ECG changes (OR, 95% CI) (1.50, 1.10-2.00), diabetes (3.57, 2.93-4.35), hypertension (2.30, 1.85-2.86), female sex (0.61, 0.53-0.69), age (1.09, 1.05-1.13, for each 5-year increase) and eGFR (0.80, 0.78-0.81, for each 10 mg/g increase) were independently associated with MA. The presence of MA increased the prevalence of major ECG changes from 21 to 31% in those with hypertension (44.9%), 15 to 28% among those with diabetes (21.4%), 14 to 26% among those with overweight or obesity (68.4%) and 14 to 26% among current users of tobacco (38.7%) (P < 0.001) each. The strong association between MA and major ECG changes underscores the importance of screening Indo-Asian subjects for MA for unmasking underlying CVD, especially those with hypertension, diabetes, obesity, and tobacco users.


Cognitive impairment and intracranial calcification in chronic hypoparathyroidism.
Intracranial calcification is associated with chronic hypoparathyroidism. The relationship between intracranial calcification, neurological abnormalities and cognitive deficits in this disorder is unknown. The purpose of this study was to determine whether chronic hypoparathyroidism is associated with cognitive impairment. We studied 11 hypoparathyroid patients and compared them with a sex-, age-, and education-matched control group. The hypoparathyroidism was postsurgical in nine and idiopathic in two. All patients underwent nonenhanced head computed tomography, detailed neurological examinations, and a battery of cognitive tests. These tests were performed separately and individual examiners were blinded to the results of the other components of the study. The mean age of the patients was 55 years; the duration of hypoparathyroidism was at least 9 years. Neuropsychological testing revealed cognitive impairment in 65% of hypoparathyroid subjects, and the presence of significant differences between the hypoparathyroid and control groups. Computed tomography showed intracranial calcification in 6 of 10 hypoparathyroid subjects tested, and neurological (motor) examination revealed 5 of 11 with abnormal findings. There were positive correlations between the presence of cognitive deficits and cerebral calcification (r = 0.59, P = 0.07), between abnormal motor findings and cerebral calcification (r = 0.77, P < 0.01) and between abnormal motor findings and the degree of cognitive deficit (r = 0.83, P < 0.01). We conclude that cognitive and neurological deficits commonly occur in patients with chronic hypoparathyroidism and may be pathophysiologically related to the presence of intracranial calcification.


Rational design of carboxyl groups perpendicularly attached to a graphene sheet: a platform for enhanced biosensing applications.
Graphene oxide (GO)-based materials offer great potential for biofunctionalization with applications ranging from biosensing to drug delivery. Such biofunctionalization utilizes specific functional groups, typically a carboxyl moiety, as anchoring points for biomolecule. However, due to the fact that the exact chemical structure of GO is still largely unknown and poorly defined (it was postulated to consist of various oxygen-containing groups, such as epoxy, hydroxyl, carboxyl, carbonyl, and peroxy in varying ratios), it is challenging to fabricate highly biofunctionalized GO surfaces. The predominant anchoring sites (i.e., carboxyl groups) are mainly present as terminal groups on the edges of GO sheets and thus account for only a fraction of the oxygen-containing groups on GO. Herein, we suggest a direct solution to the long-standing problem of limited abundance of carboxyl groups on GO; GO was first reduced to graphene and consequently modified with only carboxyl groups grafted perpendicularly to its surface by a rational synthesis using free-radical addition of isobutyronitrile with subsequent hydrolysis. Such grafted graphene oxide can contain a high amount of carboxyl groups for consequent biofunctionalization, at which the extent of grafting is limited only by the number of carbon atoms in the graphene plane; in contrast, the abundance of carboxyl groups on "classical" GO is limited by the amount of terminal carbon atoms. Such a graphene platform embedded with perpendicularly grafted carboxyl groups was characterized in detail by X-ray photoelectron spectroscopy, cyclic voltammetry, and electrochemical impedance spectroscopy, and its application was exemplified with single-nucleotide polymorphism detection. It was found that the removal of oxygen functionalities after the chemical reduction enhanced the electron-transfer rate of the graphene. More importantly, the introduction of carboxyl groups promoted a more efficient immobilization of DNA probes on the electrode surface and improved the performance of graphene as a biosensor in comparison to GO. The proposed material can be used as a universal platform for biomolecule immobilization to facilitate rapid and sensitive detection of DNA or proteins for point-of-care investigations. Such reactive carboxyl groups grafted perpendicularly on GO holds promise for a highly efficient tailored biofunctionalization for applications in biosensing or drug delivery.


Mg2+ and Ca2+ differentially regulate DNA binding and dimerization of DREAM.
DREAM (calsenilin/KChIP3) is an EF-hand calcium-binding protein that represses transcription of prodynorphin and c-fos genes. Here we present structural and binding studies on single-site mutants of DREAM designed to disable Ca(2+) binding to each of the functional EF-hands (EF-2: D150N; EF-3: E186Q; and EF-4: E234Q). Isothermal titration calorimetry (ITC) analysis of Ca(2+) binding to the various mutants revealed that, in the absence of Mg(2+), Ca(2+) binds independently and sequentially to EF-3 (DeltaH = -2.4 kcal/mol), EF-4 (DeltaH = +5.2 kcal/mol), and EF-2 (DeltaH = +1 kcal/mol). By contrast, only two Ca(2+) bind to DREAM in the presence of physiological levels of Mg(2+) for both wild-type and D150N, suggesting that EF-2 binds constitutively to Mg(2+). ITC measurements demonstrate that one Mg(2+) binds enthalpically with high affinity (K(d) = 13 mum and DeltaH = -0.79 kcal/mol) and two or more Mg(2+) bind entropically in the millimolar range. Size-exclusion chromatography studies revealed that Mg(2+) stabilizes DREAM as a monomer, whereas Ca(2+) induces protein dimerization. Electrophoretic mobility shift assays indicated that Mg(2+) is essential for sequence-specific binding of DREAM to DNA response elements (DREs) in prodynorphin and c-fos genes. The EF-hand mutants bind specifically to DRE, suggesting they are functionally intact. None of the EF-hand mutants bind DRE at saturating Ca(2+) levels, suggesting that binding of a single Ca(2+) at either EF-3 or EF-4 is sufficient to drive conformational changes that abolish DNA binding. NMR structural analysis indicates that metal-free DREAM adopts a folded yet flexible molten globule-like structure. Both Ca(2+) and Mg(2+) induce distinct conformational changes, which stabilize tertiary structure of DREAM. We propose that Mg(2+) binding at EF-2 may structurally bridge DREAM to DNA targets and that Ca(2+)-induced protein dimerization disrupts DNA binding.


Mucosal inflammation in severe glucocorticoid-dependent asthma.
To improve our understanding of the inflammatory mechanisms underlying severe disease, a biopsy study was performed comparing 15 clinically unstable glucocorticoid-dependent asthmatics, 10 mild asthmatics, and 10 control subjects. Compared with mild asthma, severe asthma was characterized by reduced mucosal eosinophilia. Whilst no significant differences were found in the numbers of mast cells, neutrophils, CD3+ and CD4+ T-cells between the three groups, up to a 4-fold increase In the numbers of activated T-lymphocytes bearing the interleukin (IL)-2 receptor (IL-2R) was found in the mucosa in severe asthma compared to mild asthma (p = 0.03) and control subjects (p = 0.003). Compared to control subjects, the mucosa of severe asthmatics contained significantly (p = 0.02) higher numbers of IL-5+ cells, with no differences between mild and severe disease. In contrast, staining for the anti-IL-4 monoclonal antibody 3H4 revealed that biopsies from mild asthmatics contained more IL4+ cells than biopsies from severe asthmatics and control subjects (p = 0.0008). In the severe asthmatics, a close correlation (r(s) = 0.76, p = 0.005) was found between the numbers of IL-2R-bearing cells and the variability in peak expiratory flow. In conclusion, persistent T-cell activation is a prominent feature of severe asthma. These results also indicate that interleukin-5, and not interleukin-4, is upregulated in severe disease.


Affinity screening using competitive binding with fluorine-19 hyperpolarized ligands.
Fluorine-19 NMR and hyperpolarization form a powerful combination for drug screening. Under a competitive equilibrium with a selected fluorinated reporter ligand, the dissociation constant (K(D)) of other ligands of interest is measurable using a single-scan Carr-Purcell-Meiboom-Gill (CPMG) experiment, without the need for a titration. This method is demonstrated by characterizing the binding of three ligands with different affinities for the serine protease trypsin. Monte Carlo simulations show that the highest accuracy is obtained when about one-half of the bound reporter ligand is displaced in the binding competition. Such conditions can be achieved over a wide range of affinities, allowing for rapid screening of non-fluorinated compounds when a single fluorinated ligand for the binding pocket of interest is known.


Gene expression profiling revealed novel mechanism of action of Taxotere and Furtulon in prostate cancer cells.
Both Taxotere and Capecitabine have shown anti-cancer activity against various cancers including prostate cancer. In combination, Taxotere plus Capecitabine has demonstrated higher anti-cancer activity in advanced breast cancers. However, the molecular mechanisms of action of Taxotere and Capecitabine have not been fully elucidated in prostate cancer. The total RNA from PC3 and LNCaP prostate cells untreated and treated with 2 nM Taxotere, 110 microM Furtulon (active metabolite of Capecitabine), or 1 nM Taxotere plus 50 microM Furtulon for 6, 36, and 72 hours, was subjected to Affymetrix Human Genome U133A Array analysis. Real-time PCR and Western Blot analysis were conducted to confirm microarray data. Taxotere and Furtulon down-regulated some genes critical for cell proliferation, cell cycle progression, transcription factor, cell signaling, and oncogenesis, and up-regulated some genes related to the induction of apoptosis, cell cycle arrest, and differentiation in both cell lines. Taxotere and Furtulon also up-regulated some genes responsible for chemotherapeutic resistance, suggesting the induction of cancer cell resistance to these agents. Taxotere and Furtulon caused the alternation of a large number of genes, many of which may contribute to the molecular mechanisms by which Taxotere and Furtulon inhibit the growth of prostate cancer cells. This information could be utilized for further mechanistic research and for devising optimized therapeutic strategies against prostate cancer.


Generation of pseudonondiffracting optical beams with superlattice structures.
We demonstrate an approach to generate a class of pseudonondiffracting optical beams with the transverse shapes related to the superlattice structures. For constructing the superlattice waves, we consider a coherent superposition of two identical lattice waves with a specific relative angle in the azimuthal direction. We theoretically derive the general conditions of the relative angles for superlattice waves. In the experiment, a mask with multiple apertures which fulfill the conditions for superlattice structures is utilized to generate the pseudonondiffracting superlattice beams. With the analytical wave functions and experimental patterns, the pseudonondiffracting optical beams with a variety of structures can be generated systematically.