[Behavioral and electroencephalographic effects of lormetazepam].
Behavioral and electroencephalographic effects of lormetazepam were investigated in rats, mice and rabbits, in comparison with those of diazepam, nitrazepam and flurazepam. Locomotor activity of mice in an open-field situation was decreased with large doses of lormetazepam and diazepam, while it was increased with nitrazepam and flurazepam. The anticonflict effect of lormetazepam in rats was much more potent than those of diazepam, nitrazepam and flurazepam. In suppressing the muricide of olfactory bulbectomized and raphe lesioned rats, lormetazepam was more potent than diazepam, nitrazepam and flurazepam. Lormetazepam, diazepam and flurazepam prevented both maximal electroshock and pentetrazol convulsions in mice, the effects on the latter being much more potent than those on the former. Lormetazepam was more potent than diazepam and flurazepam in potentiating thiopental-, ether- and ethanol anesthesia, impairing rotarod performance and in muscle relaxant activity. In conscious rabbits with chronically implanted electrodes, lormetazepam induced a drowsy EEG pattern and suppressed the EEG arousal responses not only to auditory stimulation but also to electrical stimulation of the midbrain reticular formation or hypothalamus. Lormetazepam also inhibited afterdischarges induced by electrical stimulation of the hippocampus and amygdala. These results indicate that lormetazepam has pharmacological properties characteristic of benzodiazepines and that the activity is more potent than those of diazepam, nitrazepam and flurazepam.


The countervailing actions of myeloid and plasmacytoid dendritic cells control autoimmune diabetes in the nonobese diabetic mouse.
Islet Ag-specific CD4(+) T cells receive antigenic stimulation from MHC class II-expressing APCs. Herein, we delineate the direct in vivo necessity for distinct subsets of macrophages and dendritic cells (DC) in type 1 diabetes mellitus of the NOD mouse by using diphtheria toxin-mediated cell ablation. The ablation of macrophages had no impact on islet Ag presentation or on the induction of insulitis or diabetes in either transfer or spontaneous models. However, the ablation of CD11b(+)CD11c(+) DC led to the loss of T cell activation, insulitis, and diabetes mediated by CD4(+) T cells. When the specific myeloid DC subset was "added-back" to mice lacking total DC, insulitis and diabetes were restored. Interestingly, when NOD mice were allowed to progress to the insulitis phase, the ablation of DC led to accelerated insulitis. This accelerated insulitis was mediated by the loss of plasmacytoid DC (pDC). When pDC were returned to depleted mice, the localized regulation of insulitis was restored. The loss of pDC in the pancreas itself was accompanied by the localized loss of IDO and the acceleration of insulitis. Thus, CD11c(+)CD11b(+) DC and pDC have countervailing actions in NOD diabetes, with myeloid DC providing critical antigenic stimulation to naive CD4(+) T cells and pDC providing regulatory control of CD4(+) T cell function in the target tissue.


Phosphorylation of the voltage-gated potassium channel Kv2.1 by AMP-activated protein kinase regulates membrane excitability.
Firing of action potentials in excitable cells accelerates ATP turnover. The voltage-gated potassium channel Kv2.1 regulates action potential frequency in central neurons, whereas the ubiquitous cellular energy sensor AMP-activated protein kinase (AMPK) is activated by ATP depletion and protects cells by switching off energy-consuming processes. We show that treatment of HEK293 cells expressing Kv2.1 with the AMPK activator A-769662 caused hyperpolarizing shifts in the current-voltage relationship for channel activation and inactivation. We identified two sites (S440 and S537) directly phosphorylated on Kv2.1 by AMPK and, using phosphospecific antibodies and quantitative mass spectrometry, show that phosphorylation of both sites increased in A-769662-treated cells. Effects of A-769662 were abolished in cells expressing Kv2.1 with S440A but not with S537A substitutions, suggesting that phosphorylation of S440 was responsible for these effects. Identical shifts in voltage gating were observed after introducing into cells, via the patch pipette, recombinant AMPK rendered active but phosphatase-resistant by thiophosphorylation. Ionomycin caused changes in Kv2.1 gating very similar to those caused by A-769662 but acted via a different mechanism involving Kv2.1 dephosphorylation. In cultured rat hippocampal neurons, A-769662 caused hyperpolarizing shifts in voltage gating similar to those in HEK293 cells, effects that were abolished by intracellular dialysis with Kv2.1 antibodies. When active thiophosphorylated AMPK was introduced into cultured neurons via the patch pipette, a progressive, time-dependent decrease in the frequency of evoked action potentials was observed. Our results suggest that activation of AMPK in neurons during conditions of metabolic stress exerts a protective role by reducing neuronal excitability and thus conserving energy.


Effects of physical exercise on depressive symptoms and biomarkers in depression.
Regular physical exercise/activity has been shown repeatedly to promote positive benefits in cognitive, emotional and motor domains concomitant with reductions in distress and negative affect. It exerts a preventative role in anxiety and depressive states and facilitates psychological well-being in both adolescents and adults. Not least, several meta-analyses attest to improvements brought about by exercise. In the present treatise, the beneficial effects of exercise upon cognitive, executive function and working memory, emotional, self-esteem and depressed mood, motivational, anhedonia and psychomotor retardation, and somatic/physical, sleep disturbances and chronic aches and pains, categories of depression are discussed. Concurrently, the amelioration of several biomarkers associated with depressive states: hypothalamic-pituitary-adrenal (HPA) axis homeostasis, anti-neurodegenerative effects, monoamine metabolism regulation and neuroimmune functioning. The notion that physical exercise may function as "scaffolding" that buttresses available network circuits, anti-inflammatory defences and neuroreparative processes, e.g. brain-derived neurotrophic factor (BDNF), holds a certain appeal.


Signature sequence validation of human papillomavirus type 16 (HPV-16) in clinical specimens.
Persistent infection indicated by detection of human papillomavirus 16 (HPV-16) on repeat testing over a period of time poses the greatest cervical cancer risk. However, variants of HPV-16, HPV-31 and HPV-33 may share several short sequence homologies in the hypervariable L1 gene commonly targeted for HPV genotyping. The purpose of this study was to introduce a robust laboratory procedure to validate HPV-16 detected in clinical specimens, using the GenBank sequence database as the standard reference for genotyping. A nested PCR with two pairs of consensus primers was used to amplify the HPV DNA released in crude proteinase K digest of the cervicovaginal cells in liquid-based Papanicolaou cytology specimens. The positive nested PCR products were used for direct automated DNA sequencing. A 48-base sequence downstream of the GP5+ priming site, or a 34-base sequence upstream thereof, was needed for unequivocal validation of an HPV-16 isolate. Selection of a 45-base, or shorter, sequence immediately downstream of the GP5+ site for Basic Local Alignment Search Tool sequence analysis invariably led to ambiguous genotyping results. DNA sequence analysis may be used for differential genotyping of HPV-16, HPV-31 and HPV-33 in clinical specimens. However, selection of the signature sequence for Basic Local Alignment Search Tool algorithms is crucial to distinguish certain HPV-16 variants from other closely related HPV genotypes.


Regulation of neuronal mRNA translation by CaM-kinase I phosphorylation of eIF4GII.
Ca²⁺/calmodulin-dependent kinases (CaMKs) are essential for neuronal development and plasticity, processes requiring de novo protein synthesis. Roles for CaMKs in modulating gene transcription are well established, but their involvement in mRNA translation is evolving. Here we report that activity-dependent translational initiation in cultured rat hippocampal neurons is enhanced by CaMKI-mediated phosphorylation of Ser1156 in eukaryotic initiation factor eIF4GII (4GII). Treatment with bicuculline or gabazine to enhance neuronal activity promotes recruitment of wild-type 4GII, but not the 4GII S1156A mutant or 4GI, to the heterotrimeric eIF4F (4F) complex that assembles at the 5' cap structure (m⁷GTP) of mRNA to initiate ribosomal scanning. Recruitment of 4GII to 4F is suppressed by pharmacological inhibition (STO-609) of CaM kinase kinase, the upstream activator of CaMKI. Post hoc in vitro CaMKI phosphorylation assays confirm that activity promotes phosphorylation of S1156 in transfected 4GII in neurons. Changes in cap-dependent and cap-independent translation were assessed using a bicistronic luciferase reporter transfected into neurons. Activity upregulates cap-dependent translation, and RNAi knockdown of CaMKIβ and γ isoforms, but not α or δ, led to its attenuation as did blockade of NMDA receptors. Furthermore, RNAi knockdown of 4GII attenuates cap-dependent translation and reduces density of dendritic filopodia and spine formation without effect on dendritic arborization. Together, our results provide a mechanistic link between Ca²⁺ influx due to neuronal activity and regulation of cap-dependent RNA translation via CaMKI activation and selective recruitment of phosphorylated 4GII to the 4F complex, which may function to regulate activity-dependent changes in spine density.


Adenoviral gene delivery elicits distinct pulmonary-associated T helper cell responses to the vector and to its transgene.
Replication-deficient adenovirus (Ad) vectors are effective to specifically target the respiratory epithelium for either corrective gene therapy such as cystic fibrosis or for mucosal immunization. As a consequence of transducing the lower respiratory tract with an E1/E3 deleted Ad5 vector, host responses have been characterized by the duration of transgene expression and by the induction of CTL responses. However, limited emphasis has been devoted to understanding the contribution of CD4+ T cell responses to the Ad vector. Both CD4+ and CD8+ T cells migrate into the lung following sequential intratracheal Ad5 transgene instillations. Isolated CD3+ T lymphocytes from the lungs were predominantly of the Th2 type, and after cell sorting, the IL-4-producing T cells were largely CD4+, while IFN-gamma expression was associated with both CD4+ and CD8+ T cells. Ab responses to the Ad5 vector and to the expressed transgene beta-galactosidase (beta gal) revealed elevated bronchial and serum IgA and IgG Abs with low neutralization titers. Analysis of serum IgG subclass responses showed IgG1 and IgG2b with lower IgG2a Abs to Ad5 and IgG2a and IgG2b Ab responses to beta gal. Ad5-specifc CD4+ T cells produced both Th1 (IFN-gamma and IL-2)- and Th2 (IL-4, IL-5, IL-6)-type cytokines, while beta gal-specific CD4+ T cells secreted IFN-gamma and IL-6. This study provides direct evidence for the concomitant induction of Th2- with Th1-type responses in both the pulmonary systemic and mucosal immune compartments to the Ad5 vector as well as a Th1-dominant response to the transgene.


Knowledge of gestational alcohol drinking risks: results from a survey among Italian secondary school students.
Ethanol is the most widely used drug worldwide. Its consumption has been increasing, and it is reported even during childbearing. Prenatal exposure to ethanol can lead to irreversible damages of the fetus. Knowledge about this risk could prevent these damages. There is no information about knowledge of the Italian students on this issue. Therefore the aim of this study was to describe the awareness of the Italian students attending the last year of secondary school about risk of gestational alcohol drinking for the delivering mother and the fetus. An online multiple-choice anonymous survey for students was used and e-mailed to the all Italian secondary schools. The respondents were 9.921 and the obtained results evidenced that that young females are more informed than males, and students in Northern and Central Italy are better informed than those in the South, especially on general aspects. The most of respondents knows that alcohol consumption during pregnancy can damage the fetus. However, many youngsters failed to translate this belief into the practice. In conclusion, interventions are needed to enhance knowledge and prevent these damages, and health professionals, with nurses in the first row are entitled to provide education on this topic.


Poisoning by Medical Plants.
Herbal medications are becoming increasingly popular with the impression that they cause fewer side effects in comparison with synthetic drugs; however, they may considerably contribute to acute or chronic poisoning incidents. Poison centers receive more than 100000 patients exposed to toxic plants. Most of these cases are inconsiderable toxicities involving pediatric ingestions of medicinal plants in low quantity. In most cases of serious poisonings, patients are adults who have either mistakenly consumed a poisonous plant as edible or ingested the plant regarding to its medicinal properties for therapy or toxic properties for illegal aims. In this article, we review the main human toxic plants causing mortality or the ones which account for emergency medical visits. Articles addressing "plant poisoning" in online databases were listed in order to establish the already reported human toxic cases. The current review introduces herbal plants toxicity and herb-drug interactions to warn the health professionals about possible consequences of unconscious uses of medicinal plants. The reported cases extracted from our prepared database were classified on the basis of the main toxic effects of plants, and the most prominent constituents of the plants which are responsible for specific toxic effects. Considering the long history of consumption of herbal medicines in different societies, people may wrongly think that medicinal plants are fully harmless and nontoxic. Prescription, preparation and consumption regulations of medicinal plants are not clear and strict as well as their marketing regulations that differ from country to country. The extensive and various consumption of medicinal plants without adequate observation is the most important reason for poisoning by medicinal plants.


Bridging the Gap between Social Acceptance and Ethical Acceptability.
New technology brings great benefits, but it can also create new and significant risks. When evaluating those risks in policymaking, there is a tendency to focus on social acceptance. By solely focusing on social acceptance, we could, however, overlook important ethical aspects of technological risk, particularly when we evaluate technologies with transnational and intergenerational risks. I argue that good governance of risky technology requires analyzing both social acceptance and ethical acceptability. Conceptually, these two notions are mostly complementary. Social acceptance studies are not capable of sufficiently capturing all the morally relevant features of risky technologies; ethical analyses do not typically include stakeholders' opinions, and they therefore lack the relevant empirical input for a thorough ethical evaluation. Only when carried out in conjunction are these two types of analysis relevant to national and international governance of risky technology. I discuss the Rawlsian wide reflective equilibrium as a method for marrying social acceptance and ethical acceptability. Although the rationale of my argument is broadly applicable, I will examine the case of multinational nuclear waste repositories in particular. This example will show how ethical issues may be overlooked if we focus only on social acceptance, and will provide a test case for demonstrating how the wide reflective equilibrium can help to bridge the proverbial acceptance-acceptability gap.


Primary culture of duck salt gland. II. Neurohormonal stimulation of active transport.
Primary cultures of structurally polarized sheets of avian salt gland secretory cells were mounted in Lucite chambers for transmural electrophysiological analysis. Transmural resistance values increased during the first 3 days of culture to 293 +/- 35 omega X cm2 and then decreased slowly thereafter. There was little short-circuit current (Isc) in the absence of secretagogues. Serosal addition of either carbachol or epinephrine resulted in a Isc consistent with positive charge flow from mucosa to serosa, thus demonstrating that these cell layers were capable of active ion transport in response to either cholinergic or adrenergic neurohormonal stimulation. Serosal ouabain or furosemide abolished the response to either agonist, while theophylline enhanced the response. Receptor specificity for the electrical responses was shown by selective inhibition of carbachol- and epinephrine-induced Isc by atropine and propranolol, respectively. The results demonstrate that these primary epithelial cell cultures are capable of active ion transport and are sensitive to known inhibitors of secretory transport, and suggest that intracellular coupling mechanisms for hormonal control are retained in culture. These cultures should be useful for studying mechanisms of ion secretory transport and their regulatory control.


Do the same genes underlie parallel phenotypic divergence in different Littorina saxatilis populations?
Parallel patterns of adaptive divergence and speciation are cited as powerful evidence for the role of selection driving these processes. However, it is often not clear whether parallel phenotypic divergence is underlain by parallel genetic changes. Here, we asked about the genetic basis of parallel divergence in the marine snail Littorina saxatilis, which has repeatedly evolved coexisting ecotypes adapted to either crab predation or wave action. We sequenced the transcriptome of snails of both ecotypes from three distant geographical locations (Spain, Sweden and United Kingdom) and mapped the reads to the L. saxatilis reference genome. We identified genomic regions potentially under divergent selection between ecotypes within each country, using an outlier approach based on F(ST) values calculated per locus. In line with previous studies indicating that gene reuse is generally common, we expected to find extensive sharing of outlier loci due to recent shared ancestry and gene flow between at least two of the locations in our study system. Contrary to our expectations, we found that most outliers were country specific, suggesting that much of the genetic basis of divergence is not shared among locations. However, we did find that more outliers were shared than expected by chance and that differentiation of shared outliers is often generated by the same SNPs. We discuss two mechanisms potentially explaining the limited amount of sharing we observed. First, a polygenic basis of divergent traits might allow for multiple distinct molecular mechanisms generating the same phenotypic patterns. Second, additional, location-specific axes of selection that we did not focus on in this study may produce distinct patterns of genetic divergence within each site.


A Survey of American Chiropractic Association Members' Experiences, Attitudes, and Perceptions of Practice in Integrated Health Care Settings.
The purpose of this study is to examine the self-report of experiences, attitudes, and perceived educational needs of American Chiropractic Association members regarding practice in integrated health care settings. This was a descriptive observational study of the American Chiropractic Association members. Participants completed an electronic survey reporting their current participation and interest in chiropractic integrated practice. The survey was completed in 2011 by 1142 respondents, for a response rate of 11.8%. The majority of respondents (82.9%) did not currently practice in an integrated setting, whereas 17.1% did. Those practicing in various integrated medical settings reported delivering a range of diagnostic, therapeutic, and case management services. Participation in administrative and scholarly activities was less common. Respondents not practicing in integrated settings reported being interested in delivering a very similar array of clinical services. Doctors of chiropractic practicing in hospital or outpatient medical facilities reported frequent engagement in interprofessional collaboration. Both nonintegrated and integrated respondents reported very similar educational interests on a range of clinical topics. The findings of this survey provide insight into the experiences, participation, and interests in integrated clinical practice for members of the American Chiropractic Association.


Development of quantum-based adaptive neuro-fuzzy networks.
In this study, we are concerned with a method for constructing quantum-based adaptive neuro-fuzzy networks (QANFNs) with a Takagi-Sugeno-Kang (TSK) fuzzy type based on the fuzzy granulation from a given input-output data set. For this purpose, we developed a systematic approach in producing automatic fuzzy rules based on fuzzy subtractive quantum clustering. This clustering technique is not only an extension of ideas inherent to scale-space and support-vector clustering but also represents an effective prototype that exhibits certain characteristics of the target system to be modeled from the fuzzy subtractive method. Furthermore, we developed linear-regression QANFN (LR-QANFN) as an incremental model to deal with localized nonlinearities of the system, so that all modeling discrepancies can be compensated. After adopting the construction of the linear regression as the first global model, we refined it through a series of local fuzzy if-then rules in order to capture the remaining localized characteristics. The experimental results revealed that the proposed QANFN and LR-QANFN yielded a better performance in comparison with radial basis function networks and the linguistic model obtained in previous literature for an automobile mile-per-gallon prediction, Boston Housing data, and a coagulant dosing process in a water purification plant.


Role of lipid type on morphine-stimulated diet selection in rats.
Administration of morphine is said to increase fat consumption among rats allowed to self-select nutrients. However, fats represent a diverse group of molecules, differing in metabolic and sensory properties. Despite this, lipid has yet to be manipulated as a variable in drug-stimulated nutrient selection studies. To determine whether lipid source can impact daily and morphine-stimulated (1, 3, and 10 mg/kg) diet intake, rats were provided with a choice between a high-fat and high-carbohydrate diet in three regimens in which the source of fat was varied between vegetable shortening, lard, or corn oil. Daily and morphine-stimulated diet selections were determined under all conditions. Under daily feeding conditions, rats ate more of the high-lipid diet compared with the high-carbohydrate diet when vegetable shortening or lard was the main lipid alternative, but lipid and carbohydrate intake did not differ when corn oil was the main lipid alternative. When rats were stimulated with morphine, the percentage of lipid increased relative to baseline intake only when the lipid diets were the preferred alternatives (i.e., vegetable shortening or lard). When preference between lipid and carbohydrate diets was neutral (i.e., corn oil condition), morphine did not enhance lipid consumption. These results indicate that morphine increases consumption of total energy or preferred diets and not lipid per se.


Use of non-parametric item response theory to develop a shortened version of the Positive and Negative Syndrome Scale (PANSS).
Nonparametric item response theory (IRT) was used to examine (a) the performance of the 30 Positive and Negative Syndrome Scale (PANSS) items and their options ((levels of severity), (b) the effectiveness of various subscales to discriminate among differences in symptom severity, and (c) the development of an abbreviated PANSS (Mini-PANSS) based on IRT and a method to link scores to the original PANSS. Baseline PANSS scores from 7,187 patients with Schizophrenia or Schizoaffective disorder who were enrolled between 1995 and 2005 in psychopharmacology trials were obtained. Option characteristic curves (OCCs) and Item Characteristic Curves (ICCs) were constructed to examine the probability of rating each of seven options within each of 30 PANSS items as a function of subscale severity, and summed-score linking was applied to items selected for the Mini-PANSS. The majority of items forming the Positive and Negative subscales (i.e. 19 items) performed very well and discriminate better along symptom severity compared to the General Psychopathology subscale. Six of the seven Positive Symptom items, six of the seven Negative Symptom items, and seven out of the 16 General Psychopathology items were retained for inclusion in the Mini-PANSS. Summed score linking and linear interpolation was able to produce a translation table for comparing total subscale scores of the Mini-PANSS to total subscale scores on the original PANSS. Results show scores on the subscales of the Mini-PANSS can be linked to scores on the original PANSS subscales, with very little bias. The study demonstrated the utility of non-parametric IRT in examining the item properties of the PANSS and to allow selection of items for an abbreviated PANSS scale. The comparisons between the 30-item PANSS and the Mini-PANSS revealed that the shorter version is comparable to the 30-item PANSS, but when applying IRT, the Mini-PANSS is also a good indicator of illness severity.


Stability evaluation of implants integrated in grafted and nongrafted maxillary bone: a clinical study from implant placement to abutment connection.
Clinical studies have shown a higher degree of implant failures in grafted bone compared with normal nongrafted maxillary bone. Additionally, a prolonged time for integration of titanium implants in grafted block bone has been shown by means of resonance frequency analysis (RFA). The aim of this prospective study was to compare the stability of implants placed in particulate bone, onlay block bone, interpositional bone, and nongrafted maxillary bone during the early phase of osseointegration using RFA and implant failure. Thirty-five patients with edentulism in the maxilla were included in the study. In all, 260 Astra Tech TiOblast implants (Astra Tech AB, Mölndal, Sweden) were installed. Twenty-five of these patients had severe maxillary atrophy and were treated with iliac bone grafts 5 to 6 months prior to implant placement, 19 with lateral onlay block grafts on one side (group A, 38 implants) and particulate bone for lateral augmentation on the other (group B, 38 implants). These 19 patients also got bilateral sinus floor augmentation with particulate bone (group C, 76 implants). Six patients had an unfavorable sagittal relation between the jaws and underwent a LeFort I operation with interpositional bone blocks grafted to the nasal and sinus floors (group D, 48 implants). The remaining 10 patients could be treated with implants without bone augmentation and served as control (group E, 60 implants). RFA was performed at implant placement and abutment connection 6 months later and an implant stability quotient (ISQ) value was given for each implant. Four implants (1.5%) were found mobile at abutment connection and removed (two in group A and two in group D). RFA showed a slight increase in stability from installation to abutment connection but the differences were not statistically significant in any of the groups (Wilcoxon signed rank test for comparison of paired data). Implants installed in group D had a significantly lower ISQ value at both measurements compared with the other groups (Wilcoxon Rank Sum test for comparisons of independent samples, p = .05). It is concluded that TiO(2)-blasted implants placed in nongrafted and grafted maxillary bone using a two-staged protocol show similar stability during the early phase of osseointegration. Patients reconstructed with interpositional bone graft after a LeFort I osteotomy showed lower implant stability values than nongrafted patients and other grafting techniques.


[Diabetic nephropathy. Study protocol for pre-kidney transplantation].
Between May 1990 and October 1998, 67 diabetic patients with end-stage renal disease, on dialysis, were submitted to a standardized protocol in order to assess the coexistence and degree of other diabetic and nondiabetic complications that could affect transplantation. Some of the results were analysed. Type 2 diabetic patients had more abnormal results on the lower limbs doppler ultrasound and on the lower limbs arteriography than type 1 (p < 0.05). Type 2 diabetic patients had more cardiovascular complications so the decision to transplant should be taken on a case by case basis.


Mesodermal subdivision along the mediolateral axis in chicken controlled by different concentrations of BMP-4.
Molecular mechanisms by which the mesoderm is subdivided along the mediolateral axis in early chicken embryos have been studied. When the presomitic mesoderm (medial mesoderm) was transplanted into the lateral plate, the graft was transformed into lateral plate tissue, indicating that the primitive somite was not fully committed and that the lateral plate has a cue for mesodermal lateralization. Since the lateral plate expresses a high level of BMP-4 mRNA, a member of the TGF-beta family, we hypothesized that it is the molecule responsible for the lateralization of the somite. To test this, we transplanted COS cells producing BMP-4 into the presomitic region. Those cells locally prevented the presomitic cells from differentiating into somites, converting them instead into lateral plate mesoderm, which was revealed by expression of cytokeratin mRNA, a marker for the lateral plate. The effect was dependent on the level of effective BMP-4: with a high level of BMP-4, the somite was transformed completely to lateral plate; with a low level, the somite formed but was occupied by the lateral somitic component expressing cSim 1, a marker for the lateral somite. These results suggest that different thresholds of effective BMP-4 determine distinct subtypes of the mesoderm as a lateralizer during early development.


Addressing the sexual health needs of patients with gynecologic cancers.
Research indicates that sexual health and function frequently are overlooked by healthcare professionals despite being identified as an essential aspect of patient care. This article discusses the topic of sexuality as it pertains to women with gynecologic cancers, highlights barriers to addressing sexuality with patients with cancer, and provides nurses with a framework to guide them in providing information and support to patients who are experiencing sexual difficulties or who need information about sexual function. A case study addresses the challenges a woman with cervical cancer faces and the role of the nurse in supporting the patient and her partner within the PLISSIT model.


Locally Advanced Pancreatic Cancer: Work-Up, Staging, and Local Intervention Strategies.
Locally advanced pancreatic cancer (LAPC) has several definitions but essentially is a nonmetastasized pancreatic cancer, in which upfront resection is considered not beneficial due to extensive vascular involvement and consequent high chance of a nonradical resection. The introduction of FOLFIRINOX chemotherapy and gemcitabine-nab-paclitaxel (gem-nab) has had major implications for the management and outcome of patients with LAPC. After 4-6 months induction chemotherapy, the majority of patients have stable disease or even tumor-regression. Of these, 12 to 35% are successfully downstaged to resectable disease. Several studies have reported a 30-35 months overall survival after resection; although it currently remains unclear if this is a result of the resection or the good response to chemotherapy. Following chemotherapy, selection of patients for resection is difficult, as contrast-enhanced computed-tomography (CT) scan is unreliable in differentiating between viable tumor and fibrosis. In case a resection is not considered possible but stable disease is observed, local ablative techniques are being studied, such as irreversible electroporation, radiofrequency ablation, and stereotactic body radiation therapy. Pragmatic, multicenter, randomized studies will ultimately have to confirm the exact role of both surgical exploration and ablation in these patients. Since evidence-based guidelines for the management of LAPC are lacking, this review proposes a standardized approach for the treatment of LAPC based on the best available evidence.


A prospective study of plasma vitamin D metabolites, vitamin D receptor polymorphisms, and prostate cancer.
Vitamin D insufficiency is a common public health problem nationwide. Circulating 25-hydroxyvitamin D3 (25[OH]D), the most commonly used index of vitamin D status, is converted to the active hormone 1,25 dihydroxyvitamin D3 (1,25[OH]2D), which, operating through the vitamin D receptor (VDR), inhibits in vitro cell proliferation, induces differentiation and apoptosis, and may protect against prostate cancer. Despite intriguing results from laboratory studies, previous epidemiological studies showed inconsistent associations of circulating levels of 25(OH)D, 1,25(OH)2D, and several VDR polymorphisms with prostate cancer risk. Few studies have explored the joint association of circulating vitamin D levels with VDR polymorphisms. During 18 y of follow-up of 14,916 men initially free of diagnosed cancer, we identified 1,066 men with incident prostate cancer (including 496 with aggressive disease, defined as stage C or D, Gleason 7-10, metastatic, and fatal prostate cancer) and 1,618 cancer-free, age- and smoking-matched control participants in the Physicians' Health Study. We examined the associations of prediagnostic plasma levels of 25(OH)D and 1,25(OH)2D, individually and jointly, with total and aggressive disease, and explored whether relations between vitamin D metabolites and prostate cancer were modified by the functional VDR FokI polymorphism, using conditional logistic regression. Among these US physicians, the median plasma 25(OH)D levels were 25 ng/ml in the blood samples collected during the winter or spring and 32 ng/ml in samples collected during the summer or fall. Nearly 13% (summer/fall) to 36% (winter/spring) of the control participants were deficient in 25(OH)D (<20 ng/ml) and 51% (summer/fall) and 77% (winter/spring) had insufficient plasma 25(OH)D levels (<32 ng/ml). Plasma levels of 1,25(OH)2D did not vary by season. Men whose levels for both 25(OH)D and 1,25(OH)2D were below (versus above) the median had a significantly increased risk of aggressive prostate cancer (odds ratio [OR] = 2.1, 95% confidence interval [CI] 1.2-3.4), although the interaction between the two vitamin D metabolites was not statistically significant (pinteraction = 0.23). We observed a significant interaction between circulating 25(OH)D levels and the VDR FokI genotype (pinteraction < 0.05). Compared with those with plasma 25(OH)D levels above the median and with the FokI FF or Ff genotype, men who had low 25(OH)D levels and the less functional FokI ff genotype had increased risks of total (OR = 1.9, 95% CI 1.1-3.3) and aggressive prostate cancer (OR = 2.5, 95% CI 1.1-5.8). Among men with plasma 25(OH)D levels above the median, the ff genotype was no longer associated with risk. Conversely, among men with the ff genotype, high plasma 25(OH)D level (above versus below the median) was related to significant 60% approximately 70% lower risks of total and aggressive prostate cancer. Our data suggest that a large proportion of the US men had suboptimal vitamin D status (especially during the winter/spring season), and both 25(OH)D and 1,25(OH)2D may play an important role in preventing prostate cancer progression. Moreover, vitamin D status, measured by 25(OH)D in plasma, interacts with the VDR FokI polymorphism and modifies prostate cancer risk. Men with the less functional FokI ff genotype (14% in the European-descent population of this cohort) are more susceptible to this cancer in the presence of low 25(OH)D status.


Alterations in Whole-Body Insulin Sensitivity Resulting From Repeated Eccentric Exercise of a Single Muscle Group: A Pilot Investigation.
Unaccustomed eccentric exercise using large muscle groups elicits soreness, decrements in physical function and impairs markers of whole-body insulin sensitivity; although these effects are attenuated with a repeated exposure. Eccentric exercise of a small muscle group (elbow flexors) displays similar soreness and damage profiles in response to repeated exposure. However, it is unknown whether damage to small muscle groups impacts upon whole-body insulin sensitivity. This pilot investigation aimed to characterize whole-body insulin sensitivity in response to repeated bouts of eccentric exercise of the elbow flexors. Nine healthy males completed two bouts of eccentric exercise separated by 2 weeks. Insulin resistance (updated homeostasis model of insulin resistance, HOMA2-IR) and muscle damage profiles (soreness and physical function) were assessed before, and 48 h after exercise. Matsuda insulin sensitivity indices (ISI Matsuda) were also determined in 6 participants at the same time points as HOMA2-IR. Soreness was elevated, and physical function impaired, by both bouts of exercise (both p < .05) but to a lesser extent following bout 2 (time x bout interaction, p < .05). Eccentric exercise decreased ISI Matsuda after the first but not the second bout of eccentric exercise (time x bout interaction p < .05). Eccentric exercise performed with an isolated upper limb impairs whole-body insulin sensitivity after the first, but not the second, bout.


The human anterior tympanic artery. A nutrient artery of the middle ear with highly variable origin.
The variability of the origin of the anterior tympanic artery was investigated in 104 individuals of both sexes. A surprising laterality was found: thus, while the left anterior tympanic artery originated as a singular vessel from either the maxillary or the superficial temporal artery with almost equal frequencies (44.7 and 45.9%, respectively), the right anterior tympanic artery predominantly branched off from the maxillary artery (77.8% of cases). Besides the origin from either the maxillary artery or the superficial temporal artery, also anterior tympanic arteries branching off from the external carotid artery were found to occur (4% on the left and 1% on the right side). Although in the majority of individuals, a singular anterior tympanic artery occurred within the infratemporal fossa, duplications of the anterior tympanic artery were found to be present: in one case on the right and in 8 cases on their left side. In 1 female individual, a triplet of left anterior tympanic arteries was found to supply the tympanic cavity. Also in these cases, the anterior tympanic artery arose from either the external carotid, the superficial temporal or the maxillary artery. In singular cases, even several other branches of the maxillary artery, viz. the deep auricular, middle, and accessory meningeal, as well as the posterior deep temporal, inferior alveolar and masseteric arteries were found to form common trunks with the anterior tympanic artery.


Gut microbiota regulates NKG2D ligand expression on intestinal epithelial cells.
Intestinal epithelial cells (IECs) are one of a few cell types in the body with constitutive surface expression of natural killer group 2 member D (NKG2D) ligands, although the magnitude of ligand expression by IECs varies. Here, we investigated whether the gut microbiota regulates the NKG2D ligand expression on small IECs. Germ-free and ampicillin-treated mice were shown to have a significant increase in NKG2D ligand expression. Interestingly, vancomycin treatment, which propagated the bacterium Akkermansia muciniphila and reduced the level of IFN-γ and IL-15 in the intestine, decreased the NKG2D ligand expression on IECs. In addition, a similar increase in A. muciniphila and a decreased NKG2D ligand expression was seen after feeding with dietary xylooligosaccharides. A pronounced increase in NKG2D ligand expression was furthermore observed in IL-10-deficient mice. In summary, our results suggest that the constitutive levels of NKG2D ligand expression on IECs are regulated by microbial signaling in the gut and further disfavor the intuitive notion that IEC NKG2D ligand expression is caused by low-grade immune reaction against commensal bacteria. It is more likely that constitutively high IEC NKG2D ligand expression is kept in check by an intestinal regulatory immune milieu induced by members of the gut microbiota, for example A. muciniphila.


Proteasomal degradation of herpes simplex virus capsids in macrophages releases DNA to the cytosol for recognition by DNA sensors.
The innate immune system is important for control of infections, including herpesvirus infections. Intracellular DNA potently stimulates antiviral IFN responses. It is known that plasmacytoid dendritic cells sense herpesvirus DNA in endosomes via TLR9 and that nonimmune tissue cells can sense herpesvirus DNA in the nucleus. However, it remains unknown how and where myeloid cells, such as macrophages and conventional dendritic cells, detect infections with herpesviruses. In this study, we demonstrate that the HSV-1 capsid was ubiquitinated in the cytosol and degraded by the proteasome, hence releasing genomic DNA into the cytoplasm for detection by DNA sensors. In this context, the DNA sensor IFN-γ-inducible 16 is important for induction of IFN-β in human macrophages postinfection with HSV-1 and CMV. Viral DNA localized to the same cytoplasmic regions as did IFN-γ-inducible 16, with DNA sensing being independent of viral nuclear entry. Thus, proteasomal degradation of herpesvirus capsids releases DNA to the cytoplasm for recognition by DNA sensors.


Molecular mechanisms of EAST/SeSAME syndrome mutations in Kir4.1 (KCNJ10).
Inwardly rectifying potassium channel Kir4.1 is critical for glial function, control of neuronal excitability, and systemic K(+) homeostasis. Novel mutations in Kir4.1 have been associated with EAST/SeSAME syndrome, characterized by mental retardation, ataxia, seizures, hearing loss, and renal salt waste. Patients are homozygous for R65P, G77R, C140R or T164I; or compound heterozygous for A167V/R297C or R65P/R199Stop, a deletion of the C-terminal half of the protein. We investigated the functional significance of these mutations by radiotracer efflux and inside-out membrane patch clamping in COSm6 cells expressing homomeric Kir4.1 or heteromeric Kir4.1/Kir5.1 channels. All of the mutations compromised channel function, but the underlying mechanisms were different. R65P, T164I, and R297C caused an alkaline shift in pH sensitivity, indicating that these positions are crucial for pH sensing and pore gating. In R297C, this was due to disruption of intersubunit salt bridge Glu(288)-Arg(297). C140R breaks the Cys(108)-Cys(140) disulfide bond essential for protein folding and function. A167V did not affect channel properties but may contribute to decreased surface expression in A167V/R297C. In G77R, introduction of a positive charge within the bilayer may affect channel structure or gating. R199Stop led to a dramatic decrease in surface expression, but channel activity was restored by co-expression with intact subunits, suggesting remarkable tolerance for truncation of the cytoplasmic domain. These results provide an explanation for the molecular defects that underlie the EAST/SeSAME syndrome.


Adult open heart surgery in New York State. An analysis of risk factors and hospital mortality rates.
This study analyzes data from New York State's new Cardiac Surgery Reporting System, which contains information about cardiac preoperative risk factors, postoperative complications, and hospital discharge. The purposes of the study were to determine the set of significant clinical risk factors and to identify cardiac surgical centers most likely to have serious quality-of-care problems. Significant risk factors for in-hospital death were age, gender, ejection fraction, previous myocardial infarction, number of open heart operations in previous admissions, diabetes requiring medication, dialysis dependence, disasters (acute structural defect, renal failure, cardiogenic shock, gunshot), unstable angina, intractable congestive heart failure, left main trunk narrowed more than 90%, and type of operation performed. Four of the 28 hospitals had significantly higher mortality rates than expected, given the risk factors of their patients. Subsequent site visits and medical record reviews confirmed that these facilities had high percentages of quality-of-care problems among cases resulting in mortality.


Chewing of betel, areca and tobacco: perceptions and knowledge regarding their role in head and neck cancers in an urban squatter settlement in Pakistan.
The link of betel, areca and chewable tobacco with head and neck cancers is clearly established. Fifty eight percent of the global head and neck cancers occur in South and Southeast Asia, where chewing of betel, areca and tobacco are common. This study was carried out to establish the pattern of use of Paan, Chaalia, Gutka, Niswar, Tumbaku and Naas among population of squatter settlement of Karachi and to determine the perceptions and knowledge regarding their role in the etiology of head and neck cancers. It was a cross-sectional study, performed at Bilal colony in Karachi. Through systematic sampling, 425 subjects [a male and female from a household] were interviewed with a structured questionnaire. Knowledge regarding etiology of head and neck cancers was classified in ordinals of "good", "some" and "poor", for each substance separately, while practices were classified into "daily user", "occasional user" and "never user". About 40% of the participants were chewing at least one item [betel, areca or tobacco products] on daily basis. This prevalence was 2.46 times higher among males than females and 1.39 times higher among adolescents than adults. At least 79% of the participants were classified as having poor knowledge about the carcinogenicity of each of these items. Knowledge increased with age and level of education. Health hazards of these items were poorly recognized and about 20% perceived at least one of these items to be beneficial. Positive attitudes were seen regarding the steps to curb the production, business and consumption of these substances. In conclusion, prevalence of chewing of betel, areca and tobacco among adults and adolescents is high. Deficiency in knowledge and wrong perception of favorable effect of chewing products is common. Besides curtailing the availability of chewing products, correct knowledge regarding its ill-effects should be inculcated among population to decrease the burden of head and neck cancers.


[Effect of phenolic compounds of virgin olive oil on LDL oxidation resistance].
Several epidemiological and experimental studies have associated the intake of antioxidants, which are abundant in the Mediterranean diet, with a low incidence of cardiovascular disease. One possible mechanism of this action is the oxidative protection in low density lipoproteins (LDL). The aim of our study was to compare the antioxidative activity of diverse phenolic compounds present in virgin olive oil on these lipoproteins. LDL was isolated from blood plasma of healthy volunteers by sequential ultracentrifugation. This was followed by oxidation with CuC12 in the presence of different concentrations of phenolic compounds and virgin olive oil extract. Production of conjugated dienes was determined by the continuous monitoring of increased absorbency at 234 nm as an indicator of LDL oxidation. Virgin olive oil extract prolonged the latency phase and significantly lowered the progression rate (p < 0.05) at low concentrations (2 g/ml). This antioxidative effect was also observed with low concentrations (2 M) of caffeic acid and oleuropein (p < 0.05). However, it was necessary to increase the concentration of flavone up to 50 times to observe a similar effect (p < 0.05). Both virgin olive oil extract enriched in phenolic compounds and phenolic compounds present in olive oil (caffeic acid and oleuropein) are potent antioxidants at very low concentrations. Thus, the beneficial effects of a Mediterranean diet may be partly due to the protective action of these compounds.


An endogenous F-box protein regulates ARGONAUTE1 in Arabidopsis thaliana.
ARGONAUTE1 (AGO1) mediates microRNA- and small interfering RNA-directed posttranscriptional gene silencing in Arabidopsis thaliana. Mutant alleles of SQUINT (SQN) slightly reduce AGO1 activity and have weak effects on shoot morphology. A screen for mutations that suppress the sqn phenotype produced loss-of-function mutations in the F-box gene FBW2. Mutations in FBW2 not only suppress sqn but also suppress many of the developmental phenotypes of weak, but not null, alleles of AGO1 by increasing AGO1 protein levels. Conversely, over-expression of FBW2 decreases the abundance of the AGO1 protein but not AGO1 messenger RNA, further indicating that FBW2 regulates AGO1 protein levels. fbw2 mutants have no obvious morphological phenotype, but display a reduced sensitivity to abscisic acid (ABA) that can be attributed to increased AGO1 activity. Our results indicate that FBW2 is a novel negative regulator of AGO1 and suggest that it plays a role in ABA signalling and/or response.


Nox4 and redox signaling mediate TGF-β-induced endothelial cell apoptosis and phenotypic switch.
Transforming growth factor-β (TGF-β) triggers apoptosis in endothelial cells, while the mechanisms underlying this action are not entirely understood. Using genetic and pharmacological tools, we demonstrated that TGF-β induced a moderate apoptotic response in human cultured endothelial cells, which was dependent upon upregulation of the Nox4 NADPH oxidase and production of reactive oxygen species (ROS). In contrast, we showed that ectopic expression of Nox4 via viral vectors (vNox4) produced an antiapoptotic effect. TGF-β caused ROS-dependent p38 activation, whereas inhibition of p38 blunted TGF-β-induced apoptosis. However, vNox4, but not TGF-β, activated Akt, and inhibition of Akt attenuated the antiapoptotic effect of vNox4. Akt activation induced by vNox4 was accompanied by inactivation of the protein tyrosine phosphatase-1B (PTP1B) function and enhanced vascular endothelial growth factor receptor (VEGFR)-2 phosphorylation. Moreover, we showed that TGF-β enhanced Notch signaling and increased expression of the arterial marker EphrinB2 in a redox-dependent manner. In summary, our results suggest that Nox4 and ROS have pivotal roles in mediating TGF-β-induced endothelial apoptosis and phenotype specification. Redox mechanisms may influence endothelial cell functions by modulating p38, PTP1B/VEGFR/Akt and Notch signaling pathways.


Transfer of newborns to neonatal care unit: a registry based study in Northern Tanzania.
Reduction in neonatal mortality has been slower than anticipated in many low income countries including Tanzania. Adequate neonatal care may contribute to reduced mortality. We studied factors associated with transfer of babies to a neonatal care unit (NCU) in data from a birth registry at Kilimanjaro Christian Medical Centre (KCMC) in Tanzania. A total of 21 206 singleton live births registered from 2000 to 2008 were included. Multivariable analysis was carried out to study neonatal transfer to NCU by socio-demographic factors, pregnancy complications and measures of the condition of the newborn. A total of 3190 (15%) newborn singletons were transferred to the NCU. As expected, neonatal transfer was strongly associated with specific conditions of the baby including birth weight above 4000 g (relative risk (RR) = 7.2; 95% confidence interval (CI) 6.5-8.0) or below 1500 g (RR = 3.0; 95% CI: 2.3-4.0), five minutes Apgar score less than 7 (RR = 4.0; 95% CI: 3.4-4.6), and preterm birth before 34 weeks of gestation (RR = 1.8; 95% CI: 1.5-2.1). However, pregnancy- and delivery-related conditions like premature rupture of membrane (RR = 2.3; 95% CI: 1.9-2.7), preeclampsia (RR = 1.3; 95% CI: 1.1-1.5), other vaginal delivery (RR = 2.2; 95% CI: 1.7-2.9) and caesarean section (RR = 1.9; 95% CI: 1.8-2.1) were also significantly associated with transfer. Birth to a first born child was associated with increased likelihood of transfer (relative risk (RR) 1.4; 95% CI: 1.2-1.5), while the likelihood was reduced (RR = 0.5; 95% CI: 0.3-0.9) when the father had no education. In addition to strong associations between neonatal transfer and classical neonatal risk factors for morbidity and mortality, some pregnancy-related and demographic factors were predictors of neonatal transfer. Overall, transfer was more likely for babies with signs of poor health status or a complicated pregnancy. Except for a possibly reduced use of transfer for babies of non-educated fathers and a high transfer rate for first born babies, there were no signs that transfer was based on non-medical indications.


Identification of human, rat and chicken ribosomal proteins by a combination of two-dimensional polyacrylamide gel electrophoresis and mass spectrometry.
To identify the exact spot position of human, rat and chicken ribosomal proteins (RP) separated by two-dimensional polyacrylamide gel electrophoresis (2-DE), a 2-DE system was designed to separate RP with a pI>8.6 according to their charge in the first dimension and to their molecular mass in the second dimension. Individual proteins were excised from the gels and identified by mass spectrometry after digestion by trypsin. In addition, a mixture of purified RP from these three species was also analyzed by tandem mass tag spectrometry. By combining those two methods 74 RP from human, 76 from rat and 67 from chicken were identified according to the nomenclature initially defined for rat liver RP and by using the Swiss-Prot/trEMBL databases. Whereas human and rat RP were well described, most of RP from chicken were not characterized in databases, since 35 out of 67 chicken RP identified in this study were not listed yet. We propose here the first comprehensive description of chicken RP and their comparison to those from human and rat.


Structural Analysis using SHALiPE to Reveal RNA G-Quadruplex Formation in Human Precursor MicroRNA.
RNA G-quadruplex (rG4) structures are of fundamental importance to biology. A novel approach is introduced to detect and structurally map rG4s at single-nucleotide resolution in RNAs. The approach, denoted SHALiPE, couples selective 2'-hydroxyl acylation with lithium ion-based primer extension, and identifies characteristic structural fingerprints for rG4 mapping. We apply SHALiPE to interrogate the human precursor microRNA 149, and reveal the formation of an rG4 structure in this non-coding RNA. Additional analyses support the SHALiPE results and uncover that this rG4 has a parallel topology, is thermally stable, and is conserved in mammals. An in vitro Dicer assay shows that this rG4 inhibits Dicer processing, supporting the potential role of rG4 structures in microRNA maturation and post-transcriptional regulation of mRNAs.


Mitochondrial DNA mutations in respiratory complex-I in never-smoker lung cancer patients contribute to lung cancer progression and associated with EGFR gene mutation.
Mitochondrial DNA (mtDNA) mutations were reported in different cancers. However, the nature and role of mtDNA mutation in never-smoker lung cancer patients including patients with epidermal growth factor receptor (EGFR) and KRAS gene mutation are unknown. In the present study, we sequenced entire mitochondrial genome (16.5 kb) in matched normal and tumors obtained from 30 never-smoker and 30 current-smoker lung cancer patients, and determined the mtDNA content. All the patients' samples were sequenced for KRAS (exon 2) and EGFR (exon 19 and 21) gene mutation. The impact of forced overexpression of a respiratory complex-I gene mutation was evaluated in a lung cancer cell line. We observed significantly higher (P = 0.006) mtDNA mutation in the never-smokers compared to the current-smoker lung cancer patients. MtDNA mutation was significantly higher (P = 0.026) in the never-smoker Asian compared to the current-smoker Caucasian patients' population. MtDNA mutation was significantly (P = 0.007) associated with EGFR gene mutation in the never-smoker patients. We also observed a significant increase (P = 0.037) in mtDNA content among the never-smoker lung cancer patients. The majority of the coding mtDNA mutations targeted respiratory complex-I and forced overexpression of one of these mutations resulted in increased in vitro proliferation, invasion, and superoxide production in lung cancer cells. We observed a higher prevalence and new relationship between mtDNA alterations among never-smoker lung cancer patients and EGFR gene mutation. Moreover, a representative mutation produced strong growth effects after forced overexpression in lung cancer cells. Signature mtDNA mutations provide a basis to develop novel biomarkers and therapeutic strategies for never-smoker lung cancer patients.


Dorsolateral and ventromedial prefrontal cortex orchestrate normative choice.
Humans are noted for their capacity to over-ride self-interest in favor of normatively valued goals. We examined the neural circuitry that is causally involved in normative, fairness-related decisions by generating a temporarily diminished capacity for costly normative behavior, a 'deviant' case, through non-invasive brain stimulation (repetitive transcranial magnetic stimulation) and compared normal subjects' functional magnetic resonance imaging signals with those of the deviant subjects. When fairness and economic self-interest were in conflict, normal subjects (who make costly normative decisions at a much higher frequency) displayed significantly higher activity in, and connectivity between, the right dorsolateral prefrontal cortex (DLPFC) and the posterior ventromedial prefrontal cortex (pVMPFC). In contrast, when there was no conflict between fairness and economic self-interest, both types of subjects displayed identical neural patterns and behaved identically. These findings suggest that a parsimonious prefrontal network, the activation of right DLPFC and pVMPFC, and the connectivity between them, facilitates subjects' willingness to incur the cost of normative decisions.


Evaluation of three parasite lactate dehydrogenase-based rapid diagnostic tests for the diagnosis of falciparum and vivax malaria.
In areas where non-falciparum malaria is common rapid diagnostic tests (RDTs) capable of distinguishing malaria species reliably are needed. Such tests are often based on the detection of parasite lactate dehydrogenase (pLDH). In Dawei, southern Myanmar, three pLDH based RDTs (CareStart Malaria pLDH (Pan), CareStart Malaria pLDH (Pan, Pf) and OptiMAL-IT)were evaluated in patients presenting with clinically suspected malaria. Each RDT was read independently by two readers. A subset of patients with microscopically confirmed malaria had their RDTs repeated on days 2, 7 and then weekly until negative. At the end of the study, samples of study batches were sent for heat stability testing. Between August and November 2007, 1004 patients aged between 1 and 93 years were enrolled in the study. Slide microscopy (the reference standard) diagnosed 213 Plasmodium vivax (Pv) monoinfections, 98 Plasmodium falciparum (Pf) mono-infections and no malaria in 650 cases. The sensitivities (sens) and specificities (spec), of the RDTs for the detection of malaria were- CareStart Malaria pLDH (Pan) test: sens 89.1% [CI95 84.2-92.6], spec 97.6% [CI95 96.5-98.4]. OptiMal-IT: Pf+/- other species detection: sens 95.2% [CI95 87.5-98.2], spec 94.7% [CI95 93.3-95.8]; non-Pf detection alone: sens 89.6% [CI95 83.6-93.6], spec 96.5% [CI95 94.8-97.7]. CareStart Malaria pLDH (Pan, Pf): Pf+/- other species: sens 93.5% [CI95 85.4-97.3], spec 97.4% [95.9-98.3]; non-Pf: sens 78.5% [CI95 71.1-84.4], spec 97.8% [CI95 96.3-98.7]. Inter-observer agreement was excellent for all tests (kappa > 0.9). The median time for the RDTs to become negative was two days for the CareStart Malaria tests and seven days for OptiMAL-IT. Tests were heat stable up to 90 days except for OptiMAL-IT (Pf specific pLDH stable to day 20 at 35 degrees C). None of the pLDH-based RDTs evaluated was able to detect non-falciparum malaria with high sensitivity, particularly at low parasitaemias. OptiMAL-IT performed best overall and would perform best in an area of high malaria prevalence among screened fever cases. However, heat stability was unacceptable and the number of steps to perform this test is a significant drawback in the field. A reliable, heat-stable, highly sensitive RDT, capable of diagnosing all Plasmodium species has yet to be identified.


Comparison of height acceleration curves in the Fels, Zurich, and Berkeley growth data.
A new type of smoothing spline designed for nonparametric estimation of growth acceleration curves was applied to longitudinal measures of height from the Fels, Zurich, and Berkeley growth studies. Landmark features of the individual acceleration curves were located and examined for site effects and site by sex interaction. The curves in each group were structurally averaged by transforming the age scale of each child to align the landmarks with overall means. The modal shapes of the structural average curves were found to be highly similar in the three studies, although the relative number of children who conformed to the modal pattern differed among the studies. Significant site by sex interactions were found in the timing of the landmarks associated with the pubertal growth spurt and with the prepubertal 'midspurt'.


A Cellular Mechanism to Detect and Alleviate Reductive Stress.
Metazoan organisms rely on conserved stress response pathways to alleviate adverse conditions and preserve cellular integrity. Stress responses are particularly important in stem cells that provide lifetime support for tissue formation and repair, but how these protective systems are integrated into developmental programs is poorly understood. Here we used myoblast differentiation to identify the E3 ligase CUL2FEM1B and its substrate FNIP1 as core components of the reductive stress response. Reductive stress, as caused by prolonged antioxidant signaling or mitochondrial inactivity, reverts the oxidation of invariant Cys residues in FNIP1 and allows CUL2FEM1B to recognize its target. The ensuing proteasomal degradation of FNIP1 restores mitochondrial activity to preserve redox homeostasis and stem cell integrity. The reductive stress response is therefore built around a ubiquitin-dependent rheostat that tunes mitochondrial activity to redox needs and implicates metabolic control in coordination of stress and developmental signaling.


Large-Scale Off-Target Identification Using Fast and Accurate Dual Regularized One-Class Collaborative Filtering and Its Application to Drug Repurposing.
Target-based screening is one of the major approaches in drug discovery. Besides the intended target, unexpected drug off-target interactions often occur, and many of them have not been recognized and characterized. The off-target interactions can be responsible for either therapeutic or side effects. Thus, identifying the genome-wide off-targets of lead compounds or existing drugs will be critical for designing effective and safe drugs, and providing new opportunities for drug repurposing. Although many computational methods have been developed to predict drug-target interactions, they are either less accurate than the one that we are proposing here or computationally too intensive, thereby limiting their capability for large-scale off-target identification. In addition, the performances of most machine learning based algorithms have been mainly evaluated to predict off-target interactions in the same gene family for hundreds of chemicals. It is not clear how these algorithms perform in terms of detecting off-targets across gene families on a proteome scale. Here, we are presenting a fast and accurate off-target prediction method, REMAP, which is based on a dual regularized one-class collaborative filtering algorithm, to explore continuous chemical space, protein space, and their interactome on a large scale. When tested in a reliable, extensive, and cross-gene family benchmark, REMAP outperforms the state-of-the-art methods. Furthermore, REMAP is highly scalable. It can screen a dataset of 200 thousands chemicals against 20 thousands proteins within 2 hours. Using the reconstructed genome-wide target profile as the fingerprint of a chemical compound, we predicted that seven FDA-approved drugs can be repurposed as novel anti-cancer therapies. The anti-cancer activity of six of them is supported by experimental evidences. Thus, REMAP is a valuable addition to the existing in silico toolbox for drug target identification, drug repurposing, phenotypic screening, and side effect prediction. The software and benchmark are available at https://github.com/hansaimlim/REMAP.


SNP calling by sequencing pooled samples.
Performing high throughput sequencing on samples pooled from different individuals is a strategy to characterize genetic variability at a small fraction of the cost required for individual sequencing. In certain circumstances some variability estimators have even lower variance than those obtained with individual sequencing. SNP calling and estimating the frequency of the minor allele from pooled samples, though, is a subtle exercise for at least three reasons. First, sequencing errors may have a much larger relevance than in individual SNP calling: while their impact in individual sequencing can be reduced by setting a restriction on a minimum number of reads per allele, this would have a strong and undesired effect in pools because it is unlikely that alleles at low frequency in the pool will be read many times. Second, the prior allele frequency for heterozygous sites in individuals is usually 0.5 (assuming one is not analyzing sequences coming from, e.g. cancer tissues), but this is not true in pools: in fact, under the standard neutral model, singletons (i.e. alleles of minimum frequency) are the most common class of variants because P(f) ∝ 1/f and they occur more often as the sample size increases. Third, an allele appearing only once in the reads from a pool does not necessarily correspond to a singleton in the set of individuals making up the pool, and vice versa, there can be more than one read - or, more likely, none - from a true singleton. To improve upon existing theory and software packages, we have developed a Bayesian approach for minor allele frequency (MAF) computation and SNP calling in pools (and implemented it in a program called snape): the approach takes into account sequencing errors and allows users to choose different priors. We also set up a pipeline which can simulate the coalescence process giving rise to the SNPs, the pooling procedure and the sequencing. We used it to compare the performance of snape to that of other packages. We present a software which helps in calling SNPs in pooled samples: it has good power while retaining a low false discovery rate (FDR). The method also provides the posterior probability that a SNP is segregating and the full posterior distribution of f for every SNP. In order to test the behaviour of our software, we generated (through simulated coalescence) artificial genomes and computed the effect of a pooled sequencing protocol, followed by SNP calling. In this setting, snape has better power and False Discovery Rate (FDR) than the comparable packages samtools, PoPoolation, Varscan : for N = 50 chromosomes, snape has power ≈ 35%and FDR ≈ 2.5%. snape is available at http://code.google.com/p/snape-pooled/ (source code and precompiled binaries).


Crystal structure of a sulfur carrier protein complex found in the cysteine biosynthetic pathway of Mycobacterium tuberculosis.
The structure of the protein complex CysM-CysO from a new cysteine biosynthetic pathway found in the H37Rv strain of Mycobacterium tuberculosis has been determined at 1.53 A resolution. CysM (Rv1336) is a PLP-containing beta-replacement enzyme and CysO (Rv1335) is a sulfur carrier protein with a ubiquitin-like fold. CysM catalyzes the replacement of the acetyl group of O-acetylserine by CysO thiocarboxylate to generate a protein-bound cysteine that is released in a subsequent proteolysis reaction. The protein complex in the crystal structure is asymmetric with one CysO protomer binding to one end of a CysM dimer. Additionally, the structures of CysM and CysO were determined individually at 2.8 and 2.7 A resolution, respectively. Sequence alignments with homologues and structural comparisons with CysK, a cysteine synthase that does not utilize a sulfur carrier protein, revealed high conservation of active site residues; however, residues in CysM responsible for CysO binding are not conserved. Comparison of the CysM-CysO binding interface with other sulfur carrier protein complexes revealed a similarity in secondary structural elements that contribute to complex formation in the ThiF-ThiS and MoeB-MoaD systems, despite major differences in overall folds. Comparison of CysM with and without bound CysO revealed conformational changes associated with CysO binding.


Effect of the time of diagnosis on outcome in patients with acute venous thromboembolism. Findings from the RIETE Registry.
The influence of the day of diagnosis (weekends vs. weekdays) on outcome in patients with acute venous thromboembolism (VTE) has not been thoroughly studied. We used the RIETE database to compare the clinical characteristics, treatment details, and mortality rate at 7 and 30 days, of all patients diagnosed with acute VTE on weekends versus those diagnosed on weekdays. Up to January 2010, 30,394 patients were included in RIETE, of whom 5,479 (18%) were diagnosed on weekends. Most clinical characteristics were similar in both groups, but patients diagnosed on weekends had less often cancer (20% vs. 22%; p=0.004), and presented more likely with pulmonary embolism (PE) than those diagnosed on weekdays (52% vs. 47%; p <0.001). Most patients in both groups received initial therapy with low-molecular-weight heparin (90% and 91%, respectively; p=0.01), then switched to vitamin K antagonists (72% and 71%, respectively; p=0.007). The 7-day mortality rate in patients presenting with PE was 2.75% in those diagnosed on weekends versus 3.00% in those diagnosed on weekdays (p=0.49). At 30 days, the mortality rate was 6.51% versus 6.06%, respectively (p=0.38). In patients presenting with deep vein thrombosis alone, the 7-day mortality rate in those diagnosed on weekends was 1.04% versuss 0.66% in those diagnosed on weekdays (p=0.053). The mortality rate at 30 days was of 3.41% versus 2.88% (p=0.14), respectively. In RIETE, the clinical characteristics, treatment strategies, and 7- and 30-day mortality rates of patients diagnosed on weekends were similar to those in patients diagnosed on weekdays.


A systematic review of co-responder models of police mental health 'street' triage.
Police mental health street triage is an increasingly common intervention when dealing with police incidents in which there is a suspected mental health component. We conducted a systematic review of street triage interventions with three aims. First, to identify papers reporting on models of co-response police mental health street triage. Second, to identify the characteristics of service users who come in to contact with these triage services. Third, to evaluate the effectiveness of co-response triage services. We conducted a systematic review. We searched the following databases: Ovid MEDLINE, Embase, PsycINFO, EBSCO CINAHL, Scopus, Thompson Reuters Web of Science Core Collection, The Cochrane Library, ProQuest National Criminal Justice Reference Service Abstracts, ProQuest Dissertations & Theses, EThoS, and OpenGrey. We searched reference and citation lists. We also searched for other grey literature through Google, screening the first 100 PDFs of each of our search terms. We performed a narrative synthesis of our results. Our search identified 11,553 studies. After screening, 26 were eligible. Over two-thirds (69%) had been published within the last 3 years. We did not identify any randomised control trials. Results indicated that street triage might reduce the number of people taken to a place of safety under S136 of the Mental Health Act where that power exists, or reduce the use of police custody in other jurisdictions. There remains a lack of evidence to evaluate the effectiveness of street triage and the characteristics, experience, and outcomes of service users. There is also wide variation in the implementation of the co-response model, with differences in hours of operation, staffing, and incident response.


Behavioural contributions to the regulated intake of plant secondary metabolites in koalas.
In a given period of time, herbivores often eat less as dietary plant secondary metabolite (PSM) concentrations increase. This reduction in total food intake is interpreted as a need for the herbivore to regulate PSM ingestion in order to avoid toxication. However, regulation of PSM ingestion involves more than the reduction of total intake; it involves an alteration of meal patterns, through a reduction in the number and/or the size of the meals eaten. Despite this, studies of how herbivores alter their meal patterns when offered varying concentrations of PSMs are rare. We investigated whether koalas adjust the number and/or the size of their meals when offered eucalypt foliage varying naturally in concentrations of formylated phloroglucinol compounds (FPCs), a group of PSMs that have previously been shown to inhibit total food intake. High FPC concentrations caused koalas to eat more slowly, eat shorter meals and eat less per meal, which resulted in a reduced total intake. Because increasing FPC concentrations did not cause koalas to alter the number of meals that they ate, clear individual differences between koalas were observed, where some consistently ate fewer larger meals and others ate many smaller meals. Thus, different feeding strategies may still achieve the same outcome of a regulated intake of PSMs. The changes observed match the meal patterns of other herbivores ingesting PSMs known to stimulate nausea and emetic pathways, supporting the idea that feedback signals from nausea are an important way that koalas avoid toxication when eating eucalypt foliage.


Cultural diversity and dementia in Scottish care homes.
theoretical guidance and policy directives emphasize the necessity of considering the culturally-specific needs of health and social care service users as part of 'person-centred' approaches to care. This article outlines the ideas underpinning an ongoing PhD study of the interactions between care staff and residents which is aimed at gaining insights into how person-centred care is fulfilled for people with dementia in Scottish care homes. certain dynamics may constrain the deployment of culturally-meaningful care approaches for certain service users in Scottish care homes. These include limited understandings of the nature of cultural diversity in modern Scotland, failure to recognize the import of 'inter-generational' cultural change, lack of research around the interactions between staff and service users from different cultural backgrounds, and a sketchy demographic picture of the ethnic/cultural composition of Scottish care home populations. Groups of service users in particular settings (such as older people with dementia in long-term care) may be especially vulnerable to the effects of care processes constituted under such circumstances. poorly recognized gaps in understanding of the sociocultural and historical influences which help constitute 'the person' in many Scottish care home residents may exist. If these are found to constrain 'knowing the person', can the term 'person-centred' be meaningfully applied to care?


Cryo DualBeam Focused Ion Beam-Scanning Electron Microscopy to Evaluate the Interface Between Cells and Nanopatterned Scaffolds.
With the advance of nanotechnology in biomaterials science and tissue engineering, it is essential that new techniques become available to observe processes that take place at the direct interface between tissue and scaffold materials. Here, Cryo DualBeam focused ion beam-scanning electron microscopy (FIB-SEM) was used as a novel approach to observe the interactions between frozen hydrated cells and nanometric structures in high detail. Through a comparison of images acquired with transmission electron microscopy (TEM), conventional FIB-SEM operated at ambient temperature, and Cryo DualBeam FIB-SEM, the advantages and disadvantages of each technique were evaluated. Ultrastructural details of both (extra)cellular components and cell organelles were best observe with TEM. However, processing artifacts such as shrinkage of cells at the substrate interface were introduced in both TEM and conventional FIB-SEM. In addition, the cellular contrast in conventional FIB-SEM was low; consequently, cells were difficult to distinguish from the adjoining substrate. Cryo DualBeam FIB-SEM did preserve (extra)cellular details like the contour, cell membrane, and mineralized matrix. The three described techniques have proven to be complementary for the evaluation of processes that take place at the interface between tissue and substrate.


Partly conditional survival models for longitudinal data.
It is common in longitudinal studies to collect information on the time until a key clinical event, such as death, and to measure markers of patient health at multiple follow-up times. One approach to the joint analysis of survival and repeated measures data adopts a time-varying covariate regression model for the event time hazard. Using this standard approach, the instantaneous risk of death at time t is specified as a possibly semi-parametric function of covariate information that has accrued through time t. In this manuscript, we decouple the time scale for modeling the hazard from the time scale for accrual of available longitudinal covariate information. Specifically, we propose a class of models that condition on the covariate information through time s and then specifies the conditional hazard for times t, where t > s. Our approach parallels the "partly conditional" models proposed by Pepe and Couper (1997, Journal of the American Statistical Association 92, 991-998) for pure repeated measures applications. Estimation is based on the use of estimating equations applied to clusters of data formed through the creation of derived survival times that measure the time from measurement of covariates to the end of follow-up. Patient follow-up may be terminated either by the occurrence of the event or by censoring. The proposed methods allow a flexible characterization of the association between a longitudinal covariate process and a survival time, and facilitate the direct prediction of survival probabilities in the time-varying covariate setting.


Hypnosis decouples cognitive control from conflict monitoring processes of the frontal lobe.
Hypnosis can profoundly alter sensory awareness and cognitive processing. While the cognitive and behavioral phenomena associated with hypnosis have long been thought to relate to attentional processes, the neural mechanisms underlying susceptibility to hypnotic induction and the hypnotic condition are poorly understood. Here, we tested the proposal that highly hypnotizable individuals are particularly adept at focusing attention at baseline, but that their attentional control is compromised following hypnosis due to a decoupling between conflict monitoring and cognitive control processes of the frontal lobe. Employing event-related fMRI and EEG coherence measures, we compared conflict-related neural activity in the anterior cingulate cortex (ACC) and control-related activity in the lateral frontal cortex (LFC) during Stroop task performance between participants of low and high hypnotic susceptibility, at baseline and after hypnotic induction. The fMRI data revealed that conflict-related ACC activity interacted with hypnosis and hypnotic susceptibility, in that highly susceptible participants displayed increased conflict-related neural activity in the hypnosis condition compared to baseline, as well as with respect to subjects with low susceptibility. Cognitive-control-related LFC activity, on the other hand, did not differ between groups and conditions. These data were complemented by a decrease in functional connectivity (EEG gamma band coherence) between frontal midline and left lateral scalp sites in highly susceptible subjects after hypnosis. These results suggest that individual differences in hypnotic susceptibility are linked with the efficiency of the frontal attention system, and that the hypnotized condition is characterized by a functional dissociation of conflict monitoring and cognitive control processes.


Metabolic and structural properties of human obestatin {1-23} and two fragment peptides.
Obestatin is a peptide produced in the oxyntic mucosa of the stomach and co-localizes with ghrelin on the periphery of pancreatic islets. Several studies demonstrate that obestatin reduces food and water intake, decreases body weight gain, inhibits gastrointestinal motility, and modulates glucose-induced insulin secretion. In this study we evaluated the acute metabolic effects of human obestatin {1-23} and fragment peptides {1-10} or {11-23} in high-fat fed mice, and then investigated their solution structure by NMR spectroscopy and molecular modelling. Obestatins {1-23} and {11-23} significantly reduced food intake (86% and 90% respectively) and lowered glucose responses to feeding, whilst leaving insulin responses unchanged. No metabolic changes could be detected following the administration of obestatin {1-10}. In aqueous solution none of the obestatin peptides possessed secondary structural features. However, in a 2,2,2-trifluoroethanol (TFE-d(3))-H(2)O solvent mixture, the structure of obestatin {1-23} was characterized by an alpha-helix followed by a single turn helix conformation between residues Pro(4) and Gln(15) and His(19) and Ala(22) respectively. Obestatin {1-10} showed no structural components whereas {11-23} contained an alpha-helix between residues Val(14) and Ser(20) in a mixed solvent. These studies are the first to elucidate the structure of human obestatin and provide clear evidence that the observed alpha-helical structures are critical for in vivo activity. Future structure/function studies may facilitate the design of novel therapeutic agents based on the obestatin peptide structure.


Cannabinoids infused mouthwash products are as effective as chlorhexidine on inhibition of total-culturable bacterial content in dental plaque samples.
Dental plaque is a global health problem affecting people of various age groups. Cannabinoids are gaining enormous research attention due to its beneficial properties for various applications. A preliminary observation on antimicrobial property of cannabinoids against dental plaque bacteria has been reported recently. As a follow-up research, here we report the in vitro evaluation of cannabinoids infused mouthwash products against total culturable (aerobic) bacterial content from dental plaque samples. We tested two cannabinoid-infused mouthwash products containing cannabidiol (CBD) and cannabigerol (CBG) respectively (each mouthwash containing < 1% cannabinoid by weight) in vitro against total-culturable bacteria from dental plaque samples collected from 72 adults aged between 18 and 83 years. The participants were grouped on the basis of Dutch periodontal screening index (DPSI) score. To compare the efficacy of our products, we included two most commonly available products over the counter (Product A and Product B) to represent commercially available mouthwash products and the gold standard chlorhexidine digluconate 0.2% as a positive control. The product A represents mouthwash containing essential oils and alcohol, and Product B represents alcohol-free mouthwash that contains fluoride. All the mouthwash products were evaluated directly as such without any dilution through disc diffusion and agar well diffusion approaches and the diameter of zone of inhibition was measured. The limitation in methodology was that, the samples were open-label and the person who performed the manual measurements was unblind to test and control products used. On average, the cannabinoids infused mouthwash products showed the similar bactericidal efficacy as that of chlorhexidine 0.2%. Both chlorhexidine 0.2% and cannabinoids infused mouthwash products were effective against all the samples tested. Product A did not show any significant antimicrobial activity in any of the samples tested, except that a very marginal inhibition with a zone of 7-8 mm was observed only in 9 samples. Product B did not show any detectable inhibition zone at all in any of the samples tested. The ranges of zones of inhibition (and their average) were 8-25 mm (18.1 mm) for CBD-mouthwash, 8-25 mm (17.7 mm) for CBG-mouthwash; 12-25 mm (16.8 mm) for chlorhexidine 0.2%; 0-8 mm (0.1 mm) for Product A; and 0 mm for Product B. Although the difference in performance was slightly higher than chlorhexidine in both the cases, the difference was statistically significant for CBD-mouthwash and near significant for CBG-mouthwash. No significant difference was observed between CBD- and CBG-mouthwash. No significant difference in performance was found between DPSI score groups for any of the product tested. To our knowledge this is the first report on such efficient mouthwash product with natural key ingredients including cannabinoids and without any kind of fluoride or alcohol. Our in vitro results demonstrate the potential of cannabinoids in developing efficient and safer mouthwash products and next generation oral care products without fluoride and alcohol.


Decentralized robust adaptive control for the multiagent system consensus problem using neural networks.
A robust adaptive control approach is proposed to solve the consensus problem of multiagent systems. Compared with the previous work, the agent's dynamics includes the uncertainties and external disturbances, which is more practical in real-world applications. Due to the approximation capability of neural networks, the uncertain dynamics is compensated by the adaptive neural network scheme. The effects of the approximation error and external disturbances are counteracted by employing the robustness signal. The proposed algorithm is decentralized because the controller for each agent only utilizes the information of its neighbor agents. By the theoretical analysis, it is proved that the consensus error can be reduced as small as desired. The proposed method is then extended to two cases: Agents form a prescribed formation, and agents have the higher order dynamics. Finally, simulation examples are given to demonstrate the satisfactory performance of the proposed method.


Ten years of change to coral communities off Mona and Desecheo Islands, Puerto Rico, from disease and bleaching.
Remote reefs off southwest Puerto Rico have experienced recent losses in live coral cover of 30 to 80%, primarily due to the decline of Montastraea annularis and M. faveolata from disease and bleaching. These species were formerly the largest, oldest, and most abundant corals on these reefs, constituting over 65% of the living coral cover and 40 to 80% of the total number of colonies. From 1998 to 2001, outbreaks of yellow band disease (YBD) and white plague (WP) affected 30 to 60% of the M. annularis (complex) colonies. Disease prevalence declined beginning in 2002, and then increased immediately following the 2005 mass bleaching event. Colonies of M. annularis (complex) have been reduced in abundance by 24 to 32%, and remaining colonies are missing more than half their tissue. Both M. annularis and M. faveolata have failed to recruit, resheeting has been minimal, and exposed skeletal surfaces are being colonized by macroalgae, bioeroding sponges, and hydrozoans. Other scleractinian corals were smaller in size (mean = 28 cm diameter) and exhibited lower levels of partial mortality; these taxa were affected to a lesser extent by coral diseases and bleaching-associated tissue loss over the last decade. The numbers of small colonies (1 to 9 cm) of these species identified since 2005 also exceeded numbers of larger colonies that died. These reefs appear to be exhibiting shifts in species assemblages, with replacement of M. annularis (complex) by shorter-lived brooding species and other massive and plating corals (Agaricia, Porites, Meandrina, Eusmilia, Diploria, and Siderastrea spp.). To avoid a catastrophic and permanent loss of the dominant, slow-growing reef-building corals, the causes and effects of diseases need to be better understood, and possible control mechanisms must be developed. In particular, steps must be taken to mitigate environmental and anthropogenic stressors that increase the spread and severity of disease.


A Novel Method for Continuous Determination of the Intracellular pH in Bacteria with the Internally Conjugated Fluorescent Probe 5 (and 6-)-Carboxyfluorescein Succinimidyl Ester.
A novel method based on the intracellular conjugation of the fluorescent probe 5 (and 6-)-carboxyfluorescein succinimidyl ester (cFSE) was developed to determine the intracellular pH of bacteria. cFSE can be taken up by bacteria in the form of its diacetate ester, 5 (and 6-)-carboxyfluorescein diacetate succinimidyl ester, which is subsequently hydrolyzed by esterases to cFSE in the cytoplasm. When Lactococcus lactis cells were permeabilized with ethanol, a significant proportion of cFSE was retained in the cells, which indicated that cFSE was bound intracellularly. Unbound probe could be conveniently extruded by a short incubation of the cells in the presence of a fermentable sugar, most likely by exploiting an active transport system. Such a transport system for cFSE was identified in L. lactis, Listeria innocua, and Bacillus subtilis. The intracellular pH in bacteria can be determined from the ratio of the fluorescence signal at the pH-sensitive wavelength (490 nm) and the fluorescence signal at the pH-insensitive wavelength (440 nm). This cFSE ratio method significantly reduced problems due to the efflux of fluorescent probe from the cells during the measurement. Moreover, the method described was successfully used to determine the intracellular pH in bacteria under stress conditions, such as elevated temperatures and the presence of detergents.


Identification of a novel marine fish virus, Singapore grouper iridovirus-encoded microRNAs expressed in grouper cells by Solexa sequencing.
MicroRNAs (miRNAs) are ubiquitous non-coding RNAs that regulate gene expression at the post-transcriptional level. An increasing number of studies has revealed that viruses can also encode miRNAs, which are proposed to be involved in viral replication and persistence, cell-mediated antiviral immune response, angiogenesis, and cell cycle regulation. Singapore grouper iridovirus (SGIV) is a pathogenic iridovirus that has severely affected grouper aquaculture in China and Southeast Asia. Comprehensive knowledge about the related miRNAs during SGIV infection is helpful for understanding the infection and the pathogenic mechanisms. To determine whether SGIV encoded miRNAs during infection, a small RNA library derived from SGIV-infected grouper (GP) cells was constructed and sequenced by Illumina/Solexa deep-sequencing technology. We recovered 6,802,977 usable reads, of which 34,400 represented small RNA sequences encoded by SGIV. Sixteen novel SGIV-encoded miRNAs were identified by a computational pipeline, including a miRNA that shared a similar sequence to herpesvirus miRNA HSV2-miR-H4-5p, which suggests miRNAs are conserved in far related viruses. Generally, these 16 miRNAs are dispersed throughout the SGIV genome, whereas three are located within the ORF057L region. Some SGIV-encoded miRNAs showed marked sequence and length heterogeneity at their 3' and/or 5' end that could modulate their functions. Expression levels and potential biological activities of these viral miRNAs were examined by stem-loop quantitative RT-PCR and luciferase reporter assay, respectively, and 11 of these viral miRNAs were present and functional in SGIV-infected GP cells. Our study provided a genome-wide view of miRNA production for iridoviruses and identified 16 novel viral miRNAs. To the best of our knowledge, this is the first experimental demonstration of miRNAs encoded by aquatic animal viruses. The results provide a useful resource for further in-depth studies on SGIV infection and iridovirus pathogenesis.


Synthesis of Stable Nanographenes with OBO-Doped Zigzag Edges Based on Tandem Demethylation-Electrophilic Borylation.
A tandem demethylation-aryl borylation strategy was developed to synthesize OBO-doped tetrabenzo[a,f,j,o]perylenes (namely "bistetracenes") and tetrabenzo[bc,ef,kl,no]coronenes (namely "peritetracenes"). The OBO-doped bistetracene analogues exhibited excellent stability and strong fluorescence, in contrast to the unstable all-carbon bistetracene. Single-crystal X-ray analysis for OBO-doped bistetracene revealed a twisted double [5]helicene structure, indicating that this synthesis is applicable to new heterohelicenes. Importantly, cyclodehydrogenation of the bistetracene analogues successfully produced the unprecedented heteroatom-doped peritetracenes, which opened up a new avenue to periacene-type nanographenes with stable zigzag edges.


Omcg1 is critically required for mitosis in rapidly dividing mouse intestinal progenitors and embryonic stem cells.
Recent studies have shown that factors involved in transcription-coupled mRNA processing are important for the maintenance of genome integrity. How these processes are linked and regulated in vivo remains largely unknown. In this study, we addressed in the mouse model the function of Omcg1, which has been shown to participate in co-transcriptional processes, including splicing and transcription-coupled repair. Using inducible mouse models, we found that Omcg1 is most critically required in intestinal progenitors. In absence of OMCG1, proliferating intestinal epithelial cells underwent abnormal mitosis followed by apoptotic cell death. As a consequence, the crypt proliferative compartment of the small intestine was quickly and totally abrogated leading to the rapid death of the mice. Lack of OMCG1 in embryonic stem cells led to a similar cellular phenotype, with multiple mitotic defects and rapid cell death. We showed that mutant intestinal progenitors and embryonic stem cells exhibited a reduced cell cycle arrest following irradiation, suggesting that mitotic defects may be consecutive to M phase entry with unrepaired DNA damages. These findings unravel a crucial role for pre-mRNA processing in the homeostasis of the small intestine and point to a major role of OMCG1 in the maintenance of genome integrity.


Synthesis of some novel organic nitrates and comparative in vitro study of their vasodilator profile.
Synthesis and structural characterization of the 4-phenylbutane-1,2-diyl dinitrate and of the erythro and threo diastereoisomers of 4-phenylbutane-1,2,3-triyl trinitrate as well as the HPLC chiral separation of the corresponding racemic mixtures are reported. Vasodilator activity of the single enantiomers of these products, of 4-phenylbutyl nitrate, and of the previously described phenylpropyl analogues were assessed on rat aorta strips precontracted with phenylephrine. The compounds were able to relax the contracted tissue in a concentration dependent manner. In the couples of antipodes, a complete lack of enantioselectivity was observed as far as the vasodilator potency is concerned. The concentration response curves of the products, with the exception of those of all the trinitrooxy substituted models, were rightward shifted in the presence of ALDH-2 inhibitors. Mono and dinitrates, but not trinitrates, displayed in vitro cross-tolerance with GTN. This new series of nitric acid esters is an interesting tool that can help to shed light on the unresolved puzzle of nitrate pharmacology. Selected members are worthy of additional study as potential drugs.


Outcome comparison of two surgical methods used for the treatment of acute thrombosis of arteriovenous fistulas: thrombectomy versus de novo creation of arteriovenous fistula.
The aim of this study was to evaluate efficacy of two surgical methods used for the treatment of acute arteriovenous fistula (AVF) thrombosis. Twenty two out of twenty five patients that were admitted at the Clinic for vascular surgery in Sarajevo received successful surgical treatment for the salvage of acutely thromboses AVF from 2007-2009. They were included in retrospective, descriptive clinical study. Based on the type of surgical procedures performed, 22 patients were divided into two groups. The first group included 10 patients and they had successful thrombectomy of thromboses AVF while 12 patients in second group underwent de novo creation of AVF using blood vessels already exploited for construction of thromboses AVF. Patency rate of salvaged AVF in analyzed groups was compared one month and 6 months after intervention. In the postoperative follow up there was no statistically significant difference in patency rate of salvaged AVF between analyzed groups after one month, (80% vs 100%, Fisher exact test value =2,520, p= 0.195). Patency rate of salvaged AVF after six months of the follow up was significantly better in group that received de novo construction of AVF when compared to thrombectomy group (25% vs. 91%, Fisher exact test value = 1,062, p=0.002). De novo construction of AVF in case of acutely thrombosed AVF offered better patency rate of salvaged AVF when compared to surgical thrombectomy in the follow up period of six months.