β-Amyrin synthase from Euphorbia tirucalli. Steric bulk, not the π-electrons of Phe, at position 474 has a key role in affording the correct folding of the substrate to complete the normal polycyclization cascade.
β-Amyrin, a triterpene, is widely distributed in plants and its glycosides confer important biological activities. Mutagenesis studies on β-amyrin synthase are very limited as compared with those of squalene-hopene cyclase and lanosterol synthase. This study was conducted to elucidate the function of the F474 residue of Euphorbia tirucalli β-amyrin cyclase, which is highly conserved in the superfamily of oxidosqualene cyclases. Nine site-specific variants with Gly, Ala, Val, Leu, Met, Tyr, Trp, His, and Thr were constructed. We isolated 9 products from these mutants in addition to β-amyrin and determined the chemical structures. The Gly and Ala mutants produced significantly larger amounts of the bicyclic products and a decreased amount of β-amyrin, indicating that the F474 residue was located near the B-ring formation site. Surprisingly, the Ala variant produced (9βH)-polypoda-7,13,17,21-tetraen-3β-ol and (9βH)-polypoda-8(26),13,17,21-tetraen-3β-ol, which are generated from a chair-boat folding conformation. This is the first report describing the conformational change from the chair-chair into the chair-boat folding conformation among the reported mutagenesis studies of oxidosqualene cyclases. Substitution with aliphatic amino acids lacking π-electrons such as Val, Leu, and Met led to a significantly decreased production of bicyclic compounds, and in turn exhibited a higher production of β-amyrin. Furthermore, the Leu and Met variants exhibited high enzymatic activities: ca. 74% for Leu and ca. 91% for Met variants as compared to the wild-type. These facts unambiguously demonstrate that the major role of Phe474 is not to stabilize the transient cation via cation-π interaction, but is to confer the appropriate steric bulk near the B-ring formation site, leading to the completion of the normal polycyclization pathway without accumulation of abortive cyclization products.


Gene expression patterns in heterozygous Plk4 murine embryonic fibroblasts.
The polo-like kinases (Plks) are a group of serine/threonine kinases which have roles in many aspects of cellular function including the regulation of mitotic activity and cellular stress responses. This study focuses on Plk4, the most divergent member of the Plk family, which is necessary for proper cellular proliferation. More specifically, alterations in Plk4 levels cause significantly adverse mitotic defects including abnormal centrosome duplication and aberrant mitotic spindle formation. We sought to clarify the effect of reduced Plk4 levels on the cell by examining transcript profiles of Plk4 wild-type and heterozygous mouse embryonic fibroblasts (MEFs). Subsequently, the levels of several key proteins involved in the DNA damage response were examined. 143 genes were found to be significantly up-regulated in the heterozygous MEFs compared to their wild-type counterparts, while conversely, 9 genes were down-regulated. Numerous genes with increased transcript levels in heterozygous MEFs were identified to be involved in p53-dependent pathways. Furthermore, examination of the promoter regions of all up- and down-regulated genes revealed that the majority contained putative p53 responsive elements. An analysis of transcript levels in MEFs after exposure to either ionizing or ultraviolet radiation revealed a significant change between wild type and heterozygous MEFS for Plk4 transcript levels upon only UV exposure. Furthermore, changes in protein levels of several important cell check-point and apoptosis regulators were examined, including p53, Chk1, Chk2, Cdc25C and p21. In heterozygous MEFs, p53, p21 and Chk2 protein levels were at significantly higher levels. Furthermore, p53 activity was increased 5 fold in the Plk4 heterozygous MEFs. Global transcript profiles and levels of key proteins involved in cellular proliferation and DNA damage pathways were examined in wild-type and Plk4 heterozygous MEFs. It was determined that Plk4 haploinsufficiency leads to changes in the levels of RNA accumulation for a number of key cellular genes as well as changes in protein levels for several important cell cycle/DNA damage proteins. We propose a model in which reduced Plk4 levels invoke an increase in p53 levels that leads to the aforementioned changes in global transcription profiles.


Burnout and satisfaction with work-life balance among US physicians relative to the general US population.
Despite extensive data about physician burnout, to our knowledge, no national study has evaluated rates of burnout among US physicians, explored differences by specialty, or compared physicians with US workers in other fields. We conducted a national study of burnout in a large sample of US physicians from all specialty disciplines using the American Medical Association Physician Masterfile and surveyed a probability-based sample of the general US population for comparison. Burnout was measured using validated instruments. Satisfaction with work-life balance was explored. Of 27 276 physicians who received an invitation to participate, 7288 (26.7%) completed surveys. When assessed using the Maslach Burnout Inventory, 45.8% of physicians reported at least 1 symptom of burnout. Substantial differences in burnout were observed by specialty, with the highest rates among physicians at the front line of care access (family medicine, general internal medicine, and emergency medicine). Compared with a probability-based sample of 3442 working US adults, physicians were more likely to have symptoms of burnout (37.9% vs 27.8%) and to be dissatisfied with work-life balance (40.2% vs 23.2%) (P < .001 for both). Highest level of education completed also related to burnout in a pooled multivariate analysis adjusted for age, sex, relationship status, and hours worked per week. Compared with high school graduates, individuals with an MD or DO degree were at increased risk for burnout (odds ratio [OR], 1.36; P < .001), whereas individuals with a bachelor's degree (OR, 0.80; P = .048), master's degree (OR, 0.71; P = .01), or professional or doctoral degree other than an MD or DO degree (OR, 0.64; P = .04) were at lower risk for burnout. Burnout is more common among physicians than among other US workers. Physicians in specialties at the front line of care access seem to be at greatest risk.


Systematic analyses of the cytotoxic effects of compound 11a, a putative synthetic agonist of photoreceptor-specific nuclear receptor (PNR), in cancer cell lines.
Photoreceptor cell-specific receptor (PNR/NR2E3) is an orphan nuclear receptor that plays a critical role in retinal development and photoreceptor maintenance. The disease-causing mutations in PNR have a pleiotropic effect resulting in varying retinal diseases. Recently, PNR has been implicated in control of cellular functions in cancer cells. PNR was reported to be a novel regulator of ERα expression in breast cancer cells, and high PNR expression correlates with favorable response to tamoxifen treatment. Moreover, PNR was shown to increase p53 stability in HeLa cells, implying that PNR may be a therapeutic target in this and other cancers that retain a wild type p53 gene. To facilitate further understanding of PNR functions in cancer, we characterized compound 11a, a synthetic, putative PNR agonist in several cell-based assays. Interestingly, we showed that 11a failed to activate PNR and its cytotoxicity was independent of PNR expression, excluding PNR as a mediator for 11a cytotoxicity. Systematic analyses of the cytotoxic effects of 11a in NCI-60 cell lines revealed a strong positive correlation of cytotoxicity with p53 status, i.e., p53 wild type cell lines were significantly more sensitive to 11a than p53 mutated or null cell lines. Furthermore, using HCT116 p53+/+ and p53-/- isogenic cell lines we revealed that the mechanism of 11a-induced cytotoxicity occurred through G1/S phase cell cycle arrest rather than apoptosis. In conclusion, we observed a correlation of 11a sensitivity with p53 status but not with PNR expression, suggesting that tumors expressing wild type p53 might be responsive to this compound.


Calcium/calmodulin transduces thrombin-stimulated secretion: studies in intact and minimally permeabilized human umbilical vein endothelial cells.
Thrombin stimulates cultured endothelial cells (EC) to secrete stored von Willebrand factor (vWF), but the signal transduction pathways are poorly defined. Thrombin is known to elevate the concentration of intracellular calcium ([Ca2+]i) and to activate protein kinase C (PKC) in EC. Since both calcium ionophores and phorbol esters release vWF, both second messenger pathways have been postulated to participate in vWF secretion in response to naturally occurring agonists. We find that in intact human EC, vWF secretion stimulated by either thrombin or by a thrombin receptor activating peptide, TR(42-55), can be correlated with agonist-induced elevations of [Ca2+]i. Further evidence implicating calcium in the signal transduction pathway is suggested by the finding that MAPTAM, a cell-permeant calcium chelator, in combination with the extracellular calcium chelator EGTA, can inhibit thrombin-stimulated secretion. In contrast, the observation that staurosporine (a pharmacological inhibitor of PKC) blocks phorbol ester- but not thrombin-stimulated secretion provides evidence against PKC-mediated signal transduction. To examine further the signal transduction pathway initiated by thrombin, we developed novel conditions for minimal permeabilization of EC with saponin (4-8 micrograms/ml for 5-15 min at 37 degrees C) which allow the introduction of small extracellular molecules without the loss of large intracellular proteins and which retain thrombin-stimulated secretion. These minimally permeabilized cells secrete vWF in response to exogenous calcium, and EGTA blocks thrombin-induced secretion. Moreover, in these cells, thrombin-stimulated secretion is blocked by a calmodulin-binding inhibitory peptide but not by a PKC inhibitory peptide. Taken together, these findings demonstrate that thrombin-stimulated vWF secretion is transduced by a rise in [Ca2+]i and provide the first evidence for the role of calmodulin in this process.


Concurrent activity of anammox and denitrifying bacteria in the Black Sea.
After the discovery of ANaerobic AMMonium OXidation (anammox) in the environment, the role of heterotrophic denitrification as the main marine pathway for fixed N loss has been questioned. A 3 part, 15 month time series investigating nitrite reductase (nirS) mRNA transcripts at a single location in the Black Sea was conducted in order to better understand the activity of anammox and denitrifying bacteria. Here we show that both of these groups were active, as well as being concurrent in the lower suboxic zone over this time span. Their distributions, however, differed in that only expression of denitrification-type nirS was seen in the upper suboxic zone, where geochemistry was variable. Depth profiles covering the suboxic zone showed that the four groups of anammox-type sequences were expressed consistently in the lower suboxic zone, and were consistent with anammox 16 S rDNA gene profiles. By contrast, denitrifier-type nirS sequence groups were mixed; some groups exhibited consistent expression in the lower suboxic zone, while others appeared less consistent. Co-occurrence of both anammox and denitrifier expression was common and ongoing. Both types of transcripts were also found in samples with low concentrations of sulfide (>2 μM). Six major groups of denitrifier-type nirS transcripts were identified, and several groups of denitrifier-type nirS transcripts were closely related to sequences from the Baltic Sea. An increase in denitrifier-type nirS transcript diversity and depth range in October 2007 corresponded to a small increase in mixed layer net community productivity (NCP) as measured by O(2)/Ar gas ratios, as well as to an increase in N(2) concentrations in the suboxic zone. Taken together, the variations in expression patterns between anammox and denitrification provide one possible explanation as to how near instantaneous rate measurements, such as isotope spike experiments, may regularly detect anammox activity but underreport denitrification.


Diffusive growth of fullerenes and carbon nanotubes.
The video of a growing fullerene within a carbon nanotube, initiated by a tungsten catalyst, provides a dramatic realization of a complex nanoscale process. While there may be many detailed models which can account for this growth, we propose one of the simplest possible models which is consistent with the major observed features of the growth process. In particular, we assume that the fullerene is immersed in a carbon vapor environment, and that the growth occurs as a consequence of the diffusion of the carbon vapor into the fullerene. Moreover, we assume that the classical diffusion equation applies in the region exterior to the fullerene and that a standard Stefan condition applies at the moving fullerene surface. We assume that the gaseous medium through which the carbon atoms diffuse is represented through the value of the diffusion coefficient D appearing in the classical diffusion equation. We also assume that the influence of the catalyst is felt through the value of the constant alpha appearing in the Stefan condition. Based on these assumptions, we derive simple similarity solutions for both spherical and ellipsoidal fullerenes which are entirely consistent with the observations. A corresponding analysis is provided for the longitudinal growth of a carbon nanotube.


Nonylphenol induces pancreatic damage in rats through mitochondrial dysfunction and oxidative stress.
The organic alkylphenol 4-nonylphenol (NP) is regarded to be an endocrine disrupting chemical (EDC), one of the widely diffused and stable environmental contaminants. Due to its hydrophobicity and long half-life, NP can easily accumulate in living organisms, including humans, where it displays a series of toxic effects. It has been widely reported that NP affects male reproduction. In addition, there is increasing evidence suggesting that NP is detrimental to various organs, including the pancreas. This study investigated the adverse effects of NP exposure on the pancreas. Sprague-Dawley rats were treated with different doses of NP for 90 consecutive days. The data suggested that the body weights of the rats treated with NP decreased, and the highest dose of NP treatment (180 mg kg-1) dramatically increased water consumption by rats. Meanwhile, H&E staining and immunohistochemistry indicated that islets in the pancreases shrunk when the rats were treated with the indicated doses of NP. TUNEL staining demonstrated that NP exposure up-regulated the level of apoptosis in the pancreases in a dose-dependent manner. Besides this, NP exposure inhibited the secretion of insulin and disrupted glucose tolerance. The levels of reactive oxygen species (ROS) and intracellular calcium ([Ca2+]i) in the islets were up-regulated in the groups of rats treated with NP, but the levels of Mitochondrial Membrane Potential (MMP) were down-regulated. These results suggest that NP-induced pancreatic damage in rats occurs through mitochondrial dysfunction and oxidative stress, which causes disruption of glucose tolerance and decrease in insulin secretion.


Aminoacyl-tRNA formation in the extreme thermophile Thermus thermophilus.
Thermophilic organisms must be capable of accurate translation at temperatures in which the individual components of the translation machinery and also specific amino acids are particularly sensitive. Thermus thermophilus is a good model organism for studies of thermophilic translation because many of the components in this process have undergone structural and biochemical characterization. We have focused on the pathways of aminoacyl-tRNA synthesis for glutamine, asparagine, proline, and cysteine. We show that the T. thermophilus prolyl-tRNA synthetase (ProRS) exhibits cysteinyl-tRNA synthetase (CysRS) activity although the organism also encodes a canonical CysRS. The ProRS requires tRNA for cysteine activation, as is known for the characterized archaeal prolyl-cysteinyl-tRNA synthetase (ProCysRS) enzymes. The heterotrimeric T. thermophilus aspartyl-tRNA(Asn) amidotransferase can form Gln-tRNA in addition to Asn-tRNA: however, a 13-amino-acid C-terminal truncation of the holoenzyme A subunit is deficient in both activities when assayed with homologous substrates. A survey of codon usage in completed prokaryotic genomes identified a higher Glu:Gln ratio in proteins of thermophiles compared to mesophiles.


Transovarial transmission efficiency of Babesia bovis tick stages acquired by Rhipicephalus (Boophilus) microplus during acute infection.
The protozoan parasite Babesia bovis, a reemerging threat to U.S. cattle, is acquired by adult female ticks of the subgenus Boophilus and is transovarially transmitted as the kinete stage to developing larval offspring. Sporozoites develop within larvae and are transmitted during larval feeding on a bovine host. This study evaluated the efficiency of B. bovis infection within Rhipicephalus (Boophilus) microplus following acquisition feeding on acutely parasitemic cattle. Parasite levels were quantified in blood from experimentally infected cattle and within hemolymph and larvae derived from acquisition-fed female B. microplus. There was a positive correlation between blood parasite levels in acutely parasitemic cattle and kinete levels in the hemolymph of adult female Boophilus ticks following acquisition feeding; however, there was no relationship between kinete levels in females and infection rates of larval progeny. Boophilus microplus females that acquisition fed produced larval progeny with infection rates of 12% to 48%. Importantly, larvae derived from replete females with very low levels of kinete infection, as demonstrated by microscopy and PCR, had infection rates of 22% to 30% and transmitted B. bovis during transmission feeding. These data demonstrate that although hemolymph infection may be undetectable, transmission to larval progeny occurs at a level which ensures transmission to the bovine host.


Role of the Paraventricular Nucleus of the Hypothalamus in the Sympathoexcitatory Effects of Leptin.
Leptin binds to receptors in multiple hypothalamic nuclei to increase sympathetic nerve activity; however, the neurocircuitry is unclear. Here, using anesthetized male Sprague-Dawley rats, we investigated the role of the paraventricular nucleus of the hypothalamus. Intracerebroventricular injection of leptin slowly increased lumbar sympathetic nerve activity (LSNA), heart rate, mean arterial pressure, and baroreflex control of LSNA and heart rate. Inhibition of the paraventricular nucleus with muscimol completely reversed leptin's effects. Blockade of paraventricular melanocortin 3/4 receptors with SHU9119 or ionotropic glutamate receptors with kynurenate, alone or together, each partially reversed the effects of leptin, implicating increased activation of glutamate and melanocortin 3/4 receptors. Conversely, although blockade of neuropeptide Y Y1 receptors in the paraventricular nucleus increased LSNA, mean arterial pressure, and heart rate, these responses were prevented by intracerebroventricular or arcuate nucleus injections of leptin, suggesting that, at least in part, leptin also increases sympathetic nerve activity by suppression of tonic neuropeptide Y inhibitory inputs from the arcuate nucleus. Injection of the melanocortin 3/4 receptor agonist melanotan-II into the paraventricular nucleus increased LSNA, mean arterial pressure, and heart rate only after blockade of neuropeptide Y Y1 receptors. Therefore, we conclude that leptin increases LSNA in part via increased glutamatergic and α-melanocyte-stimulating hormone drive of paraventricular sympathoexcitatory neurons, the latter of which requires simultaneous withdrawal of tonic neuropeptide Y inhibition.


Mathematical modeling of tuberculosis bacillary counts and cellular populations in the organs of infected mice.
Mycobacterium tuberculosis is a particularly aggressive microorganism and the host's defense is based on the induction of cellular immunity, in which the creation of a granulomatous structure has an important role. We present here a new 2D cellular automata model based on the concept of a multifunctional process that includes key factors such as the chemokine attraction of the cells; the role of innate immunity triggered by natural killers; the presence of neutrophils; apoptosis and necrosis of infected macrophages; the removal of dead cells by macrophages, which induces the production of foamy macrophages (FMs); the life cycle of the bacilli as a determinant for the evolution of infected macrophages; and the immune response. The results obtained after the inclusion of two degrees of tolerance to the inflammatory response triggered by the infection shows that the model can cover a wide spectrum, ranging from highly-tolerant (i.e. mice) to poorly-tolerant hosts (i.e. mini-pigs or humans). This model suggest that stopping bacillary growth at the onset of the infection might be difficult and the important role played by FMs in bacillary drainage in poorly-tolerant hosts together with apoptosis and innate lymphocytes. It also shows the poor ability of the cellular immunity to control the infection, provides a clear protective character to the granuloma, due its ability to attract a sufficient number of cells, and explains why an already infected host can be constantly reinfected.


Scaling up integrated management of childhood illness to the national level: achievements and challenges in Peru.
This paper presents the first published report of a national-level effort to implement the Integrated Management of Childhood Illness (IMCI) strategy at scale. IMCI was introduced in Peru in late 1996, the early implementation phase started in 1997, with the expansion phase starting in 1998. Here we report on a retrospective evaluation designed to describe and analyze the process of taking IMCI to scale in Peru, conducted as one of five studies within the Multi-Country Evaluation of IMCI Effectiveness, Cost and Impact (MCE) coordinated by the World Health Organization. Trained surveyors visited each of Peru's 34 districts, interviewed district health staff and reviewed district records. Findings show that IMCI was not institutionalized in Peru: it was implemented parallel to existing programmes to address acute respiratory infections and diarrhoea, sharing budget lines and management staff. The number of health workers trained in IMCI case management increased until 1999 and then decreased in 2000 and 2001, with overall coverage levels among doctors and nurses calculated to be 10.3%. Efforts to implement the community component of IMCI began with the training of community health workers in 2000, but expected synergies between health facility and community interventions were not realized because districts where clinical training was most intense were not those where community IMCI training was strongest. We summarize the constraints to scaling up IMCI, and examine both the methodological and policy implications of the findings. Few monitoring data were available to document IMCI implementation in Peru, limiting the potential of retrospective evaluations to contribute to programme improvement. Even basic indicators recommended for national monitoring could not be calculated at either district or national levels. The findings document weaknesses in the policy and programme supports for IMCI that would cripple any intervention delivered through the health service delivery system. The Ministry of Health in Peru is now working to address these weaknesses; other countries working to achieve high and equitable coverage with essential child survival interventions can learn from their experience.


Extraction and analysis of multiple periodic motions in video sequences.
The analysis of periodic or repetitive motions is useful in many applications, such as the recognition and classification of human and animal activities. Existing methods for the analysis of periodic motions first extract motion trajectories using spatial information and then determine if they are periodic. These approaches are mostly based on feature matching or spatial correlation, which are often infeasible, unreliable, or computationally demanding. In this paper, we present a new approach, based on the time-frequency analysis of the video sequence as a whole. Multiple periodic trajectories are extracted and their periods are estimated simultaneously. The objects that are moving in a periodic manner are extracted using the spatial domain information. Experiments with synthetic and real sequences display the capabilities of this approach.


Nationwide school malaria parasitaemia survey in public primary schools, the United Republic of Tanzania.
A nationwide, school, malaria survey was implemented to assess the risk factors of malaria prevalence and bed net use among primary school children in mainland Tanzania. This allowed the mapping of malaria prevalence at council level and assessment of malaria risk factors among school children. A cross-sectional, school, malaria parasitaemia survey was conducted in 25 regions, 166 councils and 357 schools in three phases: (1) August to September 2014; (2) May 2015; and, (3) October 2015. Children were tested for malaria parasites using rapid diagnostic tests and were interviewed about household information, parents' education, bed net indicators as well as recent history of fever. Multilevel mixed effects logistic regression models were fitted to estimate odds ratios of risk factors for malaria infection and for bed net use while adjusting for school effect. In total, 49,113 children were interviewed and tested for malaria infection. The overall prevalence of malaria was 21.6%, ranging from < 0.1 to 53% among regions and from 0 to 76.4% among councils. The malaria prevalence was below 5% in 62 of the 166 councils and above 50% in 18 councils and between 5 and 50% in the other councils. The variation of malaria prevalence between schools was greatest in regions with a high mean prevalence, while the variation was marked by a few outlying schools in regions with a low mean prevalence. Overall, 70% of the children reported using mosquito nets, with the highest percentage observed among educated parents (80.7%), low land areas (82.7%) and those living in urban areas (82.2%). The observed prevalence among school children showed marked variation at regional and sub-regional levels across the country. Findings of this survey are useful for updating the malaria epidemiological profile and for stratification of malaria transmission by region, council and age groups, which is essential for guiding resource allocation, evaluation and prioritization of malaria interventions.


Double tracer gas single-breath washout: reproducibility in healthy subjects and COPD.
The applicability and interpretation of inert tracer gas washout tests is hampered by the lack of feasible protocols and reproducibility data. We assessed feasibility, variability and reproducibility of a new easy to perform double tracer gas (DTG) single-breath washout (SBW) test and compared this with conventional nitrogen washouts. In 40 healthy nonsmokers and 20 patients with stable chronic obstructive pulmonary disease (COPD), we performed three N2 vital capacity SBWs, three N2 multiple-breath washouts and three tidal DTG-SBW tests. Follow-up was after 1 week, 1 month and 6 months. Main outcomes were the lung clearance index (LCI) (N2 multiple-breath washout), slope of phase III (dN2) (N2 vital capacity SBW) and slope of phase III (SIIIDTG) (DTG-SBW). In healthy subjects, mean±sd LCI at baseline was 6.94±0.61, dN2 0.99±0.42% N2 per litre and SIIIDTG -0.206±0.108 g·mol(-1)·L(-1). In COPD, LCI and dN2 were significantly higher (LCI 12.23±2.67, dN2 7.43±5.38% N2 per litre; p<0.001) and SIIIDTG significantly steeper (-0.653±0.428 g·mol(-1)·L(-1), p<0.001). Reproducibility was high for main outcome parameters: the intraclass correlation coefficient over 6 months was 0.77 (0.86 in COPD) for LCI, 0.82 (0.89) for dN2 and 0.83 (0.93) for SIIIDTG. The tidal DTG-SBW is a reproducible test in healthy and COPD subjects that seems attractive for use in routine clinical settings.


Synergistic Potentiation of Cystic Fibrosis Transmembrane Conductance Regulator Gating by Two Chemically Distinct Potentiators, Ivacaftor (VX-770) and 5-Nitro-2-(3-Phenylpropylamino) Benzoate.
Cystic fibrosis (CF) is caused by loss-of-function mutations of the cystic fibrosis transmembrane conductance regulator (CFTR) gene encoding a phosphorylation-activated but ATP-gated chloride channel. Previous studies suggested that VX-770 [ivacaftor, N-(2,4-di-tert-butyl-5-hydroxyphenyl)-4-oxo-1,4-dihydroquinoline-3-carboxamide], a CFTR potentiator now used in clinics, increases the open probability of CFTR by shifting the gating conformational changes to favor the open channel configuration. Recently the chloride channel blocker and CFTR potentiator 5-nitro-2-(3-phenylpropylamino) benzoate (NPPB) has been reported to enhance CFTR activity by a mechanism that exploits the ATP hydrolysis-driven, nonequilibrium gating mechanism unique to CFTR. Surprisingly however, NPPB increased the activity of nonhydrolytic G551D-CFTR, the third most common disease-associated mutation. Here, we further investigated the mechanism of NPPB's effects on CFTR gating by assessing its interaction with well-studied VX-770. Interestingly, once G551D-CFTR was maximally potentiated by VX-770, NPPB further increased its activity. However, quantitative analysis of this drug-drug interaction suggests that this pharmacologic synergism is not due to independent actions of NPPB and VX-770 on CFTR gating; instead, our data support a dependent mechanism involving two distinct binding sites. This latter idea is further supported by the observation that the locked-open time of a hydrolysis-deficient mutant K1250A was shortened by NPPB but prolonged by VX-770. In addition, the effectiveness of NPPB, but not of VX-770, was greatly diminished in a mutant whose second nucleotide-binding domain was completely removed. Interpreting these results under the framework of current understanding of CFTR gating not only reveals insights into the mechanism of action for different CFTR potentiators but also brings us one step forward to a more complete schematic for CFTR gating.


Rapid functional assays of recombinant IP3 receptors.
Functional assays of inositol 1,4,5-trisphosphate receptors (IP3R) currently use 45Ca2+ release methods, fluorescent Ca2+ indicators within either the ER or cytosol, or electrophysiological analyses of IP3R in the nuclear envelope or artificial bilayers. None of the methods is presently amenable to the rapid, high-throughput quantitative analyses of IP3R function needed to address the structural determinants of IP3R behavior. We use a low-affinity Ca2+ indicator (Mag-fluo-4) to measure free [Ca2+] within the ER of permeabilized DT40 cells expressing only rat type 1 IP(3)R, and establish that the indicator is capable of reliably reporting the Ca(2+) release evoked by IP3. A 96-well fluorescence plate reader equipped for automated fluid additions (FlexStation, Molecular Devices) is used to monitor IP3-evoked Ca2+ release. The method allows quick and economical functional assays of recombinant IP3R in small volumes (< or = 100 microl).


Potentially pathogenic autoantibodies associated with epilepsy and encephalitis in children and adults.
Autoantibodies to surface proteins that influence neuronal excitability are increasingly found in different forms of epilepsy or encephalitis in adults, and are also beginning to be identified in children. The conditions are often refractory to traditional antiepileptic drugs. Detection of these antibodies can help to identify forms of epilepsy that may respond to immunotherapies.


Virulence Dynamics and Regional Structuring of Puccinia striiformis f. sp. tritici in France Between 1984 and 2009.
Understanding of long-term virulence dynamics of pathogen populations in response to host resistance gene deployment is of major importance for disease management and evolutionary biology. We monitored the virulence dynamics of Puccinia striiformis f. sp. tritici, the causal agent of wheat stripe rust, over 25 years in France. Virulence dynamics was explained by estimates of area associated with resistance genes carried by farmers' cultivars. The epidemics assessed through disease severity significantly correlated with the number of P. striiformis f. sp. tritici isolates collected each year, used to describe virulence dynamics. In the south, the dominance of the Mediterranean pathotype 6E16 and the cultivation of a susceptible cultivar were associated with an epidemic from 1997 to 1999. In the north, five epidemics occurred due to successive acquisition of virulence to the resistance genes Yr7, Yr6, Yr9, Yr17, and Yr32, either by acquisition of the virulence in the previous dominant pathotype or by incursion or selection of one or two new pathotypes. Frequency of pathotypes with Vr7 and Vr6 declined with the reduction in the cultivation of corresponding Yr gene cultivars, whereas the virulence Vr9 persisted longer than the cultivation of Yr9 cultivars. Although the first pathotypes carrying Vr9 decreased, this virulence persisted in other pathotypes even in the absence of Yr9 cultivars. At the regional level, Yr9 cultivars in the north caused a shift from high Vr6 frequency to high Vr9 frequency whereas, in the central region, where Yr9 cultivars were rare, Vr6 remained prevalent.


An assessment of wheat yield sensitivity and breeding gains in hot environments.
Genetic improvements in heat tolerance of wheat provide a potential adaptation response to long-term warming trends, and may also boost yields in wheat-growing areas already subject to heat stress. Yet there have been few assessments of recent progress in breeding wheat for hot environments. Here, data from 25 years of wheat trials in 76 countries from the International Maize and Wheat Improvement Center (CIMMYT) are used to empirically model the response of wheat to environmental variation and assess the genetic gains over time in different environments and for different breeding strategies. Wheat yields exhibited the most sensitivity to warming during the grain-filling stage, typically the hottest part of the season. Sites with high vapour pressure deficit (VPD) exhibited a less negative response to temperatures during this period, probably associated with increased transpirational cooling. Genetic improvements were assessed by using the empirical model to correct observed yield growth for changes in environmental conditions and management over time. These 'climate-corrected' yield trends showed that most of the genetic gains in the high-yield-potential Elite Spring Wheat Yield Trial (ESWYT) were made at cooler temperatures, close to the physiological optimum, with no evidence for genetic gains at the hottest temperatures. In contrast, the Semi-Arid Wheat Yield Trial (SAWYT), a lower-yielding nursery targeted at maintaining yields under stressed conditions, showed the strongest genetic gains at the hottest temperatures. These results imply that targeted breeding efforts help us to ensure progress in building heat tolerance, and that intensified (and possibly new) approaches are needed to improve the yield potential of wheat in hot environments in order to maintain global food security in a warmer climate.


Exploring family and social context through the electronic health record: Physicians' experiences.
There is ample evidence that social and familial context significantly impacts health. However, family and social history templates typically used in clinical practice exclude prompts to explore important contextual information, such as family dynamics, health beliefs, housing, and neighborhood environment. At the Residency Program in Social Medicine at Montefiore Medical Center/Albert Einstein College of Medicine in Bronx, NY, we developed and piloted an expanded family and social information (FSI) template in our electronic health record (EHR) system. After 10 physicians used the FSI template during routine clinical practice, we conducted and qualitatively analyzed transcripts of semistructured interviews to assess their experiences. The major themes of physician's experiences using the FSI template included: expanded thinking (promoted thinking and discussion about contextual information), relevancy to care (highlighted important life events, helpful resource for future care or for team-based care), fragmentation (patient narrative difficult to categorize into discrete sections), and abstract phrasing (prompts too abstract and/or too complex to explore well). Some minor themes (themes that were repeated often, but not consistent in all interviews) were also noted: practice demands, educational purpose, and wish list. Within our small sample, we found that an expanded family and social information template broadened physicians' clinical thinking during routine visits. We also found that the structure of the EHR and practice demands create significant barriers to exploring a patient's narrative effectively. Themes identified in this research will inform national efforts to prioritize a contextual and biopsychosocial perspective in EHR systems. (PsycINFO Database Record


"Click for Closer Care": A Content Analysis of Community Pharmacy Websites in Four Countries.
Combinations of professional and commercial communication are typically very controversial, particularly in health care communication on the Internet. Websites of licensed community pharmacies on the other hand tend to raise remarkably little controversy, although they typically contain controversial combinations of clinical and commercial services previously unprecedented in professional health care communication. The aim of this study was to fill the void of knowledge about the combination of clinical and commercial services presented on the websites of licensed community pharmacies. A content analysis of clinical and commercial services presented in a random sample of 200 licensed community pharmacy websites from Great Britain, the Netherlands, the Canadian provinces British Columbia and Manitoba, and the Australian states New South Wales and Western Australia was conducted. The top five specific services mentioned on the community pharmacy websites were cosmetic products (126/200, 63.0%), medication refill request options (124/200, 62.0%), over-the-counter medicine (115/200, 57.5%), complementary and alternative medicine (107/200, 53.5%), and home medical aids (98/200, 49.0%). On average, 72.5% (145/200) of the community pharmacy websites across the 4 countries included a combination of clinical and commercial services. A combination of clinical and commercial services was more often present on chain pharmacy websites (120/147, 82.8%) than single pharmacy websites (25/53, 47%; P<.001), and most often on the Canadian community pharmacy websites, followed by the Australian, British, and Dutch pharmacy websites, respectively (P<.02). Furthermore, more than half of the pharmacies' homepages contained a combination of clinical and commercial images (107/200, 53.5%), and almost half of the homepage menus contained a combination of clinical and commercial items (99/200, 49.5%). The latter were, again, more common on chain pharmacy than single pharmacy websites (P<.001), with significant differences between countries (P<.001). A considerable share of websites of licensed community pharmacies in Great Britain, the Netherlands, Canada, and Australia combine clinical services with commercial services. Previous research into the presence of a combination of commercial and professional services suggests that such a combination may lead to increased interest in commercial services that may be unnecessary or inappropriate to patients' health.


Residual Curarization and Postoperative Respiratory Complications Following Laparoscopic Sleeve Gastrectomy. The Effect of Reversal Agents: Sugammadex vs. Neostigmine.
Incomplete muscle relaxant reversal or re-curarization may be associated with postoperative respiratory complications. In this retrospective study we compared the incidence of postoperative residual curarization and respiratory complications in association with the type of muscle relaxant reversal agent, sugammadex or neostigmine, in patients undergoing laparoscopic sleeve gastrectomy. We reviewed the charts of all patients (179) undergoing laparoscopic sleeve gastrectomy from July 2012 to July 2013 at Wolfson Medical Center. Sugammadex 1.5-2 mg/kg (112 patients) or neostigmine 2.5 mg (67 patients) were used as reversal agents. Results were compared by the type of reversal agent employed. Compared parameters included demographic and anaesthetic data, residual curarization, oxyhemoglobin saturation (SpO2) in the recovery room (PACU), episodes of SpO2 lower than 90% in PACU, unexpected intensive care (ICU) admissions, incidence of atelectasis and pneumonia, re-intubation and duration of hospitalization. Obstructive sleep apnea syndrome (OSAS) was more frequent in the sugammadex group (19% vs. 8%; p = 0.026). Total intravenous anesthesia (TIVA) was more frequently associated with sugammadex (33% vs. 16%; p = 0.007). There were no differences in postoperative residual curarization, SpO2 < 90% episodes, reintubation, ICU admissions, pulmonary complications and duration of hospitalization. With the inherent limitations of a retrospective study, the use of sugammadex following laparoscopic sleeve gastrectomy showed no advantage over neostigmine in terms of residual curarization and respiratory complications.


Schistosoma mansoni treatment reduces HIV entry into cervical CD4+ T cells and induces IFN-I pathways.
Schistosoma mansoni (Sm) infection has been linked with an increased risk of HIV acquisition in women. Therefore, defining the mechanism(s) by which Sm alters HIV susceptibility might lead to new HIV prevention strategies. Here, we analyze the impact of standard Sm therapy in HIV-uninfected Sm+ Ugandan adult women on genital HIV susceptibility and mucosal and systemic immunology. Schistosomiasis treatment induces a profound reduction of HIV entry into cervical and blood CD4+ T cells that is sustained for up to two months, despite transient systemic and mucosal immune activation and elevated genital IL-1α levels. Genital IFN-α2a levels are also elevated post-treatment, and IFN-α2a blocks HIV entry into primary CD4+ T cells ex vivo. Transcriptomic analysis of blood mononuclear cells post-Sm treatment shows IFN-I pathway up-regulation and partial reversal of Sm-dysregulated interferon signaling. These findings indicate that Sm therapy may reduce HIV susceptibility for women with Sm infection, potentially through de-repression of IFN-I pathways.


Future directions for the development of virtual reality within an automotive manufacturer.
Virtual Reality (VR) can reduce time and costs, and lead to increases in quality, in the development of a product. Given the pressure on car companies to reduce time-to-market and to continually improve quality, the automotive industry has championed the use of VR across a number of applications, including design, manufacturing, and training. This paper describes interviews with 11 engineers and employees of allied disciplines from an automotive manufacturer about their current physical and virtual properties and processes. The results guided a review of research findings and scientific advances from the academic literature, which formed the basis of recommendations for future developments of VR technologies and applications. These include: develop a greater range of virtual contexts; use multi-sensory simulation; address perceived differences between virtual and real cars; improve motion capture capabilities; implement networked 3D technology; and use VR for market research.


The Canine Morbillivirus Strain Associated with An Epizootic in Caspian Seals Provides New Insights into the Evolutionary History of this Virus.
Canine morbillivirus (canine distemper virus; CDV) is a worldwide distributed morbillivirus that causes sporadic cases and recurrent epizootics among an increasing number of wild, feral, and domestic animal species. We investigated the evolutionary history of CDV strains involved in the 1988 Lake Baikal (CDVPS88) and the 2000 Caspian Sea (CDVPC00) seal die-offs by recovery of full-length sequences from archived material using next-generation sequencing. Bayesian phylogenetic analyses indicated that CDVPC00 constitutes a novel strain in a separate clade (tentatively termed "Caspian") from the America-1 clade, which is comprised of older vaccine strains. The America-1/Caspian monophyletic group is positioned most basally with respect to other clades and is estimated to have separated from other CDV clades around 1832. Our results indicate that CDVPC00 recovered from the epizootic in the Caspian Sea in 2000 belongs to a previously undetected novel clade and constitutes the most ancestral wild-type CDV clade.


Gel versus capillary electrophoresis genotyping for categorizing treatment outcomes in two anti-malarial trials in Uganda.
Molecular genotyping is performed in anti-malarial trials to determine whether recurrent parasitaemia after therapy represents a recrudescence (treatment failure) or new infection. The use of capillary instead of agarose gel electrophoresis for genotyping offers technical advantages, but it is unclear whether capillary electrophoresis will result in improved classification of anti-malarial treatment outcomes. Samples were genotyped using both gel and capillary electrophoresis from randomized trials of artemether-lumefantrine (AL) vs. dihydroartemisinin-piperaquine (DP) performed in two areas of Uganda: Kanungu, where transmission is moderate, and Apac, where transmission is very high. Both gel and capillary methods evaluated polymorphic regions of the merozoite surface protein 1 and 2 and glutamine rich protein genes. Capillary electrophoresis detected more alleles and provided higher discriminatory power than agarose gel electrophoresis at both study sites. There was only moderate agreement between classification of outcomes with the two methods in Kanungu (kappa = 0.66) and poor agreement in Apac (kappa = 0.24). Overall efficacy results were similar when using gel vs. capillary methods in Kanungu (42-day risk of treatment failure for AL: 6.9% vs. 5.5%, p = 0.4; DP 2.4% vs. 2.9%, p = 0.5). However, the measured risk of recrudescence was significantly higher when using gel vs. capillary electrophoresis in Apac (risk of treatment failure for AL: 17.0% vs. 10.7%, p = 0.02; DP: 8.5% vs. 3.4%, p = 0.03). Risk differences between AL and DP were not significantly different whether gel or capillary methods were used. Genotyping with gel electrophoresis overestimates the risk of recrudescence in anti-malarial trials performed in areas of high transmission intensity. Capillary electrophoresis provides more accurate outcomes for such trials and should be performed when possible. In areas of moderate transmission, gel electrophoresis appears adequate to estimate comparative risks of treatment failure.


Outcome following aminosalicylate therapy in children newly diagnosed as having ulcerative colitis.
Despite a paucity of published supporting data, 5-aminosalicylate (5-ASA) use in pediatric ulcerative colitis (UC) is common. The present study describes the use and outcome of a large multicenter inception cohort of children with UC treated with 5-ASA. Data were obtained from the Pediatric Inflammatory Bowel Disease Collaborative Research Group Registry, a prospective North American observational study of children newly diagnosed as having inflammatory bowel disease ages 16 years or younger. Patient data are recorded at diagnosis, 30 days, and then quarterly. Patients are managed by physician dictate, not protocol. Disease activity is classified by physician global assessment. The primary outcome examined was corticosteroid (CS) free, inactive UC at 1 year following initiation of 5-ASA within 30 days of diagnosis (with or without concomitant CS use) without the need for rescue therapy (immunomodulators, biologics, or colectomy). Study subjects included 213 patients newly diagnosed as having UC who received oral 5-ASA compounds (115 of whom also received CS) during the first 30 days after diagnosis, and no other oral therapies for the treatment of UC. Of these 213 patients, 86 (40%) were CS free and physician global assessment inactive at 1 year without rescue. Outcome was not associated with disease severity at diagnosis, demographic or laboratory factors examined, or initial dose of 5-ASA used. Forty percent of children taking 5-ASA as primary maintenance therapy at diagnosis are in CS-free remission after 1 year of treatment. Further pediatric studies will be needed to address whether increased adherence and/or higher dosing schedules will improve outcomes.


Maximization of mutual information for offline Thai handwriting recognition.
This paper aims to improve the performance of an HMM-based offline Thai handwriting recognition system through discriminative training and the use of fine-tuned feature extraction methods. The discriminative training is implemented by maximizing the mutual information between the data and their classes. The feature extraction is based on our proposed block-based PCA and composite images, shown to be better at discriminating Thai confusable characters. We demonstrate significant improvements in recognition accuracies compared to the classifiers that are not discriminatively optimized.


Griottes: a generalist tool for network generation from segmented tissue images.
Microscopy techniques and image segmentation algorithms have improved dramatically this decade, leading to an ever increasing amount of biological images and a greater reliance on imaging to investigate biological questions. This has created a need for methods to extract the relevant information on the behaviors of cells and their interactions, while reducing the amount of computing power required to organize this information. This task can be performed by using a network representation in which the cells and their properties are encoded in the nodes, while the neighborhood interactions are encoded by the links. Here, we introduce Griottes, an open-source tool to build the "network twin" of 2D and 3D tissues from segmented microscopy images. We show how the library can provide a wide range of biologically relevant metrics on individual cells and their neighborhoods, with the objective of providing multi-scale biological insights. The library's capacities are demonstrated on different image and data types. This library is provided as an open-source tool that can be integrated into common image analysis workflows to increase their capacities.


Monozygotic twins with NASH cirrhosis: cumulative effect of multiple single nucleotide polymorphisms?
Multiple genetic and environmental factors interact to determine an individual's predisposition to non-alcoholic fatty liver disease and its phenotypic characteristics. Association studies have found a number of alleles associated with the development of non-alcoholic steatohepatitis. Our aim was to investigate whether multiple risk-associated alleles may be present in affected monozygotic twins, indicating underlying genetic predisposition to non-alcoholic steatohepatitis. We determined the genotype of 14 candidate gene polymorphisms (at 11 unlinked loci) in a set of monozygotic twins who presented with cirrhosis within 18 months of each other. Genotyping revealed multiple single nucleotide polymorphisms at 9 independent loci in genes PNPLA3, APOC3, GCKR, TRIB1, LYPLAL1, PPP1R3B, COL13A1, and EFCAB4B, previously implicated in contributing to non-alcoholic steatohepatitis pathogenesis. In conclusion, this case series illustrates the potential cumulative effect of multiple polymorphisms in the development and potential progression of a complex trait such as NASH cirrhosis.


Mitigating losses: how scientific organisations can help address the impact of the COVID-19 pandemic on early-career researchers.
Scientific collaborations among nations to address common problems and to build international partnerships as part of science diplomacy is a well-established notion. The international flow of people and ideas has played an important role in the advancement of the 'Sciences' and the current pandemic scenario has drawn attention towards the genuine need for a stronger role of science diplomacy, science advice and science communication. In dealing with the COVID-19 pandemic, visible interactions across science, policy, science communication to the public and diplomacy worldwide have promptly emerged. These interactions have benefited primarily the disciplines of knowledge that are directly informing the pandemic response, while other scientific fields have been relegated. The effects of the COVID-19 pandemic on scientists of all disciplines and from all world regions are discussed here, with a focus on early-career researchers (ECRs), as a vulnerable population in the research system. Young academies and ECR-driven organisations could suggest ECR-powered solutions and actions that could have the potential to mitigate these effects on ECRs working on disciplines not related to the pandemic response. In relation with governments and other scientific organisations, they can have an impact on strengthening and creating fairer scientific systems for ECRs at the national, regional, and global level.


Association of Oral Corticosteroid Bursts With Severe Adverse Events in Children.
The adverse effects from the long-term use of oral corticosteroids are known, but, to our knowledge, few studies have reported the risk of corticosteroid bursts, particularly among children. To quantify the associations of corticosteroid bursts with severe adverse events, including gastrointestinal (GI) bleeding, sepsis, pneumonia, and glaucoma, in children. This study used data derived from the National Health Insurance Research Database in Taiwan from January 1, 2013, to December 31, 2017, on children younger than 18 years of age and used a self-controlled case series design. Data were analyzed from January 1 to July 30, 2020. Oral corticosteroid bursts (defined as oral corticosteroid use for ≤14 days). Incidence rates were calculated of 4 severe adverse events (GI bleeding, sepsis, pneumonia, and glaucoma) in children who did or did not receive corticosteroid bursts. Conditional fixed-effect Poisson regression was used to estimate incidence rate ratios (IRRs) of severe adverse events within 5 to 30 days and 31 to 90 days after initiation of corticosteroid bursts. Among 4 542 623 children, 23% (1 064 587; 544 268 boys [51.1%]; mean [SD] age, 9.7 [5.8] years) were prescribed a single corticosteroid burst. The most common indications were acute respiratory tract infections and allergic diseases. The incidence rate differences per 1000 person-years between children administered a single corticosteroid burst and those not prescribed corticosteroids were 0.60 (95% CI, 0.55-0.64) for GI bleeding, 0.03 (95% CI, 0.02-0.05) for sepsis, 9.35 (95% CI, 9.19-9.51) for pneumonia, and 0.01 (95% CI, 0.01-0.03) for glaucoma. The IRRs within 5 to 30 days after initiating corticosteroid bursts were 1.41 (95% CI, 1.27-1.57) for GI bleeding, 2.02 (95% CI, 1.55-2.64) for sepsis, 2.19 (95% CI, 2.13-2.25) for pneumonia, and 0.98 (95% CI, 0.85-1.13) for glaucoma; the IRRs within the subsequent 31 to 90 days were 1.10 (95% CI, 1.02-1.19) for GI bleeding, 1.08 (95% CI, 0.88-1.32) for sepsis, 1.09 (95% CI, 1.07-1.11) for pneumonia, and 0.95 (95% CI, 0.85-1.06) for glaucoma. This study suggests that corticosteroid bursts, which are commonly prescribed for children with respiratory and allergic conditions, are associated with a 1.4- to 2.2-fold increased risk of GI bleeding, sepsis, and pneumonia within the first month after initiation of corticosteroid therapy that is attenuated during the subsequent 31 to 90 days.


Unintentional injuries in the home in the United States Part I: mortality.
Unintentional injuries are a leading cause of death in the United States. It is unclear, however, what proportion of these injuries occur in the home. The purpose of this paper is to quantify and describe fatal unintentional injuries that take place in the home environment. Data from the National Vital Statistics System (NVSS) were used to calculate average annual rates for unintentional home injury deaths, with 95% confidence intervals from 1992 to 1999 for the United States overall, and by mechanism of injury, gender, and age group. From 1992 to 1999, an average of 18,048 unintentional home injury deaths occurred annually in the United States (6.83 deaths per 100,000). Home injury deaths varied by age and gender, with males having higher rates of home injury death than females (8.78 vs 4.97 per 100,000), and older adults (>/=70 years) having higher rates than all other age groups. Falls (2.25 per 100,000), poisoning (1.83 per 100,000), and fire/burn injuries (1.29 per 100,000) were the leading causes of home injury death. Rates of fall death were highest for older adults, poisoning deaths were highest among middle-aged adults, and fire/burn death rates were highest among children. Inhalation/suffocation and drowning deaths were important injury issues for young children. Unintentional injury in the home is a significant problem. Specific home injury issues include falls among older adults, poisonings among middle-aged adults, fire/burn injuries among older adults and children, and inhalation/suffocation and drowning among young children. In addition, recommendations are presented for improvements to the NVSS.


UK Biobank: a globally important resource for cancer research.
UK Biobank is a large-scale prospective study with deep phenotyping and genomic data. Its open-access policy allows researchers worldwide, from academia or industry, to perform health research in the public interest. Between 2006 and 2010, the study recruited 502,000 adults aged 40-69 years from the general population of the United Kingdom. At enrolment, participants provided information on a wide range of factors, physical measurements were taken, and biological samples (blood, urine and saliva) were collected for long-term storage. Participants have now been followed up for over a decade with more than 52,000 incident cancer cases recorded. The study continues to be enhanced with repeat assessments, web-based questionnaires, multi-modal imaging, and conversion of the stored biological samples to genomic and other '-omic' data. The study has already demonstrated its value in enabling research into the determinants of cancer, and future planned enhancements will make the resource even more valuable to cancer researchers. Over 26,000 researchers worldwide are currently using the data, performing a wide range of cancer research. UK Biobank is uniquely placed to transform our understanding of the causes of cancer development and progression, and drive improvements in cancer treatment and prevention over the coming decades.


Mobile phone exposure does not induce apoptosis on spermatogenesis in rats.
Some studies have reported that microwave radiation can have adverse effects on reproduction. Therefore, the purpose of this study was to investigate the apoptosis-inducing effect of mobile phone exposure on spermatogonia in seminiferous tubules. The study was carried out on 31 Wistar albino adult male rats. The rats were separated into three groups in this study (cage control: 10, sham group: 7, and exposed group: 14). For the study group, rats were exposed to radiation 2 h/day (7 days/week) for 10 months. For the sham group, rats were placed into the carousel and the same procedure was applied except that the generator was turned off. For the cage control, nothing was applied to the rats in this group and they completed their life cycle in the cage during the study period. In this study, rats were sacrificed after 10 months of exposure and their testes were taken. Testes tissue was immunohistochemically stained for the active (cleaved) caspase-3. Positively stained cells were counted in up to ten different areas, and the frequency of positive cells was determined in percentage. Scoring was done by taking into account both the intensity of staining and the distribution of positively stained cells. Therefore, protein expression was evaluated by a semiquantitative scoring system. The final score for apoptosis of testes in the exposed group was not statistically significant according to the sham and the cage control groups (p>0.05). The results of this study showed that 2 h/day (7 days/week) exposure of 900 MHz radiation over a period of 10 months does not affect the active (cleaved) caspase-3 levels in testes, a well-known feature of typical apoptosis.


Structural basis for translational stalling by human cytomegalovirus and fungal arginine attenuator peptide.
Specific regulatory nascent chains establish direct interactions with the ribosomal tunnel, leading to translational stalling. Despite a wealth of biochemical data, structural insight into the mechanism of translational stalling in eukaryotes is still lacking. Here we use cryo-electron microscopy to visualize eukaryotic ribosomes stalled during the translation of two diverse regulatory peptides: the fungal arginine attenuator peptide (AAP) and the human cytomegalovirus (hCMV) gp48 upstream open reading frame 2 (uORF2). The C terminus of the AAP appears to be compacted adjacent to the peptidyl transferase center (PTC). Both nascent chains interact with ribosomal proteins L4 and L17 at tunnel constriction in a distinct fashion. Significant changes at the PTC were observed: the eukaryotic-specific loop of ribosomal protein L10e establishes direct contact with the CCA end of the peptidyl-tRNA (P-tRNA), which may be critical for silencing of the PTC during translational stalling. Our findings provide direct structural insight into two distinct eukaryotic stalling processes.


Role of cooling and drying in hyperventilation induced asthma.
Respiratory heat loss has been proposed as a mechanism of exercise induced asthma. Whether the predominant stimulus is airway drying or cooling remains unclear. We have measured changes in FEV1 after isocapnic cold air hyperventilation (CAH) (-23.4 degrees (SD 0.43 degrees) C) and dry ambient air hyperventilation (AAH) (18.7 degrees (0.52 degrees)C) in seven asthmatic patients (mean age 31 (SD 9) years and baseline FEV1 3.2(0.9)1) and in seven normal subjects (age 28(6) years and FEV1 3.6(0.7)1). The inspired water content in both cases was 0.3 mg/l air. The rate of respiratory heat exchange per breath was calculated in watts (W) with microcomputer based equipment. Cold air hyperventilation caused a fall in FEV1 almost twice that of ambient air hyperventilation at each level of ventilation: CAH v AAH (% fall) 8.0 (5.1) v 3.9 (4.0) at 15 l/min, 11.6 (7.8) v 7.0 (4.4) at 30 l/min, and 20.7 (10.9) v 12.4 (6.3) at 60 l/min. Identical latent heat loss (evaporative drying) was imposed on the airway during the two challenges. Sensible heat loss (convective cooling) in cold air hyperventilation was 41 W at 15 l/min, 63 W at 30 l/min, and 114 W at 60 l/min; whereas in ambient air hyperventilation the loss was 6, 13, and 23 W respectively. It is concluded that the rate of cooling of the upper airway is the predominant stimulus in hyperventilation induced asthma.


A novel submerged Rotala rotundifolia, its growth characteristics and remediation potential for eutrophic waters.
The vegetative growth and remediation potential of Rotala rotundifolia, a novel submerged aquatic plant, for eutrophic waters were investigated on different sediments, and under a range of nitrogen concentrations. Rotala Rotundifolia grew better on silt than on sand and gravel in terms of plant height, tiller number and biomass accumulation. Percent increment of biomass was enhanced at low water nitrogen (ammonium nitrogen concentration ≤10 mg/L). The maximum total nitrogen and total phosphorus removals in the overlying water were between 54% to 66% and 42% to 57%, respectively. Nitrogen contents in the sediments increased with increasing water nitrogen levels, whereas, nitrogen contents in the plant tissues showed no apparent regularity, and the greatest value was obtained at ammonium nitrogen concentration 15 mg/L. Both phosphorus contents in the sediments and tissues of plants were not affected significantly by additional nitrogen supply. Direct nitrogen uptake by plants was in the range of 16% to 39% when total phosphorus concentration was 1.0 mg/L. These results suggested that Rotala Rotundifolia can be used to effectively remove nitrogen and phosphorus in eutrophic waters.


Longitudinal trends in prescribing for elderly patients: two surveys four years apart.
Elderly people are prescribed more drugs than younger people. The consequences of excessive or unwise prescribing, such as drug interactions, are well known. A longitudinal study was undertaken to examine levels and patterns of prescribed drug use among a group of elderly people. Use of prescribed drugs by a sample of elderly people in Nottingham aged 65 years and over was assessed on two occasions four years apart, in 1985 and 1989. Complete drug data were available for 1003 respondents in 1985 and 662 respondents in 1989 (with attrition due mainly to mortality). Drug use increased significantly with age. Women took significantly more drugs than men. Approximately half of respondents were taking at least two drugs. The overall number of drugs per person being taken in 1989 was significantly greater than in 1985. This difference remained significant when age and mortality were controlled, suggesting that changes in drug use over time within this sample may reflect genuine changes in prescribing practice (rather than simply the effects of ageing). The most commonly prescribed drug classes on each occasion were drugs for the cardiovascular system, central nervous system, musculoskeletal system, gastrointestinal tract and respiratory system. The subgroups of drugs most commonly reported at each interview were diuretics, hypnotics and anxiolytics, analgesics and non-steroidal anti-inflammatory drugs. Drugs within the category 'hypnotics and anxiolytics' showed clear and differential trends over time, with the use of anxiolytics falling, while the use of hypnotics increased. Among those respondents admitted to residential care during the course of the study higher levels of psychotropic drug use, and an increase in antipsychotic medication, were observed. It is important that the drug regimens of elderly people are frequently reviewed to ensure that only the minimum number of effective drugs, in the simplest regimen, are prescribed.


Capacity of MIMO free space optical communications using multiple partially coherent beams propagation through non-Kolmogorov strong turbulence.
We study the average capacity performance for multiple-input multiple-output (MIMO) free-space optical (FSO) communication systems using multiple partially coherent beams propagating through non-Kolmogorov strong turbulence, assuming equal gain combining diversity configuration and the sum of multiple gamma-gamma random variables for multiple independent partially coherent beams. The closed-form expressions of scintillation and average capacity are derived and then used to analyze the dependence on the number of independent diversity branches, power law α, refractive-index structure parameter, propagation distance and spatial coherence length of source beams. Obtained results show that, the average capacity increases more significantly with the increase in the rank of MIMO channel matrix compared with the diversity order. The effect of the diversity order on the average capacity is independent of the power law, turbulence strength parameter and spatial coherence length, whereas these effects on average capacity are gradually mitigated as the diversity order increases. The average capacity increases and saturates with the decreasing spatial coherence length, at rates depending on the diversity order, power law and turbulence strength. There exist optimal values of the spatial coherence length and diversity configuration for maximizing the average capacity of MIMO FSO links over a variety of atmospheric turbulence conditions.


A Retrospective Review of Upper Gastrointestinal Bleed Outcomes During Hospital Admission While on Oral Anticoagulation.
Introduction Direct-acting oral anticoagulants (DOACs) are approved for stroke prevention in non-valvular atrial fibrillation and treatment of venous thromboembolism. Most recent guidelines recommend DOACs over warfarin for most diagnoses given their predictable pharmacodynamics, lack of required monitoring, and safety profile. Specific outcomes such as shock, acute renal failure, and blood transfusion requirement while on oral anticoagulation compared to no anticoagulation remain unknown in patients with upper gastrointestinal (GI) bleeds. Methods This retrospective study used the HCA Healthcare Enterprise Data Warehouse (EDW) to analyze 13,440 patients aged >18 years that were admitted with an upper GI bleed from January 2017 to December 2019. The patients were categorized based on oral anticoagulant (i.e. rivaroxaban, apixaban, dabigatran and warfarin). The control group was patients admitted with an upper GI bleed not on oral anticoagulation. We evaluated the severity of upper GI bleeds while on oral anticoagulation based on the outcomes: mortality rate, length of stay, acute renal failure, shock, and need for packed red blood cell transfusions (pRBC). Comorbid conditions assessed were coronary artery disease (CAD), chronic obstructive pulmonary disease (COPD), heart failure (HF), atrial fibrillation (AF), venous thromboembolism (VTE), peripheral vascular disease (PVD), tobacco abuse, alcohol abuse, and chronic kidney disease (CKD). Home use of proton pump inhibitors (PPI), aspirin, and P2Y12 inhibitors were also evaluated. Results Patients on a DOAC without home PPI have a mortality odds ratio of 3.066 with a confidence interval (CI) greater than 95% (1.48-6.26, p<0.05) compared to patients on a DOAC and home PPI. Patients on warfarin and no home PPI have a mortality odds ratio of 5.55 (95% CI (1.02-30.35), p<0.05) compared to those on warfarin with home PPI use. In the no anticoagulation group, those not on PPI have an odds ratio of 3.28 (95% CI (2.54-4.24), p<0.05) of death compared to home PPI use. There was no statistical difference in mortality between each DOAC and warfarin. There was no difference in the presence of acute renal failure or shock when comparing each DOAC, warfarin, and no medication. For patients presenting with GI bleed, 0.8414 units of pRBC were transfused. Patients not on oral anticoagulation were found to have statistically significant decrease in pRBC transfusion if they did not report alcohol use, CKD, HF, AF, VTE, PVD. Patients on DOACs and alcohol use have an average pRBC transfusion count that is 0.922 units more than those without reported alcohol use (p=0.006). In the warfarin group, there was no statistical significance noted when comparing pRBC transfusions and also when comparing to baseline comorbidities. Conclusion The retrospective study leads us to conclude that overall, patients taking the DOACs or warfarin had no statistically significant increase in RBC transfusions, length of stay, shock, acute renal failure, or mortality rate compared to patients who were not on oral anticoagulation. Home PPI use was shown to lower odds of mortality in patients on anticoagulation who presented with upper GI bleeding. PPI use had no effect on the need for transfusion or length of stay in patients on anticoagulation. These results can help predict which patients are likely to have higher mortality based on the use of home PPIs.


Association between size of the lateral ventricle and asymmetry of the fornix in patients with temporal lobe epilepsy.
Our goal was to determine whether the size of the lateral ventricle influences asymmetry of the fornix in patients with mesial temporal sclerosis. The cross-sectional area of the lateral ventricle was measured along with the thickness of the fornix in 18 patients with pathologically proved mesial temporal sclerosis. Two additional patients with ventricular asymmetry and seizures but without mesial sclerosis were also studied. Ten of 18 patients with mesial temporal sclerosis had equally sized ventricular areas or less than a 25% difference between the two sides. In seven of these 10, the left and right fornices were equal; in the other three patients, the fornix was thinner on the side of mesial sclerosis. Among the eight patients with ventricular area differences greater than 25%, the larger ventricle was on the side of mesial sclerosis in five cases. In all of these, the ipsilateral fornix was thinner. In three of eight cases, the contralateral ventricle was larger; in two, the left and right fornix were equal; and in one, the fornix contralateral to the side of mesial sclerosis was smaller. The greatest difference in fornix thickness was noted in patients with ventricular asymmetry greater than 40%, regardless of side or presence of mesial temporal sclerosis. The size of the lateral ventricle may influence the apparent thickness of the fornix.


An integrated workflow for charting the human interaction proteome: insights into the PP2A system.
Protein complexes represent major functional units for the execution of biological processes. Systematic affinity purification coupled with mass spectrometry (AP-MS) yielded a wealth of information on the compendium of protein complexes expressed in Saccharomyces cerevisiae. However, global AP-MS analysis of human protein complexes is hampered by the low throughput, sensitivity and data robustness of existing procedures, which limit its application for systems biology research. Here, we address these limitations by a novel integrated method, which we applied and benchmarked for the human protein phosphatase 2A system. We identified a total of 197 protein interactions with high reproducibility, showing the coexistence of distinct classes of phosphatase complexes that are linked to proteins implicated in mitosis, cell signalling, DNA damage control and more. These results show that the presented analytical process will substantially advance throughput and reproducibility in future systematic AP-MS studies on human protein complexes.


Why a Confirmation Strategy Dominates Psychological Science.
Our research explored the incidence and appropriateness of the much-maligned confirmatory approach to testing scientific hypotheses. Psychological scientists completed a survey about their research goals and strategies. The most frequently reported goal is to test the non-absolute hypothesis that a particular relation exists in some conditions. As expected, few scientists reported testing universal hypotheses. Most indicated an inclination to use a confirmation strategy to test the non-absolute hypotheses that a particular relation sometimes occurs or sometimes does not occur, and a disconfirmation strategy to test the absolute hypotheses that a particular relation always occurs or never occurs. The confirmatory search that dominates the field was found to be associated with the testing of non-absolute hypotheses. Our analysis indicates that a confirmatory approach is the normatively correct test of the non-absolute hypotheses that are the starting point of most studies. It also suggests that the strategy of falsification that was once proposed by Popper is generally incorrect given the infrequency of tests of universal hypotheses.


Categorical colour perception occurs in both signalling and non-signalling colour ranges in a songbird.
Although perception begins when a stimulus is transduced by a sensory neuron, numerous perceptual mechanisms can modify sensory information as it is processed by an animal's nervous system. One such mechanism is categorical perception, in which (1) continuously varying stimuli are labelled as belonging to a discrete number of categories and (2) there is enhanced discrimination between stimuli from different categories as compared with equally different stimuli from within the same category. We have shown previously that female zebra finches ( Taeniopygia guttata) categorically perceive colours along an orange-red continuum that aligns with the carotenoid-based coloration of male beaks, a trait that serves as an assessment signal in female mate choice. Here, we demonstrate that categorical perception occurs along a blue-green continuum as well, suggesting that categorical colour perception may be a general feature of zebra finch vision. Although we identified two categories in both the blue-green and the orange-red ranges, we also found that individuals could better differentiate colours from within the same category in the blue-green as compared with the orange-red range, indicative of less clear categorization in the blue-green range. We discuss reasons why categorical perception may vary across the visible spectrum, including the possibility that such differences are linked to the behavioural or ecological function of different colour ranges.


DNA methylation patterns facilitate the identification of microRNA transcription start sites: a brain-specific study.
Predicting the transcription start sites (TSSs) of microRNAs (miRNAs) is important for understanding how these small RNA molecules, known to regulate translation and stability of protein-coding genes, are regulated themselves. Previous approaches are primarily based on genetic features, trained on TSSs of protein-coding genes, and have low prediction accuracy. Recently, a support vector machine based technique has been proposed for miRNA TSS prediction that uses known miRNA TSS for training the classifier along with a set of existing and novel CpG island based features. Current progress in epigenetics research has provided genomewide and tissue-specific reports about various phenotypic traits. We hypothesize that incorporating epigenetic characteristics into statistical models may lead to better prediction of primary transcripts of human miRNAs. In this paper, we have tested our hypothesis on brain-specific miRNAs by using epigenetic as well as genetic features to predict the primary transcripts. For this, we have used a sophisticated feature selection technique and a robust classification model. Our prediction model achieves an accuracy of more than 80% and establishes the potential of epigenetic analysis for in silico prediction of TSSs.


Computational Psychiatry: towards a mathematically informed understanding of mental illness.
Computational Psychiatry aims to describe the relationship between the brain's neurobiology, its environment and mental symptoms in computational terms. In so doing, it may improve psychiatric classification and the diagnosis and treatment of mental illness. It can unite many levels of description in a mechanistic and rigorous fashion, while avoiding biological reductionism and artificial categorisation. We describe how computational models of cognition can infer the current state of the environment and weigh up future actions, and how these models provide new perspectives on two example disorders, depression and schizophrenia. Reinforcement learning describes how the brain can choose and value courses of actions according to their long-term future value. Some depressive symptoms may result from aberrant valuations, which could arise from prior beliefs about the loss of agency ('helplessness'), or from an inability to inhibit the mental exploration of aversive events. Predictive coding explains how the brain might perform Bayesian inference about the state of its environment by combining sensory data with prior beliefs, each weighted according to their certainty (or precision). Several cortical abnormalities in schizophrenia might reduce precision at higher levels of the inferential hierarchy, biasing inference towards sensory data and away from prior beliefs. We discuss whether striatal hyperdopaminergia might have an adaptive function in this context, and also how reinforcement learning and incentive salience models may shed light on the disorder. Finally, we review some of Computational Psychiatry's applications to neurological disorders, such as Parkinson's disease, and some pitfalls to avoid when applying its methods.


The 'glial' glutamate transporter, EAAT2 (Glt-1) accounts for high affinity glutamate uptake into adult rodent nerve endings.
The excitatory amino acid transporters (EAAT) removes neurotransmitters glutamate and aspartate from the synaptic cleft. Most CNS glutamate uptake is mediated by EAAT2 into glia, though nerve terminals show evidence for uptake, through an unknown transporter. Reverse-transcriptase PCR identified the expression of EAAT1, EAAT2, EAAT3 and EAAT4 mRNAs in primary cultures of mouse cortical or striatal neurones. We have used synaptosomes and glial plasmalemmal vesicles (GPV) from adult mouse and rat CNS to identify the nerve terminal transporter. Western blotting showed detectable levels of the transporters EAAT1 (GLAST) and EAAT2 (Glt-1) in both synaptosomes and GPVs. Uptake of [3H]D-aspartate or [3H]L-glutamate into these preparations revealed sodium-dependent uptake in GPV and synaptosomes which was inhibited by a range of EAAT blockers: dihydrokainate, serine-o-sulfate, l-trans-2,4-pyrrolidine dicarboxylate (PDC) (+/-)-threo-3-methylglutamate and (2S,4R )-4-methylglutamate. The IC50 values found for these compounds suggested functional expression of the 'glial, transporter, EAAT2 in nerve terminals. Additionally blockade of the majority EAAT2 uptake sites with 100 micro m dihydrokainate, failed to unmask any functional non-EAAT2 uptake sites. The data presented in this study indicate that EAAT2 is the predominant nerve terminal glutamate transporter in the adult rodent CNS.


Mining and characterization of ubiquitin E3 ligases expressed in the mouse testis.
Ubiquitin-mediated protein modification and degradation are believed to play important roles in mammalian spermatogenesis. The catalogues of ubiquitin activating enzymes, conjugating enzymes, and ligases (E3s) have been known for mammals such as mice and humans. However, a systematic characterization of E3s expressed during spermatogenesis has not been carried out. In present study, we set out to mine E3s from the mouse genome and to characterize their expression pattern, subcellular localization, and enzymatic activities based on microarray data and biochemical assays. We identified 398 putative E3s belonging to the RING, U-box, and HECT subfamilies and found that most genes were conserved between mice and humans. We discovered that 73 of them were highly or specifically expressed in the testes based on the microarray expression data. We selected 10 putative E3 genes to examine their mRNA expression pattern, and several genes to study their subcellular localization and E3 ligase activity. RT-PCR results showed that all the selected genes were predominately expressed in the testis. Some putative E3s were localized in the cytoplasm while others were in both the cytoplasm and the nucleus. Moreover, all the selected proteins were enzymatically active as demonstrated by in vitro and in vivo assays. We have identified a large number of putative E3s that are expressed during mouse spermatogenesis. Among these, a significant portion is highly or specifically expressed in the testis. Subcellular localization and enzymatic activity assays suggested that these E3s might execute diverse functions in mammalian spermatogenesis. Our results may serve as an initial guide to the field for further functional analysis.


The secretory route of the leaderless protein interleukin 1beta involves exocytosis of endolysosome-related vesicles.
Interleukin 1beta (IL-1beta), a secretory protein lacking a signal peptide, does not follow the classical endoplasmic reticulum-to-Golgi pathway of secretion. Here we provide the evidence for a "leaderless" secretory route that uses regulated exocytosis of preterminal endocytic vesicles to transport cytosolic IL-1beta out of the cell. Indeed, although most of the IL-1beta precursor (proIL-1beta) localizes in the cytosol of activated human monocytes, a fraction is contained within vesicles that cofractionate with late endosomes and early lysosomes on Percoll density gradients and display ultrastructural features and markers typical of these organelles. The observation of organelles positive for both IL-1beta and the endolysosomal hydrolase cathepsin D or for both IL-1beta and the lysosomal marker Lamp-1 further suggests that they belong to the preterminal endocytic compartment. In addition, similarly to lysosomal hydrolases, secretion of IL-1beta is induced by acidotropic drugs. Treatment of monocytes with the sulfonylurea glibenclamide inhibits both IL-1beta secretion and vesicular accumulation, suggesting that this drug prevents the translocation of proIL-1beta from the cytosol into the vesicles. A high concentration of extracellular ATP and hypotonic medium increase secretion of IL-1beta but deplete the vesicular proIL-1beta content, indicating that exocytosis of proIL-1beta-containing vesicles is regulated by ATP and osmotic conditions.


Diazotrophy in alluvial meadows of subarctic river systems.
There is currently limited understanding of the contribution of biological N2 fixation (diazotrophy) to the N budget of large river systems. This natural source of N in boreal river systems may partially explain the sustained productivity of river floodplains in Northern Europe where winter fodder was harvested for centuries without fertilizer amendments. In much of the world, anthropogenic pollution and river regulation have nearly eliminated opportunities to study natural processes that shaped early nutrient dynamics of large river systems; however, pristine conditions in northern Fennoscandia allow for the retrospective evaluation of key biochemical processes of historical significance. We investigated biological N2 fixation (diazotrophy) as a potential source of nitrogen fertility at 71 independent floodplain sites along 10 rivers and conducted seasonal and intensive analyses at a subset of these sites. Biological N2 fixation occurred in all floodplains, averaged 24.5 kg N ha(-1) yr(-1) and was down regulated from over 60 kg N ha(-1) yr(-1) to 0 kg N ha(-1) yr(-1) by river N pollution. A diversity of N2-fixing cyanobacteria was found to colonize surface detritus in the floodplains. The data provide evidence for N2 fixation to be a fundamental source of new N that may have sustained fertility at alluvial sites along subarctic rivers. Such data may have implications for the interpretation of ancient agricultural development and the design of contemporary low-input agroecosystems.


Advanced MRI findings in patients with breast hamartomas.
Although it has been stated that breast hamartomas are rare tumors, radiologists frequently encounter them in their daily practices. Fat, glandular and fibrous tissues all produce a mass of disorganized but mature specialized cells. Because hamartomas do not have specific diagnostic histological features, the clinical and radiological findings are important in their diagnosis. The aim of this study is to present the advanced magnetic resonance imaging (MRI) findings of breast hamartomas. Eight patients with breast hamartomas were examined using MRI techniques in addition to ultrasonographic and/or mammographic findings. Each of the lesions examined showed a gradual enhancement pattern in its time-signal intensity curve on dynamic contrast-enhanced MRI. On MR spectroscopy, water and lipid peaks were detected that resembled normal breast tissue. The diffusion features of the lesions were variable due to the different ratios of the tissue elements constituting them on diffusion-weighted imaging (DWI). Advanced MRI findings may clarify diagnoses by providing additional information following sonography, especially in lactating or pregnant women, in whom mammographic examination is not preferred.


Global phenotypic analysis and transcriptional profiling defines the weak acid stress response regulon in Saccharomyces cerevisiae.
Weak organic acids such as sorbate are potent fungistatic agents used in food preservation, but their intracellular targets are poorly understood. We thus searched for potential target genes and signaling components in the yeast genome using contemporary genome-wide functional assays as well as DNA microarray profiling. Phenotypic screening of the EUROSCARF collection revealed the existence of numerous sorbate-sensitive strains. Sorbate hypersensitivity was detected in mutants of the shikimate biosynthesis pathway, strains lacking the PDR12 efflux pump or WAR1, a transcription factor mediating stress induction of PDR12. Using DNA microarrays, we also analyzed the genome-wide response to acute sorbate stress, allowing for the identification of more than 100 genes rapidly induced by weak acid stress. Moreover, a novel War1p- and Msn2p/4p-independent regulon that includes HSP30 was identified. Although induction of the majority of sorbate-induced genes required Msn2p/4p, weak acid tolerance was unaffected by a lack of Msn2p/4p. Ectopic expression of PDR12 from the GAL1-10 promoter fully restored sorbate resistance in a strain lacking War1p, demonstrating that PDR12 is the major target of War1p under sorbic acid stress. Interestingly, comparison of microarray data with results from the phenotypic screening revealed that PDR12 remained as the only gene, which is both stress inducible and required for weak acid resistance. Our results suggest that combining functional assays with transcriptome profiling allows for the identification of key components in large datasets such as those generated by global microarray analysis.


Management of irritable bowel syndrome in primary care: feasibility randomised controlled trial of mebeverine, methylcellulose, placebo and a patient self-management cognitive behavioural therapy website. (MIBS trial).
IBS affects 10-22% of the UK population. Abdominal pain, bloating and altered bowel habit affect quality of life, social functioning and time off work. Current GP treatment relies on a positive diagnosis, reassurance, lifestyle advice and drug therapies, but many suffer ongoing symptoms.A recent Cochrane review highlighted the lack of research evidence for IBS drugs. Neither GPs, nor patients have good evidence to inform prescribing decisions. However, IBS drugs are widely used: In 2005 the NHS costs were nearly £10 million for mebeverine and over £8 million for fibre-based bulking agents. CBT and self-management can be helpful, but poor availability in the NHS restricts their use. We have developed a web-based CBT self-management programme, Regul8, based on an existing evidence based self-management manual and in partnership with patients. This could increase access with minimal increased costs. The aim is to undertake a feasibility factorial RCT to assess the effectiveness of the commonly prescribed medications in UK general practice for IBS: mebeverine (anti-spasmodic) and methylcellulose (bulking-agent) and Regul8, the CBT based self-management website.135 patients aged 16 to 60 years with IBS symptoms fulfilling Rome III criteria, recruited via GP practices, will be randomised to 1 of 3 levels of the drug condition: mebeverine, methylcellulose or placebo for 6 weeks and to 1 of 3 levels of the website condition, Regul8 with a nurse telephone session and email support, Regul8 with minimal email support, or no website, thus creating 9 groups. Irritable bowel symptom severity scale and IBS-QOL will be measured at baseline, 6 and 12 weeks as the primary outcomes. An intention to treat analysis will be undertaken by ANCOVA for a factorial trial. This pilot will provide valuable information for a larger trial. Determining the effectiveness of commonly used drug treatments will help patients and doctors make informed treatment decisions regarding drug management of IBS symptoms, enabling better targeting of treatment. A web-based self-management CBT programme for IBS developed in partnership with patients has the potential to benefit large numbers of patients with low cost to the NHS. Assessment of the amount of email or therapist support required for the website will enable economic analysis to be undertaken.


Medical futility in the natural attitude.
Medical futility has a long history going back to Plato but continues to be a controversial topic. Patients, families, and health professionals are faced with decisions about which treatments and interventions may be futile, but such questions as who decides; how do competing values get resolved; what value is placed on human life; how are decisions balanced according to reason and, in some circumstances, faith; and who decides effect, benefit, and burdens of treatments are often difficult to answer. The naïve reality of medical futility is explored in the natural attitude to expose presumptions and facts related to both physiologic (fact-based) and evaluative (value-based) futility components. Highlights from the bioethical and clinical literature, a review of 3 landmark cases, and implications for nursing practice are presented. This natural attitude description could serve as what ought to be bracketed for a future phenomenology.


Exercise pressor reflex function is altered in spontaneously hypertensive rats.
In hypertension, exercise elicits excessive elevations in mean arterial pressure (MAP) and heart rate (HR) increasing the risk for adverse cardiac events and stroke during physical activity. The exercise pressor reflex (a neural drive originating in skeletal muscle), central command (a neural drive originating in cortical brain centres) and the tonically active arterial baroreflex contribute importantly to cardiovascular control during exercise. Each of these inputs potentially mediates the heightened cardiovascular response to physical activity in hypertension. However, given that exercise pressor reflex overactivity is known to elicit enhanced circulatory responses to exercise in disease states closely related to hypertension (e.g. heart failure), we tested the hypothesis that the exaggerated cardiovascular response to exercise in hypertension is mediated by an overactive exercise pressor reflex. To test this hypothesis, we used a rat model of exercise recently developed in our laboratory that selectively stimulates the exercise pressor reflex independent of central command and/or the arterial baroreflex. Activation of the exercise pressor reflex during electrically induced static muscle contraction in the absence of input from central command resulted in significantly larger increases in MAP and HR in male spontaneously hypertensive rats as compared to normotensive Wistar-Kyoto rats over a wide range of exercise intensities. Similar findings were obtained in animals in which input from both central command and the arterial baroreflex were eliminated. These findings suggest that the enhanced cardiovascular response to exercise in hypertension is mediated by an overactive exercise pressor reflex. Potentially, effective treatment of exercise pressor reflex dysfunction may reduce the cardiovascular risks associated with exercise in hypertension.


A novel enterovirus and parechovirus multiplex one-step real-time PCR-validation and clinical experience.
As the number of new enteroviruses and human parechoviruses seems ever growing, the necessity for updated diagnostics is relevant. We have updated an enterovirus assay and combined it with a previously published assay for human parechovirus resulting in a multiplex one-step RT-PCR assay. The multiplex assay was validated by analysing the sensitivity and specificity of the assay compared to the respective monoplex assays, and a good concordance was found. Furthermore, the enterovirus assay was able to detect 42 reference strains from all 4 species, and an additional 9 genotypes during panel testing and routine usage. During 15 months of routine use, from October 2008 to December 2009, we received and analysed 2187 samples (stool samples, cerebrospinal fluids, blood samples, respiratory samples and autopsy samples) were tested, from 1546 patients and detected enteroviruses and parechoviruses in 171 (8%) and 66 (3%) of the samples, respectively. 180 of the positive samples could be genotyped by PCR and sequencing and the most common genotypes found were human parechovirus type 3, echovirus 9, enterovirus 71, Coxsackievirus A16, and echovirus 25. During 2009 in Denmark, both enterovirus and human parechovirus type 3 had a similar seasonal pattern with a peak during the summer and autumn. Human parechovirus type 3 was almost invariably found in children less than 4 months of age. In conclusion, a multiplex assay was developed allowing simultaneous detection of 2 viruses, which can cause similar clinical symptoms.