Consequences of ADHD medication use for children's outcomes.
This paper estimates effects of early ADHD medication use on key human capital outcomes for children diagnosed with ADHD while using rarely available register based data on diagnoses and prescription drug purchases. Our main identification strategy exploits plausible exogenous assignment of children to hospitals with specialist physicians, while our analysis of health outcomes also allows for an individual level panel data strategy. We find that the behavior of specialist physicians varies considerably across hospitals and that the prescribing behavior does affect the probability that a given child is treated. Results show that children diagnosed with ADHD in pharmacological treatment have fewer hospital contacts if treated and that treatment to some extent protects against criminal behavior.


Inhibition of Ebola virus entry by a C-peptide targeted to endosomes.
Ebola virus (EboV) and Marburg virus (MarV) (filoviruses) are the causative agents of severe hemorrhagic fever. Infection begins with uptake of particles into cellular endosomes, where the viral envelope glycoprotein (GP) catalyzes fusion between the viral and host cell membranes. This fusion event is thought to involve conformational rearrangements of the transmembrane subunit (GP2) of the envelope spike that ultimately result in formation of a six-helix bundle by the N- and C-terminal heptad repeat (NHR and CHR, respectively) regions of GP2. Infection by other viruses employing similar viral entry mechanisms (such as HIV-1 and severe acute respiratory syndrome coronavirus) can be inhibited with synthetic peptides corresponding to the native CHR sequence ("C-peptides"). However, previously reported EboV C-peptides have shown weak or insignificant antiviral activity. To determine whether the activity of a C-peptide could be improved by increasing its intracellular concentration, we prepared an EboV C-peptide conjugated to the arginine-rich sequence from HIV-1 Tat, which is known to accumulate in endosomes. We found that this peptide specifically inhibited viral entry mediated by filovirus GP proteins and infection by authentic filoviruses. We determined that antiviral activity was dependent on both the Tat sequence and the native EboV CHR sequence. Mechanistic studies suggested that the peptide acts by blocking a membrane fusion intermediate.


27-hydroxycholesterol is an endogenous selective estrogen receptor modulator.
Selective estrogen receptor (ER) modulators (SERMs) are ER ligands whose relative agonist/antagonist activities vary in a cell- and promoter-dependent manner. The molecular basis underlying this selectivity can be attributed to the ability of these ligands to induce distinct alterations in ER structure leading to differential recruitment of coactivators and corepressors. Whether SERM activity is restricted to synthetic ligands or whether molecules exist in vivo that function in an analogous manner remains unresolved. However, the recent observation that oxysterols bind ER and antagonize the actions of 17beta-estradiol (E2) on the vascular wall suggests that this class of ligands may possess SERM activity. We demonstrate here that 27-hydroxycholesterol (27HC), the most prevalent oxysterol in circulation, functions as a SERM, the efficacy of which varies when assessed on different endpoints. Importantly, 27HC positively regulates both gene transcription and cell proliferation in cellular models of breast cancer. Using combinatorial peptide phage display, we have determined that 27HC induces a unique conformational change in both ERalpha and ERbeta, distinguishing it from E2 and other SERMs. Thus, as with other ER ligands, it appears that the unique pharmacological activity of 27HC relates to its ability to impact ER structure and modulate cofactor recruitment. Cumulatively, these data indicate that 27HC is an endogenous SERM with partial agonist activity in breast cancer cells and suggest that it may influence the pathology of breast cancer. Moreover, given the product-precursor relationship between 27HC and cholesterol, our findings have implications with respect to breast cancer risk in obese/hypercholesteremic individuals.


Odon device for instrumental vaginal deliveries: results of a medical device pilot clinical study.
A prolonged and complicated second stage of labour is associated with serious perinatal complications. The Odon device is an innovation intended to perform instrumental vaginal delivery presently under development. We present an evaluation of the feasibility and safety of delivery with early prototypes of this device from an early terminated clinical study. Hospital-based, multi-phased, open-label, pilot clinical study with no control group in tertiary hospitals in Argentina and South Africa. Multiparous and nulliparous women, with uncomplicated singleton pregnancies, were enrolled during the third trimester of pregnancy. Delivery with Odon device was attempted under non-emergency conditions during the second stage of labour. The feasibility outcome was delivery with the Odon device defined as successful expulsion of the fetal head after one-time application of the device. Of the 49 women enrolled, the Odon device was inserted successfully in 46 (93%), and successful Odon device delivery as defined above was achieved in 35 (71%) women. Vaginal, first and second degree perineal tears occurred in 29 (59%) women. Four women had cervical tears. No third or fourth degree perineal tears were observed. All neonates were born alive and vigorous. No adverse maternal or infant outcomes were observed at 6-weeks follow-up for all dyads, and at 1 year for the first 30 dyads. Delivery using the Odon device is feasible. Observed genital tears could be due to the device or the process of delivery and assessment bias. Evaluating the effectiveness and safety of the further developed prototype of the BD Odon Device™ will require a randomized-controlled trial. ANZCTR ACTRN12613000141741 Registered 06 February 2013. Retrospectively registered.


Optoelectronic oscillator for a measurement of acoustic velocity in acousto-optic device.
We report a novel means of measuring the acoustic velocity based on a well-known acousto-optic interaction. With an acousto-optic modulator (AOM), we construct an optoelectronic oscillator (OEO) that can measure the acoustic velocity in the AOM directly. The free spectral range between the modes is a function of the total loop length of the OEO, which is mainly dependent on the propagation time of the acoustic wave through the AOM. By changing the propagation time, we measured the acoustic velocity from the variation of the free spectral range. The results are reported and compared with earlier results. This method is insensitive to the variation of the optical phase shift. In addition, the high frequency-stability and microwave spectral purity of the OEO allow reliable and precise measurements.


The MAP kinase MpkA controls cell wall integrity, oxidative stress response, gliotoxin production and iron adaptation in Aspergillus fumigatus.
The saprophytic fungus Aspergillus fumigatus is the most important air-borne fungal pathogen. The cell wall of A. fumigatus has been studied intensively as a potential target for development of effective antifungal agents. A major role in maintaining cell wall integrity is played by the mitogen-activated protein kinase (MAPK) MpkA. To gain a comprehensive insight into this central signal transduction pathway, we performed a transcriptome analysis of the ΔmpkA mutant under standard and cell wall stress conditions. Besides genes involved in cell wall remodelling, protection against ROS and secondary metabolism such as gliotoxin, pyomelanin and pseurotin A, also genes involved in siderophore biosynthesis were regulated by MpkA. Consistently, northern and western blot analyses indicated that iron starvation triggers phosphorylation and thus activation of MpkA. Furthermore, localization studies indicated that MpkA accumulates in the nucleus under iron depletion. Hence, we report the first connection between a MAPK pathway and siderophore biosynthesis. The measurement of amino acid pools and of the pools of polyamines indicated that arginine was continuously converted into ornithine to fuel the siderophore pool in the ΔmpkA mutant strain. Based on our data, we propose that MpkA fine-tunes the balance between stress response and energy consuming cellular processes.


Pex8p: an intraperoxisomal organizer of the peroxisomal import machinery.
Peroxisomes transport folded and oligomeric proteins across their membrane. Two cytosolic import receptors, Pex5p and Pex7p, along with approximately 12 membrane-bound peroxins participate in this process. While interactions among individual peroxins have been described, their organization into functional units has remained elusive. We have purified and defined two core complexes of the peroxisomal import machinery: the docking complex comprising Pex14p and Pex17p, with the loosely associated Pex13p, and the RING finger complex containing Pex2p, Pex10p, and Pex12p. Association of both complexes into a larger import complex requires Pex8p, an intraperoxisomal protein. We conclude that Pex8p organizes the formation of the larger import complex from the trans side of the peroxisomal membrane and thus might enable functional communication between both sides of the membrane.


Conformation-specific infrared and ultraviolet spectroscopy of tyrosine-based protonated dipeptides.
We present the spectroscopy and photofragmentation dynamics of two isomeric protonated dipeptides, H+AlaTyr and H+TyrAla, in a cold ion trap. By a combination of infrared-ultraviolet double resonance experiments and density functional theory calculations, we establish the conformations present at low temperature. Interaction of the charge at the N-terminus with the carbonyl group and the tyrosine pi-cloud seems to be critical in stabilizing the low-energy conformations. H+AlaTyr has the flexibility to allow a stronger interaction between the charge and the aromatic ring than in H+TyrAla, and this interaction may be responsible for many of the differences we observe in the former: a significant redshift in the ultraviolet spectrum, a much larger photofragmentation yield, fewer stable conformations, and the absence of fragmentation in excited electronic states.


Evaluation of a city-wide school-located influenza vaccination program in Oakland, California, with respect to vaccination coverage, school absences, and laboratory-confirmed influenza: A matched cohort study.
It is estimated that vaccinating 50%-70% of school-aged children for influenza can produce population-wide indirect effects. We evaluated a city-wide school-located influenza vaccination (SLIV) intervention that aimed to increase influenza vaccination coverage. The intervention was implemented in ≥95 preschools and elementary schools in northern California from 2014 to 2018. Using a matched cohort design, we estimated intervention impacts on student influenza vaccination coverage, school absenteeism, and community-wide indirect effects on laboratory-confirmed influenza hospitalizations. We used a multivariate matching algorithm to identify a nearby comparison school district with pre-intervention characteristics similar to those of the intervention school district and matched schools in each district. To measure student influenza vaccination, we conducted cross-sectional surveys of student caregivers in 22 school pairs (2017 survey, N = 6,070; 2018 survey, N = 6,507). We estimated the incidence of laboratory-confirmed influenza hospitalization from 2011 to 2018 using surveillance data from school district zip codes. We analyzed student absenteeism data from 2011 to 2018 from each district (N = 42,487,816 student-days). To account for pre-intervention differences between districts, we estimated difference-in-differences (DID) in influenza hospitalization incidence and absenteeism rates using generalized linear and log-linear models with a population offset for incidence outcomes. Prior to the SLIV intervention, the median household income was $51,849 in the intervention site and $61,596 in the comparison site. The population in each site was predominately white (41% in the intervention site, 48% in the comparison site) and/or of Hispanic or Latino ethnicity (26% in the intervention site, 33% in the comparison site). The number of students vaccinated by the SLIV intervention ranged from 7,502 to 10,106 (22%-28% of eligible students) each year. During the intervention, influenza vaccination coverage among elementary students was 53%-66% in the comparison district. Coverage was similar between the intervention and comparison districts in influenza seasons 2014-2015 and 2015-2016 and was significantly higher in the intervention site in seasons 2016-2017 (7%; 95% CI 4, 11; p < 0.001) and 2017-2018 (11%; 95% CI 7, 15; p < 0.001). During seasons when vaccination coverage was higher among intervention schools and the vaccine was moderately effective, there was evidence of statistically significant indirect effects: The DID in the incidence of influenza hospitalization per 100,000 in the intervention versus comparison site was -17 (95% CI -30, -4; p = 0.008) in 2016-2017 and -37 (95% CI -54, -19; p < 0.001) in 2017-2018 among non-elementary-school-aged individuals and -73 (95% CI -147, 1; p = 0.054) in 2016-2017 and -160 (95% CI -267, -53; p = 0.004) in 2017-2018 among adults 65 years or older. The DID in illness-related school absences per 100 school days during the influenza season was -0.63 (95% CI -1.14, -0.13; p = 0.014) in 2016-2017 and -0.80 (95% CI -1.28, -0.31; p = 0.001) in 2017-2018. Limitations of this study include the use of an observational design, which may be subject to unmeasured confounding, and caregiver-reported vaccination status, which is subject to poor recall and low response rates. A city-wide SLIV intervention in a large, diverse urban population was associated with a decrease in the incidence of laboratory-confirmed influenza hospitalization in all age groups and a decrease in illness-specific school absence rate among students in 2016-2017 and 2017-2018, seasons when the vaccine was moderately effective, suggesting that the intervention produced indirect effects. Our findings suggest that in populations with moderately high background levels of influenza vaccination coverage, SLIV programs are associated with further increases in coverage and reduced influenza across the community.


In vivo and in vitro efficacy of sainfoin (Onobrychis viciifolia) against Eimeria spp in lambs.
The effect of sainfoin (Onobrychis viciifolia) against ovine coccidia was evaluated in vivo and in vitro. In 3 in vivo trials weaned lambs were allocated into two treatment groups receiving diets with either lucerne (Medicago sativa) or sainfoin. During the trials, which lasted for 7 (trial 1) or 8 weeks (trials 2 and 3), oocysts per gram of faeces (OPGs), faecal scores and weight gain were recorded. In two of the experiments (trials 1 and 3) a reduction in the mean oocyst excretion rates was observed, starting three to four weeks after sainfoin hay feeding. This reduction ranged between 21.3% (trial 1) and 61.7% (trial 3) compared to the control values. As a result, a decrease in the total number of oocysts excreted (expressed as the mean area under the curve of the OPG) was observed from week 4 to the end of the two trials, respectively (trial 1: 42.6% reduction, p=0.05; trial 3: 52.4% reduction, p=0.06). The results did not show any significant diet effect on lamb growth rates and faecal scores. In the in vitro experiments the effect of 39 sainfoin extracts were tested in an oocyst sporulation inhibition assay. The Eimeria oocysts sporulation inhibition throughout the experiments did not exceed 10.7%, showing that extracts of this forages do not have a significant inhibitory effect on Eimeria oocyst sporulation. This was an initial attempt to investigate a possible anticoccidial effect of sainfoin and further studies are needed in order to better understand its mode of action against Eimeria.


TORC2 signaling is antagonized by protein phosphatase 2A and the Far complex in Saccharomyces cerevisiae.
The target of rapamycin (TOR) kinase, a central regulator of eukaryotic cell growth, exists in two essential, yet distinct, TOR kinase complexes in the budding yeast Saccharomyces cerevisiae: rapamycin-sensitive TORC1 and rapamycin-insensitive TORC2. Lst8, a component of both TOR complexes, is essential for cell viability. However, it is unclear whether the essential function of Lst8 is linked to TORC1, TORC2, or both. To that end, we carried out a genetic screen to isolate lst8 deletion suppressor mutants. Here we report that mutations in SAC7 and FAR11 suppress lethality of lst8Δ and TORC2-deficient (tor2-21) mutations but not TORC1 inactivation, suggesting that the essential function of Lst8 is linked only to TORC2. More importantly, characterization of lst8Δ bypass mutants reveals a role for protein phosphatase 2A (PP2A) in the regulation of TORC2 signaling. We show that Far11, a member of the Far3-7-8-9-10-11 complex involved in pheromone-induced cell cycle arrest, interacts with Tpd3 and Pph21, conserved components of PP2A, and deletions of components of the Far3-7-8-9-10-11 complex and PP2A rescue growth defects in lst8Δ and tor2-21 mutants. In addition, loss of the regulatory B' subunit of PP2A Rts1 or Far11 restores phosphorylation to the TORC2 substrate Slm1 in a tor2-21 mutant. Mammalian Far11 orthologs FAM40A/B exist in a complex with PP2A known as STRIPAK, suggesting a conserved functional association of PP2A and Far11. Antagonism of TORC2 signaling by PP2A-Far11 represents a novel regulatory mechanism for controlling spatial cell growth of yeast.


Characterisation of antithrombin-dependent anticoagulants through clot waveform analysis to potentially distinguish them from antithrombin-independent inhibitors targeting activated coagulation factors.
While antithrombin (AT)-independent inhibitors targeting thrombin or activated factor X have been assessed through clot waveform (CWA), there are no reports on assessment with respect to AT-dependent anticoagulants. The present study aims to characterise AT-dependent anticoagulants through CWA to distinguish them from AT-independent inhibitors. CWA was applied to the activated partial thromboplastin time (APTT) assay of plasma samples spiked with each of AT-dependent drugs (unfractionated heparin, enoxaparin and fondaparinux) and AT-independent drugs (rivaroxaban, apixaban, edoxaban, dabigatran, argatroban, hirudin and bivalirudin), which was performed using the CS-5100 or CN-6000 (Sysmex). The APTT-CWA data were automatically gained by the analyser program. The positive mode of clotting reaction curves was defined as the direction towards fibrin generation. Regarding dose-response curves in AT-dependent anticoagulants, the maximum positive values of the first and secondary derivatives (Max1 and Maxp2, respectively) and the maximum negative values of the secondary derivative (Maxn2) seemed to drop to zero without making an asymptotic line, consistent with the irreversibility. Such a feature was observed also in hirudin, as reported previously. Notably, the symmetric property of Max1 peaks in the waveforms was distorted dose dependently in AT independent but not AT-dependent drugs. A plot of Maxp2 logarithm versus Maxn2 logarithm was linear. The slope was about 1 in AT-dependent drugs while that was more than 1 in AT-independent drugs. These features made it possible to distinguish AT-dependent and AT-independent drugs. The results aid in further understanding of the pharmacological aspects of anticoagulation and in screening of candidates for novel anticoagulants.


Developmental dynamics of the neural crest-mesenchymal axis in creating the thymic microenvironment.
The thymic stroma is composed of epithelial and nonepithelial cells providing separate microenvironments controlling homing, differentiation, and selection of hematopoietic precursor cells to functional T cells. Here, we explore at single-cell resolution the complex composition and dynamic changes of the nonepithelial stromal compartment across different developmental stages in the human and mouse thymus, and in an experimental model of the DiGeorge syndrome, the most common form of human thymic hypoplasia. The detected gene expression signatures identify previously unknown stromal subtypes and relate their individual molecular profiles to separate differentiation trajectories and functions, revealing an unprecedented heterogeneity of different cell types that emerge at discrete developmental stages and vary in their expression of key regulatory signaling circuits and extracellular matrix components. Together, these findings highlight the dynamic complexity of the nonepithelial thymus stroma and link this to separate instructive roles essential for normal thymus organogenesis and tissue maintenance.


Refining the definition of mandibular osteoradionecrosis in clinical trials: The cancer research UK HOPON trial (Hyperbaric Oxygen for the Prevention of Osteoradionecrosis).
Mandibular osteoradionecrosis (ORN) is a common and serious complication of head and neck radiotherapy for which there is little reliable evidence for prevention or treatment. The diagnosis and classification of ORN have been inconsistently and imprecisely defined, even in clinical trials. A systematic review of diagnosis and classifications of ORN with specific focus on clinical trials is presented. The most suitable classification was evaluated for consistency using blinded independent review of outcome data (clinical photographs and radiographs) in the HOPON trial. Of 16 ORN classifications found, only one (Notani) appeared suitable as an endpoint in clinical trials. Clinical records of 217 timepoints were analysed amongst 94 randomised patients in the HOPON trial. The only inconsistency in classification arose where minor bone spicules (MBS) were apparent, which occurred in 19% of patients. Some trial investigators judged MBS as clinically unimportant and not reflecting ORN, others classified as ORN based on rigid definitions in common clinical use. When MBS was added as a distinct category to the Notani classification this ambiguity was resolved and agreement between observers was achieved. Most definitions and clinical classifications are based on retrospective case series and may be unsuitable for prospective interventional trials of ORN prevention or treatment. When ORN is used as a primary or secondary outcome in prospective clinical trials, the use of Notani classification with the additional category of MBS is recommended as it avoids subjectivity and enhances reliability and consistency of reporting.


Discovery and validation of a three-gene signature to distinguish COVID-19 and other viral infections in emergency infectious disease presentations: a case-control and observational cohort study.
Emergency admissions for infection often lack initial diagnostic certainty. COVID-19 has highlighted a need for novel diagnostic approaches to indicate likelihood of viral infection in a pandemic setting. We aimed to derive and validate a blood transcriptional signature to detect viral infections, including COVID-19, among adults with suspected infection who presented to the emergency department. Individuals (aged ≥18 years) presenting with suspected infection to an emergency department at a major teaching hospital in the UK were prospectively recruited as part of the Bioresource in Adult Infectious Diseases (BioAID) discovery cohort. Whole-blood RNA sequencing was done on samples from participants with subsequently confirmed viral, bacterial, or no infection diagnoses. Differentially expressed host genes that met additional filtering criteria were subjected to feature selection to derive the most parsimonious discriminating signature. We validated the signature via RT-qPCR in a prospective validation cohort of participants who presented to an emergency department with undifferentiated fever, and a second case-control validation cohort of emergency department participants with PCR-positive COVID-19 or bacterial infection. We assessed signature performance by calculating the area under receiver operating characteristic curves (AUROCs), sensitivities, and specificities. A three-gene transcript signature, comprising HERC6, IGF1R, and NAGK, was derived from the discovery cohort of 56 participants with bacterial infections and 27 with viral infections. In the validation cohort of 200 participants, the signature differentiated bacterial from viral infections with an AUROC of 0·976 (95% CI 0·919-1·000), sensitivity of 97·3% (85·8-99·9), and specificity of 100% (63·1-100). The AUROC for C-reactive protein (CRP) was 0·833 (0·694-0·944) and for leukocyte count was 0·938 (0·840-0·986). The signature achieved higher net benefit in decision curve analysis than either CRP or leukocyte count for discriminating viral infections from all other infections. In the second validation analysis, which included SARS-CoV-2-positive participants, the signature discriminated 35 bacterial infections from 34 SARS-CoV-2-positive COVID-19 infections with AUROC of 0·953 (0·893-0·992), sensitivity 88·6%, and specificity of 94·1%. This novel three-gene signature discriminates viral infections, including COVID-19, from other emergency infection presentations in adults, outperforming both leukocyte count and CRP, thus potentially providing substantial clinical utility in managing acute presentations with infection. National Institute for Health Research, Medical Research Council, Wellcome Trust, and EU-FP7.


Evolutionary responses to climate change in a range expanding plant.
To understand the biological effects of climate change, it is essential to take into account species' evolutionary responses to their changing environments. Ongoing climate change is resulting in species shifting their geographical distribution ranges poleward. We tested whether a successful range expanding plant has rapidly adapted to the regional conditions in its novel range, and whether adaptation could be driven by herbivores. Furthermore, we investigated if enemy release occurred in the newly colonized areas and whether plant origins differed in herbivore resistance. Plants were cloned and reciprocally transplanted between three experimental sites across the range. Effects of herbivores on plant performance were tested by individually caging plants with either open or closed cages. There was no indication of (regional) adaptation to abiotic conditions. Plants originating from the novel range were always larger than plants from the core distribution at all experimental sites, with or without herbivory. Herbivore damage was highest and not lowest at the experimental sites in the novel range, suggesting no release from enemy impact. Genotypes from the core were more damaged compared to genotypes from newly colonized areas at the most northern site in the novel range, which was dominated by generalist slug herbivory. We also detected subtle shifts in chemical defenses between the plant origins. Genotypes from the novel range had more inducible defenses. Our results suggest that plants that are expanding their range with climate change may evolve increased vigor and altered herbivore resistance in their new range, analogous to invasive plants.


Pain as an indication for rib fixation: a bi-institutional pilot study.
In trauma patients, open reduction and internal fixation of rib fractures remain controversial. We hypothesized that patients who have open reduction and internal fixation of rib fractures would experience less pain compared with controls and thus require fewer opiates. Further, we hypothesized that improved pain control would result in fewer pulmonary complications and decreased length of stay. This is a retrospective bi-institutional matched case-control study. Cases were matched 1:2 by age, injury severity Score, chest abbreviated injury severity score, head abbreviated injury severity score, pulmonary contusion score, and number of fractured ribs. The daily total doses of analgesic drugs were converted to equianalgesic intravenous morphine doses, and the primary outcome was inpatient narcotic administration. Sixteen patients between July 2005 and June 2009 underwent rib fixation in 5 ± 3 days after injury using an average of 3 (1-5) metallic plates. Morphine requirements decreased from 110 mg ± 98 mg preoperatively to 63 ± 57 mg postoperatively (p = 0.01). There were no significant differences between cases and controls in the mean morphine dose (79 ± 63 vs. 76 ± 55 mg, p = 0.65), hospital stay (18 ± 12 vs. 16 ± 11 days, p = 0.67), intensive care unit stay (9 ± 8 vs. 7 ± 10 days, p = 0.75), ventilation days (7 ± 8 vs. 6 ± 10, p = 0.44), and pneumonia rates (31% vs. 38%, p = 0.76). The need for analgesia was significantly reduced after rib fixation in patients with multiple rib fractures. However, no difference in outcomes was observed when these patients were compared with matched controls in this pilot study. Further study is required to investigate these preliminary findings.


Host genetic control of natural killer cell diversity revealed in the Collaborative Cross.
Natural killer (NK) cells are innate effectors armed with cytotoxic and cytokine-secreting capacities whose spontaneous antitumor activity is key to numerous immunotherapeutic strategies. However, current mouse models fail to mirror the extensive immune system variation that exists in the human population which may impact on NK cell-based therapies. We performed a comprehensive profiling of NK cells in the Collaborative Cross (CC), a collection of novel recombinant inbred mouse strains whose genetic diversity matches that of humans, thereby providing a unique and highly diverse small animal model for the study of immune variation. We demonstrate that NK cells from CC strains displayed a breadth of phenotypic and functional variation reminiscent of that reported for humans with regards to cell numbers, key marker expression, and functional capacities. We took advantage of the vast genetic diversity of the CC and identified nine genomic loci through quantitative trait locus mapping driving these phenotypic variations. SNP haplotype patterns and variant effect analyses identified candidate genes associated with lung NK cell numbers, frequencies of CD94+ NK cells, and expression levels of NKp46. Thus, we demonstrate that the CC represents an outstanding resource to study NK cell diversity and its regulation by host genetics.


Health outcomes during the 2008 financial crisis in Europe: systematic literature review.
To systematically identify, critically appraise, and synthesise empirical studies about the impact of the 2008 financial crisis in Europe on health outcomes. Systematic literature review. Structural searches of key databases, healthcare journals, and organisation based websites. Empirical studies reporting on the impact of the financial crisis on health outcomes in Europe, published from January 2008 to December 2015, were included. All selected studies were assessed for risk of bias. Owing to the heterogeneity of studies in terms of study design and analysis and the use of overlapping datasets across studies, studies were analysed thematically per outcome, and the evidence was synthesised on different health outcomes without formal meta-analysis. 41 studies met the inclusion criteria, and focused on suicide, mental health, self rated health, mortality, and other health outcomes. Of those studies, 30 (73%) were deemed to be at high risk of bias, nine (22%) at moderate risk of bias, and only two (5%) at low risk of bias, limiting the conclusions that can be drawn. Although there were differences across countries and groups, there was some indication that suicides increased and mental health deteriorated during the crisis. The crisis did not seem to reverse the trend of decreasing overall mortality. Evidence on self rated health and other indicators was mixed. Most published studies on the impact of financial crisis on health in Europe had a substantial risk of bias; therefore, results need to be cautiously interpreted. Overall, the financial crisis in Europe seemed to have had heterogeneous effects on health outcomes, with the evidence being most consistent for suicides and mental health. There is a need for better empirical studies, especially those focused on identifying mechanisms that can mitigate the adverse effects of the crisis.


Epidermal growth factor receptor (EGFR) regulates mechanical ventilation-induced lung injury in mice.
Mechanical ventilation (MV) is used as therapy to support critically ill patients; however, the mechanisms by which MV induces lung injury and inflammation remain unclear. Epidermal growth factor receptor (EGFR)-mediated signaling plays a key role in various physiologic and pathologic processes, which include those modulated by mechanical and shear forces, in various cell types. We hypothesized that EGFR-activated signaling plays a key role in ventilator-induced lung injury and inflammation (VILI). To test this hypothesis, we assessed lung vascular and alveolar permeability as well as inflammation, which are cardinal features of VILI, in mice treated with the EGFR inhibitor AG1478. Inhibition of EGFR activity greatly diminished MV-induced lung alveolar permeability and neutrophil accumulation in the bronchoalveolar lavage (BAL) fluid, as compared with vehicle-treated controls. Similarly, AG1478 inhibition diminished lung vascular leak (as assessed by Evans blue extravasation), but it did not affect interstitial neutrophil accumulation. Inhibition of the EGFR pathway also blocked expression of genes induced by MV. However, intratracheal instillation of EGF alone failed to induce lung injury. Collectively, our findings suggest that EGFR-activated signaling is necessary but not sufficient to produce acute lung injury in mice.


Albumin, a new biomarker of organophosphorus toxicant exposure, identified by mass spectrometry.
The classical laboratory tests for exposure to organophosphorus toxicants (OP) are inhibition of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) activity in blood. In a search for new biomarkers of OP exposure, we treated mice with a biotinylated organophosphorus agent, FP-biotin. The biotinylated proteins in muscle were purified by binding to avidin-Sepharose, separated by gel electrophoresis, digested with trypsin, and identified from their fragmentation patterns on a quadrupole time-of-flight mass spectrometer. Albumin and ES1 carboxylesterase (EC 126.96.36.199) were found to be major targets of FP-biotin. These FP-biotinylated proteins were also identified in mouse plasma by comparing band patterns on nondenaturing gels stained for albumin and carboxylesterase activity, with band patterns on blots hybridized with Streptavidin Alexa-680. Two additional FP-biotin targets, AChE (EC 18.104.22.168) and BChE (EC 22.214.171.124), were identified in mouse plasma by finding that enzyme activity was inhibited 50-80%. Mouse plasma contained eight additional FP-biotinylated bands whose identity has not yet been determined. In vitro experiments with human plasma showed that chlorpyrifos oxon, echothiophate, malaoxon, paraoxon, methyl paraoxon, diazoxon, diisopropylfluorophosphate, and dichlorvos competed with FP-biotin for binding to human albumin. Though experiments with purified albumin have previously shown that albumin covalently binds OP, this is the first report of OP binding to albumin in a living animal. Carboxylesterase is not a biomarker in man because humans have no carboxylesterase in blood. It is concluded that OP bound to albumin could serve as a new biomarker of OP exposure in man.


High Royal Jelly-Producing Honeybees (Apis mellifera ligustica) (Hymenoptera: Apidae) in China.
China is the largest producer and exporter of royal jelly (RJ) in the world, supplying >90% of the global market. The high production of RJ in China is principally owing to the high RJ-producing lineage of honeybees (Apis mellifera ligustica Spinola, 1806) established by beekeepers in the 1980s. We describe the development of high royal jelly-producing honeybees and the management of this lineage today. Previous research and recent advances in the genetic characterization of this lineage, and the molecular markers and mechanisms associated with high RJ production are summarized. The gaps in our knowledge and prospects for future research are also highlighted.


Salt tolerance conferred by overexpression of a vacuolar Na+/H+ antiport in Arabidopsis.
Agricultural productivity is severely affected by soil salinity. One possible mechanism by which plants could survive salt stress is to compartmentalize sodium ions away from the cytosol. Overexpression of a vacuolar Na+/H+ antiport from Arabidopsis thaliana in Arabidopsis plants promotes sustained growth and development in soil watered with up to 200 millimolar sodium chloride. This salinity tolerance was correlated with higher-than-normal levels of AtNHX1 transcripts, protein, and vacuolar Na+/H+ (sodium/proton) antiport activity. These results demonstrate the feasibility of engineering salt tolerance in plants.


Negative play contagion in calves.
Play is a strong outwardly directed, emotional behaviour and can contagiously spread between individuals. It has been suggested that high-playing animals could 'seed' play in others, spreading positive affective states. Despite the current interest in play contagion there has been no previous attempt to measure the strength of the play contagion effect. The calf (Bos taurus) is ideal for testing the strength of play contagion as play in calves is strongly related to energy intake from milk. We manipulated play in calves through their milk allowances and housed the calves in uniform groups all on the same milk allowance (high = UHigh or low = ULow) or in mixed groups with calves in the same group receiving either a high (= MHigh) or low (= MLow) milk allowance. We measured locomotor play using accelerometers on two consecutive days when calves were four and eight weeks old, in order to study play contagion over a protracted developmental window. We anticipated that differences in the level of play contagion between treatment groups would result in difference in the play levels observed in the MLow and ULow individuals. Contrary to our expectations we found that spontaneous play was suppressed in the high-milk calves housed in mixed groups (MHigh), in comparison to calves housed with group mates all receiving high-milk (UHigh). These results are the first to quantify a negative play contagion effect, particularly in a situation of long-term contact, and may suggest that negative contagion has a stronger effect on play behaviour than positive contagion.


Lifetime cost-effectiveness simulation of once-weekly exenatide in type 2 diabetes: A cost-utility analysis based on the EXSCEL trial.
The Exenatide Study of Cardiovascular Event Lowering (EXSCEL) trial assessed once-weekly exenatide (EQW) vs. placebo, added to usual care in 14,752 patients with type 2 diabetes mellitus (Clinicaltrials.gov: NCT01144338). We assessed the lifetime cost-effectiveness of adding EQW vs. usual care alone from a healthcare perspective. Medical resource use and EQ-5D utilities were collected throughout the study. Within-trial results were extrapolated to a lifetime horizon using the UK Prospective Diabetes Study Outcomes Model version 2 (UKPDS-OM2), predicting predict cardiovascular and microvascular events. Cost-effectiveness was evaluated separately for US and UK settings, with outcomes measured in quality-adjusted life-years (QALYs). EQW plus usual care gained 0.162 QALYs at an additional cost of $41,545/patient, compared with usual care in a US setting. The incremental cost-effectiveness ratio (ICER) was $259,223/QALY. In a UK setting, the QALY gain was 0.151 at an additional cost of £6357: an ICER of £42,589/QALY. Sensitivity analyses ranged between $34,369-$269,571 and £3430-£46,560 per QALY gained. In a lifetime extrapolation, adding EQW to usual care increased QALYs and costs compared with usual care alone. The base-case ICERs exceeded the commonly-cited cost-effectiveness thresholds of $100,000/QALY and £20,000/QALY. However, ICERs were considerably lower in some subgroups, and in sensitivity analyses.


Achieving benefit for patients in primary care informatics: the report of a international consensus workshop at Medinfo 2007.
Landmark reports suggest that sharing health data between clinical computer systems should improve patient safety and the quality of care. Enhancing the use of informatics in primary care is usually a key part of these strategies. To synthesise the learning from the international use of informatics in primary care. The workshop was attended by 21 delegates drawn from all continents. There were presentations from USA, UK and the Netherlands, and informal updates from Australia, Argentina, and Sweden and the Nordic countries. These presentations were discussed in a workshop setting to identify common issues. Key principles were synthesised through a post-workshop analysis and then sorted into themes. Themes emerged about the deployment of informatics which can be applied at health service, practice and individual clinical consultation level: 1 At the health service or provider level, success appeared proportional to the extent of collaboration between a broad range of stakeholders and identification of leaders. 2 Within the practice much is currently being achieved with legacy computer systems and apparently outdated coding systems. This includes prescribing safety alerts, clinical audit and promoting computer data recording and quality. 3 In the consultation the computer is a 'big player' and may make traditional models of the consultation redundant. We should make more efforts to share learning; develop clear internationally acceptable definitions; highlight gaps between pockets of excellence and real-world practice, and most importantly suggest how they might be bridged. Knowledge synthesis from different health systems may provide a greater understanding of how the third actor (the computer) is best used in primary care.


Characterization of 12 silent alleles of the human butyrylcholinesterase (BCHE) gene.
The silent phenotype of human butyrylcholinesterase (BChE), present in most human populations in frequencies of approximately 1/100,000, is characterized by the complete absence of BChE activity or by activity <10% of the average levels of the usual phenotype. Heterogeneity in this phenotype has been well established at the phenotypic level, but only a few silent BCHE alleles have been characterized at the DNA level. Twelve silent alleles of the human butyrylcholinesterase gene (BCHE) have been identified in 17 apparently unrelated patients who were selected by their increased sensitivity to the muscle relaxant succinylcholine. All of these alleles are characterized by single nucleotide substitutions or deletions leading to distinct changes in the structure of the BChE enzyme molecule. Nine of the nucleotide substitutions result in the replacement of single amino acid residues. Three of these variants, BCHE*33C, BCHE*198G, and BCHE*201T, produce normal amounts of immunoreactive but enzymatically inactive BChE protein in the plasma. The other six amino acid substitutions, encoded by BCHE*37S, BCHE*125F, BCHE*170E, BCHE*471R, and BCHE*518L, seem to cause reduced expression of BChE protein, and their role in determining the silent phenotype was confirmed by expression in cell culture. The other four silent alleles, BCHE*271STOP, BCHE*500STOP, BCHE*FS6, and BCHE*I2E3-8G, encode BChES truncated at their C-terminus because of premature stop codons caused by nucleotide substitutions, a frame shift, or altered splicing. The large number of different silent BCHE alleles found within a relatively small number of patients shows that the heterogeneity of the silent BChE phenotype is high. The characterization of silent BChE variants will be useful in the study of the structure/function relationship for this and other closely related enzymes.


Cellular mechanisms governing synaptic development in Drosophila melanogaster.
The neuromuscular connections of Drosophila are ideally suited for studying synaptic function and development. Hypotheses about cell recognition can be tested in a simple array of pre- and postsynaptic elements. Drosophila muscle fibers are multiply innervated by individually identifiable motoneurons. The neurons express several synaptic cotransmitters, including glutamate, proctolin, and octopamine, and are specialized by their synaptic morphology, neurotransmitters, and connectivity. During larval development the initial motoneuron endings grow extensively over the surface of the muscle fibers, and differentiate synaptic boutons of characteristic morphology. While considerable growth occurs postembryonically, the initial wiring of motoneurons to muscle fibers is accomplished during mid-to-late embryogenesis (stages 15-17). Efferent growth cones sample multiple muscle fibers with rapidly moving filopodia. Upon reaching their target muscle fibers, the growth cones rapidly differentiate into synaptic contacts whose morphology prefigures that of the larval junction. Mismatch experiments show that growth cones recognize specific muscle fibers, and can do so when the surrounding musculature is radically altered. However, when denied their normal targets, motoneurons can establish functional synapses on alternate muscle fibers. Blocking synaptic activity with either injected toxins or ion channel mutants does not derange synaptogenesis, but may influence the number of motor ending processes. The molecular mechanisms governing cellular recognition during synaptogenesis remain to be identified. However, several cell surface glycoproteins known to mediate cellular adhesion events in vitro are expressed by the developing synapses. Furthermore, enhancer detector lines have identified genes with expression restricted to small subsets of muscle fibers and/or motoneurons during the period of synaptogenesis. These observations suggest that in Drosophila a mechanism of target chemoaffinity may be involved in the genesis of stereotypic synaptic wiring.


Exploring walking path quality as a factor for urban elementary school children's active transport to school.
Path quality has not been well studied as a correlate of active transport to school. We hypothesize that for urban-dwelling children the environment between home and school is at least as important as the environment immediately surrounding their homes and/or schools when exploring walking to school behavior. Tools from spatial statistics and geographic information systems (GIS) were applied to an assessment of street blocks to create a walking path quality measure based on physical and social disorder (termed "incivilities") for each child. Path quality was included in a multivariate regression analysis of walking to school status for a sample of 362 children. The odds of walking to school for path quality was 0.88 (95% CI: 0.72-1.07), which although not statistically significant is in the direction supporting our hypothesis. The odds of walking to school for home street block incivility suggests the counter intuitive effect (OR = 1.10, 95% CI: 1.08-1.19). Results suggest that urban children living in communities characterized by higher incivilities are more likely to walk to school, potentially placing them at risk for adverse health outcomes because of exposure to high incivility areas along their route. Results also support the importance of including path quality when exploring the influence of the environment on walking to school behavior.


The epidemiology of HIV infection in Zambia.
Population surveys of health and fertility are an important source of information about demographic trends and their likely impact on the HIV/AIDS epidemic. In contrast to groups sampled at health facilities they can provide nationally and regionally representative estimates of a range of variables. Data on HIV-sero-status were collected in the 2001 Zambia Demographic and Health Survey (ZDHS) and made available in a separate data file in which HIV status was linked to a very limited set of demographic variables. We utilized this data set to examine associations between HIV prevalence, gender, age and geographical location. We applied the generalized geo-additive semi-parametric model as an alternative to the common linear model, in the context of analyzing the prevalence of HIV infection. This model enabled us to account for spatial auto-correlation, non-linear, location effects on the prevalence of HIV infection at the disaggregated provincial level (nine provinces) and assess temporal and geographical variation in the prevalence of HIV infection, while simultaneously controlling for important risk factors. Of the overall sample of 3950, 54% was female. The overall HIV-positivity rate was 565 (14.3%). The mean age at HIV diagnosis for male was 30.3 (SD=11.2) and 27.7 (SD=9.3) for female respectively. Lusaka and Copperbelt have the first and second highest prevalence of AIDS/HIV (marginal odds ratios of 3.24 and 2.88, respectively) but when the younger age of the urban population and the spatial auto-correlation was taken into account, Lusaka and Copperbelt were no longer among the areas with the highest prevalence. Non-linear effects of age at HIV diagnosis are also discussed and the importance of spatial residual effects and control of confounders on the prevalence of HIV infection. The study was conducted to assess the spatial pattern and the effect of confounding risk factors on AIDS/HIV prevalence and to develop a means of adjusting estimates of AIDS/HIV prevalence on the important risk factors. Controlling for important risk factors, such as geographical location (spatial auto-correlation), age structure of the population and gender, gave estimates of prevalence that are statistically robust. Researchers should be encouraged to use all available information in the data to account for important risk factors when reporting AIDS/HIV prevalence. Where this is not possible, correction factors should be applied, particularly where estimates of AIDS/HIV prevalence are pooled in systematic reviews. Our maps can be used for policy planning and management of AIDS/HIV in Zambia.


Noninvasive determination of the optical properties of two-layered turbid media.
Light propagation in two-layered turbid media having an infinitely thick second layer is investigated in the steady-state, frequency, and time domains. A solution of the diffusion approximation to the transport equation is derived by employing the extrapolated boundary condition. We compare the reflectance calculated from this solution with that computed with Monte Carlo simulations and show good agreement. To investigate if it is possible to determine the optical coefficients of the two layers and the thickness of the first layer, the solution of the diffusion equation is fitted to reflectance data obtained from both the diffusion equation and the Monte Carlo simulations. Although it is found that it is, in principle, possible to derive the optical coefficients of the two layers and the thickness of the first layer, we concentrate on the determination of the optical coefficients, knowing the thickness of the first layer. In the frequency domain, for example, it is shown that it is sufficient to make relative measurements of the phase and the steady-state reflectance at three distances from the illumination point to obtain useful estimates of the optical coefficients. Measurements of the absolute steady-state spatially resolved reflectance performed on two-layered solid phantoms confirm the theoretical results.


Complete genome sequence of Fer-de-Lance virus reveals a novel gene in reptilian paramyxoviruses.
The complete RNA genome sequence of the archetype reptilian paramyxovirus, Fer-de-Lance virus (FDLV), has been determined. The genome is 15,378 nucleotides in length and consists of seven nonoverlapping genes in the order 3' N-U-P-M-F-HN-L 5', coding for the nucleocapsid, unknown, phospho-, matrix, fusion, hemagglutinin-neuraminidase, and large polymerase proteins, respectively. The gene junctions contain highly conserved transcription start and stop signal sequences and tri-nucleotide intergenic regions similar to those of other Paramyxoviridae. The FDLV P gene expression strategy is like that of rubulaviruses, which express the accessory V protein from the primary transcript and edit a portion of the mRNA to encode P and I proteins. There is also an overlapping open reading frame potentially encoding a small basic protein in the P gene. The gene designated U (unknown), encodes a deduced protein of 19.4 kDa that has no counterpart in other paramyxoviruses and has no similarity with sequences in the National Center for Biotechnology Information database. Active transcription of the U gene in infected cells was demonstrated by Northern blot analysis, and bicistronic N-U mRNA was also evident. The genomes of two other snake paramyxovirus genotypes were also found to have U genes, with 11 to 16% nucleotide divergence from the FDLV U gene. Pairwise comparisons of amino acid identities and phylogenetic analyses of all deduced FDLV protein sequences with homologous sequences from other Paramyxoviridae indicate that FDLV represents a new genus within the subfamily Paramyxoviridae. We suggest the name Ferlavirus for the new genus, with FDLV as the type species.


Common complaints, difficult diagnosis: multiple myeloma.
To review the presenting signs and symptoms of multiple myeloma, its pathophysiology, diagnostic evaluation, and treatment options. A literature review of research articles and publications by oncology experts who specialize in multiple myeloma, supplemented by a case study. Multiple myeloma is a cancer of the geriatric population, with the average age at diagnosis between 65 and 68 years. As the population of those over age 65 is predicted to double by the year 2050, the incidence of myeloma is expected to increase. Nurse practitioners (NPs) must become familiar with the signs, symptoms, and complications of myeloma for patients to be diagnosed and referred to specialists in a timely manner. Patients with multiple myeloma often present with vague, common symptoms such as back pain, bony pain, fatigue, and anemia. These symptoms may be treated as separate medical conditions if NPs fail to include multiple myeloma in their differential diagnosis. If NPs are educated on this malignancy, they will have the expertise to look for other signs of the disease such as hypercalcemia, pathological fractures, osteopenia, or renal failure. Without early recognition of multiple myeloma and referrals to oncology specialists, patients are left with a delayed diagnosis and poor symptom control.


Functional organization of human nuclei.
1. Despite much progress in deciphering nuclear functions at the molecular level, our understanding of how these processes occur in vivo has been limited by the technologies presently available. I have used and developed a permeabilized cell system that retains most of the RNA- and DNA-polymerizing activities of HeLa cells. 2. Focal sites of transcription were visualized after incubation with bromouridine-UTP and immunolabelling with an antibody that reacts with bromouridine-RNA. Focal sites of replication were directly visualized by incubation with fluorochrome-dUTP conjugates. Approximately 300 transcription and 150 replication fluorescent foci were visualized in human cells. Foci resisted nucleolytic removal of 90% of chromatin. Experiments using laser scanning confocal microscopy show co-localization of sites of transcription with both splicing and replication sites, the latter particularly at the onset of S phase. Sites of replication were localized to discrete ovoid bodies when chromatin-depleted nuclei were visualized by thick section (resinless) electron microscopy. 3. These results suggest that active polymerases are focally concentrated (approximately 40 per focus) in 'factories' within nuclei. This higher-order organization may be important for both the initiation of replication and transcription in vivo.


Leveraging European infrastructures to access 1 million human genomes by 2022.
Human genomics is undergoing a step change from being a predominantly research-driven activity to one driven through health care as many countries in Europe now have nascent precision medicine programmes. To maximize the value of the genomic data generated, these data will need to be shared between institutions and across countries. In recognition of this challenge, 21 European countries recently signed a declaration to transnationally share data on at least 1 million human genomes by 2022. In this Roadmap, we identify the challenges of data sharing across borders and demonstrate that European research infrastructures are well-positioned to support the rapid implementation of widespread genomic data access.


A preliminary report on stem cell therapy for neuropathic pain in humans.
Mesenchymal stem cells (MSCs) have been shown in animal models to attenuate chronic neuropathic pain. This preliminary study investigated if: i) injections of autologous MSCs can reduce human neuropathic pain and ii) evaluate the safety of the procedure. Ten subjects with symptoms of neuropathic trigeminal pain underwent liposuction. The lipoaspirate was digested with collagenase and washed with saline three times. Following centrifugation, the stromal vascular fraction was resuspended in saline, and then transferred to syringes for local injections into the pain fields. Outcome measures at 6 months assessed reduction in: i) pain intensity measured by standard numerical rating scale from 0-10 and ii) daily dosage requirements of antineuropathic pain medication. Subjects were all female (mean age 55.3 years ± standard deviation [SD] 14.67; range 27-80 years) with pain symptoms lasting from 4 months to 6 years and 5 months. Lipoaspirate collection ranged from 102-214 g with total cell numbers injected from 33 million to 162 million cells. Cell viability was 62%-91%. There were no systemic or local tissue side effects from the stem cell therapy (n=41 oral and facial injection sites). Clinical pain outcomes showed that at 6 months, 5/9 subjects had reduced both pain intensity scores and use of antineuropathic medication. The mean pain score pre-treatment was 7.5 (SD 1.58) and at 6 months had decreased to 4.3 (SD 3.28), P=0.018, Wilcoxon signed-rank test. Antineuropathic pain medication use showed 5/9 subjects reduced their need for medication (gabapentin, P=0.053, Student's t-test). This preliminary open-labeled study showed autologous administration of stem cells for neuropathic trigeminal pain significantly reduced pain intensity at 6 months and is a safe and well tolerated intervention.


The psychosocial impact of food allergy and food hypersensitivity in children, adolescents and their families: a review.
Food allergy affects 6% of children but there is no cure, and strict avoidance of index allergens along with immediate access to rescue medication is the current best management. With specialist care, morbidity from food allergy in children is generally low, and mortality is very rare. However, there is strong evidence that food allergy and food hypersensitivity has an impact on psychological distress and on the quality of life (QoL) of children and adolescents, as well as their families. Until recently, the measurement of QoL in allergic children has proved difficult because of the lack of investigative tools available. New instruments for assessing QoL in food allergic children have recently been developed and validated, which should provide further insights into the problems these children encounter and will enable us to measure the effects of interventions in patients. This review examines the published impact of food allergy on affected children, adolescents and their families. It considers influences such as gender, age, disease severity, co-existing allergies and external influences, and examines how these may impact on allergy-related QoL and psychological distress including anxiety and depression. Implications of the impact are considered alongside avenues for future research.


Differential mechanisms underlie trace and delay conditioning in Drosophila.
Two forms of associative learning-delay conditioning and trace conditioning-have been widely investigated in humans and higher-order mammals1. In delay conditioning, an unconditioned stimulus (for example, an electric shock) is introduced in the final moments of a conditioned stimulus (for example, a tone), with both ending at the same time. In trace conditioning, a 'trace' interval separates the conditioned stimulus and the unconditioned stimulus. Trace conditioning therefore relies on maintaining a neural representation of the conditioned stimulus after its termination (hence making distraction possible2), to learn the conditioned stimulus-unconditioned stimulus contingency3; this makes it more cognitively demanding than delay conditioning4. Here, by combining virtual-reality behaviour with neurogenetic manipulations and in vivo two-photon brain imaging, we show that visual trace conditioning and delay conditioning in Drosophila mobilize R2 and R4m ring neurons in the ellipsoid body. In trace conditioning, calcium transients during the trace interval show increased oscillations and slower declines over repeated training, and both of these effects are sensitive to distractions. Dopaminergic activity accompanies signal persistence in ring neurons, and this is decreased by distractions solely during trace conditioning. Finally, dopamine D1-like and D2-like receptor signalling in ring neurons have different roles in delay and trace conditioning; dopamine D1-like receptor 1 mediates both forms of conditioning, whereas the dopamine D2-like receptor is involved exclusively in sustaining ring neuron activity during the trace interval of trace conditioning. These observations are similar to those previously reported in mammals during arousal5, prefrontal activation6 and high-level cognitive learning7,8.


Qualitative research in health care: II. A structured review and evaluation of studies.
A hand search of the original papers in seven medical journals over 5 years was conducted in order to identify those reporting qualitative research. A total of 210 papers were initially identified, of which 70 used qualitative methods of both data collection and analysis. These papers were evaluated by the researchers using a checklist which specified the criteria of good practice. Overall, 2% of the original papers published in the journals reported qualitative studies. Papers were more frequently positively assessed in terms of having clear aims, reporting research for which a qualitative approach was appropriate and describing their methods of data collection. Papers were less frequently positively assessed in relation to issued of data analysis such as validity, reliability and providing representative supporting evidence. It is concluded that the full potential of qualitative research has yet to be realized in the field of health care.


Transmetallation from CCC-NHC pincer Zr complexes in the synthesis of air-stable CCC-NHC pincer Co(iii) complexes and initial hydroboration trials.
Development of CCC-NHC pincer Co complexes via transmetalation from Zr is reported. Formation of these air-stable Co(iii) complexes was achieved through use of a CoCl2 or Co(acac)3in situ or with a discrete CCC-NHC pincer Zr transmetallating agent. Preliminary activity in the hydroboration of styrene is reported. This facile methodology will further the development of CCC-NHC pincer first-row transition metal complexes.


Movement Disorder Society Task Force report on the Hoehn and Yahr staging scale: status and recommendations.
The Movement Disorder Society Task Force for Rating Scales for Parkinson's disease (PD) prepared a critique of the Hoehn and Yahr scale (HY). Strengths of the HY scale include its wide utilization and acceptance. Progressively higher stages correlate with neuroimaging studies of dopaminergic loss, and high correlations exist between the HY scale and some standardized scales of motor impairment, disability, and quality of life. Weaknesses include the scale's mixing of impairment and disability and its non-linearity. Because the HY scale is weighted heavily toward postural instability as the primary index of disease severity, it does not capture completely impairments or disability from other motor features of PD and gives no information on nonmotor problems. Direct clinimetric testing of the HY scale has been very limited, but the scale fulfills at least some criteria for reliability and validity, especially for the midranges of the scale (Stages 2-4). Although a "modified HY scale" that includes 0.5 increments has been adopted widely, no clinimetric data are available on this adaptation. The Task Force recommends that: (1) the HY scale be used in its original form for demographic presentation of patient groups; (2) when the HY scale is used for group description, medians and ranges should be reported and analysis of changes should use nonparametric methods; (3) in research settings, the HY scale is useful primarily for defining inclusion/exclusion criteria; (4) to retain simplicity, clinicians should "rate what you see" and therefore incorporate comorbidities when assigning a HY stage; and (5) because of the wide usage of the modified HY scale with 0.5 increments, this adaptation warrants clinimetric testing. Without such testing, however, the original five-point scales should be maintained.


Direct rate assessment of laccase catalysed radical formation in lignin by electron paramagnetic resonance spectroscopy.
Laccases (EC 126.96.36.199) catalyse removal of an electron and a proton from phenolic hydroxyl groups, including phenolic hydroxyls in lignins, to form phenoxy radicals during reduction of O2. We employed electron paramagnetic resonance spectroscopy (EPR) for real time measurement of such catalytic radical formation activity on three types of lignin (two types of organosolv lignin, and a lignin rich residue from wheat straw hydrolysis) brought about by two different fungal laccases, derived from Trametes versicolor (Tv) and Myceliophthora thermophila (Mt), respectively. Laccase addition to suspensions of the individual lignin samples produced immediate time and enzyme dose dependent increases in intensity in the EPR signal with g-values in the range 2.0047-2.0050 allowing a direct quantitative monitoring of the radical formation and thus allowed laccase enzyme kinetics assessment on lignin. The experimental data verified that the laccases acted upon the insoluble lignin substrates in the suspensions. When the action on the lignin substrates of the two laccases were compared on equal enzyme dosage levels (by activity units on syringaldazine) the Mt laccase exerted a significantly faster radical formation than the Tv laccase on all three types of lignin substrates. When comparing the equal laccase dose rates on the three lignin substrates the enzymatic radical formation rate on the wheat straw lignin residue was consistently higher than those of the organosolv lignins. The pH-temperature optimum for the radical formation rate in organosolv lignin was determined by response surface methodology to pH 4.8, 33°C and pH 5.8, 33°C for the Tv laccase and the Mt laccase, respectively. The results verify direct radical formation action of fungal laccases on lignin without addition of mediators and the EPR methodology provides a new type of enzyme assay of laccases on lignin.


Representation of number in animals and humans: a neural model.
This article addresses the representation of numerical information conveyed by nonsymbolic and symbolic stimuli. In a first simulation study, we show how number-selective neurons develop when an initially uncommitted neural network is given nonsymbolic stimuli as input (e.g., collections of dots) under unsupervised learning. The resultant network is able to account for the distance and size effects, two ubiquitous effects in numerical cognition. Furthermore, the properties of the network units conform in detail to the characteristics of recently discovered number-selective neurons. In a second study, we simulate symbol learning by presenting symbolic and nonsymbolic input simultaneously. The same number-selective neurons learn to represent the numerical meaning of symbols. In doing so, they show properties reminiscent of the originally available number-selective neurons, but at the same time, the representational efficiency of the neurons is increased when presented with symbolic input. This finding presents a concrete proposal on the linkage between higher order numerical cognition and more primitive numerical abilities and generates specific predictions on the neural substrate of number processing.


Creating a CD-ROM program for cancer-related patient education.
To describe the process of developing a cancer-related patient and family education CD-ROM program and initial evaluation results. Published research, theory, practice, and personal experience. CD-ROM programs can be far more comprehensive than the booklets and videotapes used more commonly in patient education. Developing CD-ROM programs requires funding, organizational skills, access to content experts, and a team composed of people who have the varied skills required for a finished multimedia product. The time frame for CD-ROM production is often longer than that of other patient-education formats. Published reports and this institution's experience confirm that patients accept this medium. Evaluation to date suggests that CD-ROMs may be more useful to patients and their families than any other single information source. CD-ROM technology is more expensive than videotapes and booklets, but it allows for greater depth of content and may satisfy a broader range of educational needs than other media. Funding often can be obtained through foundations and with unrestricted educational grants from pharmaceutical companies. Nurses can lead multidisciplinary teams to produce CD-ROMs for their patient populations. These programs can be used before a patient has a first consultation to introduce a cancer or treatment and anytime during cancer diagnosis and treatment. They can reinforce one-on-one teaching or provide greater depth of content than ever could be provided in individualized teaching sessions. They can facilitate patients' self-directed learning and may allow nurses and doctors to teach on a different level. These programs also can complement patients' Internet searches either by creating a solid foundation for further investigation or by confirming the reliability of information gained through a variety of Internet sources.


Risk Factors for Dysplasia in Recurrent Respiratory Papillomatosis in an Adult and Pediatric Population.
Recurrent respiratory papillomatosis (RRP) is classically described as a benign neoplasm of the larynx caused by the low-risk human papillomavirus (HPV) viral subtypes. Nevertheless, transformation to dysplasia and invasive carcinoma can occur. We aimed to assess the prevalence of dysplasia and carcinoma-ex-papilloma in both adult-onset and juvenile-onset RRP and identify patient risk factors for this dysplastic transformation. Ten-year retrospective chart review of a tertiary otolaryngology referral center. Patients with papilloma were identified from a review of a pathology database and clinical records. Patient demographics, pathologic data, and treatment history, including use of cidofovir as an adjunctive therapy for papilloma, were extracted from electronic medical records. One hundred fifty-nine RRP patients were identified, 96 adult-onset (AORRP) and 63 juvenile-onset (JORRP) cases. Of this cohort, 139 (87%) had only benign papilloma as a pathologic diagnosis. In the AORRP cohort, 10 patients (10%) were diagnosed with dysplasia or carcinoma in situ in addition to papilloma, and 5 patients (5%) had malignant transformation to invasive carcinoma-ex-papilloma. There was a significantly higher age of disease onset for those with dysplasia or carcinoma versus those without dysplasia or carcinoma (56 vs 45 years old; P = .0005). Of the 63 JORRP patients, there were no cases of dysplasia but 3 (5%) cases of invasive carcinoma-ex-papilloma, all involving pulmonary disease. The JORRP patients with carcinoma-ex-papilloma had a younger average disease onset (2 vs 6 years old; P = .009) and a higher rate of tracheal involvement than those without carcinoma. Gender, smoking history, number of operations, or use of cidofovir showed no association with the development of dysplasia or carcinoma-ex-papillomatosis in either the AORRP or JORRP population. In a large series of RRP, age of disease onset is the strongest predictor of dysplastic transformation in the adult and pediatric population. Carcinoma-ex-papillomatosis was uniformly associated with pulmonary disease in the JORRP population in this series. No other demographic or behavioral factors, including adjunctive therapy with cidofovir, were statistically associated with dysplasia or carcinoma-ex-papilloma.


Caregiver-reported religious beliefs and complementary and alternative medicine use among children admitted to an epilepsy monitoring unit.
Complementary and alternative medicine (CAM) includes a wide range of practices and products that are generally outside the use of conventional medicine as practiced in Western cultures. Use of CAM in persons with epilepsy is high, even compared to individuals with other chronic health conditions. In this study, we surveyed caregivers of children admitted to a regional epilepsy monitoring unit (EMU) in the southeast United States to assess CAM use among patients (N=225). Thirteen percent of respondents indicated current use of CAM by their child, 16% reported past use, and 43% reported interest in future use, most commonly in marijuana as a potential treatment (23%). Over 25% of respondents expressed interest in CAM use related to side effects of anti-epileptic medications. Regarding prayer as a form of CAM, a large majority of respondents in this sample identified as Christian and actively prayed for their child's illness, revealing a high prevalence of spiritual practices in this population. Eighty-one percent of respondents reported that they had not discussed CAM use with their doctor. Discussing CAM use with a health care provider was significantly related to past CAM use (p<.02), but not current use or willingness to try CAM in the future (p>.05). These results have important implications for future practice and support increased communication and patient education, as many anti-epileptic medications interact with certain herbs and supplements, posing a potential health risk and treatment barrier in this population.


[The relations of the mandibular canal with the outer surfaces of the body of the mandible and the risks which arise from its injury].
The investigations of relations and position of the mandibular canal (CM) were carried out on 80 mandibles (33 dentulous, 27 partly edentulous and 20 completely edentulous). The studies of relations of CM to the sides of mandibular body were accomplished by morphometric analysis of CM on consecutive transversal sections of mandibular body (54 preparations). The relation of the CM to be buccal or to the lingual side is expressed as the distance from the surface of the correspondent side of the mandibular body, which is shown in Table 1. According to these data, the position of the canal is at first proximate to the lingual side, and from the first molar tooth (M1) it approaches the buccal surface of the mandibular body. The position of the entire CM was determined by analysis of mandibular preparations (26) after removing the buccal osseous lamina. These investigations revealed that the position of the CM is predominantly buccal. Reconstruction of relations of the CM to the sides of mandibular body, according to the data obtained from transversal sections, and the real position of the CM are presented in Scheme I. The difference in direction lines of relations and of predominant position of the CM is a consequence of morphological characteristics of mandibular body.


Accumulation of endogenous peptides triggers a pathogen stress response in Arabidopsis thaliana.
The stepwise degradation of peptides to amino acids in plant mitochondria and chloroplasts is catalyzed by a network of oligopeptidases (presequence protease PreP, organellar oligopeptidase OOP) and aminopeptidases. In the present report, we show that the lack of oligopeptidase activity in Arabidopsis thaliana results in the accumulation of endogenous free peptides, mostly of chloroplastic origin (targeting peptides and degradation products). Using mRNA sequencing and deep coverage proteomics, allowing for the identification of 17 000 transcripts and 11 000 proteins, respectively, we uncover a peptide-stress response occurring in plants lacking PreP and OOP oligopeptidase activity. The peptide-stress response results in the activation of the classical plant defense pathways in the absence of pathogenic challenge. The constitutive activation of the pathogen-defense pathways imposes a strong growth penalty and a reduction of the plants reproductive fitness. Our results indicate that the absence of organellar oligopeptidases PreP1/2 and OOP results in the accumulation of peptides that are perceived as pathogenic effectors and activate the signaling pathways of plant-defense response.


Conservative surgery for mitral valve disease: clinical and echocardiographic analysis of results.
Eighty-five patients underwent mitral valve reconstruction by the Carpentier method from January 1976 to December 1981. Concomitant procedures were performed in 30 patients (aortic valve replacement in 23, coronary revascularisation in six, and tricuspid valve repair in seven). Before operation 76 patients (89%) were in clinical class II or III (New York Heart Association) and atrial fibrillation was present in 50. Thirty-six patients had valvular incompetence, while 26 had pure stenosis. The aetiology was rheumatic in 57 cases and dysplastic in 21. The patients were assessed for clinical improvement, durability of valve repair, thromboembolism, and survival. There was one death, an operative mortality rate of 1.2%, and 63 of 74 patients followed for one to six years were in clinical class I after operation. The actuarial survival was 92% with a 93% incidence of freedom from thromboemboli at five years. Six patients had embolic episodes, four of whom had aortic valve replacement. Three patients had a repeat operation 16-20 months later, a valve failure rate of 6.7%. Nineteen patients with ruptured chordae had postoperative echocardiographic assessment of myocardial and mitral valve functions; the peak rates of dimension changes of the left ventricular cavity (indicative of flow across the mitral valve) fell to normal in most patients, and the left ventricular end-diastolic dimensions decreased significantly from 6.4 (1.53) to 5.09 (1.31) cm (mean and SD)--p less than 0.05. Our results confirm that reconstructive mitral surgery is able to restore and maintain normal valve function in addition to providing satisfactory relief of symptoms.


A Call to Action: A Specialty-Specific Course to Support the Next Generation of Clinician Scientists in Anesthesiology.
Clinical production pressure is a significant problem for faculty of anesthesiology departments who seek to remain involved in research. Lack of protected time to dedicate to research and insufficient external funding add to this long-standing issue. Recent trends in funding to the departments of anesthesiology and their academic output validate these concerns. A 2022 study examining National Institutes of Health (NIH) grant recipients associated with anesthesiology departments across 10 years (2011-2020) outlines total awarded funds at $1,676,482,440, with most of the funds awarded to only 10 departments in the United States. Of note, the total 1-year NIH funding in 2021 for academic internal medicine departments was 3 times higher than the 10-year funding of anesthesiology departments. Additionally, American Board of Anesthesiology (ABA) diplomats represent a minority (37%) of the anesthesiology researchers obtaining grant funding, with a small number of faculty members receiving a prevalence of monies. Overall, the number of publications per academic anesthesiologist across the United States remains modest as does the impact of the scholarly work. Improving environments in which academic anesthesiologists thrive may be paramount to successful academic productivity. In fact, adding to the lack of academic time is the limited bandwidth of senior academic physicians to mentor and support aspiring physician scientists. Given then the challenges for individual departments and notable successes of specialty-specific collaborative efforts (eg Foundation for Anesthesia Education and Research [FAER]), additional pooled-resource approaches may be necessary to successfully support and develop clinician scientists. It is in this spirit that the leadership of Anesthesia and Analgesia and the Journal of Education in Perioperative Medicine, unified with the Association of University Anesthesiologists, aim to sponsor the Introduction to Clinical Research for Academic Anesthesiologists (ICRAA) Course. Directed toward early career academic anesthesiologists who wish to gain competency specifically in the fundamentals of clinical research and receive mentorship to develop an investigative project, the yearlong course will provide participants with the skills necessary to design research initiatives, ethically direct research teams, successfully communicate ideas with data analysts, and write and submit scientific articles. Additionally, the course, articulated in a series of interactive lectures, mentored activities, and workshops, will teach participants to review articles submitted for publication to medical journals and to critically appraise evidence in published research. It is our hope that this initiative will be of interest to junior faculty of academic anesthesiology departments nationally and internationally.


Porin threading drives receptor disengagement and establishes active colicin transport through Escherichia coli OmpF.
Bacteria deploy weapons to kill their neighbours during competition for resources and to aid survival within microbiomes. Colicins were the first such antibacterial system identified, yet how these bacteriocins cross the outer membrane (OM) of Escherichia coli is unknown. Here, by solving the structures of translocation intermediates via cryo-EM and by imaging toxin import, we uncover the mechanism by which the Tol-dependent nuclease colicin E9 (ColE9) crosses the bacterial OM. We show that threading of ColE9's disordered N-terminal domain through two pores of the trimeric porin OmpF causes the colicin to disengage from its primary receptor, BtuB, and reorganises the translocon either side of the membrane. Subsequent import of ColE9 through the lumen of a single OmpF subunit is driven by the proton-motive force, which is delivered by the TolQ-TolR-TolA-TolB assembly. Our study answers longstanding questions, such as why OmpF is a better translocator than OmpC, and reconciles the mechanisms by which both Tol- and Ton-dependent bacteriocins cross the bacterial outer membrane.


A TetR-family transcription factor regulates fatty acid metabolism in the archaeal model organism Sulfolobus acidocaldarius.
Fatty acid metabolism and its regulation are known to play important roles in bacteria and eukaryotes. By contrast, although certain archaea appear to metabolize fatty acids, the regulation of the underlying pathways in these organisms remains unclear. Here, we show that a TetR-family transcriptional regulator (FadRSa) is involved in regulation of fatty acid metabolism in the crenarchaeon Sulfolobus acidocaldarius. Functional and structural analyses show that FadRSa binds to DNA at semi-palindromic recognition sites in two distinct stoichiometric binding modes depending on the operator sequence. Genome-wide transcriptomic and chromatin immunoprecipitation analyses demonstrate that the protein binds to only four genomic sites, acting as a repressor of a 30-kb gene cluster comprising 23 open reading frames encoding lipases and β-oxidation enzymes. Fatty acyl-CoA molecules cause dissociation of FadRSa binding by inducing conformational changes in the protein. Our results indicate that, despite its similarity in overall structure to bacterial TetR-family FadR regulators, FadRSa displays a different acyl-CoA binding mode and a distinct regulatory mechanism.


Large urban outbreak of orally acquired acute Chagas disease at a school in Caracas, Venezuela.
Trypanosoma cruzi oral transmission is possible through food contamination by vector's feces. Little is known about the epidemiology and clinical features of microepidemics of orally acquired acute Chagas disease (CD). A case-control, cohort-nested, epidemiological study was conducted during an outbreak of acute CD that affected a school community. Structured interviews were designed to identify symptoms and sources of infection. Electrocardiograms were obtained for all patients. Specific serum antibodies were assessed by immunoenzimatic and indirect hemagglutination tests. In some cases, parasitemia was tested directly or by culture, animal inoculation, and/or a polymerase chain reaction technique. Infection was confirmed in 103 of 1000 exposed individuals. Of those infected, 75% were symptomatic, 20.3% required hospitalization, 59% showed ECG abnormalities, parasitemia was documented in 44, and 1 child died. Clinical features differed from those seen in vectorial transmission. The infection rate was significantly higher among younger children. An epidemiological investigation incriminated contaminated fresh guava juice as the sole source of infection. This outbreak was unique, because it affected a large, urban, predominantly young, middle-class, otherwise healthy population and resulted in an unprecedented public health emergency. Rapid diagnosis and treatment avoided higher lethality. Food-borne transmission of T. cruzi may occur more often than is currently recognized.


The yin and yang of nrf2-regulated selenoproteins in carcinogenesis.
The NF-E2-related factor-2 (Nrf2) is a transcription factor which regulates the major cellular defense systems and thereby contributes to the prevention of many diseases including cancer. Selenium deficiency is associated with a higher cancer risk making also this essential trace element a promising candidate for cancer prevention. Two selenoproteins, thioredoxin reductase-1 (TrxR1) and glutathione peroxidase-2 (GPx2), are targets for Nrf2. Selenium deficiency activates Nrf2 as does a TrxR1 knockout making a synergism between both systems plausible. Although this might hold true for healthy cells, the interplay may turn into the opposite in cancer cells. The induction of the detoxifying and antioxidant enzymes by Nrf2 will make cancer cells chemoresistant and will protect them against oxidative damage. The essential role of TrxR1 in maintaining proliferation makes its upregulation in cancer cells detrimental. The anti-inflammatory potential of GPx2 will help to inhibit cancer initiation and inflammation-triggered promotion, but its growth supporting potential will also support tumor growth. This paper considers beneficial and adverse consequences of the activation of Nrf2 and the selenoproteins which appear to depend on the cancer stage.


Student engagement, assessed using heart rate, shows no reset following active learning sessions in lectures.
Heart rate can be used as a measure of cognitive engagement. We measured average student heart rates during medical school lecture classes using wristwatch-style monitors. Analysis of 42 classes showed a steady decline in heart rate from the beginning to end of a lecture class. Active learning sessions (peer-discussion based problem solving) resulted in a significant uptick in heart rate, but this returned to the average level immediately following the active learning period. This is the first statistically robust assessment of changes in heart rate during the course of college lecture classes and indicates that personal heart rate monitors may be useful tools for assessment of different teaching modalities. The key findings suggest that the value of active learning within the classroom resides in the activity itself and not in an increase in engagement or reset in attention during the didactic period following an active learning session.


An extensive denture-induced hyperplasia of maxilla.
Denture-induced hyperplasia is a reactive lesion arising from excessive and chronic mechanical pressure on the vestibular oral mucosa. It has a female predilection and it is mostly seen in the maxilla. The size of the lesion may be as small as a few millimeters to massive lesion involving the entire vestibule. It is usually asymptomatic but sometimes severe inflammation and ulceration can occur. Elimination of the inflammation and excision of the lesion is the treatment of choice. Denture induced hyperplasia in right maxillary buccal vestibule in a middle age old female patient was presented. Surgical excision was done and new denture was fabricated.


Switching photochromic molecules adsorbed on optical microfibres.
The internal state of organic photochromic spiropyran molecules adsorbed on optical microfibres is optically controlled and measured by state-dependent light absorption. Repeated switching between the states is achieved by exposure to the evanescent field of a few nanowatts of light guided in the microfibre. By adjusting the microfibre evanescent field strength the dynamic equilibrium state of the molecules is controlled. Time-resolved photoswitching dynamics are measured and modelled with a rate equation model. We also study how many times the photochromic system can be switched before undergoing significant photochemical degradation.


Ventral Hernia Management: Expert Consensus Guided by Systematic Review.
To achieve consensus on the best practices in the management of ventral hernias (VH). Management patterns for VH are heterogeneous, often with little supporting evidence or correlation with existing evidence. A systematic review identified the highest level of evidence available for each topic. A panel of expert hernia-surgeons was assembled. Email questionnaires, evidence review, panel discussion, and iterative voting was performed. Consensus was when all experts agreed on a management strategy. Experts agreed that complications with VH repair (VHR) increase in obese patients (grade A), current smokers (grade A), and patients with glycosylated hemoglobin (HbA1C) ≥ 6.5% (grade B). Elective VHR was not recommended for patients with BMI ≥ 50 kg/m (grade C), current smokers (grade A), or patients with HbA1C ≥ 8.0% (grade B). Patients with BMI= 30-50 kg/m or HbA1C = 6.5-8.0% require individualized interventions to reduce surgical risk (grade C, grade B). Nonoperative management was considered to have a low-risk of short-term morbidity (grade C). Mesh reinforcement was recommended for repair of hernias ≥ 2 cm (grade A). There were several areas where high-quality data were limited, and no consensus could be reached, including mesh type, component separation technique, and management of complex patients. Although there was consensus, supported by grade A-C evidence, on patient selection, the safety of short-term nonoperative management, and mesh reinforcement, among experts; there was limited evidence and broad variability in practice patterns in all other areas of practice. The lack of strong evidence and expert consensus on these topics has identified gaps in knowledge where there is need of further evidence.


Serum nitric oxide metabolites in patients with multiple sclerosis.
Multiple sclerosis (MS) is an autoimmune disease of the central nervous system characterized by myelin breakdown. The free radical nitric oxide (NO), which is considered to be a major metabolite in immune function and in autoimmune disorders, is among the possible mediators causing the inflammatory reactions in MS. Consequently, NO has been implicated in the pathogenesis of MS and its animal model experimental allergic encephalomyelitis (EAE). In this study, stable metabolites of NO (NO(2-)+NO(3-)) levels were determined in sera of MS patients (n=23) and control subjects (n=16). NO(2-)+NO(3-) levels were higher in MS patients when compared to control subjects. However, there was not any correlation with serum NO(2-)+NO(3-) values and clinical features of the disease such as duration of sickness, the time elapsed from the last attack and EDSS values. Our results imply that nitric oxide may be involved in the pathogenesis of MS although further studies are required to elucidate underlying mechanisms.


Retinal vascular caliber measurements: clinical significance, current knowledge and future perspectives.
The retinal vasculature provides a unique window to assess vascular health noninvasively and directly in vivo. Advances in fundus photography and retinal image analysis techniques have enabled the objective and accurate assessment of quantitative retinal vascular caliber measurement. Over the last decade, large population-based studies have shown that retinal vascular calibers are associated with a wide range of subclinical (e.g. atherosclerosis, inflammation and endothelial dysfunction) and clinical cardiovascular diseases (hypertension, diabetes mellitus, stroke, kidney and heart diseases). However, while retinal image analysis provided exciting possibilities to study the pathogenesis of these diseases, its direct applicability in a clinical setting as a 'test' to predict cardiovascular diseases is yet to be established, particularly within the context of being used as a population screening tool. Nevertheless, with continual development of retinal imaging techniques and newer understanding of the clinical significance of these retinal changes, there remains scope for the development of retinal vascular caliber measurements as a biomarker for vascular disease risk assessment in targeted areas and patient subgroups (e.g. patients with diabetes, suspected hypertension and stroke).


Multiple, ligand-dependent routes from the active site of cytochrome P450 2C9.
The active site of liver-specific, drug-metabolizing cytochrome P450 (CYP) monooxygenases is deeply buried in the protein and is connected to the protein surface through multiple tunnels, many of which were found open in different CYP crystal structures. It has been shown that different tunnels could serve as ligand passage routes in different CYPs. However, it is not understood whether one CYP uses multiple routes for substrate access and product release and whether these routes depend on ligand properties. From 300 ns of molecular dynamics simulations of CYP2C9, the second most abundant CYP in the human liver we found four main ligand exit routes, the occurrence of each depending on the ligand type and the conformation of the F-G loop, which is likely to be affected by the CYP-membrane interaction. A non-helical F-G loop favored exit towards the putative membrane-embedded region. Important protein features that direct ligand exit include aromatic residues that divide the active site and whose motions control access to two pathways. The ligands interacted with positively charged residues on the protein surface through hydrogen bonds that appear to select for acidic substrates. The observation of multiple, ligand-dependent routes in a CYP aids understanding of how CYP mutations affect drug metabolism and provides new possibilities for CYP inhibition.


Management of erosion of inflatable penile prosthesis reservoir into bladder. A different approach.
We report a rare case of erosion of an inflatable penile prosthesis reservoir into the bladder that was managed with a different approach from the literature by preserving the existing reservoir. Inflatable penile implant was applied to a 54-year-old male patient who had undergone with a robot-assisted radical prostatectomy operation due to localized prostate cancer 2 years before. Two months after the operation, the patient referred to our clinic with predominant symptoms of lower urinary tract system associated with scrotal pain and swelling. The urinary system ultrasonography (USG) and the lower abdomen magnetic resonance imaging (MRI) demonstrated that the reservoir of the penile prosthesis was in the bladder. Cystoscopy confirmed that the reservoir was in the bladder. According to literature the reservoir was surgically removed from bladder. After bladder repair, the rectus muscles were repaired creating a space between the rectus muscle and the skin, where the reservoir was placed. After postoperative observation, the patient was discharged without any infection and regression of the lower urinary tract symptoms. No problem was referred by using the penile prosthesis when at 1-month and 3-month follow up and the patient was not uncomfortable in this regard. In conclusion no drawback occurred by using the old reservoir.


A description of the methods of the Nulliparous Pregnancy Outcomes Study: monitoring mothers-to-be (nuMoM2b).
The primary aim of the "Nulliparous Pregnancy Outcomes Study: monitoring mothers-to-be" is to determine maternal characteristics, which include genetic, physiologic response to pregnancy, and environmental factors that predict adverse pregnancy outcomes. Nulliparous women in the first trimester of pregnancy were recruited into an observational cohort study. Participants were seen at 3 study visits during pregnancy and again at delivery. We collected data from in-clinic interviews, take-home surveys, clinical measurements, ultrasound studies, and chart abstractions. Maternal biospecimens (serum, plasma, urine, cervicovaginal fluid) at antepartum study visits and delivery specimens (placenta, umbilical cord, cord blood) were collected, processed, and stored. The primary outcome of the study was defined as pregnancy ending at <37+0 weeks' gestation. Key study hypotheses involve adverse pregnancy outcomes of spontaneous preterm birth, preeclampsia, and fetal growth restriction. We recruited 10,037 women to the study. Basic characteristics of the cohort at screening are reported. The "Nulliparous Pregnancy Outcomes Study: monitoring mothers-to-be" cohort study methods and procedures can help investigators when they plan future projects.