Developing and applying a gene functional association network for anti-angiogenic kinase inhibitor activity assessment in an angiogenesis co-culture model.
Tumor angiogenesis is a highly regulated process involving intercellular communication as well as the interactions of multiple downstream signal transduction pathways. Disrupting one or even a few angiogenesis pathways is often insufficient to achieve sustained therapeutic benefits due to the complexity of angiogenesis. Targeting multiple angiogenic pathways has been increasingly recognized as a viable strategy. However, translation of the polypharmacology of a given compound to its antiangiogenic efficacy remains a major technical challenge. Developing a global functional association network among angiogenesis-related genes is much needed to facilitate holistic understanding of angiogenesis and to aid the development of more effective anti-angiogenesis therapeutics. We constructed a comprehensive gene functional association network or interactome by transcript profiling an in vitro angiogenesis model, in which human umbilical vein endothelial cells (HUVECs) formed capillary structures when co-cultured with normal human dermal fibroblasts (NHDFs). HUVEC competence and NHDF supportiveness of cord formation were found to be highly cell-passage dependent. An enrichment test of Biological Processes (BP) of differentially expressed genes (DEG) revealed that angiogenesis related BP categories significantly changed with cell passages. Built upon 2012 DEGs identified from two microarray studies, the resulting interactome captured 17226 functional gene associations and displayed characteristics of a scale-free network. The interactome includes the involvement of oncogenes and tumor suppressor genes in angiogenesis. We developed a network walking algorithm to extract connectivity information from the interactome and applied it to simulate the level of network perturbation by three multi-targeted anti-angiogenic kinase inhibitors. Simulated network perturbation correlated with observed anti-angiogenesis activity in a cord formation bioassay. We established a comprehensive gene functional association network to model in vitro angiogenesis regulation. The present study provided a proof-of-concept pilot of applying network perturbation analysis to drug phenotypic activity assessment.


Social equity shapes zone-selection: Balancing aquatic biodiversity conservation and ecosystem services delivery in the transboundary Danube River Basin.
Freshwater biodiversity is declining, despite national and international efforts to manage and protect freshwater ecosystems. Ecosystem-based management (EBM) has been proposed as an approach that could more efficiently and adaptively balance ecological and societal needs. However, this raises the question of how social and ecological objectives can be included in an integrated management plan. Here, we present a generic model-coupling framework tailored to address this question for freshwater ecosystems, using three components: biodiversity, ecosystem services (ESS), and a spatial prioritisation that aims to balance the spatial representation of biodiversity and ESS supply and demand. We illustrate this model-coupling approach within the Danube River Basin using the spatially explicit, potential distribution of (i) 85 fish species as a surrogate for biodiversity as modelled using hierarchical Bayesian models, and (ii) four estimated ESS layers produced by the Artificial Intelligence for Ecosystem Services (ARIES) platform (with ESS supply defined as carbon storage and flood regulation, and demand specified as recreation and water use). These are then used for (iii) a joint spatial prioritisation of biodiversity and ESS employing Marxan with Zones, laying out the spatial representation of multiple management zones. Given the transboundary setting of the Danube River Basin, we also run comparative analyses including the country-level purchasing power parity (PPP)-adjusted gross domestic product (GDP) and each country's percent cover of the total basin area as potential cost factors, illustrating a scheme for balancing the share of establishing specific zones among countries. We demonstrate how emphasizing various biodiversity or ESS targets in an EBM model-coupling framework can be used to cost-effectively test various spatially explicit management options across a multi-national case study. We further discuss possible limitations, future developments, and requirements for effectively managing a balance between biodiversity and ESS supply and demand in freshwater ecosystems.


In vivo 3D human vocal fold imaging with polarization sensitive optical coherence tomography and a MEMS scanning catheter.
We present in-vivo 3D human vocal fold images with polarization sensitive optical coherence tomography (PS-OCT). Characterizing the extent and location of vocal fold lesions provides useful information in guiding surgeons during phonomicrosurgery. Previous studies showed that PS-OCT imaging can distinguish vocal fold lesions from normal tissue, but these studies were limited to 2D cross-sectional imaging and were susceptible to sampling error. In-vivo 3D endoscopic imaging was performed by using a recently developed 2-axis MEMS scanning catheter and a spectral domain OCT (SD-OCT), running at 18.5 frames/s. Imaging was performed in the operating room with patients under general anesthesia and 3D images were acquired either by 2D scanning of the scanner on the sites of interest or by combining 1D scanning and manual sliding to capture whole length of the vocal fold. Vocal fold scar, polyps, nodules, papilloma and malignant lesions were imaged and characteristics of individual lesions were analyzed in terms of spatial distribution and variation of tissue structure and birefringence. The 3D large sectional PS-OCT imaging showed that the spatial extent of vocal fold lesions can be found non-invasively with good contrast from normal tissue.


Bowel function after minimally invasive urogynecologic surgery: a prospective randomized controlled trial.
The goals of this study were to assess the effect of a standardized postoperative bowel regimen of over-the-counter medications on (1) time to first bowel movement (BM) and (2) pain level associated with first BM in subjects undergoing minimally invasive urogynecologic surgery. Eligible patients scheduled to undergo minimally invasive urogynecologic surgery were offered participation. Enrolled subjects were randomized by computerized schedule. Demographic and perioperative data were collected. Subjects completed a validated questionnaire preoperatively and postoperatively assessing preexisting constipation, frequency and consistency of bowel movements, use of pain medications, mean daily pain level (using visual analog scale), stool consistency, and pain associated with first postoperative bowel movement. The control group was instructed to take docusate sodium twice daily postoperatively. The treatment group took docusate sodium plus Miralax, fiber wafers, and bisacodyl suppositories as directed by protocol. Wilcoxon or t testing was used to compare continuous variables; χ testing was used for categorical relationships, and backward-elimination multiple regression was used to assess independent effects. Seventy-two subjects were enrolled and randomized. Twelve subjects withdrew, leaving 60 (30 per group) completing the study. There were no statistically significant differences between groups in baseline characteristics. Mean (SD) age was 63 (9) years for the control group and 58 (10) for the study group (P = 0.06). Mean pelvic organ prolapse stage was III in each group. The mean (SD) operating room time was 198 (65) minutes for the controls and 216 (74) for the study subjects. Sixty-five percent underwent robot-assisted surgery (50% hysterectomy and 63% sacrocolpopexy). Ninety-eight percent of surgeries were performed under general anesthesia.Before adjustment, the mean (SD) time to first BM was 77 (24) hours in controls versus 64 (21) in the study patients (P = 0.03). Using multiple regression, baseline frequency of defecation (1-2 BMs/wk) was directly associated with the time to first BM (added 25.2 hours; P = 0.009) and being in the study group was inversely associated (first BM, 11.7 hours sooner; P = 0.04). No other variables were retained.There was no difference in pain associated with first postoperative BM (visual analog scale, 3.6 (3.2) vs 3.7 (2.8); P = 0.98), but those with prior complaints of vaginal or rectal splinting had higher pain scores (1.9 and 2.8 points higher, respectively; P = 0.04 for both). There was a trend toward higher pain scores with higher postoperative daily narcotic intake (P = 0.06). No other variables were retained.There was a significant difference in recorded compliance between control versus study regimens (94% vs 81%, respectively; P = 0.002). Mean time to first postoperative BM after minimally invasive urogynecologic surgery is more than 3.5 days with use of docusate sodium alone and is only slightly shorter when combination therapy is used. First BM after surgery is considered to be painful despite the use of medications. Future studies targeting postoperative discomfort/pain with defecation could target preoperative bowel regimens or more aggressive postoperative interventions. Regimens should remain simple to increase compliance.


Transformation of cystic forms of Borrelia burgdorferi to normal, mobile spirochetes.
The purpose of this study was to evaluate the behaviour of Borrelia burgdorferi under controlled conditions. The occurrence of cystic forms of Borrelia burgdorferi in vitro was noted, and these cysts were able to be transformed to normal, mobile spirochetes. B. burgdorferi was cultivated in a commercial culture medium without serum. The spirochetes multiplied only slowly in this medium, and transformation to encysted forms was observed after 1 week. When these cysts were transferred to the same culture medium with rabbit serum, the encysted forms developed into regular, mobile spirochetes after 6 weeks, and their regeneration time was normal. Examination of these cysts in the transmission electron microscope revealed transverse fission inside the cysts. It is probable that similar phenomena may occur in vivo under conditions unfavourable for spirochetes. These observations may help to explain why diagnosis and treatment of B. burgdorferi infections in humans can be difficult.


Resonant quantum transitions in trapped antihydrogen atoms.
The hydrogen atom is one of the most important and influential model systems in modern physics. Attempts to understand its spectrum are inextricably linked to the early history and development of quantum mechanics. The hydrogen atom's stature lies in its simplicity and in the accuracy with which its spectrum can be measured and compared to theory. Today its spectrum remains a valuable tool for determining the values of fundamental constants and for challenging the limits of modern physics, including the validity of quantum electrodynamics and--by comparison with measurements on its antimatter counterpart, antihydrogen--the validity of CPT (charge conjugation, parity and time reversal) symmetry. Here we report spectroscopy of a pure antimatter atom, demonstrating resonant quantum transitions in antihydrogen. We have manipulated the internal spin state of antihydrogen atoms so as to induce magnetic resonance transitions between hyperfine levels of the positronic ground state. We used resonant microwave radiation to flip the spin of the positron in antihydrogen atoms that were magnetically trapped in the ALPHA apparatus. The spin flip causes trapped anti-atoms to be ejected from the trap. We look for evidence of resonant interaction by comparing the survival rate of trapped atoms irradiated with microwaves on-resonance to that of atoms subjected to microwaves that are off-resonance. In one variant of the experiment, we detect 23 atoms that survive in 110 trapping attempts with microwaves off-resonance (0.21 per attempt), and only two atoms that survive in 103 attempts with microwaves on-resonance (0.02 per attempt). We also describe the direct detection of the annihilation of antihydrogen atoms ejected by the microwaves.


Cancer mortality in a northern Italian cohort of rubber workers.
An analysis of the mortality of a cohort of 6629 workers employed from 1906 to 1981 in a rubber tyre factory in northern Italy (978 deaths and over 133,000 man-years at risk) showed that the all cause mortality ratio was slightly lower than expected (0.91). Overall cancer mortality was close to expected (275 v 259.4) but there were significant excess rates for two cancer sites: pleura (9 observed v 0.8 expected, which may be due to the use of fibre containing talc) and bladder (16 observed v 8.8 expected). Death rates were not raised for other sites previously associated with employment in the rubber industry, such as cancers of the lung and brain, leukaemias, or lymphomas. The substantially reduced relative risk of pleural cancer among workers first employed after 1940 (RR = 0.05 compared with before 1940) probably reflected improvements in working conditions over more recent periods. For cancer of the bladder, the relative risk was also lower for workers first engaged after 1940. Thus no appreciable risk for any disease was apparent for workers employed over the past four decades. Analysis for each of the 27 job categories showed a substantial excess for cancer of the pleura in the mechanical maintenance workers (4 observed v 0.17 expected); an excess of cancer of the lung (21 v 13.48) was also present in this job category.


Concurrent infection with Leishmania donovani and Leishmania major in a Kenyan patient: clinical description and parasite characterization.
Leishmania isolates aspirated a few months apart from the spleen of an indigenous adult male kala-azar patient from Baringo District, Kenya, were biochemically characterized and compared. The patient lived within a dual focus of L. donovani kalazar and L. major cutaneous leishmaniasis. A primary Leishmania isolate from splenic aspirates was cryopreserved (NLB-294). The patient was treated with sodium stibogluconate for kala-azar and discharged. Three months later, he had clinical relapse and returned for retreatment. During his second visit, the patient participated in a diagnostic study in which urine and nasopharyngeal samples were cultured for leishmaniasis. Urine, nasopharyngeal, and splenic samples were positive for Leishmania. Secondary isolates from splenic (NLB-294-I) and urine (NLB-318) cultures were cryopreserved and characterized by cellulose acetate electrophoresis (CAE) using 20 enzymes. Whereas the urine isolate was typed as L. donovani, the splenic aspirate culture revealed a mixed infection with L. donovani and L. major. The primary isolate (NLB-294) was then characterized and also showed a mixed infection. To exclude the possibility of protein post-translational modifications in electrophoretic assays, the primary and secondary isolates were grown and processed under identical cultural and lysis conditions, and compared using CAE. The results were identical to the first electrophoretic assays showing mixed promastigote banding patterns. Stationary-phase promastigotes of the secondary splenic isolate (NLB-294-I) inoculated subcutaneously, intraperitoneally, and intracardially into Syrian hamsters and BALB/c mice produced both kala-azar and cutaneous leishmaniasis within 6.5 months.(ABSTRACT TRUNCATED AT 250 WORDS)


What explains between-school differences in rates of smoking?
Schools have the potential to influence their pupils' behaviour through the school's social organisation and culture (non-formal school characteristics), as well as through the formal curriculum. This paper examines whether these school characteristics (which include a measure of quality of social relationships) can account for school differences in smoking rates. This study uses a longitudinal survey involving 5,092 pupils in 24 Scottish schools. Pupils' smoking (at age 15/16), cognitive measures, attitude to school and pupils' rating of teacher pupil relationships (at age 13/14) were linked to school level data comprising teacher assessed quality of pupil-staff relationships, school level deprivation, staying on rates and attendance. Analysis involved multi-level modelling. Overall, 25% of males and 39% of females reported smoking, with rates by school ranging from 8% to 33% for males and from 28% to 49% for females. When individual socio-economic and socio-cultural factors were controlled for there was still a large school effect for males and a smaller (but correlated) school effect for females at 15/16 years. For girls their school effect was explained by their rating of teacher-pupil relationships and attitude to school. These variables were also significant in predicting smoking among boys. However, the school effect for boys was most radically attenuated and became insignificant when the interaction between poor quality of teacher - pupil relationships and school level affluence was fitted, explaining 82% of the variance between schools. In addition, researchers' rating of the schools' focus on caring and inclusiveness was also significantly associated with both male and female smoking rates. School-level characteristics have an impact on male and female pupils' rates of smoking up to 15/16 years of age. The size of the school effect is greater for males at this age. The social environment of schools, in particular the quality of teacher-pupil relationships, pupils' attitude to school and the school's focus on caring and inclusiveness, can influence both boys' and girls' smoking. This provides support for the school-wide or "Health Promoting School" approach to smoking prevention.


The effect of sex-biased dispersal on opposite-sexed spatial genetic structure and inbreeding risk.
Natal sex-biased dispersal has long been thought to reduce the risk of inbreeding by spatially separating opposite-sexed kin. Yet, comprehensive and quantitative evaluations of this hypothesis are lacking. In this study, we quantified the effectiveness of sex-biased dispersal as an inbreeding avoidance strategy by combining spatially explicit simulations and empirical data. We quantified the extent of kin clustering by measuring the degree of spatial autocorrelation among opposite-sexed individuals (FM structure). This allowed us to systematically explore how the extent of sex-biased dispersal, generational overlap, and mate searching distance, influenced both kin clustering, and the resulting inbreeding in the absence of complementary inbreeding avoidance strategies. Simulations revealed that when sex-biased dispersal was limited, positive FM genetic structure developed quickly and increased as the mate searching distance decreased or as generational overlap increased. Interestingly, complete long-range sex-biased dispersal did not prevent the development of FM genetic structure when generations overlapped. We found a very strong correlation between FM genetic structure and both FIS under random mating, and pedigree-based measures of inbreeding. Thus, we show that the detection of FM genetic structure can be a strong indicator of inbreeding risk. Empirical data for two species with different life history strategies yielded patterns congruent with our simulations. Our study illustrates a new application of spatial genetic autocorrelation analysis that offers a framework for quantifying the risk of inbreeding that is easily extendable to other species. Furthermore, our findings provide other researchers with a context for interpreting observed patterns of opposite-sexed spatial genetic structure.


Symptomatic Exposures Among California Inmates 2011-2013.
Prisoners have a high prevalence of substance misuse and abuse, but few studies have examined symptomatic exposures among incarcerated populations. We sought to further characterize the nature of these exposures among this population using the California Poison Control System data. Keyword searches identified inmate cases in 2011-2013 for patients 20+ years old exposed to a single substance and taken to hospital from jail, prison, or police custody. Comparisons were made with non-inmate cases during the same period, using similar limitations. Body stuffers and body packers were analyzed as a subgroup. Seven hundred four inmate cases were compared to 106,260 non-inmate cases. Inmates were more likely to be younger, male, and to have engaged in drug misuse or abuse. They most commonly ingested methamphetamine, heroin, acetaminophen, and anticonvulsants. Inmates were more likely to receive activated charcoal (OR 9.87, 8.20-11.88), whole bowel irrigation (OR 44.50, 33.83-58.54), undergo endotracheal intubation (OR 4.09, 2.91-5.73), and to experience a major clinical outcome or death (OR 1.41, 1.05-1.89). When body stuffers and packers were removed, clinical findings were similar, though the odds of a major outcome or death became statistically non-significant. Body stuffers and body packers primarily used methamphetamine and heroin, and compared with other inmates had significantly higher odds of both adverse clinical effects and poor outcome. This large series provides a profile of symptomatic exposures among inmates, a little-studied population. The potential for high morbidity among body stuffers and packers suggests that a high index of suspicion of such ingestions be maintained when evaluating patients prior to incarceration.


Studies of the effects of FK506 on renal allografting in the beagle dog.
The immunosuppressive activities of a newly discovered macrolide extracted from Streptomyces tsukubaensis, FK506, were examined using 38 renal allografts in the beagle dog. The median survival time was 15.5 days in dogs without treatment, 61 days with a dose of 0.08 mg/kg/day and 176 days with a dose of 0.16 mg/kg/day of intramuscularly administered FK506. Prolongation of survival was statistically significant when compared with controls (P = 0.02, 0.0044, respectively). None of 6 recipient dogs receiving the agent at a dose of 0.16 mg/kg/day encountered rejection during the treatment course. Three of them survived over 200 days. Oral administration of FK506 at a dose of 0.32 mg/kg/day did not prolong the median survival time (20.5 days) compared with the placebo treated control (16.5 days), but oral treatment with 1.0 mg/kg/day resulted in all of the recipient dogs surviving over 130 days. Histological studies of 7 kidney graft biopsy specimens of the dogs surviving over 3 months revealed no cell infiltration or only some degree of reversible interstitial cell infiltration, but vascular and glomerular changes were not observed in any of the specimens. Irregularity of nuclear shape and cytoplasmic vacuolation of the pars recta of the proximal tubules were observed in one dog each. Liver biopsy specimens showed no consistent evidence of hepatocellular damage. Three dogs died of intussusception 2-3 weeks posttransplant. The dogs treated intramuscularly with 0.32 mg/kg/day suffered from anorexia. Two dogs receiving oral treatment at a dose of 1.0 mg/kg developed papilloma of the skin around day 60, but the tumors disappeared by day 120. We conclude that FK506 is a powerful immunosuppressant in the dog with tolerable side effects.


The role of rheumatoid arthritis (RA) flare and cumulative burden of RA severity in the risk of cardiovascular disease.
To examine the role of rheumatoid arthritis (RA) flare, remission and RA severity burden in cardiovascular disease (CVD). In a population-based cohort of patients with RA without CVD (age ≥30 years; 1987 American College of Rheumatology criteria met in 1988-2007), we performed medical record review at each clinical visit to estimate flare/remission status. The previously validated RA medical Records-Based Index of Severity (RARBIS) and Claims-Based Index of RA Severity (CIRAS) were applied. Age- and sex-matched non-RA subjects without CVD comprised the comparison cohort. Cox models were used to assess the association of RA activity/severity with CVD, adjusting for age, sex, calendar year of RA, CVD risk factors and antirheumatic medications. Study included 525 patients with RA and 524 non-RA subjects. There was a significant increase in CVD risk in RA per time spent in each acute flare versus remission (HR 1.07 per 6-week flare, 95% CI 1.01 to 1.15). The CVD risk for patients with RA in remission was similar to the non-RA subjects (HR 0.90, 95% CI 0.51 to 1.59). Increased cumulative moving average of daily RARBIS (HR 1.16, 95% CI 1.03 to 1.30) and CIRAS (HR 1.38, 95% CI 1.12 to 1.70) was associated with CVD. CVD risk was higher in patients with RA who spent more time in medium (HR 1.08, 95% CI 0.98 to 1.20) and high CIRAS tertiles (HR 1.18, 95% CI 1.06 to 1.31) versus lower tertile. Our findings show substantial detrimental role of exposure to RA flare and cumulative burden of RA disease severity in CVD risk in RA, suggesting important cardiovascular benefits associated with tight inflammation control and improved flare management in patients with RA.


Evaluation of a Single Dose of Azithromycin for Trachoma in Low-Prevalence Communities.
Trachoma, caused by repeated ocular infection with Chlamydia trachomatis, is the leading infectious cause of blindness worldwide and is targeted for elimination as a public health problem. We sought to determine whether a one-time azithromycin mass treatment would reduce trachomatous inflammation-follicular (TF) levels below the elimination threshold of 5% in communities with disease prevalence between 5 and 9.9%. The study was conducted in 96 sub-village units (balozis) in the Kongwa district of Tanzania which were predicted from prior prevalence surveys to have TF between 5 and 9.9%. Balozis were randomly assigned to the intervention and control arms. The intervention arm received a single mass drug administration of azithromycin. At baseline and 12-month follow-up, ocular exams for trachoma, ocular swabs for detection of chlamydial DNA, and finger prick blood for analysis of anti-chlamydial antibody were taken. Comparison of baseline and 12-month follow-up showed no significant difference in the overall TF1-9 prevalence by balozi between control and treatment arms. In the treatment arm there was a significant reduction of ocular infection 12 months after treatment (p = 0.004) but no change in the control arm. No change in Pgp3-specific antibody responses were observed after treatment in the control or treatment arms. Anti-CT694 responses increased in both study arms (p = 0.009 for control arm and p = 0.04 for treatment arm). These data suggest that a single round of MDA may not be sufficient to decrease TF levels below 5% when TF1-9 is between 5 and 9.9% at baseline.


First study of microdeletions in the Y chromosome of Algerian infertile men with idiopathic oligo- or azoospermia.
The human Y chromosome is essential for human sex determination and spermatogenesis. The long arm contains the azoospermia factor (AZF) region. Microdeletions in this region are responsible for male infertility. The objective of this study was to determine the frequency of Y microdeletions in Algerian infertile males with azoospermia and oligoasthenoteratozoospermia syndrome (OATS) and to compare the prevalence of these abnormalities with other countries and regions worldwide. A sample of 80 Algerian infertile males with a low sperm count (1-20 × 10(6) sperms/ml) as well as 20 fertile male controls was screened for Y chromosome microdeletions. 49 men were azoospermic and 31 men had OATS. Genomic DNA was isolated from blood and polymerase chain reaction was carried out with a set of 6 AZFa, AZFb and AZFc STS markers to detect the microdeletions as recommended by the European Academy of Andrology. Among the 80 infertile men screened for microdeletion, 1 subject was found to have microdeletions in the AZFc (sY254 and sY255) region. The deletion was found in azoospermic subjects (1/49, 2%). The overall AZF deletion frequency was low (1/80, 1.3%). AZF microdeletions were observed neither in the OATS group nor in the control group. The frequency of AZF microdeletions in infertile men from Algeria was comparable to those reported in the literature. We suggest analyzing 6 STS in the first step to detect Y microdeletions in our population.


Assessing the Clinical and Socioeconomic Burden of Respiratory Syncytial Virus in Children Aged Under 5 Years in Primary Care: Protocol for a Prospective Cohort Study in England and Report on the Adaptations of the Study to the COVID-19 Pandemic.
Respiratory syncytial virus (RSV) commonly causes lower respiratory tract infections and hospitalization in children. In 2019-2020, the Europe-wide RSV ComNet standardized study protocol was developed to measure the clinical and socioeconomic disease burden of RSV infections among children aged <5 years in primary care. RSV has a recognized seasonality in England. We aimed to describe (1) the adaptations of the RSV ComNet standardized study protocol for England and (2) the challenges of conducting the study during the COVID-19 pandemic. This study was conducted by the Oxford-Royal College of General Practitioners Research and Surveillance Centre-the English national primary care sentinel network. We invited all (N=248) general practices within the network that undertook virology sampling to participate in the study by recruiting eligible patients (registered population: n=3,056,583). Children aged <5 years with the following case definition of RSV infection were included in the study: those consulting a health care practitioner in primary care with symptoms meeting the World Health Organization's definition of acute respiratory illness or influenza-like illness who have laboratory-confirmed RSV infection. The parents/guardians of these cases were asked to complete 2 previously validated questionnaires (14 and 30 days postsampling). A sample size of at least 100 RSV-positive cases is required to estimate the percentage of children that consult in primary care who need hospitalization. Assuming a swab positivity rate of 20% in children aged <5 years, we estimated that 500 swabs are required. We adapted our method for the pandemic by extending sampling planned for winter 2020-2021 to a rolling data collection, allowing verbal consent and introducing home swabbing because of increased web-based consultations during the COVID-19 pandemic. The preliminary results of the data collection between International Organization for Standardization (ISO) weeks 1-41 in 2021 are described. There was no RSV detected in the winter of 2020-2021 through the study. The first positive RSV swab collected through the sentinel network in England was collected in ISO week 17 and then every week since ISO week 25. In total, 16 (N=248, 6.5%) of the virology-sampling practices volunteered to participate; these were high-sampling practices collecting the majority of eligible swabs across the sentinel network-200 (43.8%) out of 457 swabs, of which 54 (N=200, 27%) were positive for RSV. Measures to control the COVID-19 pandemic meant there was no circulating RSV last winter; however, RSV has circulated out of season, as detected by the sentinel network. The sentinel network practices have collected 40% (200/500) of the required samples, and 27% (54/200) were RSV positive. We have demonstrated the feasibility of implementing a European-standardized RSV disease burden study protocol in England during a pandemic, and we now need to recruit to this adapted protocol. DERR1-10.2196/38026.


Effect assessment in work environment interventions: a methodological reflection.
This paper addresses a number of issues for work environment intervention (WEI) researchers in light of the mixed results reported in the literature. If researchers emphasise study quality over intervention quality, reviews that exclude case studies with high quality and multifactorial interventions may be vulnerable to 'quality criteria selection bias'. Learning from 'failed' interventions is inhibited by both publication bias and reporting lengths that limit information on relevant contextual and implementation factors. The authors argue for the need to develop evaluation approaches consistent with the complexity of multifactorial WEIs that: a) are owned by and aimed at the whole organisation; and b) include intervention in early design stages where potential impact is highest. Context variety, complexity and instability in and around organisations suggest that attention might usefully shift from generalisable 'proof of effectiveness' to a more nuanced identification of intervention elements and the situations in which they are more likely to work as intended. STATEMENT OF RELEVANCE: This paper considers ergonomics interventions from perspectives of what constitutes quality and 'proof". It points to limitations of traditional experimental intervention designs and argues that the complexity of organisational change, and the need for multifactorial interventions that reach deep into work processes for greater impact, should be recognised.


Interannual changes in zooplankton echo subtropical and high latitude climate effects in the southern East China Sea.
Climate variability plays a central role in the dynamics of marine pelagic ecosystems shaping the structure and abundance changes of plankton communities, thereby affecting energy pathways and biogeochemical fluxes in the ocean. Here we have investigated complex interactions driven a climate-hydrology-plankton system in the southern East China Sea over the period 2000 to 2012. In particular, we aimed at quantifying the influence of climate phenomena playing out in tropical (El Nino 3.4) and middle-high latitudes (East Asia Winter Monsoon, EAWM, and Pacific Decadal Oscillation, PDO) on pelagic copepods. We found that the EAWM and El Nino 3.4 showed a non-stationary and non-linear relationship with local temperature variability. In the two cases, the strength of the relationship, as indexed by the wavelet coherence analysis, decreased along with the positive phase of the PDO. Likewise, the influence of EAWM and El Nino3.4 on copepods exhibited a non-stationary link that changed along with the PDO state. Indeed, copepods and EAWM were closely related during the positive phase, while the link copepods-El Nino 3.4 was stronger during the negative phase. Our results pointed out cascading effects from climate to plankton driven by the positive phase of the PDO through its effect on temperature conditions, and likely through a larger southward transport of nutrient-rich water masses to northern Taiwan and the Taiwan Strait. We suggest a chain of mechanisms whereby the PDO shapes interannual dynamics of pelagic copepods and highlight that these results have implications for integrative management measures, as pelagic copepods plays a prominent role in food web dynamics and for harvested fish in the East China Sea.


Advantages of genome sequencing by long-read sequencer using SMRT technology in medical area.
PacBio RS II is the first commercialized third-generation DNA sequencer able to sequence a single molecule DNA in real-time without amplification. PacBio RS II's sequencing technology is novel and unique, enabling the direct observation of DNA synthesis by DNA polymerase. PacBio RS II confers four major advantages compared to other sequencing technologies: long read lengths, high consensus accuracy, a low degree of bias, and simultaneous capability of epigenetic characterization. These advantages surmount the obstacle of sequencing genomic regions such as high/low G+C, tandem repeat, and interspersed repeat regions. Moreover, PacBio RS II is ideal for whole genome sequencing, targeted sequencing, complex population analysis, RNA sequencing, and epigenetics characterization. With PacBio RS II, we have sequenced and analyzed the genomes of many species, from viruses to humans. Herein, we summarize and review some of our key genome sequencing projects, including full-length viral sequencing, complete bacterial genome and almost-complete plant genome assemblies, and long amplicon sequencing of a disease-associated gene region. We believe that PacBio RS II is not only an effective tool for use in the basic biological sciences but also in the medical/clinical setting.


Concomitant fungal and Mycobacterium bovis infections in beef cattle in Kenya.
Bovine tuberculosis is an important zoonosis and accurate diagnosis is important for its surveillance. Post-mortem diagnosis may, however, be compromised by lesions caused by other pathogens. In an investigation on its prevalence in slaughter cattle in Kenya, Mycobacterium bovis and dimorphic fungi were inadvertently identified separately or concurrently in tuberculous lesions. Beef carcasses were inspected for lesions in two abattoirs in Nairobi. Tissues with lesions were collected and transported to the laboratory. Smears of lesions were stained by acid-fast procedure and examined microscopically. Lesions were cultured in Löwenstein-Jensen (LJ) and in BBL™ Mycobacterium growth indicator tubes (MGIT) media. Mycobacteria isolates in LJ medium were identified by DNA typing. Smears of BBLTM MGIT cultures were acid-fast stained and examined microscopically. Tissue sections were stained with periodic acid-Schiff reagent before examination. Of the 929 carcasses examined, 176 had granulomatous lesions. Dimorphic fungi were detected as acid-fast negative cells in 58 (32.9; 33.5%) of the lesion smears, either alone (29.0; 16.4%) or concurrently with acid-fast bacilli (29.0; 16.4%). The fungi were also detected in some BBL TM MGIT-culture smears and lesioned tissue sections. The fungi were identified, by means of cellular morphology, as Paracoccidioides brasiliensis and Blastomyces dermatitidis. A total of 64 isolates of mycobacteria were recovered in LJ medium, 19 of which were identified as M. bovis. The present report documents native P. brasiliensis infections outside the presumed endemic region and B. dermatitidis infections in a livestock animal. The findings further indicate the importance of dimorphic fungi as a differential diagnosis of bovine tuberculosis in the region.


Preventive treatment in migraine and the new US guidelines.
Migraine headaches are among the most common headache disorders seen in various practices. The prevalence of migraine headaches is 18% in women and 6% in men. While millions of Americans suffer from migraine headaches, roughly 3%-13% of identified migraine patients are on preventive therapy, while an estimated 38% actually need a preventive agent. The challenge among physicians is not only when to start a daily preventive agent but which preventive agent to choose. Circumstances warranting prevention have been described in the past, and in 2012, a new set of guidelines with an evidence review on preventive medications was published. A second set of guidelines provided evidence on nonsteroidal anti-inflammatory drugs, herbs, minerals, and vitamins for prevention of episodic migraine. This article describes the updated US guidelines for the prevention of migraines and also outlines the major studies from which these guidelines were derived.


Mutation analysis of novel human liver-related putative tumor suppressor gene in hepatocellular carcinoma.
To find the point mutations meaningful for inactivation of liver-related putative tumor suppressor gene (LPTS) gene, a human novel liver-related putative tumor suppressor gene and telomerase inhibitor in hepatocellular carcinoma. The entire coding sequence of LPTS gene was examined for mutations by single strand conformation polymorphism (SSCP) assay and PCR products direct sequencing in 56 liver cancer cell lines, 7 ovarian cancer and 7 head neck tumor cell lines and 70 pairs of HCC tissues samples. The cDNA fragment coding for the most frequent mutant protein was subcloned into GST fusion expression vector. The product was expressed in E.coli and purified by glutathione-agarose column. Telomeric repeat amplification protocol (TRAP) assays were performed to study the effect of point mutation to telomerase inhibitory activity. SSCP gels showed the abnormal shifting bands and DNA sequencing found that there were 5 different mutations and/or polymorphisms in 12 tumor cell lines located at exon2, exon5 and exon7. The main alterations were A(778)A/G and A(880)T in exon7. The change in site of 778 could not be found in HCC tissue samples, while the mutation in position 880 was seen in 7 (10 %) cases. The mutation in the site of 880 had no effect on telomerase inhibitory activity. Alterations identified in this study are polymorphisms of LPTS gene. LPTS mutations occur in HCC but are infrequent and of little effect on the telomerase inhibitory function of the protein. Epigenetics, such as methylation, acetylation, may play the key role in inactivation of LPTS.


Persistence of biomarker ATP and ATP-generating capability in bacterial cells and spores contaminating spacecraft materials under earth conditions and in a simulated martian environment.
Most planetary protection research has concentrated on characterizing viable bioloads on spacecraft surfaces, developing techniques for bioload reduction prior to launch, and studying the effects of simulated martian environments on microbial survival. Little research has examined the persistence of biogenic signature molecules on spacecraft materials under simulated martian surface conditions. This study examined how endogenous adenosine-5'-triphosphate (ATP) would persist on aluminum coupons under simulated martian conditions of 7.1 mbar, full-spectrum simulated martian radiation calibrated to 4 W m(-2) of UV-C (200 to 280 nm), -10 degrees C, and a Mars gas mix of CO(2) (95.54%), N(2) (2.7%), Ar (1.6%), O(2) (0.13%), and H(2)O (0.03%). Cell or spore viabilities of Acinetobacter radioresistens, Bacillus pumilus, and B. subtilis were measured in minutes to hours, while high levels of endogenous ATP were recovered after exposures of up to 21 days. The dominant factor responsible for temporal reductions in viability and loss of ATP was the simulated Mars surface radiation; low pressure, low temperature, and the Mars gas composition exhibited only slight effects. The normal burst of endogenous ATP detected during spore germination in B. pumilus and B. subtilis was reduced by 1 or 2 orders of magnitude following, respectively, 8- or 30-min exposures to simulated martian conditions. The results support the conclusion that endogenous ATP will persist for time periods that are likely to extend beyond the nominal lengths of most surface missions on Mars, and planetary protection protocols prior to launch may require additional rigor to further reduce the presence and abundance of biosignature molecules on spacecraft surfaces.


Hepatic immunohistochemistry of bile transporters in progressive familial intrahepatic cholestasis.
Background. Diagnosis of progressive familial intrahepatic cholestasis (PFIC) is a challenging matter that involves the summation of clinical, laboratory, radiological, and liver histological parameters; in addition to specific investigations to exclude other causes of neonatal cholestasis. The aim of this study was to evaluate liver tissue immunohistochemistry of bile salt export pump (BSEP) and multidrug resistance 3 (MDR3) proteins in differentiating PFIC from other causes of neonatal cholestasis, particularly, when genotyping is unavailable. The study included 25 patients diagnosed phenotypically as PFIC including 2 with PFIC1, 17 with PFIC2 and 6 with PFIC3. A second group of 25 cholestatic newborns with confirmed etiologies other than PFIC, termed as non-PFIC, included as controls. Liver biopsies from all patients were obtained and immunostained for BSEP and MDR3. Negative immunoreaction of BSEP and MDR3 was found in the majority of PFIC group (76 and 64% respectively). Nonetheless, the negative immunoreaction was demonstrated in a considerable number of the non-PFIC group. BSEP immunoreaction was negative in the majority (82.4%) of PFIC2 but in none of the two patients with PFIC1. In addition, negative MDR3 immunoreaction was more frequently associated with PFIC3 compared to non-PFIC group. MDR3 and BSEP immunostaining would be a helpful tool in supporting the phenotypic diagnosis of PFIC subtypes and in differentiating PFIC from other causes of neonatal cholestasis.


The Keap1/Nrf2 pathway in health and disease: from the bench to the clinic.
The transcription factor nuclear factor-erythroid 2 p45-related factor 2 (Nrf2, with gene called NFE2L2) is a master regulator of the antioxidant response. In the last decade, interest has intensified in this research area as its importance in several physiological and pathological processes has become widely recognized; these include redox signalling and redox homoeostasis, drug metabolism and disposition, intermediary metabolism, cellular adaptation to stress, chemoprevention and chemoresistance, toxicity, inflammation, neurodegeneration, lipogenesis and aging. Regulation of Nrf2 is complex and although much attention has focussed on its repression by Kelch-like ECH-associated protein-1 (Keap1), recently it has become increasingly apparent that it is also controlled by cross-talk with other signalling pathways including the glycogen synthase kinase-3 (GSK-3)-β-transducin repeat-containing protein (β-TrCP) axis, ERAD (endoplasmic reticulum-associated degradation)-associated E3 ubiquitin-protein ligase (Hrd1, also called synoviolin), nuclear factor-kappa B (NF-κB), Notch and AMP kinase. Due to its beneficial role in several diseases, Nrf2 has become a major therapeutic target, with novel natural, synthetic and targeted small molecules currently under investigation to modulate the pathway and in clinical trials.


Cyanoacrylate microbial sealants for skin preparation prior to surgery.
Surgical site infections are a continuing concern in health care. Microbial sealant is a liquid applied to the skin immediately before surgery. It is thought to contribute to reducing surgical site infections by sealing in the skin flora to prevent contamination and infection of the surgical site. To assess the effects of the preoperative application of microbial sealants (compared with no microbial sealant) on the rates of surgical site infection in people undergoing clean surgery. We searched the Cochrane Wounds Group Specialised Register (searched 10 May 2010), the Cochrane Central Register of Controlled Trials (CENTRAL) (The Cochrane Library 2010, Issue 2), Ovid MEDLINE (1950 to April Week 3 2010), Ovid MEDLINE - In-Process & Other Non-Indexed Citations (searched 10 May 2010), Ovid EMBASE (1980 to 2010 Week 18) and EBSCO CINAHL (1982 to 10 May 2010). We searched bibliographies and contacted manufacturers of microbial sealants for unpublished studies. There were no restrictions based on language, date or publication status. Randomised controlled trials (RCTs) were eligible for inclusion if they involved people undergoing clean surgery in an operating theatre and compared the use of preoperative microbial sealants with no microbial sealant. All review authors independently extracted data on the characteristics, risk of bias and outcomes of the eligible trial. One small trial (177 participants undergoing hernia repair) met the inclusion criteria. There was no statistically significant difference in the rates of surgical site infection (three patients in the control group developed a surgical site infection compared with none in the intervention group; risk ratio (RR) 0.17, 95% CI 0.01 to 3.19, P = 0.23). There is currently insufficient evidence as to whether the use of microbial sealants reduces the risk of surgical site infection in people undergoing clean surgery and further rigorous RCTs are required.


A systematic review of interventions for anxiety in people with HIV.
People with human immunodeficiency virus (HIV) show elevated anxiety levels compared to the general population. Anxiety can predate HIV infection or be triggered by HIV diagnosis and the many stresses that emerge during the course of HIV disease. Many psychological and pharmacological therapies have been shown to treat anxiety in the general population but a systematic understanding of which interventions have been tested in and are effective with HIV-seropositive individuals is needed. This review examines all published intervention studies on anxiety and HIV from 1980 to 2009 covered by the databases MedLine (1980-2009) and PsycINFO (1980-2009) for a definitive account of effectiveness of interventions and an indication of prevalence of HIV-related anxiety and measurement within studies. Standard systematic research methods were used to gather quality published papers on HIV and anxiety, searching published data bases according to quality inclusion criteria. From the search, 492 papers were generated and hand searched resulting in 39 studies meeting adequacy inclusion criteria for analysis. Of these, 30 (76.9%) were implemented in North America (the USA and Canada), with little representation from developing countries. Thirty-three (84.6%) studies recruited only men or mostly men. A total of 50 interventions were investigated by the 39 studies; 13 targeted HIV, symptoms or associated outcomes/conditions, 20 directly targeted anxiety and another 17 indirectly targeted anxiety. Twenty-four (48%) interventions were effective in reducing anxiety (including 11 indirect interventions), 16 (32%) were ineffective and 10 (20%) had an unknown effect on anxiety. Sixty-five percent of interventions directly targeting anxiety were effective. Psychological interventions (especially cognitive behavioural stress management interventions and cognitive behavioural therapy) were generally more effective than pharmacological interventions. Only three studies provided prevalence rates - these ranged from 13% to 80%. Anxiety was measured using 16 different instruments. Our detailed data suggest that interventions are both effective and available, although further research into enhancing efficacy would be valuable. Also, the vast majority of studies were Western-based, no studies looked at children or adolescents and few looked specifically at women. An international effort to harmonise measurement of anxiety is also missing. There is a need to routinely log anxiety in those with HIV infection during the course of their disease, to provide specific data on women, young people and those in diverse geographic areas and incorporate management into care protocols.


Evaluation of a new electronic spirometer: the vitalograph "Escort" spirometer.
The "Escort" spirometer is a lightweight, hand held spirometer employing a Fleisch pneumotachograph. Measurements of forced expiratory volume in one second (FEV1), forced vital capacity (FVC), and peak expiratory flow (PEF) are obtained from a single FVC manoeuvre. Results are displayed on a small liquid crystal display, but there is no graphical display. The performance of the Escort spirometer has been compared with that of a wedge bellows spirometer (Vitalograph S model) and a Wright PEF meter. One hundred and thirteen subjects performed three FVC manoeuvres on the wedge bellows and Escort spirometers and three PEF manoeuvres on the Wright meter. The best reading for each index was recorded. In 21 of the subjects comparison of a Wright manoeuvre with an FVC manoeuvre on the Escort spirometer was performed, whilst in three subjects the effect of repeated blows was studied. The FEV1 ranged from 0.5 to 5.4 litres, FVC from 1.05 to 6.2 litres, and PEF from 100 to 725 l/min. The mean (SD) difference for the FEV1 was -0.05 (0.15) (95% confidence interval (95% CI) -0.07 to -0.02) litres, for FVC 0.03 (0.28) (95% CI -0.02 to +0.08) litres, and for PEF 1.68 (50.6) (95% CI -7.7 to +11.1) l/min. The differences were positively correlated with the mean reading for PEF and FVC but not for FEV1. The Wright PEF manoeuvre performed on the Escort produced significantly higher PEF readings (mean difference -22.9 litres). There was no significant effect of repeated FVC manoeuvres on any of the indices. The Escort spirometer compares extremely well with a wedge bellows spirometer for measurement of FEV1 and FVC, whilst yielding results of PEF from an FVC manoeuvre which are comparable to those obtained from a Wright meter. It can be recommended for use as a portable hand held spirometer.


Recent Development in Big Data Analytics for Business Operations and Risk Management.
"Big data" is an emerging topic and has attracted the attention of many researchers and practitioners in industrial systems engineering and cybernetics. Big data analytics would definitely lead to valuable knowledge for many organizations. Business operations and risk management can be a beneficiary as there are many data collection channels in the related industrial systems (e.g., wireless sensor networks, Internet-based systems, etc.). Big data research, however, is still in its infancy. Its focus is rather unclear and related studies are not well amalgamated. This paper aims to present the challenges and opportunities of big data analytics in this unique application domain. Technological development and advances for industrial-based business systems, reliability and security of industrial systems, and their operational risk management are examined. Important areas for future research are also discussed and revealed.


Volunteering as a Meaningful Occupation in the Process of Recovery From Serious Mental Illness: A Qualitative Study.
Volunteering is an activity that facilitates social welfare, a sense of connection to others, and the construction of links with other people and the community. To describe the experience of a group of people with serious mental illness (SMI) who engaged in community volunteer work. Qualitative, phenomenological study using purposeful sampling and an inductive thematic analysis. Several mental health community centers in Madrid and Catalonia, Spain. People with SMI engaged in community volunteer work. Outcomes and Measures: In-depth interviews were used to collect data. Two themes, with four subthemes each, emerged from the data: (1) engaging in a meaningful occupation enhances the recovery process (subthemes: undergoing positive experiences, performing or resuming a meaningful occupation, living in the present moment, and relating with one's social and family environment) and (2) a process of mutual support (subthemes: in helping others, you help yourself; helping others on the basis of one's first-person experience; recover before you can help others; and role of professionals in volunteering). Volunteering is perceived as a meaningful occupation. Helping others generates a reciprocal benefit that favors the recovery process and contributes to the establishment of personal and social bonds. What This Article Adds: This study provides a basis for understanding the potential of volunteering to be a meaningful occupation in the recovery process of people with SMI. It can help occupational therapy practitioners develop volunteer-based intervention programs that support the recovery process.


Vaginal Myomectomy and Total Vaginal Hysterectomy on Large Prolapsed Cervical Myoma: A Case Report.
Cervical myomas are benign tumors originating from cervical muscle tissue with a very rare incidence of only about 8% of all myomas. The surgical approach depends on the position of cervical myoma. This case report discusses a 44-year-old woman who complained of a lump discharge from her birth canal 6 months ago, and currently discharging from her vagina. We performed vaginal myomectomy, and the cervical myoma measuring 8 × 8 × 6 cm with solid consistency was removed. We continued with total vaginal hysterectomy. Post-operative recovery was progressing well. The histopathology report was consistent with leiomyoma. Large prolapsed cervical myoma can be disturbing and discomforting for many patients. It is relatively rare and can be successfully removed vaginally with minimal morbidity.


Tricyclic and related drugs for nocturnal enuresis in children.
Enuresis (bedwetting) affects up to 20% of five year-olds and 2% of adults. Although spontaneous remission often occurs, the social, emotional and psychological costs can be great. Tricyclics have been used to treat enuresis since the 1960s. To assess the effects of tricyclic and related drugs compared with other interventions for treating children with enuresis. We searched the Cochrane Incontinence Group Specialised Trials Register (containing trials identified from the Cochrane Central Register of Controlled Trials (CENTRAL), MEDLINE, MEDLINE in process, ClinicalTrials.gov, WHO ICTRP and handsearching of journals and conference proceedings), on 30 November 2015, and reference lists of relevant articles. We included all randomised and quasi-randomised trials comparing a tricyclic or related drug with another intervention for treating enuresis. We also included combination therapies that included tricyclics. We excluded trials for treating daytime wetting. Two review authors independently assessed the quality of the eligible trials, and extracted data. We settled differences by discussion with a third review author. Sixty-four trials met the inclusion criteria, involving 4071 children. The quality of many trials was poor, with comparisons addressed by single studies. Minor adverse effects were common, and reported in 30 trials. These included dizziness, headache, mood changes, gastrointestinal discomforts and neutropenia. More serious side-effects can occur but were not reported. Seven trials reported no adverse effects.Tricyclics are more effective than placebo, particularly for short-term outcomes. Compared to placebo, imipramine resulted in one fewer wet nights per week (mean difference (MD) -0.95, 95% confidence interval (CI) -1.40 to -0.50; 4 trials, 347 children), with fewer failing to achieve 14 consecutive dry nights (78% versus 95% for placebo, RR 0.74, 95% CI 0.61 to 0.90; 12 trials, 831 children). Amitriptyline and desipramine were more effective than placebo, but nortriptyline and mianserin showed no difference. Most tricyclics did not have a sustained effect after ceasing treatment, with 96% wetting at follow-up for imipramine versus 97% for placebo.Imipramine combined with oxybutynin is also more effective than placebo, with 33% failing to achieve 14 consecutive dry nights at the end of treatment versus 78% for placebo (RR 0.43, 95% CI 0.23 to 0.78; 1 trial, 47 children) and 45% wetting at follow-up versus 79% for placebo (RR 0.58, 95% CI 0.34 to 0.99; 1 trial, 36 children).There was insufficient evidence to judge the effect between different doses of tricyclics, and between different tricyclics. Treatment outcomes between tricyclic and desmopressin were similar, but were mixed when tricyclic was compared with an anticholinergic. However, when imipramine was compared with desmopressin plus oxybutynin (1 trial, 45 children), the combination therapy was more effective, with one fewer wet nights per week (MD 1.07, 95% CI 0.06 to 2.08) and 36% failing to achieve 14 consecutive dry nights versus 87% for imipramine (RR 2.39, 95% CI 1.35 to 4.25). Tricyclics were also more effective or showed no difference in response when compared to other drugs which are no longer used for enuresis.Tricyclics were less effective than alarms. Although there was no difference in the number of wet nights, 67% failed to achieve 14 consecutive dry nights for imipramine versus only 17% for alarms (RR 4.00, 95% CI 1.06 to 15.08; 1 trial, 24 children). Alarm therapy also had a more sustained effect after ceasing treatment with 100% on imipramine versus 58% on alarms wetting at follow-up (RR 1.67, 95% CI 1.03 to 2.69; 1 trial, 24 children).Imipramine was more effective than simple behavioural therapies during treatment, with one fewer wet nights per week compared with star chart plus placebo (MD -0.80, 95% CI -1.33 to -0.27; 1 trial, 250 children). At follow-up 40% were wet with imipramine versus 80% with fluids and avoiding punishment (RR 0.50, 95% CI 0.28 to 0.89; 1 trial, 40 children). However, imipramine was less effective than complex behavioural therapies, with 61% failing to achieve 14 consecutive dry nights for imipramine versus 33% for the three-step programme (RR 1.83, 95% CI 1.08 to 3.12; 1 trial, 72 children) and 16% for the three-step programme combined with motivational therapy and computer-led education (RR 3.91, 95% CI 2.30 to 6.66; 1 trial, 132 children) at the end of treatment, with similar results at follow-up.Tricyclics were more effective than restricted diet, with 99% failing to achieve 14 consecutive dry nights versus 84% for imipramine (RR 0.84, 95% CI 0.75 to 0.93; 1 trial, 147 children).There was insufficient evidence to judge the effect of tricyclics compared to the other miscellaneous interventions studied.At the end of treatment there were about two fewer wet nights for imipramine plus oxybutynin compared with imipramine monotherapy (MD -2.10, 95% CI -2.99 to -1.21; 1 trial, 63 children) and 48% on imipramine plus oxybutynin failed to achieve 14 consecutive dry nights compared with 74% on imipramine monotherapy (RR 0.68, 95% CI 0.50 to 0.92; 2 trials, 101 children). At follow-up, 45% on imipramine plus oxybutynin were wetting versus 83% on imipramine monotherapy (RR 0.55, 95% CI 0.32 to 0.92; 1 trial, 36 children).When imipramine combined with desmopressin was compared with imipramine monotherapy, there was no difference in outcomes. However, when imipramine plus desmopressin was compared with desmopressin monotherapy, the combination was more effective, with 15% not achieving 14 consecutive dry nights at the end of treatment for imipramine plus desmopressin versus 40% for desmopressin monotherapy (RR 0.38, 95% CI 0.17 to 0.83; 1 trial, 86 children). Tricyclics combined with alarm therapy were not more effective than alarm monotherapy, alarm combined with desmopressin or alarm combined with nortriptyline. The addition of a tricyclic to other behavioural therapies did not alter treatment response. There was evidence that tricyclics are effective at reducing the number of wet nights during treatment, but do not have a sustained effect after treatment stops, with most children relapsing. In contrast, there was evidence that alarm therapy has better short- and long-term outcomes. There was some evidence that tricyclics combined with anticholinergics may be more effective that tricyclic monotherapy.


Contraction-sensitive skeletal muscle afferents inhibit arterial baroreceptor signalling in the nucleus of the solitary tract: role of intrinsic GABA interneurons.
Arterial baroreceptor and skeletal muscle receptor afferents relay sensory information to the nucleus of the solitary tract (NTS) during exercise. Previous studies have suggested that skeletal muscle afferent input inhibits baroreflex function; however, detailed information on the role of muscle afferents and GABAergic mechanisms in the NTS is limited. Furthermore, identification of specific afferent modalities that activate GABAergic neurons in the NTS remains unknown. In the present study, we examined the neuroanatomical and physiological interactions between spinal dorsal horn cells that transmit contraction-sensitive input from skeletal muscle and GABAergic interneurons in the NTS. Biotinylated dextran amine (BDA, 10%, 25-100 nL) microinjection into dorsal horn of the cervical spinal cord was combined with glutamate decarboxylase (GAD) immunohistochemistry to visualize the nature of the relationship of BDA-labeled fibers in the NTS with GAD immunoreactivity (GAD-ir). BDA-labeled axons and terminal processes were localized in the medial, commissural, dorsomedial and dorsolateral subdivisions of the caudal NTS. Moreover, BDA-labeled fibers were observed in close proximity to GAD-ir structures throughout these regions of the NTS. The physiological interaction between skeletal muscle receptor and arterial baroreceptor afferents was investigated using an arterially perfused, decerebrate rat preparation. Activation of skeletal muscle afferents by electrically evoked twitch contraction of the forelimb attenuated baroreflex responsiveness (BR, calculated as the ratio of changes in heart rate to systemic pressure) from -1.5+/-0.3 bpm.mm Hg(-1) to -0.1+/-0.1 bpm.mm Hg(-1) (control versus contraction, P<0.05, n=15). However, forelimb contraction failed to inhibit the reflex bradycardia evoked by activation of peripheral chemoreceptor afferents, indicating a reflex-specific action. Bilateral microinjection of bicuculline methiodide (BIC, 10 microM, 40-60 nL) into the caudal NTS restored baroreflex responsiveness during contraction (-1.6+/-0.2 versus -0.1+/-0.1 versus -1.5+/-0.2 bpm.mmHg(-1), control versus contraction versus contraction+BIC P<0.05, n=8). We conclude that activation of ascending spinal neurons from the cervical dorsal horn by contraction-sensitive skeletal muscle afferents selectively inhibits arterial baroreceptor signaling in the NTS via activation of a GABAergic mechanism.


Identification of target-binding peptide motifs by high-throughput sequencing of phage-selected peptides.
High-throughput sequencing was previously applied to phage-selected peptides in order to gain insight into the abundance and diversity of isolated peptides. Herein we developed a procedure to efficiently compare the sequences of large numbers of phage-selected peptides for the purpose of identifying target-binding peptide motifs. We applied the procedure to analyze bicyclic peptides isolated against five different protein targets: sortase A, urokinase-type plasminogen activator, coagulation factor XII, plasma kallikrein and streptavidin. We optimized sequence data filters to reduce biases originating from the sequencing method and developed sequence correction algorithms to prevent identification of false consensus motifs. With our strategy, we were able to identify rare target-binding peptide motifs, as well as to define more precisely consensus sequences and sub-groups of consensus sequences. This information is valuable to choose peptide leads for drug development and it facilitates identification of epitopes. We furthermore show that binding motifs can be identified after a single round of phage selection. Such a selection regimen reduces propagation-related bias and may facilitate application of phage display in non-specialized laboratories, as procedures such as bacterial infection, phage propagation and purification are not required.


Analysis of Refractive Index Distributions in Cylindrical, Graded-Index Glass Rods (GRIN Rods) Used as Image Relays.
Cylindrical transparent media whose refractive index decreases with increasing cylinder radius have been investigated in connection with coherent light propagation in gas waveguides. Recently, graded index glass rods (trade named SELFOC rods) have been used as imaging devices. We report here on a geometrical optical study of graded index systems used for relaying images at unit magnification. We have found that two index distributions previously studied result in large image aberrations when the presence of skew rays is taken into account. We have derived an index distribution which is "ideal" for helical skew rays. Using ray tracing methods we have examined image aberrations for various index distributions and for various rod geometries. We find that (1) no one refractive index distribution can be "ideal" for both meridional and skew rays, (2) image resolution is generally low, reaching about 1000 spots per field, and (3) the optimal index distribution varies with the ratio of rod length to radius and the relative aperture and is intermediate between the helically ideal and the meridionally ideal distributions.


High-dose megestrol acetate therapy of ovarian carcinoma: a phase II study by the Northern California Oncology Group.
The activity of high-dose megestrol acetate was studied in 47 patients with epithelial ovarian cancers after failure of initial chemotherapy. The dose of megestrol acetate was 800 mg/d orally (PO) for 4 weeks and then 400 mg/d until tumor progression. Patients generally had far-advanced disease. Prior therapy included cisplatin, doxorubicin, and cyclophosphamide (PAC) or other cisplatin-containing regimens in 37, other combinations in eight, and single agents in only two patients. Seventeen patients (36%) developed intestinal obstructions within the first 2 months on study. Tumor histology was serous in 37, endometrioid in six, and clear-cell in two. Two thirds of the tumors were histologic grade 3, and the others were grade 2. Complete remission was obtained in one patient, with time to progression of 4 months. There were three partial remissions, with times to progression of 4, 5, and 18 months. The overall response rate (complete and partial) was 8%. Three additional patients had minor remissions (3, 5, and 8 months), and five had stable disease, for 3, 4, 5, 6, and 9 months. There was no correlation of response with grade, histologic type, or site of disease, but responding patients had a longer survival from diagnosis to protocol entry and from protocol failure to death than did nonresponding patients. The major side effect of megestrol acetate was increased appetite, which caused one patient to withdraw from the study, and resulted in a 10- to 20-kg weight gain in five patients. Plasma levels of megestrol acetate averaged 600 ng/mL in the first month of therapy and decreased to approximately 400 ng/mL at 8 and 12 weeks, after the drug dosage had been reduced. Serum follicle-stimulating hormone (FSH) and luteinizing hormone (LH) levels were markedly lower during megestrol therapy compared with pretreatment values. Megestrol acetate at 1 microgram/mL in vitro inhibited soft agar colony formation from one of 17 specimens of ovarian carcinomas. We conclude that megestrol acetate in high doses has modest, but definite, palliative effects in some patients with advanced ovarian carcinoma in whom chemotherapy has failed. A controlled trial of megestrol plus combination chemotherapy as first-line treatment of advanced ovarian carcinoma should be considered.


Multiple-micronutrient supplementation: Evidence from large-scale prenatal programmes on coverage, compliance and impact.
Micronutrient deficiencies during pregnancy pose important challenges for public-health, given the potential adverse outcomes not only during pregnancy but across the life-course. Provision of iron-folic acid (IFA) supplements is the strategy most commonly practiced and recommended globally. How to successfully implement IFA and multiple micronutrient supplementation interventions among pregnant women and to achieve sustainable/permanent solutions to prenatal micronutrient deficiencies remain unresolved issues in many countries. This paper aims to analyse available experiences of prenatal IFA and multiple micronutrient interventions to distil learning for their effective planning and large-scale implementation. Relevant articles and programme-documentation were comprehensively identified from electronic databases, websites of major-agencies and through hand-searching of relevant documents. Retrieved documents were screened and potentially relevant reports were critically examined by the authors with the aim of identifying a set of case studies reflecting regional variation, a mix of implementation successes and failures, and a mix of programmes and large-scale experimental studies. Information on implementation, coverage, compliance, and impact was extracted from reports of large-scale interventions in Central America, Southeast Asia, South Asia, and Sub-Saharan Africa. The WHO/CDC Logic-Model for Micronutrient Interventions in Public Health was used as an organizing framework for analysing and presenting the evidence. Our findings suggest that to successfully implement supplementation interventions and achieve sustainable-permanent solutions efforts must focus on factors and processes related to quality, cost-effectiveness, coverage, utilization, demand, outcomes, impacts, and sustainability of programmes including strategic analysis, management, collaborations to pilot a project, and careful monitoring, midcourse corrections, supervision and logistical-support to gradually scaling it up.


Optimization of a femtosecond ellipsometer for gold photoreflectance studies.
I develop a quantitative optimization procedure for femtosecond reflectance ellipsometer measurements. The femtosecond ellipsometer eliminates the need for separate thin-film reflectance and transmittance measurements and allows self-consistent determination of dielectric index changes of optically thick films on arbitrary substrates. I verify the optimization procedure and use the femtosecond ellipsometric measurements to investigate the photoreflectance of an optically thick gold film. A comparison of the extracted real and imaginary dielectric function components reveals evidence of thermalized and nonthermalized hot-electron populations.


Emergency surgery data and documentation reporting forms for sudden-onset humanitarian crises, natural disasters and the existing burden of surgical disease.
Following large-scale disasters and major complex emergencies, especially in resource-poor settings, emergency surgery is practiced by Foreign Medical Teams (FMTs) sent by governmental and non-governmental organizations (NGOs). These surgical experiences have not yielded an appropriate standardized collection of data and reporting to meet standards required by national authorities, the World Health Organization, and the Inter-Agency Standing Committee's Global Health Cluster. Utilizing the 2011 International Data Collection guidelines for surgery initiated by Médecins Sans Frontières, the authors of this paper developed an individual patient-centric form and an International Standard Reporting Template for Surgical Care to record data for victims of a disaster as well as the co-existing burden of surgical disease within the affected community. The data includes surgical patient outcomes and perioperative mortality, along with referrals for rehabilitation, mental health and psychosocial care. The purpose of the standard data format is fourfold: (1) to ensure that all surgical providers, especially from indigenous first responder teams and others performing emergency surgery, from national and international (Foreign) medical teams, contribute relevant and purposeful reporting; (2) to provide universally acceptable forms that meet the minimal needs of both national authorities and the Health Cluster; (3) to increase transparency and accountability, contributing to improved humanitarian coordination; and (4) to facilitate a comprehensive review of services provided to those affected by the crisis.


Nurses' Perceptions of Workload Burden in Pediatric Critical Care.
Quantifying nurses' perceptions of workload burden when managing critically ill patients is essential for designing interventions to ease nurses' workday. To explore pediatric intensive care unit (PICU) nurses' perceptions of their workload when caring for critically ill patients and managing protocolized therapies. This study was embedded in a multicenter randomized clinical trial where participants were assigned to receive either lower-target or higher-target glucose control. Nurses from 35 participating PICUs completed a baseline survey containing questions about their perceptions of PICU workload in general. They completed an intervention survey after caring for a study patient. Two workload measurement instruments, the Subjective Workload Assessment Technique (SWAT) and the National Aeronautics and Space Administration-Task Load Index (NASA-TLX), were embedded in these surveys. Baseline surveys were completed by 1476 PICU nurses, predominantly female with a bachelor's degree and a median (interquartile range) of 6 (3-11) years of nursing experience and 4 (2-9) years of PICU experience. Most nurses (65%) rated time burden as the most important component of their workload, followed by cognitive (22%) or psychological stress (13%) burden. Work performance was selected most often as contributing to workload, followed by cognitive demand, time pressure, effort, and physical demand. Intervention surveys were completed by 73% of enrolled participants (505 of 693). Nurses managing the lower glucose target group reported higher levels of workload burden as measured by the SWAT (P = .002) and NASA-TLX (P < .001). This study describes the workload burden perceived by PICU nurses when managing critically ill patients in general and when managing protocolized therapies.


Imaging activation of peptidergic spinal afferent varicosities within visceral organs using novel CGRPα-mCherry reporter mice.
In vertebrates, visceral pain from internal organs is detected by spinal afferents, whose cell bodies lie in dorsal root ganglia (DRG). Until now, all recordings from spinal afferents have been restricted to recording transmission of action potentials along axons, or from cell bodies lying outside their target organ, which is not where sensory transduction occurs. Our aim was to record directly from a major class of spinal afferent within visceral organs, where transduction of sensory stimuli into action potentials occurs. Using novel calcitonin gene-related peptide (CGRP)α reporter mice, DRG neurons expressed mCherry, including nerve axons within viscera. In colon, a minority of total CGRP immunoreactivity was attributed CGRPα. In isolated unstretched colon, calcium imaging from CGRPα-expressing varicose axons did not detect resolvable calcium transients. However, noxious levels of maintained circumferential stretch to the colon induced repetitive calcium transients simultaneously in multiple neighboring varicosities along single mCherry-expressing axons. Discrete varicosities could generate unitary calcium transients independently of neighboring varicosities. However, axons expressing mCherry only generated coordinated calcium transients when accompanied by simultaneous activation of multiple varicosities along that axon. Simultaneous imaging from different classes of myenteric neurons at the same time as mCherry-expressing axons revealed coordinated calcium transients in multiple myenteric neurons, independent of activity in mCherry-expressing axons. CGRPα-expressing axon terminals preferentially responded to heat, capsaicin, and low pH. We show that direct recordings can be made from the major class of peptidergic spinal afferent that contributes to visceral nociception. This approach can provide powerful insights into transduction of stimuli in viscera.


Safety and efficacy of hydromorphone as an analgesic alternative to morphine in acute pain: a randomized clinical trial.
We compare a standard weight-based dose of intravenous hydromorphone (Dilaudid) to a standard weight-based dose of intravenous morphine in adults presenting to the ED with acute severe pain. This was a prospective, randomized, double-blind, clinical trial conducted in an academic medical center. Of the 198 adult patients presenting to the ED with acute severe pain who were randomized to receive either intravenous hydromorphone at 0.015 mg/kg or intravenous morphine at 0.1 mg/kg, 191 patients had sufficient data for analysis. The main outcome measure was the difference between the 2 groups in pain reduction at 30 minutes as measured on a validated numeric rating scale. Adverse effects, pain reduction at 5 minutes and 2 hours postbaseline, and additional analgesics and antiemetics were tracked as secondary outcome measures. The mean change of pain from baseline to 30 minutes postbaseline in patients allocated to intravenous hydromorphone was -5.5 numeric rating scale units versus -4.1 in patients allocated to intravenous morphine (difference -1.3; 95% confidence interval -2.2 to -0.5). Adverse effects were similar in both groups, with the exception of pruritus, which did not occur in patients receiving hydromorphone (0% versus 6% [difference -6%; 95% confidence interval -11% to -1%]). No patient required naloxone. For the treatment of acute, severe pain in the emergency department, intravenous hydromorphone at 0.015 mg/kg represents a feasible alternative to intravenous morphine at 0.1 mg/kg.


Markers of insulin resistance and colorectal cancer mortality.
A link between insulin metabolism and colorectal cancer has been hypothesized, supported by a series of potential physiological mechanisms, and confirmed by a number of reports in experimental animals. However, the evidence in humans is limited and mostly indirect. The aim of the present report is to analyze whether individuals with a cluster of metabolic abnormalities associated with abnormalities in insulin metabolism experience higher mortality for colorectal cancer than those without this cluster of metabolic abnormalities. A total of 21,311 men and 15,991 women 20-69 years of age were followed-up for an average of 7 years as part of the Risk Factors and Life Expectancy Project, a pooling of a number of epidemiological studies conducted in Italy. Our analyses indicate that participants with high levels of blood glucose and a cluster of metabolic abnormalities linked to insulin resistance experienced a significant increased risk of colorectal cancer mortality compared with participants without the cluster. For the presence of the cluster of metabolic abnormalities, the calculated hazard ratios and 95% CIs were 2.96 (1.05-8.31) for men, 2.71 (0.59-12.50) for women, and 2.99 (1.27-7.01) when both sexes were combined. These associations were independent from the potential confounding effect of age, drinking of alcoholic beverages, and smoking. Our findings are supportive of the hypotheses that glucose metabolism hyperinsulinemia, insulin resistance, and metabolic abnormalities associated with it may play a significant role in the etiology of colorectal cancer.


Engineering Aspergillus niger for galactaric acid production: elimination of galactaric acid catabolism by using RNA sequencing and CRISPR/Cas9.
meso-Galactaric acid is a dicarboxylic acid that can be produced by the oxidation of D-galacturonic acid, the main constituent of pectin. Mould strains can be engineered to perform this oxidation by expressing the bacterial enzyme uronate dehydrogenase. In addition, the endogenous pathway for D-galacturonic acid catabolism has to be inactivated. The filamentous fungus Aspergillus niger would be a suitable strain for galactaric acid production since it is efficient in pectin hydrolysis, however, it is catabolizing the resulting galactaric acid via an unknown catabolic pathway. In this study, a transcriptomics approach was used to identify genes involved in galactaric acid catabolism. Several genes were deleted using CRISPR/Cas9 together with in vitro synthesized sgRNA. As a result, galactaric acid catabolism was disrupted. An engineered A. niger strain combining the disrupted galactaric and D-galacturonic acid catabolism with an expression of a heterologous uronate dehydrogenase produced galactaric acid from D-galacturonic acid. The resulting strain was also converting pectin-rich biomass to galactaric acid in a consolidated bioprocess. In the present study, we demonstrated the use of CRISPR/Cas9 mediated gene deletion technology in A. niger in an metabolic engineering application. As a result, a strain for the efficient production of galactaric acid from D-galacturonic acid was generated. The present study highlights the usefulness of CRISPR/Cas9 technology in the metabolic engineering of filamentous fungi.


RING Dimerization Links Higher-Order Assembly of TRIM5α to Synthesis of K63-Linked Polyubiquitin.
Members of the tripartite motif (TRIM) protein family of RING E3 ubiquitin (Ub) ligases promote innate immune responses by catalyzing synthesis of polyubiquitin chains linked through lysine 63 (K63). Here, we investigate the mechanism by which the TRIM5α retroviral restriction factor activates Ubc13, the K63-linkage-specific E2. Structural, biochemical, and functional characterization of the TRIM5α:Ubc13-Ub interactions reveals that activation of the Ubc13-Ub conjugate requires dimerization of the TRIM5α RING domain. Our data explain how higher-order oligomerization of TRIM5α, which is promoted by the interaction with the retroviral capsid, enhances the E3 Ub ligase activity of TRIM5α and contributes to its antiretroviral function. This E3 mechanism, in which RING dimerization is transient and depends on the interaction of the TRIM protein with the ligand, is likely to be conserved in many members of the TRIM family and may have evolved to facilitate recognition of repetitive epitope patterns associated with infection.


Effects of Inulin Propionate Ester Incorporated into Palatable Food Products on Appetite and Resting Energy Expenditure: A Randomised Crossover Study.
Supplementation with inulin-propionate ester (IPE), which delivers propionate to the colon, suppresses ad libitum energy intake and stimulates the release of satiety hormones acutely in humans, and prevents weight gain. In order to determine whether IPE remains effective when incorporated into food products (FP), IPE needs to be added to a widely accepted food system. A bread roll and fruit smoothie were produced. Twenty-one healthy overweight and obese humans participated. Participants attended an acclimatisation visit and a control visit where they consumed un-supplemented food products (FP). Participants then consumed supplemented-FP, containing 10 g/d inulin or IPE for six days followed by a post-supplementation visit in a randomised crossover design. On study visits, supplemented-FP were consumed for the seventh time and ad libitum energy intake was assessed 420 min later. Blood samples were collected to assess hormones and metabolites. Resting energy expenditure (REE) was measured using indirect calorimetry. Taste and appearance ratings were similar between FP. Ad libitum energy intake was significantly different between treatments, due to a decreased intake following IPE-FP. These observations were not related to changes in blood hormones and metabolites. There was an increase in REE following IPE-FP. However, this effect was lost after correcting for changes in fat free mass. Our results suggest that IPE suppresses appetite and may alter REE following its incorporation into palatable food products.


Lodestone Compass: Chinese or Olmec Primacy?: Multidisciplinary analysis of an Olmec hematite artifact from San Lorenzo, Veracruz, Mexico.
Considering the unique morphology (purposefully shaped polished bar with a groove) and composition (magnetic mineral with magnetic moment vector in the floating plane) of M-160, and acknowledging that the Olmec were a sophisticated people who possessed advanced knowledge and skill in working iron ore minerals, I would suggest for consideration that the Early Formative artifact M-160 was probably manufactured and used as what I have called a zeroth-order compass, if not a first-order compass. The data I have presented in this article support this hypothesis, although they are not sufficient to prove it. That M-160 could be used today as a geomagnetically directed pointer is undeniable. The original whole bar may indeed have pointed close to magnetic north-south. The groove functions well as a sighting mark, and the slight angle it makes with the axis of the bar appears to be the result of calibration rather than accident. A negative supporting argument is that M-160 looks utilitarian rather than decorative, and no function for the object other than that of a compass pointer has been suggested by anyone who has examined it critically. Whether such a pointer would have been used to point to something astronomical (zeroth-order compass) or to geomagnetic north-south (first-order compass) is entirely open to speculation. The observation of the family of Olmec site alignments 8 degrees west of north is a curiosity in its own right, and the possibility that these alignments have an astronomical or geomagnetic origin should be explored. I also believe that it is constructive to compare the first millennium Chinese, who used the lodestone compass for geomancy, with the Gulf Coast Olmec since both were agrarian-terrestrial societies. The Olmec's apparent concern with orientation and skillful use of magnetic minerals also stimulates one to draw cross-cultural parallels. The evidence and analysis offered in this article provide a basis for hypotheses of parallel cultural developments in China and the Olmec New World. If the Olmec did discover the geomagnetic orienting properties of lodestone, as did the Han Chinese, it is most reasonable to speculate that they would have used their compass for comparable geomantic purposes. It should, however, be recognized that the Olmec claim, if documented, predates the Chinese discovery of the geomagnetic lodestone compass by more than a millennium. At present, M-160 is a unique artifact and San Lorenzo a unique site: "The first civilized center of Mesoamerica and probably of the New World" (19, p. 89). Further documentation of the Olmec claim must await the discovery of similar artifacts in museums, private collections, or as yet undiscovered Olmec sites. I would welcome communications from anyone possessing information relating to such artifacts. Regardless of shape, purposefully grooved and highly polished specimens of magnetic minerals are of particular interest. It would also be useful for the archeologist excavating Olmec burials and offerings to carefully note their alignments and consider them in a geomantic context. In addition to the discovery of supporting artifacts, establishment of Olmec primacy of the lodestone compass depends on the acquisition of the archeomagnetic data for the Early Formative period. I appeal to archeologists who find good archeomagnetic samples (burned hearths and post-holes) from the Formative periods to convey this information to R. DuBois of the University of Oklahoma. In a few years, the archeomagnetic data should be available for the last three millennia and the possibilities are very exciting.


Blind testing of routine, fully automated determination of protein structures from NMR data.
The protocols currently used for protein structure determination by nuclear magnetic resonance (NMR) depend on the determination of a large number of upper distance limits for proton-proton pairs. Typically, this task is performed manually by an experienced researcher rather than automatically by using a specific computer program. To assess whether it is indeed possible to generate in a fully automated manner NMR structures adequate for deposition in the Protein Data Bank, we gathered 10 experimental data sets with unassigned nuclear Overhauser effect spectroscopy (NOESY) peak lists for various proteins of unknown structure, computed structures for each of them using different, fully automatic programs, and compared the results to each other and to the manually solved reference structures that were not available at the time the data were provided. This constitutes a stringent "blind" assessment similar to the CASP and CAPRI initiatives. This study demonstrates the feasibility of routine, fully automated protein structure determination by NMR.


Agonist binding to multiple muscarinic receptors.
The binding of agonists to muscarinic cholinergic receptors is well described by a binding model of multiple affinity states (superhigh, high, and low) in most central and peripheral tissues. Although previous studies of the influences by divalent cations, guanine nucleotides, and sulfhydryl reagents support the concept that these regulators act through closely related sites to alter the relative proportions of muscarinic agonist affinity states, it has become apparent that muscarinic receptor subtypes (as defined with the nonclassical antagonist pirenzepine) are differentially affected by the regulators. For example, in tissues that have few high-affinity [3H]pirenzepine-binding sites (heart, ileum, cerebellum), magnesium ions promote the formation of a high agonist affinity state, whereas exposure of these tissues to the sulfhydryl reagent N-ethylmaleimide (NEM) or guanine nucleotides promotes the formation of a low agonist affinity state. Conversely, tissues rich in high-affinity [3H]pirenzepine-binding sites (cerebral cortex, corpus striatum, hippocampus) show little, if any, change in agonist binding site affinity when magnesium ions or guanine nucleotides are present. Furthermore, NEM enhances the muscarinic binding site affinity for agonists in these tissues. Taken together, these results support the concept of muscarinic receptor heterogeneity, as proposed from previous physiological studies, and indicate that the aforementioned regulators (guanine nucleotides, magnesium ions, NEM) differentially alter the agonist-binding properties of these muscarinic receptor subtypes.


Treatment of hypertension with clonidine.
Clonidine, in a daily dosage of 0.15-4.8 mg., effectively lowered systolic and diastolic pressures in 26 out of 28 impatients with moderate to severe hypertension, including five with primary renal disease. The action of the drug did not depend on posture and was not associated with reduction in renal function. Side-effects were not severe, but mental changes occurred in four patients.Clonidine is a useful alternative to currently available antihypertensive drugs, but further evaluation of its longterm efficacy is required.


Alterations in piglet small intestinal structure at weaning.
The small intestinal mucosa of 112 piglets aged between 21 and 32 days was examined to identify the effects on its structure of weaning and of consumption of creep food. In unweaned control pigs a gradual increase in crypt depth occurred with age, while villus height altered little. In contrast, weaned pigs showed a highly significant increase in crypt depth and an increase in the complexity of villus morphology with a dramatic reduction in villus height. Crypts elongated most in the distal small intestine, while the loss of villus height was greatest proximally. No inflammatory reaction was seen and the alterations were not influenced by consumption of creep food before weaning. This failed to support the hypothesis that hypersensitivity to dietary antigens is involved in the aetiology of such intestinal changes. The reduction in small intestinal absorptive area and the appearance of a less mature enterocyte population help to explain the increased susceptibility of the pig to diarrhoea and growth checks in the post-weaning period.


Telomeres, tethers and trypanosomes.
Temporal and spatial organization of the nucleus is critical for the control of transcription, mRNA processing and the assembly of ribosomes. This includes the occupancy of specific territories by mammalian chromosomes, the presence of subnuclear compartments such as the nucleolus and Cajal bodies and the division of chromatin between active and inactive states. These latter are commonly associated with the location of DNA within euchromatin and heterochromatin respectively; critically these distinctions arise through modifications to chromatin-associated proteins, including histones, as well as the preferential localization of heterochromatin at the nuclear periphery. Most research on nuclear organization has focused on metazoa and fungi; however, recent technical advances have made more divergent eukaryotes accessible to study, with some surprising results. For example, the organization of heterochromatin is mediated in metazoan nuclei in large part by lamins, the prototypical intermediate filament proteins. Despite the presence of heterochromatin, detected both biochemically and by EM in most eukaryotic organisms, until this year lamins were thought to be restricted to metazoan taxa, and the proteins comprising the lamina in other lineages were unknown. Recent work indicates the presence of lamin orthologs in amoeba, while trypanosomatids possess a large coiled-coil protein, NUP-1, that performs functions analogous to lamins. These data indicate that the presence of a nuclear lamina is substantially more widespread than previously thought, with major implications for the evolution of eukaryotic gene expression mechanisms. We discuss these and other recent findings on the organization of nuclei in diverse organisms, and the implications of these findings for the evolutionary origin of eukaryotes.


Bone morphogenetic protein (BMP) ligands and receptors in bovine ovarian follicle cells: actions of BMP-4, -6 and -7 on granulosa cells and differential modulation of Smad-1 phosphorylation by follistatin.
Given the paucity of information on the potential roles of bone morphogenetic proteins (BMPs) in the ruminant ovary we conducted immunolocalization and functional studies on cells isolated from bovine antral follicles. Immunocytochemistry revealed expression of BMP-4 and -7 in isolated theca cells whereas granulosa cells and oocytes selectively expressed BMP-6. All three cell types expressed a range of BMP-responsive type-I (BMPRIB, ActRI) and type-II (BMPRII, ActRII, ActRIIB) receptors supporting autocrine/paracrine roles within the follicle. This was reinforced by functional experiments on granulosa cells which showed that BMP-4, -6 and -7 promoted cellular accumulation of phosphorylated Smad-1 but not Smad-2 and enhanced 'basal' and IGF-stimulated secretion of oestradiol (E2), inhibin-A, activin-A and follistatin (FS). Concomitantly, each BMP suppressed 'basal' and IGF-stimulated progesterone secretion, consistent with an action to prevent or delay atresia and/or luteinization. BMPs also increased viable cell number under 'basal' (BMP-4 and -7) and IGF-stimulated (BMP-4, -6 and -7) conditions. Since FS, a product of bovine granulosa cells, has been shown to bind several BMPs, we used the Biacore technique to compare its binding affinities for activin-A (prototype FS ligand) and BMP-4, -6 and -7. Compared with activin-A (K(d) 0.28 +/- 0.02 nM; 100%), the relative affinities of FS for BMP-4, -6 and -7 were 10, 5 and 1% respectively. Moreover, studies on granulosa cells showed that preincubation of ligand with excess FS abolished activin-A-induced phosphorylation of Smad-2 and BMP-4-induced phosphorylation of Smad-1. However, FS only partially reversed BMP-6-induced Smad-1 phosphorylation and had no inhibitory effect on BMP-7-induced Smad-1 phosphorylation. These findings support functional roles for BMP-4, -6 and -7 as paracrine/autocrine modulators of granulosa cell steroidogenesis, peptide secretion and proliferation in bovine antral follicles. The finding that FS can differentially modulate BMP-induced receptor activation and that this correlates with the relative binding affinity of FS for each BMP type implicates FS as a potential modulator of BMP action in the ovary.