Subpopulations of liver coated vesicles resolved by preparative agarose gel electrophoresis.

Rat liver clathrin coated vesicles (CVs) were separated into several distinct subpopulations using non-sieving concentrations of agarose, which allowed the separation of species differing primarily in surface charge. Using preparative agarose electrophoresis (Kedersha, N. L., and L. H. Rome, 1986, Anal. Biochem., in press), the CVs were recovered and analyzed for differences in morphology, coat protein composition, and stripped vesicle protein composition. Coat proteins from different populations appeared identical on SDS PAGE, and triskelions stripped from the different populations showed the same mobility on the agarose gel, suggesting that the mobility differences observed in intact CVs were due to differences in the surface charge of underlying vesicles rather than to variations in their clathrin coats. Several non-coat polypeptides appeared to segregate exclusively with different populations as resolved by two-dimensional electrophoresis. Stripped CVs also exhibited considerable heterogeneity when analyzed by Western blotting: the fast-migrating population was enriched in the mannose 6-phosphate receptor, secretory acetylcholine esterase, and an Mr 195,000 glycoprotein. The slow-migrating population of CVs was enriched in the asialoglycoprotein receptor, and it appeared to contain all detectable concanavalin A-binding polypeptides as well as the bulk of detectable WGA-binding proteins. When CVs were prepared from 125I-asialoorosomucoid-perfused rat liver, ligand was found in the slow-migrating CVs, suggesting that these were endocytic in origin. Morphological differences were also observed: the fast-migrating population was enriched in smaller CVs, whereas the slow-migrating population exhibited an enrichment in larger CVs. As liver consists largely of hepatocytes, these subpopulations appear to originate from the same cell type and probably represent CVs of different intracellular origin and destination

Exploring the effects of demonstration and enactment in facilitating recall of instructions in working memory

Across the lifespan the ability to follow instructions is essential for the successful completion of a multitude of daily activities. This ability will often rely on the storage and processing of information in working memory, and previous research in this domain has found that self-enactment at encoding or observing other-enactment at encoding (demonstration) improves performance at recall. However, no working memory research has directly compared these manipulations. Experiment 1 explored the effects of both self-enactment and demonstration on young adults’ (N=48) recall of action-object instruction sequences (e.g. ‘spin the circle, tap the square’). Both manipulations improved recall, with demonstration providing relatively larger boosts to performance across conditions. More detailed analyses suggested that this improvement was driven by improving the representations of actions, rather than objects, in these action-object sequences. Experiment 2 (N=24) explored this further, removing the objects from the physical environment and comparing partial demonstration (i.e. action-only or object-only) with no or full demonstration. The results showed that partial demonstration only benefitted features that were demonstrated, while full demonstration improved memory for actions, objects and their pairings. Overall these experiments indicate how self-enactment, and particularly demonstration, can benefit verbal recall of instruction sequences through the engagement of visuo-motor processes that provide additional forms of coding to support working memory performance

The validity and reliability of observational assessment tools available to measure fundamental movement skills in school-age children: A systematic review

Background Fundamental Movement Skills (FMS) play a critical role in ontogenesis. Many children have insufficient FMS, highlighting the need for universal screening in schools. There are many observational FMS assessment tools, but their psychometric properties are not readily accessible. A systematic review was therefore undertaken to compile evidence of the validity and reliability of observational FMS assessments, to evaluate their suitability for screening. Methods A pre-search of ‘fundamental movement skills’ OR ‘fundamental motor skills’ in seven online databases (PubMed, Ovid MEDLINE, Ovid Embase, EBSCO CINAHL, EBSCO SPORTDiscus, Ovid PsycINFO and Web of Science) identified 24 assessment tools for school-aged children that: (i) assess FMS; (ii) measure actual motor competence and (iii) evaluate performance on a standard battery of tasks. Studies were subsequently identified that: (a) used these tools; (b) quantified validity or reliability and (c) sampled school-aged children. Study quality was assessed using COnsensus-based Standards for the selection of health Measurement INstruments (COSMIN) checklists. Results Ninety studies were included following the screening of 1863 articles. Twenty-one assessment tools had limited or no evidence to support their psychometric properties. The Test of Gross Motor Development (TGMD, n = 34) and the Movement Assessment Battery for Children (MABC, n = 37) were the most researched tools. Studies consistently reported good evidence for validity, reliability for the TGMD, whilst only 64% of studies reported similarly promising results for the MABC. Twelve studies found good evidence for the reliability and validity of the Bruininks-Oseretsky Test of Motor Proficiency but poor study quality appeared to inflate results. Considering all assessment tools, those with promising psychometric properties often measured limited aspects of validity/reliability, and/or had limited feasibility for large scale deployment in a school-setting. Conclusion There is insufficient evidence to justify the use of any observational FMS assessment tools for universal screening in schools, in their current form

Coronary-artery bypass surgery in patients with ischemic cardiomyopathy

BACKGROUND The survival benefit of a strategy of coronary-artery bypass grafting (CABG) added to guideline-directed medical therapy, as compared with medical therapy alone, in patients with coronary artery disease, heart failure, and severe left ventricular systolic dysfunction remains unclear. METHODS From July 2002 to May 2007, a total of 1212 patients with an ejection fraction of 35% or less and coronary artery disease amenable to CABG were randomly assigned to undergo CABG plus medical therapy (CABG group, 610 patients) or medical therapy alone (medical-therapy group, 602 patients). The primary outcome was death from any cause. Major secondary outcomes included death from cardiovascular causes and death from any cause or hospitalization for cardiovascular causes. The median duration of follow-up, including the current extended-follow-up study, was 9.8 years. RESULTS A primary outcome event occurred in 359 patients (58.9%) in the CABG group and in 398 patients (66.1%) in the medical-therapy group (hazard ratio with CABG vs. medical therapy, 0.84; 95% confidence interval [CI], 0.73 to 0.97; P=0.02 by log-rank test). A total of 247 patients (40.5%) in the CABG group and 297 patients (49.3%) in the medical-therapy group died from cardiovascular causes (hazard ratio, 0.79; 95% CI, 0.66 to 0.93; P=0.006 by log-rank test). Death from any cause or hospitalization for cardiovascular causes occurred in 467 patients (76.6%) in the CABG group and in 524 patients (87.0%) in the medical-therapy group (hazard ratio, 0.72; 95% CI, 0.64 to 0.82; P<0.001 by log-rank test). CONCLUSIONS In a cohort of patients with ischemic cardiomyopathy, the rates of death from any cause, death from cardiovascular causes, and death from any cause or hospitalization for cardiovascular causes were significantly lower over 10 years among patients who underwent CABG in addition to receiving medical therapy than among those who received medical therapy alone. (Funded by the National Institutes of Health; STICH [and STICHES] ClinicalTrials.gov number, NCT00023595.

Prevalence of contraindications to mefloquine use among USA military personnel deployed to Afghanistan

<p>Abstract</p> <p>Background</p> <p>Mefloquine has historically been considered safe and well-tolerated for long-term malaria chemoprophylaxis, but its prescribing requires careful attention to rule out contraindications to its use, including a history of certain psychiatric and neurological disorders. The prevalence of these disorders has not been defined in cohorts of U.S. military personnel deployed to areas where long-term malaria chemoprophylaxis is indicated.</p> <p>Methods</p> <p>Military medical surveillance and pharmacosurveillance databases were utilized to identify contraindications to mefloquine use among a cohort of 11,725 active duty U.S. military personnel recently deployed to Afghanistan.</p> <p>Results</p> <p>A total of 9.6% of the cohort had evidence of a contraindication. Females were more than twice as likely as males to have a contraindication (OR = 2.48, P < 0.001).</p> <p>Conclusion</p> <p>These findings underscore the importance of proper systematic screening prior to prescribing and dispensing mefloquine, and the need to provide alternatives to mefloquine suitable for long-term administration among deployed U.S. military personnel.</p

Survey of patient satisfaction with the Breastfeeding Education and Support Services of The Royal Women's Hospital, Melbourne

<p>Abstract</p> <p>Background</p> <p>The Breastfeeding Education and Support Services (BESS) is a unit of The Royal Women's Hospital in Melbourne, Australia, staffed by International Board Certified Lactation Consultants (IBCLCs), providing day/short-stay and an outpatient clinic for mothers and infants with breastfeeding problems. It is important to measure women's experience of visiting the service as part of quality assurance. The aim of this project was to conduct an anonymous postal survey of clients' satisfaction with BESS.</p> <p>Methods</p> <p>An anonymous survey was posted on 16 November 2005 and again on 31 January 2006, to all women who had attended BESS in September 2005.</p> <p>Results</p> <p>The response rate was 60.5% (78/129). Eighty percent (62/78) of respondents attended day-stay, 33% (26/78) attended short-stay and 15% (12/78) attended the outpatient clinic. The percentage of women who responded "strongly agree" to the statement "Overall, I am satisfied with the services" was 49% (35/72) and 50% (6/12) for those who went to day/short-stay and the outpatient clinic respectively. Overall, 56% of all respondents responded that the quality of BESS was "better than expected". The most common breastfeeding problem reported was difficulty attaching the baby to the breast, followed by nipple damage, low milk supply and painful feeding.</p> <p>Conclusion</p> <p>BESS seems to have provided a satisfactory service to most clients. Most respondents were clearly satisfied with the support given by the IBCLCs and have also responded that the staff were professional and knowledgeable in their field of work.</p

Developing and validating a school-based screening tool of Fundamental Movement Skills (FUNMOVES) using Rasch analysis

Background A large proportion of children are not able to perform age-appropriate fundamental movement skills (FMS). Thus, it is important to assess FMS so that children needing additional support can be identified in a timely fashion. There is great potential for universal screening of FMS in schools, but research has established that current assessment tools are not fit for purpose. Objective To develop and validate the psychometric properties of a FMS assessment tool designed specifically to meet the demands of universal screening in schools. Methods A working group consisting of academics from developmental psychology, public health and behavioural epidemiology developed an assessment tool (FUNMOVES) based on theory and prior evidence. Over three studies, 814 children aged 4 to 11 years were assessed in school using FUNMOVES. Rasch analysis was used to evaluate structural validity and modifications were then made to FUNMOVES activities after each study based on Rasch results and implementation fidelity. Results The initial Rasch analysis found numerous psychometric problems including multidimensionality, disordered thresholds, local dependency, and misfitting items. Study 2 showed a unidimensional measure, with acceptable internal consistency and no local dependency, but that did not fit the Rasch model. Performance on a jumping task was misfitting, and there were issues with disordered thresholds (for jumping, hopping and balance tasks). Study 3 revealed a unidimensional assessment tool with good fit to the Rasch model, and no further issues, once jumping and hopping scoring were modified. Implications The finalised version of FUNMOVES (after three iterations) meets standards for accurate measurement, is free and able to assess a whole class in under an hour using resources available in schools. Thus FUNMOVES has the potential to allow schools to efficiently screen FMS to ensure that targeted support can be provided and disability barriers removed

Canine respiratory coronavirus employs caveolin-1-mediated pathway for internalization to HRT-18G cells

Canine respiratory coronavirus (CRCoV), identified in 2003, is a member of the Coronaviridae family. The virus is a betacoronavirus and a close relative of human coronavirus OC43 and bovine coronavirus. Here, we examined entry of CRCoV into human rectal tumor cells (HRT-18G cell line) by analyzing co-localization of single virus particles with cellular markers in the presence or absence of chemical inhibitors of pathways potentially involved in virus entry. We also targeted these pathways using siRNA. The results show that the virus hijacks caveolin-dependent endocytosis to enter cells via endocytic internalization

Towards an In Vitro Model of Plasmodium Hypnozoites Suitable for Drug Discovery

Contains fulltext : 96475.pdf (publisher's version ) (Open Access)BACKGROUND: Amongst the Plasmodium species in humans, only P. vivax and P. ovale produce latent hepatic stages called hypnozoites, which are responsible for malaria episodes long after a mosquito bite. Relapses contribute to increased morbidity, and complicate malaria elimination programs. A single drug effective against hypnozoites, primaquine, is available, but its deployment is curtailed by its haemolytic potential in glucose-6-phosphate dehydrogenase deficient persons. Novel compounds are thus urgently needed to replace primaquine. Discovery of compounds active against hypnozoites is restricted to the in vivo P. cynomolgi-rhesus monkey model. Slow growing hepatic parasites reminiscent of hypnozoites had been noted in cultured P. vivax-infected hepatoma cells, but similar forms are also observed in vitro by other species including P. falciparum that do not produce hypnozoites. METHODOLOGY: P. falciparum or P. cynomolgi sporozoites were used to infect human or Macaca fascicularis primary hepatocytes, respectively. The susceptibility of the slow and normally growing hepatic forms obtained in vitro to three antimalarial drugs, one active against hepatic forms including hypnozoites and two only against the growing forms, was measured. RESULTS: The non-dividing slow growing P. cynomolgi hepatic forms, observed in vitro in primary hepatocytes from the natural host Macaca fascicularis, can be distinguished from similar forms seen in P. falciparum-infected human primary hepatocytes by the differential action of selected anti-malarial drugs. Whereas atovaquone and pyrimethamine are active on all the dividing hepatic forms observed, the P. cynomolgi slow growing forms are highly resistant to treatment by these drugs, but remain susceptible to primaquine. CONCLUSION: Resistance of the non-dividing P. cynomolgi forms to atovaquone and pyrimethamine, which do not prevent relapses, strongly suggests that these slow growing forms are hypnozoites. This represents a first step towards the development of a practical medium-throughput in vitro screening assay for novel hypnozoiticidal drugs

A novel malaria vaccine candidate antigen expressed in Tetrahymena thermophila

Development of effective malaria vaccines is hampered by the problem of producing correctly folded Plasmodium proteins for use as vaccine components. We have investigated the use of a novel ciliate expression system, Tetrahymena thermophila, as a P. falciparum vaccine antigen platform. A synthetic vaccine antigen composed of N-terminal and C-terminal regions of merozoite surface protein-1 (MSP-1) was expressed in Tetrahymena thermophila. The recombinant antigen was secreted into the culture medium and purified by monoclonal antibody (mAb) affinity chromatography. The vaccine was immunogenic in MF1 mice, eliciting high antibody titers against both N- and C-terminal components. Sera from immunized animals reacted strongly with P. falciparum parasites from three antigenically different strains by immunofluorescence assays, confirming that the antibodies produced are able to recognize parasite antigens in their native form. Epitope mapping of serum reactivity with a peptide library derived from all three MSP-1 Block 2 serotypes confirmed that the MSP-1 Block 2 hybrid component of the vaccine had effectively targeted all three serotypes of this polymorphic region of MSP-1. This study has successfully demonstrated the use of Tetrahymena thermophila as a recombinant protein expression platform for the production of malaria vaccine antigens