Inertial-Based Human Motion Capture: A Technical Summary of Current Processing Methodologies for Spatiotemporal and Kinematic Measures.
Inertial-based motion capture (IMC) has been suggested to overcome many of the limitations of traditional motion capture systems. The validity of IMC is, however, suggested to be dependent on the methodologies used to process the raw data collected by the inertial device. The aim of this technical summary is to provide researchers and developers with a starting point from which to further develop the current IMC data processing methodologies used to estimate human spatiotemporal and kinematic measures. The main workflow pertaining to the estimation of spatiotemporal and kinematic measures was presented, and a general overview of previous methodologies used for each stage of data processing was provided. For the estimation of spatiotemporal measures, which includes stride length, stride rate, and stance/swing duration, measurement thresholding and zero-velocity update approaches were discussed as the most common methodologies used to estimate such measures. The methodologies used for the estimation of joint kinematics were found to be broad, with the combination of Kalman filtering or complimentary filtering and various sensor to segment alignment techniques including anatomical alignment, static calibration, and functional calibration methods identified as being most common. The effect of soft tissue artefacts, device placement, biomechanical modelling methods, and ferromagnetic interference within the environment, on the accuracy and validity of IMC, was also discussed. Where a range of methods have previously been used to estimate human spatiotemporal and kinematic measures, further development is required to reduce estimation errors, improve the validity of spatiotemporal and kinematic estimations, and standardize data processing practices. It is anticipated that this technical summary will reduce the time researchers and developers require to establish the fundamental methodological components of IMC prior to commencing further development of IMC methodologies, thus increasing the rate of development and utilisation of IMC.


Identifying research priorities in eating disorders: A Delphi study building consensus across clinicians, researchers, consumers, and carers in Australia.
Eating disorders are underrepresented among successful applications for medical research funding. Developing agreement on the top research priorities may assist in fostering collaborations, innovation, and meaningful consumer and carer involvement. This study aimed to develop consensus among Australian clinicians, researchers, consumers, carers, and interested members of affiliated industries, on the priorities for eating disorders research. The Delphi expert consensus method was used. Participants were 291 members of the Australia New Zealand Academy for Eating Disorders (ANZAED) or the National Eating Disorders Collaboration (NEDC). Three panels were formed based on participant expertise: "ED Specialists," Consumers/Carers, and Affiliates. A total of 32 potential research areas (29 in Survey 1, 3 in Survey 2) were rated on a 5-point scale of importance, with 80% agreement across all three panels required for endorsement as a priority. Participants also ranked 7 broad research domains in order of priority. Research areas describing early intervention and evidence-based treatments were the most highly rated, with more than 90% endorsement from each of the three panels. The research domains of accessible evidence-based treatments, early intervention and detection, and origins of eating disorders, were the most highly ranked. The ED specialists and Consumers/Carers panels had very similar patterns of responding, while the Affiliates panel showed small differences. Using the Delphi expert consensus method resulted in a collaborative consensus driven eating disorders research agenda for the Australian context and forms a model upon which other countries may also develop their funding priorities.


Evaluation of the repeatability of rhinomanometry and its use in assessing transnasal resistance and pressure in dogs.
To evaluate a modified posterior rhinomanometric method for clinical application in dogs. 15 healthy Beagles and 8 Bulldogs (4 healthy and 4 with respiratory problems). Rhinomanometry was performed 3 times within a 15-minute period in anesthetized dogs. Transnasal pressure (P(NA)) and nasal resistance (R(NA)) were determined by use of artificial airflow (adjusted for body weight) for inspiration (P(NAin) and R(NAin), respectively) and expiration (P(NAout) and R(NAout)). Procedures were repeated for the Beagles 7 days later. For the Beagles, mean +/- SD of P(NAin) for both days (0.162 +/- 0.042 kPa) was significantly lower than P(NAout) (0.183 +/- 0.053 kPa). Similarly, R(NAin) (1.47 +/- 0.41 kPa/[L/s]) was significantly lower than R(NAout) (1.64 +/- 0.46 kPa/[L/s]). Pairwise comparison of values for P(NA) and R(NA) for the 2 days revealed no significant difference. Repeatability of the method (estimated as within-day variation) for R(NA) was +/- 0.19 kPa/(L/s), whereas variation between the days was +/- 0.36 kPa/(L/s) for R(NAin) and +/- 0.44 kPa/(L/s) for R(NAout). The 4 clinically normal Bulldogs had R(NA) values ranging from 1.69 to 3.48 kPa/(L/s), whereas in the 4 Bulldogs with respiratory problems, R(NA) ranged from 9.83 to 20.27 kPa/(L/s). R(NA) is inversely dependent on body size and nonlinearly associated with airflow. We propose that R(NA) in dogs should be determined for airflows standardized on the basis of body size. The P(NA) and R(NA) in Beagles can be measured with sufficient repeatability for clinical use and nasal obstructions are detectable.


Molecular characterization of Coriolus versicolor PSP-induced apoptosis in human promyelotic leukemic HL-60 cells using cDNA microarray.
Proteins and peptide bound polysaccharides (PSP) extracted from Basidiomycetous fungi are widely used in cancer immunotherapy and recently demonstrated to induce apoptosis in cancer cells in vitro. In order to provide the molecular pharmacological mechanisms of PSP on human cancer cells, we investigated the gene expression profiles of PSP-treated apoptotic human promyelotic leukemic HL-60 cells using ResGen 40k IMAGE printed cDNA microarray. In total 378 and 111 transcripts were identified as differentially expressed in the apoptotic cells by at least a factor of 2 or 3, respectively. Our data show that PSP-induced apoptosis in HL-60 cells might be mediated by up-regulation of early transcription factors such as AP-1, EGR1, IER2 and IER5, and down-regulation of NF-kappaB transcription pathways. Other gene expression changes, including the increase of several apoptotic or anti-proliferation genes, such as GADD45A/B and TUSC2, and the decrease of a batch of phosphatase and kinase genes, may also provide further evidences in supporting the process of PSP induced apoptosis in cancer cells. Some of the well-characterized carcinogenesis-related gene transcripts such as SAT, DCT, Melan-A, uPA and cyclin E1 were also alternated by PSP in the HL-60 cells. These transcripts can be employed as markers for quality control of PSP products on functional levels. The present study provides new insight into the molecular mechanisms involved in PSP-induced apoptosis in leukemic HL-60 cells analyzed by cDNA microarray.


Laboratory evaluation of a flow cytometric BCR-ABL immunobead assay.
A new flow cytometric (FC) BCR-ABL immunobead assay has been developed recently. Here we present the laboratory evaluation of the commercially available kit. Mononuclear cells were isolated, lysed and processed according to the instructions of the manufacturer. Anti-BCR antibodies adsorbed to capture beads bind the BCR-ABL fusion proteins of the lysed cells, a phycoerythrin (PE)-conjugated anti-ABL antibody is the detector reagent and mean fluorescence intensity (MFI) signals were recorded by flow cytometry. Detection of t(9;22)(q34;q11) translocation was carried out with a quantitative PCR assay. MFI results of 20 normal peripheral blood samples were 88±8 (mean±SD), CV 9%. K562 cells were used as positive control. Within-batch imprecision was excellent (3.7% in the normal and 10% in the pathological range). Cut-off was chosen at MFI 112, where both sensitivity and specificity were 100%. Altogether 17 chronic myeloid leukemia (CML) and 16 acute leukemia samples were analyzed. All PCR positive samples (n=14) were positive with the FC method and negative results were also concordant (n=15). Frozen cell lysates can be stored up to 4 weeks without significant decrease of MFI signal. The FC BCR-ABL assay is a fast, reproducible and reliable method that may be incorporated into standard flow cytometric protocols to help clinical decision-making.


Fc-disabled anti-mouse CD40L antibodies retain efficacy in promoting transplantation tolerance.
CD40L antibodies have proven to be powerful immunosuppressive agents in nonhuman primates but unfortunately perturb blood coagulation. Neither the therapeutic nor the prothrombotic mechanism of anti-CD40L is defined sufficiently to determine whether these effects can be uncoupled. Recent evidence suggests that the Fc region of anti-CD40L antibodies interacting with Fc receptors plays an important role in stabilizing platelet aggregates. An Fc-disabled, aglycosylated anti-CD40L heavy chain variant was therefore created to determine whether it might still be useful in promoting transplantation tolerance. In a number of mouse models an engineered aglycosyl anti-CD40L recapitulated the effects of the intact anti-CD40L antibody in tolerance protocols involving transplantation of allogeneic bone marrow and skin. In contrast, another anti-CD40L variant with a conventional rat gamma2b heavy chain was less effective in ensuring long-term skin graft survival, possibly associated with its faster clearance from the circulation. These results show that short pulses of anti-CD40L antibody therapy may still be useful in tolerance protocols even when the Fc region is disabled.


Overexpression of cytosolic glutamine synthetase. Relation to nitrogen, light, and photorespiration.
In plants, ammonium released during photorespiration exceeds primary nitrogen assimilation by as much as 10-fold. Analysis of photorespiratory mutants indicates that photorespiratory ammonium released in mitochondria is reassimilated in the chloroplast by a chloroplastic isoenzyme of glutamine synthetase (GS2), the predominant GS isoform in leaves of Solanaceous species including tobacco (Nicotiana tabacum). By contrast, cytosolic GS1 is expressed in the vasculature of several species including tobacco. Here, we report the effects on growth and photorespiration of overexpressing a cytosolic GS1 isoenzyme in leaf mesophyll cells of tobacco. The plants, which ectopically overexpress cytosolic GS1 in leaves, display a light-dependent improved growth phenotype under nitrogen-limiting and nitrogen-non-limiting conditions. Improved growth was evidenced by increases in fresh weight, dry weight, and leaf soluble protein. Because the improved growth phenotype was dependent on light, this suggested that the ectopic expression of cytosolic GS1 in leaves may act via photosynthetic/photorespiratory process. The ectopic overexpression of cytosolic GS1 in tobacco leaves resulted in a 6- to 7-fold decrease in levels of free ammonium in leaves. Thus, the overexpression of cytosolic GS1 in leaf mesophyll cells seems to provide an alternate route to chloroplastic GS2 for the assimilation of photorespiratory ammonium. The cytosolic GS1 transgenic plants also exhibit an increase in the CO(2) photorespiratory burst and an increase in levels of photorespiratory intermediates, suggesting changes in photorespiration. Because the GS1 transgenic plants have an unaltered CO(2) compensation point, this may reflect an accompanying increase in photosynthetic capacity. Together, these results provide new insights into the possible mechanisms responsible for the improved growth phenotype of cytosolic GS1 overexpressing plants. Our studies provide further support for the notion that the ectopic overexpression of genes for cytosolic GS1 can potentially be used to affect increases in nitrogen use efficiency in transgenic crop plants.


Prevalence and characteristics of meticillin-resistant Staphylococcus aureus in humans in contact with farm animals, in livestock, and in food of animal origin, Switzerland, 2009.
A total of 2,662 samples, collected from March to September 2009 in Switzerland, were tested for the presence of meticillin-resistant Staphylococcus aureus (MRSA). The collection comprised nasal swabs from 148 pig farmers, 133 veterinarians, 179 slaughterhouse employees, 800 pigs, 300 calves, 400 cattle, 100 pooled neck skin swabs from chicken carcasses, and 460 food samples of animal origin. Moreover, 142 S. aureus strains, isolated from bovine mastitis milk, were included in the study. Twenty samples (< 1%; four veterinarians, 10 pigs, three calves, one young bull, and two mastitis milk samples) tested positive for MRSA. Genotyping of the MRSA strains was performed by multilocus sequence typing, spa- and SCCmec-typing, and revealed ST398 (n=18), ST8 (n=1), ST 1 (n=1), spa types t011 (n=7), t034 (n=11), t064 (n=1), t127 (n=1), and SCCmec types IV (n=4) and V (n=16). The 20 MRSA strains were subjected to antibiotic susceptibility testing and pulsed-field gel electrophoresis using the restriction enzyme EagI. Supplementary PCR reactions were performed to investigate the presence of Panton-Valentine leukocidin and staphylococcal enterotoxins A to D.


Sexual abuse history and pelvic floor disorders in women.
Sexual abuse rates in the general female population range between 15% and 25%, and sexual abuse is known to have a long-term impact on a woman's health. The aim of this study was to report the prevalence of sexual abuse history in women presenting to clinicians for pelvic floor disorders (PFD) and to determine whether a history of sexual abuse is associated with a specific type of PFD. We conducted a retrospective chart review of new urogynecology patients seen at the University of New Mexico Hospital. All women underwent a standardized history and physical examination and completed symptom severity and quality-of-life measures. Univariate and multivariable analyses were conducted to determine which PFDs were associated with a history of sexual abuse among women with and without a history of sexual abuse. A total of 1899 new urogynecology patients with complete information were identified from January 2007 and October 2011; 1260 (66%) were asked about a history of sexual abuse. The prevalence of sexual abuse was 213/1260 (17%). In the multivariable analysis, only chronic pelvic pain remained significantly associated with a history of sexual abuse. A history of sexual abuse is common among women with PFDs, and these women were more likely to have chronic pelvic pain.


Comparison of chemical treatments to kill Salmonella on alfalfa seeds destined for sprout production.
Outbreaks of salmonellosis in the US, Canada and Finland linked to alfalfa sprouts have been attributed to Salmonella stanley in 1995 and Salmonella newport in 1996. This study was undertaken to compare the efficacy of chemical treatments in killing a mixture of five Salmonella serovars inoculated onto alfalfa seeds. Solutions containing calcium hypochlorite or sodium hypochlorite at concentrations of 1800 and 2000 micrograms/ml active (available) chlorine respectively, as well as 6% hydrogen peroxide or 80% ethanol were effective in reducing Salmonella populations by more than 1000 fold. However, viable Salmonella cells were detected in seeds treated for 10 min in these solutions. The inaccessibility of Salmonella cells in crevices and between the cotyledon and testa of seeds to lethal concentrations of these chemicals are thought to be the reason for the lack of effectiveness.


Crosstalk between mesenchymal stem cells and endothelial cells leads to downregulation of cytokine-induced leukocyte recruitment.
Mesenchymal stem cells (MSC) have immunomodulatory properties, but their effects on endothelial cells (EC) and recruitment of leukocytes are unknown. We cocultured human bone marrow-derived MSC with EC and found that MSC could downregulate adhesion of flowing neutrophils or lymphocytes and their subsequent transendothelial migration. This applied for EC treated with tumor necrosis factor-α (TNF), interleukin-1β (IL-1), or TNF and interferon-γ combined. Supernatant from cocultures also inhibited endothelial responses. This supernatant had much higher levels of IL-6 than supernatant from cultures of the individual cells, which also lacked inhibitory functions. Addition of neutralizing antibody against IL-6 removed the bioactivity of the supernatant and also the immunomodulatory effects of coculture. Studies using siRNA showed that IL-6 came mainly from the MSC in coculture, and reduction in production in MSC alone was sufficient to impair the protective effects of coculture. Interestingly, siRNA knockdown of IL-6-receptor expression in MSC as well as EC inhibited anti-inflammatory effects. This was explained when we detected soluble IL-6R receptor in supernatants and showed that receptor removal reduced the potency of supernatant. Neutralization of transforming growth factor-β indicated that activation of this factor in coculture contributed to IL-6 production. Thus, crosstalk between MSC and EC caused upregulation of production of IL-6 by MSC which in turn downregulated the response of EC to inflammatory cytokines, an effect potentiated by MSC release of soluble IL-6R. These studies establish a novel mechanism by which MSC might have protective effects against inflammatory pathology and cardiovascular disease.


Design, analysis, and interpretation of method-comparison studies.
Clinicians often need to know whether a new method of measurement is equivalent to an established one already in clinical use. This article reviews the methodology of a method-comparison study to assist the clinician with the conduct and evaluation of such studies. Temperature data from 1 subject are used to illustrate the procedures. Although one would not make decisions on the basis of the findings from 1 subject, the large number of paired measurements in the data set permits its use for illustrative purposes. Currently available software eliminates the need for tedious statistical computation but does not reduce the burden of understanding the concepts underlying a method-comparison study and accurate interpretation of the findings.


Regulation of microphthalmia-associated transcription factor MITF protein levels by association with the ubiquitin-conjugating enzyme hUBC9.
The basic helix-loop-helix/leucine zipper (bHLH/ZIP) microphthalmia-associated transcription factor (MITF) regulates transcription of genes encoding enzymes essential for melanin biosynthesis in melanocytes and retinal pigmented epithelial cells. To determine how MITF activity is regulated, we used the yeast two-hybrid system to identify proteins expressed by human melanoma cells that interact with MITF. The majority of clones that showed positive interaction with a 158-amino-acid region of MITF containing the bHLH/ZIP domain (aa 168-325) encoded the ubiquitin conjugating enzyme hUBC9. The association of MITF with hUBC9 was further confirmed by an in vitro GST pull-down assay. Although hUBC9 is known to interact preferentially with SENTRIN/SUMO1, in vitro transcription/translation analysis demonstrated greater association of MITF with ubiquitin than with SENTRIN. Importantly, cotransfection of MITF and hUBC9 expression vectors resulted in MITF protein degradation. MITF protein was stabilized by the proteasome inhibitor MG132, indicating the role of the ubiquitin-proteasome system in MITF degradation. Serine 73, which is located in a region rich in proline, glutamic acid, serine, and threonine (PEST), regulates MITF protein stability, since a serine to alanine mutation prevented hUBC9-mediated MITF (S73A) degradation. Furthermore, we identified lysine 201 as a potential ubiquitination site. A lysine to arginine mutation abolished MITF (K201R) degradation by hUBC9 in vivo. Our experiments indicate that by targeting MITF for proteasome degradation, hUBC9 is a critical regulator of melanocyte differentiation.


Serum metabolite profile associated with incident type 2 diabetes in Koreans: findings from the Korean Genome and Epidemiology Study.
The identification of metabolic alterations in type 2 diabetes (T2D) is useful for elucidating the pathophysiology of the disease and in classifying high-risk individuals. In this study, we prospectively examined the associations between serum metabolites and T2D risk in a Korean community-based cohort (the Ansan-Ansung cohort). Data were obtained from 1,939 participants with available metabolic profiles and without diabetes, cardiovascular disease, or cancer at baseline. The acylcarnitine, amino acid, amine, and phospholipid levels in fasting serum samples were analyzed by targeted metabolomics. During the 8-year follow-up period, we identified 282 cases of incident T2D. Of all metabolites measured, 22 were significantly associated with T2D risk. Specifically, serum levels of alanine, arginine, isoleucine, proline, tyrosine, valine, hexose and five phosphatidylcholine diacyls were positively associated with T2D risk, whereas lyso-phosphatidylcholine acyl C17:0 and C18:2 and other glycerophospholipids were negatively associated with T2D risk. The associated metabolites were further correlated with T2D-relevant risk factors such as insulin resistance and triglyceride indices. In addition, a healthier diet (as measured by the modified recommended food score) was independently associated with T2D risk. Alterations of metabolites such as amino acids and choline-containing phospholipids appear to be associated with T2D risk in Korean adults.


Genetic Data from Nearly 63,000 Women of European Descent Predicts DNA Methylation Biomarkers and Epithelial Ovarian Cancer Risk.
DNA methylation is instrumental for gene regulation. Global changes in the epigenetic landscape have been recognized as a hallmark of cancer. However, the role of DNA methylation in epithelial ovarian cancer (EOC) remains unclear. In this study, high-density genetic and DNA methylation data in white blood cells from the Framingham Heart Study (N = 1,595) were used to build genetic models to predict DNA methylation levels. These prediction models were then applied to the summary statistics of a genome-wide association study (GWAS) of ovarian cancer including 22,406 EOC cases and 40,941 controls to investigate genetically predicted DNA methylation levels in association with EOC risk. Among 62,938 CpG sites investigated, genetically predicted methylation levels at 89 CpG were significantly associated with EOC risk at a Bonferroni-corrected threshold of P < 7.94 × 10-7. Of them, 87 were located at GWAS-identified EOC susceptibility regions and two resided in a genomic region not previously reported to be associated with EOC risk. Integrative analyses of genetic, methylation, and gene expression data identified consistent directions of associations across 12 CpG, five genes, and EOC risk, suggesting that methylation at these 12 CpG may influence EOC risk by regulating expression of these five genes, namely MAPT, HOXB3, ABHD8, ARHGAP27, and SKAP1. We identified novel DNA methylation markers associated with EOC risk and propose that methylation at multiple CpG may affect EOC risk via regulation of gene expression. SIGNIFICANCE: Identification of novel DNA methylation markers associated with EOC risk suggests that methylation at multiple CpG may affect EOC risk through regulation of gene expression.


The role of calmodulin-binding sites in the regulation of the Drosophila TRPL cation channel expressed in Xenopus laevis oocytes by ca2+, inositol 1,4,5-trisphosphate and GTP-binding proteins.
The roles of calmodulin-binding sites in the regulation by Ca2+, inositol 1,4,5-trisphosphate (InsP3) and GTP-binding regulatory proteins (G-proteins) of the Drosophila melanogaster TRPL (transient-receptor-potential-like) non-specific Ca2+ channel were investigated. Wild-type TRPL protein and two mutant forms, TRPL (W713G) and TRPL (W814G), in which a key tryptophan residue in each of the two putative calmodulin-binding sites (Sites 1 and 2, respectively) was replaced by glycine, were expressed heterologously in Xenopus laevis oocytes. Immunofluorescence studies indicated that the expressed TRPL, TRPL (W713G) and TRPL (W814G) proteins are located at the plasma membrane. TRPL oocytes (oocytes injected with trpl cRNA) and TRPL (W814G) oocytes [oocytes injected with trpl (W814G) cRNA] exhibited substantially greater rates of basal (constitutive) Ca2+ inflow (measured using fluo-3 and the Ca2+ add-back protocol) than mock-injected oocytes (mock oocytes). In TRPL (W713G) oocytes, this difference was abolished. In TRPL and TRPL (W814G) [oocytes injected with trpl (W713G) cRNA], but not in TRPL (W713G) oocytes, basal Ca2+ inflow was inhibited by W13, an inhibitor of calmodulin action. Calmodulin (3 muM intracellular) inhibited basal Ca2+ inflow in TRPL but not in TRPL (W713G) or TRPL (W814G) oocytes. Staurosporin, an inhibitor of protein kinase C (PKC), inhibited, while PMA (an activator of PKC) stimulated, basal Ca2+ inflow in TRPL oocytes. In oocytes incubated in the presence of PMA (to suppress Ca2+ inflow through endogenous receptor-activated Ca2+ channels), the InsP3-induced stimulation of Ca2+ inflow through TRPL channels was more clearly evident than in oocytes incubated in the absence of PMA. InsP3 caused a significant stimulation of Mn2+ inflow in TRPL but not in mock oocytes. Rates of InsP3-stimulated Ca2+ inflow through the TRPL, TRPL (W713G) and TRPL (W814G) channels were similar. The ability of GTPgammaS to stimulate Ca2+ inflow through TRPL channels was inhibited by 50% in TRPL (W713G) oocytes but was unaffected in TRPL (W814G) oocytes. It is concluded that, in the environment of the Xenopus oocyte, the Drosophila TRPL channel is activated by (a) interaction with Ca2+/calmodulin at calmodulin-binding Site 1; (b) PKC; (c) InsP3 in a process that does not involve Ca2+ and calmodulin; and (d) a trimeric G-protein(s) through both a Ca2+/calmodulin-dependent and a Ca2+/calmodulin-independent mechanism.


Detonative propagation and accelerative expansion of the Crab Nebula shock front.
The accelerative expansion of the Crab Nebula's outer envelope is a mystery in dynamics, as a conventional expanding blast wave decelerates when bumping into the surrounding interstellar medium. Here we show that the strong relativistic pulsar wind bumping into its surrounding nebula induces energy-generating processes and initiates a detonation wave that propagates outward to form the current outer edge, namely, the shock front, of the nebula. The resulting detonation wave, with a reactive downstream, then provides the needed power to maintain propagation of the shock front. Furthermore, relaxation of the curvature-induced reduction of the propagation velocity from the initial state of formation to the asymptotic, planar state of Chapman-Jouguet propagation explains the observed accelerative expansion. Potential richness in incorporating reactive fronts in the description of various astronomical phenomena is expected.


Characterization of a novel ubiquitin-conjugating enzyme that regulates beta1,4-galactosyltransferase-1 in embryonic stem cells.
In this study we identified a novel galactosyltransferase 1-associating protein (GTAP) by cDNA cloning from a murine embryonic cDNA library using the two-hybrid yeast system. GTAP is expressed in early embryonic tissues, as well as in adult tissues with active cell turnover, and belongs to the class III ubiquitin-conjugating (E2) enzyme family. Its COOH-terminal domain contains a consensus sequence for ubiquitin binding shared by all the ubiquitin-conjugating enzymes, whereas its NH(2)-terminal domain appears critical for the binding and internalization of cell surface galactosyltransferase 1 (GalT1) in embryonic stem cells through a monensin- and MG132-dependent pathway. We have found that GTAP regulates GalT1-associated, laminin-dependent embryonic cell adhesion and the formation of embryoid bodies. Thus, GTAP functions as an evolutionarily conserved E2 enzyme, which may participate in intercellular adhesion and embryonic development. Disclosure of potential conflicts of interest is found at the end of this article.


2013 White Paper on recent issues in bioanalysis: 'hybrid'--the best of LBA and LCMS.
The 2013 7th Workshop on Recent Issues in Bioanalysis was held in Long Beach, California, USA, where close to 500 professionals from pharmaceutical and biopharmaceutical companies, CROs and regulatory agencies convened to discuss current topics of interest in bioanalysis. These 'hot' topics, which covered both small and large molecules, were the starting point for fruitful exchanges of knowledge, and sharing of ideas among speakers, panelists and attendees. The discussions led to specific recommendations pertinent to bioanalytical science. Such as the previous editions, this 2013 White Paper addresses important bioanalytical issues and provides practical answers to the topics presented, discussed and agreed upon by the global bioanalytical community attending the 7th Workshop on Recent Issues in Bioanalysis.


Mitral valve prolapse complicated by acute cerebral embolism, arrhythmias and painless myocardial infarction. A case presentation and overview.
A case of 'primary' mitral valve prolapse is documented. The patient was admitted with right-sided hemiplegia of sudden onset, probably caused by a cerebral embolus from the mitral valve. He also had a painless transmural inferior myocardial infarction (MI) of indeterminate age which was diagnosed electrocardiographically and on left ventricular cine angiography. Since selective coronary arteriography delineated the absence of fixed obstructive atherosclerotic disease, and since coronary vasospasm could not be provoked with the ergonovine (ergometrine) maleate test, it is further postulated that a coronary embolus from the abnormal mitral valve apparatus was responsible for the painless MI. A percutaneous right ventricular endomyocardial biopsy specimen displayed findings not indicative of a 'cardiomyopathy'.


Do subjective cognitive complaints correlate with cognitive impairment in systemic lupus erythematosus? A Danish outpatient study.
This study examined the prevalence of cognitive impairment and its association with depressive symptoms and self-reported cognitive complaints in Danish outpatients with systemic lupus erythematosus (SLE). Fifty-seven consecutive female SLE-outpatients were examined with a comprehensive neuropsychological test-battery, a 20-item self-administered Perceived Deficits Questionnaire (PDQ) and a self-rated depression scale (Major Depression Inventory). Twenty-two patients (38.5%) were classified as cognitively impaired, mostly with deficits in executive functions and attention. Among cognitively impaired patients only 18.2% had significantly higher PDQ scores than the normal range. PDQ scores were highly correlated to depressive symptoms (r = 0.67, p < 0.001). Only two neuropsychological tests were significantly correlated with subjective cognitive complaints. When these variables and self-rated depression score were entered into a regression model both depression score and Symbol Digit Modalities Test performances were significantly associated with the PDQ score. In conclusion, cognitive impairments were common in this group of (mild) SLE outpatients, but the level of significant subjective cognitive complaints was low even among patients with cognitive impairment. Affective status may influence subjective experience of cognitive functions even more than cognitive functioning itself, and absence of subjective cognitive complaints did not exclude the presence of cognitive impairments.


1910s' brains revisited. Cortical complexity in early 20th century patients with intellectual disability or with dementia praecox.
The idea of cortical surface anomalies in subjects with intellectual disability (mental retardation) and schizophrenia can be traced back to early 20th century qualitative observations. Since it is unknown whether modern quantitative measures of cortical complexity and folding would retrieve those early empirical observations, we measured fractal dimension and sulcal span index in photographs of human brains taken in the 1910's. Brain photographs were compared between 36 patients with mental retardation and 21 patients with dementia praecox for the fractal dimension and sulcal span index. Also, a mental retardation subgroup with no-or-non-understandable speech (n = 12) was compared with a subgroup with comprehensible speech (n = 23). Mental retardation group had a lower whole-brain fractal dimension than dementia praecox, and a higher sulcal span index in left posterior cortex. The mental retardation subgroup with comprehensible speech had a lower fractal dimension in left hemisphere than the subgroup with no-or-non-understandable speech and a lower sulcal index in left posterior cortex. Measures of cortical complexity and folding suggest differences between mental retardation and dementia praecox, and regional variations according to language abilities in mental retardation. The findings provide a unique picture of cortical surface changes in their original untreated form, one century ago.


Microwave-dried or air-dried? Consumers' stated preferences and attitudes for organic dried strawberries. A multi-country investigation in Europe.
Non-thermal food processing technologies are becoming more important in the organic food sector because, beyond preserving the organic feature, they could offer organic products with additional benefits in terms of enhanced nutritional content and healthiness as well as better sensory properties that could satisfy the more complex demands of organic consumers. Berries have a well-known health benefits and show increasing market shares in European markets while dehydration can increase the food convenience in terms of extended shelf-life. This study investigates for the first time organic consumers' stated preferences, attitudes and individual differences for a non-thermal organic processing technology. Specifically, we investigated consumers' preferences for organic dried strawberries varying in drying technology used, such as the most conventional (i.e. thermal) air drying and the most innovative (i.e. non-thermal) microwave drying, origin, price levels, and nutrient contents in three European countries: Norway, Romania and Turkey. Data from a total of 614 consumers were collected through an online choice experiment. Results show that on average consumers prefer organic dried strawberries produced with air drying technology that have national origin, with natural nutrient content and at low price, but country and individual differences are identified. Consumers who showed least rejection for microwave dried products are young, mostly from Norway and have higher positive attitudes towards new food technologies. Consumers who showed most rejection for microwave dried products are older, mostly from Turkey and have higher positive attitudes for organic, natural and ecological products. Organic producers who adopt microwave drying might better inform consumers about the characteristics, the process and highlight the nutritional benefits of such technology. Finally, this research informs policy makers about the need to define and regulate more clearly microwave drying as an organic technology, as well as to regulate labelling to ensure that consumers are not misled and correctly informed about the new technology.


A heliocentric view of leptin.
Leptin is significantly broadening our understanding of the mechanisms underlying neuroendocrine function. Initially, based on a rather static view of the hormone, most investigations focused on the effects of leptin on food intake control and body-weight homeostasis, with attention primarily focused on the implications of leptin as a lipostatic factor and central satiety agent. However, the almost ubiquitous distribution of leptin receptors in peripheral tissues provided a fertile area for investigation and a more dynamic view of leptin started to unfold. This adipocyte-derived circulating peptidic hormone, with a tertiary structure resembling that of members of the long-chain helical cytokine family, has generated an enormous interest in the interaction as well as integration between brain targets and peripheral signals. Considerable evidence for systemic effects of leptin on specific tissues and metabolic pathways indicates that leptin operates both directly and indirectly to orchestrate complex pathophysiological processes. Disentangling the biochemical and molecular mechanisms in which leptin is involved represents one of the major challenges ahead.


Decay-accelerating factor is present on paroxysmal nocturnal hemoglobinuria erythroid progenitors and lost during erythropoiesis in vitro.
A glycoprotein that regulates the deposition of C3b on the erythrocyte surface, called decay-accelerating factor or DAF, is absent from the red blood cells (RBC) of patients with paroxysmal nocturnal hemoglobinuria (PNH), explaining in part their abnormal sensitivity to complement. We used a specific antiserum to DAF, flow microfluorometry, and clonogenic assays for erythroid progenitor cells to study PNH erythropoiesis in vitro. By fluorescence-activated cell sorter analysis, all RBC from normal individuals are DAF+. In contrast, the RBC of six patients with PNH showed discrete populations of DAF- cells (10-44%; x +/- SEM = 31 +/- 6%). The DAF- RBC population was partly eliminated by prior acidified serum lysis. To determine whether erythropoietic progenitors expressed DAF, bone marrow cells were sorted by flow microfluorometry and the separated DAF+ and DAF- populations then cultured in vitro. In two normal individuals, but also in six patients with PNH, erythroid colonies formed only from cells in the DAF+ fraction. However, a variable proportion of the normoblast progeny of these DAF+ progenitor cells from patients with PNH was DAF-. Individual bursts removed from cultures of PNH bone marrow showed two discrete populations by fluorescence; the majority of normoblasts were DAF-, only 3 of 27 individual bursts had greater than 50% DAF+ cells, and in three patients, DAF- normoblasts averaged 79%. In contrast, the progeny of individual bursts from normal individuals comprised a unimodal DAF+ population. In each PNH patient, one normal burst (greater than 80% DAF+ normoblasts) was detected, possibly reflecting a normal residual population of erythroid progenitors. By the criterion of DAF expression, there was no evidence of separate populations of normal and PNH type progenitor cells. The phenotypically normal erythroid progenitors of PNH bone marrow acquire the PNH characteristics during differentiation in vitro.


Genome instability drives epistatic adaptation in the human pathogen Leishmania.
How genome instability is harnessed for fitness gain despite its potential deleterious effects is largely elusive. An ideal system to address this important open question is provided by the protozoan pathogen Leishmania, which exploits frequent variations in chromosome and gene copy number to regulate expression levels. Using ecological genomics and experimental evolution approaches, we provide evidence that Leishmania adaptation relies on epistatic interactions between functionally associated gene copy number variations in pathways driving fitness gain in a given environment. We further uncover posttranscriptional regulation as a key mechanism that compensates for deleterious gene dosage effects and provides phenotypic robustness to genetically heterogenous parasite populations. Finally, we correlate dynamic variations in small nucleolar RNA (snoRNA) gene dosage with changes in ribosomal RNA 2'-O-methylation and pseudouridylation, suggesting translational control as an additional layer of parasite adaptation. Leishmania genome instability is thus harnessed for fitness gain by genome-dependent variations in gene expression and genome-independent compensatory mechanisms. This allows for polyclonal adaptation and maintenance of genetic heterogeneity despite strong selective pressure. The epistatic adaptation described here needs to be considered in Leishmania epidemiology and biomarker discovery and may be relevant to other fast-evolving eukaryotic cells that exploit genome instability for adaptation, such as fungal pathogens or cancer.


LincRNA-p21 activates endoplasmic reticulum stress and inhibits hepatocellular carcinoma.
LincRNA-p21 is a downstream long non-coding RNA (lncRNA) transcript of p53. LincRNA-p21 serves as a repressor in p53-dependent transcriptional responses and participates in diverse biological processes, including apoptosis, cell cycle, metabolism and pluripotency. However, the role of lincRNA-p21 in human hepatocellular carcinoma remains to be defined. Here in this work, we demonstrated that lincRNA-p21 acted as a tumor suppressive lncRNA in human hepatocellular carcinoma. We firstly found the downregulation of lincRNA-p21 level in human hepatocellular carcinoma tissues, and showed that low expression of lincRNA-p21 was associated with high disease stage and predicted poor survival. Further we showed that lincRNA-p21 knockdown promoted proliferation and colony formation of HepG2, Huh7 and Bel-7042 cells in vitro, while lincRNA-p21 overexpression obtained oppose results. Using tumor xenograft experiments, we also demonstrated that lincRNA-p21 inhibited HepG2 cell growth in vivo and lincRNA-p21 contributed to sorafenib-induced growth regression of HepG2 cell in vivo. Further mechanism analysis revealed that lincRNA-p21 promoted ER stress both in vitro and in vivo, which facilitated apoptosis of hepatocellular carcinoma cells. Finally, we demonstrated that ER stress accounted for lincRNA-p21 effects on apoptosis, proliferation and in vivo growth of hepatocellular carcinoma. These findings implicate that lincRNA-p21 is a potential prognostic factor and therapeutic target for human hepatocellular carcinoma.


Comparing high and low acculturated mothers and physical activity in Hispanic children.
Parents represent a key ecological component in influencing their child's physical activity. The aim of this exploratory study was to assess the relationship between maternal acculturation and physical activity in Hispanic children. 102 Hispanic mothers (mean age 36.2 yrs; +SD 7.3 yrs) and their children (mean age 10.0 yrs, +SD 0.8 yrs) participated. Most of the mothers (74%) were foreign-born, with 62% classified as low acculturated and 38% high acculturated. Demographic, acculturation, and anthropometric measures were completed by mothers and children. Physical activity was measured using accelerometers. Relationships between maternal acculturation and demographic variables and children's physical activity were examined using chi-square, Analysis of Variance, and simple regression. Children had higher physical activity levels than their mothers (t(49) = -7.87, P < .0001). Significant correlations between maternal and child's physical activity levels were observed in moderate (r2 = 0.13, P = .001), vigorous (r2 = 0.08, P = .05), and moderate-to-vigorous physical activity (r2 = 0.17, P = .002). Low acculturated mothers were more likely to have active children compared with high acculturated mothers. Maternal BMI and other demographic characteristics were not significantly associated with child's physical activity. Findings from this study revealed an association among maternal acculturation, role modeling, and child's physical activity.


Analyzing Somatic Genome Rearrangements in Human Cancers by Using Whole-Exome Sequencing.
Although exome sequencing data are generated primarily to detect single-nucleotide variants and indels, they can also be used to identify a subset of genomic rearrangements whose breakpoints are located in or near exons. Using >4,600 tumor and normal pairs across 15 cancer types, we identified over 9,000 high confidence somatic rearrangements, including a large number of gene fusions. We find that the 5' fusion partners of functional fusions are often housekeeping genes, whereas the 3' fusion partners are enriched in tyrosine kinases. We establish the oncogenic potential of ROR1-DNAJC6 and CEP85L-ROS1 fusions by showing that they can promote cell proliferation in vitro and tumor formation in vivo. Furthermore, we found that ∼4% of the samples have massively rearranged chromosomes, many of which are associated with upregulation of oncogenes such as ERBB2 and TERT. Although the sensitivity of detecting structural alterations from exomes is considerably lower than that from whole genomes, this approach will be fruitful for the multitude of exomes that have been and will be generated, both in cancer and in other diseases.


Functional role of A-type potassium currents in rat presympathetic PVN neurones.
Despite the fact that paraventricular nucleus (PVN) neurones innervating the rostral ventrolateral medulla (RVLM) play important roles in the control of sympathetic function both in physiological and pathological conditions, the precise mechanisms controlling their activity are still incompletely understood. In the present study, we evaluated whether the transient outward potassium current I(A) is expressed in PVN-RVLM neurones, characterized its biophysical and pharmacological properties, and determined its role in shaping action potentials and firing discharge in these neurones. Patch-clamp recordings obtained from retrogradely labelled, PVN-RVLM neurones indicate that a 4-AP sensitive, TEA insensitive current, with biophysical properties consistent with I(A), is present in these neurones. Pharmacological blockade of I(A) depolarized resting V(m) and prolonged Na(+) action potential duration, by increasing its width and by slowing down its decay time course. Interestingly, blockade of I(A) either increased or decreased the firing activity of PVN-RVLM neurones, supporting the presence of subsets of PVN-RVLM neurones differentially modulated by I(A). In all cases, the effects of I(A) on firing activity were prevented by a broad spectrum Ca(2+) channel blocker. Immunohistochemical studies suggest that I(A) in PVN-RVLM neurons is mediated by Kv1.4 and/or Kv4.3 channel subunits. Overall, our results demonstrate the presence of I(A) in PVN-RVLM neurones, which actively modulates their action potential waveform and firing activity. These studies support I(A) as an important intrinsic mechanism controlling neuronal excitability in this central presympathetic neuronal population.


Enzymatic hydrolysis of wheat arabinoxylan by a recombinant "minimal" enzyme cocktail containing beta-xylosidase and novel endo-1,4-beta-xylanase and alpha-l-arabinofuranosidase activities.
This study describes the identification of the key enzyme activities required in a "minimal" enzyme cocktail able to catalyze hydrolysis of water-soluble and water-insoluble wheat arabinoxylan and whole vinasse, a fermentation effluent resulting from industrial ethanol manufacture from wheat. The optimal arabinose-releasing and xylan-depolymerizing enzyme activities were identified from data obtained when selected, recombinant enzymes were systematically supplemented to the different arabinoxylan substrates in mixtures; this examination revealed three novel alpha-l-arabinofuranosidase activities: (i) one GH51 enzyme from Meripilus giganteus and (ii) one GH51 enzyme from Humicola insolens, both able to catalyze arabinose release from singly substituted xylose; and (iii) one GH43 enzyme from H. insolens able to catalyze the release of arabinose from doubly substituted xylose. Treatment of water-soluble and water-insoluble wheat arabinoxylan with an enzyme cocktail containing a 20%:20%:20%:40% mixture and a 25%:25%:25%:25% mixture, respectively, of the GH43 alpha-l-arabinofuranosidase from H. insolens (Abf II), the GH51 alpha-l-arabinofuranosidase from M. giganteus (Abf III), a GH10 endo-1,4-beta-xylanase from H. insolens (Xyl III), and a GH3 beta-xylosidase from Trichoderma reesei (beta-xyl) released 322 mg of arabinose and 512 mg of xylose per gram of water-soluble wheat arabinoxylan dry matter and 150 mg of arabinose and 266 mg of xylose per gram of water-insoluble wheat arabinoxylan dry matter after 24 h at pH 5, 50 degrees C. A 10%:40%:50% mixture of Abf II, Abf III, and beta-xyl released 56 mg of arabinose and 91 mg of xylose per gram of vinasse dry matter after 24 h at pH 5, 50 degrees C. The optimal dosages of the "minimal" enzyme cocktails were determined to be 0.4, 0.3, and 0.2 g enzyme protein per kilogram of substrate dry matter for the water-soluble wheat arabinoxylan, the water-insoluble wheat arabinoxylan, and the vinasse, respectively. These enzyme protein dosage levels were approximately 14, approximately 18, and approximately 27 times lower than the dosages used previously, when the same wheat arabinoxylan substrates were hydrolyzed with a combination of Ultraflo L and Celluclast 1.5 L, two commercially available enzyme preparations produced by H. insolens and T. reesei.


Beta-cell destruction in NOD mice correlates with Fas (CD95) expression on beta-cells and proinflammatory cytokine expression in islets.
A mechanism of autoimmune destruction of islet beta-cells in type 1 diabetes has been proposed to be the binding of Fas ligand (FasL) on T-cells to Fas receptors on beta-cells. We investigated this proposal by examining the expression of FasL and Fas on islet-infiltrating T-cells and beta-cells in relation to beta-cell destruction in a syngeneic islet transplant model in NOD mice. Diabetic NOD mice were transplanted with syngeneic islets and injected with complete Freund's adjuvant, which prevented diabetes recurrence (nondestructive insulitis), and with phosphate-buffered saline, which did not (beta-cell destructive insulitis). Two-color immunohistochemical assays revealed that FasL was expressed on CD4+ T-cells, CD8+ T-cells, and beta-cells in islet grafts from both diabetic and normoglycemic mice, and the percentage of each type of cell that expressed FasL was greater in islet grafts from normoglycemic compared with diabetic mice. In contrast, Fas was expressed on CD4+ T-cells, CD8+ T-cells, and beta-cells in islet grafts from diabetic mice, but it was nearly or totally absent on these cells in islet grafts from normoglycemic mice. Similarly, polymerase chain reaction analysis of islet grafts revealed that Fas mRNA expression was significantly lower in islet grafts from normoglycemic compared with diabetic mice. Also, mRNA levels of interleukin (IL)-1alpha, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma were significantly lower in islet grafts from normoglycemic mice. Finally, Fas was induced on NOD islet cells by incubation with IL-1beta, IFN-gamma, and the combination of IL-1beta, TNF-alpha, and IFN-gamma. These findings support the concept that cytokine-induced Fas receptor expression on islet beta-cells is a mechanism for their destruction by FasL-expressing CD4+ and CD8+ T-cells and, possibly, by FasL-expressing beta-cells themselves.


Empathy and altruism.
The psychophysiological responses of 60 subjects were measured as they observed a performer play a roulette game. Half of the subjects were led to believe that they were similar to the performer in personality and values, and half were led to believe that they were dissimilar. Half of the subjects in each condition believed that the performer won money and experienced pain as he played the game, and half believed that he performed a cognitive and motor skill task. Subjects who observed a performer who ostensibly experienced pleasure and pain exhibited greater psychophysiological reactions than subjects who did not. Subjects who believed they were similar to the performer tended to react more strongly than subjects who believed they were different from him. Similar subjects also reported identifying most with the performer and feeling the worst while he waited to receive shocks. It was concluded that the similar subjects empathized most with the performer who appeared to experience pleasure and pain. When required to make a choice between helping themselves at a cost to the performer or helping the performer at a cost to themselves, the subjects who reacted most empathically behaved most altruistically. The results were interpreted as casting some light on century-old questions about the human capacity for altruism.


Genetically Determined ABO Blood Group and its Associations With Health and Disease.
To determine the spectrum of phenotypes linked to the ABO blood group system, using genetic determinants of the ABO blood group system. Approach and Results: We assessed the risk of 41 health and disease outcomes, and 36 linear traits associated with the ABO blood group system in the UK Biobank cohort. A total of 406 755 unrelated individuals were included in this study. Blood groups A, B, and O were determined based on allele combinations of previously established single-nucleotide polymorphisms rs8176746, rs8176719 in the ABO gene. Group AB was excluded because of its relative small sample size. Overall, 187 387 (46%) were male with a mean (SD) age of 57±8.1 years and a median total exposure of 64 person-years (interquartile range, 57-70). Of 406 755 individuals, 182 621 (44.9%) participants had blood group O, 182 786 (44.9%) had blood group A, and 41 348 (10.2%) had blood group B. ABO blood groups were associated with 11 health and disease outcomes (P<2.19×10-4). ABO blood groups were primarily associated with cardiovascular outcomes. Compared with individuals with blood group O, blood groups A and B were associated with increased odds of up to 1.56 (95% CI, 1.43-1.69) for thromboembolic events and decreased odds for hypertension (0.94 [95% CI, 0.92-0.97]). The ABO blood group system is associated with several parameters of healthy aging and disease development. Knowledge of ABO blood groups might be of interest for more personalized approaches towards health maintenance and the prevention of diseases.


High flow nasal cannula in acute hypercapnic exacerbation of chronic obstructive pulmonary disease: an emerging utility.
High flow nasal cannula (HFNC) provides warmed and humidified air with flow rates up to 60 liters/min with relatively fixed oxygen content (FiO2). It has been extensively evaluated for hypoxemic respiratory failure and has been used in mild acute respiratory distress syndrome, pre-intubation, bronchoscopy and pediatric obstructive sleep apnea. Recent data has suggested a role in stable hypercapnic chronic obstructive pulmonary disease (COPD) and even in acute exacerbations, though, the use has not been advocated by any guidelines yet. We present a case of acute hypercapnic exacerbation of COPD, intolerant to non-invasive ventilation, showing response and improvement on use of HFNC. This case highlights this potential mechanisms and prospects for the same.


Long-term blood pressure variability throughout young adulthood and cognitive function in midlife: the Coronary Artery Risk Development in Young Adults (CARDIA) study.
Whether long-term blood pressure (BP) variability throughout young adulthood is associated with cognitive function in midlife remains uncertain. Using data from the Coronary Artery Risk Development in Young Adults (CARDIA), which recruited healthy young adults aged 18 to 30 years (mean age, 25 years) at baseline (Y(0)), we assessed BP variability by SD and average real variability (ARV) for 25 years (8 visits). Cognitive function was assessed with the Digit Symbol Substitution Test (psychomotor speed test), the Rey Auditory Verbal Learning Test (verbal memory test), and the modified Stroop test (executive function test) at follow-up (Y(25)). At the Y(25) examination, participants (n=2326) had a mean age of 50.4 years, 43% were men, and 40% were black. In multivariable-adjusted linear regression models, higher ARV(SBP), ARV(DBP), and SD(DBP) were significantly associated with lower scores of Digit Symbol Substitution Test (β [SE]: -0.025 [0.006], -0.029 [0.007], and -0.029 [0.007], respectively; all P<0.001) and Rey Auditory Verbal Learning Test (β [SE]: -0.016 [0.006], -0.021 [0.007], and -0.019 [0.007], respectively; all P<0.05) after adjustment for demographic and clinical characteristics and with cumulative exposure to BP through Y(0) to Y(25). Neither SDBP nor ARV(BP) was associated with the Stroop score. The associations between ARV(BP) or SD(BP) and each cognitive function test were similar between blacks and whites except for 1 significant interaction between race and SDS(BP) on the Digit Symbol Substitution Test (P<0.05). Long-term BP variability for 25 years beginning in young adulthood was associated with worse psychomotor speed and verbal memory tests in midlife, independent of cumulative exposure to BP during follow-up.


Salivary glutathione in bipolar disorder: A pilot study.
Glutathione (GSH) is an important cellular antioxidant and its levels are decreased in some studies of bipolar patients. Saliva provides a simple and feasible means of measuring GSH but has not yet been applied to the study of bipolar disorder. The purpose of the study was to compare salivary levels of GSH and oxidized glutathione (GSSG) in bipolar patients and healthy controls. Saliva was sampled from 22 medicated, euthymic patients with bipolar disorder and 20 healthy controls. GSH and GSSG were measured using an enzyme kinetic essay. GSH and GSSG were significantly higher in saliva from bipolar patients relative to controls. The ratio of GSH:GSSG was unchanged. There was no correlation between the measured clinical characteristics of the patients and GSH levels. The main limitation of the study was the small sample size. Patients were medicated which may have influenced saliva production and hence GSH levels. In addition, salivary GSH may not reflect GSH status in tissues more directly involved in the pathophysiology of bipolar disorder. Salivary GSH can be readily measured in bipolar patients. Relative to controls, salivary levels of GSH and GSSG were increased in bipolar patients but their ratio was unchanged. The origin and significance of these change requires further study.


Signal intensity analysis and optimization for in vivo imaging of Cherenkov and excited luminescence.
During external beam radiotherapy (EBRT), in vivo Cherenkov optical emissions can be used as a dosimetry tool or to excite luminescence, termed Cherenkov-excited luminescence (CEL) with microsecond-level time-gated cameras. The goal of this work was to develop a complete theoretical foundation for the detectable signal strength, in order to provide guidance on optimization of the limits of detection and how to optimize near real time imaging. The key parameters affecting photon production, propagation and detection were considered and experimental validation with both tissue phantoms and a murine model are shown. Both the theoretical analysis and experimental data indicate that the detection level is near a single photon-per-pixel for the detection geometry and frame rates commonly used, with the strongest factor being the signal decrease with the square of distance from tissue to camera. Experimental data demonstrates how the SNR improves with increasing integration time, but only up to the point where the dominance of camera read noise is overcome by stray photon noise that cannot be suppressed. For the current camera in a fixed geometry, the signal to background ratio limits the detection of light signals, and the observed in vivo Cherenkov emission is on the order of 100× stronger than CEL signals. As a result, imaging signals from depths <15 mm is reasonable for Cherenkov light, and depths <3 mm is reasonable for CEL imaging. The current investigation modeled Cherenkov and CEL imaging of two oxygen sensing phosphorescent compounds, but the modularity of the code allows for easy comparison of different agents or alternative cameras, geometries or tissues.


Immunoglobulin heavy variable (IGHV) genes and alleles: new entities, new names and implications for research and prognostication in chronic lymphocytic leukaemia.
Νext generation sequencing studies in Homo sapiens have identified novel immunoglobulin heavy variable (IGHV) genes and alleles necessitating changes in the international ImMunoGeneTics information system (IMGT) GENE-DB and reference directories of IMGT/V-QUEST. In chronic lymphocytic leukaemia (CLL), the somatic hypermutation (SHM) status of the clonotypic rearranged IGHV gene is strongly associated with patient outcome. Correct determination of this parameter strictly depends on the comparison of the nucleotide sequence of the clonotypic rearranged IGHV gene with that of the closest germline counterpart. Consequently, changes in the reference directories could, in principle, affect the correct interpretation of the IGHV mutational status in CLL. To this end, we analyzed 8066 productive IG heavy chain (IGH) rearrangement sequences from our consortium both before and after the latest update of the IMGT/V-QUEST reference directory. Differences were identified in 405 cases (5 % of the cohort). In 291/405 sequences (71.9 %), changes concerned only the IGHV gene or allele name, whereas a change in the percent germline identity (%GI) was noted in 114/405 (28.1 %) sequences; in 50/114 (43.8 %) sequences, changes in the %GI led to a change in the mutational set. In conclusion, recent changes in the IMGT reference directories affected the interpretation of SHM in a sizeable number of IGH rearrangement sequences from CLL patients. This indicates that both physicians and researchers should consider a re-evaluation of IG sequence data, especially for those IGH rearrangement sequences that, up to date, have a GI close to 98 %, where caution is warranted.


AACR White Paper: Shaping the Future of Cancer Prevention - A Roadmap for Advancing Science and Public Health.
The recent pace, extent, and impact of paradigm-changing cancer prevention science has been remarkable. The American Association for Cancer Research (AACR) convened a 3-day summit, aligned with five research priorities: (i) Precancer Atlas (PCA). (ii) Cancer interception. (iii) Obesity-cancer linkage, a global epidemic of chronic low-grade inflammation. (iv) Implementation science. (v) Cancer disparities. Aligned with these priorities, AACR co-led the Lancet Commission to formally endorse and accelerate the NCI Cancer Moonshot program, facilitating new global collaborative efforts in cancer control. The expanding scope of creative impact is perhaps most startling-from NCI-funded built environments to AACR Team Science Awarded studies of Asian cancer genomes informing global primary prevention policies; cell-free epigenetic marks identifying incipient neoplastic site; practice-changing genomic subclasses in myeloproliferative neoplasia (including germline variant tightly linked to JAK2 V617F haplotype); universal germline genetic testing for pancreatic cancer; and repurposing drugs targeting immune- and stem-cell signals (e.g., IL-1β, PD-1, RANK-L) to cancer interception. Microbiota-driven IL-17 can induce stemness and transformation in pancreatic precursors (identifying another repurposing opportunity). Notable progress also includes hosting an obesity special conference (connecting epidemiologic and molecular perspectives to inform cancer research and prevention strategies), co-leading concerted national implementation efforts in HPV vaccination, and charting the future elimination of cancer disparities by integrating new science tools, discoveries and perspectives into community-engaged research, including targeted counter attacks on e-cigarette ad exploitation of children, Hispanics and Blacks. Following this summit, two unprecedented funding initiatives were catalyzed to drive cancer prevention research: the NCI Cancer Moonshot (e.g., PCA and disparities); and the AACR-Stand Up To Cancer bold "Cancer Interception" initiative.


Toll/IL-1 receptor domain-containing adaptor inducing IFN-beta (TRIF) associates with TNF receptor-associated factor 6 and TANK-binding kinase 1, and activates two distinct transcription factors, NF-kappa B and IFN-regulatory factor-3, in the Toll-like receptor signaling.
We previously reported a new Toll/IL-1R (TIR)-containing molecule, named TIR domain-containing adaptor inducing IFN-beta (TRIF). Although initial study indicated that TRIF possesses the ability to activate not only the NF-kappaB-dependent but also the IFN-beta promoters, the molecular mechanisms of TRIF-induced signaling are poorly understood. In this study, we investigated the signaling cascades through TRIF. TNF receptor-associated factor (TRAF)6 interacted with TRIF through the TRAF domain of TRAF6 and TRAF6-binding motifs found in the N-terminal portion of TRIF. Disruption of TRAF6-binding motifs of TRIF disabled it from associating with TRAF6, and resulted in a reduction in the TRIF-induced activation of the NF-kappaB-dependent but not IFN-beta promoter. TANK-binding kinase (TBK)-1, which was recently reported to be a kinase of IFN regulatory factor-3, which is an essential transcription factor for IFN-beta expression, also associated with the N-terminal region of TRIF. Moreover, the association between TRIF and TBK1 appeared to require the kinase activity of TBK1, as well as phosphorylation of TRIF. Because TRAF6 and TBK1 bind close the region of TRIF, it seems that TRAF6 physically prevents the association between TRIF and TBK1. Taken together, these results demonstrate that TRIF associates with TRAF6 and TBK1 independently, and activates two distinct transcription factors, NF-kappaB and IFN regulatory factor-3, respectively.


Possible relevance of pigeons as an indicator species for monitoring air pollution.
Wild city pigeons were caught at four different locations in the Netherlands to represent areas of high (Amsterdam-high), moderate (Amsterdam-medium), and low (Maastricht and Assen) traffic density. It is assumed that local ambient air pollution decreases as a function of traffic density. In these pigeons levels of polycyclic aromatic hydrocarbon (PAH)-DNA adducts, oxidative DNA damage, and heavy metal residues were determined in kidney, lung, liver, and blood (no adduct analysis in blood). The contribution of leaded gasoline to total body lead content was estimated by measuring concentrations of Pb and its isotopes in blood. We also analyzed samples of ambient air particulate matter for PAH and heavy metal concentrations at the four different locations. Interregional differences in heavy metals in ambient air particulate matter were reflected relatively well by pigeon body loads. The higher lead and cadmium concentrations in blood, kidney, liver, and lung were found in the Amsterdam high traffic density area, followed by Amsterdam medium, Assen, and Maastricht. A high Pb concentration in blood coincided with relatively low 206Pb/207Pb values, indicating a high contribution of leaded gasoline to total blood Pb concentrations in pigeons from the Amsterdam high traffic density area. Significantly enhanced blood zinc values were found in pigeons from both locations in Amsterdam compared to pigeons from the other two areas. However, no differences in Zn tissue levels between the four different groups were found. Oxidative DNA damage, determined as the ratio of 7-Hydro-8-oxo-2'-deoxyguanosine/ deoxyguanosine, in pigeon liver was highest in Amsterdam-high, followed by Assen (low traffic density). Pb content, but not the Cd content, was positively associated with oxidative DNA damage in liver tissue. In lung tissue, a negative correlation was found between oxidative DNA damage and Zn content. These results indicate that the carcinogenic potential of Pb might be ascribed to oxygen radical formation, whereas Zn plays a protective role against oxidative DNA damage. Places with high and medium traffic density could be clearly discriminated on the basis of PAH levels in the ambient air. The PAH content in particulate air samples was not, however, reflected in total PAH-related DNA adduct levels because no differences could be observed in tissue adduct levels in pigeons from the four different locations. Our results indicate that wild city pigeons can be used as biological indicators of exposure to heavy metal pollution in outdoor air.


Experiences of partners of professional nurses venting traumatic information.
Professional nurses employed in trauma units encounter numerous stressors in their practice environment. They use different strategies to cope with this stress, including venting traumatic information to their partners and other family members. To describe how partners of professional nurses cope with traumatic information being vented to them. A qualitative research method with an interpretive descriptive inquiry design was used to explore, interpret and describe the coping experiences of the nurses' partners. Purposive sampling was used to select a total of 14 partners, but only ten participated in semi-structured interviews. Tesch's eight steps of open coding were used for data analysis. Four main themes were identified indicating adaptive and maladaptive coping skills, namely partners' experiences of traumatic information vented to them; partners' coping activities; reciprocal communication and relationship support between partners and nurses; and resilience of partners to deal with the nursing profession. Partners employed different ways to cope with traumatic information. It was essential for partners and nurses to be supported by nurses' practice environments and to develop resilience to fulfil reciprocal supportive roles in their relationships.


Preventing Complications in Pregnant Women With Cardiac Disease.
Pregnancy can lead to complications in women with heart disease, and these complications can be life threatening. Understanding serious complications and how they can be prevented is important. The primary objectives were to determine the incidence of serious cardiac events (SCEs) in pregnant women with heart disease, whether they were preventable, and their impact on fetal and neonatal outcomes. Serious obstetric events were also examined. A prospectively assembled cohort of 1,315 pregnancies in women with heart disease was studied. SCEs included cardiac death or arrest, ventricular arrhythmias, congestive heart failure or arrhythmias requiring admission to an intensive care unit, myocardial infarction, stroke, aortic dissection, valve thrombosis, endocarditis, and urgent cardiac intervention. The Harvard Medical Study criteria were used to adjudicate preventability. Overall, 3.6% of pregnancies (47 of 1,315) were complicated by SCEs. The most frequent SCEs were cardiac death or arrest, heart failure, arrhythmias, and urgent interventions. Most SCEs (66%) occurred in the antepartum period. Almost one-half of SCEs (49%) were preventable; the majority of preventable SCEs (74%) were secondary to provider management factors. Adverse fetal and neonatal events were more common in pregnancies with SCEs compared with those without cardiac events (62% vs. 29%; p < 0.001). Serious obstetric events were less common (1.7%) and were primarily due to pre-eclampsia with severe features. Pregnant women with heart disease are at risk for serious cardiac complications, and approximately one-half of all SCEs are preventable. Strategies to prevent serious cardiac complications in this high-risk cohort of women need to be developed.


The DNA sequence of the structural gene of gonococcal protein III and the flanking region containing a repetitive sequence. Homology of protein III with enterobacterial OmpA proteins.
The insert of a lambda gt11 clone expressing gonococcal protein III was sequenced. The deduced amino acid sequence showed a coding frame of 236 amino acids with a typical 22-amino-acid signal peptide, followed by the known NH2-terminal sequence of PIII. The mature protein has a molecular weight of 23,298. It was found that PIII had extensive and very striking homology to the carboxy-terminal portion of enterobacterial OmpA proteins. The homology encompasses the OmpA domain that is believed to be located in the periplasmic space. If the disposition of PIII across the OM is analogous, then the surface-exposed domain consists of less than 40 amino acids. These include a potential 15-amino-acid disulfide loop, a feature not found in OmpA proteins. Hybridization studies with the sequenced insert indicated that it contained a repetitive sequence that occurred at least 20 times in the genome. By additional hybridization studies the area containing the repetitive sequence was narrowed to a region of 43 bp. This region contained an exact copy of the consensus sequence of a 26-bp repetitive sequence recently described. An analogous sequence recurs in an inverted orientation 53 bp downstream.


Beneficial effects of green tea--a review.
Tea is the most consumed drink in the world after water. Green tea is a 'non-fermented' tea, and contains more catechins, than black tea or oolong tea. Catechins are in vitro and in vivo strong antioxidants. In addition, its content of certain minerals and vitamins increases the antioxidant potential of this type of tea. Since ancient times, green tea has been considered by the traditional Chinese medicine as a healthful beverage. Recent human studies suggest that green tea may contribute to a reduction in the risk of cardiovascular disease and some forms of cancer, as well as to the promotion of oral health and other physiological functions such as anti-hypertensive effect, body weight control, antibacterial and antivirasic activity, solar ultraviolet protection, bone mineral density increase, anti-fibrotic properties, and neuroprotective power. Increasing interest in its health benefits has led to the inclusion of green tea in the group of beverages with functional properties. However, although all the evidence from research on green tea is very promising, future studies are necessary to fully understand its contributions to human health, and advise its regular consumption in Western diets, in which green tea consumption is nowadays limited and sporadic.


Prevalence and correlates of habitual snoring in high school students.
To examine the prevalence and correlates of habitual snoring in senior high school students in Korea. A cross-sectional survey. Ten high schools in the southern part of Seoul, Korea. A total of 3,871 high school students (2,703 male students and 1,168 female students; age range, 15 to 18 years; mean age: male students, 16.8 years; female students, 16.9 years) who were attending the 11th grade. Data were collected on sociodemographic characteristics, school performance, and patterns of sleep and sleep-related disturbances. The overall prevalence of habitual snoring was 11.2% (boys, 12.4%; girls, 8.5%). The mean total sleep time was similar in habitual snorers vs nonsnorers (6.4 and 6.3 h per day, respectively). Frequency of snoring increased significantly with body mass index (BMI) [p < 0.001], cigarette smoking (p < 0.01), prevalence of witnessed apnea (p < 0.001), and Epworth sleepiness scale score (p < 0.001). The frequency of snoring increased with a decline in school performance (p < 0.001). For those whose school performance was low, there was a 35% excess in the odds of habitual snoring vs those whose school performance was high (odds ratio, 1.35; 95% confidence interval, 1.01 to 1.78). These findings suggest that chronic habitual snoring is associated with multiple factors in adolescents. Whether interventions to modify BMI and smoking can alter snoring habits and related clinical problems warrants further study, particularly as it also may improve academic performance in high school students.


Disturbed glutathione metabolism and decreased antioxidant levels in human immunodeficiency virus-infected patients during highly active antiretroviral therapy--potential immunomodulatory effects of antioxidants.
Oxidative stress has been implicated in the pathogenesis of human immunodeficiency virus (HIV) infection. We examined the effect of highly active antiretroviral therapy (HAART) on plasma levels of several antioxidants and intracellular glutathione-redox status in CD4+ T cells, in 20 HIV-infected patients. HAART was accompanied by both an improvement of glutathione-redox status and an increase in levels of antioxidant vitamins, without full normalization. Glutathione supplementation in vitro increases T cell proliferation and suppresses the spontaneous release of tumor necrosis factor-alpha from peripheral blood mononuclear cells, in HIV-infected patients receiving HAART. Our findings suggest that therapeutic intervention aimed at normalization of oxidative disturbances in HIV infection could be of interest, in addition to HAART.


Time-resolved fluorometry in end-point and real-time PCR quantification of nucleic acids.
Two time-resolved fluorescence-based methods for nucleic acid quantification are described and their results are compared. Both methods use an exogenous internal standard to eliminate errors arising from different steps of the assay. The first method is a competitive end-point assay, where the standard competes for the same primers with the actual target sequence, prostate-specific antigen (PSA) cDNA. The standard and target are quantified in a dual-label plate hybridization with lanthanide-labelled probes after a fixed number of PCR cycles. The second method is based on real-time monitoring of PCR and on the use of a novel homogeneous signal generation principle that relies on the use of a 5'-->3' exonucleolytic DNA polymerase and a probe labelled with an environment sensitive, stable and fluorescent lanthanide chelate. In this assay, a non-competitive, exogenous internal standard is used. Both assays have a wide linear range (50-5 x 10(6) and 10-5 x 10(7) input PSA cDNA molecules for the end-point and real-time assays, respectively) and there is a strong correlation between the results obtained with the two assays (r = 1.0). Being somewhat faster to perform, the real-time format is better suited for assays that require high throughput.


Heart tumors in Puerto Rico de novo atrial fibrillation as clinical presentation in a subgroup of patients.
Primary cardiac tumors are rare neoplasms in humans, of which the most common is the atrial Myxoma. The objective of this study was to find the incidence of these tumors at the Heart Center of Puerto Rico and the Caribbean. This study was approved by the Institutional Review Board of the Medical Sciences Campus University of Puerto Rico to review the records at the Heart Center of patients with heart tumors in the last 14 years. The sample consists of 55 patients (78.9% were females and 24.1% were male) with a median age of 52 years. Sixty-five percent of patients lived in rural areas. Clinical presentations included shortness of breath (43.1%), chest pain (37.9%), asymptomatic (25%), palpitations (20.7%), neurologic symptoms (10.3%) and dizziness (6.9%). Electrocardiographic findings included normal sinus rhythm (53.4%), non-specific ST-T changes (32.8%), sinus tachycardia (20.7%), left atrial enlargement (10.3%) and atrial fibrillation (8.6%). A subgroup presenting with atrial fibrillation prior to diagnosis had left atrial myxoma. The tumors found, in descending order of frequency are: left atrial myxoma, right atrial myxoma, papillary fibroelastoma, hamartoma, lipoma and rhabdomyoma. We found a correlation between large left atrial myxoma and atrial fibrillation. The most frequent heart tumor was atrial myxoma. The larger myxomas were associated with atrial fibrillation.


Metastasis-associated protein 1 interacts with NRIF3, an estrogen-inducible nuclear receptor coregulator.
The transcriptional activity of estrogen receptor alpha (ER-alpha) is modified by regulatory action and interactions of coactivators and corepressors. Recent studies have shown that the metastasis-associated protein 1 (MTA1) represses estrogen receptor element (ERE)-driven transcription in breast cancer cells. With a yeast two-hybrid screen to clone MTA1-interacting proteins, we identified a known nuclear receptor coregulator (NRIF3) as an MTA1-binding protein. NRIF3 interacted with MTA1 both in vitro and in vivo. NRIF3 bound to the C-terminal region of MTA1, while MTA1 bound to the N-terminal region of NRIF3, containing one nuclear receptor interaction LXXLL motif. We showed that NRIF3 is an ER coactivator, hyperstimulated ER transactivation functions, and associated with the endogenous ER and its target gene promoter. MTA1 repressed NRIF3-mediated stimulation of ERE-driven transcription and interfered with NRIF3's association with the ER target gene chromatin. In addition, NRIF3 deregulation enhanced the responsiveness of breast cancer cells to estrogen-induced stimulation of growth and anchorage independence. Furthermore, we found that NRIF3 is an estrogen-inducible gene and activated ER associated with the ER response element in the NRIF3 gene promoter. These findings suggest that NRIF3, an MTA1-interacting protein, is an estrogen-inducible gene and that regulatory interactions between MTA1 and NRIF3 might be important in modulating the sensitivity of breast cancer cells to estrogen.


Traditional Invasive and Synchrotron-Based Noninvasive Assessments of Three-Dimensional-Printed Hybrid Cartilage Constructs In Situ.
Three-dimensional (3D)-printed constructs made of polycaprolactone and chondrocyte-impregnated alginate hydrogel (hybrid cartilage constructs) can mimic the biphasic nature of articular cartilage, thus offering promise for cartilage tissue engineering applications. Notably, the regulatory pathway for medical device development requires validation of such constructs through in vitro bench tests and in vivo preclinical examinations for premarket approval. For this, noninvasive imaging techniques are required for effective evaluation of the progress of these cartilage constructs, especially when implanted in animal models or human subjects. However, characterization of the individual components of the hybrid cartilage constructs and their associated time-dependent structural changes by currently available noninvasive techniques is challenging as these constructs contain a combination of hydrophobic and hydrophilic biomaterials with different refractive indices. In this study, we report the use of a novel synchrotron radiation inline phase contrast imaging computed tomography (SR-inline-PCI-CT) approach for noninvasive (in situ) characterization of 3D-printed hybrid cartilage constructs that has been implanted subcutaneously in mice over a 21-day period. In parallel, traditional invasive assays were used to evaluate the in vivo performance of the implanted hybrid cartilage constructs with respect to their cell viability and secretion of cartilage-specific extracellular matrix over the 21-day period postimplantation in mice. SR-inline-PCI-CT allowed striking visualization of the individual components within the 3D-printed hybrid cartilage constructs, as well as characterization of the time-dependent structural changes after implantation. In addition, the relationship between the implanted constructs and the surrounding tissues was delineated. Furthermore, traditional assays showed that cell viability within the cartilage constructs was at least 70% at all three time points, and secretion of alcian blue- and collagen type 2-positive matrices increased progressively over the 21-day period postimplantation. Overall, these results demonstrate that the 3D-printed hybrid cartilage constructs have good in vivo performance and validate their potential for regeneration of articular cartilage in vivo. In addition, SR-inline-PCI-CT has demonstrated potential for longitudinal and noninvasive monitoring of the functionality of 3D-printed hybrid cartilage constructs in a way that is translatable to other soft tissue engineering applications.


Effectiveness of "shifting boundaries" teen dating violence prevention program for subgroups of middle school students.
We examine whether the Shifting Boundaries (SB) intervention, a primary intervention to prevent youth dating violence and sexual harassment (DV/H), is differentially effective for girls compared with boys or for youth with a history of DV/H experiences. We randomly assigned SB to 30 public middle schools in New York City, enrolling 117 sixth and seventh grade classes to receive a classroom, building, combined, or neither intervention. The SB classroom intervention included six sessions emphasizing the laws/consequences of DV/H, establishing boundaries and safe relationships. The SB schoolwide/building intervention included the use of school-based restraining orders, greater faculty/security presence in unsafe "hot spots" mapped by students, and posters to increase DV/H awareness and reporting. Student surveys were implemented at baseline, immediately after intervention, and 6 months after intervention. At 6 months after intervention, the SB building-level intervention was associated with significant reductions in the frequency of sexual harassment (SH) perpetration and victimization; the prevalence and frequency of sexual dating violence victimization; and the frequency of total dating violence victimization and perpetration. We also had one anomalous finding that the interventions were associated with an increase in the prevalence of SH victimization. These results were consistent for girls and boys, and those with or without a history of DV/H, with the one exception for those exposed to the SB building condition who had earlier reported perpetrating SH had a significantly lower frequency of perpetrating SH at the follow-up than those without such a history. SB can provide effective universal prevention of middle school DV/H experiences, regardless of students' prior exposure histories, and for boys and girls.


Integrating the teaching role into one's identity: a qualitative study of beginning undergraduate medical teachers.
Beginning medical teachers often see themselves as doctors or researchers rather than as teachers. Using both figured worlds theory and dialogical self theory, this study explores how beginning teachers in the field of undergraduate medical education integrate the teacher role into their identity. A qualitative study was performed, involving 18 beginning medical teachers at a Dutch medical school. The teachers were interviewed twice and kept a logbook over a period of 7 months. The study shows that the integration of the teacher role into the teachers' identity was hampered by the idea that teaching is perceived by others as a low status occupation. Some teachers experienced significant tension because of this, while others showed resilience in resisting the negative associations that were thought to exist regarding teaching. The teachers used five different identity narratives in order to integrate the teacher role into their identity, in which the positions of teacher and doctor or researcher were found to be combined, adopted or rejected in diverse ways. The five identity narratives were: (1) coalition between the I-position of teacher and other I-positions; (2) no integration of the I-position of teacher: holding on to other I-positions; (3) construction of the I-position of teacher and other I-positions as opposites; (4) coalition between the I-position of teacher and a third position of coordinator; and (5) meta-position: trivialising the importance of status. These identity narratives offer starting points for supporting undergraduate teachers during their early professional years.


Tricistronic viral vectors co-expressing interleukin-12 (1L-12) and CD80 (B7-1) for the immunotherapy of cancer: preclinical studies in myeloma.
Synergy between interleukin-12 (IL-12) and B7-1 (CD80) for cancer immunotherapy has previously been demonstrated in animal models of breast cancer, lymphoma, and multiple myeloma. With a view to human clinical application, tricistronic retroviral and adenovirus vectors co-expressing IL-12 (IL-12p40 plus IL-12p35) and CD80 were constructed by utilizing two internal ribosome entry site (IRES) sequences to link the three cDNAs. A murine stem cell virus (MSCV)-based retroviral vector (MSCV-hIL12.B7) utilized distinct IRES sequences from the encephalomyocarditis virus (EMCV) and the foot-and-mouth disease virus (FMCV), whereas Ad5-based adenovirus vectors contained transcriptional units with two EMCV IRES sequences under the control of murine (AdMh12.B7) or human (AdHh12.B7) cytomegalovirus promoters. AdMh12.B7 was found to consistently direct higher levels of IL-12 and CD80 expression than AdHh12.B7 following infection of a number of human tumor cell lines. In preclinical studies, the human myeloma cell line U266 was infected with MSCV-hIL12.B7 and a resulting clonal cell line, U/MSCV-h12.B7, was generated with stable expression of CD80 and secreting IL-12 at 1 ng/24 h/10(6) cells. By comparison, following AdMh12.B7 infection, 81% of infected U266 cells (U/AdMh12.B7) expressed CD80 and secreted IL-12 at 25-50 ng/24 h/10(6) cells. Both engineered myeloma cell lines stimulated enhanced allogeneic mixed lymphocyte proliferation and provoked increases in cytotoxic T-lymphocyte responses and gamma-interferon release from normal donor lymphocytes exposed to parental U266 cells. These results suggest potential clinical utility of AdMh12.B7 in immunotherapy strategies for the treatment of multiple myeloma and other cancers.


Is Crohn's disease rare in India?
Crohn's disease (CD) is believed to be rare in India. To analyze the data pertaining to patients with CD seen in a tertiary referral center. Data on patients with chronic inflammatory bowel disease attending our Unit over a 5-year period were analyzed. The diagnosis of CD was established by the presence of characteristic segmental bowel involvement, consistent histological picture, exclusion of infectious causes, relapsing nature of the disease, response to appropriate drug therapy, and lack of evidence for another etiological factor. Of the 25 patients (age range 12-52, mean 31.7, years) 13 were men. Abdominal pain was present in 21 (84%) patients, diarrhea in 20 (80%), blood per rectum in 11 (44%) and fever in 4 (16%). The ileocecal region was involved in 6 (24%) patients and anal canal in 4 (16%); 19 (76%) had patchy involvement of different segments of the large bowel. Colonoscopic biopsies revealed granulomas in 12 (48%). During a mean follow up of 36.4 (range 6-54) months, 15 (60%) patients had 21 relapses. Despite initial response to 5-aminosalicylic acid in five patients, 23 needed glucocorticoid therapy at least once. CD may not be rare in India. Because of the high prevalence of intestinal tuberculosis there is a possibility that CD may be under-recognized in India.


MCSS functionality maps for a flexible protein.
The Multiple Copy Simultaneous Search (MCSS) methodology for finding energetically favorable positions and orientations of small functional groups in a binding site is extended to include flexibility of the target. This makes possible the finding of novel minima not present in a fixed structure and so extends the diversity of inhibitors that can be constructed starting with the MCSS procedure. Quenched molecular dynamics is used to generate energetically favorable positions and orientations of the functional groups in the field of a flexible protein. The method is applied to the viral protein HIV-1 protease with methanol and methyl ammonium as a test case. If the protein is quenched with many copies of functional groups randomly distributed in the binding site, the resulting minima have ligand-protein interaction energies that are, on average, less favorable than those obtained with standard MCSS. This is a consequence of the renormalized potential function employed in the Locally Enhanced Sampling (LES) approximation. However, local optimizations of existing MCSS minima with a flexible protein results in lower energy minima in regions of the protein that are of particular interest. Their use in constructing a consensus protein model for ligand design is discussed.


Reliability of the Canadian emergency department triage and acuity scale: interrater agreement.
To determine the rate of interobserver reliability of the Canadian Emergency Department Triage and Acuity Scale (CTAS). Ten physicians and 10 nurses were randomly selected to review and assign a triage level on 50 ED case summaries containing presenting complaint, mode of arrival, vital signs, and a verbatim triage note. The rate of agreement within and between groups of raters was determined using kappa statistics. One-way, 2-way analysis of variance (ANOVA) and combined ANOVA were used to quantify reliability coefficients for intraclass and interclass correlations. The overall chance-corrected agreement kappa for all observers was.80 (95% confidence interval [CI] .79 to .81), and the probability of agreement between 2 random observers on a random case was.539. For nurses alone, kappa=.84 (95% CI .83 to .85, P = .598), and for doctors alone, kappa= .83 (95% CI .81 to .85, P = .566). The 1-way, 2-way ANOVA and combined ANOVA showed that the reliability coefficients (84%) for both nurses and physicians were similar to the kappa values. A combined ANOVA showed there was a. 2-point difference with physicians assigning a higher triage level. The high rate of interobserver agreement has important implications for case mix comparisons and suggests that this scale is understood and interpreted in a similar fashion by nurses and physicians.


Application of differential display, with in situ hybridization verification, to microscopic samples of breast cancer tissue.
The technique of differential display (DD) has been used widely to identify potentially interesting overexpressed or repressed genes in a variety of compared samples. When used in studying tissue samples, it inevitably confronts problems of limited amount of input material and cell-type heterogeneity. We report here the application of in situ hybridization as a method of confirmatory test for DD as well as definition of cell type expressing differential cDNA. This procedure employed material derived from a single case of human mammary, grade III, infiltrating ductal carcinoma, using free-hand microdissection, where we have compared gene expression profiles in invasive tumour with those in adjacent normal tissue. A total of 21 cDNAs were found to be differentially expressed between the two tissue types; 11 upregulated in the tumour sample and 10 upregulated in the normal sample. Six cDNAs were utilized as probes for in situ hybridization analysis of a further five cases of comparably staged breast cancer. One of these clones, 11AT1, which was found to be homologous to Hsc70, was shown to be overexpressed in tumour cells relative to adjacent normal stroma and to benign glandular epithelium in all five cases; an increase in expression was further confirmed at protein level by immunohistochemistry. The study demonstrated the applicability of in situ hybridization as a screening test in DD strategy for studying tissue material and a reasonable technique combination of identifying changes in gene expression associated with tumour development.


18F-FSPG PET/CT Imaging of System xC- Transporter Activity in Patients with Primary and Metastatic Brain Tumors.
Background The PET tracer (4S)-4-(3-[18F]fluoropropyl)-l-glutamate (18F-FSPG) targets the system xC- cotransporter, which is overexpressed in various tumors. Purpose To assess the role of 18F-FSPG PET/CT in intracranial malignancies. Materials and Methods Twenty-six patients (mean age, 54 years ± 12; 17 men; 48 total lesions) with primary brain tumors (n = 17) or brain metastases (n = 9) were enrolled in this prospective, single-center study (ClinicalTrials.gov identifier: NCT02370563) between November 2014 and March 2016. A 30-minute dynamic brain 18F-FSPG PET/CT scan and a static whole-body (WB) 18F-FSPG PET/CT scan at 60-75 minutes were acquired. Moreover, all participants underwent MRI, and four participants underwent fluorine 18 (18F) fluorodeoxyglucose (FDG) PET imaging. PET parameters and their relative changes were obtained for all lesions. Kinetic modeling was used to estimate the 18F-FSPG tumor rate constants using the dynamic and dynamic plus WB PET data. Imaging parameters were correlated to lesion outcomes, as determined with follow-up MRI and/or pathologic examination. The Mann-Whitney U test or Student t test was used for group mean comparisons. Receiver operating characteristic curve analysis was used for performance comparison of different decision measures. Results 18F-FSPG PET/CT helped identify all 48 brain lesions. The mean tumor-to-background ratio (TBR) on the whole-brain PET images at the WB time point was 26.6 ± 24.9 (range: 2.6-150.3). When 18F-FDG PET was performed, 18F-FSPG permitted visualization of non-18F-FDG-avid lesions or allowed better lesion differentiation from surrounding tissues. In participants with primary brain tumors, the predictive accuracy of the relative changes in influx rate constant Ki and maximum standardized uptake value to discriminate between poor and good lesion outcomes were 89% and 81%, respectively. There were significant differences in the 18F-FSPG uptake curves of lesions with good versus poor outcomes in the primary brain tumor group (P < .05) but not in the brain metastases group. Conclusion PET/CT imaging with (4S)-4-(3-[18F]fluoropropyl)-l-glutamate (18F-FSPG) helped detect primary brain tumors and brain metastases with a high tumor-to-background ratio. Relative changes in 18F-FSPG uptake with multi-time-point PET appear to be helpful in predicting lesion outcomes. Clinical trial registration no. NCT02370563 © RSNA, 2022 Online supplemental material is available for this article.