Proteomic characterization of macro-, micro- and nano-extracellular vesicles derived from the same first trimester placenta: relevance for feto-maternal communication.
What proteins are carried by extracellular vesicles (EVs) released from normal first trimester placentae? One thousand five hundred and eighty-five, 1656 and 1476 proteins were characterized in macro-, micro- and nano-vesicles, respectively, from first trimester placentae, with all EV fractions being enriched for proteins involved in vesicle transport and inflammation. Placental EVs are being increasingly recognized as important mediators of both healthy and pathological pregnancies. However, current research has focused on detecting changes in specific proteins in particular fractions of vesicles during disease. This is the first study to investigate the full proteome of different-sized fractions of EVs from the same first trimester placenta and highlights the differences/similarities between the vesicle fractions. A well-established ex vivo placental explant culture model was used to generate macro-, micro- and nano-vesicles from 56 first trimester placentae. Vesicle fractions were collected by differential ultracentrifugation, quantified and characterized. Placental macro-, micro- and nano-vesicles were characterized by microscopy, dynamic light scattering and nanoparticle tracking analysis. The proteome of each EV fraction was interrogated using liquid chromatography-coupled tandem mass spectrometry. Results were validated by semi-quantitative western blotting. A total of 1585, 1656 and 1476 proteins were identified in macro-, micro- and nano-vesicles, respectively. One thousand one hundred and twenty-five proteins were shared between all three fractions while up to 223 proteins were unique to each fraction. Gene Ontology pathway analysis showed an enrichment of proteins involved in vesicle transport and inflammation in all three fractions of EVs. The expression levels of proteins involved in internalization of vesicles (annexin V, calreticulin, CD31, CD47), the complement pathway [C3, decay-accelerating factor (DAF), membrane cofactor protein (MCP), protectin] and minor histocompatibility antigens [ATP-dependent RNA helicase (DDX3), ribosomal protein S4 (RPS4)] were different between different-sized EVs. This study is largely hypothesis-generating in nature. It is important to validate these findings using EVs isolated from maternal plasma and the function of the different EV fractions would need further investigation. Our results support the concept that various EV factions can interact with different maternal cells and have unique effects to mediate feto-maternal communication during early pregnancy. This study also provides a list of candidate proteins, which may inform the identification of robust markers that can be used to isolate placental vesicles from the maternal blood in the future. M.T. is a recipient of the University of Auckland Health Research Doctoral Scholarship and the Freemasons Postgraduate Scholarship. This project was supported by a School of Medicine Performance-based research fund (PBRF) grant awarded to L.W.C. No authors have any conflicts of interest to disclose.


D-amino acids govern stationary phase cell wall remodeling in bacteria.
In all known organisms, amino acids are predominantly thought to be synthesized and used as their L-enantiomers. Here, we found that bacteria produce diverse D-amino acids as well, which accumulate at millimolar concentrations in supernatants of stationary phase cultures. In Vibrio cholerae, a dedicated racemase produced D-Met and D-Leu, whereas Bacillus subtilis generated D-Tyr and D-Phe. These unusual D-amino acids appear to modulate synthesis of peptidoglycan, a strong and elastic polymer that serves as the stress-bearing component of the bacterial cell wall. D-Amino acids influenced peptidoglycan composition, amount, and strength, both by means of their incorporation into the polymer and by regulating enzymes that synthesize and modify it. Thus, synthesis of D-amino acids may be a common strategy for bacteria to adapt to changing environmental conditions.


Cell lineage and the differentiation of corneal epithelial cells.
Studies were designed to determine whether cell division and cell differentiation are linked directly in the corneal epithelium. To obtain these data, corneal basal epithelial cells were labeled during DNA synthesis, and the resultant daughter cells were followed for as long as 2 weeks. Adult rats were injected with 5-bromo-deoxyuridine (BrdU), a thymidine analog, and were killed 6 hours to 14 days later. Corneas were fixed and permeabilized, and BrdU-labeled nuclei were detected with a monoclonal antibody to BrdU and a fluorescent-labeled secondary antibody. Fluorescent nuclei were visualized in three dimensions in corneal whole-mounts using a laser scanning confocal microscope. Optical sections were collected and displayed as serial images, three-dimensional anaglyphs, color-encoded projections, or three-dimensional reconstructions. Data were confirmed using 3H-thymidine autoradiography of epithelia sectioned parallel to the corneal surface. Cells synthesizing DNA at the time of injection incorporated BrdU into their DNA. Pairs of labeled nuclei were produced by the division of cells that had been labeled with BrdU. These daughter cells remained in the basal layer of the epithelium for a variable period of time. Some daughter cells continued to divide, producing clusters of labeled basal cells. When labeled daughter cells left the basal layer and began the process of terminal differentiation, they nearly always did so together. The synchronous differentiation of daughter cells was evident from the pairs of labeled nuclei seen throughout the depth of the epithelium from 2 to 14 days after labeling. Cell division and differentiation are not linked directly in the corneal epithelium. After cell division, daughter cells either remain in the basal layer, where they may undergo additional rounds of cell division, or both cells differentiate synchronously. When the daughter cells of a mitosis differentiate, the time between the previous cell division and differentiation is highly variable. This suggests that the coordination of cell division and differentiation in the corneal epithelium involves a complex regulatory network.


Beta blockers for elective surgery in elderly patients: population based, retrospective cohort study.
To test whether atenolol (a long acting beta blocker) and metoprolol (a short acting beta blocker) are associated with equivalent reductions in risk for elderly patients undergoing elective surgery. Population based, retrospective cohort analysis. Acute care hospitals in Ontario, Canada, over one decade. Consecutive patients older than 65 who were admitted for elective surgery, without symptomatic coronary disease. Death or myocardial infarction. 37,151 patients were receiving atenolol or metoprolol before surgery, of which the most common operations were orthopaedic or abdominal procedures. As expected, the two groups were similar in demographic characteristics, medical therapy, and type of surgery. 1038 patients experienced a myocardial infarction or died, a rate that was significantly lower for patients receiving atenolol than for those receiving metoprolol (2.5% v 3.2%, P < 0.001). The decreased risk with atenolol persisted after adjustment for measured demographic, medical, and surgical factors; extended to comparisons of other long acting and short acting beta blockers; was accentuated in analyses that focused on patients with the clearest evidence of beta blocker treatment; and reflected the immediate postoperative interval. Patients receiving metoprolol do not have as low a perioperative cardiac risk as patients receiving atenolol, in accord with possible acute withdrawal after missed doses.


HIV infection and arterial inflammation assessed by (18)F-fluorodeoxyglucose (FDG) positron emission tomography (PET): a prospective cross-sectional study.
HIV-infected patients are at increased risk of myocardial infarction and arterial inflammation has been suggested as a pathophysiological explanation. We compared the uptake of (18)F-fluorodeoxyglucose (FDG) by PET in four arterial regions, and factors associated with FDG uptake in well-treated HIV-infected patients without cardiovascular disease (CVD) and healthy controls. We prospectively scanned 26 HIV-infected patients on stable antiretroviral therapy and 25 healthy volunteers with FDG PET/CT, measuring standardized uptake values (SUV) in the carotid arteries, the ascending, descending, and abdominal aorta. We performed correlation analyses between FDG uptake and intima-media thickness (IMT), and soluble biomarkers of inflammation. We found no difference in arterial FDG uptake between the HIV-infected patients and healthy controls quantified either as mean SUVmax or target-to background ratio in the carotid region, the ascending aorta, the descending aorta, or the abdominal aorta. Correlations between SUV, IMT, and soluble biomarkers were scarce in both groups. In a group of optimally treated HIV-infected patients with full viral suppression, low Framingham risk score and no known CVD, we found no evidence of increased arterial inflammation as assessed by FDG PET/CT compared to healthy volunteers.


Clinical outcomes of patients with critical limb ischemia who undergo routine coronary angiography and subsequent percutaneous coronary intervention.
Critical limb ischemia (CLI) is associated with a high risk of cardiovascular ischemic events. We assessed the strategy of routine coronary angiography and subsequent coronary revascularization, if clinically indicated, in patients with CLI who underwent percutaneous transluminal angioplasty (PTA). Of a total 286 consecutive CLI patients treated by PTA, 252 patients who underwent coronary angiography before or after PTA were enrolled. Coronary artery disease (CAD) was defined as angiographic stenosis ≥50% and significant CAD as ≥70% stenosis. Of the 252 patients with CLI who underwent coronary angiography, a total of 167 patients (66.3%) had CAD and 85 patients (33.7%) did not have CAD. Patients in the CAD group were older, had a higher prevalence of diabetes and cerebrovascular disease, and had a lower mean ejection fraction. In the CAD group, of the 145 patients with significant CAD, percutaneous coronary intervention (PCI) was performed in 114 patients (78.6%). At 1 year, the CAD and non-CAD groups had no statistically significant differences in mortality (7.1% vs 4.7%; P=.45), myocardial infarction (1.1% vs 0%; P=.31), and PCI (4.7% vs 1.1%; P=.31). These outcomes were similar after the adjustment of baseline confounders. At 1 year, the CAD and non-CAD groups had similar rates of repeat PTA (16.7% vs 17.6%; P=.86), target lesion revascularization (13.7% vs 14.1%; P=.94), and amputation (19.1% vs 16.4%; P=.60). A strategy of routine coronary angiography and coronary revascularization may be a reasonable treatment option for these patients who have high risk for severe CAD. A randomized trial is needed to determine if this is the preferred strategy for CLI patients.


Cellular distribution of insulin-degrading enzyme gene expression. Comparison with insulin and insulin-like growth factor receptors.
Insulin-degrading enzyme (IDE) hydrolyzes both insulin and IGFs and has been proposed to play a role in signal termination after binding of these peptides to their receptors. In situ hybridization was used to investigate the cellular distribution of IDE mRNA and to compare it with insulin receptor (IR) and IGF-I receptor (IGFR) gene expression in serial thin sections from a variety of tissues in embryonic and adult rats. IDE mRNA is highly abundant in kidney and liver, tissues known to play a role in insulin degradation. IDE and IR mRNAs are highly coexpressed in brown fat and liver. The highest level IDE gene expression, on a per cell basis, is found in germinal epithelium. IDE and IGFR mRNAs are colocalized in oocytes, while IDE is colocalized with the IGF-II receptor in spermatocytes, suggesting that IDE may be involved with degradation of IGF-II in the testis. In summary, IDE expression demonstrates significant anatomical correlation with insulin/IGF receptors. These data are compatible with a role for IDE in degrading insulin and IGFs after they bind to and are internalized with their respective receptors and may also suggest a novel role for IDE in germ cells.


Colonization of dodder, Cuscuta indecora, by 'Candidatus Liberibacter asiaticus' and 'Ca. L. americanus'.
Huanglongbing, or citrus greening, threatens the global citrus industry. The presumptive pathogens, 'Candidatus Liberibacter asiaticus' and 'Ca. L. americanus' can be transferred from citrus to more easily studied experimental hosts by using holoparasitic dodder plants. However, the interaction between 'Candidatus Liberibacter' spp. and the dodder has not been studied. We combined quantitative polymerase chain reaction with electron microscopy to show that only 65% of tendrils of Cuscuta indecora grown on 'Ca. Liberibacter' spp.-infected host plants had detectable levels of the pathogen. Among tendrils that were colonized by Liberibacter in at least one 2 cm segment, most were not colonized in all segments. Furthermore, the estimated population levels of the pathogen present in serial 2 cm segments of dodder tendrils varied widely and without any consistent pattern. Thus, there was generally not a concentration gradient of the pathogen from the source plant towards the recipient and populations of the pathogen were sometimes found in the distal segments of the dodder plant but not in the proximal or middle segments. Populations of the pathogens ranged from 2 x 10(2) to 3.0 x 10(8) cells per 2 cm segment. On a fresh weight basis, populations as high as 1.4 x 10(10) cells per g of tissue were observed demonstrating that 'Ca. Liberibacter' spp. multiplies well in Cuscuta indecora. However, 55% of individual stem segments did not contain detectable levels of the pathogen, consistent with a pattern of nonuniform colonization similar to that observed in the much more anatomically complex citrus tree. Colonization of dodder by the pathogen is also nonuniform at the ultrastructural level, with adjacent phloem vessel elements being completely full of the pathogen or free of the pathogen. We also observed bacteria in the phloem vessels that belonged to two distinct size classes based on the diameters of cross sections of cells. In other sections from the same tendrils we observed single bacterial cells that were apparently in the process of differentiating between the large and round forms to the long and thin forms (or vice versa). The process controlling this morphological differentiation of the pathogen is not known. The highly reduced and simplified anatomy of the dodder plant as well as its rapid growth rate compared with citrus, and the ability of the plant to support multiplication of the pathogen to high levels, makes it an interesting host plant for further studies of host-pathogen interactions.


A surprising cross-species conservation in the genomic landscape of mouse and human oral cancer identifies a transcriptional signature predicting metastatic disease.
Improved understanding of the molecular basis underlying oral squamous cell carcinoma (OSCC) aggressive growth has significant clinical implications. Herein, cross-species genomic comparison of carcinogen-induced murine and human OSCCs with indolent or metastatic growth yielded results with surprising translational relevance. Murine OSCC cell lines were subjected to next-generation sequencing (NGS) to define their mutational landscape, to define novel candidate cancer genes, and to assess for parallels with known drivers in human OSCC. Expression arrays identified a mouse metastasis signature, and we assessed its representation in four independent human datasets comprising 324 patients using weighted voting and gene set enrichment analysis. Kaplan-Meier analysis and multivariate Cox proportional hazards modeling were used to stratify outcomes. A quantitative real-time PCR assay based on the mouse signature coupled to a machine-learning algorithm was developed and used to stratify an independent set of 31 patients with respect to metastatic lymphadenopathy. NGS revealed conservation of human driver pathway mutations in mouse OSCC, including in Trp53, mitogen-activated protein kinase, phosphoinositide 3-kinase, NOTCH, JAK/STAT, and Fat1-4. Moreover, comparative analysis between The Cancer Genome Atlas and mouse samples defined AKAP9, MED12L, and MYH6 as novel putative cancer genes. Expression analysis identified a transcriptional signature predicting aggressiveness and clinical outcomes, which were validated in four independent human OSCC datasets. Finally, we harnessed the translational potential of this signature by creating a clinically feasible assay that stratified patients with OSCC with a 93.5% accuracy. These data demonstrate surprising cross-species genomic conservation that has translational relevance for human oral squamous cell cancer. Clin Cancer Res; 20(11); 2873-84. ©2014 AACR.


Associations of gender and etiology with outcomes in heart failure with systolic dysfunction: a pooled analysis of 5 randomized control trials.
This study sought to explore the gender-related differences in etiology and outcomes in chronic heart failure (HF) patients from 5 randomized trials. Each year, 550,000 new cases of HF are identified; however, there remain limited data on gender-related differences in etiology and outcomes among patients with HF with systolic dysfunction. We analyzed data from 8,791 men and 2,851 women randomized in 5 clinical trials (PRAISE [Prospective Randomized Amlodipine Survival Evaluation], PRAISE-2, MERIT-HF [Metoprolol Extended Release Randomized Intervention Trial in Heart Failure], VEST [Vesnarinone Trial], and PROMISE [Prospective Randomized Milrinone Survival Evaluation]) to explore gender-related differences in etiology (ischemic vs. nonischemic) and outcomes (all-cause mortality and death or all-cause hospitalization). Hazard ratios (HR), 95% confidence intervals (CIs), and Kaplan-Meier survival curves were generated by gender and etiology. A total of 18% of ischemic and 31% of nonischemic patients were women. Irrespective of etiology, women were older, more ethnically diverse, and had higher systolic blood pressures, more diabetes, and severe HF symptoms, but less often smoked or had prior myocardial infarctions than men. Mean ejection fractions were similar between women (23.6%) and men (23.2%). The 1-year Kaplan-Meier survival estimates varied by gender and etiology (female nonischemics, HR 0.88 [95% CI 0.85 to 0.89]; female ischemics, HR 0.83 [95% CI 0.81 to 0.85]; male nonischemics, HR 0.84 [95% CI 0.83 to 0.85]; male ischemics, HR 0.79 [95% CI 0.78 to 0.81]). After adjustment, female gender (HR 0.77 [95% CI 0.69 to 0.85]) and nonischemic etiology (HR 0.80 [95% CI 0.72 to 0.89]) were associated with longer survival time. Time to death or hospitalization was longer among nonischemics (HR 0.83 [95% CI 0.78 to 0.89], p < 0.0001); however, female gender was not significantly associated with the composite outcome (HR 1.01 [95% CI 0.95 to 1.08]). Our data clarify that outcomes differ by both gender and etiology among patients with HF with systolic dysfunction. Understanding these differences may lead to better management of HF patients and improved overall prognosis.


Obesogenic television food advertising to children in Malaysia: sociocultural variations.
Food advertising on television (TV) is well known to influence children's purchasing requests and models negative food habits in Western countries. Advertising of unhealthy foods is a contributor to the obesogenic environment that is a key driver of rising rates of childhood obesity. Children in developing countries are more at risk of being targeted by such advertising, as there is a huge potential for market growth of unhealthy foods concomitant with poor regulatory infrastructure. Further, in developing countries with multi-ethnic societies, information is scarce on the nature of TV advertising targeting children. To measure exposure and power of TV food marketing to children on popular multi-ethnic TV stations in Malaysia. Ethnic-specific popular TV channels were identified using industry data. TV transmissions were recorded for each channel from November 2012 to August 2013 (16 hr/day) for randomly selected weekdays and weekend days during normal days and repeated during school holidays (n=88 days). Coded food/beverage advertisements were grouped into core (healthy), non-core (non-healthy), or miscellaneous (unclassified) food categories. Peak viewing time (PVT) and persuasive marketing techniques were identified. Non-core foods were predominant in TV food advertising, and rates were greater during school holidays compared to normal days (3.51 vs 1.93 food ads/hr/channel, p<0.001). During normal days' PVT, the ratio of non-core to core food advertising was higher (3.25 food ads/hr/channel), and this more than trebled during school holidays to 10.25 food ads/hr/channel. Popular channels for Indian children had the lowest rate of food advertising relative to other ethnic groups. However, sugary drinks remained a popular non-core product advertised across all broadcast periods and channels. Notably, promotional characters doubled for non-core foods during school holidays compared to normal days (1.91 vs 0.93 food ads/hr/channel, p<0.001). This study highlights non-core food advertising, and predominantly sugary drinks are commonly screened on Malaysian TV channels. The majority of these sugary drinks were advertised by multinational companies, and this observation warrants regulatory attention.


Human adenosine deaminases ADA1 and ADA2 bind to different subsets of immune cells.
At sites of inflammation and tumor growth, the local concentration of extracellular adenosine rapidly increases and plays a role in controlling the immune responses of nearby cells. Adenosine deaminases ADA1 and ADA2 (ADAs) decrease the level of adenosine by converting it to inosine, which serves as a negative feedback mechanism. Mutations in the genes encoding ADAs lead to impaired immune function, which suggests a crucial role for ADAs in immune system regulation. It is not clear why humans and other mammals possess two enzymes with adenosine deaminase activity. Here, we found that ADA2 binds to neutrophils, monocytes, NK cells and B cells that do not express CD26, a receptor for ADA1. Moreover, the analysis of CD4+ T-cell subset revealed that ADA2 specifically binds to regulatory T cells expressing CD39 and lacking the receptor for ADA1. Also, it was found that ADA1 binds to CD16- monocytes, while CD16+ monocytes preferably bind ADA2. A study of the blood samples from ADA2-deficient patients showed a dramatic reduction in the number of lymphocyte subsets and an increased concentration of TNF-α in plasma. Our results suggest the existence of a new mechanism, where the activation and survival of immune cells is regulated through the activities of ADA2 or ADA1 anchored to the cell surface.


Informative priors based on transcription factor structural class improve de novo motif discovery.
An important problem in molecular biology is to identify the locations at which a transcription factor (TF) binds to DNA, given a set of DNA sequences believed to be bound by that TF. In previous work, we showed that information in the DNA sequence of a binding site is sufficient to predict the structural class of the TF that binds it. In particular, this suggests that we can predict which locations in any DNA sequence are more likely to be bound by certain classes of TFs than others. Here, we argue that traditional methods for de novo motif finding can be significantly improved by adopting an informative prior probability that a TF binding site occurs at each sequence location. To demonstrate the utility of such an approach, we present priority, a powerful new de novo motif finding algorithm. Using data from TRANSFAC, we train three classifiers to recognize binding sites of basic leucine zipper, forkhead, and basic helix loop helix TFs. These classifiers are used to equip priority with three class-specific priors, in addition to a default prior to handle TFs of other classes. We apply priority and a number of popular motif finding programs to sets of yeast intergenic regions that are reported by ChIP-chip to be bound by particular TFs. priority identifies motifs the other methods fail to identify, and correctly predicts the structural class of the TF recognizing the identified binding sites. Supplementary material and code can be found at http://www.cs.duke.edu/~amink/.


A randomized clinical trial of theory-based activities for the behavioral symptoms of dementia in nursing home residents.
To test the main and interactive effects of activities derived from the Need-Driven Dementia-Compromised Behavior model for responding to behavioral symptoms in nursing home residents. Randomized double-blind clinical trial. Nine community-based nursing homes. One hundred twenty-eight cognitively impaired residents randomly assigned to activities adjusted to functional level (FL) (n=32), personality style of interest (PSI) (n=33), functional level and personality style of interest (FL+PSI) (n=31), or active control (AC) (n=32). Three weeks of activities provided twice daily. Agitation, passivity, engagement, affect, and mood assessed from video recordings and real-time observations during baseline, intervention, random times outside of intervention, and 1 week after intervention. All treatments improved outcomes during intervention except mood, which worsened under AC. During intervention the PSI group demonstrated greater engagement, alertness, and attention than the other groups; the FL+PSI group demonstrated greater pleasure. During random times, engagement returned to baseline levels except in the FL group in which it decreased. There was also less agitation and passivity in groups with a component adjusted to PSI. One week after the intervention, mood, anxiety, and passivity improved over baseline; significantly less pleasure was displayed after withdrawal of treatment. The hypothesis that activities adjusted to FL+PSI would improve behavioral outcomes to a greater extent than partially adjusted or nonadjusted activities was partially supported. PSI is a critical component of individualized activity prescription.


A new metric for measuring condition in large predatory sharks.
A simple metric (span condition analysis; SCA) is presented for quantifying the condition of sharks based on four measurements of body girth relative to body length. Data on 104 live sharks from four species that vary in body form, behaviour and habitat use (Carcharhinus leucas, Carcharhinus limbatus, Ginglymostoma cirratum and Galeocerdo cuvier) are given. Condition shows similar levels of variability among individuals within each species. Carcharhinus leucas showed a positive relationship between condition and body size, whereas the other three species showed no relationship. There was little evidence for strong differences in condition between males and females, although more male sharks are needed for some species (e.g. G. cuvier) to verify this finding. SCA is potentially viable for other large marine or terrestrial animals that are captured live and then released.


In search of causal pathways in diabetes: a study using proteomics and genotyping data from a cross-sectional study.
The pathogenesis of type 2 diabetes is not fully understood. We investigated whether circulating levels of preselected proteins were associated with the outcome 'diabetes' and whether these associations were causal. In 2467 individuals of the population-based, cross-sectional EpiHealth study (45-75 years, 50% women), 249 plasma proteins were analysed by the proximity extension assay technique. DNA was genotyped using the Illumina HumanCoreExome-12 v1.0 BeadChip. Diabetes was defined as taking glucose-lowering treatment or having a fasting plasma glucose of ≥7.0 mmol/l. The associations between proteins and diabetes were assessed using logistic regression. To investigate causal relationships between proteins and diabetes, a bidirectional two-sample Mendelian randomisation was performed based on large, genome-wide association studies belonging to the DIAGRAM and MAGIC consortia, and a genome-wide association study in the EpiHealth study. Twenty-six proteins were positively associated with diabetes, including cathepsin D, retinal dehydrogenase 1, α-L-iduronidase, hydroxyacid oxidase 1 and galectin-4 (top five findings). Three proteins, lipoprotein lipase, IGF-binding protein 2 and paraoxonase 3 (PON-3), were inversely associated with diabetes. Fourteen of the proteins are novel discoveries. The Mendelian randomisation study did not disclose any significant causal effects between the proteins and diabetes in either direction that were consistent with the relationships found between the protein levels and diabetes. The 29 proteins associated with diabetes are involved in several physiological pathways, but given the power of the study no causal link was identified for those proteins tested in Mendelian randomisation. Therefore, the identified proteins are likely to be biomarkers for type 2 diabetes, rather than representing causal pathways.


Disentangling data discrepancies with integrated population models.
A common challenge for studying wildlife populations occurs when different survey methods provide inconsistent or incomplete inference on the trend, dynamics, or viability of a population. A potential solution to the challenge of conflicting or piecemeal data relies on the integration of multiple data types into a unified modeling framework, such as integrated population models (IPMs). IPMs are a powerful approach for species that inhabit spatially and seasonally complex environments. We provide guidance on exploiting the capabilities of IPMs to address inferential discrepancies that stem from spatiotemporal data mismatches. We illustrate this issue with analysis of a migratory species, the American Woodcock (Scolopax minor), in which individual monitoring programs suggest differing population trends. To address this discrepancy, we synthesized several long-term data sets (1963-2015) within an IPM to estimate continental-scale population trends, and link dynamic drivers across the full annual cycle and complete extent of the woodcock's geographic range in eastern North America. Our analysis reveals the limiting portions of the life cycle by identifying time periods and regions where vital rates are lowest and most variable, as well as which demographic parameters constitute the main drivers of population change. We conclude by providing recommendations for resolving conflicting population estimates within an integrated modeling approach, and discuss how strategies (e.g., data thinning, expert opinion elicitation) from other disciplines could be incorporated into ecological analyses when attempting to combine multiple, incongruent data types.


ESCMID* guideline for the diagnosis and management of Candida diseases 2012: prevention and management of invasive infections in neonates and children caused by Candida spp.
Invasive candidiasis (IC) is a relatively common syndrome in neonates and children and is associated with significant morbidity and mortality. These guidelines provide recommendations for the prevention and treatment of IC in neonates and children. Appropriate agents for the prevention of IC in neonates at high risk include fluconazole (A-I), nystatin (B-II) or lactoferrin ± Lactobacillus (B-II). The treatment of IC in neonates is complicated by the high likelihood of disseminated disease, including the possibility of infection within the central nervous system. Amphotericin B deoxycholate (B-II), liposomal amphotericin B (B-II), amphotericin B lipid complex (ABLC) (C-II), fluconazole (B-II), micafungin (B-II) and caspofungin (C-II) can all be potentially used. Recommendations for the prevention of IC in children are largely extrapolated from studies performed in adults with concomitant pharmacokinetic data and models in children. For allogeneic HSCT recipients, fluconazole (A-I), voriconazole (A-I), micafungin (A-I), itraconazole (B-II) and posaconazole (B-II) can all be used. Similar recommendations are made for the prevention of IC in children in other risk groups. With several exceptions, recommendations for the treatment of IC in children are extrapolated from adult studies, with concomitant pharmacokinetic studies. Amphotericin B deoxycholate (C-I), liposomal amphotericin B (A-I), ABLC (B-II), micafungin (A-I), caspofungin (A-I), anidulafungin (B-II), fluconazole (B-I) and voriconazole (B-I) can all be used.


One-step synthesis, electronic structure, and photocatalytic activity of earth-abundant visible-light-driven FeAl2O4.
The development of inexpensive visible-light-driven photocatalysts is an important prerequisite for realizing the industrial application of photocatalysis technology. In this paper, an earth-abundant FeAl2O4 photocatalyst is prepared via facile solution combustion synthesis. Density functional theory and the scanning Kelvin probe technique are employed to ascertain the positions of the energy bands and the Fermi level. Phenol is taken as a model pollutant to evaluate the photocatalytic activity of FeAl2O4. The scavenger experiment results, ˙OH-trapping fluorescence technique, and electron spin resonance measurements confirm that the superoxide anion radical is the main active species generated in the photocatalytic process, which also further corroborates the proposed electronic structure of FeAl2O4. The degradation experiments and O2 temperature programmed desorption results over various samples verify that the crystallinity degree is a more important factor than the oxygen adsorption ability in determining photocatalytic activity.


Screening for colorectal cancer: reasons for refusal of faecal occult blood testing in a general practice in England.
To ascertain reasons for non-compliance with faecal occult blood tests in colorectal cancer screening programmes. A standard interview by a trained nurse of a random sample of those who declined screening. The Leicestershire town of Market Harborough, where most of the 25,000 population are served by a single general practice of 10 partners. Altogether 4185 residents aged 51 to 70 years were invited to receive a free faecal occult blood test (Haemoccult). Eighty one subjects from a sample of 351 who wrote declining the offer were interviewed. Non-compliers were divided into those who did not request a test kit and those who returned an unused kit. In the former group the commonest reasons given were intercurrent illness (39%), fear of further tests and surgery (24%), and feeling well (22%). For those who returned unused kits the commonest reasons were the unpleasantness of the stool collection procedure (65%), feeling well (30%), intercurrent illness (23%), and fear of further tests and surgery (20%). In both groups the main concern of those who did not comply were fear of further diagnostic tests and surgery rather than concern at the lack of effective treatment for cancer. To increase compliance, education and publicity must explain the concept of asymptomatic illness and allay people's fear of hospital investigation and treatment. The benefits of screening should be particularly emphasised to those who return kits so they may overcome their reservations.


Radiation therapy for diabetes insipidus caused by Langerhans cell histiocytosis.
Hypothalamic-pituitary radiation therapy has been the standard treatment for the diabetes insipidus of Langerhans cell histiocytosis. The goal of this study was to assess the role of radiation therapy in Langerhans cell histiocytosis-associated diabetes insipidus and to compare the results with nonirradiated controls. Forty-seven patients with pathologically confirmed Langerhans cell histiocytosis were diagnosed with diabetes insipidus between 1950 and 1989 and were treated at the Mayo Clinic. These patients were divided into two groups on the basis of treatment for the diabetes insipidus: The first group (radiation group) included 30 patients (28 of whom were evaluable for response) who received hypothalamic-pituitary radiation therapy, and the second group (control group) included 17 patients who did not. A partial response to treatment was defined as a reduction in vasopressin dosage or improvement in computed tomography (CT) or magnetic resonance imaging (MRI). A complete response was defined as no further need for vasopressin therapy or normalization of CT or MRI. End points analyzed included treatment response, patient characteristics, morbidity, dose-response relationship, and survival. Patient characteristics of the two groups were similar except for age and lung involvement, both of which were significantly less in the radiation group. Thirty-six percent of patients (10 of 28) in the radiation group responded to hypothalamic-pituitary radiation therapy (22% complete response and 14% partial response), whereas none in the control group responded. Five of the six complete responders were irradiated within 14 days of the diagnosis of diabetes insipidus. The mean dose used in the responding and nonresponding patients was 11.2 and 10 Gy, respectively. Three of five patients (60%) treated with more than 15 Gy responded compared to seven of 23 (30%) treated with less than 15 Gy. Eight of the 10 responders (80%), compared to 16 of 35 nonresponders (46%), were female. Only one in 20 patients with concomitant lung histiocytosis responded. Complications of therapy may include insufficiency in other hypothalamic-pituitary axes in the treated patients. Actuarial survivals at 5, 10, 20, and 40 years for the entire group were 80%, 78%, 75%, and 65%, respectively, with a median follow-up in living patients of 14.7 years.


GPs' role in caring for children and young people with life-limiting conditions: a retrospective cohort study.
GPs are rarely actively involved in healthcare provision for children and young people (CYP) with life-limiting conditions (LLCs). This raises problems when these children develop minor illness or require management of other chronic diseases. To investigate the association between GP attendance patterns and hospital urgent and emergency care use. Retrospective cohort study using a primary care data source (Clinical Practice Research Datalink) in England. The cohort numbered 19 888. CYP aged 0-25 years with an LLC were identified using Read codes (primary care) or International Classification of Diseases 10 th Revision (ICD-10) codes (secondary care). Emergency inpatient admissions and accident and emergency (A&E) attendances were separately analysed using multivariable, two-level random intercept negative binomial models with key variables of consistency and regularity of GP attendances. Face-to-face GP surgery consultations reduced, from a mean of 7.12 per person year in 2000 to 4.43 in 2015. Those consulting the GP less regularly had 15% (95% confidence interval [CI] = 10% to 20%) more emergency admissions and 5% more A&E visits (95% CI = 1% to 10%) than those with more regular consultations. CYP who had greater consistency of GP seen had 10% (95% CI = 6% to 14%) fewer A&E attendances but no significant difference in emergency inpatient admissions than those with lower consistency. There is an association between GP attendance patterns and use of urgent secondary care for CYP with LLCs, with less regular GP attendance associated with higher urgent secondary healthcare use. This is an important area for further investigation and warrants the attention of policymakers and GPs, as the number of CYP with LLCs living in the community rises.


Effects of ferric citrate supplementation on advanced glycation end products in a rat model of streptozotocin/nicotinamide-induced diabetes.
Diabetes is associated with the increased risks of anemia and activation of advanced glycation end products (AGEs) and the receptor for AGEs (RAGE). However, the effects of pharmacological doses of iron supplementation on AGE metabolism are less clear. The aim was to investigate the effect of ferric citrate supplementation on AGE metabolism. Diabetes was induced in overnight starved rats by intraperitoneal injections of 40 mg/kg streptozotocin and 120 mg/kg nicotinamide. Diabetic rats were fed a standard diet or pharmacological doses of ferric citrate (0.5, 1, 2, and 3 g of ferric iron/kg diet) for 10 weeks. Ferric citrate supplementation showed a dose-related effect on the hepatic steatosis score, malondialdehyde, cathepsin D, and glyoxalase I. A Western blot analysis revealed that >1 g of ferric iron suppressed hepatic AGE receptor 1 and high-mobility group-box 1 expressions but increased heme oxygenase-1 and RAGE expressions. Further analysis showed that high doses of ferric iron triggered sterol regulatory element-binding protein 1c, p38-mitogen-activated protein kinase, and nuclear factor-κB protein expressions. Overall, the present results suggest a dose-related effect of ferric citrate supplementation on AGE metabolism, and this effect was more evident at high iron doses (>1 g of ferric iron/kg diet).


[A case of malignant paraganglioma presenting with skull metastases].
A non-functioning paraganglioma is usually benign, however, it may cause distant metastases. There is no histological appearance for the diagnosis of malignancy or absolute criteria for predicting malignant potential. Bony metastases from paraganglioma are known to occur, but, skull metastases are very rare. We report a case of intracranial metastases from a renal paraganglioma. A 61-year-old male presented with temporal headache and exophthalmos on the left side. Seven years prior, he underwent surgery to remove a mass in the right renal hilum, which was diagnosed as renal cell carcinoma at that time. Computed tomography and magnetic resonance imaging showed a ring-like enhanced mass in the left middle fossa, which destroyed the sphenoid bone and the lateral wall of the orbit. Another osteolytic lesion was revealed in the occipital bone. The fragile tumor was totally resected. Histopathological study revealed the Zellballen pattern with extensive coagulation necrosis. No apparent nuclear atypia or mitosis were present. Immunohistochemistry showed reactivity for synaptophysin and chromogranin A in the tumor cells. Review of the surgical specimen of the previously resected renal tumor revealed the same pathological and immunohistochemical findings as those of chief cells in the middle fossa tumor. Thus, this tumor was diagnosed as a malignant paraganglioma metastasized from renal paraganglioma. After six cycle chemotherapy with cyclophosphamide and vincristine, his condition was stable for two years, however, he died four years after the diagnosis of malignancy.


Mortality among Japanese Zen priests.
A cohort study was done on 1396 deaths seen among 4352 Japanese male Zen priests during a follow up period from 1 January 1955 to 31 December 1978. Standardised mortality ratios were computed for major causes of death by comparing with the counterparts of the general Japanese male population. The SMR for all causes of death was 0.82 (p less than 0.001) and the SMR values for cerebrovascular diseases, pneumonia and bronchitis, peptic ulcer, liver cirrhosis, cancer of the respiratory organs, and cancer of the lung were all significantly smaller than unity. Taking regional mortality differences into account, a similar computation was made dividing the cohort into two subcohorts--that is, the priests living in eastern Japan and those in western Japan. Both subcohorts showed a highly significantly smaller SMR than unity for all causes of death. With the exception of only a few causes of death for which the observed number of deaths was small, they also showed such reduced SMRs for nearly all of the causes of death tested. A questionnaire survey on the current life style of active priests showed that they smoke less, eat less, meat and fish as they follow the more traditional Japanese dietary habits, and live in less polluted areas, but their drinking habits do not differ much from that of the average Japanese adult man. Possible reasons for their reduced mortality are discussed.


Stress increases the risk of type 2 diabetes onset in women: A 12-year longitudinal study using causal modelling.
Type 2 diabetes is associated with significant morbidity and mortality. Modifiable risk factors have been found to contribute up to 60% of type 2 diabetes risk. However, type 2 diabetes continues to rise despite implementation of interventions based on traditional risk factors. There is a clear need to identify additional risk factors for chronic disease prevention. The aim of this study was to examine the relationship between perceived stress and type 2 diabetes onset, and partition the estimates into direct and indirect effects. Women born in 1946-1951 (n = 12,844) completed surveys for the Australian Longitudinal Study on Women's Health in 1998, 2001, 2004, 2007 and 2010. The total causal effect was estimated using logistic regression and marginal structural modelling. Controlled direct effects were estimated through conditioning in the regression model. A graded association was found between perceived stress and all mediators in the multivariate time lag analyses. A significant association was found between hypertension, as well as physical activity and body mass index, and diabetes, but not smoking or diet quality. Moderate/high stress levels were associated with a 2.3-fold increase in the odds of diabetes three years later, for the total estimated effect. Results were only slightly attenuated when the direct and indirect effects of perceived stress on diabetes were partitioned, with the mediators only explaining 10-20% of the excess variation in diabetes. Perceived stress is a strong risk factor for type 2 diabetes. The majority of the effect estimate of stress on diabetes risk is not mediated by the traditional risk factors of hypertension, physical activity, smoking, diet quality, and body mass index. This gives a new pathway for diabetes prevention trials and clinical practice.


Ribosomal history reveals origins of modern protein synthesis.
The origin and evolution of the ribosome is central to our understanding of the cellular world. Most hypotheses posit that the ribosome originated in the peptidyl transferase center of the large ribosomal subunit. However, these proposals do not link protein synthesis to RNA recognition and do not use a phylogenetic comparative framework to study ribosomal evolution. Here we infer evolution of the structural components of the ribosome. Phylogenetic methods widely used in morphometrics are applied directly to RNA structures of thousands of molecules and to a census of protein structures in hundreds of genomes. We find that components of the small subunit involved in ribosomal processivity evolved earlier than the catalytic peptidyl transferase center responsible for protein synthesis. Remarkably, subunit RNA and proteins coevolved, starting with interactions between the oldest proteins (S12 and S17) and the oldest substructure (the ribosomal ratchet) in the small subunit and ending with the rise of a modern multi-subunit ribosome. Ancestral ribonucleoprotein components show similarities to in vitro evolved RNA replicase ribozymes and protein structures in extant replication machinery. Our study therefore provides important clues about the chicken-or-egg dilemma associated with the central dogma of molecular biology by showing that ribosomal history is driven by the gradual structural accretion of protein and RNA structures. Most importantly, results suggest that functionally important and conserved regions of the ribosome were recruited and could be relics of an ancient ribonucleoprotein world.


The post-operative analgesic efficacy of celecoxib compared with placebo and parecoxib after total hip or knee arthroplasty.
Nonsteroidal antiinflammatory drugs (NSAIDs) in combination with opioids is a model of multimodal analgesia. NSAIDs have the oral and parenteral forms. The aim of the present study was to evaluate the efficacy of celecoxib compared with placebo and parecoxib after total hip or knee arthroplasty. A total of 120, ASA 1-2, aged 18-75 years, patients were randomly assigned to receive one of the three groups: Group I (control) received placebo (n=40), group II received 400 mg celecoxib orally (n=40) and group III received 40 mg parecoxib intravenously (n=40). The present study medication was administered I hour before surgery. All patients had access to patient-controlled analgesia (PCA) with intravenous morphine. Patients were studied at 0, 1, 6, 12 and 24 hours postoperatively for verbal numerical rating scale (VNRS), morphine consumption, satisfaction score and side effects. The intraoperatively fentayl requirement were similar among the three groups (p < 0.00). Celecoxib and parecoxib significantly decreased the amount of morphine requirement after total hip or knee arthroplasty compared to placebo at 1, 6, 12 and 24 hours (p < 0.00). The celecoxib group required more morphine than the parecoxib group at 1, 6, 12 and 24 hours (p < 0.00). The VNRS score in parecoxib group was significantly lower than the celecoxib and control groups at 1, 6, 12 but not at 24 hours. The VNRS score was lower in the celecoxib group compared to the control group at I and 6 hours postoperatively (p = 0.01, p < 0. 01 respectively). The placebo group had a higher sedation score (p = 0.008) but not for nausea vomiting (p = 0.36) and pruritus (p = 0.12) compared to the treatment groups. Within 12 hours after total hip and knee arthroplasty, pre-operative administration of parenteral parecoxib 40 mg was more effective than oral celecoxib 400 mg and placebo in terms of morphine consumption and VNRS score.


Body mapping of cutaneous wetness perception across the human torso during thermo-neutral and warm environmental exposures.
Sensing skin wetness is linked to inputs arising from cutaneous cold-sensitive afferents. As thermosensitivity to cold varies significantly across the torso, we investigated whether similar regional differences in wetness perception exist. We also investigated the regional differences in thermal pleasantness and whether these sensory patterns are influenced by ambient temperature. Sixteen males (20 ± 2 yr) underwent a quantitative sensory test under thermo-neutral [air temperature (Tair) = 22°C; relative humidity (RH) = 50%] and warm conditions (Tair = 33°C; RH = 50%). Twelve regions of the torso were stimulated with a dry thermal probe (25 cm(2)) with a temperature of 15°C below local skin temperature (Tsk). Variations in Tsk, thermal, wetness, and pleasantness sensations were recorded. As a result of the same cold-dry stimulus, the skin-cooling response varied significantly by location (P = 0.003). The lateral chest showed the greatest cooling (-5 ± 0.4°C), whereas the lower back showed the smallest (-1.9 ± 0.4°C). Thermal sensations varied significantly by location and independently from regional variations in skin cooling with colder sensations reported on the lateral abdomen and lower back. Similarly, the frequency of perceived skin wetness was significantly greater on the lateral and lower back as opposed to the medial chest. Overall wetness perception was slightly higher under warm conditions. Significantly more unpleasant sensations were recorded when the lateral abdomen and lateral and lower back were stimulated. We conclude that humans present regional differences in skin wetness perception across the torso, with a pattern similar to the regional differences in thermosensitivity to cold. These findings indicate the presence of a heterogeneous distribution of cold-sensitive thermo-afferent information.


Development of bovine fetal testis tissue after ectopic xenografting in mice.
Testis tissue xenografting represents a versatile model to study testis biology, and to preserve fertility in immature animals. To evaluate whether bovine fetal testes can mature when grafted into mouse hosts, small fragments of testes from midgestation (125 to 145 days of gestation) bovine fetuses were grafted ectopically into immunodeficient castrated male mice. At grafting, donor tissue displayed the typical seminiferous cords composed of gonocytes and primitive Sertoli cells. At 5 or 10 months after grafting, weight of the seminal vesicles in recipient mice was indicative of production of bioactive testosterone by xenografts. Xenografts showed similar development regardless of donor age. At 5 months, tubule formation occurred but germ cell differentiation had not proceeded beyond the spermatogonia stage. At 10 months, an increase in tubule size was evident and pachytene spermatocytes were observed as the most advanced type of germ cells in the xenografts of 2 donors. The number of tubules with germ cells was reduced in xenografts compared to donor tissue, but at 10 months the number of germ cells per tubule was higher than in donors. Germ cell proliferation was similar in donor tissue and xenografts. However, Sertoli cells showed a higher proliferation rate in xenografts collected at 5 months than in donor fetal testes and xenografts collected at 10 months. Sertoli cells in xenografts showed a progressive but incomplete loss of expression of Müllerian inhibiting substance and weak androgen receptor expression, indicating an incomplete Sertoli cell maturation. In conclusion, fetal testis tissue developed partially, qualitatively similar to pubertal testes in situ.


Expression of complement components differs between kidney allografts from living and deceased donors.
A disparity remains between graft survival of renal allografts from deceased donors and from living donors. A better understanding of the molecular mechanisms that underlie this disparity may allow the development of targeted therapies to enhance graft survival. Here, we used microarrays to examine whole genome expression profiles using tissue from 53 human renal allograft protocol biopsies obtained both at implantation and after transplantation. The gene expression profiles of living-donor kidneys and pristine deceased-donor kidneys (normal histology, young age) were significantly different before reperfusion at implantation. Deceased-donor kidneys exhibited a significant increase in renal expression of complement genes; posttransplantation biopsies from well-functioning, nonrejecting kidneys, regardless of donor source, also demonstrated a significant increase in complement expression. Peritransplantation phenomena, such as donor death and possibly cold ischemia time, contributed to differences in complement pathway gene expression. In addition, complement gene expression at the time of implantation was associated with both early and late graft function. These data suggest that complement-modulating therapy may improve graft outcomes in renal transplantation.


Network Properties for Ranking Predicted miRNA Targets in Breast Cancer.
MicroRNAs control the expression of their target genes by translational repression and transcriptional cleavage. They are involved in various biological processes including development and progression of cancer. To uncover the biological role of miRNAs it is important to identify their target genes. The small number of experimentally validated target genes makes computer prediction methods very important. However, state-of-the-art prediction tools result in a great number of putative targets with an unpredictable number of false positives. In this paper, we propose and evaluate two approaches for ranking the biological relevance of putative targets of miRNAs which are associated with breast cancer.


The employment costs of caregiving in Norway.
Informal eldercare is an important pillar of modern welfare states and the ongoing demographic transition increases the demand for it while social trends reduce the supply. Substantial opportunity costs of informal eldercare in terms of forgone labor opportunities have been identified, yet the effects seem to differ substantially across states and there is a controversy on the effects in the Nordic welfare states. In this study, the effects of informal care on the probability of being employed, the number of hours worked, and wages in Norway are analyzed using data from the Life cOurse, Generation, and Gender survey. New and previously suggested instrumental variables are used to control for the potential endogeneity existing between informal care and employment-related outcomes. In total, being an informal caregiver in Norway is found to entail substantially less costs in terms of forgone formal employment opportunities than in non-Nordic welfare states.


Community-Based Lifestyle Intervention in Patients With Coronary Artery Disease: The RESPONSE-2 Trial.
Among patients with coronary artery disease (CAD), improvement of lifestyle-related risk factors (LRFs) reduces cardiovascular morbidity and mortality. However, modification of LRFs is highly challenging. This study sought to evaluate the impact of combining community-based lifestyle programs with regular hospital-based secondary prevention. The authors performed a randomized controlled trial of nurse-coordinated referral of patients and their partners to 3 widely available community-based lifestyle programs, in 15 hospitals in the Netherlands. Patients admitted for acute coronary syndrome and/or revascularization, with ≥1 LRF (body mass index >27 kg/m2, self-reported physical inactivity, and/or smoking) were included. All patients received guideline-based usual care. The intervention was based on 3 lifestyle programs for weight reduction, increasing physical activity, and smoking cessation. The primary outcome was the proportion of success at 12 months, defined as improvement in ≥1 qualifying LRF using weight (≥5% reduction), 6-min-walking distance (≥10% improvement), and urinary cotinine (200 ng/ml detection limit) without deterioration in the other 2. The authors randomized 824 patients. Complete data on the primary outcome were available in 711 patients. The proportion of successful patients in the intervention group was 37% (133 of 360) compared with 26% (91 of 351) in the control group (p = 0.002; risk ratio: 1.43; 95% confidence interval: 1.14 to 1.78). In the intervention group, partner participation was associated with a significantly greater success rate (46% vs. 34%; p = 0.03). Among patients with coronary artery disease, nurse-coordinated referral to a comprehensive set of community-based, widely available lifestyle interventions, with optional partner participation, leads to significant improvements in LRFs. (RESPONSE-2: Randomised Evaluation of Secondary Prevention by Outpatient Nurse SpEcialists 2; NTR3937).


Global Metal Use Targets in Line with Climate Goals.
Metals underpin essential functions in modern society, yet their production currently intensifies climate change. This paper develops global targets for metal flows, stocks, and use intensity in the global economy out to 2100. These targets are consistent with emissions pathways to achieve a 2 °C climate goal and cover six major metals (iron, aluminum, copper, zinc, lead, and nickel). Results indicate that despite advances in low-carbon metal production, a transformative system change to meet the society's needs with less metal is required to remain within a 2 °C pathway. Globally, demand for goods and services over the 21st century needs to be met with approximately 7 t/capita of metal stock-roughly half the current level in high-income countries. This systemic change will require a peak in global metal production by 2030 and deep decoupling of economic growth from both metal flows and stocks. Importantly, the identified science-based targets are theoretically achievable through such measures as efficient design, more intensive use, and longer product lifetime, but immediate action is crucial before middle- and low-income countries complete full-scale urbanization.


Proeryptotic Activity of 4-Hydroxynonenal: A New Potential Physiopathological Role for Lipid Peroxidation Products.
Eryptosis is a physiological, apoptosis-like death of injured erythrocytes crucial to prevent premature haemolysis and the pathological sequalae generated by cell-free haemoglobin. When dysregulated, the process is associated to several inflammatory-based pathologies. 4-Hydroxy-trans-2-nonenal (HNE) is an endogenous signalling molecule at physiological levels and, at higher concentrations, is involved in the pathogenesis of several inflammatory-based diseases. This work evaluated whether HNE could induce eryptosis in human erythrocytes. Measurements of phosphatidylserine, cell volume, intracellular oxidants, Ca++, glutathione, ICAM-1, and ceramide were assessed by flow cytometry. Scanning electron microscopy evaluated morphological alterations of erythrocytes. Western blotting assessed caspases. PGE2 was measured by ELISA. Adhesion of erythrocytes on endothelial cells was evaluated by gravity adherence assay. HNE in the concentration range between 10-100 µM induces eryptosis, morphological alterations correlated to caspase-3 activation, and increased Ca++ levels. The process is not mediated by redox-dependent mechanisms; rather, it strongly depends on PGE2 and ceramide. Interestingly, HNE induces significant increase of erythrocytes adhesion to endothelial cells (ECs) that are in turn dysfunctionated as evident by overexpression of ICAM-1. Our results unveil a new physiopathological role for HNE, provide mechanistic details of the HNE-induced eryptosis, and suggest a novel mechanism through which HNE could exert pro-inflammatory effects.


SERCaMP: a carboxy-terminal protein modification that enables monitoring of ER calcium homeostasis.
Endoplasmic reticulum (ER) calcium homeostasis is disrupted in diverse pathologies, including neurodegeneration, cardiovascular diseases, and diabetes. Temporally defining calcium dysregulation during disease progression, however, has been challenging. Here we describe secreted ER calcium-monitoring proteins (SERCaMPs), which allow for longitudinal monitoring of ER calcium homeostasis. We identified a carboxy-terminal modification that is sufficient to confer release of a protein specifically in response to ER calcium depletion. A Gaussia luciferase (GLuc)-based SERCaMP provides a simple and sensitive method to monitor ER calcium homeostasis in vitro or in vivo by analyzing culture medium or blood. GLuc-SERCaMPs revealed ER calcium depletion in rat primary neurons exposed to various ER stressors. In vivo, ER calcium disruption in rat liver was monitored over several days by repeated sampling of blood. Our results suggest that SERCaMPs will have broad applications for the long-term monitoring of ER calcium homeostasis and the development of therapeutic approaches to counteract ER calcium dysregulation.


Telephone follow-up to a mail survey: when to offer an interview compared to a reminder call.
Using a different mode of contact on the final follow-up to survey non-respondents is an identified strategy to increase response rates. This study was designed to determine if a reminder phone call or a phone interview as a final mode of contact to a mailed survey works better to increase response rates and which strategy is more cost effective. A randomized study was embedded within a survey study of individuals treated with ulcerative colitis conducted in March 2009 in Olmsted County, Minnesota. After two mail contacts, non-respondents were randomly assigned to either a reminder telephone call or a telephone interview. Average cost per completed interview and response rates were compared between the two experimental conditions. The response rate in the reminder group and the interview did not differ where we considered both a completed survey and a signed form a complete (24% vs. 29%, p = 0.08). However, if such a signed form was not required, there was a substantial advantage to completing the interview over the phone (24% vs. 43%, p < 0.0001). The reminder group on average cost $27.00 per completed survey, while the interview group on average cost $53.00 per completed survey when a signed form was required and $36.00 per complete when a signed form was not required. The additional cost of completing an interview is worth it when an additional signed form is not required of the respondent. However, when such a signed form is required, offering an interview instead of a reminder phone call as a follow up to non-respondents does not increase response rates enough to outweigh the additional costs.


Secretome analysis identifies novel signal Peptide peptidase-like 3 (Sppl3) substrates and reveals a role of Sppl3 in multiple Golgi glycosylation pathways.
Signal peptide peptidase-like 3 (Sppl3) is a Golgi-resident intramembrane-cleaving protease that is highly conserved among multicellular eukaryotes pointing to pivotal physiological functions in the Golgi network which are only beginning to emerge. Recently, Sppl3 was shown to control protein N-glycosylation, when the key branching enzyme N-acetylglucosaminyltransferase V (GnT-V) and other medial/trans Golgi glycosyltransferases were identified as first physiological Sppl3 substrates. Sppl3-mediated endoproteolysis releases the catalytic ectodomains of these enzymes from their type II membrane anchors. Protein glycosylation is a multistep process involving numerous type II membrane-bound enzymes, but it remains unclear whether only few of them are Sppl3 substrates or whether Sppl3 cleaves many of them and thereby controls protein glycosylation at multiple levels. Therefore, to systematically identify Sppl3 substrates we used Sppl3-deficient and Sppl3-overexpression cell culture models and analyzed them for changes in secreted membrane protein ectodomains using the proteomics "secretome protein enrichment with click sugars (SPECS)" method. SPECS analysis identified numerous additional new Sppl3 candidate glycoprotein substrates, several of which were biochemically validated as Sppl3 substrates. All novel Sppl3 substrates adopt a type II topology. The majority localizes to the Golgi network and is implicated in Golgi functions. Importantly, most of the novel Sppl3 substrates catalyze the modification of N-linked glycans. Others contribute to O-glycan and in particular glycosaminoglycan biosynthesis, suggesting that Sppl3 function is not restricted to N-glycosylation, but also functions in other forms of protein glycosylation. Hence, Sppl3 emerges as a crucial player of Golgi function and the newly identified Sppl3 substrates will be instrumental to investigate the molecular mechanisms underlying the physiological function of Sppl3 in the Golgi network and in vivo. Data are available via ProteomeXchange with identifier PXD001672.


Perampanel, a selective, noncompetitive α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor antagonist, as adjunctive therapy for refractory partial-onset seizures: interim results from phase III, extension study 307.
To evaluate safety, tolerability, and seizure outcome data during long-term treatment with once-daily adjunctive perampanel (up to 12 mg/day) in patients with refractory partial-onset seizures. Study 307 was an extension study for patients completing the double-blind phase of three pivotal phase III trials (studies 304, 305, and 306). The study consisted of two phases: an open-label treatment phase (including a 16-week blinded conversion period and a planned 256-week maintenance period) and a 4-week follow-up phase. Patients were blindly titrated during the conversion period to their individual maximum tolerated dose (maximum 12 mg/day). Adverse events (AEs) were monitored throughout the study and seizure frequency recorded. The interim data cutoff date for analyses was December 1, 2010. In total, 1,218 patients were enrolled in the study. At the interim cutoff date, 1,186 patients were in the safety analysis set; 1,089 (91.8%) patients had >16 weeks of exposure to perampanel, 580 (48.9%) patients had >1 year of exposure, and 19 (1.6%) patients had >2 years of exposure. At the interim analysis, 840 (70.8%) patients remained on perampanel treatment. The large majority of patients (n = 1,084 [91%]) were titrated to 10 mg or 12 mg/day. Median (range) duration of exposure was 51.4 (1.1-128.1) weeks. Treatment-emergent AEs were reported in 87.4% of patients. The most frequent were dizziness (43.9%), somnolence (20.2%), headache (16.7%), and fatigue (12.1%). Serious AEs were reported in 13.2% of patients. In the intent-to-treat analysis set (n = 1,207), the frequency of all seizures decreased over the first 26 weeks of perampanel treatment in patients with at least 26 weeks of exposure to perampanel (n = 1,006 [83.3%]); this reduction was maintained in patients with at least 1 year of exposure (n = 588 [48.7%]). The overall median percent changes in seizure frequency in patients included in each 13-week interval of perampanel treatment were -39.2% for weeks 14-26 (n = 1,114), -46.5% for weeks 40-52 (n = 731), and -58.1% for weeks 92-104 (n = 59). Overall responder rates in patients included in each 13-week interval of perampanel treatment were 41.4% for weeks 14-26 (n = 1,114), 46.9% for weeks 40-52 (n = 731), and 62.7% for weeks 92-104 (n = 59). During the blinded conversion period, the reduction in seizure frequency in patients previously randomized to placebo (-42.4%, n = 369) was similar to that in patients previously randomized to perampanel (-41.5%, n = 817). Consistent with pivotal phase III trials, these interim results demonstrated that perampanel had a favorable tolerability profile in patients with refractory partial-onset seizures over the longer term. The decrease in seizure frequency was consistent and maintained in those patients over at least 1 year of perampanel exposure.


Transcatheter umbrella closure of congenital heart defects.
Between October 1984 and September 1986, we attempted transcatheter umbrella closure, using the Rashkind PDA occluder, of 12 congenital or postoperative cardiovascular defects (other than patent ductus arteriosus [PDA]) in 11 patients. In each, we used the umbrella for closure because the defect was too short and/or too large to close with conventional transcatheter methods. The defects included three post-Glenn venous communications (superior vena cava-right atrium, n = 2; azygos vein to inferior vena cava), four congenital "interatrial defects" producing cyanosis ("coronary sinus" septal defect, left superior vena cava to left atrium, patent foramen ovale, left inferior vena cava to left atrium), and five non-PDA systemic-to-pulmonary arterial communications (two congenital and three postoperative). Ten of 12 defects were embolized successfully; nine had complete or subtotal closure, and one was partially closed. The first attempted closure resulted in embolization of a 12 mm device to a lower-lobe pulmonary artery, without clinical sequelae. No other complications occurred. Clinical improvement was most dramatic in those patients whose cyanosis was relieved and less obvious when pulmonary blood flow was reduced. The Rashkind umbrella device, originally designed for closure of PDA, considerably expands the list of congenital or operative defects that can be closed nonsurgically.


Illuminati: a form of gene expression plasticity in Drosophila neural stem cells.
While testing for genome instability in Drosophila as reported by unscheduled upregulation of UAS-GFP in cells that co-express GAL80 and GAL4, we noticed that, as expected, background levels were low in most developing tissues. However, GFP-positive clones were frequent in the larval brain. Most of these clones originated from central brain neural stem cells. Using imaging-based approaches and genome sequencing, we show that these unscheduled clones do not result from chromosome loss or mutations in GAL80. We have named this phenomenon 'Illuminati'. Illuminati is strongly enhanced in brat tumors and is also sensitive to environmental conditions such as food content and temperature. Illuminati is suppressed by Su(var)2-10, but it is not significantly affected by several modifiers of position effect variegation or Gal4::UAS variegation. We conclude that Illuminati identifies a previously unknown type of functional instability that may have important implications in development and disease.


Evaluation of the preclinical efficacy of four antivenoms, distributed in sub-Saharan Africa, to neutralize the venom of the carpet viper, Echis ocellatus, from Mali, Cameroon, and Nigeria.
Snakebite envenoming causes a heavy toll in sub-Saharan Africa in terms of mortality and sequelae. In the West African savannah, the viperid Echis ocellatus is responsible for the vast majority of bites. In the last decades, several new antivenoms have been introduced for the treatment of these envenomings, although the assessment of their preclinical efficacy against the venom of E. ocellatus has been studied only for some of them. This work analyzed comparatively the ability of four antivenoms (FAV Afrique, EchiTAb G, EchiTAB-Plus-ICP(®), and Inoserp™ Panafricain) to neutralize lethal, hemorrhagic, and in vitro coagulant activities of the venoms of E. ocellatus from Mali, Cameroon, and Nigeria. In addition, an immunoaffinity chromatography antivenomic protocol was used to assess the ability of the four antivenoms to bind to the proteins of these venoms. Results showed that all the antivenoms were effective in the neutralization of the three effects investigated, and were able to immunocapture, completely or partially, the most abundant components in the E. ocellatus venoms from the geographical origins sampled. Our observations also highlighted quantitative differences between antivenoms in their neutralizing and antivenomics profiles, especially regarding neutralization of in vitro coagulant activity, suggesting that different doses of these antivenoms are probably needed for an effective treatment of human envenomings by this species.


Potential for false positive HIV test results with the serial rapid HIV testing algorithm.
Rapid HIV tests provide same-day results and are widely used in HIV testing programs in areas with limited personnel and laboratory infrastructure. The Uganda Ministry of Health currently recommends the serial rapid testing algorithm with Determine, STAT-PAK, and Uni-Gold for diagnosis of HIV infection. Using this algorithm, individuals who test positive on Determine, negative to STAT-PAK and positive to Uni-Gold are reported as HIV positive. We conducted further testing on this subgroup of samples using qualitative DNA PCR to assess the potential for false positive tests in this situation. Of the 3388 individuals who were tested, 984 were HIV positive on two consecutive tests, and 29 were considered positive by a tiebreaker (positive on Determine, negative on STAT-PAK, and positive on Uni-Gold). However, when the 29 samples were further tested using qualitative DNA PCR, 14 (48.2%) were HIV negative. Although this study was not primarily designed to assess the validity of rapid HIV tests and thus only a subset of the samples were retested, the findings show a potential for false positive HIV results in the subset of individuals who test positive when a tiebreaker test is used in serial testing. These findings highlight a need for confirmatory testing for this category of individuals.


Separation and characterization of two chemically distinct lipopolysaccharides in two Pectinatus species.
Lipopolysaccharides (LPS) from the type strains of the anaerobic beer spoilage bacteria Pectinatus cerevisiiphilus and P. frisingensis were extracted with the 5:5:8 volume ratio modification of the phenolchloroform-petroleum ether method (H. Brade and C. Galanos, Eur. J. Biochem. 122:233-237, 1982). Sequential precipitations of LPS with water and acetone from the phenol phase yielded LPS which differed in that water-precipitable material (LPS-H2O; 0.1 to 0.4% of the dry weight of the cells) was rough-type LPS, whereas acetone-precipitable material (LPS-Ac; 4.6 to 5.8% of the dry weight) contained both rough-type LPS and high-molecular-weight material resembling smooth LPS. The LPS were chemically characterized, and they contained D-glucosamine, 4-amino-4-deoxy-L-arabinose, 3-deoxy-D-manno-2-octulosonic acid, D-fucose, D-galactose, D-glucose, D-mannose, and phosphate. D-Fucose was present mostly in LPS-Ac, suggesting that it is a constituent of the O antigen. The major fatty acids were ester- and amide-linked (R)-3-hydroxytridecanoic and ester-linked undecanoic acids, with minor amounts of ester-linked tridecanoic and (R)-3-hydroxyundecanoic acids. The chemical compositions of LPS-H2O and LPS-Ac suggested that they differ not only in their smooth or rough nature but also in the structure of their core regions. This may explain their different precipitabilities from the extraction mixture. The extraction method was also shown to be applicable to the isolation of smooth-type LPS from Salmonella enterica serovar Typhimurium. Extraction of two Typhimurium strains carrying chemically different O antigens resulted in high yields (8% of the dry weight) of LPS. Strain SH2183, which contains the relatively hydrophobic O-4,5,12 antigen yielded almost exclusively LPS-Ac, whereas the LPS of strain SH5770, which has a hydrophilic O-6,7 antigen, was exclusively LPS-H2O. No fractionation to smooth and rough LPS occurred with the Typhimurium strains.


HLA associations in the antitumor response against malignant melanoma.
In this study we analyzed the human leukocyte antigen (HLA) pattern of North American Caucasian patients with metastatic melanoma as compared with the North American Caucasian (NAC) population. We also investigated whether the HLA type of melanoma patients had an effect on their tolerance and response to interleukin-2 (IL-2)-based therapy. Four hundred twelve serologic phenotypes of Caucasian melanoma patients referred to the National Cancer Institute, National Institutes of Health, from February 1989 through December 1993 were collected by typing the patient's peripheral blood lymphocytes. Furthermore, 74 melanoma patients were typed for HLA class II by high-resolution sequence specific primer-polymerase chain reaction. Response rate and treatment-related toxicity in those patients receiving IL-2-based treatment (N = 272) were compared with HLA serologic types. The frequency of four HLA-B alleles was significantly different in the melanoma compared with the NAC population: of these, HLA-B5, -B8, and -B15 had a frequency falling between the NAC and the Northern European population. No other significant differences between melanoma patients and NAC population were noted for other HLA loci. A correlation was noted between HLA-DR3 and -DR4 alleles and decreased tolerance to IL-2, whereas homozygosity for HLA-DR decreased the chance of response. There were no significant associations between HLA type and response. It is unlikely that the associations noted between some HLA-B alleles and melanoma bear significantly on the etiology of the disease. The differences seen between American melanoma patients and the NAC population are probably best explained by geographical ancestry. The association between HLA-DR and tolerance to IL-2 therapy noted in this study may offer insight toward the understanding of mechanisms regulating the cascade of events after the systemic administration of IL-2.


Spectral analysis of muscle sympathetic nerve activity in heat-stressed humans.
Whole body heating increases muscle sympathetic nerve activity (MSNA); however, the effect of heat stress on spectral characteristics of MSNA is unknown. Such information may provide insight into mechanisms of heat stress-induced MSNA activation. The purpose of the present study was to test the hypothesis that heat stress-induced changes in systolic blood pressure variability parallel changes in MSNA variability. In 13 healthy subjects, MSNA, electrocardiogram, arterial blood pressure (via Finapres), and respiratory activity were recorded under both normothermic and heat stress conditions. Spectral characteristics of integrated MSNA, R-R interval, systolic blood pressure, and respiratory excursions were assessed in the low (LF; 0.03-0.15 Hz) and high (HF; 0.15-0.45 Hz) frequency components. Whole body heating significantly increased skin and core body temperature, MSNA burst rate, and heart rate, but not mean arterial blood pressure. Systolic blood pressure and R-R interval variability were significantly reduced in both the LF and HF ranges. Compared with normothermic conditions, heat stress significantly increased the HF component of MSNA, while the LF component of MSNA was not altered. Thus the LF-to-HF ratio of MSNA oscillatory components was significantly reduced. These data indicate that the spectral characteristics of MSNA are altered by whole body heating; however, heat stress-induced changes in MSNA do not parallel changes in systolic blood pressure variability. Moreover, the reduction in LF component of systolic blood pressure during heat stress is unlikely related to spectral changes in MSNA.


Motivated Cue-Integration and Emotion Regulation: Awareness of the Association Between Interoceptive and Exteroceptive Embodied Cues and Personal Need Creates an Emotion Goal.
Research on emotion suggests that individuals regulate their emotions to attain hedonic or instrumental goals. However, little is known of emotion regulation under low emotional clarity. The theory of motivated cue integration (MCI) suggests that emotion regulation under low emotional clarity should be understood as dissociation between a high-level individual hierarchical system of goals and low level interoceptive and exteroceptive embodied cues. MCI conceptualizes low emotional clarity as the product of low access to signals of emotion that result in prediction error associated with mismatch between the current bodily state and the predicted state. This deficit in emotional processing could be understood as a problem of means substitution, suggesting that use of alternative multisensory data may facilitate situational evaluation. Based on this reasoning, a new perspective on emotion regulation under low emotional clarity is presented, according to which interchangeable attention to multisensory data associated with words, associations, and images may help in cue integration, enabling the creation of a link between concrete bodily cues, abstract mental representation, and a more accurate prediction. Based on the idea that emotional episodes are conceptualized as special types of goal-directed action episodes, this process will lead to the creation of broader integrative meaning, results in the development of emotion goal.


Accessory veins in nonmaturing autogenous arteriovenous fistulae: analysis of anatomic features and impact on fistula maturation.
The appropriate management of nonmaturing arteriovenous (AV) fistulae continues to be a controversial issue. While coil embolization of accessory side-branch veins can be performed to encourage maturation of nonmaturing AV fistulae, the true efficacy and optimal patient population are not well understood. Fistulagrams performed on nonmaturing AV fistulae were retrospectively reviewed in 145 patients (86 males, median age 63 years) for the presence of accessory veins. Fistula and accessory vein measurements were obtained, as were rates of eventual fistula maturation after accessory vein coil embolization. Of 145 nonmaturing fistulae, 49 (34%) had a stenosis without any accessory veins, 76 (52%) had a stenosis and one or more accessory veins, and 20 (14%) had an accessory vein without concurrent stenosis. Eighteen AV fistulae had one or more accessory veins without coexisting stenosis. Nine fistulae had a caliber decrease immediately downstream from the accessory vein. Coil embolization of dominant accessory veins with a caliber decrease immediately downstream (n = 6) resulted in a 100% eventual fistula maturation rate versus 67% for fistulae without this configuration (n = 6, p = 0.15). Accessory vein size was not correlated with maturation rates (p = 0.51). The majority of nonmaturing fistulae with accessory veins had a coexisting stenosis. Higher maturation rates may result with selected anatomic parameters, although additional studies with more robust sample sizes are needed prior to definitive conclusions.


Characterization of six flagellin genes in the H3, H53 and H54 standard strains of Escherichia coli.
Six flagellin genes in three H standard Escherichia coli strains for H3, H53 and H54 were characterized. Each strain has two flagellin genes, one of which is expressed as its standard H antigen. A pair of flagellin genes flkA3 (encoding for H3 antigen) and fliC16 (H16) was cloned from Bi7327-41, flkA53 (H53) and fliC-53 from E480-68, and flmA54 (H54) and fliC-54 from E223-69. Two fliC genes, fliC-53 and fliC-54, are nonfunctional owing to the insertions of IS1 and IS1222, respectively. The flkA and flmA regions are located in the 3' end of the rnpB gene and near the nlpA gene, respectively. Each of them is followed by a gene homologous to fljA, which is known to repress the expression of fliC(i) in Salmonella enterica serovar Typhimurium. These results suggest that they are derived from the same origin of the fljBA operon. However, these regions contain neither the hin gene nor the invertible H segment. The four flagellin genes, fliC16, flkA3, flkA53 and flmA54, share high homology in nucleotide and amino-acid sequences with one another and with the S. enterica serovar Typhimurium flagellin genes. The promoter sequence of fliC16 is homologous to that of fliC(i), whereas the promoter sequences of flkA and flmA are homologous to that of fljB. The terminator sequences of the fliC16, fliC-53 and fliC-54 genes are conserved among themselves and identical with that of the E. coli fliC48 gene. Three FljA repressors, FljA3, FljA53 and FljA54, are homologous highly with one another and moderately with FljA of Salmonella. These results indicate that six flagellin genes analyzed are markedly similar to the Salmonella flagellin genes, suggesting their lateral transfer from Salmonella.


Failure of nonoperative management of splenic injuries: causes and consequences.
Presenting patient characteristics can predict which patients will fail nonoperative therapy of blunt splenic injuries. Retrospective descriptive population study. All patients admitted with blunt splenic trauma were identified from a statewide trauma registry between January 1, 1995, and December 31, 2001. Patients were characterized as requiring immediate intervention or successful or failed nonoperative management based on time from emergency department arrival to intervention (surgery or angioembolectomy). Injury and patient characteristics included age, the presence of hypotension, Injury Severity Score, and the Glasgow Coma Scale score. Risk factors for the failure of nonoperative management were evaluated using the chi(2) statistic. The failure of nonoperative management associated with the admitting hospital's trauma designation level was evaluated using logistic regression. None. Determine factors associated with failure of nonoperative management of blunt splenic injuries. Two thousand two hundred forty-three patients met criteria for inclusion in the study. Six hundred ten patients (27%) underwent immediate splenectomy, splenorrhaphy, or splenic embolization (within 4 hours). Of the remaining 1633 patients who were admitted with planned nonoperative management, 252 patients (15%) failed. Of the injury and patient characteristics reviewed, being older than 55 years and having an ISS higher than 25 were significantly associated with failure. Risk of failure also increased with admission to a level III or IV trauma hospital compared with a level I trauma hospital. Being older than 55 years and having an ISS higher than 25 along with admission to a level III or IV trauma hospital were associated with a significant risk of failure of nonoperative management of splenic injuries. The Glasgow Coma Scale score, associated injuries, and presenting hemodynamics were not predictive of failure in this large retrospective review.


Intensification of bioactive compounds extraction from medicinal plants using ultrasonic irradiation.
Extraction processes are largely used in many chemical, biotechnological and pharmaceutical industries for recovery of bioactive compounds from medicinal plants. To replace the conventional extraction techniques, new techniques as high-pressure extraction processes that use environment friendly solvents have been developed. However, these techniques, sometimes, are associated with low extraction rate. The ultrasound can be effectively used to improve the extraction rate by the increasing the mass transfer and possible rupture of cell wall due the formation of microcavities leading to higher product yields with reduced processing time and solvent consumption. This review presents a brief survey about the mechanism and aspects that affecting the ultrasound assisted extraction focusing on the use of ultrasound irradiation for high-pressure extraction processes intensification.


Extensive diversity of Ig-superfamily proteins in the immune system of insects.
The extensive somatic diversification of immune receptors is a hallmark of higher vertebrates. However, whether molecular diversity contributes to immune protection in invertebrates is unknown. We present evidence that Drosophila immune-competent cells have the potential to express more than 18,000 isoforms of the immunoglobulin (Ig)-superfamily receptor Down syndrome cell adhesion molecule (Dscam). Secreted protein isoforms of Dscam were detected in the hemolymph, and hemocyte-specific loss of Dscam impaired the efficiency of phagocytic uptake of bacteria, possibly due to reduced bacterial binding. Importantly, the molecular diversity of Dscam transcripts generated through a mechanism of alternative splicing is highly conserved across major insect orders, suggesting an unsuspected molecular complexity of the innate immune system of insects.


The association of working alliance, outcome expectation, adherence and self-efficacy with clinical outcomes for Achilles tendinopathy: A feasibility cohort study (the MAP study).
This study evaluated the feasibility of a large longitudinal cohort study utilizing an online platform to investigate the association and predictive relationship of working alliance, outcome expectations, adherence and self-efficacy with outcome in Achilles tendinopathy. The objectives were: (1) to determine the recruitment and retention rate and (2) to carry out preliminary data analysis of the selected variables and clinical outcomes. A multi-centred, longitudinal feasibility cohort study was used. Eligible participants were directed to a bespoke online platform hosting study information and the outcome measures in the form of an online questionnaire. Responses from the online questionnaire were collected on three occasions: at baseline, at 6 and at 12 weeks following completion of the first questionnaire. Feasibility outcomes (recruitment and retention rates) were described using descriptive statistics. The website recorded a total 55 views. These 55 views resulted in 24 participants consenting to join the study. The questionnaire at baseline was started 63 times and completed on 60 separate occasions resulting in a 95% conversion rate. Retainment for completion of the questionnaire for a second time was 83.3% and for the third time was 66.6%. All questionnaires were completed fully yielding a missing data indicator of 0%. Feasibility studies ask the question 'can this be done'? Based on the data from recruitment and rates and exploratory correlation analysis a future study can be done; this previously untested online platform appears feasible, but changes could be useful before proceeding to a much larger study.


Microaggressions, Feelings of Difference, and Resilience Among Adopted Children with Sexual Minority Parents.
Limited research exists about the unique experiences and possible marginalization of children with sexual minority parents. From a larger longitudinal project of diverse adoptive families, we examined cross-sectional data using mixed methods from interviews with 49 adopted children (M age = 8 years; 47% female) in 27 two-father and 22 two-mother families. Using thematic analysis, we coded themes of awareness of difference, microaggressions, and resilience (i.e., coping and positive family conceptualizations). Children experienced "feeling different" and microaggressions from peers, but generally at a low to medium intensity and with neutral (not negative) emotion. More instances of resilience and positive family conceptualizations were reported than microaggressions or feelings of difference, suggesting that children develop positive perceptions of their family and navigate experiences of difference with resilience. Filling important gaps in the literature, we discuss implications of our results for practice and policy.


Identification of a cell-surface antigen associated with activated T lymphoblasts and activated platelets.
We have identified and biochemically characterized an antigen, 8A3, which is expressed on activated T lymphoblasts and activated platelets. Monoclonal antibodies to 8A3 were raised against the primitive lymphoid/myeloid cell line KG1a and additionally bound to the erythroleukemia-derived cell line HEL, whilst exhibiting little or no reactivity with a panel of other hematopoietic cell lines. The 8A3 antigen was expressed on poorly differentiated T-cell leukemias and on phytohemagglutinin-activated T-cells maintained in interleukin-2 (7,000 sites/cell). This antigen, though not detected on resting platelets, was expressed on thrombin-activated platelets (2,000 sites/platelet). Antibodies to 8A3 identified polypeptides of Mr 170,000 and 150,000 in lysates of surface-iodinated KG1a cells, T lymphoblasts, and activated platelets under both reducing and nonreducing conditions. However, peptide mapping and susceptibily to glycosidases indicated that the 8A3 antigen was a monomeric glycoprotein of Mr 170,000 which contained two N-linked endoglycosidase H-sensitive glycans, and that the Mr 150,000 structure was derived from it by proteolytic degradation. The 8A3 antigen was not detectably phosphorylated in KG1a cells in vivo, nor did immune complexes containing it exhibit kinase activity in vitro. Structural and serologic characteristics of the 8A3 antigen indicate that it is different from other previously described leukocyte activation antigens including transferrin receptors, interleukin-2 receptors, members of the integrin family of adhesion molecules, or "restricted" members of the leukocyte-common antigen/CD45 cluster. Furthermore, the 8A3 antigen does not appear to be related to the other previously described activation-specific platelet molecule, GMP140/PADGEM. This antibody may be useful in monitoring T-cell activation status in some clinical situations and in characterizing clinically relevant activation-associated platelet membrane alterations.


Intrinsic and extrinsic factors in muscle aging.
The regenerative potential of skeletal muscle, and overall muscle mass, decline with age. This regenerative potential may be influenced by autocrine growth factors intrinsic to the muscle itself. Extrinsic host factors that may influence muscle regeneration include hormones, growth factors secreted in a paracrine manner by accessory cells, innervation, and antioxidant mechanisms. Unaccustomed exercise, which involves mechanical overload of myofibers, provides a convenient method for studying muscle regeneration in both humans and animal models. An inflammatory response ensues in which distinctive populations of macrophages infiltrate the affected tissue: some of these macrophages are involved in phagocytosis of damaged fibers; other macrophages arriving at later times may deliver growth factors or cytokines that promote regeneration. These include fibroblast growth factor and insulin-like growth factor, which are important regulators of muscle precursor cell growth and differentiation, as well as nerve growth factor, which is essential for maintenance or reestablishment of neuronal contact. Other cytokines, including interleukin-1, tumor necrosis factor, interleukin-15, and ciliary neurotrophic factor, have a strong influence on the balance between muscle protein synthesis and breakdown. The functional activity of invading macrophages can be influenced by age, by factors in myofibers and extracellular matrix, and can be influenced systemically by the antioxidant status of the host.