Tripled Readout Slices in Multi Time-Point pCASL Using Multiband Look-Locker EPI.
Multi time-point pseudo-continuous arterial spin labelling (pCASL) with a Look-Locker EPI readout can sample the signal curve of blood kinetics at multiple time points after the labelling pulse. However, due to signal relaxation of labelled blood, the number of readout slices is limited. The aim of this study is to employ a multiband excitation technique to triple the number of readout slices in multi time-point pCASL. The multiband technique, along with 2-fold in-plane parallel imaging, was incorporated into the Look-Locker EPI for the multi time-point sampling of blood kinetic behaviour following the pCASL labelling scheme. The performance evaluation of the multiband and the single-band techniques were performed on four healthy subjects using a 32-channel head RF coil at 3T. Quantitative perfusion maps were analysed using a combination of labelling with and without flow suppression gradients. The perfusion maps provided by the multiband accelerated multi time-point pCASL were in good agreement with the conventional single-band technique. Multiband acceleration caused SNR loss but offered quantitative perfusion maps in 6.23 min with 18 slices compared with 6 slices within the same time period for the single-band method. As conclusion, the multiband technique can successfully triple the number of readout slices while achieving comparable perfusion data in the same measurement time as the conventional single-band readout.


Adjuvant chemotherapy for patients with primary hepatocellular carcinoma: a meta-analysis.
Numerous studies have investigated the effects of adjuvant chemotherapy for primary hepatocellular carcinoma (HCC) patients. We conducted this analysis to evaluate the efficacy of adjuvant chemotherapy in HCC patients after hepatectomy. PubMed/MEDLINE, EMBASE, Cochrane, and other databases were searched for eligible studies. The major endpoints were overall survival (OS) and disease-free survival (DFS). The pooled odds ratio (OR) was calculated using a random-effects model to summarize the results. In the meta-analysis of 13 randomized control trials (RCTs) and 35 observational studies with 4747 patients, hepatectomy plus adjuvant chemotherapy showed superiority over hepatectomy alone in 1-year DFS (OR = 1.86, 1.38-2.51, p < 0.001), 3-year DFS (OR = 2.37, 1.73-3.24, p < 0.001) and 5-year DFS (OR = 1.99, 1.55-2.55, p < 0.001), as well as 1-year OS (OR = 2.16, 95% confidence interval 1.75-2.68, p < 0.001), 3-year OS (OR = 1.77, 1.48-2.13, p < 0.001) and 5-year OS (OR = 1.92, 1.44-2.56, p < 0.001). Subgroup and sensitivity analysis revealed that only adjuvant TACE had significant survival benefits. The meta-analysis of studies involving patients with portal vein tumor thrombus (PVTT), but not other factors related to recurrence risk, revealed favorable outcomes of the Treatment arm over the Control arm. The present study shows that adjuvant chemotherapy can improve outcomes for HCC patients. The benefits of adjuvant TACE have been confirmed whereas the effects of other adjuvant chemotherapy modalities remain uncertain. Adjuvant chemotherapy is likely to be more applicable to certain patient populations for instance those with PVTT, but further research in identifying these patient factors is of importance for tailoring adjuvant therapies to individual patients in the future.


Common SNPs in FTO gene are associated with obesity related anthropometric traits in an island population from the eastern Adriatic coast of Croatia.
Multiple studies have provided compelling evidence that the FTO gene variants are associated with obesity measures. The objective of the study was to investigate whether FTO variants are associated with a broad range of obesity related anthropometric traits in an island population. We examined genetic association between 29 FTO SNPs and a comprehensive set of anthropometric traits in 843 unrelated individuals from an island population in the eastern Adriatic coast of Croatia. The traits include 11 anthropometrics (height, weight, waist circumference, hip circumference, bicondilar upper arm width, upper arm circumference, and biceps, triceps, subscapular, suprailiac and abdominal skin-fold thicknesses) and two derived measures (BMI and WHR). Using single locus score tests, 15 common SNPs were found to be significantly associated with "body fatness" measures such as weight, BMI, hip and waist circumferences with P-values ranging from 0.0004 to 0.01. Similar but less significant associations were also observed between these markers and bicondilar upper arm width and upper arm circumference. Most of these significant findings could be explained by a mediating effect of "body fatness". However, one unique association signal between upper arm width and rs16952517 (P-value = 0.00156) could not be explained by this mediating effect. In addition, using a principle component analysis and conditional association tests adjusted for "body fatness", two novel association signals were identified between upper arm circumference and rs11075986 (P-value = 0.00211) and rs16945088 (P-value = 0.00203). The current study confirmed the association of common variants of FTO gene with "body fatness" measures in an isolated island population. We also observed evidence of pleiotropic effects of FTO gene on fat-free mass, such as frame size and muscle mass assessed by bicondilar upper arm width and upper arm circumference respectively and these pleiotropic effects might be influenced by variants that are different from the ones associated with "body fatness".


Will the NHS continue to function in an influenza pandemic? A survey of healthcare workers in the West Midlands, UK.
If UK healthcare services are to respond effectively to pandemic influenza, levels of absenteeism amongst healthcare workers (HCWs) must be minimised. Current estimates of the likelihood that HCWs will continue to attend work during a pandemic are subject to scientific and predictive uncertainty, yet an informed evidence base is needed if contingency plans addressing the issues of HCW absenteeism are to be prepared. This paper reports the findings of a self-completed survey of randomly selected HCWs across three purposively sampled healthcare trusts in the West Midlands. The survey aimed to identify the factors positively or negatively associated with willingness to work during an influenza pandemic, and to evaluate the acceptability of potential interventions or changes to working practice to promote the continued presence at work of those otherwise unwilling or unable to attend. 'Likelihood' and 'persuadability' scores were calculated for each respondent according to indications of whether or not they were likely to work under different circumstances. Binary logistic regression was used to compute bivariate and multivariate odds ratios to evaluate the association of demographic variables and other respondent characteristics with the self-described likelihood of reporting to work. The survey response rate was 34.4% (n = 1032). Results suggest absenteeism may be as high as 85% at any point during a pandemic, with potential absence particularly concentrated amongst nursing and ancillary workers (OR 0.3; 95% CI 0.1 to 0.7 and 0.5; 95% CI 0.2 to 0.9 respectively). Levels of absenteeism amongst HCWs may be considerably higher than official estimates, with potential absence concentrated amongst certain groups of employees. Although interventions designed to minimise absenteeism should target HCWs with a low stated likelihood of working, members of these groups may also be the least receptive to such interventions. Changes to working conditions which reduce barriers to the ability to work may not address barriers linked to willingness to work, and may fail to overcome HCWs' reluctance to work in the face of what may still be deemed unacceptable risk to self and/or family.


Elevated serum pancreatic enzyme levels after hemorrhagic shock predict organ failure and death.
Intraluminal pancreatic enzymes have been shown in animal models to be associated with multiple organ failure after hemorrhagic shock, independent of pancreatitis. The translocation of these enzymes into the circulation may serve as a marker of hemorrhagic shock-induced gut ischemia in critically injured trauma patients. We hypothesized that serum amylase and lipase would be significantly elevated in patients presenting in hemorrhagic shock and in those who develop organ failure. : Review of a prospective database at a level-1 trauma center from 2000 to 2005. Two thousand seven hundred eleven critically injured trauma patients without pancreatic injuries were evaluated for shock (systolic pressure <90 mm Hg in the emergency department), massive transfusion (10 units of packed red blood cells within the first 24 hours), and organ failure (standard criteria for acute pulmonary, cardiovascular, renal, and hepatic system failure were used). Serum levels >2 times the upper limit of normal for amylase (30-130 U/L) and lipase (7-60 U/L) were defined as elevated. Univariate analyses were performed with the Pearson's chi, and binary logistic regression was used to determine significant risk factors for organ failure. Results with a p value <0.05 were considered significant and are reported. : Patients with elevated amylase (n = 481, 18%) were more likely to present in shock (16% vs. 8%), require massive transfusion (19% vs. 9%), develop organ failure (34% vs. 16%), and die (23% vs. 13%). Patients with elevated lipase (n = 288, 11%) were more likely to require massive transfusion (18% vs. 10%) and develop organ failure (43% vs. 16%). Independent predictors of organ failure were age (odds ratio [OR] = 1.016), Injury Severity Score (OR = 1.02), massive transfusion (OR = 3.1), elevated amylase (OR = 1.9), and elevated lipase (OR = 3.2). Elevated amylase was also an independent predictor of mortality (OR = 1.3). : Serum levels of pancreatic enzymes are elevated in patients who present in shock or require a massive transfusion and are independent predictors of organ failure. Whether these elevations are caused by ischemic pancreatitis or the translocation of intraluminal enteric pancreatic enzymes is uncertain and future studies are needed. Trauma patients with elevated pancreatic enzymes in the absence of a pancreatic injury have an increased risk of morbidity and mortality.


Initiation and Development of a Toxic and Persistent Pseudo-nitzschia Bloom off the Oregon Coast in Spring/Summer 2015.
In spring/summer 2015, a toxic bloom by the diatom Pseudo-nitzschia (PN) occurred along the west coast of the United States which led to closures of the harvest of razor clams and Dungeness crabs. Twice monthly observations of temperature, salinity, nutrients, chlorophyll and phytoplankton species composition allowed us to track oceanographic conditions preceding and during the development of the bloom. PN cells were first detected during late winter 2015. A PN bloom was initiated following the onset of coastal upwelling in mid-April; subsequent peaks in May and June were sustained by episodic upwelling events and reached magnitudes of 105 cells/L and 106 cells/L, 40% and 90% of the total diatom abundance, respectively. The bloom temporarily crashed in July due to a lack of upwelling, but PN cells increased again in August due to a resumption of upwelling, albeit with lower magnitude. Macronutrient conditions prior to this bloom likely played a critical role in triggering the bloom and its toxicity (particularly silicic acid limitation stress). Nutrient stress preceding the toxic bloom was related to two oceanographic events: an anomalously warm and thick water mass that occupied the northern North Pacific from September 2014 through 2015 leading to a highly-stratified water column, and the drawdown of nitrate and silicic acid during an unusually intense winter phytoplankton bloom in February and early March 2015.


Lunar surface: changes in 31 months and micrometeoroid flux.
Comparison of pictures of the lunar surface taken 31 months apart by Surveyor 3 and Apollo 12 show only one change in the areas disturbed by Surveyor: a 2-millimeter particle, in a footpad imprint, that may have fallen in from the rim or been kicked in by an approaching astronaut. Vertical walls 6 centimeters high did not collapse and dark ejecta remained dark. No meteorite craters as large as 1.5 millimeters in diameter were seen on a smooth soil surface 20 centimeters in diameter; this indicates a micrometeoroid flux lower than 4 x 10(-7) micrometeoroids per square meter-second at an energy equivalent to about 3 x 10(-8) gram at 20 kilometers per second. This flux is near the lower limit of previous determinations.


STAT3, STAT4, NFATc1, and CTCF regulate PD-1 through multiple novel regulatory regions in murine T cells.
Programmed death-1 (PD-1) is a crucial negative regulator of CD8 T cell development and function, yet the mechanisms that control its expression are not fully understood. Through a nonbiased DNase I hypersensitivity assay, four novel regulatory regions within the Pdcd1 locus were identified. Two of these elements flanked the locus, bound the transcriptional insulator protein CCCTC-binding factor, and interacted with each other, creating a potential regulatory compartmentalization of the locus. In response to T cell activation signaling, NFATc1 bound to two of the novel regions that function as independent regulatory elements. STAT binding sites were identified in these elements as well. In splenic CD8 T cells, TCR-induced PD-1 expression was augmented by IL-6 and IL-12, inducers of STAT3 and STAT4 activity, respectively. IL-6 or IL-12 on its own did not induce PD-1. Importantly, STAT3/4 and distinct chromatin modifications were associated with the novel regulatory regions following cytokine stimulation. The NFATc1/STAT regulatory regions were found to interact with the promoter region of the Pdcd1 gene, providing a mechanism for their action. Together these data add multiple novel distal regulatory regions and pathways to the control of PD-1 expression and provide a molecular mechanism by which proinflammatory cytokines, such as IL-6 or IL-12, can augment PD-1 expression.


[Antioxidant properties of lactic acid bacteria--probiotic and yogurt strains].
Antioxidant properties of 14 strains of lactic acid bacteria were evaluated in vitro using FRAP assay, inhibition of luminol oxidation in Hb-H2O2 system and inhibition of NADPH-Fe2+ induced microsomal lipid peroxidation. All strains demonstrated high reducing properties, but only L. casei spp. (including L. casei 114001) and L. fermentum ME-3 revealed pronounced ability to suppress oxidation of luminol (by 43-65,8%) and microsomal lipid peroxidation (by 57,9-89,5%). Either L. casei 114001 (10(8) CFU suspended in physiological solution) or fermented dairy drink containing equivalent amount of L. casei 114001 were daily administered orally to male Wistar rats. Antioxidant capacity of blood plasma, liver and intestines of animals elevated while MDA content in blood plasma decreased.


Sulfonimidamides in Medicinal and Agricultural Chemistry.
The synthesis and evaluation of structural analogues and isosteres are of central importance in medicinal and agricultural chemistry. The sulfonamide functional group represents one of the most important amide isosteres in contemporary drug design, and about 500 such compounds have overcome both the pharmacological and regulatory hurdles that precede studies in humans. The mono aza analogues of sulfonamides, that is, sulfonimidamides, are rapidly gaining popularity as a novel functional group among synthetic chemists involved in the design of biologically active compounds for both pharmaceutical and agrochemical applications. Herein, we review these recent developments to showcase the promise of this functional group.


Satisfaction With the Outcome of Physical Therapist-Prescribed Exercise in Chronic Whiplash-Associated Disorders: Secondary Analysis of a Randomized Clinical Trial.
Study Design Secondary analysis of a randomized clinical trial. Background Patient perception of the benefits gained from treatment is important, yet satisfaction with the outcome of treatment for chronic whiplash-associated disorders (WADs) has not been investigated. Objectives To investigate whether satisfaction with the outcome of treatment for chronic WAD changed over time, and whether there were group differences. Methods Two hundred sixteen people with chronic WAD (66% women; mean age, 40.4 years) participated in a 3-month program of physical therapist-led neck-specific exercises with or without a behavioral approach, or received a prescription of general physical activity. The main outcome was satisfaction with the outcome of treatment, assessed at baseline and 3, 6, and 12 months later. Additional outcomes were enablement and expectation fulfillment. Results Satisfaction improved over time in the 3 groups (odds ratio = 1.15; 95% confidence interval: 1.10, 1.20; P<.001). There was a significant group-by-time interaction (P<.001), with increased odds of being satisfied in the groups receiving neck-specific exercises compared to general physical activity. Enablement increased after completion of the intervention in all groups (P<.001). People who received neck-specific exercises reported greater enablement and expectation fulfillment than people prescribed general physical activity (P<.01). Conclusion Exercise interventions for chronic WAD led to increased satisfaction for 12 months following treatment that was unrelated to the type of exercise intervention received. Level of Evidence Therapy, level 1b. Registered January 22, 2012 at www.ClinicalTrials.gov (NCT01528579). J Orthop Sports Phys Ther 2016;46(8):640-649. Epub 3 Jul 2016. doi:10.2519/jospt.2016.6136.


Molecular typing of Borrelia burgdorferi sensu lato by PCR-restriction fragment length polymorphism analysis.
The etiologic agent of Lyme borreliosis, Borrelia burgdorferi sensu lato, has been isolated from many biologic sources in North America and Eurasia, and isolates have been divided into three distinct genospecies (B. burgdorferi sensu stricto, Borrelia garinii, and Borrelia afzelii). In order to explore the possible association of genospecies with disease manifestation, 60 isolates of B. burgdorferi sensu lato were subjected to 5S rDNA-linked restriction fragment length polymorphism (RFLP) analysis. The results confirmed earlier studies which indicated that virtually all North American isolates are B. burgdorferi sensu stricto, whereas Eurasian strains fall into all three genospecies. Thirty-five isolates were further characterized by PCR amplification of a region of the 16S-23S rDNA spacer and HinfI digestion of the products. This method resulted in the subdivision of B. burgdorferi sensu stricto into two distinct PCR-RFLP types. In contrast, B. garinii isolates all displayed an identical pattern. Additionally, a number of previously unclassified North American isolates (25015, DN127, 19857, 24330) showed distinctively different PCR-RFLP patterns. The application of this method for the typing of uncultured B. burgdorferi directly in biologic samples was demonstrated by analysis of several field-collected Ixodes scapularis tick specimens. The described PCR-RFLP technique should allow for the direct and rapid molecular typing of B. burgdorferi-containing samples and facilitate studies of the relationship between spirochete genotype and clinical disease.


Deficiency of Mitochondrial Glycerol 3-Phosphate Dehydrogenase Exacerbates Podocyte Injury and the Progression of Diabetic Kidney Disease.
Mitochondrial function is essential for bioenergetics, metabolism, and signaling and is compromised in diseases such as proteinuric kidney diseases, contributing to the global burden of kidney failure, cardiovascular morbidity, and death. The key cell type that prevents proteinuria is the terminally differentiated glomerular podocyte. In this study, we characterized the importance of mitochondrial glycerol 3-phosphate dehydrogenase (mGPDH), located on the inner mitochondrial membrane, in regulating podocyte function and glomerular disease. Specifically, podocyte-dominated mGPDH expression was downregulated in the glomeruli of patients and mice with diabetic kidney disease and adriamycin nephropathy. Podocyte-specific depletion of mGPDH in mice exacerbated diabetes- or adriamycin-induced proteinuria, podocyte injury, and glomerular pathology. RNA sequencing revealed that mGPDH regulated the receptor for the advanced glycation end product (RAGE) signaling pathway, and inhibition of RAGE or its ligand, S100A10, protected against the impaired mitochondrial bioenergetics and increased reactive oxygen species generation caused by mGPDH knockdown in cultured podocytes. Moreover, RAGE deletion in podocytes attenuated nephropathy progression in mGPDH-deficient diabetic mice. Rescue of podocyte mGPDH expression in mice with established glomerular injury significantly improved their renal function. In summary, our study proposes that activation of mGPDH induces mitochondrial biogenesis and reinforces mitochondrial function, which may provide a potential therapeutic target for preventing podocyte injury and proteinuria in diabetic kidney disease.


Long-term treatment with ruxolitinib for patients with myelofibrosis: 5-year update from the randomized, double-blind, placebo-controlled, phase 3 COMFORT-I trial.
The randomized, double-blind, placebo-controlled, phase 3 COMFORT-I trial evaluated the JAK1/JAK2 inhibitor ruxolitinib in patients with intermediate-2/high-risk myelofibrosis. The primary and planned 3-year analyses of COMFORT-I data demonstrated that ruxolitinib-the first myelofibrosis-approved therapy-reduced splenomegaly and prolonged overall survival versus placebo. Here, we present the final 5-year results. Patients managed in Australia, Canada, and the USA were randomized centrally (interactive voice response system) 1:1 to oral ruxolitinib twice daily (15 or 20 mg per baseline platelet counts) or placebo. Investigators and patients were blinded to treatment. The secondary endpoints evaluated in this analysis were durability of a ≥35% reduction from baseline in spleen volume (spleen response) and overall survival, evaluated in the intent-to-treat population. Safety was evaluated in patients who received study treatment. Patients were randomized (September 2009-April 2010) to ruxolitinib (n = 155) or placebo (n = 154). At termination, 27.7% of ruxolitinib-randomized patients and 25.2% (28/111) who crossed over from placebo were on treatment; no patients remained on placebo. Patients randomized to ruxolitinib had a median spleen response duration of 168.3 weeks and prolonged median overall survival versus placebo (ruxolitinib group, not reached; placebo group, 200 weeks; HR, 0.69; 95% CI, 0.50-0.96; P = 0.025) despite the crossover to ruxolitinib. The ruxolitinib safety profile remained consistent with previous analyses. The most common new-onset all-grade nonhematologic adverse events starting <12 versus ≥48 months after ruxolitinib initiation were fatigue (29.0 vs 33.3%) and diarrhea (27.8 vs 14.6%). New-onset grade 3 or 4 anemia and thrombocytopenia both primarily occurred within the first 6 months, with no cases after 42 months. The most common treatment-emergent adverse event-related deaths in the ruxolitinib-randomized group were sepsis (2.6%), disease progression (1.9%), and pneumonia (1.9%). The final COMFORT-I results continue to support ruxolitinib as an effective treatment for patients with intermediate-2/high-risk MF. ClinicalTrials.gov, NCT00952289.


[What makes residents of care centers feel at home and satisfied: a Delphi study].
A major event in the human life course is the move to a care center. In order for this transition to be successful, it is important that a care center creates the right conditions to achieve that residents feel at home and satisfied. The aim of this study is to investigate which characteristics are important for this, according to the employees of care centers in the Netherlands. First a limited list of 15 characteristics has been constructed on the basis of literature research. With the help of the Delphi method, in three rounds, employees of Dutch care institutions were then asked about the importance of these characteristics for a successful adaptation of residents to the new living environment and they were asked to provide additional comments. Based on this, characteristics have been reformulated, added or removed. The result after the third round was a high degree of agreement on the characteristics that were considered (very) important. The attribute 'respect', scored in all three rounds the highest, followed directly by the characteristics 'approach' and 'safety'. There was a high degree of agreement about the most important characteristics between the different job levels (such as employee care or supervisor).


Characterisation of phases in nanostructured, multilayered titanium alloys by analytical and high-resolution electron microscopy.
Surface processing of a Ti-6Al-4V alloy led to a complex multilayered microstructure containing several phases of the Ni-Ti-P-Al-O system, which improves the mechanical and tribological surface properties. The microstructure, chemical and phase compositions of the hard layer formed on the surface were investigated by LM, XRD, SEM as well as analytical/high-resolution TEM, STEM, EDS, electron diffraction and FIB. Phase identification based on electron diffraction, HRTEM and EDS microanalysis revealed the presence of several binary and ternary phases in the system Ti-Ni-P, sometimes with partial substitution of Ti by Al. However some phases, mainly nanoparticles, still remain not identified satisfactorily. Electron microscopy techniques used for identification of phases present in surface multilayers and some practical limits to their routine application are reminded here.


Epigenetic CpG demethylation of the promoter and reactivation of the expression of Neurog1 by curcumin in prostate LNCaP cells.
Curcumin (CUR), a major bioactive polyphenolic component from turmeric curry, Curcuma longa, has been shown to be a potent anti-cancer phytochemical with well-established anti-inflammatory and anti-oxidative stress effects. Chromatin remodeling-related epigenetic regulation has emerged as an important mechanism of carcinogenesis, chemoprevention, and chemotherapy. CUR has been found to inhibit histone acetyltransferase activity, and it was also postulated to be a potential DNA methyltransferase (DNMT) and histone deacetylase (HDAC) inhibitor. In this study, we show that when human prostate LNCaP cells were treated with CUR, it led to demethylation of the first 14 CpG sites of the CpG island of the Neurog1 gene and restored the expression of this cancer-related CpG-methylation epigenome marker gene. At the protein level, CUR treatment had limited effects on the expression of epigenetic modifying proteins MBD2, MeCP2, DNMT1, and DNMT3a. Using ChIP assay, CUR decreased MeCP2 binding to the promoter of Neurog1 dramatically. CUR treatment showed different effects on the protein expression of HDACs, increasing the expression of HDAC1, 4, 5, and 8 but decreasing HDAC3. However, the total HDAC activity was decreased upon CUR treatment. Further analysis of the tri-methylation of histone 3 at lysine 27 (H3K27me3) showed that CUR decreased the enrichment of H3K27me3 at the Neurog1 promoter region as well as at the global level. Taken together, our present study provides evidence on the CpG demethylation ability of CUR on Neurog1 while activating its expression, suggesting a potential epigenetic modifying role for this phytochemical compound in human prostate cancer cells.


Crowdsourcing image annotation for nucleus detection and segmentation in computational pathology: evaluating experts, automated methods, and the crowd.
The development of tools in computational pathology to assist physicians and biomedical scientists in the diagnosis of disease requires access to high-quality annotated images for algorithm learning and evaluation. Generating high-quality expert-derived annotations is time-consuming and expensive. We explore the use of crowdsourcing for rapidly obtaining annotations for two core tasks in com- putational pathology: nucleus detection and nucleus segmentation. We designed and implemented crowdsourcing experiments using the CrowdFlower platform, which provides access to a large set of labor channel partners that accesses and manages millions of contributors worldwide. We obtained annotations from four types of annotators and compared concordance across these groups. We obtained: crowdsourced annotations for nucleus detection and segmentation on a total of 810 images; annotations using automated methods on 810 images; annotations from research fellows for detection and segmentation on 477 and 455 images, respectively; and expert pathologist-derived annotations for detection and segmentation on 80 and 63 images, respectively. For the crowdsourced annotations, we evaluated performance across a range of contributor skill levels (1, 2, or 3). The crowdsourced annotations (4,860 images in total) were completed in only a fraction of the time and cost required for obtaining annotations using traditional methods. For the nucleus detection task, the research fellow-derived annotations showed the strongest concordance with the expert pathologist- derived annotations (F-M =93.68%), followed by the crowd-sourced contributor levels 1,2, and 3 and the automated method, which showed relatively similar performance (F-M = 87.84%, 88.49%, 87.26%, and 86.99%, respectively). For the nucleus segmentation task, the crowdsourced contributor level 3-derived annotations, research fellow-derived annotations, and automated method showed the strongest concordance with the expert pathologist-derived annotations (F-M = 66.41%, 65.93%, and 65.36%, respectively), followed by the contributor levels 2 and 1 (60.89% and 60.87%, respectively). When the research fellows were used as a gold-standard for the segmentation task, all three con- tributor levels of the crowdsourced annotations significantly outperformed the automated method (F-M = 62.21%, 62.47%, and 65.15% vs. 51.92%). Aggregating multiple annotations from the crowd to obtain a consensus annotation resulted in the strongest performance for the crowd-sourced segmentation. For both detection and segmentation, crowd-sourced performance is strongest with small images (400 × 400 pixels) and degrades significantly with the use of larger images (600 × 600 and 800 × 800 pixels). We conclude that crowdsourcing to non-experts can be used for large-scale labeling microtasks in computational pathology and offers a new approach for the rapid generation of labeled images for algorithm development and evaluation.


Molecular dynamics simulations elucidate conformational selection and induced fit mechanisms in the binding of PD-1 and PD-L1.
Blockage of the interactions between immunologic checkpoint protein PD-1 and its ligand PD-L1 showed efficacy for cancer treatment. X-ray structures have captured static conformational snapshots of PD-1 and revealed that the CC' loop adopts an open conformation in the apo-protein but turns into a closed form and interacts with PD-L1 in the complex. This structural heterogeneity brings difficulties for structure-based drug discovery targeting PD-1. To gain insights into the role of the CC' loop in molecular recognition, we have undertaken a comparative study between the open and closed conformations in apo-PD-1 and the PD-1/PD-L1 complex using molecular dynamics simulations. Results show that the moderate stability of intramolecular hydrogen bonds between SER71 and THR120 allows the CC' loop to sample both the open and closed states in apo-PD-1. Binding of PD-L1 accelerates the open-to-closed switch and locks the loop in the closed state through four newly formed intermolecular hydrogen bonds. Thus, we suggest a complex binding mechanism between PD-1 and PD-L1 where both the conformational selection and induced fit theories play a role.


Factors Influencing Clinical After Effects of Post Orthognathic Surgery - An Observational Clinical Study.
For maintaining the occlusion, screws to anchor bones are needed to be used in transalveolar manner to get the intermaxillary fixation in participants with no preoperative orthodontic treatment or participants with loose or broken appliances. The present clinical trial was hence aimed to assess the postoperative complications following orthognathic surgical repair of skeletal malocclusion. Forty-two participants were divided into two groups (n = 22). In Group I, predrill was done to create the holes in transalveolar position before screw insertion. For Group II, self-cutting screws were used without the drills. The radiographs were then taken to assess the associated root injuries. To evaluate the effect of different steroid doses on the pain, nerve healing, and swelling, the participants were divided into three groups (n = 14). Plate removal and associated factors were also evaluated. Collected data were statistically analyzed. In Group where no predrill was done, no root injuries were seen. Considerably less facial edema was observed in Group II and III compared to control Group I. This difference was statistically significant with a P value of 0.2057. At 1 week, 3 months, and 6-month postoperatively in Group II and Group III, no significant difference was seen. No significant difference in the postoperative pain between the groups was seen (P = 0.85103). Neurosensory Visual Analog Score measurement revealed no significant difference between three groups at 6 months with the P value of 0.81821. The present study concludes that risk for the root injury is possessed by the screws that require predrill, whereas the self-drilling screws had no risk for root injury.


Release from prison--a high risk of death for former inmates.
The U.S. population of former prison inmates is large and growing. The period immediately after release may be challenging for former inmates and may involve substantial health risks. We studied the risk of death among former inmates soon after their release from Washington State prisons. We conducted a retrospective cohort study of all inmates released from the Washington State Department of Corrections from July 1999 through December 2003. Prison records were linked to the National Death Index. Data for comparison with Washington State residents were obtained from the Wide-ranging OnLine Data for Epidemiologic Research system of the Centers for Disease Control and Prevention. Mortality rates among former inmates were compared with those among other state residents with the use of indirect standardization and adjustment for age, sex, and race. Of 30,237 released inmates, 443 died during a mean follow-up period of 1.9 years. The overall mortality rate was 777 deaths per 100,000 person-years. The adjusted risk of death among former inmates was 3.5 times that among other state residents (95% confidence interval [CI], 3.2 to 3.8). During the first 2 weeks after release, the risk of death among former inmates was 12.7 (95% CI, 9.2 to 17.4) times that among other state residents, with a markedly elevated relative risk of death from drug overdose (129; 95% CI, 89 to 186). The leading causes of death among former inmates were drug overdose, cardiovascular disease, homicide, and suicide. Former prison inmates were at high risk for death after release from prison, particularly during the first 2 weeks. Interventions are necessary to reduce the risk of death after release from prison.


Complete Sequence and Organization of pFR260, the Bacillus thuringiensis INTA Fr7-4 Plasmid Harboring Insecticidal Genes.
We report the complete sequence and analysis of pFR260, a novel megaplasmid of 260,595 bp from the Bacillus thuringiensis strain INTA Fr7-4 isolated in Argentina. It carries 7 insecticidal genes: 3 cry8 copies previously reported, 2 vip1, and 2 vip2. Also, it carries a gene encoding a putative atypical Cry protein. These genes are arranged in a region of approximately 105 kbp in size with characteristics of a pathogenicity island with a potential coleopteran-specific insecticide profile. DNA strand composition asymmetry, as determined by GC skew analysis, and the presence of a Rep protein involved in the initiation of replication suggest a bidirectional theta mechanism of replication. In addition, many genes involved in conjugation and a CRISPR-Cas system were detected. The pFR260 sequence was deposited in GenBank under accession number KX258624.


Prognostic interaction between expression of p53 and estrogen receptor in patients with node-negative breast cancer: results from IBCSG Trials VIII and IX.
The prognostic significance of p53 protein expression in early breast cancer remains uncertain, with some but not all studies finding an association with poorer outcomes. Estrogen receptor (ER) expression is both a positive prognostic marker and predictive of response to endocrine therapies. The relationship between these biomarkers is unknown. We constructed tissue microarrays (TMAs) from available pathological material from 1113 patients participating in two randomized clinical trials comparing endocrine therapy alone versus chemo-endocrine therapy in node-negative breast cancer. Expression of p53 defined as >10% positive nuclei was analyzed together with prior immunohistochemical assays of ER performed at central pathological review of whole tumor sections. ER was present (i.e. >1% positive tumor cell nuclei) in 80.1% (880/1092). p53 expression was significantly more frequent when ER was absent, 125/212 (59%) than when ER was present, 171/880 (19%), p <0.0001. A significant qualitative interaction was observed such that p53 expression was associated with better disease-free survival (DFS) and overall survival (OS) among patients whose tumors did not express ER, but worse DFS and OS among patients whose tumors expressed ER. The interaction remained significant after allowance for pathologic variables, and treatment. Similar effects were seen when luminal and non-luminal intrinsic subtypes were compared. Interpretation of the prognostic significance of p53 expression requires knowledge of concurrent expression of ER. The reason for the interaction between p53 and ER is unknown but may reflect qualitatively different p53 mutations underlying the p53 expression in tumors with or without ER expression. Current Controlled Trials ACTRN12607000037404 (Trial VIII) and ACTRN12607000029493 (Trial IX).


Leisure activity associated with cognitive ability level, but not cognitive change.
Although activity participation is promoted as cognitively protective, critical questions of causality remain. In a cohort followed every 5 years from age 75 to 85 years, potential reciprocal associations between level and change in leisure activity participation and level and change in cognitive abilities were examined. Participants in the Glostrup 1914 Cohort, a longitudinal study of aging, completed standardized cognitive ability tests and reported their leisure activity participation (11 activities defined a leisure activity score) at ages 75, 80, and 85. Higher leisure activity was associated with higher cognitive ability (significant correlations ranged from 0.15 to 0.31, p < 0.05). Between ages 75 and 85, participation in leisure activities and cognitive ability declined significantly. Growth curve models, which provided latent variables for level of and 10-year change in both leisure activity and cognitive ability, confirmed the positive association between levels of leisure activity and cognitive ability (path coefficient = 0.36, p < 0.001); however, neither leisure activity level nor change in leisure activity were associated with cognitive change. Although a positive association between leisure activity and cognitive ability was reported-the likely precedents of this are discussed-there was no evidence that a higher level or maintenance of leisure activity was protective against cognitive decline across a 10-year follow-up.


Is the skillset obtained in surgical simulation transferable to the operating theatre?
Simulated surgical training offers a safe and accessible way of learning surgical procedures outside the operating room. Training programs have been developed using simulated laboratories to train surgical trainees to proficiency outside the operating room. Despite the global enthusiasm among educators to enhance training through simulation-based learning, it remains to be elucidated whether the skill set obtained is transferrable to the operating room. Using standardized search methods, the authors searched the Cochrane Central Register of Controlled Trials, PubMed, Embase, and Web-Based Knowledge, as well as the reference lists of relevant articles, and retrieved all published randomized controlled trials. Sixteen randomized controlled trials involving 309 participants were identified to be suitable for qualitative analysis using the Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines. The mean Consolidated Standards of Reporting Trials score was 16 (range, 12-22). The studies showed considerable clinical and methodologic diversity. Operative time improved consistently in all trials after training and was the only objective parameter measurable in the live setting. Studies that used the Objective Structured Assessment of Technical Skills as their primary outcome showed improved scores in 80% of trials, and studies that used performance indicators to assess participants all showed improved scores after simulation training in all of the trials, with 88% showing statistical significance. The current literature consistently demonstrates the positive impact of simulation on operative time and predefined performance scores. However, these reproducible measures alone are insufficient to demonstrate transferability of skills from the laboratory to the operating room. The authors advocate a multimodal assessment, including metrics, the Objective Structured Assessment of Technical Skills, and critical step completion. This may provide a more complete assessment of operative performance. Only then can it be concluded that simulation skills are transferable to the live operative setting.


Regional variations in home canning practices and the risk of foodborne botulism in the Republic of Georgia, 2003.
Foodborne botulism is a severe, paralytic illness caused by ingestion of preformed neurotoxins produced by Clostridium botulinum. In 2003, we conducted a population-based household survey of home canning practices to explore marked regional variations in botulism incidence in the Republic of Georgia (ROG). We designed a cluster sampling scheme and subdivided each of the 10 regions of the ROG into a variable number of strata. Households were selected from each stratum using a two-step cluster sampling methodology. We administered a questionnaire about home canning practices to household members responsible for food preparation. Using multivariate logistic regression analysis, we modeled high (eastern ROG) against low (western ROG) incidence areas. Overall, we surveyed 2,742 households nationwide. Home canning with a capping device hermetically sealing the lid covering the jar was practiced by 1,909 households (65.9%; 95% confidence interval [CI]: 59.8 to 72.1%). Canning was more prevalent in regions of low botulism incidence (34 versus 32%; P < 0.01). When compared with low-botulism areas, the following practices were associated with an increased risk in high-botulism areas: ≥ 6 months between canning vegetables and consuming them (adjusted odds ratio [aOR] = 2.1; 95% CI: 1.3 to 3.5) and adding any of the following ingredients to the jar at time of preparation: >1 tablespoon of salt per liter (aOR = 5.1; 95% CI: 1.2 to 22.6); vinegar (aOR = 2.2; 95% CI: 1.3 to 3.7), and greens (aOR = 5.6; 95% CI: 1.7 to 18.2). The following practices were associated with a decreased risk in high-botulism areas: >57 jars canned per household annually (aOR = 0.5; 95% CI: 0.3 to 0.9), covering or immersing vegetables in boiling water before placing them into the jar (aOR = 0.3 95% CI: 0.2 to 0.6), covering or immersing vegetables in boiling water after placing them into the jar (aOR = 0.4; 95% CI: 0.2 to 0.9), or adding garlic (aOR = 0.2; 95% CI: 0.1 to 0.5) or aspirin (aOR = 0.1; 95% CI: 0.1 to 0.2) to the jar at the time of preparation.


Design of Treatment Trials for Functional Gastrointestinal Disorders.
This article summarizes recent progress and regulatory guidance on design of trials to assess the efficacy of new therapies for functional gastrointestinal disorders (FGIDs). The double-masked, placebo-controlled, parallel-group design remains the accepted standard for evaluating treatment efficacy. A control group is essential, and a detailed description of the randomization process and concealed allocation method must be included in the study report. The control will most often be placebo, but for therapeutic procedures and for behavioral treatment trials, respectively, a sham procedure and control intervention with similar expectation of benefit, but lacking the treatment principle, are recommended. Investigators should be aware of, and attempt to minimize, expectancy effects (placebo, nocebo, precebo). The primary analysis should be based on the proportion of patients in each treatment arm who satisfy a treatment responder definition or a prespecified clinically meaningful change in a patient-reported outcome measure. Data analysis should use the intention-to-treat principle. Reporting of results should follow the Consolidated Standards for Reporting Trials guidelines and include secondary outcome measures to support or explain the primary outcome and an analysis of harms data. Trials should be registered in a public location before initiation and results should be published regardless of outcome.


Insulin sensitivity and body weight changes in young white carriers of the codon 64 amino acid polymorphism of the beta 3-adrenergic receptor gene.
Recently, a missense mutation replacing tryptophan with arginine at codon 64 of the beta 3-adrenergic receptor gene was shown to be associated with insulin resistance in nondiabetic subjects and to an earlier onset of NIDDM in Pima Indians. We studied whether the codon 64 amino acid polymorphism of the beta 3-adrenergic receptor gene in a cohort of young healthy Danes was associated with high birth weight, accelerated weight gain during childhood and adolescence, present obesity, or impaired insulin sensitivity. The protocol included 380 unrelated white subjects in whom insulin sensitivity and secretion were measured during a combined intravenous glucose and tolbutamide tolerance test. A number of biochemical and anthropometric characteristics were determined for each subject. The subjects were genotyped for the codon 64 polymorphism by applying polymerase chain reaction restriction fragment-length polymorphism screening with the use of endonuclease BstN1. The allelic frequency of the mutated allele was 7% (95% CI: 5-10%), and it was similar in obese and nonobese subjects. The beta 3-adrenergic receptor gene variant was not related to birth weight or weight gain during childhood or adolescence. In its heterozygous form, the gene variant was not associated with an altered insulin sensitivity index (SI) or other features of the insulin resistance syndrome (BMI, blood pressure, fasting serum lipid levels, or fasting serum fibrinolytic variables). Three homozygous carriers of the polymorphism were identified, and each had a significantly higher BMI (27.4 +/- 1.3 vs. 23.5 +/- 3.7 kg/m2 [mean +/- SD]; P = 0.032), lower SI [4.9 +/- 2.9 vs. 15.4 +/- 9.0 10(-5) x (min x pmol/l)-1; P = 0.013], and higher fasting serum C-peptide (730 +/- 155 vs. 471 +/- 158 pmol/l; P = 0.016) than the wild-type carriers. The homozygous carriers also had significantly higher levels of fasting serum triglyceride (P = 0.042) and serum LDL cholesterol (P = 0.013). When adjustments were made for age, sex, BMI, and VO2max in a multiple regression analysis, a significantly negative association was found between homozygosity for the codon 64 variant and the SI (P = 0.009). We conclude that in young healthy Danes, the homozygous form but not the heterozygous form of the codon 64 amino acid polymorphism of the beta 3-adrenergic receptor may be associated with obesity and, independent of BMI, with a low SI. Since only three homozygous carriers were identified among 380 subjects, the results must be interpreted with caution, and studies of larger population samples are needed.


Antioxidants modulate mitochondrial PKA and increase CREB binding to D-loop DNA of the mitochondrial genome in neurons.
The protein kinase A (PKA) and the cAMP response element (CRE) binding protein (CREB) signaling pathways mediate plasticity and prosurvival responses in neurons through their ability to regulate gene expression. The PKA-CREB signaling mechanism has been well characterized in terms of nuclear gene expression. We show that the PKA catalytic and regulatory subunits and CREB are localized to the mitochondrial matrix of neurons. Mitochondrial CRE sites were identified by using both serial analyses of chromatin occupancy and chromatin immunoprecipitation. Deferoxamine (DFO), an antioxidant and iron chelator known to inhibit oxidative stress-induced death, activated mitochondrial PKA and increased mitochondrial CREB phosphorylation (Ser-133). DFO increased CREB binding to CRE in the mitochondrial D-loop DNA and D-loop CRE-driven luciferase activity. In contrast, KT5720, a specific inhibitor of PKA, reduced DFO-mediated neuronal survival against oxidative stress induced by glutathione depletion. Neuronal survival by DFO may be, in part, mediated by the mitochondrial PKA-dependent pathway. These results suggest that the regulation of mitochondrial function via the mitochondrial PKA and CREB pathways may underlie some of the salutary effects of DFO in neurons.


Deletion of TnAbaR23 results in both expected and unexpected antibiogram changes in a multidrug-resistant Acinetobacter baumannii strain.
Since the 2006 discovery of the Acinetobacter baumannii strain AYE AbaR1 resistance island, similar elements have been reported in numerous members of this species. As AbaR1 is distantly related to Tn7, we have renamed it TnAbaR1. TnAbaR transposons are known to carry multiple antibiotic resistance- and efflux-associated genes, although none have been experimentally studied en bloc. We deleted the TnAbaR transposon in A. baumannii A424, which we have designated TnAbaR23, and characterized independent deletion mutants DCO163 and DCO174. The NotI pulsed-field gel electrophoresis (PFGE) profile of strain DCO174 was consistent with targeted deletion of TnAbaR23 alone, but strain DCO163 apparently harbored a second large genomic deletion. Nevertheless, "subtractive amplification" targeting 52 TnAbaR and/or resistance-associated loci yielded identical results for both mutants and highlighted genes lost relative to strain A424. PCR mapping and genome sequencing revealed the entire 48.3-kb sequence of TnAbaR23. Consistent with TnAbaR23 carrying two copies of sul1, both mutants exhibited markedly increased susceptibility to sulfamethoxazole. In contrast, loss of tetAR(A) resulted in only a minor and variable increase in tetracycline susceptibility. Despite not exhibiting a growth handicap, strain DCO163 was more susceptible than strain DCO174 to 9 of 10 antibiotics associated with mutant-to-mutant variation in susceptibility, suggesting impairment of an undefined resistance-associated function. Remarkably, despite all three strains sharing identical gyrA and parC sequences, the ciprofloxacin MIC of DCO174 was >8-fold that of DCO163 and A424, suggesting a possible paradoxical role for TnAbaR23 in promoting sensitivity to ciprofloxacin. This study highlights the importance of experimental scrutiny and challenges the assumption that resistance phenotypes can reliably be predicted from genotypes alone.


Efficient directional excitation of surface plasmons by a single-element nanoantenna.
Directional light scattering is important in basic research and real applications. This area has been successfully downscaled to wavelength and subwavelength scales with the development of optical antennas, especially single-element nanoantennas. Here, by adding an auxiliary resonant structure to a single-element plasmonic nanoantenna, we show that the highly efficient lowest-order antenna mode can be effectively transferred into inactive higher-order modes. On the basis of this mode conversion, scattered optical fields can be well manipulated by utilizing the interference between different antenna modes. Both broadband directional excitation of surface plasmon polaritons (SPPs) and inversion of SPP launching direction at different wavelengths are experimentally demonstrated as typical examples. The proposed strategy based on mode conversion and mode interference provides new opportunities for the design of nanoscale optical devices, especially directional nanoantennas.


Synovial explant inflammatory mediator production corresponds to rheumatoid arthritis imaging hallmarks: a cross-sectional study.
Despite the widespread use of magnetic resonance imaging (MRI) and Doppler ultrasound for the detection of rheumatoid arthritis (RA) disease activity, little is known regarding the association of imaging-detected activity and synovial pathology. The purpose of this study was to compare site-specific release of inflammatory mediators and evaluate the corresponding anatomical sites by examining colour Doppler ultrasound (CDUS) and MRI scans. RA patients were evaluated on the basis of CDUS and 3-T MRI scans and subsequently underwent synovectomy using a needle arthroscopic procedure of the hand joints. The synovial tissue specimens were incubated for 72 hours, and spontaneous release of monocyte chemoattractant protein 1 (MCP-1), interleukin 6 (IL-6), macrophage inflammatory protein 1β (MIP-1β) and IL-8 was measured by performing multiplex immunoassays. Bone marrow oedema (BME), synovitis and erosion scores were estimated on the basis of the rheumatoid arthritis magnetic resonance imaging score (RAMRIS). Mixed models were used for the statistical analyses. Parsimony was achieved by omitting covariates with P > 0.1 from the statistical model. Tissue samples from 58 synovial sites were obtained from 25 patients. MCP-1 was associated with CDUS activity (P = 0.009, approximate Spearman's ρ = 0.41), RAMRIS BME score (P = 0.01, approximate Spearman's ρ = 0.42) and RAMRIS erosion score (P = 0.03, approximate Spearman's ρ = 0.31). IL-6 was associated with RAMRIS synovitis score (P = 0.04, approximate Spearman's ρ = 0.50), BME score (P = 0.04, approximate Spearman's ρ = 0.31) and RAMRIS erosion score (P = 0.03, approximate Spearman's ρ = 0.35). MIP-1β was associated with CDUS activity (P = 0.02, approximate Spearman's ρ = 0.38) and RAMRIS synovitis scores (P = 0.02, approximate Spearman's ρ = 0.63). IL-8 associations with imaging outcome measures did not reach statistical significance. The association between imaging activity and synovial inflammatory mediators underscores the high sensitivity of CDUS and MRI in the evaluation of RA disease activity. The associations found in our present study have different implications for synovial mediator releases and corresponding imaging signs. For example, MCP-1 and IL-6 were associated with both general inflammation and bone destruction, in contrast to MIP-1β, which was involved solely in general synovitis. The lack of association of IL-8 with synovitis was likely underestimated because of a large proportion of samples above assay detection limits among the patients with the highest synovitis scores.


Gain and losses in THz quantum cascade laser with metal-metal waveguide.
Coupling of broadband terahertz pulses into metal-metal terahertz quantum cascade lasers is presented. Mode matched terahertz transients are generated on the quantum cascade laser facet of subwavelength dimension. This method provides a full overlap of optical mode and active laser medium. A longitudinal optical-phonon depletion based active region design is investigated in a coupled cavity configuration. Modulation experiments reveal spectral gain and (broadband) losses. The observed gain shows high dynamic behavior when switching from loss to gain around threshold and is clamped at total laser losses.


Resource heterogeneity, diet shifts and intra-cohort competition: effects on size divergence in YOY fish.
Most organisms exhibit a substantial size variation among individuals due to individual differences in experienced biotic and abiotic environmental conditions and because individuals undergo growth and development during most of their life time. One important issue in this context is how size variation within cohorts may develop over time. Here, we tested the hypothesis, in gape-limited animals such as fish, that size divergence among individuals within a cohort depends on the opportunity to undergo size-dependent diet shifts, by allowing initially larger individuals to make an early diet shift when the first resource becomes limiting. We used young-of-the-year perch (Perca fluviatilis) as our study organism. Competitive intensity and the opportunity to undergo a diet shift from zooplankton to macroinvertebrates affected both mean growth rates and the extent to which inter-individual variation in growth was manifested. As predicted, increased competition combined with the presence of both zooplankton and benthic macroinvertebrates increased the degree of size variation. However, size divergence was also observed among individuals when only the initial resource, zooplankton, was available. We argue that only non-exploitative interactions, such as dominance structures and social interactions, could have caused this latter pattern, as exploitative competition is expected to lead to size convergence due to the superior competitive ability of smaller individuals. Our results suggest that diet shifts are not a prerequisite for size divergence in animal cohorts, and that dominance and social interactions may have similar effects on size variation within cohorts. Finally, development of size variation is suggested to have strong implications for overall cohort performance.


A role for seed storage proteins in Arabidopsis seed longevity.
Proteomics approaches have been a useful tool for determining the biological roles and functions of individual proteins and identifying the molecular mechanisms that govern seed germination, vigour and viability in response to ageing. In this work the dry seed proteome of four Arabidopsis thaliana genotypes, that carry introgression fragments at the position of seed longevity quantitative trait loci and as a result display different levels of seed longevity, was investigated. Seeds at two physiological states, after-ripened seeds that had the full germination ability and aged (stored) seeds of which the germination ability was severely reduced, were compared. Aged dry seed proteomes were markedly different from the after-ripened and reflected the seed longevity level of the four genotypes, despite the fact that dry seeds are metabolically quiescent. Results confirmed the role of antioxidant systems, notably vitamin E, and indicated that protection and maintenance of the translation machinery and energy pathways are essential for seed longevity. Moreover, a new role for seed storage proteins (SSPs) was identified in dry seeds during ageing. Cruciferins (CRUs) are the most abundant SSPs in Arabidopsis and seeds of a triple mutant for three CRU isoforms (crua crub cruc) were more sensitive to artificial ageing and their seed proteins were highly oxidized compared with wild-type seeds. These results confirm that oxidation is involved in seed deterioration and that SSPs buffer the seed from oxidative stress, thus protecting important proteins required for seed germination and seedling formation.


The definition and diagnosis of asthma.
The diagnosis of asthma depends on what we mean by the word. Its definition continues to be controversial because there is no single genetic or environmental cause. Addressed from a descriptive point of view, the disease components include airway inflammation, symptoms, variable airflow limitation and chronic airflow limitation. The essentialist definition conveys the message that asthma is a separate disease entity, fails to identify a primary defining characteristic which separates it from other diseases and is long winded. These disadvantages are overcome by the nominalist definition of asthma in which the word 'asthma'refers to an abnormality of airway function, specifically to wide variations in airflow limitation over short periods of time. In patients with asthma the other components of airway disease need to be considered. These have separate nominalist definitions and especially include different types of bronchitis for airway inflammation and chronic obstructive pulmonary disease for chronic airflow limitation. What is present will vary between and within patients. The accurate diagnosis of asthma and of other components of disease all require objective measurements. Currently spirometry and airway responsiveness should be available to the general practitioner, who sees milder disease, and additional quantitative sputum cell counts in specialist practice, where moderate to severe disease is more prevalent. Such measurements characterize the patient, identify heterogeneity and allow treatment to be personalized.


Generalized anetoderma: An unusual manifestation of secondary syphilis treated with injection penicillin.
Anetoderma also called macular atrophy is a rare, benign disorder characterized microscopically by the pan-dermal loss of elastic fibers in the dermis and presenting clinically as circumscribed, skin-colored or gray-white atrophic macules and/or patches on the trunk and/or extremities. Lesions are described as having a "sac-like" appearance, since they bulge or herniate upon palpation. It is a rare benign condition of diverse etiology; whose characteristic is the diminution or absence of the dermal elastic fibers. Anetoderma is divided into primary (idiopathic) and secondary anetoderma, with the former occurring in areas of previously normal skin and the latter developing in areas of prior skin pathology. Both may occur in association with underlying systemic conditions and warrant evaluation for associated disorders. There are no effective treatment options for anetoderma at present. We report here an unusual case of generalized anetoderma occurring in association with secondary syphilis treated with injection benzathine penicillin.


Noise Tailoring in Memristive Filaments.
In this study, the possibilities of noise tailoring in filamentary resistive switching memory devices are investigated. To this end, the resistance and frequency scaling of the low-frequency 1/f-type noise properties are studied in representative mainstream material systems. It is shown that the overall noise floor is tailorable by the proper material choice, as demonstrated by the order-of-magnitude smaller noise levels in Ta2O5 and Nb2O5 transition-metal oxide memristors compared to Ag-based devices. Furthermore, the variation of the resistance states allows orders-of-magnitude tuning of the relative noise level in all of these material systems. This behavior is analyzed in the framework of a point-contact noise model highlighting the possibility for the disorder-induced suppression of the noise contribution arising from remote fluctuators. These findings promote the design of multipurpose resistive switching units, which can simultaneously serve as analog-tunable memory elements and tunable noise sources in probabilistic computing machines.


Caring and Uncaring Encounters between Assistant Nurses and Immigrants with Dementia Symptoms in Two Group Homes in Sweden-an Observational Study.
The total number of people with dementia symptoms is expected to double every 20 years and there will also be an increase in the number of older immigrants in several countries. There are considerable deficiencies in the present knowledge of how to conduct well-functioning health care for immigrants with dementia symptoms. The aim of this study was to explore caring and uncaring encounters between assistant nurses and immigrants in two group homes for persons with dementia symptoms in Sweden: a Finnish-speaking as well as a Swedish-speaking context. In addition, this study aims to describe how caring and uncaring encounters are manifested in these two contexts according to Halldórsdóttir's theory of "Caring and Uncaring encounters". Descriptive field notes from 30 separate observations were analyzed using qualitative deductive content analysis. The main category "caring encounters" focused on reaching out to initiate connection through communication, removing masks of anonymity by acknowledging the unique person, acknowledgment of connection by being personal. Reaching a level of truthfulness by being present and showing respect, raising the level of solidarity by equality and true negotiation of care, based on the residents' needs. The main category, uncaring encounters, focused on disinterest in and insensitivity towards the other, coldness in the connection and lack of humanity in care situations. The observations showed that caring encounters occurred more in the Finnish-speaking context and uncaring encounters more often in the Swedish context. Encounters could be caring, uncaring, and carried out using a person-centered approach. Communication and relationships could be facilitated using the same language but also through learning to interpret residents' needs and desires.


"A bit more truthful": the validity of adolescent sexual behaviour data collected in rural northern Tanzania using five methods.
To assess the validity of sexual behaviour data collected from African adolescents using five methods. 9280 Tanzanian adolescents participated in a biological marker and face to face questionnaire survey and 6079 in an assisted self-completion questionnaire survey; 74 participated in in-depth interviews and 56 person weeks of participant observation were conducted. 38% of males and 59% of females reporting sexual activity did so in only one of the two 1998 questionnaires. Only 58% of males and 29% of females with biological markers consistently reported sexual activity in both questionnaires. Nine of 11 (82%) in-depth interview respondents who had had biological markers provided an invalid series of responses about sex in the survey and in-depth interview series. Only one of six female in-depth interview respondents with an STI reported sex in any of the four surveys, but five reported it in the in-depth interviews. In this low prevalence population, biological markers on their own revealed that a few adolescents had had sex, but in combination with in-depth interviews they may be useful in identifying risk factors for STIs. Self-reported sexual behaviour data were fraught with inconsistencies. In-depth interviews seem to be more effective than assisted self-completion questionnaires and face to face questionnaires in promoting honest responses among females with STIs. Participant observation was the most useful method for understanding the nature, complexity, and extent of sexual behaviour.


Family care conferences in long-term care: Exploring content and processes in end-of-life communication.
ABSTRACTObjective:End-of-life (EoL) communication in long-term care (LTC) homes is often inadequate and delayed, leaving residents dying with unknown preferences or goals of care. Poor communication with staff contributes to families feeling unprepared, distressed, and dissatisfied with care. Family care conferences (FCCs) aim to increase structured systematic communication around goals and plans for the end of life. As part of the Strengthening a Palliative Approach to Care (SPA-LTC) project, FCCs were implemented in four LTC sites in Ontario, Canada. The purpose of this substudy was to examine FCC content and such guiding processes as documentation and multidisciplinary staff participation. A total of 24 FCCs were held for residents with a Palliative Performance Scale score of 40% (nearing death). Data were collected from conference forms (i.e., Family Questionnaires, Care Plan Conference Summaries), site-specific electronic chart documents, and fieldnotes. Directed content analysis of data was informed by the Canadian Hospice Palliative Care Association's Square of Care Model, which describes eight domains of care: disease management, physical, psychological, social, practical, spiritual, EoL, and loss/bereavement. The FCCs addressed an average of 71% of the content domains, with physical and EoL care addressed most frequently and loss/bereavement addressed the least. Two goals and five interventions were documented and planned on average per FCC. Examination of the processes supporting EoL communication found: (1) advantages to using FCC forms versus electronic charts; and (2) high levels of multidisciplinary participation overall but limited participation of personal support workers (PSWs) and physicians. Communication around the end of life in LTC can be supported through the use of FCCs. Description of content and FCC processes provides guidance to persons implementing FCCs. Recommendations for tailoring conferences to optimize communication include use of specific conference forms, increased bereavement discussion, and further engagement of PSWs and physicians.


Global surgical package reimbursement and the acute care surgeon: a threat to optimal care.
Emergency surgery patients risk greater mortality and morbidity than elective patients undergoing the same procedure. However, the differential effort required for the care of emergency surgical patients is poorly defined. This study sought to characterize costs and outcomes of elective versus emergent right hemicolectomy. 2006 Nationwide Inpatient Sample data were used to compare mortality, total charges, and length of stay (LOS) in patients undergoing emergency versus elective right hemicolectomy (International Classification of Diseases-9th procedure code 45.73). Mann-Whitney tests examined total costs and LOS; multivariable regression modeled inhospital mortality controlling for age, gender, insurance status, and comorbidities. Among 8,074,825 inpatient admissions, 7,767 emergent and 10,399 elective right hemicolectomies were identified. Emergent patients were similar in age (66.9 years vs. 67.6 years; p=0.129), more likely women (43.7% vs. 42.3%; p=0.048) and had greater comorbidity (Charlson score 3.37 vs. 3.01; p<0.001) compared with elective patients. Emergent patient LOS was approximately double that of elective patients (13 days vs. 7 days; p<0.001). Mean total charges were $78,118 for emergent versus $39,265 elective patients (p<0.001). Emergent patients had greater odds of inhospital mortality (odds ratio, 5.86; 95% confidence intervals, 4.80-7.14). Emergent right hemicolectomy patients have greater comorbidity, experience longer stays, accrue twice the charges, and have higher mortality risk. This reflects a heightened effort required to care for emergent patients unrecognized by Global Surgical Package reimbursement. The concentration of emergency surgical patients in acute care surgery services necessitates accounting for the additional effort associated with these predictable risks to ensure optimal care.


The association between weekly work hours, crew familiarity, and occupational injury and illness in emergency medical services workers.
Emergency Medical Services (EMS) workers are shift workers in a high-risk, uncontrolled occupational environment. EMS-worker fatigue has been associated with self-reported injury, but the influence of extended weekly work hours is unknown. A retrospective cohort study was designed using historical shift schedules and occupational injury and illness reports. Using multilevel models, we examined the association between weekly work hours, crew familiarity, and injury or illness. In total, 966,082 shifts and 950 reports across 14 EMS agencies were obtained over a 1-3 year period. Weekly work hours were not associated with occupational injury or illness. Schedule characteristics that yield decreased exposure to occupational hazards, such as part-time work and night work, conferred reduced risk of injury or illness. Extended weekly work hours were not associated with occupational injury or illness. Future work should focus on transient exposures and agency-level characteristics that may contribute to adverse work events.


Assessing the effects of high methionine intake on DNA methylation.
Methylation of DNA occurs at cytosines within CpG (cytosine-guanine) dinucleotides and is 1 of several epigenetic mechanisms that serve to establish and maintain tissue-specific patterns of gene expression. The methyl groups transferred in mammalian DNA methylation reactions are ultimately derived from methionine. High dietary methionine intake might therefore be expected to increase DNA methylation. Because of the circular nature of the methionine cycle, however, methionine excess may actually impair DNA methylation by inhibiting remethylation of homocysteine. Although little is known regarding the effect of dietary methionine supplementation on mammalian DNA methylation, the available data suggest that methionine supplementation can induce hypermethylation of DNA in specific genomic regions. Because locus-specific DNA hypomethylation is implicated in the etiology of various cancers and developmental syndromes, clinical trials of "promethylation" dietary supplements are already under way. However, aberrant hypermethylation of DNA could be deleterious. It is therefore important to determine whether dietary supplementation with methionine can effectively support therapeutic maintenance of DNA methylation without causing excessive and potentially adverse locus-specific hypermethylation. In the viable yellow agouti (Avy) mouse, maternal diet affects the coat color distribution of offspring by perturbing the establishment of methylation at the Avy metastable epiallele. Hence, the Avy mouse can be employed as a sensitive epigenetic biosensor to assess the effects of dietary methionine supplementation on locus-specific DNA methylation. Recent developments in epigenomic approaches that survey locus-specific DNA methylation on a genome-wide scale offer broader opportunities to assess the effects of high methionine intake on mammalian epigenomes.


Ethanol consumption affects stress response and insulin binding in tissues of rats.
The mechanisms of harmful effect of excessive chronic ethanol intake on cardiovascular and neuronal systems, metabolic processes and protective effects of low ethanol intake on cerebrovascular and cardiovascular disturbances are not yet known. The aim of present study was to investigate the effect of short-term and long-term ethanol consumption on food intake, plasma levels of corticosterone (CS), glucose (G) and insulin (INS) in intact rats and in animals exposed to immobilization stress (IMO) or restrain stress (RESTR). The insulin binding to specific membrane receptors in adipocytes, muscle and liver was also determined. Adult male rats were fed liquid diet without and with ethanol (5% per weight) for 9-12 days (short-term intake) and at the end the animals were exposed to acute IMO stress. The second group of rats was fed solid diet and without or with ethanol in drinking water (6% per volume) for 52 days (long-term ethanol intake). A part of these animals was exposed to repeated restrain stress during 42 days. The groups of pair fed rats with the same food intake as in ethanol diet fed animals were in both experiments. The food intake, plasma glucose, insulin and corticoster-one levelswere determined. Plasma cell membranes were isolated from adipose, liver and skeletal muscle tissues and insulin binding to specific receptors was determined. Decreased food consumption was observed after ethanol intake. Increased plasma G and CS were noted in rats fed ethanol diet for a short time. Plasma insulin levels were not affected by ethanol intake. Exposure to IMO stress resulted in increase of plasma G levels in controls and pair fed groups. A higher increase of CS plasma levels after IMO stress was noted in rats with ethanol intake for a short-time, however, no changes of plasma CS were noted after repeated exposure to restrain stress. Plasma levels of insulin were decreased after IMO and restrain stresses, while in rats fed ethanol diet for a short-time insulin decrease was deeper as compared to controls. The binding capacity of insulin receptors in adipose tissue was elevated in rats fed ethanol diet and deep decrease of insulin binding was noted in rats exposed to stress. In liver insulin binding was elevated after short-term ethanol intake and stress exposure resulted in decrease of insulin binding in ethanol fed rats. The binding capacity of insulin receptors in skeletal muscle was not changed in rats fed ethanol diet. The results showed 1. differences in short term and long term ethanol intake on basal glucose, insulin and corticosterone levels, 2. effects of ethanol intake on changes of insulin plasma levels and insulin binding in adipose and liver tissues after exposure to stress, 3. effects of short term ethanol intake on the response of plasma hormones to immobilization stress.


Systemically Administered Plant Recombinant Holo-Intrinsic Factor Targets the Liver and is not Affected by Endogenous B12 levels.
Precision targeting imaging agents and/or treatment agents to select cells or organs in the body remains a significant need and is an area of intense research. It has been hypothesized that the vitamin B12 (B12) dietary pathway, or components thereof, may be exploitable in this area. The question of whether gastric Intrinsic factor (IF), critical for B12 absorption in the GI tract via the cubilin receptor, could be used as a targeting moiety for the cubilin receptor systemically, has not been investigated. Cubilin is the only known receptor for holo-IF and is found primarily in the kidney and ear (outside of the ileum of the GI) offering significant scope for specific targeting. We utilized plant derived human gastric IF in fluorescent cell and PET based in vivo imaging and biodistribution studies and demonstrated that plant derived IF primarily targets the liver, likely a consequence of the unique glycosylation profile of the IF, and is not affected by endogenous B12 levels.


Duodenal intraepithelial lymphocytosis with normal villous architecture in pediatric patients: Mayo Clinic experience, 2000-2009.
Small bowel intraepithelial lymphocytosis (IELs) with normal villous architecture is a relatively common finding, often of uncertain significance. The aims of our study were to determine the prevalence of this finding, its clinical associations, its specificity for celiac disease (CD), and whether histologic clues exist that increase the specificity for CD in this setting, all in the pediatric population. The Mayo Clinic electronic pathology database was searched between January 1, 2000 and December 31, 2009 for patients younger than 18 years who had the terms "normal villi" and "increased intraepithelial lymphocytes" in their small bowel biopsy reports. All of the slides were reviewed to confirm the histologic findings. Demographic, serologic, pathologic, and clinical informations were obtained. Among 1290 duodenal biopsies obtained from children during the years 2000 and 2009, 56 (4.3%) were noted to have "normal villous architecture with increased intraepithelial lymphocytes." In the 54 patients not known to have CD before biopsy, 48 (89%) had serologic testing for CD. Of these 48 patients, 9 were labeled with CD, although only 5 of 9 met the definite criteria for the diagnosis, based on a combination of serologic markers, human leukocyte antigen haplotyping, and response to gluten-free diet. No clinical features pointed to a diagnosis of CD. There was no correlation between CD and number of IELs, but patients with newly diagnosed CD were more likely to have a tip-heavy lymphocyte distribution. Other diagnoses made during the study period and in follow-up were inflammatory bowel disease (5), Helicobacter pylori infection (3), medication-related injury (10), and systemic autoimmune disorders (2). Increased IELs with normal villous architecture in small bowel biopsies are clinically important in children, and are associated with a new definite diagnosis of CD in 9% of pediatric patients. Even at this low sensitivity, clinical work-up for CD is mandated in all of the patients with this finding.


CT depiction of pulmonary emboli: display window settings.
To compare computed tomographic (CT) window settings selected by radiologists with those determined by using two alternative approaches for depiction of pulmonary emboli (PE). Institutional review board approval was obtained; informed consent was not required. This study was compliant with the Health Insurance Portability and Accountability Act. Twenty-five clinical chest CT studies were obtained with a standardized PE protocol and retrospectively evaluated by five chest and two body CT radiologists. Of these studies, 13 were positive for PE, and 12 were negative. At the main pulmonary artery (PA), mean attenuations (MPA) and standard deviations (SDPA) were measured. Initially, images were displayed with a standard mediastinal window setting (window width, W = 400 HU; window center, C = 30 HU), and each observer adjusted the setting to a personally preferred setting (eg, "personal") for PE detection. Images displayed at this setting were compared in a side-by-side fashion with the "modified" (W = MPA + 2 . SDPA, C = W/2) and "double-half" (W = 2 . MPA, C = MPA/2) window setting. Each observer rated images from 1 (ie, most preferred) to 3 (ie, least preferred). For quantitative analysis, window width and center value of each setting were divided by corresponding MPA to compute a width ratio and a center ratio. Window settings and ratings were compared with repeated-measures analysis of variance, paired t tests, and Wilcoxon signed-rank tests. Ratings for all three types of window settings were significantly different (P < .001). Observers preferred their personal settings the most and the modified settings the least. Mean ratios for the seven observers were 1.68 +/- 0.20 for window width and 0.47 +/- 0.08 for window center. Window width ratios for all settings were significantly different from each other (P < .001). Window center ratios were significantly higher for the modified setting than for the double-half setting (P = .013). Values for mean PA attenuation were correlated with window width ratios for six (86%) observers (mean r2 value = 0.29 +/- 0.19, P < or = .03) and with window center ratios for four (57%) observers (mean r2 value = 0.16 +/- 0.14, P < or = .02), thus indicating a trend of setting window width and window center higher when contrast enhancement is lower and vice versa. On average, observers selected CT window settings for PE detection at a window width of slightly less than twice the mean PA attenuation and at a window center of about half the mean PA attenuation. Observers tended to use larger window widths and centers as the degree of PA enhancement was lower.


Mechanical tension increases CCN2/CTGF expression and proliferation in gingival fibroblasts via a TGFβ-dependent mechanism.
Unlike skin, oral gingival do not scar in response to tissue injury. Fibroblasts, the cell type responsible for connective tissue repair and scarring, are exposed to mechanical tension during normal and pathological conditions including wound healing and fibrogenesis. Understanding how human gingival fibroblasts respond to mechanical tension is likely to yield valuable insights not only into gingival function but also into the molecular basis of scarless repair. CCN2/connective tissue growth factor is potently induced in fibroblasts during tissue repair and fibrogenesis. We subjected gingival fibroblasts to cyclical strain (up to 72 hours) using the Flexercell system and showed that CCN2 mRNA and protein was induced by strain. Strain caused the rapid activation of latent TGFβ, in a fashion that was reduced by blebbistatin and FAK/src inhibition, and the induction of endothelin (ET-1) mRNA and protein expression. Strain did not cause induction of α-smooth muscle actin or collagen type I mRNAs (proteins promoting scarring); but induced a cohort of pro-proliferative mRNAs and cell proliferation. Compared to dermal fibroblasts, gingival fibroblasts showed reduced ability to respond to TGFβ by inducing fibrogenic mRNAs; addition of ET-1 rescued this phenotype. Pharmacological inhibition of the TGFβ type I (ALK5) receptor, the endothelin A/B receptors and FAK/src significantly reduced the induction of CCN2 and pro-proliferative mRNAs and cell proliferation. Controlling TGFβ, ET-1 and FAK/src activity may be useful in controlling responses to mechanical strain in the gingiva and may be of value in controlling fibroproliferative conditions such as gingival hyperplasia; controlling ET-1 may be of benefit in controlling scarring in response to injury in the skin.


The contribution of hospital nursing leadership styles to 30-day patient mortality.
Nursing work environment characteristics, in particular nurse and physician staffing, have been linked to patient outcomes (adverse events and patient mortality). Researchers have stressed the need for nursing leadership to advance change in healthcare organizations to create safer practice environments for patients. The relationship between styles of nursing leadership in hospitals and patient outcomes has not been well examined. The purpose of this study was to examine the contribution of hospital nursing leadership styles to 30-day mortality after controlling for patient demographics, comorbidities, and hospital factors. Ninety acute care hospitals in Alberta, Canada, were categorized into five styles of nursing leadership: high resonant, moderately resonant, mixed, moderately dissonant, and high dissonant. In the secondary analysis, existing data from three sources (nurses, patients, and institutions) were used to test a hypothesis that the styles of nursing leadership at the hospital level contribute to patient mortality rates. Thirty-day mortality was 7.8% in the study sample of 21,570 medical patients; rates varied across hospital categories: high resonant (5.2%), moderately resonant (7.4%), mixed (8.1%), moderately dissonant (8.8%), and high dissonant (4.3%). After controlling for patient demographics, comorbidities, and institutional and hospital nursing characteristics, nursing leadership styles explained 5.1% of 72.2% of total variance in mortality across hospitals, and high-resonant leadership was related significantly to lower mortality. Hospital nursing leadership styles may contribute to 30-day mortality of patients. This relationship may be moderated by homogeneity of leadership styles, clarity of communication among leaders and healthcare providers, and work environment characteristics.


Electrical stimulation to accelerate wound healing.
There are several applications of electrical stimulation described in medical literature to accelerate wound healing and improve cutaneous perfusion. This is a simple technique that could be incorporated as an adjunctive therapy in plastic surgery. The objective of this review was to evaluate the results of randomized clinical trials that use electrical stimulation for wound healing. We identified 21 randomized clinical trials that used electrical stimulation for wound healing. We did not include five studies with treatment groups with less than eight subjects. Electrical stimulation was associated with faster wound area reduction or a higher proportion of wounds that healed in 14 out of 16 wound randomized clinical trials. The type of electrical stimulation, waveform, and duration of therapy vary in the literature. Electrical stimulation has been shown to accelerate wound healing and increase cutaneous perfusion in human studies. Electrical stimulation is an adjunctive therapy that is underutilized in plastic surgery and could improve flap and graft survival, accelerate postoperative recovery, and decrease necrosis following foot reconstruction.


Synthesized Population Databases: A US Geospatial Database for Agent-Based Models.
Agent-based models simulate large-scale social systems. They assign behaviors and activities to "agents" (individuals) within the population being modeled and then allow the agents to interact with the environment and each other in complex simulations. Agent-based models are frequently used to simulate infectious disease outbreaks, among other uses.RTI used and extended an iterative proportional fitting method to generate a synthesized, geospatially explicit, human agent database that represents the US population in the 50 states and the District of Columbia in the year 2000. Each agent is assigned to a household; other agents make up the household occupants.For this database, RTI developed the methods for generating synthesized households and personsassigning agents to schools and workplaces so that complex interactions among agents as they go about their daily activities can be taken into accountgenerating synthesized human agents who occupy group quarters (military bases, college dormitories, prisons, nursing homes).In this report, we describe both the methods used to generate the synthesized population database and the final data structure and data content of the database. This information will provide researchers with the information they need to use the database in developing agent-based models.Portions of the synthesized agent database are available to any user upon request. RTI will extract a portion (a county, region, or state) of the database for users who wish to use this database in their own agent-based models.


A conserved region of engrailed, shared among all en-, gsc-, Nk1-, Nk2- and msh-class homeoproteins, mediates active transcriptional repression in vivo.
The engrailed homeoprotein is a dominantly acting or 'active' transcriptional repressor both in cultured cells and in vivo. When retargeted via a homeodomain swap to the endogenous fushi tarazu gene (ftz), it actively represses it, resulting in a ftz mutant phenocopy. We have mapped functional regions of engrailed using this in vivo repression assay. In addition to a region containing an active repression domain identified in cell culture assays (K. Han and J. L. Manley (1993) EMBO J. 12, 2723-2733), we find that two evolutionarily conserved regions contribute to activity. The one of these that does not flank the HD is particularly crucial to repression activity in vivo. We find that this domain is present not only in all engrailed-class homeoproteins but also in all known members of several other classes, including goosecoid, Nk1, Nk2 and msh. Thus engrailed's active repression function in vivo is dependent on a highly conserved interaction that was established early in the evolution of the homeobox gene superfamily. We further show using rescue transgenes that the widely conserved in vivo repression domain is required for the normal function of engrailed in the embryo.


A new subhaplogroup of native American Y-Chromosomes from the Andes.
The human Y chromosome contains highly informative markers for making historical inferences about the pre-Columbian peopling of Americas. However, the scarcity of these markers has limited its use in the inference of shared ancestry and past migrations relevant to the origin of the culturally and biologically diverse Native Americans. To identify new single nucleotide polymorphisms (SNPs) and increase the phylogenetic resolution of the major haplogroup Q found in the Americas, we have performed a search for new polymorphisms based on sequencing divergent Y chromosomes identified by microsatellite haplotype analysis. Using this approach, a new Y-SNP (SA01) has been identified in the Andean populations of South America, allowing for the detection of a new sublineage of Q1a3a. This sublineage displays a less complex phylogeographic network of associated microsatellites and more restricted geographic occurrence, and is given the designation Q1a3a4. This result indicates that our approach can be successfully used to identify sublineages of interest in a specific region that allow the investigation of particular histories of human populations.


Preliminary results of a new antibiotic susceptibility test against biofilm installation in device-associated infections: the Antibiofilmogram®.
Biofilms are complex communities of microorganisms embedded in an extracellular matrix and adherent to a surface. The development was described as a four-stage process leading to the formation of a mature biofilm which was resistant to immune system and antibiotic actions. In bone and joint infections (BJIs), the formation of biofilms is a leading cause of treatment failure. Here we study the capacity of 11 antibiotics commonly used in the treatment of BJIs to inhibit the biofilm formation on 29 clinical Staphylococcus aureus isolates by a new test called Antibiofilmogram(®) The minimal inhibitory concentration (MIC) and biofilm MIC (bMIC) were determined in vitro and showed similar values for clindamycin, fusidic acid, linezolid and rifampin. Reversely, daptomycin, fosfomycin, gentamicin and ofloxacin showed a bMIC distribution different from MIC with bMIC above breakpoint. Finally, cloxacillin, teicoplanin and vancomycin revealed an intermediate bMIC distribution with a strain-dependent pattern. A murine in vivo model of catheter-associated S. aureus infection was made and showed a significant reduction, but not total prevention, of catheter colonization with cloxacillin at bMIC, and no or limited reduction with cloxacillin at MIC. Antibiofilmogram(®) could be of great interest after surgical operations on contaminated prostheses and after bacteremia in order to prevent the colonization of the device.


The effects of epidural anesthesia on the Doppler velocimetry of umbilical and uterine arteries in normal term labor.
To study the possible effects of epidural anesthesia on the Doppler velocimetry of the fetal umbilical and maternal uterine arteries, 15 normal term parturients were examined during active labor. Each Doppler study, conducted over 90 minutes after a 500-mL intravenous volume pre-load, was divided into three phases: 30 minutes of pre-anesthetic control, 30 minutes during epidural catheter placement and dosing, and 30 minutes after establishing effective regional blockade. During each phase, maternal blood pressure and pulse were monitored every 5 minutes, and continuous-wave Doppler recordings of the umbilical and uterine arteries were made at three separate intervals. Epidural placement and dosage techniques were similar for all patients; 0.25% bupivacaine solution was used and sensory levels of T6 obtained. The mean maternal systolic and diastolic blood pressures and pulse rate declined significantly, but no woman experienced hypotension. Mean systolic/diastolic ratios and pulsatility indices of the umbilical and uterine arteries did not change significantly during the study. We conclude that effective epidural anesthesia does not have a significant impact on Doppler flow characteristics of either the maternal or fetal umbilical vasculature, despite lowered maternal blood pressure and pulse rate.


Comparative genomics using Fugu reveals insights into regulatory subfunctionalization.
A major mechanism for the preservation of gene duplicates in the genome is thought to be mediated via loss or modification of cis-regulatory subfunctions between paralogs following duplication (a process known as regulatory subfunctionalization). Despite a number of gene expression studies that support this mechanism, no comprehensive analysis of regulatory subfunctionalization has been undertaken at the level of the distal cis-regulatory modules involved. We have exploited fish-mammal genomic alignments to identify and compare more than 800 conserved non-coding elements (CNEs) that associate with genes that have undergone fish-specific duplication and retention. Using the abundance of duplicated genes within the Fugu genome, we selected seven pairs of teleost-specific paralogs involved in early vertebrate development, each containing clusters of CNEs in their vicinity. CNEs present around each Fugu duplicated gene were identified using multiple alignments of orthologous regions between single-copy mammalian orthologs (representing the ancestral locus) and each fish duplicated region in turn. Comparative analysis reveals a pattern of element retention and loss between paralogs indicative of subfunctionalization, the extent of which differs between duplicate pairs. In addition to complete loss of specific regulatory elements, a number of CNEs have been retained in both regions but may be responsible for more subtle levels of subfunctionalization through sequence divergence. Comparative analysis of conserved elements between duplicated genes provides a powerful approach for studying regulatory subfunctionalization at the level of the regulatory elements involved.


Exome sequencing as a diagnostic tool for pediatric-onset ataxia.
Ataxia demonstrates substantial phenotypic and genetic heterogeneity. We set out to determine the diagnostic yield of exome sequencing in pediatric patients with ataxia without a molecular diagnosis after standard-of-care assessment in Canada. FORGE (Finding Of Rare disease GEnes) Canada is a nation-wide project focused on identifying novel disease genes for rare pediatric diseases using whole-exome sequencing. We retrospectively selected all FORGE Canada projects that included cerebellar ataxia as a feature. We identified 28 such families and a molecular diagnosis was made in 13; a success rate of 46%. In 11 families, we identified mutations in genes associated with known neurological syndromes and in two we identified novel disease genes. Exome analysis of sib pairs and/or patients born to consanguineous parents was more likely to be successful (9/13) than simplex cases (4/15). Our data suggest that exome sequencing is an effective first line test for pediatric patients with ataxia where a specific single gene is not immediately suspected to be causative.