Autoantibody to apolipoprotein A-1 in hepatitis C virus infection: a role in atherosclerosis?
One to three per cent of the world's population has hepatitis C virus (HCV) infection, which is not only a major cause of liver disease and cancer but also associated with an increased risk of atherosclerosis, despite an ostensibly favourable lipid profile. Autoantibodies are frequent in HCV infection and emerging evidence shows that autoantibodies could be valuable for cardiovascular disease (CVD) risk stratification. This study investigated a novel independent biomarker of CVD, autoantibodies to apolipoprotein A-1 (anti-apoA-1 IgG) and lipids in patients with chronic HCV before, during and after direct-acting anti-viral (DAA) therapy. Eighty-nine blinded serum samples from 27 patients with advanced chronic HCV were assayed for lipids and anti-apoA-1 IgG by ELISA. Pre-treatment HCV viral load correlated with high-density lipoprotein cholesterol (HDL-C, r = 0.417; p = 0.042) and negatively with apolipoprotein (apo)B (r = - 0.497; p = 0.013) and markers of CVD risk, the apoB/apoA-1 ratio (r = - 0.490; p = 0.015) and triglyceride level (TG)/HDL-C ratio (r = - 0.450; p = 0.031). Fourteen (52%) of 27 patients had detectable anti-apoA-1 IgG autoantibodies pre-treatment; only two became undetectable with virological cure. Autoantibody-positive sera had lower apoA-1 (p = 0.012), HDL-C (p = 0.009) and total cholesterol (p = 0.006) levels. This is the first report of the presence of an emerging biomarker for atherosclerosis, anti-apoA-1 IgG, in some patients with HCV infection. It may be induced by apoA-1 on the surface of HCV lipoviral particles. The autoantibodies inversely correlate with apoA-1 and HDL levels and may render HDL dysfunctional. Whether these hypothesis-generating findings have clinical implications in HCV patients requires further study.


Methylation of the TERT promoter and risk stratification of childhood brain tumours: an integrative genomic and molecular study.
Identification of robust biomarkers of malignancy and methods to establish disease progression is a major goal in paediatric neuro-oncology. We investigated whether methylation of the TERT promoter can be a biomarker for malignancy and patient outcome in paediatric brain tumours. For the discovery cohort, we used samples obtained from patients with paediatric brain tumours and individuals with normal brain tissues stored at the German Cancer Research Center (Heidelberg, Germany). We used methylation arrays for genome-wide assessment of DNA. For the validation cohort, we used samples obtained from several tissues for which full clinical and follow-up data were available from two hospitals in Toronto (ON, Canada). We did methylation analysis using quantitative Sequenom and pyrosequencing of an identified region of the TERT promoter. We assessed TERT expression by real-time PCR. To establish whether the biomarker could be used to assess and predict progression, we analysed methylation in paired samples of tumours that transformed from low to high grade and from localised to metastatic, and in choroid plexus tumours of different grades. Finally, we investigated overall survival in patients with posterior fossa ependymomas in which the identified region was hypermethylated or not. All individuals responsible for assays were masked to the outcome of the patients. Analysis of 280 samples in the discovery cohort identified one CpG site (cg11625005) in which 78 (99%) of 79 samples from normal brain tissues and low-grade tumours were not hypermethylated, but 145 (72%) of 201 samples from malignant tumours were hypermethylated (>15% methylated; p<0.0001). Analysis of 68 samples in the validation cohort identified a subset of five CpG sites (henceforth, upstream of the transcription start site [UTSS]) that was hypermethylated in all malignant paediatric brain tumours that expressed TERT but not in normal tissues that did not express TERT (p<0.0001). UTSS had a positive predictive value of 1.00 (95% CI 0.95-1.00) and a negative predictive value of 0.95 (0.87-0.99). In two paired samples of paediatric gliomas, UTSS methylation increased during transformation from low to high grade; it also increased in two paired samples that progressed from localised to metastatic disease. Two of eight atypical papillomas that had high UTSS methylation progressed to carcinomas, while the other six assessed did not progress or require additional treatment. 5-year overall survival was 51% (95% CI 31-71) for 25 patients with hypermethylated UTSS posterior fossa ependymomas and 95% (86-100) for 20 with non-hypermethylated tumours (p=0.0008). 5-year progression-free survival was 86% (68-100) for the 25 patients with non-hypermethylated UTSS tumours and 30% (10-50) for those with hypermethylated tumours (p=0.0008). Hypermethylation of the UTSS region in the TERT promoter is associated with TERT expression in cancers. In paediatric brain tumours, UTSS hypermethylation is associated with tumour progression and poor prognosis. This region is easy to amplify, and the assay to establish hypermethylation can be done on most tissues in most clinical laboratories. Therefore the UTSS region is a potentially accessible biomarker for various cancers. The Canadian Institute of Health Research and the Terry Fox Foundation.


Tumour prevention by a single antibody domain targeting the interaction of signal transduction proteins with RAS.
Many disease-related processes occur via protein complexes that are considered undruggable with small molecules. An example is RAS, which is frequently mutated in cancer and contributes to initiation and maintenance of the disease by constitutive signal transduction through protein interaction with effector proteins, like PI3K, RAF and RALGDS. Such protein interactions are therefore significant targets for therapy. We describe a single immunoglobulin variable region domain that specifically binds to activated GTP-bound RAS and prevents RAS-dependent tumorigenesis in a mouse model. The crystal structure of the immunoglobulin-RAS complex shows that the variable region competitively binds to the conformationally variant regions of RAS, where its signalling effector molecules interact. This allows the plasma membrane targeted single domain intrabody to inhibit signalling by mutant RAS. This mode of action is a novel advance to directly interfere with oncogenic RAS function in human cancer and shows a universally applicable approach to develop macromolecules to combat cancer. In addition, this method illustrates a general means for interfering with protein interactions that are commonly considered intractable as conventional drug targets.


Safety of intramedullary Schwann cell transplantation for postrehabilitation spinal cord injuries: 2-year follow-up of 33 cases.
Many experimental studies on spinal cord injuries (SCIs) support behavioral improvement after Schwann cell treatment. This study was conducted to evaluate safety issues 2 years after intramedullary Schwann cell transplantation in 33 consecutively selected patients with SCI. Of 356 patients with SCIs who had completed at least 6 months of a conventional rehabilitation program and who were screened for the study criteria, 33 were enrolled. After giving their informed consent, they volunteered for participation. They underwent sural nerve harvesting and intramedullary injection of a processed Schwann cell solution. Outcome assessments included a general health questionnaire, neurological examination, and functional recordings in terms of American Spinal Injury Association (ASIA) and Functional Independence Measure scoring, which were documented by independent observers. There were 24 patients with thoracic and 9 with cervical injuries. Sixteen patients were categorized in ASIA Grade A, and the 17 remaining participants had ASIA Grade B. There were no cases of deep infection, and the follow-up MR imaging studies obtained at 2 years did not reveal any deformity related to the procedure. There was no case of permanent neurological worsening or any infectious or viral complications. No new increment in syrinx size or abnormal tissue and/or tumor formation were observed on contrast-enhanced MR imaging studies performed 2 years after the treatment. Preliminary results, especially in terms of safety, seem to be promising, paving the way for future cell therapy trials.


HIV-1 DNA Flap formation promotes uncoating of the pre-integration complex at the nuclear pore.
The HIV-1 central DNA Flap acts as a cis-acting determinant of HIV-1 genome nuclear import. Indeed, DNA-Flap re-insertion within lentiviral-derived gene transfer vectors strongly stimulates gene transfer efficiencies. In this study, we sought to understand the mechanisms by which the central DNA Flap mediates HIV-1 nuclear import. Here, we show that reverse transcription (RT degrees) occurs within an intact capsid (CA) shell, independently of the routing process towards the nuclear membrane, and that uncoating is not an immediate post-fusion event, but rather occurs at the nuclear pore upon RT degrees completion. We provide the first observation with ultrastructural resolution of intact intracellular HIV-1 CA shells by scanning electron microscopy. In the absence of central DNA Flap formation, uncoating is impaired and linear DNA remains trapped within an integral CA shell precluding translocation through the nuclear pore. These data show that DNA Flap formation, the very last event of HIV-1 RT degrees, acts as a viral promoting element for the uncoating of HIV-1 at the nuclear pore.


The Prevalence of and Factors Associated With Receiving Concurrent Controlled Substance Prescriptions.
Controlled substance prescription (CSP) use and abuse in the United States has become a public health epidemic. One common and dangerous indicator of abuse involves obtaining CSPs concurrently. The objective of this study was to identify the prevalence and factors associated with individuals receiving potentially inappropriate concurrent CSPs. This was a retrospective, cohort analysis using data from the 2013-2014 Texas prescription drug monitoring program (PDMP). Descriptive statistics and a multiple logistic regression analysis were conducted to examine the individual-level and prescription utilization factors associated with individuals obtaining concurrent CSPs. Among Texas residents, 1,640,015 individuals (6.10% of all Texas residents and 15.99% of all CSP utilizers) were identified with at least one concurrent CSP combination. Concurrent opioid prescriptions were found to be the most prevalent concurrent use combination (n = 1,574,572). Having prescriptions for opioids, being ≥18 years of age, utilizing CSPs for >30 days, utilizing greater than one CSP, having a multiple provider episode, and traveling >25 miles to obtain CSPs were all statistically significant predictors of utilizing concurrent CSPs. The study findings indicate a high prevalence of individuals utilizing concurrent CSPs. This practice is concerning when considering that many overdose deaths result from the concurrent use of CSPs. Prescribers should utilize PDMPs to ensure that aberrant prescription drug behaviors, such as the use of concurrent opioids and benzodiazepines do not readily occur.


Zika Virus Associated with Microcephaly.
A widespread epidemic of Zika virus (ZIKV) infection was reported in 2015 in South and Central America and the Caribbean. A major concern associated with this infection is the apparent increased incidence of microcephaly in fetuses born to mothers infected with ZIKV. In this report, we describe the case of an expectant mother who had a febrile illness with rash at the end of the first trimester of pregnancy while she was living in Brazil. Ultrasonography performed at 29 weeks of gestation revealed microcephaly with calcifications in the fetal brain and placenta. After the mother requested termination of the pregnancy, a fetal autopsy was performed. Micrencephaly (an abnormally small brain) was observed, with almost complete agyria, hydrocephalus, and multifocal dystrophic calcifications in the cortex and subcortical white matter, with associated cortical displacement and mild focal inflammation. ZIKV was found in the fetal brain tissue on reverse-transcriptase-polymerase-chain-reaction (RT-PCR) assay, with consistent findings on electron microscopy. The complete genome of ZIKV was recovered from the fetal brain.


Santamarine Inhibits NF-кB and STAT3 Activation and Induces Apoptosis in HepG2 Liver Cancer Cells via Oxidative Stress.
Sesquiterpene lactones have long been used in traditional Chinese medicines to treat inflammatory diseases. Recently, sesquiterpene lactone family compounds have been recognized as potential anticancer agents. Thus, it is necessary to explore new sesquiterpene lactones and their antitumor mechanism for cancer treatments. In the present study, we have explored the potential anti-cancer activity of a novel sesquiterpene lactone compound "santamarine" (STM) in HepG2 cells. It inhibited proliferation and induced apoptosis dose-dependently with IC50 ~ 70 μM. Induction of apoptosis was found to be linked with increased reactive oxygen species (ROS) generation, decreased activity of thioredoxin reductase (TrxR), glutathione (GSH) depletion, mitochondrial membrane potential (ΔΨm) dissipation, Bcl-2 family proteins modulation, cytochrome c release, caspases-9, -8 and -3 activation and PARP cleavage. Further mechanistic study demonstrated that STM inhibited the constitutive and TNF-α-induced translocation of NF-кB into nucleus by decreasing phosphorylation of IkB-α. Moreover, STM inhibited STAT3 activation by decreasing phosphorylation at tyrosine705. NAC pretreatment reversed the effect of STM-mediated cell death, NF-кB inhibition and blockage of STAT3 activity, indicating the involvement of oxidative stress in STM-mediated anticancer activity. Further studies are needed to explore the exact molecular mechanism of STM-induced apoptosis to develop it into a lead for treatment of liver cancer in future.


TEASIng apart alien species risk assessments: a framework for best practices.
Some alien species cause substantial impacts, yet most are innocuous. Given limited resources, forecasting risks from alien species will help prioritise management. Given that risk assessment (RA) approaches vary widely, a synthesis is timely to highlight best practices. We reviewed quantitative and scoring RAs, integrating > 300 publications into arguably the most rigorous quantitative RA framework currently existing, and mapping each study onto our framework, which combines Transport, Establishment, Abundance, Spread and Impact (TEASI). Quantitative models generally measured single risk components (78% of studies), often focusing on Establishment alone (79%). Although dominant in academia, quantitative RAs are underused in policy, and should be made more accessible. Accommodating heterogeneous limited data, combining across risk components, and developing generalised RAs across species, space and time without requiring new models for each species may increase attractiveness for policy applications. Comparatively, scoring approaches covered more risk components (50% examined > 3 components), with Impact being the most common component (87%), and have been widely applied in policy (> 57%), but primarily employed expert opinion. Our framework provides guidance for questions asked, combining scores and other improvements. Our risk framework need not be completely parameterised to be informative, but instead identifies opportunities for improvement in alien species RA.


Sequential batch membrane bio-reactor for wastewater treatment: The effect of increased salinity.
In this work, a sequential batch membrane bioreactor pilot plant is investigated to analyze the effect of a gradual increase in salinity on carbon and nutrient removal, membrane fouling and biomass kinetic parameters. The salinity was increased by 2gNaClL(-1) per week up to 10gNaClL(-1). The total COD removal efficiency was quite high (93%) throughout the experiment. A gradual biomass acclimation to the salinity level was observed during the experiment, highlighting the good recovery capabilities of the system. Nitrification was also influenced by the increase in salinity, with a slight decrease in nitrification efficiency (the lowest value was obtained at 10gNaClL(-1) due to lower nitrifier activity). Irreversible cake deposition was the predominant fouling mechanism observed during the experiment. Respirometric tests exhibited a stress effect due to salinity, with a reduction in the respiration rates observed (from 8.85mgO2L(-1)h(-1) to 4mgO2L(-1)h(-1)).


Humans can perceive heading without visual path information.
It has previously been reported that humans can determine their direction of 3D translation (heading) from the 2D velocity field of retinal motion experienced during self-motion through a rigid environment, as is done by current computational models of visual heading estimation from optic flow. However, these claims were supported by studies that used stimuli that contained low rotational flow rates and/or additional visual cues beyond the velocity field or a task in which observers were asked to indicate their future trajectory of self-motion (path). Thus, previous conclusions about heading estimation have been confounded by the presence of other visual factors beyond the velocity field, by the use of a path-estimation task, or both. In particular, path estimation involves an exocentric computation with respect to an environmental reference, whereas heading estimation is an egocentric computation with respect to one's line of sight. Here, we use a heading-adjustment task to demonstrate that humans can precisely estimate their heading from the velocity field, independent of visual information about path, displacement, layout, or acceleration, with accuracy robust to rotation rates at least as high as 20 deg/s. Our findings show that instantaneous velocity-field information about heading is directly available for the visual control of locomotion and steering.


Conserved indels in essential proteins that are distinctive characteristics of Chlamydiales and provide novel means for their identification.
All known chlamydiae are either proven human or animal pathogens or possess such potential. Due to increasing reports of chlamydiae diversity in the environment, it is important to develop reliable means for identifying and characterizing Chlamydiales species. The identification of environmental chlamydiae at present relies on their branching pattern in 16S rRNA trees, as well as 16S/23S consensus motifs which display variability. At present, no reliable molecular signatures are known which are unique to all Chlamydiales species. Besides the rRNAs, sequence information for different Chlamydiales is not available for any other gene sequence. In this report, a number of molecular signatures are described that consist of conserved inserts and deletions (indels), in widely distributed proteins [RNA polymerase alpha subunit (RpoA), elongation factor (EF)-Tu, EF-P, DNA gyrase B subunit (GyrB) and lysyl-tRNA synthetase (LysRS)], that are distinctive characteristics of all available chlamydiae homologues (from Chlamydiaeceae species and Parachlamydiae sp. UWE25) and not found in any other bacteria. Using PCR primers for highly conserved regions in these proteins, the corresponding fragments of these genes from Simkania negevensis, Waddlia chondrophila, and in a number of cases for Neochlamydia hartmanellae, covering all families within the phylum Chlamydiae, have been cloned and sequenced. The shared presence of the identified signatures in these species provides strong evidence that these molecular signatures are distinctive characteristics of the entire order Chlamydiales and can be used to reliably determine the presence of chlamydiae or chlamydia-related organisms in environmental samples. The sequence information for these protein fragments was also used to determine the interrelationships among chlamydiae species. In a phylogenetic tree based on a combined dataset of sequences from RpoA, EF-Tu, EF-P and GyrB, the environmental chlamydiae (i.e. Simkania, Waddlia and Parachlamydia) and the traditional Chlamydiaceae (i.e. Chlamydophila and Chlamydia) formed two distinct clades. Similar relationships were also observed in individual protein phylogenies, as well as in a 16S rRNA tree for the same species. These results provide evidence that the divergence between the traditional Chlamydiaceae species and the other chlamydiae families occurred very early in the evolution of this group of bacteria.


Impaired cardiac contractility response to hemodynamic stress in S100A1-deficient mice.
Ca(2+) signaling plays a central role in cardiac contractility and adaptation to increased hemodynamic demand. We have generated mice with a targeted deletion of the S100A1 gene coding for the major cardiac isoform of the large multigenic S100 family of EF hand Ca(2+)-binding proteins. S100A1(-/-) mice have normal cardiac function under baseline conditions but have significantly reduced contraction rate and relaxation rate responses to beta-adrenergic stimulation that are associated with a reduced Ca(2+) sensitivity. In S100A1(-/-) mice, basal left-ventricular contractility deteriorated following 3-week pressure overload by thoracic aorta constriction despite a normal adaptive hypertrophy. Surprisingly, heterozygotes also had an impaired response to acute beta-adrenergic stimulation but maintained normal contractility in response to chronic pressure overload that coincided with S100A1 upregulation to wild-type levels. In contrast to other genetic models with impaired cardiac contractility, loss of S100A1 did not lead to cardiac hypertrophy or dilation in aged mice. The data demonstrate that high S100A1 protein levels are essential for the cardiac reserve and adaptation to acute and chronic hemodynamic stress in vivo.


Do 5HTTLPR and stress interact in risk for depression and suicidality? Item response analyses of a large sample.
The reported interaction between the length polymorphism (5HTTLPR) in the serotonin transporter gene (SLC6A4) and stressful life events on depression has led to many attempts to replicate but with inconsistent results. This inconsistency may reflect, in part, small sample size and the unknown contribution of the long allele SNP, rs25531. Using a large twin sample of 3,243 individuals from 2,230 families aged 18-95 years (mean = 32.3, SD = 13.6) we investigate the interaction between 5HTTLPR (subtyped with SNP rs25531) and stressful events on risk of depression and suicidality using both ordinal regressions and item response theory analyses. Participants reported via mailed questionnaire (82% response rate) both stressful events in the preceeding 12 months and symptoms of depression. Stressful events were defined as "personal" (affecting the individual), or "network" (affecting close family or friends). One to 10 years later (mean = 4.2 years), participants completed a comprehensive clinical psychiatric telephone interview (83% response rate) which assessed DSM-IV major depression and ideation of suicidality. Self-reports of depression and an increase in depression/suicidality assessed by clinical interview are significantly associated with prior personal events (P < 0.001) after controlling for age and sex. However, they are inconsistently associated with prior network events (ranging, ns to P < 0.01) and are not significantly associated with any of the genotype main effects (5HTTLPR, 5HTTLPR + rs25531) or interactions (stress x genotype). We find no evidence to support the hypothesis of any 5HTTLPR genotype by stress interaction.


Occupational cadmium exposure-associated oxidative stress and erythrocyte fragility among jewelry workers in India.
Cadmium-induced pulmonary and renal target organ effects are well-established although its association with oxidative stress and associated hematological effects for human toxicity remain understudied. In a population of cadmium-exposed male jewelry manufacturing workers (n = 32) and referents without direct exposure (n = 21), all with urinary cadmium quantification, we measured plasma antioxidant enzymes (catalase, superoxide dismutase), lipid peroxidation (malondialdehyde), erythrocyte fragility, and surface irregularity of the erythrocyte membrane. Compared to referents, exposed workers manifested significantly lower plasma antioxidant enzymes, and increased malondialdehyde and erythrocyte fragility (for all, P < 0.01). Consistent with the exposure subcategories, activities of superoxide dismutase and catalase were reduced and lipid peroxidation and erythrocyte fragility were enhanced (P < 0.01 for all) in terms of Cd-effect indicating a strong impact on hematological system and oxidative stress. Cd exposure contributes to oxidative stress and related erythrocyte effects thus making the hematological system another end-organ target for chronic Cd toxicity.


Structural requirements for charged lipid molecules to directly increase or suppress K+ channel activity in smooth muscle cells. Effects of fatty acids, lysophosphatidate, acyl coenzyme A and sphingosine.
We determined the structural features necessary for fatty acids to exert their action on K+ channels of gastric smooth muscle cells. Examination of the effects of a variety of synthetic and naturally occurring lipid compounds on K+ channel activity in cell-attached and excised membrane patches revealed that negatively charged analogs of medium to long chain fatty acids (but not short chain analogs) as well as certain other negatively charged lipids activate the channels. In contrast, positively charged, medium to long chain analogs suppress activity, and neutral analogs are without effect. The key requirements for effective compounds seem to be a sufficiently hydrophobic domain and the presence of a charged group. Furthermore, those negatively charged compounds unable to "flip" across the bilayer are effective only when applied at the cytosolic surface of the membrane, suggesting that the site of fatty acid action is also located there. Finally, because some of the effective compounds, for example, the fatty acids themselves, lysophosphatidate, acyl Coenzyme A, and sphingosine, are naturally occurring substances and can be liberated by agonist-activated or metabolic enzymes, they may act as second messengers targeting ion channels.


Modulation of human beta-defensin-2 transcription in pulmonary epithelial cells by lipopolysaccharide-stimulated mononuclear phagocytes via proinflammatory cytokine production.
Human beta-defensin (hBD)-2, a cationic antimicrobial peptide primarily induced in epithelial cells in response to inflammatory stimuli, plays an important role in host defense. To elucidate the expression mechanism of hBD-2 in the lung, we investigated the modulation of hBD-2 transcription in pulmonary epithelial cells by mononuclear phagocytes stimulated with LPS. Coculture of A549 pulmonary epithelial cells with Mono-Mac-6 monocytic cells in the presence of Escherichia coli LPS markedly up-regulated hBD-2 promoter activity, whereas A549 alone did not respond to LPS to activate the hBD-2 promoter. Furthermore, IL-1beta and TNF-alpha in the culture supernatants from LPS-stimulated monocytic cells activated the hBD-2 promoter in A549 cells. Of note, IL-1beta was more potent than TNF-alpha in this effect. In addition, a mutation of the NF-kappaB site at -200 (pkappaB1 site) completely abolished this IL-1beta- and TNF-alpha-induced hBD-2 promoter activation, whereas NF-kappaB inhibitors (MG-132 and helenalin) strongly suppressed it. Moreover, electrophoretic mobility shift assay suggested that NF-kappaB, consisting of p65-p50 heterodimer, could bind to the pkappaB1 site in cytokine-stimulated A549 cells. Interestingly, flow cytometric analysis revealed that A549 cells expressed CD14 but lacked Toll-like receptor 4, which may account for the hyporesponsiveness of A549 cells to LPS. Taken together, these results suggest that hBD-2 expression in pulmonary epithelial cells is modulated by NF-kappaB via the actions of IL-1beta and TNF-alpha produced by LPS-stimulated mononuclear phagocytes.


Crosslinked conjugated polymers as hole transport layers in high-performance quantum dot light-emitting diodes.
Film morphologies of functional layers in all-solution-processed quantum dot light-emitting diodes (QLEDs) play a crucial role in device performance. Solvents for adjacent layers should be strictly orthogonal to prevent the preceding layer being redissolved by the processing solvent of the next layer. Herein, we use a photochemical crosslinking method to obtain solvent-resistant hole transport layers (HTLs) with photoinitiator bifunctional bis-benzophenone (BP-BP). With this method, ultra-smooth quantum dot (QD) layers can be fabricated using toluene as solvent, which is known to be a nonorthogonal solvent in common non-crosslinked HTLs. A green QLED device based on crosslinked HTLs exhibits a high external quantum efficiency of 8.93%, which is 1.9-fold higher than that of the non-crosslinked device. The improved device performance is ascribed to the well preserved film morphology of crosslinked HTLs and the prevention of QDs intermixing with HTLs during the QD deposition in toluene. This crosslinking strategy avoids high-temperature annealing, allowing the fabrication of flexible devices on plastic substrates. Moreover, it broadens the range of applicable solvents for solution-processed multilayer optoelectronic devices because non-orthogonal solvents can be used after crosslinking preceding layers.


Effects of magnesium sulphate on intraoperative anaesthetic requirements and postoperative analgesia in gynaecology patients receiving total intravenous anaesthesia.
This randomized, double-blind, prospective study was undertaken to evaluate the effects of magnesium sulphate on anaesthetic requirements and postoperative analgesia in patients undergoing total i.v. anaesthesia (TIVA). Fifty patients who underwent gynaecological surgery were randomly divided into two groups. Before induction of anaesthesia, the magnesium group (Group M) received magnesium sulphate 50 mg kg(-1) i.v. as a bolus and then 15 mg kg(-1) h(-1) i.v. by continuous infusion. The control group (Group S) received the same amount of isotonic saline. TIVA (propofol+remifentanil) was administered under bispectral index monitoring during anaesthesia induction and maintenance. Rocuronium was administered before orotracheal intubation and during surgery when the train-of-four count was 2 or more. After operation, patient-controlled analgesia with a solution of ketorolac and morphine was used and the consumption of this solution was recorded. Pain scores at rest and upon movement were evaluated 30 min, 4, 24, and 48 h after surgery. Patients in Group M required less rocuronium than those in Group S [mean (SD) 0.44 (0.09) vs 0.35 (0.07) microg kg(-1) min(-1), P<0.05]. The total amounts of propofol and remifentanil administered were similar in the two groups. Postoperative pain scores, cumulative analgesic consumption, and shivering incidents were significantly lower in Group M (P<0.05). Mean arterial pressure just after intubation and during the immediate postoperative period was also significantly lower in Group M (P<0.05). I.v. magnesium sulphate during TIVA reduced rocuronium requirement and improved the quality of postoperative analgesia.


Employment precariousness and poor mental health: evidence from Spain on a new social determinant of health.
Evidence on the health-damaging effects of precarious employment is limited by the use of one-dimensional approaches focused on employment instability. This study assesses the association between precarious employment and poor mental health using the multidimensional Employment Precariousness Scale. Cross-sectional study of 5679 temporary and permanent workers from the population-based Psychosocial Factors Survey was carried out in 2004-2005 in Spain. Poor mental health was defined as SF-36 mental health scores below the 25th percentile of the Spanish reference for each respondent's sex and age. Prevalence proportion ratios (PPRs) of poor mental health across quintiles of employment precariousness (reference: 1st quintile) were calculated with log-binomial regressions, separately for women and men. Crude PPRs showed a gradient association with poor mental health and remained generally unchanged after adjustments for age, immigrant status, socioeconomic position, and previous unemployment. Fully adjusted PPRs for the 5th quintile were 2.54 (95% CI: 1.95-3.31) for women and 2.23 (95% CI: 1.86-2.68) for men. The study finds a gradient association between employment precariousness and poor mental health, which was somewhat stronger among women, suggesting an interaction with gender-related power asymmetries. Further research is needed to strengthen the epidemiological evidence base and to inform labour market policy-making.


Association of coffee drinking with all-cause and cause-specific mortality in over 190,000 individuals: data from two prospective studies.
We examined the association of coffee drinking with all-cause and cause-specific mortality in a pooled analysis of two Korean prospective cohort studies: The Korea National Health and Nutrition Examination Survey and the Korean Genome and Epidemiology Study. We included 192,222 participants, and a total of 6057 deaths were documented. Cox proportional hazards model was used to estimate the hazard ratios (HRs) and 95% confidence intervals (CIs), and the HRs were combined using a random-effects model. Coffee drinking was associated with a lower risk of all-cause mortality [HR (95% CI) = 0.84 (0.77-0.92), for ≥3 cups/day of coffee drinking versus non-drinkers; p for trend = 0.004]. We observed the potential benefit of coffee drinking for mortality due to cardiovascular disease, respiratory disease, and diabetes, but not for cancer mortality. Overall, we found that moderate coffee drinking was associated with a lower risk of death in population-based cohort analysis of Korean adults.


Engaging family caregivers and health system partners in exploring how multi-level contexts in primary care practices affect case management functions and outcomes of patients and family caregivers at end of life: a realist synthesis.
An upstream approach to palliative care in the last 12 months of life delivered by primary care practices is often referred to as Primary Palliative Care (PPC). Implementing case management functions can support delivery of PPC and help patients and their families navigate health, social and fiscal environments that become more complex at end-of-life. A realist synthesis was conducted to understand how multi-level contexts affect case management functions related to initiating end-of-life conversations, assessing patient and caregiver needs, and patient/family centred planning in primary care practices to improve outcomes. The synthesis also explored how these functions aligned with critical community resources identified by patients/families dealing with end-of-life. A realist synthesis is theory driven and iterative, involving the investigation of proposed program theories of how particular contexts catalyze mechanisms (program resources and individual reactions to resources) to generate improved outcomes. To assess whether program theories were supported and plausible, two librarian-assisted and several researcher-initiated purposive searches of the literature were conducted, then extracted data were analyzed and synthesized. To assess relevancy, health system partners and family advisors informed the review process. Twenty-eight articles were identified as being relevant and evidence was consolidated into two final program theories: 1) Making end-of-life discussions comfortable, and 2) Creating plans that reflect needs and values. Theories were explored in depth to assess the effect of multi-level contexts on primary care practices implementing tools or frameworks, strategies for improving end-of-life communications, or facilitators that could improve advance care planning by primary care practitioners. Primary care practitioners' use of tools to assess patients/families' needs facilitated discussions and planning for end-of-life issues without specifically discussing death. Also, receiving training on how to better communicate increased practitioner confidence for initiating end-of-life discussions. Practitioner attitudes toward death and prior education or training in end-of-life care affected their ability to initiate end-of-life conversations and plan with patients/families. Recognizing and seizing opportunities when patients are aware of the need to plan for their end-of-life care, such as in contexts when patients experience transitions can increase readiness for end-of-life discussions and planning. Ultimately conversations and planning can improve patients/families' outcomes.


Curcumin suppresses increased bone resorption by inhibiting osteoclastogenesis in rats with streptozotocin-induced diabetes.
Curcumin is a potent inhibitor of the transcription factor activator protein-1 which plays an essential role in osteoclastogenesis. However, the effects of curcumin on bone metabolism have not been clarified in vivo. We reported herein the inhibitory effects of curcumin on the stimulated osteoclastic activity in insulin-dependent diabetes mellitus using rats with streptozotocin-induced diabetes. A dietary supplement of curcumin reversed the increase in levels of activity and mRNA of tartrate-resistant acid phosphatase (TRAP) and cathepsin K to control values. A histochemical analysis showed that the increase in TRAP-positive cells in the distal femur of the diabetic rats was reduced to the control level by the supplement. These results suggested that curcumin reduced diabetes-stimulated bone resorptive activity and the number of osteoclasts. When bone marrow cells were cultured with macrophage colony stimulating factor and receptor activator NF-kappaB ligand (RANKL), the increased activity to form TRAP-positive multinucleated cells and the increased levels of mRNA and protein of c-fos and c-jun in the cultured cells from diabetic rats decreased to control levels in the curcumin-supplemented rats. Similarly, the increased expression of c-fos and c-jun in the distal femur of the diabetic rats was significantly reduced by the supplement. These results suggested that curcumin suppressed the increased bone resorptive activity through the prevention of osteoclastogenesis associated with inhibition of the expression of c-fos and c-jun in the diabetic rats.


Regulation of insulin-like growth factor I messenger RNA levels in vascular smooth muscle cells.
We have previously demonstrated specific insulin-like growth factor I (IGF I) messenger RNA (mRNA) transcripts in cultured rat aortic smooth muscle cells (RASM). To define the role of IGF I in the autocrine growth program of vascular smooth muscle cells, we quantitated IGF I mRNA levels in proliferating and quiescent (serum-deprived for 48 hours) RASM. IGF I mRNA levels were markedly decreased in quiescent cells, and this effect was reversible on reexposure to serum. Since platelet-derived growth factor (PDGF) acts synergistically with IGF I to stimulate vascular smooth muscle cell growth, we exposed quiescent RASM to PDGF AB or BB and quantitated IGF I transcript levels. Both PDGF dimers caused a marked, rapid increase in IGF I message levels. To determine whether induction of IGF I mRNA levels correlated with secretion of IGF I, we measured immunoreactive IGF I in RASM conditioned medium after separation of IGF I binding proteins by gel filtration chromatography. PDGF caused a significant increase in IGF I release at 24 hours. These findings indicate that IGF I mRNA levels in vitro are regulated by serum and by growth factors such as PDGF. Serum deprivation reversibly decreases IGF I transcript levels, and exposure of quiescent cells to PDGF increases IGF I mRNA levels and IGF I release. Regulation of IGF I expression by competence growth factors such as PDGF may play an important role in the control of vascular smooth muscle cell growth.


Prevalence of antibodies against Rift Valley fever virus in Kenyan wildlife.
Rift Valley fever virus (RVFV) is an arbovirus associated with periodic outbreaks, mostly on the African continent, of febrile disease accompanied by abortion in livestock, and a severe, fatal haemorrhagic syndrome in humans. However, the maintenance of the virus during the inter-epidemic period (IEP) when there is low or no disease activity detected in livestock or humans has not been determined. This study report prevalence of RVFV-neutralizing antibodies in sera (n=896) collected from 16 Kenyan wildlife species including at least 35% that were born during the 1999-2006 IEP. Specimens from seven species had detectable neutralizing antibodies against RVFV, including African buffalo, black rhino, lesser kudu, impala, African elephant, kongoni, and waterbuck. High RVFV antibody prevalence (>15%) was observed in black rhinos and ruminants (kudu, impala, buffalo, and waterbuck) with the highest titres (up to 1:1280) observed mostly in buffalo, including animals born during the IEP. All lions, giraffes, plains zebras, and warthogs tested were either negative or less than two animals in each species had low (<or= 1:16) titres of RVFV antibodies. Of 249 sera collected from five wildlife species during the 2006-2007 outbreak, 16 out of 19 (84%) of the ruminant (gerenuk, waterbuck, and eland) specimens had RVFV-neutralizing titres >or= 1:80. These data provide evidence that wild ruminants are infected by RVFV but further studies are required to determine whether these animals play a role in the virus maintenance between outbreaks and virus amplification prior to a noticeable outbreak.


Continuity and change: paradigm shifts in neural induction.
The problem of "primary embryonic induction" was one of the first areas of developmental biology to become "molecularized." What had been seen as an intractable series of problems became amenable to the techniques of Northern blotting, ectopic RNA insertion, and in situ hybridization. These molecular analyses showed that some of the fundamental concepts of primary embryonic induction concluded by experimental embryologists were false. First, primary embryonic induction was not primary. The organizer tissue, itself, was the product of a prior induction. Second, the neural fate of cells was not being induced. Rather, the epidermal fate was induced and the neural state was the default, uninduced, fate of ectodermal tissues. Third, primary embryonic induction was not something unique to vertebrates. Rather, the ventral neural cord of insects formed using the same mechanisms as the dorsal neural tube of vertebrates. Fourth, the brain formed in a matter distinctly different from that the spinal cord. Despite these differences, there has been a clear and strong continuity between the experimental embryological tradition and the molecular genetic tradition, and these new results are seen by many contemporary developmental geneticists as strengthening, rather than destroying, the older science.


Initiating artificial nutrition support: a clinical judgement analysis.
Variations in the practice of artificial nutrition are partly a result of the judgements of clinicians. Little is known about these judgements, the decision processes used or the quality of the decisions that arise. The methodology of clinical judgement analysis describes and explains judgemental differences between clinicians and offers a window into why clinician choices sometimes fail to correspond with current best practice guidelines. A clinical judgement analysis with 27 clinicians using two judgement tasks was carried out. Clinicians expressed their 'likelihood of feeding' and 'whether or not they would initiate artificial nutrition support' in response to 54 patient scenarios. Measures of agreement between clinicians, the degree of linear reasoning employed, their judgemental consistency and concordance with National Institute for Health and Clinical Excellence (NICE) guidelines were assessed. Clinicians' judgements varied substantially both in the amount of information employed and the weighting given to information. Clinicians had only moderate agreement with NICE recommendations and overestimated the need for artificial nutrition support. Suboptimal judgements were consistently applied by clinicians. When clinicians reasoned in more linear ways, their judgements and decisions improved. Overestimating the need for artificial nutritional support may inflate health service costs and increase risks for patients unnecessarily. A better understanding by clinicians of the information needed (as well as how to use it) for artificial nutrition support initiation may help improve clinical decision-making.


Interaction of tricyclic drug analogs with synaptic plasma membranes: structure-mechanism relationships in inhibition of neuronal Na+/K(+)-ATPase activity.
Perturbations of rat brain synaptic plasma membrane (SPM) bilayer structure and Na+/K(+)-ATPase activity were correlated for drugs that are structurally related and exhibit similar toxicological side effects but belong to different pharmacological classes. Na+/K(+)-ATPase IC50 values decrease linearly with increasing octanol/water partition coefficients (log-log plot) for a series of dimethylethylamine-containing drugs (i.e., chlorpromazine, amitriptyline, imipramine, doxepin, and diphenhydramine), emphasizing hydrophobicity in inhibition. However, nortriptyline and desipramine are 1.2 log units less hydrophobic than their N-methylated parent drugs but more potent inhibitors. To investigate this, bilayer surface structure was examined by the binding of the fluorophore 1-anilinonaphthalene-8-sulfonic acid (ANS) to SPMs. The dissociation constant and wavelength maximum of ANS are invariant with drug binding; however, the limiting fluorescence intensity of ANS (F infinity) is increased. Such data indicate that these cationic drugs bind to the membrane surface, increasing the number but not the polarity of ANS binding sites by cancelling charge at anionic phospholipid groups. More importantly, there is a close linear correlation between the concentrations of drugs necessary to increase F infinity by 40% and the IC50 values, with full compensation for the N-demethylated drugs. This correlation implies that drug-induced increases in SPM-bound ANS fluorescence are a better predictor of Na+/K(+)-ATPase inhibition than are octanol/water partition coefficients and that electrostatic interactions are also involved in inhibition. Furthermore, it points toward similar mechanisms of biomembrane surface interaction governing both inhibition and fluorescence change that are common to these drugs. K(+)-dependent p-nitrophenylphosphatase activity is inhibited with the same potency as Na+/K(+)-ATPase activity, indicating that inhibition may involve drug interaction near the K+ binding sites. Furthermore, chlorpromazine, diphenhydramine, and dimethylaminopropyl chloride alter K(+)-activation of K(+)-dependent p-nitrophenylphosphatase, progressing from noncompetitive through mixed to competitive inhibition as their hydrophobicity changes, and these mechanisms are consistent with steric hindrance of K+ binding. In contrast to the ANS data, decreases in 1,6-diphenyl-1,3,5-hexatriene fluorescence anisotropy induced by these drugs do not correlate with Na+/K(+)-ATPase inhibition, and drug N-demethylation enhances inhibition without altering anisotropy; both findings indicate that Na+/K(+)-ATPase activity is not predominantly influenced by changes in bulk fluidity. Taken together, these data suggest that electrostatic interactions at the biomembrane surface between the protonated amino group of the drug and anionic groups on the enzyme and/or phospholipids near the K+binding sites are crucial to inhibition and that drug hydrophobicity modulated the number and orientation of these interactions.


Assessment of a vessel and tissue-sealing device for ovariectomy in chickens to evaluate the potential application of the procedure to other avian species.
To assess the feasibility of a novel technique involving a vessel and tissue-sealing device (VTSD) for ovariectomy in chickens to evaluate the potential application of the procedure to other avian species. 20 domestic laying hens (Gallus domesticus), of which 10 were immature (< 4 months old) and 10 were adults (> 18 months old). Ovariectomy was performed with a VTSD through a left lateral celiotomy. Birds were allowed to recover for 14 days after the procedure and then were euthanized for necropsy. A board-certified veterinary pathologist performed complete necropsies, with particular attention to identifying any remaining ovarian tissue. All birds survived the procedure. For the mature and juvenile birds, the mean ± SD durations of anesthesia (interval from intubation to extubation) were 67.2 ± 7.6 minutes and 50.5 ± 5.1 minutes, respectively, and mean durations of surgery were 45.3 ± 8.5 minutes and 31.6 ± 5.1 minutes, respectively. Three birds had severe hemorrhage during ovariectomy. At necropsy, ovarian tissue was present grossly in 2 mature birds and histologically in 6 additional birds (2 mature and 4 juvenile birds), indicating incomplete excision in 8 (40%) birds. Results indicated that the evaluated VTSD can be used to successfully perform ovariectomies in both juvenile and mature chickens, although the procedure was associated with major hemorrhage and incomplete excision of ovarian tissue in some cases. Use of this VTSD for ovariectomy in birds of other species, particularly birds with reproductive tract disease, should be investigated.


Predictors of costs related to cardiovascular disease among patients with atrial fibrillation in five European countries.
To estimate predictors of direct costs and costs of hospitalization related to cardiovascular disease (CVD) in patients with atrial fibrillation (AF) recruited to the Euro Heart Survey on AF (EHS-AF) in Greece, Italy, Poland, Spain, and the Netherlands. Annual direct costs were modelled by country using ordinary least squares (OLS) regression. For costs of hospitalization related to CVD, logistic regressions followed by conditional OLS regression were employed. In each case, effects of the following potential explanatory variables were tested: age, sex, body mass index, type of AF, diabetes, hypertension, myocardial infarction (MI), angina pectoris (AP), valvular heart disease (VHD), congestive heart failure (CHF), stroke, and/or other underlying heart disease at the time of enrolment in the EHS-AF. Estimated direct annual costs for the reference EHS-AF patient (female aged <65 years with first-detected AF and no co-morbidities at baseline) were €933 in Greece, €1383 in Italy, €698 in Poland, €1316 in Spain, and €1544 in the Netherlands. The co-morbidities identified as predictors of direct costs were VHD in Greece, Italy, and Spain, AP in Italy and Spain, diabetes and stroke in Poland, CHF in Italy, MI in Spain and other underlying heart disease in Poland and the Netherlands. For costs of CVD-related hospitalization, the most important co-morbidity identified as a predictor was VHD. The results reported in this study increase the understanding of the cost structure of CVD in AF patients and may therefore inform the targeting of intervention therapy in selected groups of at-risk patients.


Pathology, toxicology, and latency of irritant gases known to cause bronchiolitis obliterans disease: Does diacetyl fit the pattern?
Bronchiolitis obliterans (BO) is a rare disease involving concentric bronchiolar fibrosis that develops rapidly following inhalation of certain irritant gases at sufficiently high acute doses. While there are many potential causes of bronchiolar lesions involved in a variety of chronic lung diseases, failure to clearly define the clinical features and pathological characteristics can lead to ambiguous diagnoses. Irritant gases known to cause BO follow a similar pathologic process and time course of disease onset in humans. Studies of inhaled irritant gases known to cause BO (e.g., chlorine, hydrochloric acid, ammonia, nitrogen oxides, sulfur oxides, sulfur or nitrogen mustards, and phosgene) indicate that the time course between causal chemical exposures and development of clinically significant BO disease is typically limited to a few months. The mechanism of toxic action exerted by these irritant gases generally involves widespread and severe injury of the epithelial lining of the bronchioles that leads to acute respiratory symptoms which can include lung edema within days. Repeated exposures to inhaled irritant gases at concentrations insufficient to cause marked respiratory distress or edema may lead to adaptive responses that can reduce or prevent severe bronchiolar fibrotic changes. Risk of BO from irritant gases is driven substantially by toxicokinetics affecting concentrations occurring at the bronchiolar epithelium. Highly soluble irritant gases that cause BO like ammonia generally follow a threshold-dependent cytotoxic mechanism of action that at sufficiently high doses results in severe inflammation of the upper respiratory tract and the bronchiolar epithelium concurrently. This is followed by acute respiratory distress, pulmonary edema, and post inflammatory concentric fibrosis that become clinically obvious within a few months. In contrast, irritant gases with lower solubility like phosgene also follow a threshold-dependent mechanism of cytotoxicity action but can exhibit more insidious and isolated bronchiolar tissue damage with a similar latency to fibrosis. To date, animal and human studies on the highly soluble gas, diacetyl, have not identified a coherent pattern of pathology and latency that would be expected based on studies of other known causes of bronchiolitis obliterans disease.


Beta-catenin immunohistochemistry separates mesenteric fibromatosis from gastrointestinal stromal tumor and sclerosing mesenteritis.
Although separating gastrointestinal stromal tumor (GIST) from mesenteric fibromatosis and sclerosing mesenteritis is clinically important, this distinction sometimes poses problems for practicing pathologists. In the STI571 (Gleevec, Imatinib) era, the problem may be further compounded when protocol-driven staining for CD117 (c-kit) is performed on spindle cell proliferations presenting in the bowel wall and mesentery using an antibody known to react with the majority of mesenteric fibromatoses when other antibodies are more specific. Because most mesenteric fibromatoses have mutations in the pathway and hence have abnormal nuclear accumulation of beta-catenin protein, we studied beta-catenin expression among a panel of other immunohistochemical stains to distinguish mesenteric fibromatosis, GIST, and sclerosing mesenteritis. Examples of gastrointestinal stromal tumors (GIST, 11), sclerosing mesenteritis (5), and mesenteric fibromatosis (10) were retrieved from the archives of our institutions. Cases were studied with an immunohistochemical panel consisting of CD117, beta-catenin, CD34, smooth muscle actin, desmin, keratin, and S-100 protein. Cases were scored as "negative," "focally positive," or "diffusely positive." In evaluating beta-catenin, nuclear accumulation was required. GIST all had CD117 (11 of 11, diffuse) and CD34 (11 of 11, diffuse) with variable actin (5 of 11, focal) and negative desmin, keratin, S-100 protein. All GIST lacked beta-catenin (0 of 11). Mesenteric fibromatosis had CD117 (6 of 10, 3 focal, 3 diffuse), typically expressed more weakly than in GIST, actin (5 of 9, focal), and desmin (3 of 8, focal) in keeping with myofibroblastic differentiation but lacked CD34, S-100, and keratin. CD117 staining was not eliminated by use of a non-avidin-biotin technique. Nuclear beta-catenin was detected in 9 of 10 fibromatoses, including one case associated with familial adenomatous polyposis. Two of five sclerosing mesenteritis cases focally expressed CD117. None of the sclerosing mesenteritis cases had nuclear beta-catenin. Sclerosing mesenteritis cases were otherwise fibroblastic and myofibroblastic with focal actin in 5 of 5 and negative desmin, keratin, and S-100 protein but one had CD34 (1 of 5, focal). With increasing protocol-driven interest in evaluating bowel wall and mesenteric spindle cell lesions using CD117 (c-kit) antibodies, it is important for practicing pathologists to be aware that lesions other than GISTs are likely to express this antigen using certain antibodies. beta-Catenin staining identifies lesions that are, instead, mesenteric fibromatoses.


Genotype-phenotype correlation analysis for three primary angle closure glaucoma-associated genetic polymorphisms.
Recently, three genetic susceptibility loci for primary angle closure glaucoma (PACG) were identified: COL11A1 rs3753841, PCMTD1-ST18 rs1015213, and PLEKHA7 rs11024102. The purpose of this study was to investigate whether these single nucleotide polymorphisms (SNPs) affect the phenotype of PACG patients. A retrospective analysis was performed for 700 Singaporean Chinese PACG patients who had been genotyped. The associations between the three SNPs and clinical features related to severity of glaucoma were studied. For a subgroup of patients who had ≥ 5 years of follow-up and ≥ 5 reliable visual field (VF) tests, differences in glaucoma progression, as measured by the proportion of VF progression and blindness, were compared among groups with different genotypes. The minor allele frequencies at COL11A1 rs3753841, PCMTD1-ST18 rs1015213, and PLEKHA7 rs11024102 were 36%, 2.1%, and 41.5%, respectively. There were no significant differences in sex, diagnosis (acute primary angle closure [APAC] versus non-APAC), age at diagnosis, laterality of glaucoma, or need for filtration surgery among patients with different genotypes (all P > 0.05). We also found no significant difference between genotypes and the IOP at presentation, and other clinical characteristics at DNA collection (vertical cup-to-disc ratio, best corrected visual acuity, baseline VF mean deviation, or pattern standard deviation). For the subgroup analysis, we did not observe significant associations between VF progression and the proportion of blindness with any of the PACG susceptibility loci. The three genetic susceptibility loci for PACG did not underlie any major phenotypic diversity in terms of disease severity or progression.


Glycogen synthase kinase-3: cryoprotection and glycogen metabolism in the freeze-tolerant wood frog.
The terrestrial anuran Rana sylvatica tolerates extended periods of whole-body freezing during the winter. Freezing survival is facilitated by extensive glycogen hydrolysis and distribution of high concentrations of the cryoprotectant glucose into blood and all tissues. As glycogenesis is both an energy-expensive process and counter-productive to maintaining sustained high cryoprotectant levels, we proposed that glycogen synthase kinase-3 (GSK-3) would be activated when wood frogs froze and would phosphorylate its downstream substrates to inactivate glycogen synthesis. Western blot analysis determined that the amount of phosphorylated (inactive) GSK-3 decreased in all five tissues tested in 24 h frozen frogs compared with unfrozen controls. Total GSK-3 protein levels did not change, with the exception of heart GSK-3, indicating that post-translational modification was the primary regulatory mechanism for this kinase. Kinetic properties of skeletal muscle GSK-3 from control and frozen frogs displayed differential responses to a temperature change (22 versus 4°C) and high glucose. For example, when assayed at 4°C, the K(m) for the GSK-3 substrate peptide was ∼44% lower for frozen frogs than the corresponding value in control frogs, indicating greater GSK-3 affinity for its substrates in the frozen state. This indicates that at temperatures similar to the environment encountered by frogs, GSK-3 in frozen frogs will phosphorylate its downstream targets more readily than in unfrozen controls. GSK-3 from skeletal muscle of control frogs was also allosterically regulated. AMP and phosphoenolpyruvate activated GSK-3 whereas inhibitors included glucose, glucose 6-phosphate, pyruvate, ATP, glutamate, glutamine, glycerol, NH(4)Cl, NaCl and KCl. The combination of phosphorylation and allosteric control argues for a regulatory role of GSK-3 in inactivating glycogenesis to preserve high glucose cryoprotectant levels throughout each freezing bout.


Enemy release and plant invasion: patterns of defensive traits and leaf damage in Hawaii.
Invasive species may be released from consumption by their native herbivores in novel habitats and thereby experience higher fitness relative to native species. However, few studies have examined release from herbivory as a mechanism of invasion in oceanic island systems, which have experienced particularly high loss of native species due to the invasion of non-native animal and plant species. We surveyed putative defensive traits and leaf damage rates in 19 pairs of taxonomically related invasive and native species in Hawaii, representing a broad taxonomic diversity. Leaf damage by insects and pathogens was monitored in both wet and dry seasons. We found that native species had higher leaf damage rates than invasive species, but only during the dry season. However, damage rates across native and invasive species averaged only 2% of leaf area. Native species generally displayed high levels of structural defense (leaf toughness and leaf thickness, but not leaf trichome density) while native and invasive species displayed similar levels of chemical defenses (total phenolics). A defense index, which integrated all putative defense traits, was significantly higher for native species, suggesting that native species may allocate fewer resources to growth and reproduction than do invasive species. Thus, our data support the idea that invasive species allocate fewer resources to defense traits, allowing them to outperform native species through increased growth and reproduction. While strong impacts of herbivores on invasion are not supported by the low damage rates we observed on mature plants, population-level studies that monitor how herbivores influence recruitment, mortality, and competitive outcomes are needed to accurately address how herbivores influence invasion in Hawaii.


Multiple steps in the regulation of transcription-factor level and activity.
This review focuses on the regulation of transcription factors, many of which are DNA-binding proteins that recognize cis-regulatory elements of target genes and are the most direct regulators of gene transcription. Transcription factors serve as integration centres of the different signal-transduction pathways affecting a given gene. It is obvious that the regulation of these regulators themselves is of crucial importance for differential gene expression during development and in terminally differentiated cells. Transcription factors can be regulated at two, principally different, levels, namely concentration and activity, each of which can be modulated in a variety of ways. The concentrations of transcription factors, as of intracellular proteins in general, may be regulated at any of the steps leading from DNA to protein, including transcription, RNA processing, mRNA degradation and translation. The activity of a transcription factor is often regulated by (de) phosphorylation, which may affect different functions, e.g. nuclear localization DNA binding and trans-activation. Ligand binding is another mode of transcription-factor activation. It is typical for the large super-family of nuclear hormone receptors. Heterodimerization between transcription factors adds another dimension to the regulatory diversity and signal integration. Finally, non-DNA-binding (accessory) factors may mediate a diverse range of functions, e.g. serving as a bridge between the transcription factor and the basal transcription machinery, stabilizing the DNA-binding complex or changing the specificity of the target sequence recognition. The present review presents an overview of different modes of transcription-factor regulation, each illustrated by typical examples.


Genomic diversity of colorectal cancer: Changing landscape and emerging targets.
Improvements in screening and preventive measures have led to an increased detection of early stage colorectal cancers (CRC) where patients undergo treatment with a curative intent. Despite these efforts, a high proportion of patients are diagnosed with advanced stage disease that is associated with poor outcomes, as CRC remains one of the leading causes of cancer-related deaths in the world. The development of next generation sequencing and collaborative multi-institutional efforts to characterize the cancer genome has afforded us with a comprehensive assessment of the genomic makeup present in CRC. This knowledge has translated into understanding the prognostic role of various tumor somatic variants in this disease. Additionally, the awareness of the genomic alterations present in CRC has resulted in an improvement in patient outcomes, largely due to better selection of personalized therapies based on an individual's tumor genomic makeup. The benefit of various treatments is often limited, where recent studies assessing the genomic diversity in CRC have identified the development of secondary tumor somatic variants that likely contribute to acquired treatment resistance. These studies have begun to alter the landscape of treatment for CRC that include investigating novel targeted therapies, assessing the role of immunotherapy and prospective, dynamic assessment of changes in tumor genomic alterations that occur during the treatment of CRC.


Can advertising enhance consumers' desirable COVID-19 health behavioral intentions? The role of brand-pandemic fit.
This article explores the fit between the advertised brand and the pandemic as a potential influence on consumers' intentions to engage in socially responsible health behaviors (social distancing, mask wearing, and getting tested when exposed). In an advanced and emerging market setting we find that advertisements for brands that are perceived as high on brand-pandemic fit enhance consumers' socially desirable COVID-19 health behavioral intentions and changes in brand credibility is the mechanism that drives such intentions. Fit is especially beneficial on the intentions of consumers whose health beliefs reflect only low to moderate concern about COVID-19. Consumers with low or moderate (vs. high) COVID-19 health beliefs exhibit an increased susceptibility to the fit-desirable health behavioral intentions relationship. The results are also corroborated in an emerging market context. Together, the results establish links between brand-pandemic fit of advertisements, brand credibility, health beliefs, and consumers' intentions to engage in socially desirable health behaviors. The results suggest that advertising can play a role in encouraging desirable health behaviors and can promote consumer welfare via ads of high fit products and services that provide benefits during the pandemic in both advanced and emerging markets.


The drug ornidazole inhibits photosynthesis in a different mechanism described for protozoa and anaerobic bacteria.
Ornidazole of the 5-nitroimidazole drug family is used to treat protozoan and anaerobic bacterial infections via a mechanism that involves preactivation by reduction of the nitro group, and production of toxic derivatives and radicals. Metronidazole, another drug family member, has been suggested to affect photosynthesis by draining electrons from the electron carrier ferredoxin, thus inhibiting NADP+ reduction and stimulating radical and peroxide production. Here we show, however, that ornidazole inhibits photosynthesis via a different mechanism. While having a minute effect on the photosynthetic electron transport and oxygen photoreduction, ornidazole hinders the activity of two Calvin cycle enzymes, triose-phosphate isomerase (TPI) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH). Modeling of ornidazole's interaction with ferredoxin of the protozoan Trichomonas suggests efficient electron tunneling from the iron-sulfur cluster to the nitro group of the drug. A similar docking site of ornidazole at the plant-type ferredoxin does not exist, and the best simulated alternative does not support such efficient tunneling. Notably, TPI was inhibited by ornidazole in the dark or when electron transport was blocked by dichloromethyl diphenylurea, indicating that this inhibition was unrelated to the electron transport machinery. Although TPI and GAPDH isoenzymes are involved in glycolysis and gluconeogenesis, ornidazole's effect on respiration of photoautotrophs is moderate, thus raising its value as an efficient inhibitor of photosynthesis. The scarcity of Calvin cycle inhibitors capable of penetrating cell membranes emphasizes on the value of ornidazole for studying the regulation of this cycle.


Meshless Simulation of Multi-site Radio Frequency Catheter Ablation through the Fragile Points Method.
Computational models for radio frequency catheter ablation (RFCA) of cardiac arrhythmia have been developed and tested in conditions where a single ablation site is considered. However, in reality arrhythmic events are generated at multiple sites which are ablated during treatment. Under such conditions, heat accumulation from several ablations is expected and models should take this effect into account. Moreover, such models are solved using the Finite Element Method which requires a good quality mesh to ensure numerical accuracy. Therefore, clinical application is limited since heat accumulation effects are neglected and numerical accuracy depends on mesh quality. In this work, we propose a novel meshless computational model where tissue heat accumulation from previously ablated sites is taken into account. In this way, we aim to overcome the mesh quality restriction of the Finite Element Method and enable realistic multi-site ablation simulation. We consider a two ablation sites protocol where tissue temperature at the end of the first ablation is used as initial condition for the second ablation. The effect of the time interval between the ablation of the two sites is evaluated. The proposed method demonstrates that previous models that do not account for heat accumulation between ablations may underestimate the tissue heat distribution.Clinical relevance- The proposed computational model may be used to build and update a heat map for ablation guidance taking into account the contribution from previously ablated sites. Being a meshless model, it does not require significant input from the user during preprocessing. Therefore, it is suitable for application in a clinical setting.


Structure-activity relationships of macrolides against Mycobacterium tuberculosis.
Existing 14, 15 and 16-membered macrolide antibiotics, while effective for other bacterial infections, including some mycobacteria, have not demonstrated significant efficacy in tuberculosis. Therefore an attempt was made to optimize this class for activity against Mycobacterium tuberculosis through semisyntheses and bioassay. Approximately 300 macrolides were synthesized and screened for anti-TB activity. Structural modifications on erythromycin were carried out at positions 3, 6, 9, 11, and 12 of the 14-membered lactone ring; as well as at position 4'' of cladinose and position 2' of desosamine. In general, the synthesized macrolides belong to four subclasses: 9-oxime, 11,12-carbamate, 11,12-carbazate, and 6-O-substituted derivatives. Selected compounds were assessed for mammalian cell toxicity and in some cases were further assessed for CYP3A4 inhibition, microsome stability, in vivo tolerance and efficacy. The activity of 11,12-carbamates and carbazates as well as 9-oximes is highly influenced by the nature of the substitution at these positions. For hydrophilic macrolides, lipophilic substitution may result in enhanced potency, presumably by enhanced passive permeation through the cell envelope. This strategy, however, has limitations. Removal of the C-3 cladinose generally reduces the activity. Acetylation at C-2' or 4'' maintains potency of C-9 oximes but dramatically decreases that of 11,12-substituted compounds. Further significant increases in the potency of macrolides for M. tuberculosis may require a strategy for the concurrent reduction of ribosome methylation.


The evolving relationship of wound healing and tumor stroma.
The stroma in solid tumors contains a variety of cellular phenotypes and signaling pathways associated with wound healing, leading to the concept that a tumor behaves as a wound that does not heal. Similarities between tumors and healing wounds include fibroblast recruitment and activation, extracellular matrix (ECM) component deposition, infiltration of immune cells, neovascularization, and cellular lineage plasticity. However, unlike a wound that heals, the edges of a tumor are constantly expanding. Cell migration occurs both inward and outward as the tumor proliferates and invades adjacent tissues, often disregarding organ boundaries. The focus of our review is cancer associated fibroblast (CAF) cellular heterogeneity and plasticity and the acellular matrix components that accompany these cells. We explore how similarities and differences between healing wounds and tumor stroma continue to evolve as research progresses, shedding light on possible therapeutic targets that can result in innovative stromal-based treatments for cancer.


Real-time detection of pneumothorax using electrical impedance tomography.
Pneumothorax is a frequent complication during mechanical ventilation. Electrical impedance tomography (EIT) is a noninvasive tool that allows real-time imaging of regional ventilation. The purpose of this study was to 1) identify characteristic changes in the EIT signals associated with pneumothoraces; 2) develop and fine-tune an algorithm for their automatic detection; and 3) prospectively evaluate this algorithm for its sensitivity and specificity in detecting pneumothoraces in real time. Prospective controlled laboratory animal investigation. Experimental Pulmonology Laboratory of the University of São Paulo. Thirty-nine anesthetized mechanically ventilated supine pigs (31.0 +/- 3.2 kg, mean +/- SD). In a first group of 18 animals monitored by EIT, we either injected progressive amounts of air (from 20 to 500 mL) through chest tubes or applied large positive end-expiratory pressure (PEEP) increments to simulate extreme lung overdistension. This first data set was used to calibrate an EIT-based pneumothorax detection algorithm. Subsequently, we evaluated the real-time performance of the detection algorithm in 21 additional animals (with normal or preinjured lungs), submitted to multiple ventilatory interventions or traumatic punctures of the lung. Primary EIT relative images were acquired online (50 images/sec) and processed according to a few imaging-analysis routines running automatically and in parallel. Pneumothoraces as small as 20 mL could be detected with a sensitivity of 100% and specificity 95% and could be easily distinguished from parenchymal overdistension induced by PEEP or recruiting maneuvers. Their location was correctly identified in all cases, with a total delay of only three respiratory cycles. We created an EIT-based algorithm capable of detecting early signs of pneumothoraces in high-risk situations, which also identifies its location. It requires that the pneumothorax occurs or enlarges at least minimally during the monitoring period. Such detection was operator-free and in quasi real-time, opening opportunities for improving patient safety during mechanical ventilation.


Fighting outbreaks with bacterial genomics: case review and workflow proposal.
Disease outbreak investigation is a key aspect of public health. Whole-genome sequencing of bacterial pathogen based on new generation high-throughput sequencing technologies has facilitated outbreak investigations recently. Whilst the approach has become more affordable and accessible to research and clinical laboratories, a system for adequate and efficient analyses of genome data in the context of bacterial outbreak investigations is missing. We performed a literature review of timely genomic investigations performed during the course of bacterial outbreaks that are based on new generation sequencing technologies. Currently available bioinformatics tools for genomic analyses are also reviewed here. Genomic investigations in early stages of bacterial outbreaks have shown to provide timely information on evolutionary origin, transmission route, pathogenic potential, and resistance information of the outbreak strains and allow development of strain-specific typing methods. A systematic genomic analytical workflow is proposed here for the first time to facilitate efficient extraction of epidemiologically useful information from genome data of bacterial pathogens in future bacterial outbreak investigations. With the continuous reduction of genome sequencing cost and development of user-friendly analytical tools, it is expected that high-throughput genome sequencing will be applied routinely for timely genomic analysis in bacterial outbreaks in the near future.


Accurate diagnosis of thyroid follicular lesions from nuclear morphology using supervised learning.
Follicular lesions of the thyroid remain significant diagnostic challenges in surgical pathology and cytology. The diagnosis often requires considerable resources and ancillary tests including immunohistochemistry, molecular studies, and expert consultation. Visual analyses of nuclear morphological features, generally speaking, have not been helpful in distinguishing this group of lesions. Here we describe a method for distinguishing between follicular lesions of the thyroid based on nuclear morphology. The method utilizes an optimal transport-based linear embedding for segmented nuclei, together with an adaptation of existing classification methods. We show the method outputs assignments (classification results) which are near perfectly correlated with the clinical diagnosis of several lesion types' lesions utilizing a database of 94 patients in total. Experimental comparisons also show the new method can significantly outperform standard numerical feature-type methods in terms of agreement with the clinical diagnosis gold standard. In addition, the new method could potentially be used to derive insights into biologically meaningful nuclear morphology differences in these lesions. Our methods could be incorporated into a tool for pathologists to aid in distinguishing between follicular lesions of the thyroid. In addition, these results could potentially provide nuclear morphological correlates of biological behavior and reduce health care costs by decreasing histotechnician and pathologist time and obviating the need for ancillary testing.


Global distribution of the most prevalent deletions causing hypotonia-cystinuria syndrome.
Hypotonia-cystinuria syndrome (HCS) is a recessive disorder caused by microdeletions of SLC3A1 and PREPL on chromosome 2p21. Patients present with generalized hypotonia at birth, failure to thrive, growth retardation and cystinuria type I. While the initially described HCS families live in small regions in Belgium and France, we have now identified HCS alleles in patients and carriers from the Netherlands, Italy, Canada and United States of America. Surprisingly, among the nine deletions detected in those patients, only one novel deletion was found. Furthermore, one previously described deletion was found six times, another twice. Finally, we have investigated the frequency of both deletions using a random Belgian cohort. Given the global occurrence, HCS should be considered in the differential diagnosis of neonatal hypotonia.


Using FRET to monitor protein-induced DNA bending: the TBP-TATA complex as a model system.
Proteins that bind to DNA can elicit changes in DNA conformation, such as bending and looping, which are important signals for later events such as transcription. TATA-binding protein (TBP) is one example of a protein that elicits a conformational change in DNA; TBP binds and sharply bends its recognition sequence, which is thought to facilitate the recruitment of other protein factors. Here we describe the use of fluorescence resonance energy transfer (FRET) to evaluate DNA bending using TBP as a model system. FRET is a useful technique to measure changes in DNA conformation due to protein binding because small changes in the distance between two fluorophores (2-10 nm) translate into large changes in energy transfer.


Congenital valvular aortic stenosis: limited progression during childhood.
Congenital valvular aortic stenosis is a common congenital heart malformation. The rate of progression in childhood, however, remains to be established. We assessed the progression of peak aortic velocity before intervention as well as the frequency of intervention in paediatric patients with isolated congenital valvular aortic stenosis. A retrospective cohort study was performed in 245 consecutive patients with aortic stenosis. Both clinical and echocardiographic data were obtained. Over a period of 9.0 (SD 5.2) years (range 0.1-19.4), the mean annual increase in peak systolic velocity was 0.04 m/s/year (95% CI 0.028 to 0.056 m/s/year; p<0.001) as shown by ANOVA. 40 patients underwent a cardiac intervention shortly after their first echocardiogram. Another 33 patients underwent intervention during follow-up. Interventions were performed significantly more often in patients diagnosed at a younger age and/or with a higher peak velocity at diagnosis (p<0.001). Mortality was considerable in those diagnosed in infancy (5-year survival rate of 73% (SD 9%), whereas it was nearly absent in patients diagnosed after infancy. Most patients who died during infancy had progressive left ventricular dysfunction despite adequate relief of left ventricular outflow obstruction. Valvular aortic stenosis in the paediatric age group usually has a good prognosis beyond the neonatal period. Progression over time is usually limited, although a considerable proportion of patients need intervention shortly after initial diagnosis. Mortality, except for the neonatal age group, is nearly absent.


Cytotoxic effect of gambogic acid on SH-SY5Y neuroblastoma cells is mediated by intrinsic caspase-dependent signaling pathway.
Gambogic acid (GA) is the dry resin of Garcinia hanburyi (Guttiferae) with potent anti-tumor activity, various bioactivities, including detoxification, homeostasis, anti-inflammatory, and parasiticide, whereas the effect of this natural compound on cancer cells has not been clearly clarified. Here, we examined cellular cytotoxicity by cell viability assay and DNA fragmentation by DNA-ladder assay. Activation of different protein expressions were detected by western blot analyses. We first demonstrated that GA reduces the human SH-SY5Y neuroblastoma cell viability with IC50 of 1.28 μM at 6 h which has less toxicity in fibroblast cells. However, lower concentration GA significantly downregulated the expression of anti-apoptotic protein including Bcl-2, Bcl-xL, and Mcl-1, which also dramatically activated cleaved caspase-9 and -3 in a dose- and time-dependent manner. Consequently, GA-induced cytotoxicity was not mediated by the Fas/FasL and PI3 K/AKT/GSK-3β signaling pathway. In addition, GA-induced cells showed damage morphology which had become cell rounding, neurite retraction, membrane blebbing and shrunken in a dose- and time-dependent manner that clearly indicates this morphological change might be due to the process of apoptosis which shows fragmented DNA. Therefore, the findings presented in this study demonstrate that apoptotic effects of GA on SH-SY5Y cells are mediated by intrinsic mitochondrion-dependent caspase pathway which suggests this natural compound might be effective as an anti-cancer agent for neuroblastoma malignancies.


Delayed sudden death in an infant following an accidental fall: a case report with review of the literature.
Several controversies exist regarding ultimately lethal head injuries in small children. Death from short falls, timing of head injury, lucid intervals, presence of diffuse axonal injury (DAI), and subdural hematoma (SDH) as marker of DAI are the most recent controversial topics of debate in this evolving field of study. In this area of debate, we present a case of delayed death from a witnessed fall backwards off a bed in a 9-month-old black male child who struck his head on a concrete floor and was independently witnessed as "healthy" postfall for 72 hours until he was discovered dead in bed. Grandmother, babysitter, and mother all independently corroborated under police investigation that the child "acted and behaved normally" after the fall until death. Autopsy showed a linear nondisplaced parietal skull fracture, diastasis of adjacent occipital suture, subgaleal hemorrhage with evidence of aging, small posterior clotting SDH, marked cerebral edema, and a small tear of the midsuperior body of the corpus callosum consistent with focal axonal injury (FAI). No DAI was seen, and there were no retinal hemorrhages. All other causes of death were excluded upon thorough police and medical examiner investigation. Although this seems to be a rare phenomenon, a delayed, seemingly symptom-free interval can occur between a clinically apparent mild head injury and accidental death in a young child.


Systemic venous diameters, collapsibility indices, and right atrial measurements in normal pediatric subjects.
Compromise of right heart function is an important feature of many forms of congenital heart disease, and right atrial (RA) pressure is clinically relevant. Inferior vena cava (IVC) diameter and inspiratory collapse are indices of RA pressure, but pediatric data are lacking. RA measurements, systemic venous diameters, and Doppler filling fractions were prospectively investigated in healthy volunteer children and adolescents. The IVC was measured in its long axis just above the junction with the hepatic veins in the subxiphoid view and the superior vena cava at its junction with the right atrium in the right parasternal view. The changes in IVC diameter (IVCD) during quiet respiration and with a sniff were recorded. Hepatic venous systolic filling fraction was calculated from Doppler velocities in the first hepatic vein. RA major-axis length, area, and volume were measured from the apical four-chamber view. Three measurements of each parameter were averaged over at least three respiratory cycles. The IVC collapsibility index (IVCCI) was calculated as [(IVCDmax - IVCDmin)/IVCDmax] × 100. Substituting IVCDsniff for IVCDmin in the formula, the IVCCIsniff was calculated. Of 132 subjects enrolled, data in 120 (mean age, 8.3 ± 4.5 years) were analyzed. The maximal (expiratory) and minimal (inspiratory) diameters during free breathing were 12.1 ± 3.8 and 8.9 ± 3.8 mm for the IVC and 11.9 ± 3.4 and 7.9 ± 2.6 mm for the superior vena cava. IVCCImin and IVCCIsniff were 30 ± 13 and 47 ± 18, respectively. The RA major-axis length, area, and indexed maximal volume were 3.7 ± 0.7 cm, 10.3 ± 3.6 cm(2), and 22.3 ± 7.0 mL/m(2), respectively. Correlations of maximal superior vena cava and IVC dimensions with body surface area were slightly better than with age and much stronger than with RA volume. No significant correlation was found between IVCCIs and age, gender, or indexed RA volume. Measurement of systemic venous diameters, collapsibility indices, and RA volumes is feasible in healthy children and adolescents. Venous diameters increase predictably with growth and so must be interpreted in light of body surface area. IVCCIs and hepatic venous filling fraction compare closely with those reported in adults. Pediatric nomograms for these parameters are provided, and they should next be evaluated for relation to directly measured RA pressure in this age group.


Antibiotic Prescribing for Adults Hospitalized in the Etiology of Pneumonia in the Community Study.
Community-acquired pneumonia (CAP) 2007 guidelines from the Infectious Diseases Society of America (IDSA)/American Thoracic Society (ATS) recommend a respiratory fluoroquinolone or beta-lactam plus macrolide as first-line antibiotics for adults hospitalized with CAP. Few studies have assessed guideline-concordant antibiotic use for patients hospitalized with CAP after the 2007 IDSA/ATS guidelines. We examine antibiotics prescribed and associated factors in adults hospitalized with CAP. From January 2010 to June 2012, adults hospitalized with clinical and radiographic CAP were enrolled in a prospective Etiology of Pneumonia in the Community study across 5 US hospitals. Patients were interviewed using a standardized questionnaire, and medical charts were reviewed. Antibiotics prescribed were classified according to defined nonrecommended CAP antibiotics. We assessed factors associated with nonrecommended CAP antibiotics using logistic regression. Among enrollees, 1843 of 1874 (98%) ward and 440 of 446 (99%) ICU patients received ≥1 antibiotic ≤24 hours after admission. Ward patients were prescribed a respiratory fluoroquinolone alone (n = 613; 33%), or beta-lactam plus macrolide (n = 365; 19%), beta-lactam alone (n = 240; 13%), among other antibiotics, including vancomycin (n = 235; 13%) or piperacillin/tazobactam (n = 157; 8%) ≤24 hours after admission. Ward patients with known risk for healthcare-associated pneumonia (HCAP), recent outpatient antibiotic use, and in-hospital antibiotic use <6 hours after admission were significantly more likely to receive nonrecommended CAP antibiotics. Although more than half of ward patients received antibiotics concordant with IDSA/ATS guidelines, a number received nonrecommended CAP antibiotics, including vancomycin and piperacillin/tazobactam; risk factors for HCAP, recent outpatient antibiotic, and rapid inpatient antibiotic use contributed to this. This hypothesis-generating descriptive epidemiology analysis could help inform antibiotic stewardship efforts, reinforces the need to harmonize guidelines for CAP and HCAP, and highlights the need for improved diagnostics to better equip clinicians.


Population genomics of apricots unravels domestication history and adaptive events.
Among crop fruit trees, the apricot (Prunus armeniaca) provides an excellent model to study divergence and adaptation processes. Here, we obtain nearly 600 Armeniaca apricot genomes and four high-quality assemblies anchored on genetic maps. Chinese and European apricots form two differentiated gene pools with high genetic diversity, resulting from independent domestication events from distinct wild Central Asian populations, and with subsequent gene flow. A relatively low proportion of the genome is affected by selection. Different genomic regions show footprints of selection in European and Chinese cultivated apricots, despite convergent phenotypic traits, with predicted functions in both groups involved in the perennial life cycle, fruit quality and disease resistance. Selection footprints appear more abundant in European apricots, with a hotspot on chromosome 4, while admixture is more pervasive in Chinese cultivated apricots. Our study provides clues to the biology of selected traits and targets for fruit tree research and breeding.


Facile method to radiolabel glycol chitosan nanoparticles with (64)Cu via copper-free click chemistry for MicroPET imaging.
An efficient and straightforward method for radiolabeling nanoparticles is urgently needed to understand the in vivo biodistribution of nanoparticles. Herein, we investigated a facile and highly efficient strategy to prepare radiolabeled glycol chitosan nanoparticles with (64)Cu via a strain-promoted azide-alkyne cycloaddition strategy, which is often referred to as click chemistry. First, the azide (N3) group, which allows for the preparation of radiolabeled nanoparticles by copper-free click chemistry, was incorporated to glycol chitosan nanoparticles (CNPs). Second, the strained cyclooctyne derivative, dibenzyl cyclooctyne (DBCO) conjugated with a 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA) chelator, was synthesized for preparing the preradiolabeled alkyne complex with (64)Cu radionuclide. Following incubation with the (64)Cu-radiolabeled DBCO complex (DBCO-PEG4-Lys-DOTA-(64)Cu with high specific activity, 18.5 GBq/μmol), the azide-functionalized CNPs were radiolabeled successfully with (64)Cu, with a high radiolabeling efficiency and a high radiolabeling yield (>98%). Importantly, the radiolabeling of CNPs by copper-free click chemistry was accomplished within 30 min, with great efficiency in aqueous conditions. In addition, we found that the (64)Cu-radiolabeled CNPs ((64)Cu-CNPs) did not show any significant effect on the physicochemical properties, such as size, zeta potential, or spherical morphology. After (64)Cu-CNPs were intravenously administered to tumor-bearing mice, the real-time, in vivo biodistribution and tumor-targeting ability of (64)Cu-CNPs were quantitatively evaluated by microPET images of tumor-bearing mice. These results demonstrate the benefit of copper-free click chemistry as a facile, preradiolabeling approach to conveniently radiolabel nanoparticles for evaluating the real-time in vivo biodistribution of nanoparticles.


Idiopathic normal pressure hydrocephalus: increased supplementary motor activity accounts for improvement after CSF drainage.
In patients with idiopathic normal pressure hydrocephalus (INPH), the changes in brain function that take place in conjunction with improved behavioural performance after CSF drainage is still unknown. In this study, we use functional MRI (fMRI) to investigate the changes in cortical activity that accompany improved motor and cognitive performance after long-term external lumbar drainage (ELD) of CSF in patients with INPH. Eighteen INPH patients were initially included together with age- and sex-matched controls. Data from 11 INPH patients were analysed both before and after ELD. The average drain volume for these 11 patients was 400 ml/3 days. Brain activation was investigated by fMRI before and after the procedure on a 1.5T Philips scanner using protocols taxing motor performance (finger tapping and reaction time) and cognitive functioning (memory and attention). Behavioural data were compared using non-parametric tests at a significance level of 0.05, whereas fMRI data were analysed by statistical parametric mapping including conjunction analysis of areas with enhanced activity after drainage in patients and areas activated in controls (P < 0.005, uncorrected). Improved regions were defined as areas in the INPH brain that increased in activity after ELD with the requirement that the same areas were activated in control subjects. Following ELD, right-hand finger tapping improved from 104 +/- 38 to 117 +/- 25 (mean +/- SD) (P = 0.02). Left-hand finger tapping showed a tendency to improve, the number of keystrokes increasing from 91 +/- 40 to 105 +/- 20 (P = 0.12). Right-hand reaction time improved from 1630 +/- 566 ms to 1409 +/- 442 ms, whereas left-hand reaction time improved from 1760 +/- 600 ms to 1467 +/- 420 ms (both P-values = 0.01). Significant improvements in motor performance were accompanied by bilateral increased activation in the supplementary motor area. No improvement was found in cognitive functioning. The results suggest that motor function recovery in INPH patients after CSF removal is related to enhanced activity in medial parts of frontal motor areas considered crucial for motor planning; a finding consistent with INPH being a syndrome related to a reversible suppression of frontal periventricular cortico-basal ganglia-thalamo-cortical pathways.


ARID1A influences HDAC1/BRD4 activity, intrinsic proliferative capacity and breast cancer treatment response.
Using genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) screens to understand endocrine drug resistance, we discovered ARID1A and other SWI/SNF complex components as the factors most critically required for response to two classes of estrogen receptor-alpha (ER) antagonists. In this context, SWI/SNF-specific gene deletion resulted in drug resistance. Unexpectedly, ARID1A was also the top candidate in regard to response to the bromodomain and extraterminal domain inhibitor JQ1, but in the opposite direction, with loss of ARID1A sensitizing breast cancer cells to bromodomain and extraterminal domain inhibition. We show that ARID1A is a repressor that binds chromatin at ER cis-regulatory elements. However, ARID1A elicits repressive activity in an enhancer-specific, but forkhead box A1-dependent and active, ER-independent manner. Deletion of ARID1A resulted in loss of histone deacetylase 1 binding, increased histone 4 lysine acetylation and subsequent BRD4-driven transcription and growth. ARID1A mutations are more frequent in treatment-resistant disease, and our findings provide mechanistic insight into this process while revealing rational treatment strategies for these patients.