Molecular genetic analyses of the Tyrolean Ice Man.
An approximately 5000-year-old mummified human body was recently found in the Tyrolean Alps. The DNA from tissue samples of this Late Neolithic individual, the so-called "Ice Man," has been extracted and analyzed. The number of DNA molecules surviving in the tissue was on the order of 10 genome equivalents per gram of tissue, which meant the only multi-copy sequences could be analyzed. The degradation of the DNA made the enzymatic amplification of mitochondrial DNA fragments of more than 100 to 200 base pairs difficult. One DNA sequence of a hypervariable segment of the mitochondrial control region was determined independently in two different laboratories from internal samples of the body. This sequence showed that the mitochondrial type of the Ice Man fits into the genetic variation of contemporary Europeans and that it was most closely related to mitochondrial types determined from central and northern European populations.


The life of proteins under mechanical force.
Although much of our understanding of protein folding comes from studies of isolated protein domains in bulk, in the cellular environment the intervention of external molecular machines is essential during the protein life cycle. During the past decade single molecule force spectroscopy techniques have been extremely useful to deepen our understanding of these interventional molecular processes, as they allow for monitoring and manipulating mechanochemical events in individual protein molecules. Here, we review some of the critical steps in the protein life cycle, starting with the biosynthesis of the nascent polypeptide chain in the ribosome, continuing with the folding supported by chaperones and the translocation into different cell compartments, and ending with proteolysis in the proteasome. Along these steps, proteins experience molecular forces often combined with chemical transformations, affecting their folding and structure, which are measured or mimicked in the laboratory by the application of force with a single molecule apparatus. These mechanochemical reactions can potentially be used as targets for fighting against diseases. Inspired by these insightful experiments, we devise an outlook on the emerging field of mechanopharmacology, which reflects an alternative paradigm for drug design.


Stability of aminocaproic Acid injection admixtures in 5% dextrose injection and 0.9% sodium chloride injection.
The purpose of this study was to determine that stability of aminocaproic acid 10 and 100mg/mL admixed in 5% dextrose injection and 0.9% sodiium cholride injection in polyvinyl chloride bags over periods up to seven days at 4 deg and 23 deg C. The aminocaproic acid 250-mg/mL injection was filtered and admixed in filtered 5% dextrose injection and 0.9% sodium chloride injection to yield nominal aminocaproic acid concentrations of 10 and 100 mg/mL. Evaluations were performed initially and after one, three, and seven days of storage for physical and chemical stability. The admixtures were evaluated for physical stability using visual observation in normal fluorescent room light and a high-intensity monodirectional light beam, as well as electronic measurement of turbidity and particcle content. The chemical stability was evaluated by using a stability-indicating high performance liquid chromatography (HPLC) analytical technique. Aminocaproic acid admixtures were clear and colorless. However, the admixtures in 5% dextrose injection developed a slight yellowish discoloration after 24 hours at 23 deg C. This discoloration did not indicate substantial loss of aminocaproic acid. The HPLC evalurations found that an aminocaporic acid otency of at least 98% was retained throughout seven days in all solutions at both 4 deg and 23 deg C.


Enantioselective uptake of amino acids using an electromodulated column packed with carbon fibres modified with overoxidised polypyrrole.
A column packed with carbon fibres modified with overoxidized polypyrrole was utilised for the enantioselective uptake of glutamic acid. The polypyrrole prepared with L-glutamate as a dopant was electrochemically overoxidised to create a complementary cavity for the dopant. The target amino acid enantiomers were taken up and released by applying potential to the carbon fibre stationary phase. The uptake performance of an overoxidised polypyrrole modified carbon fibre electrode was investigated by optimising various important overoxidation parameters such as an overoxidation medium and applied potential. Under optimised conditions of overoxidation and with a column wash between the consecutive measurements, the reproducible uptake of L-glutamic acid was almost six times higher than that of D-glutamic acid. Further, in a comparison, the recognition ability of overoxidized polypyrrole with a pseudo-template cavity was examined.


Novel therapies in osteoporosis: PTH-related peptide analogs and inhibitors of sclerostin.
Bone-forming approaches to treat patients with severe osteoporosis are effective, but treatment options are limited, and there is an unmet clinical need for additional drugs. This review discusses two novel and advanced anabolic therapeutic concepts that have successfully completed phase 3 trials. Romosozumab is a monoclonal antibody that targets the Wnt inhibitor sclerostin. Two phase 3 trials (FRAME and ARCH) of romosozumab for the treatment of postmenopausal osteoporosis have been completed. Both trials successfully reached their primary endpoint by reducing vertebral fractures by 75% compared to placebo (FRAME trial) and 48% compared to alendronate (ARCH trial), respectively. Abaloparatide is a PTH-related protein (PTHrP) analog that has displayed bone anabolic activity. In the phase 3 ACTIVE trial, abaloparatide was compared to placebo and teriparatide for 18 months in postmenopausal women who had already experienced an osteoporotic fracture. Abaloparatide successfully reduced the rate of new vertebral fractures by 86% compared to placebo. Furthermore, abaloparatide achieved greater BMD increases at all measured sites compared to both placebo and teriparatide. Based on these results, abaloparatide was FDA approved in April 2017. This review discusses available data of both agents with regard to efficacy and safety as well as their possible future application.


Nanopore Sequencing: Electrical Measurements of the Code of Life.
Sequencing a single molecule of deoxyribonucleic acid (DNA) using a nanopore is a revolutionary concept because it combines the potential for long read lengths (>5 kbp) with high speed (1 bp/10 ns), while obviating the need for costly amplification procedures due to the exquisite single molecule sensitivity. The prospects for implementing this concept seem bright. The cost savings from the removal of required reagents, coupled with the speed of nanopore sequencing places the $1000 genome within grasp. However, challenges remain: high fidelity reads demand stringent control over both the molecular configuration in the pore and the translocation kinetics. The molecular configuration determines how the ions passing through the pore come into contact with the nucleotides, while the translocation kinetics affect the time interval in which the same nucleotides are held in the constriction as the data is acquired. Proteins like α-hemolysin and its mutants offer exquisitely precise self-assembled nanopores and have demonstrated the facility for discriminating individual nucleotides, but it is currently difficult to design protein structure ab initio, which frustrates tailoring a pore for sequencing genomic DNA. Nanopores in solid-state membranes have been proposed as an alternative because of the flexibility in fabrication and ease of integration into a sequencing platform. Preliminary results have shown that with careful control of the dimensions of the pore and the shape of the electric field, control of DNA translocation through the pore is possible. Furthermore, discrimination between different base pairs of DNA may be feasible. Thus, a nanopore promises inexpensive, reliable, high-throughput sequencing, which could thrust genomic science into personal medicine.


Measures used to quantify participation in childhood disability and their alignment with the family of participation-related constructs: a systematic review.
We aimed to identify measures used to assess the participation of disabled children and to map the measures' content to the family of participation-related constructs (fPRC) to inform future research and practice. Six databases were searched to identify measures used to assess participation in health, psychology, and education research. Included studies involved children aged 0 to 18 years with a permanent impairment or developmental disability and reported use of a quantitative measure of participation. A second search sought relevant literature about each identified measure (including published manuals) to allow a comprehensive understanding of the measure. Measurement constructs of frequently reported measures were then mapped to the fPRC. From an initial yield of 32 767 articles, 578 reported one or more of 118 participation measures. Of these, 51 measures were reported in more than one article (our criterion) and were therefore eligible for mapping to the fPRC. Twenty-one measures quantified aspects of participation attendance, 10 quantified aspects of involvement as discrete scales, and four quantified attendance and involvement in a manner that could not be separated. Improved understanding of participation and its related constructs is developing rapidly; thoughtful selection of measures in research is critical to further our knowledge base. The fPRC can support our rapidly evolving and expanding understanding of participation. Instruments selected to measure participation do not always align with emerging concepts. Matching research aims to a chosen measure's content will improve understanding of participation. Opportunities exist to develop validated participation measures, especially self-reported measures of involvement.


Development of Kupffer cell targeting type-I interferon for the treatment of hepatitis via inducing anti-inflammatory and immunomodulatory actions.
Because of its multifaceted anti-inflammatory and immunomodulatory effects, delivering type-I interferon to Kupffer cells has the potential to function as a novel type of therapy for the treatment of various types of hepatitis. We report herein on the preparation of a Kupffer cell targeting type-I interferon, an albumin-IFNα2b fusion protein that contains highly mannosylated N-linked oligosaccharide chains, Man-HSA(D494N)-IFNα2b, attached by combining albumin fusion technology and site-directed mutagenesis. The presence of this unique oligosaccharide permits the protein to be efficiently, rapidly and preferentially distributed to Kupffer cells. Likewise IFNα2b, Man-HSA(D494N)-IFNα2b caused a significant induction in the mRNA levels of IL-10, IL-1Ra, PD-L1 in RAW264.7 cells and mouse isolated Kupffer cells, and these inductions were largely inhibited by blocking the interferon receptor. These data indicate that Man-HSA(D494N)-IFNα2b retained the biological activities of type-I interferon. Man-HSA(D494N)-IFNα2b significantly inhibited liver injury in Concanavalin A (Con-A)-induced hepatitis model mice, and consequently improved their survival rate. Moreover, the post-administration of Man-HSA(D494N)-IFNα2b at 2 h after the Con-A challenge also exerted hepato-protective effects. In conclusion, this proof-of-concept study demonstrates the therapeutic effectiveness and utility of Kupffer cell targeting type-I interferon against hepatitis via its anti-inflammatory and immunomodulatory actions.


Pax1 is expressed during development of the thymus epithelium and is required for normal T-cell maturation.
Pax1 is a transcriptional regulatory protein expressed during mouse embryogenesis and has been shown to have an important function in vertebral column development. Expression of Pax1 mRNA in the embryonic thymus has been reported previously. Here we show that Pax1 protein expression in thymic epithelial cells can be detected throughout thymic development and in the adult. Expression starts in the early endodermal epithelium lining the foregut region and includes the epithelium of the third pharyngeal pouch, a structure giving rise to part of the thymus epithelium. In early stages of thymus development a large proportion of thymus cells expresses Pax1. With increasing age, the proportion of Pax1-expressing cells is reduced and in the adult mouse only a small fraction of cortical thymic stromal cells retains strong Pax1 expression. Expression of Pax1 in thymus epithelium is necessary for establishing the thymus microenvironment required for normal T cell maturation. Mutations in the Pax-1 gene in undulated mice affect not only the total size of the thymus but also the maturation of thymocytes. The number of thymocytes is reduced about 2- to 5-fold, affecting mainly the CD4+8+ immature and CD4+ mature thymocyte subsets. The expression levels of major thymocyte surface markers remains unchanged with the exception of Thy-1 which was found to be expressed at 3- to 4-fold higher levels.


Post-gel polymerisation shrinkage profiling of polymer biomaterials using a chirped fibre Bragg grating.
A strain profile measurement technique using a chirped fibre Bragg grating (CFBG) sensor by implementing an integration of differences (IOD) method is reported in this paper. Using the IOD method the spatial distribution of strain along the length of the CFBG is extracted from its power reflectance spectra. As a proof of concept demonstration, the developed technique is applied to measure the polymerisation shrinkage strain profile of a photo-cured polymer dental composite which exhibits a non-uniform strain distribution attributed to the curing lamp characteristics. The result from the CFBG technique is compared with that of an FBG array embedded in the dental composite and is correlated with the degree of conversion of the material which also depends on the curing lamp intensity distribution. This technology will have significant impact and applications in a range of medical, materials and engineering areas where strain or temperature gradient profile measurement is required in smaller scales.


NATURAL LANGUAGE PROCESSING BASED MACHINE LEARNING MODEL USING CARDIAC MRI REPORTS TO IDENTIFY HYPERTROPHIC CARDIOMYOPATHY PATIENTS.
Hypertrophic Cardiomyopathy (HCM) is the most common genetic heart disease in the US and is known to cause sudden death (SCD) in young adults. While significant advancements have been made in HCM diagnosis and management, there is a need to identify HCM cases from electronic health record (EHR) data to develop automated tools based on natural language processing guided machine learning (ML) models for accurate HCM case identification to improve management and reduce adverse outcomes of HCM patients. Cardiac Magnetic Resonance (CMR) Imaging, plays a significant role in HCM diagnosis and risk stratification. CMR reports, generated by clinician annotation, offer rich data in the form of cardiac measurements as well as narratives describing interpretation and phenotypic description. The purpose of this study is to develop an NLP-based interpretable model utilizing impressions extracted from CMR reports to automatically identify HCM patients. CMR reports of patients with suspected HCM diagnosis between the years 1995 to 2019 were used in this study. Patients were classified into three categories of yes HCM, no HCM and, possible HCM. A random forest (RF) model was developed to predict the performance of both CMR measurements and impression features to identify HCM patients. The RF model yielded an accuracy of 86% (608 features) and 85% (30 features). These results offer promise for accurate identification of HCM patients using CMR reports from EHR for efficient clinical management transforming health care delivery for these patients.


A refined prediction method for gel retardation of DNA oligonucleotides from dinucleotide step parameters: reconciliation of DNA bending models with crystal structure data.
The development and assessment of a prediction method for gel retardation and sequence dependent curvature of DNA based on dinulcleotide step parameters are described. The method is formulated using the Babcock-Olson equations for base pair step geometry (1) and employs Monte Carlo simulated annealing for parameter optimization against experimental data. The refined base pair step parameters define a stuctural construct which, when the width of observed parameter distributions is taken into account, is consistent with the results of DNA oligonucleotide crystal structures. The predictive power of the method is demonstrated and tested via comparisons with DNA bending data on sets of sequences not included in the training set, including A-tracts with and without periodic helix phasing, phased A4T4 and T4A4 motifs, a sequence with a phased GGGCCC motif, some "unconventional" helix phasing sequences, and three short fragments of kinetoplast DNA from Crithidia fasiculata that exhibit significantly different behavior on non-denaturing polyacrylamide gels. The nature of the structural construct produced by the methodology is discussed with respect to static and dynamic models of structure and representations of bending and bendability. An independent theoretical account of sequence dependent chemical footprinting results is provided. Detailed analysis of sequences with A-tract induced axis bending forms the basis for a critical discussion of the applicability of wedge models,junction models and non A-tract, general sequence models for understanding the origin of DNA curvature at the molecular level.


Novel Lethal Form of Congenital Hypopituitarism Associated With the First Recessive LHX4 Mutation.
LHX4 encodes a member of the LIM-homeodomain family of transcription factors that is required for normal development of the pituitary gland. To date, only incompletely penetrant heterozygous mutations in LHX4 have been described in patients with variable combined pituitary hormone deficiencies. To report a unique family with a novel recessive variant in LHX4 associated with a lethal form of congenital hypopituitarism that was identified through screening a total of 97 patients. We screened 97 unrelated patients with combined pituitary hormone deficiency, including 65% with an ectopic posterior pituitary, for variants in the LHX4 gene using Sanger sequencing. Control databases (1000 Genomes, dbSNP, Exome Variant Server, ExAC Browser) were consulted upon identification of variants. We identified the first novel homozygous missense variant (c.377C>T, p.T126M) in two deceased male patients of Pakistani origin with severe panhypopituitarism associated with anterior pituitary aplasia and posterior pituitary ectopia. Both were born small for gestational age with a small phallus, undescended testes, and mid-facial hypoplasia. The parents' first-born child was a female with mid-facial hypoplasia (DNA was unavailable). Despite rapid commencement of hydrocortisone and T4 in the brothers, all three children died within the first week of life. The LHX4(p.T126M) variant is located within the LIM2 domain, in a highly conserved location. The absence of homozygosity for the variant in over 65 000 controls suggests that it is likely to be responsible for the phenotype. We report, for the first time to our knowledge, a novel homozygous mutation in LHX4 associated with a lethal phenotype, implying that recessive mutations in LHX4 may be incompatible with life.


Catastrophic Risk: Waking Up to the Reality of a Pandemic?
Will a major shock awaken the US citizens to the threat of catastrophic pandemic risk? Using a natural experiment administered both before and after the 2014 West African Ebola Outbreak, our evidence suggests "no." Our results show that prior to the Ebola scare, the US citizens were relatively complacent and placed a low relative priority on public spending to prepare for a pandemic disease outbreak relative to an environmental disaster risk (e.g., Fukushima) or a terrorist attack (e.g., 9/11). After the Ebola scare, the average citizen did not over-react to the risk. This flat reaction was unexpected given the well-known availability heuristic-people tend to over-weigh judgments of events more heavily toward more recent information. In contrast, the average citizen continued to value pandemic risk less relative to terrorism or environmental risk.


The standard operating procedure of the DOE-JGI Microbial Genome Annotation Pipeline (MGAP v.4).
The DOE-JGI Microbial Genome Annotation Pipeline performs structural and functional annotation of microbial genomes that are further included into the Integrated Microbial Genome comparative analysis system. MGAP is applied to assembled nucleotide sequence datasets that are provided via the IMG submission site. Dataset submission for annotation first requires project and associated metadata description in GOLD. The MGAP sequence data processing consists of feature prediction including identification of protein-coding genes, non-coding RNAs and regulatory RNA features, as well as CRISPR elements. Structural annotation is followed by assignment of protein product names and functions.


Dichotomal functions of phosphorylated and unphosphorylated STAT1 in hepatocellular carcinoma.
Interferons (IFNs) with antiviral and immune-stimulatory functions have been widely used in prevention and treatment of hepatocellular carcinoma (HCC). Signal transducer and activator of transcription 1 (STAT1) is a key element of the IFN signaling, and the function of STAT1 is critically determined by its phosphorylation state. This study aims to understand the functions of phosphorylated (p-) and unphosphorylated (u-) STAT1 in HCC. We found that u-STAT1 is significantly elevated in patient HCC tumor tissues and predominantly expressed in cytoplasm; while p-STAT1 is absent. Loss of u-STAT1 potently arrested cell cycle and inhibited cell growth in HCC cells. Induction of p-STAT1 by IFN-α treatment effectively triggers the expression of interferon-stimulated genes (ISGs), but has moderate effect on HCC cell growth. Interestingly, both u-STAT1 and p-STAT1 are induced by IFN-α, through with distinct time-dependent process. Furthermore, the ISG induction patterns mediated by p-STAT1 and u-STAT1 are also distinct. Importantly, artificial blocking of the induction of u-STAT1, but not p-STAT1, sensitizes HCC cells to treatment of IFNs. Therefore, p-STAT1 and u-STAT1 exert dichotomal functions and coordinately regulate the responsiveness to IFN treatment in HCC. KEY MESSAGES: STAT1 is upregulated and predominantly presented as u-STAT1 in HCC, while p-STAT1 is absent. U-STAT1 sustains but p-STAT1 inhibits HCC growth. The dynamic change of phosphorylation state of STAT1 control the responsiveness to IFN treatment.


Human health implications of organic food and organic agriculture: a comprehensive review.
This review summarises existing evidence on the impact of organic food on human health. It compares organic vs. conventional food production with respect to parameters important to human health and discusses the potential impact of organic management practices with an emphasis on EU conditions. Organic food consumption may reduce the risk of allergic disease and of overweight and obesity, but the evidence is not conclusive due to likely residual confounding, as consumers of organic food tend to have healthier lifestyles overall. However, animal experiments suggest that identically composed feed from organic or conventional production impacts in different ways on growth and development. In organic agriculture, the use of pesticides is restricted, while residues in conventional fruits and vegetables constitute the main source of human pesticide exposures. Epidemiological studies have reported adverse effects of certain pesticides on children's cognitive development at current levels of exposure, but these data have so far not been applied in formal risk assessments of individual pesticides. Differences in the composition between organic and conventional crops are limited, such as a modestly higher content of phenolic compounds in organic fruit and vegetables, and likely also a lower content of cadmium in organic cereal crops. Organic dairy products, and perhaps also meats, have a higher content of omega-3 fatty acids compared to conventional products. However, these differences are likely of marginal nutritional significance. Of greater concern is the prevalent use of antibiotics in conventional animal production as a key driver of antibiotic resistance in society; antibiotic use is less intensive in organic production. Overall, this review emphasises several documented and likely human health benefits associated with organic food production, and application of such production methods is likely to be beneficial within conventional agriculture, e.g., in integrated pest management.


PDX-1 is required for pancreatic outgrowth and differentiation of the rostral duodenum.
It has been proposed that the Xenopus homeobox gene, XlHbox8, is involved in endodermal differentiation during pancreatic and duodenal development (Wright, C.V.E., Schnegelsberg, P. and De Robertis, E.M. (1988). Development 105, 787-794). To test this hypothesis directly, gene targeting was used to make two different null mutations in the mouse XlHbox8 homolog, pdx-1. In the first, the second pdx-1 exon, including the homeobox, was replaced by a neomycin resistance cassette. In the second, a lacZ reporter was fused in-frame with the N terminus of PDX-1, replacing most of the homeodomain. Neonatal pdx-1 -/- mice are apancreatic, in confirmation of previous reports (Jonsson, J., Carlsson, L., Edlund, T. and Edlund, H. (1994). Nature 371, 606-609). However, the pancreatic buds do form in homozygous mutants, and the dorsal bud undergoes limited proliferation and outgrowth to form a small, irregularly branched, ductular tree. This outgrowth does not contain insulin or amylase-positive cells, but glucagon-expressing cells are found. The rostral duodenum shows a local absence of the normal columnar epithelial lining, villi, and Brunner's glands, which are replaced by a GLUT2-positive cuboidal epithelium resembling the bile duct lining. Just distal of the abnormal epithelium, the numbers of enteroendocrine cells in the villi are greatly reduced. The PDX-1/beta-galactosidase fusion allele is expressed in pancreatic and duodenal cells in the absence of functional PDX-1, with expression continuing into perinatal stages with similar boundaries and expression levels. These results offer additional insight into the role of pdx-1 in the determination and differentiation of the posterior foregut, particularly regarding the proliferation and differentiation of the pancreatic progenitors.


Dynamic acetylation of all lysine-4 trimethylated histone H3 is evolutionarily conserved and mediated by p300/CBP.
Histone modifications are reported to show different behaviors, associations, and functions in different genomic niches and organisms. We show here that rapid, continuous turnover of acetylation specifically targeted to all K4-trimethylated H3 tails (H3K4me3), but not to bulk histone H3 or H3 carrying other methylated lysines, is a common uniform characteristic of chromatin biology in higher eukaryotes, being precisely conserved in human, mouse, and Drosophila. Furthermore, dynamic acetylation targeted to H3K4me3 is mediated by the same lysine acetyltransferase, p300/cAMP response element binding (CREB)-binding protein (CBP), in both mouse and fly cells. RNA interference or chemical inhibition of p300/CBP using a newly discovered small molecule inhibitor, C646, blocks dynamic acetylation of H3K4me3 globally in mouse and fly cells, and locally across the promoter and start-site of inducible genes in the mouse, thereby disrupting RNA polymerase II association and the activation of these genes. Thus, rapid dynamic acetylation of all H3K4me3 mediated by p300/CBP is a general, evolutionarily conserved phenomenon playing an essential role in the induction of immediate-early (IE) genes. These studies indicate a more global function of p300/CBP in mediating rapid turnover of acetylation of all H3K4me3 across the nuclei of higher eukaryotes, rather than a tight promoter-restricted function targeted by complex formation with specific transcription factors.


Sampling Strategies for Fast Updating of Gaussian Markov Random Fields.
Gaussian Markov random fields (GMRFs) are popular for modeling dependence in large areal datasets due to their ease of interpretation and computational convenience afforded by the sparse precision matrices needed for random variable generation. Typically in Bayesian computation, GMRFs are updated jointly in a block Gibbs sampler or componentwise in a single-site sampler via the full conditional distributions. The former approach can speed convergence by updating correlated variables all at once, while the latter avoids solving large matrices. We consider a sampling approach in which the underlying graph can be cut so that conditionally independent sites are updated simultaneously. This algorithm allows a practitioner to parallelize updates of subsets of locations or to take advantage of 'vectorized' calculations in a high-level language such as R. Through both simulated and real data, we demonstrate computational savings that can be achieved versus both single-site and block updating, regardless of whether the data are on a regular or an irregular lattice. The approach provides a good compromise between statistical and computational efficiency and is accessible to statisticians without expertise in numerical analysis or advanced computing.


Induced sputum in children: feasibility, repeatability, and relation of findings to asthma severity.
The collection of induced sputum provides a non-invasive method of investigating airway inflammation. Few studies have been performed in children, so a study was undertaken to determine the feasibility of sputum induction, the repeatability of eosinophil counts and sputum eosinophil cationic protein (ECP) levels, and the relation of these to current asthma severity. For comparison, serum ECP levels were also measured. In a cross sectional study of children aged 5-15 years, 27 healthy children and 60 with asthma underwent sputum induction using inhaled nebulised hypertonic saline. The whole sputum sample was used for analysis. Ten children with stable asthma repeated the procedure within 10 days. A satisfactory sample (>500 non-squamous cells) was obtained in 61% of children with asthma and in 60% of healthy controls. The limits of agreement within subjects ranged from a 0.68 to 2.8 fold difference for eosinophil differential counts and from 0.38 to 4.4 fold for sputum ECP. Despite a median of 42% squamous cells, significant differences were found between asthma and healthy controls for the eosinophil differential count (p = 0.0004), total eosinophil counts (p = 0.03), and sputum ECP level (p = 0.0001). Overall, there was no correlation between any marker of airway inflammation and asthma severity, however expressed, including lung function. Sputum induction is only possible in a proportion of children. The repeatability of sputum cell counts and ECP levels, measured in a small number of children, was similar to that reported in adults. Sputum analysis revealed no evidence of airway inflammation in a number of highly symptomatic children with asthma.


Low-Frequency Synonymous Coding Variation in CYP2R1 Has Large Effects on Vitamin D Levels and Risk of Multiple Sclerosis.
Vitamin D insufficiency is common, correctable, and influenced by genetic factors, and it has been associated with risk of several diseases. We sought to identify low-frequency genetic variants that strongly increase the risk of vitamin D insufficiency and tested their effect on risk of multiple sclerosis, a disease influenced by low vitamin D concentrations. We used whole-genome sequencing data from 2,619 individuals through the UK10K program and deep-imputation data from 39,655 individuals genotyped genome-wide. Meta-analysis of the summary statistics from 19 cohorts identified in CYP2R1 the low-frequency (minor allele frequency = 2.5%) synonymous coding variant g.14900931G>A (p.Asp120Asp) (rs117913124[A]), which conferred a large effect on 25-hydroxyvitamin D (25OHD) levels (-0.43 SD of standardized natural log-transformed 25OHD per A allele; p value = 1.5 × 10-88). The effect on 25OHD was four times larger and independent of the effect of a previously described common variant near CYP2R1. By analyzing 8,711 individuals, we showed that heterozygote carriers of this low-frequency variant have an increased risk of vitamin D insufficiency (odds ratio [OR] = 2.2, 95% confidence interval [CI] = 1.78-2.78, p = 1.26 × 10-12). Individuals carrying one copy of this variant also had increased odds of multiple sclerosis (OR = 1.4, 95% CI = 1.19-1.64, p = 2.63 × 10-5) in a sample of 5,927 case and 5,599 control subjects. In conclusion, we describe a low-frequency CYP2R1 coding variant that exerts the largest effect upon 25OHD levels identified to date in the general European population and implicates vitamin D in the etiology of multiple sclerosis.


Oropharyngeal candidiasis in patients with HIV: suggested guidelines for therapy.
The high frequency of oropharyngeal candidiasis in immunocompromised patients has led many institutions to develop protocols to guide the use of antifungal agents in the treatment of this opportunistic infection. However, few specific recommendations have been made for directing the management of oropharyngeal candidiasis in patients infected with HIV. To meet this need, a panel of experts representing a variety of disciplines met to formulate a consensus and devise a treatment strategy for clinical application. Among other recommendations, the algorithm calls for use of a topical agent for the treatment of initial and recurring oropharyngeal candidiasis in HIV-infected patients, provided there is no esophageal involvement, patients' CD4+ lymphocyte cell count is >50 cells/mm3, and they are currently receiving or expected to receive effective antiretroviral treatment. For episodes of oropharyngeal candidiasis with concurrent esophageal involvement or where patients have a CD4+ cell count of <50 cells/mm3, are not receiving or anticipating highly active antiretroviral therapy (HAART), and have a high viral load, the algorithm suggests a systemic oral azole as the more appropriate treatment choice. Acute treatment of all oropharyngeal candidiasis episodes is preferred. Chronic suppressive antifungal treatment is to be avoided in recognition of the potential for the development of drug-resistant infection.


Comparison of Prediction Rules and Clinician Suspicion for Identifying Children With Clinically Important Brain Injuries After Blunt Head Trauma.
Children with minor head trauma frequently present to emergency departments (EDs). Identifying those with traumatic brain injuries (TBIs) can be difficult, and it is unknown whether clinical prediction rules outperform clinician suspicion. Our primary objective was to compare the test characteristics of the Pediatric Emergency Care Applied Research Network (PECARN) TBI prediction rules to clinician suspicion for identifying children with clinically important TBIs (ciTBIs) after minor blunt head trauma. Our secondary objective was to determine the reasons for obtaining computed tomography (CT) scans when clinical suspicion of ciTBI was low. This was a planned secondary analysis of a previously conducted observational cohort study conducted in PECARN to derive and validate clinical prediction rules for ciTBI among children with minor blunt head trauma in 25 PECARN EDs. Clinicians recorded their suspicion of ciTBI before CT as <1, 1-5, 6-10, 11-50, or >50%. We defined ciTBI as 1) death from TBI, 2) neurosurgery, 3) intubation for more than 24 hours for TBI, or 4) hospital admission of 2 nights or more associated with TBI on CT. To avoid overfitting of the prediction rules, we performed comparisons of the prediction rules and clinician suspicion on the validation group only. On the validation group, we compared the test accuracies of clinician suspicion > 1% versus having at least one predictor in the PECARN TBI age-specific prediction rules for identifying children with ciTBIs (one rule for children <2 years [preverbal], the other rule for children >2 years [verbal]). In the parent study, we enrolled 8,627 children to validate the prediction rules, after enrolling 33,785 children to derive the prediction rules. In the validation group, clinician suspicion of ciTBI was recorded in 8,496/8,627 (98.5%) patients, and 87 (1.0%) had ciTBIs. CT scans were obtained in 2,857 (33.6%) patients in the validation group for whom clinician suspicion of ciTBI was recorded, including 2,099/7,688 (27.3%) of those with clinician suspicion of ciTBI of <1% and 758/808 (93.8%) of those with clinician suspicion >1%. The PECARN prediction rules were significantly more sensitive than clinician suspicion >1% of ciTBI for preverbal (100% [95% confidence interval {CI} = 86.3% to 100%] vs. 60.0% [95% CI = 38.7% to 78.9%]) and verbal children (96.8% [95% CI = 88.8% to 99.6%] vs. 64.5% [95% CI = 51.3% to 76.3%]). Prediction rule specificity, however, was lower than clinician suspicion >1% for preverbal children (53.6% [95% CI = 51.5% to 55.7%] vs. 92.4% [95% CI = 91.2% to 93.5%]) and verbal children (58.2% [95% CI = 56.9% to 59.4%] vs. 90.6% [95% CI = 89.8% to 91.3%]). Of the 7,688 patients in the validation group with clinician suspicion recorded as <1%, CTs were nevertheless obtained in 2,099 (27.3%). Three of 16 (18.8%) patients undergoing neurosurgery had clinician suspicion of ciTBI <1%. The PECARN TBI prediction rules had substantially greater sensitivity, but lower specificity, than clinician suspicion of ciTBI for children with minor blunt head trauma. Because CT ordering did not follow clinician suspicion of <1%, these prediction rules can augment clinician judgment and help obviate CT ordering for children at very low risk of ciTBI.


Bridging science and traditional knowledge to assess cumulative impacts of stressors on ecosystem health.
Cumulative environmental impacts driven by anthropogenic stressors lead to disproportionate effects on indigenous communities that are reliant on land and water resources. Understanding and counteracting these effects requires knowledge from multiple sources. Yet the combined use of Traditional Knowledge (TK) and Scientific Knowledge (SK) has both technical and philosophical hurdles to overcome, and suffers from inherently imbalanced power dynamics that can disfavour the very communities it intends to benefit. In this article, we present a 'two-eyed seeing' approach for co-producing and blending knowledge about ecosystem health by using an adapted Bayesian Belief Network for the Slave River and Delta region in Canada's Northwest Territories. We highlight how bridging TK and SK with a combination of field data, interview transcripts, existing models, and expert judgement can address key questions about ecosystem health when considerable uncertainty exists. SK indicators (e.g., bird counts, mercury in fish, water depth) were graded as moderate, whereas TK indicators (e.g., bird usage, fish aesthetics, changes to water flow) were graded as being poor in comparison to the past. SK indicators were predominantly spatial (i.e., comparing to other locations) while the TK indicators were predominantly temporal (i.e., comparing across time). After being populated by 16 experts (local harvesters, Elders, governmental representatives, and scientists) using both TK and SK, the model output reported low probabilities that the social-ecological system is healthy as it used to be. We argue that it is novel and important to bridge TK and SK to address the challenges of environmental change such as the cumulative impacts of multiple stressors on ecosystems and the services they provide. This study presents a critical social-ecological tool for widening the evidence-base to a more holistic understanding of the system dynamics of multiple environmental stressors in ecosystems and for developing more effective knowledge-inclusive partnerships between indigenous communities, researchers and policy decision-makers. This represents new transformational empirical insights into how wider knowledge discourses can contribute to more effective adaptive co-management governance practices and solutions for the resilience and sustainability of ecosystems in Northern Canada and other parts of the world with strong indigenous land tenure.


Differential Effects of Testosterone and Estradiol on Clitoral Function: An Experimental Study in Rats.
Female sexual response is a complex phenomenon in which psychological, neurologic, and vascular mechanisms and hormonal factors interact. During the arousal phase, they cooperate to increase genital blood flow, thus inducing engorgement of the clitoris and lubrication of the vagina. Regulation of vascular and non-vascular smooth muscle tone is the crucial event in the erectile process. Preclinical studies have suggested that nitric oxide (NO) is the main vasodilator neurotransmitter modulating, through the second messenger cyclic guanosine monophosphate (cGMP), clitoral flow vessels. To investigate the effects of sexual steroid hormones on pro-erectile and relaxant (mediated by NO and cGMP) and anti-erectile and contractile (mediated by ras homolog gene family member A [RhoA] and Rho-associated protein kinase [ROCK]) mechanisms in the clitoris using a validated animal model of female ovariectomized Sprague-Dawley rats. Subgroups of ovariectomized rats were treated with 17β-estradiol, progesterone, testosterone, or testosterone and letrozole for 6 weeks. The experimental groups were compared with a control group of intact rats. Sex steroids plasma levels were assessed and in vitro contractility studies were carried out in order to investigate the effect of ovariectomy and in vivo treatments on clitoris smooth muscle activity. Smooth muscle cells (SMCs) from rat clitoral biopsies were isolated and characterized. RhoA activity was determined in SMCs cell cultures. RNA from tissues and cells was analyzed by quantitative real-time RT-PCR. Using real-time polymerase chain reaction, testosterone treatment upregulated the expression of NO-mediated pathway genes (endothelial and neuronal NO synthase, guanylate cyclase soluble subunit-α3, guanylate cyclase soluble subunit-β3, cGMP-dependent protein kinase 1, and phosphodiesterase type 5). Conversely, estrogen replacement upregulated the expression of calcium-sensitizing RhoA-ROCK pathway genes. In vitro contractility studies were performed on phenylephrine pre-contracted clitoris strips. Ovariectomy resulted in a decreased responsiveness to Y-27632, a ROCK inhibitor, which was fully restored by 17β-estradiol supplementation. To further examine the effect of 17β-estradiol on the RhoA-ROCK pathway, smooth muscle cells were isolated from rat clitoris and their migration capacity was evaluated. Collectively, these data demonstrate that testosterone improves the relaxation of vascular smooth muscle cells through the NO-cGMP pathway, and that testosterone and 17β-estradiol are necessary to maintain a functional contractile and relaxant machinery in the clitoris. This new concept might provide support for the concomitant use of estrogen and testosterone during the treatment of sexual arousal disorders related to hormonal imbalance or insufficiency.


Assessing the labile arsenic pool in contaminated paddy soils by isotopic dilution techniques and simple extractions.
Arsenic (As) contamination of paddy soils threatens rice cultivation and the health of populations relying on rice as a staple crop. In the present study, isotopic dilution techniques were used to determine the chemically labile (E value) and phytoavailable (L value) pools of As in a range of paddy soils from Bangladesh, India, and China and two arable soils from the UK varying in the degree and sources of As contamination. The E value accounted for 6.2-21.4% of the total As, suggesting that a large proportion of soil As is chemically nonlabile. L values measured with rice grown under anaerobic conditions were generally larger than those under aerobic conditions, indicating increased potentially phytoavailable pool of As in flooded soils. In an incubation study, As was mobilized into soil pore water mainly as arsenite under flooded conditions, with Bangladeshi soils contaminated by irrigation of groundwater showing a greater potential of As mobilization than other soils. Arsenic mobilization was best predicted by phosphate-extractable As in the soils.


Relevance of aquatic environments for hominins: a case study from Trinil (Java, Indonesia).
Knowledge about dietary niche is key to understanding hominin evolution, since diet influences body proportions, brain size, cognition, and habitat preference. In this study we provide ecological context for the current debate on modernity (or not) of aquatic resource exploitation by hominins. We use the Homo erectus site of Trinil as a case study to investigate how research questions on possible dietary relevance of aquatic environments can be addressed. Faunal and geochemical analysis of aquatic fossils from Trinil Hauptknochenschicht (HK) fauna demonstrate that Trinil at approximately 1.5Ma contained near-coastal rivers, lakes, swamp forests, lagoons, and marshes with minor marine influence, laterally grading into grasslands. Trinil HK environments yielded at least eleven edible mollusc species and four edible fish species that could be procured with no or minimal technology. We demonstrate that, from an ecological point of view, the default assumption should be that omnivorous hominins in coastal habitats with catchable aquatic fauna could have consumed aquatic resources. The hypothesis of aquatic exploitation can be tested with taphonomic analysis of aquatic fossils associated with hominin fossils. We show that midden-like characteristics of large bivalve shell assemblages containing Pseudodon and Elongaria from Trinil HK indicate deliberate collection by a selective agent, possibly hominin.


Microbial community composition and endolith colonization at an Arctic thermal spring are driven by calcite precipitation.
Environmental conditions shape community composition. Arctic thermal springs provide an opportunity to study how environmental gradients can impose strong selective pressures on microbial communities and provide a continuum of niche opportunities. We use microscopic and molecular methods to conduct a survey of microbial community composition at Troll Springs on Svalbard, Norway, in the high Arctic. Microorganisms there exist under a wide range of environmental conditions: in warm water as periphyton, in moist granular materials, and in cold, dry rock as endoliths. Troll Springs has two distinct ecosystems, aquatic and terrestrial, together in close proximity, with different underlying environmental factors shaping each microbial community. Periphyton are entrapped during precipitation of calcium carbonate from the spring's waters, providing microbial populations that serve as precursors for the development of endolithic communities. This process differs from most endolith colonization, in which the rock predates the communities that colonize it. Community composition is modulated as environmental conditions change within the springs. At Troll, the aquatic environments show a small number of dominant operational taxonomic units (OTUs) that are specific to each sample. The terrestrial environments show a more even distribution of OTUs common to multiple samples.


DMRG-CASPT2 study of the longitudinal static second hyperpolarizability of all-trans polyenes.
We have implemented internally contracted complete active space second order perturbation theory (CASPT2) with the density matrix renormalization group (DMRG) as active space solver [Y. Kurashige and T. Yanai, J. Chem. Phys. 135, 094104 (2011)]. Internally contracted CASPT2 requires to contract the generalized Fock matrix with the 4-particle reduced density matrix (4-RDM) of the reference wavefunction. The required 4-RDM elements can be obtained from 3-particle reduced density matrices (3-RDM) of different wavefunctions, formed by symmetry-conserving single-particle excitations op top of the reference wavefunction. In our spin-adapted DMRG code chemps2 https://github.com/sebwouters/chemps2, we decompose these excited wavefunctions as spin-adapted matrix product states and calculate their 3-RDM in order to obtain the required contraction of the generalized Fock matrix with the 4-RDM of the reference wavefunction. In this work, we study the longitudinal static second hyperpolarizability of all-trans polyenes C2nH2n+2 [n = 4-12] in the cc-pVDZ basis set. DMRG-SCF and DMRG-CASPT2 yield substantially lower values and scaling with system size compared to RHF and MP2, respectively.


A compartmentalized mathematical model of the β1-adrenergic signaling system in mouse ventricular myocytes.
The β1-adrenergic signaling system plays an important role in the functioning of cardiac cells. Experimental data shows that the activation of this system produces inotropy, lusitropy, and chronotropy in the heart, such as increased magnitude and relaxation rates of [Ca(2+)]i transients and contraction force, and increased heart rhythm. However, excessive stimulation of β1-adrenergic receptors leads to heart dysfunction and heart failure. In this paper, a comprehensive, experimentally based mathematical model of the β1-adrenergic signaling system for mouse ventricular myocytes is developed, which includes major subcellular functional compartments (caveolae, extracaveolae, and cytosol). The model describes biochemical reactions that occur during stimulation of β1-adrenoceptors, changes in ionic currents, and modifications of Ca(2+) handling system. Simulations describe the dynamics of major signaling molecules, such as cyclic AMP and protein kinase A, in different subcellular compartments; the effects of inhibition of phosphodiesterases on cAMP production; kinetics and magnitudes of phosphorylation of ion channels, transporters, and Ca(2+) handling proteins; modifications of action potential shape and duration; magnitudes and relaxation rates of [Ca(2+)]i transients; changes in intracellular and transmembrane Ca(2+) fluxes; and [Na(+)]i fluxes and dynamics. The model elucidates complex interactions of ionic currents upon activation of β1-adrenoceptors at different stimulation frequencies, which ultimately lead to a relatively modest increase in action potential duration and significant increase in [Ca(2+)]i transients. In particular, the model includes two subpopulations of the L-type Ca(2+) channels, in caveolae and extracaveolae compartments, and their effects on the action potential and [Ca(2+)]i transients are investigated. The presented model can be used by researchers for the interpretation of experimental data and for the developments of mathematical models for other species or for pathological conditions.


Mechanism of spontaneous activation of B cells in patients with systemic lupus erythematosus. Analysis with anti-class II antibody.
The mechanism of spontaneous activation of B cells in patients with systemic lupus erythematosus (SLE) was analyzed by using anti-class II monoclonal antibodies in vitro. B cells from SLE patients showed enhanced proliferation and Ig production by in vitro culture without any stimulation. The number of Ig-producing cells increased during a 5-day culture period, but the addition of anti-class II antibodies such as anti-HLA-DR, DQ, or DP monoclonal antibodies inhibited these B cell responses in a dose-dependent manner. Anti-class I and anti-B1 antibody gave no effect. The inhibitory effect of anti-class II antibodies on B cell responses became more remarkable when B cells were cultured on a longer period. By a Percoll gradient density centrifugation, Ig-producing cells were enriched in the lower density fraction, but became depleted in the higher density fraction. However, B cells of the higher density fraction developed into Ig-producing cells after 5 days of culture and anti-class II antibodies inhibited this development. When mitomycin C- and cycloheximide-treated B cells were added to the in vitro culture of B cells as a stimulator, B cell responses were enhanced in a dose-dependent manner. T cells treated with mitomycin C and cycloheximide had no enhancing effect on B cell responses. Furthermore, the enhancing effect of the stimulator B cells was inhibited by the pretreatment of stimulator B cells with anti-class II antibodies. These results suggest that in patients with SLE the abnormality exists in B precursor cells which are easily activated by interacting with other B cells to differentiate into Ig-producing cells and anti-class II antibodies inhibit the B cell activation by interfering with this cellular interaction.


Kidney volume, blood pressure, and albuminuria: findings in an Australian aboriginal community.
Australian Aborigines are experiencing epidemic proportions of renal disease, marked by albuminuria and, variably, hematuria. They also have high rates of low birth weight, which have been associated with lower kidney volumes and higher blood pressures. The authors evaluated relationships between kidney volume, blood pressure, albuminuria, and hematuria in 1 homogeneous group. Forty-three percent (672 of 1,560) of the population in a remote coastal Australian Aboriginal community aged 4.4 to 72.1 years participated in the study. Kidney size correlated closely with body size. Systolic blood pressure (SBP) was correlated inversely with kidney length and kidney volume, after adjusting for age, sex, and body surface area (BSA); a 1-cm increase in mean kidney length was associated with a 2.2-mm Hg decrease in SBP, and a 10-mL increase in mean kidney volume was associated with a 0.6-mm Hg decrease in SBP (P = 0.001). Mean kidney volume explained 10% of the variance in SBP in a multivariate model containing age, sex, and BSA. In addition to higher SBP, adults who had the lowest quartiles of kidney volume also had the highest levels of overt albuminuria (P = 0.044). Smaller kidneys predispose to higher blood pressures and albuminuria in this population. The lower volumes possibly represent kidneys with reduced nephron numbers, which might be related to an adverse intrauterine environment. Susceptibility to renal disease could be a direct consequence of reduced nephron numbers; the higher blood pressures with which they are associated could also contribute to, as well as derive from, this association.


Cyclin-dependent kinase and Cks/Suc1 interact with the proteasome in yeast to control proteolysis of M-phase targets.
Cell cycle-specific proteolysis is critical for proper execution of mitosis in all eukaryotes. Ubiquitination and subsequent proteolysis of the mitotic regulators Clb2 and Pds1 depend on the cyclosome/APC and the 26S proteasome. We report here that components of the cell cycle machinery in yeast, specifically the cell cycle regulatory cyclin-dependent kinase Cdc28 and a conserved associated protein Cks1/Suc1, interact genetically, physically, and functionally with components of the 26S proteasome. A mutation in Cdc28 (cdc28-1N) that interferes with Cks1 binding, or inactivation of Cks1 itself, confers stabilization of Clb2, the principal mitotic B-type cyclin in budding yeast. Surprisingly, Clb2-ubiquitination in vivo and in vitro is not affected by mutations in cks1, indicating that Cks1 is not essential for cyclosome/APC activity. However, mutant Cks1 proteins no longer physically interact with the proteasome, suggesting that Cks1 is required for some aspect of proteasome function during M-phase-specific proteolysis. We further provide evidence that Cks1 function is required for degradation of the anaphase inhibitor Pds1. Stabilization of Pds1 is partially responsible for the metaphase arrest phenotype of cks1 mutants because deletion of PDS1 partially relieves the metaphase block in these mutants.


Isolation and characterization of regulatory mutants from Schizosaccharomyces pombe involved in thiamine-regulated gene expression.
Mutants from Schizosaccharomyces pombe deficient in the regulation of thiamine-repressible acid phosphatase have been isolated. Mutants expressing derepressed levels of the enzyme in the presence and absence of thiamine map in three genes, tnr1, tnr2 and tnr3. mRNA levels of the pho4 gene (coding for thiamine repressible acid phosphatase) and another thiamine-regulatable gene, thi3 (coding for a thiamine biosynthetic enzyme and corresponding to nmt1) are constitutively synthesized in the mutants. The mutants also exhibit constitutive thiamine transport which is thiamine repressible in wild type. The tnr3 mutants reveal a 10-20-fold higher intracellular thiamine level than tnr1 and tnr2 mutants and wild type. Mutants expressing repressed levels of thiamine-repressible acid phosphatase map in gene thi1. No or little amounts of pho4- and nmt1-specific mRNA can be detected. These mutants are impaired in thiamine uptake and are thiamine auxotrophic due to the inability to synthesize the thiazole moiety of the thiamine molecule. All tested tnr and thi1 alleles are recessive, and thi1 mutations are epistatic over tnr mutations. We assume that the thi1 and tnr genes are involved in thiamine-mediated transcription control.


KLF4-dependent epigenetic remodeling modulates podocyte phenotypes and attenuates proteinuria.
The transcription factor Kruppel-like factor 4 (KLF4) has the ability, along with other factors, to reprogram somatic cells into induced pluripotent stem (iPS) cells. Here, we determined that KLF4 is expressed in kidney glomerular podocytes and is decreased in both animal models and humans exhibiting a proteinuric. Transient restoration of KLF4 expression in podocytes of diseased glomeruli in vivo, either by gene transfer or transgenic expression, resulted in a sustained increase in nephrin expression and a decrease in albuminuria. In mice harboring podocyte-specific deletion of Klf4, adriamycin-induced proteinuria was substantially exacerbated, although these animals displayed minimal phenotypical changes prior to adriamycin administration. KLF4 overexpression in cultured human podocytes increased expression of nephrin and other epithelial markers and reduced mesenchymal gene expression. DNA methylation profiling and bisulfite genomic sequencing revealed that KLF4 expression reduced methylation at the nephrin promoter and the promoters of other epithelial markers; however, methylation was increased at the promoters of genes encoding mesenchymal markers, suggesting selective epigenetic regulation of podocyte gene expression. Together, these results suggest that KLF4 epigenetically modulates podocyte phenotype and function and that the podocyte epigenome can be targeted for direct intervention and reduction of proteinuria.


Functional annotation strategy for protein structures.
Whole-genome sequencing projects are a major source of unknown function proteins. However, as predicting protein function from sequence remains a difficult task, research groups recently started to use 3D protein structures and structural models to bypass it. MED-SuMo compares protein surfaces analyzing the composition and spatial distribution of specific chemical groups (hydrogen bond donor, acceptor, positive, negative, aromatic, hydrophobic, guanidinium, hydroxyl, acyl and glycine). It is able to recognize proteins that have similar binding sites and thus, may perform similar functions. We present here a fine example which points out the interest of MED-SuMo approach for functional structural annotation.


Associated anisotropy decays of ethidium bromide interacting with DNA.
Ethidium Bromide (EB) is a commonly used dye in a deoxyribonucleic acid (DNA) study. Upon an intercalation, this dye significantly increases its brightness and fluorescence lifetime. In this report we have studied the time resolved fluorescence properties of EB existing simultaneously in free and DNA-bound forms in the solution. Fluorescence intensity decays were fitted globally to a double exponential model with lifetimes corresponding to free (1.6 ns) and bound (22 ns) forms, and molar fractions were determined for all used solutions. Anisotropy decays displayed characteristic time dependence with an initial rapid decline followed by recovery and slow decay. The short-lived fraction associated with free EB molecules decreases faster than long-lived fraction associated with EB bound to DNA. Consequently, contribution from fast rotation leads to initial rapid decay in anisotropy. On the other hand bound fraction, due to slow rotation helps recover anisotropy in time. This effect of associated anisotropy decays in systems such as EB free/EB-DNA is clearly visible in a wide range of concentrations, and should be taken into account in polarization assays and biomolecule dynamics studies.


Fracture management in HIV positive individuals: a systematic review.
Human immunodeficiency virus (HIV) infection could potentially play an important role in the management of fractures as they have been shown to affect fracture healing and the post-operative risk of implant sepsis. A systematic review of the relevant literature was performed on PubMed and Scopus databases. Twenty-six studies were identified, critiqued and analysed accordingly. No randomised controlled trials were identified. HIV positivity was not shown to influence an individual's risk of early wound infection in operatively managed closed fractures. The rate of pin track infection in open injuries managed with external fixators was low. However, in open injuries managed with internal fixation, early wound infection rates were increased in the HIV-positive population compared to HIV-negative individuals. Regarding late implant infection, in closed fractures there appeared to be no increased risk of infection but there is limited evidence for open injuries. Additionally, further evidence is needed to establish if the rate of union in both open and closed fractures are influenced by HIV status. Overall, no evidence was found to suggest that surgical management of fractures in the HIV population should be avoided, and fixation of closed fractures in the HIV population appeared to be safe. The effect of anti-retroviral therapy is unclear and this should be further researched. However, based on the limited evidence, caution should be taken in the management of open fractures due to the potentially increased infection risk. The impact of anti-retroviral therapy on the outcomes of surgery needs further evaluation.


Acute leukaemia and myelodysplastic syndromes with chromosomal rearrangement involving 11q23 locus, but not MLL gene.
Chromosome 11q23 translocations, resulting in MLL (KMT2A) rearrangement, have been well characterised in acute myeloid leukaemia (AML) and acute lymphoblastic leukaemia (ALL). However, little is known of haematopoietic neoplasms associated with 11q23 translocation but without MLL rearrangement (11q23+/MLL-). The aim of this study is to characterise such cases with 11q23+/MLL-. We retrospectively searched our database for cases with haematopoietic malignancies with 11q23+/MLL-. We identified nine patients, two with AML, two with B-lymphoblastic leukaemia (B-ALL); two with T-lymphoblastic leukaemia (T-ALL), two with myelodysplastic syndrome (MDS) and one with chronic myelomonocytic leukaemia (CMML). The translocations included t(X;11)(p11.2;q23), t(2;11)(p21;q23), t(6;11)(q27;q23), t(8;9;11)(q13;q13;q23), t(11;11)(p15;q23), t(11;14)(q23;q24) and t(11;15)(q23;q14). Five of six patients with acute leukaemia had received chemotherapy and detection of 11q23 translocation occurred at time of disease relapse. Both patients with MDS and the patient with CMML had 11q23 translocation detected at time of initial diagnosis, all three patients progressed to AML after >1 year on hypomethylating agent therapy. All patients received risk-adapted therapies, including stem cell transplant in five patients. At the last follow-up, eight patients died with a median overall survival of 14 months. 11q23+/MLL- occurs rarely, involving different partner chromosomes and showing clinical and pathological features and disease subtypes different from those cases with MLL rearrangement. 11q23+/MLL- appears to be associated with clonal evolution/disease progression in acute leukaemia, a high risk for AML progression in MDS/CMML and a high incidence of disease relapse.


Signaling by insulin-like growth factors in paralyzed skeletal muscle: rapid induction of IGF1 expression in muscle fibers and prevention of interstitial cell proliferation by IGF-BP5 and IGF-BP4.
In the absence of muscle activity, muscle fibers, muscle interstitial cells, and intramuscular nerves display characteristic reactions presumably aimed at restoring a functioning neuromuscular system and avoiding degenerative events. In partially denervated muscle these include proliferation of interstitial cells, followed by nerve sprouting. The same reactions can be induced in intact muscle by elevation of intramuscular insulin-like growth factor (IGF) levels. To determine whether IGFs may participate in the initiation of restorative reactions in inactivated muscle we analyzed the expression of IGF1 and IGF2 mRNA in botulinum toxin-paralyzed and in denervated rat skeletal muscle, and interfered with the activity of IGFs by locally applying the IGF-binding proteins IGF-BP5 and IGF-BP4. To obtain a resolution of the in situ hybridization signals at the cellular level, a nonradioactive method based on digoxigenin-labeled probes was applied. We found that muscle fiber IGF1 mRNA increased rapidly and transiently in inactivated muscle. The time course of this response was similar to that reported for ACh receptor subunits and myogenins. A similar behavior was observed for the corresponding IGF1 protein. The IGF1 response coincided with the transient muscle interstitial cell proliferation reaction. Local application of recombinant IGF-BP5 or IGF-BP4, which are endogenous and specific extracellular ligands of IGFs, prevented the stimulation of interstitial cell proliferation in paralyzed muscle. Our data demonstrate the elevated production of the growth factor IGF1 among early reactions in electrically inactive skeletal muscle fibers and indicate that muscle IGFs are required for the induction of intramuscular interstitial cell proliferation. These findings support the hypothesis that in the neuromuscular system IGF1 plays an important role in initiating cellular reactions in the vicinity of inactive muscle fibers.


An outcome study comparing intravenous sedation with midazolam/fentanyl (conscious sedation) versus propofol infusion (deep sedation) for aesthetic surgery.
The purpose of this study was to determine the differences in measurable outcomes following aesthetic procedures performed under intravenous sedation with incremental doses of midazolam and fentanyl and those performed under propofol infusion. The authors' hypothesis was that the differences in these outcome parameters are not significant between these intravenous sedation protocols. All intraoperative and perioperative records of 84 consecutive patients having aesthetic surgery under a conscious sedation protocol using incremental doses of intravenous midazolam and fentanyl were retrospectively reviewed and compared with the records of a second group of 85 patients having aesthetic surgery under a deep sedation regimen based primarily on propofol infusion. All procedures were hospital based and performed by two surgeons. Twenty-eight different parameters were examined by chart review. In addition, a patient questionnaire was used to assess patient satisfaction and patient recall of operative and perioperative pain, anxiety, nausea, and vomiting. Multivariate statistical analysis was conducted. The two sedation groups were similar with regard to aesthetic procedures performed and patient demographics. The mean duration of operative time was statistically equivalent (152 minutes and 153 minutes). In both groups, there were minor adverse intraoperative events reported but no significant complications. Transient hypotension was more common in the propofol infusion group (12.9 percent versus 2.4 percent, p = 0.018), but no patient required intervention beyond reducing the sedative agent or increasing intravenous fluids. The amount of supplemental fentanyl given intraoperatively was significantly higher in the group whose primary agent for sedation was propofol infusion than the group who received midazolam/fentanyl (209 mug and 143 mug, respectively). The overall questionnaire response rate was 80 percent for both groups. The midazolam/fentanyl sedation group had more recall of "unpleasant intraoperative events" (17 percent versus 3 percent, p = 0.007). However, both groups had low recall of intraoperative pain, anxiety, and nausea. The propofol infusion group experienced significantly more nausea in the recovery room (p = 0.002), nausea at the time of discharge (p = 0.009), and nausea the evening after the operation (p = 0.013). Greater than 90 percent of the patients in both groups would have the same anesthetic in the future rather than undergo general anesthesia. Patient safety, outcomes, and satisfaction are similar in plastic surgery procedures performed under sedation protocols using either incremental doses of midazolam and fentanyl or propofol infusion. All operative and postoperative outcomes for pain, anxiety, and vomiting were similar in the two groups except for immediate postoperative nausea, which was higher in the propofol infusion group. The overall satisfaction of patients undergoing plastic surgery procedures under these intravenous sedation protocols appears very high.


Understanding fundamental principles of enhancer biology at a model locus: Analysing the structure and function of an enhancer cluster at the α-globin locus.
Despite ever-increasing accumulation of genomic data, the fundamental question of how individual genes are switched on during development, lineage-specification and differentiation is not fully answered. It is widely accepted that this involves the interaction between at least three fundamental regulatory elements: enhancers, promoters and insulators. Enhancers contain transcription factor binding sites which are bound by transcription factors (TFs) and co-factors expressed during cell fate decisions and maintain imposed patterns of activation, at least in part, via their epigenetic modification. This information is transferred from enhancers to their cognate promoters often by coming into close physical proximity to form a 'transcriptional hub' containing a high concentration of TFs and co-factors. The mechanisms underlying these stages of transcriptional activation are not fully explained. This review focuses on how enhancers and promoters are activated during differentiation and how multiple enhancers work together to regulate gene expression. We illustrate the currently understood principles of how mammalian enhancers work and how they may be perturbed in enhanceropathies using expression of the α-globin gene cluster during erythropoiesis, as a model.


Establishing a macrophage model with relevance for oral methacrylate monomer exposures: Attenuated Staphylococcus aureus-induced cytokine release from human macrophages.
Leakage of unpolymerized methacrylate monomers after placement of methacrylate-containing polymeric dental materials leads to human exposure. Based on studies using murine macrophages and LPS from Escherichia coli (E. coli), dental monomers like 2-hydroxyethyl methacrylate (HEMA) are known to inhibit lipopolysaccharide (LPS) induced cytokine release. The aim of this study was to establish a model system with relevance for human oral monomer exposure using exposure to live gram-positive bacteria, and to confirm the HEMA-induced effects on cytokine release in this model. The human THP-1 monocyte cell line was differentiated to macrophages using phorbol 12-myristate 13-acetate (PMA), before exposure to 0.5-2mM HEMA and live Staphylococcus aureus (S. aureus) in various multiplicity of infections (MOI). Cytokine release and cytotoxicity were determined after (i) 2-24h pre-exposure to HEMA followed by 2-4h S. aureus exposure and (ii) 2-4h simultaneous exposure. The 24h pre-exposure regime was also tested in primary human airway macrophages and for phagocytosis of S. aureus in THP-1 macrophages. HEMA attenuated the cytokine release more strongly in the pre-exposure than combined exposure regime, with a maximal reduction of 95% in the S. aureus-induced cytokine release. A MOI of 0.1 (corresponding to a bacteria-macrophage ratio of 1:10) was determined to be optimal in the THP-1 macrophages as it induced sufficient cytokine release and negligible cytotoxicity. Attenuated release of S. aureus-induced interleukin (IL)-1β after HEMA exposure was confirmed in primary airway macrophages, while HEMA increased the phagocytosis of S. aureus in THP-1 cells. The model was successfully established and attenuated bacteria-induced cytokine release after HEMA exposure confirmed.


The cdx-hox pathway in hematopoietic stem cell formation from embryonic stem cells.
Embryonic stem cells (ESCs) differentiated in vitro will yield a multitude of hematopoietic derivatives, yet progenitors displaying true stem cell activity remain difficult to obtain. Possible causes are a biased differentiation to primitive yolk sac-type hematopoiesis, and a variety of developmental or functional deficiencies. Recent studies in the zebrafish have identified the caudal homeobox transcription factors (cdx1/4) and posterior hox genes (hoxa9a, hoxb7a) as key regulators for blood formation during embryonic development. Activation of Cdx and Hox genes during the in vitro differentiation of mouse ESCs followed by co-culture on supportive stromal cells generates ESC-derived hematopoietic stem cells (HSCs) capable of multilineage repopulation of lethally irradiated adult mice. We show here that brief pulses of ectopic Cdx4 or HoxB4 expression are sufficient to enhance hematopoiesis during ESC differentiation, presumably by acting as developmental switches to activate posterior Hox genes. Insights into the role of the Cdx-Hox gene pathway during embryonic hematopoietic development in the zebrafish have allowed us to improve the derivation of repopulating HSCs from murine ESCs.


Reactivity of U-associated osteopontin with lactoferrin: a one-to-many complex.
Uranium is the heaviest natural element, mainly found in aqueous medium as the hexavalent uranyl ion (UO22+). Bones are the main organs in which uranium accumulates, depending on as yet unknown molecular and cellular mechanisms. Recently, it has been revealed that osteopontin (OPN), a protein involved in bio-mineralization processes, and its main naturally occurring cleaved form (fOPN), have nanomolar affinities for UO22+. The binding of UO22+ is due to both the phosphorylation sites and acidic residues of these proteins and is accompanied by a slight gain in secondary structure. OPN is an Intrinsically Disordered Protein (IDP), a family of proteins which play a crucial role in several interaction networks, where phosphorylations are thought to be key elements. OPN has been shown to bind lactoferrin (LF) and the two proteins have antagonist functions in the modulation of the bio-mineralization process. However, to date, there has been no evidence that UO22+ and LF compete in their binding to OPN or not. Based on a series of convergent experimental data, this study first addressed in detail the LF/fOPN interaction and proposed a LF:fOPN 4/1 maximal stoichiometry. Moreover the phosphorylations were demonstrated to be necessary for the stability of such complexes. The interaction of preformed UO22+/fOPN complexes with LF was also investigated and the occurrence of several entities involving the three partners was demonstrated. These complexes did not reveal any significant conformational changes compared to those obtained in the absence of UO22+. The results have shown not only that LF and UO22+ do not compete, but also that these complexes are likely to be more stable than LF/fOPN complexes, as indicated by their melting temperature (Tm) values. The potential impact of those uranyl-stabilized ternary complexes on some biological pathways now remains to be assessed. Nonetheless, this work has contributed to shedding light on the formation of stable ternary complexes involving a large structured protein, an IDP and an exogenous metal.


Modulation of synaptic transfer between retinal cones and horizontal cells by spatial contrast.
We studied the influence of steady annular light on the kinetics and sensitivity of horizontal cell (HC) responses to modulation of the intensity of small concentric spots in the turtle retina. As shown by previous investigators, when the intensity of the annulus was equal to the mean spot intensity, spot response kinetics were the same as those for the modulation of spatially uniform light. Turning off the annulus attenuated dramatically high-frequency flicker sensitivity and enhanced somewhat low-frequency sensitivity. This phenomenon reflects a modulation of synaptic transfer between cones and second-order neurons that is mediated by cones, and it will be referred to as cone-mediated surround enhancement (CMSE). Our main results are as follows: (a) The change in test-spot response sensitivity and kinetics upon dimming a steady surrounding annulus is a consequence of the change in spatial contrast rather than change in overall light level. (b) Introduction of moderate contrast between the mean spot intensity and steady surrounding light intensity causes a marked change in spot response kinetics. (c) The dependence of spot response kinetics on surrounding light can be described by a phenomenological model in which the steady state gain and the time constant of one or two single-stage, low-pass filters increase with decreasing annular light intensity (d) The effect of surrounding light on spot responses of a given HC is not determined by change in the steady component of the membrane potential of that cell. (e) Light outside the receptive field of an HC can affect that cell's spot response kinetics. (f) In an expanding annulus experiment, the distance over which steady annular light affects spot response kinetics varies among HCs and can be quite different even between two cells with closely matched receptive field sizes. (g) The degree of CMSE is correlated with HC receptive field size. This correlation suggests that part of the enhancement mechanism is located in the HC. Taken together, our results suggest the involvement of the inner retina in CMSE.


Epidemiology of primary health problems in Beirut.
As a result of 12 years of civil war in Lebanon, it has been impossible to collect regular morbidity information at the primary level. This report is based on a case-control analysis of various health problems as identified from a population based health survey in Beirut in 1983-1984. Cases of headache, backpain and peptic ulcer, as identified from this survey of 2752 households, were matched for age, sex, and neighbourhood with controls from the same sample. Cases and controls were compared for the presence of various characteristics as collected in the household interview. Headaches were more prevalent in females and in the higher educational categories, and the odds ratio was 1.3 (95 per cent confidence interval 1.01-1.68) for the married compared to the non-married. In comparisons of backpain, the odds ratio for alcohol consumption was 2.40 (1.14-5.08), and for belonging to skilled and unskilled labour categories of occupation it was 2.33 (1.05-5.15) when the analysis was limited to the employed group only. Although the peptic ulcer cases were of lower educational background compared to their controls, no other findings were identified in this third case-control comparison. The methodological shortcomings of such studies and the various interpretations of the findings are presented in the discussion.


Long-term complete response of advanced hepatocellular carcinoma treated with multidisciplinary therapy including reduced dose of sorafenib: case report and review of the literature.
An 83-year-old man underwent computed tomography during a routine check-up due to a history of surgical treatment for pancreatic cancer. Two tumors were detected in the anterior segment of the liver. A needle biopsy of the larger tumor was performed, and pathological examination showed that the tumor was a poorly differentiated hepatocellular carcinoma. Resection was not performed considering the patient's poor physical condition. Thus, transcatheter arterial chemoembolization and radiofrequency ablation of the tumors were performed. Three months later, residual tumor of the larger lesion and multiple pulmonary metastases were detected. This time, continuous hepatic arterial infusion chemotherapy was performed. Although the pulmonary metastases markedly reduced, tumor thrombi appeared in the right portal vein on computed tomography. Finally, sorafenib was administered, which led to disappearance of the tumor thrombi and no other signs of recurrence 8 months after initiation of sorafenib on computed tomography. Although sorafenib administration has continued at reduced doses of 200 mg per day or less due to hypertension, complete response has persisted for the past 34 months. It is noteworthy that sorafenib has been given at reduced doses, but a long-term complete response is maintained in a patient who had portal tumor thrombi and distant metastasis. Herein, we present this rare case of advanced hepatocellular carcinoma controlled with reduced doses of sorafenib following multidisciplinary therapy, describe our single center experience with sorafenib use in patients with hepatocellular carcinoma, and review previous reports that focused on dose reduction of sorafenib.


Significance of settling model structures and parameter subsets in modelling WWTPs under wet-weather flow and filamentous bulking conditions.
Current research focuses on predicting and mitigating the impacts of high hydraulic loadings on centralized wastewater treatment plants (WWTPs) under wet-weather conditions. The maximum permissible inflow to WWTPs depends not only on the settleability of activated sludge in secondary settling tanks (SSTs) but also on the hydraulic behaviour of SSTs. The present study investigates the impacts of ideal and non-ideal flow (dry and wet weather) and settling (good settling and bulking) boundary conditions on the sensitivity of WWTP model outputs to uncertainties intrinsic to the one-dimensional (1-D) SST model structures and parameters. We identify the critical sources of uncertainty in WWTP models through global sensitivity analysis (GSA) using the Benchmark simulation model No. 1 in combination with first- and second-order 1-D SST models. The results obtained illustrate that the contribution of settling parameters to the total variance of the key WWTP process outputs significantly depends on the influent flow and settling conditions. The magnitude of the impact is found to vary, depending on which type of 1-D SST model is used. Therefore, we identify and recommend potential parameter subsets for WWTP model calibration, and propose optimal choice of 1-D SST models under different flow and settling boundary conditions. Additionally, the hydraulic parameters in the second-order SST model are found significant under dynamic wet-weather flow conditions. These results highlight the importance of developing a more mechanistic based flow-dependent hydraulic sub-model in second-order 1-D SST models in the future.


The Research Diagnostic Criteria for Temporomandibular Disorders. IV: evaluation of psychometric properties of the Axis II measures.
To evaluate the psychometric properties of the Research Diagnostic Criteria for Temporomandibular Disorders (RDC/TMD) biobehavioral (Axis II) screening instruments. Participants with Axis I TMD diagnoses (n = 626) completed the Axis II instruments (Depression, Nonspecific Physical Symptoms, Graded Chronic Pain) and other instruments assessing psychological distress, pain, and disability at three study sites. Internal consistency, temporal stability, and convergent/discriminant validity of the Axis II measures were assessed. To assess criterion validity of Nonspecific Physical Symptoms and Depression instruments as screeners, 170 participants completed a structured psychiatric diagnostic interview. The Axis II instruments showed very good to excellent internal consistency (Cronbach's alpha coefficients = 0.80 to 0.95). Their convergent (correlation range 0.3 to 0.9) and discriminant (range 0.0 to 0.6) validity were generally supported, although Nonspecific Physical Symptoms was more strongly associated with depressive than with somatic symptoms. Temporal stability was high for characteristic pain intensity (Lin's correlation concordance coefficient [CCC] = 0.91), interference (CCC = 0.89), and chronic pain grade (weighted kappa = 0.87), and fair to good for Depression and Nonspecific Physical Symptoms (CCC = 0.63 to 0.78). The Depression instrument normal versus moderate to severe cutoff point was good at identifying current-year depression and dysthymia diagnoses (sensitivity 87%, specificity 53%). Nonspecific Physical Symptoms did not have high utility for detecting psychiatric disorders (sensitivity 86%, specificity 31%). The Axis-II Depression and Graded Chronic Pain instruments have clinically relevant and acceptable psychometric properties for reliability and validity and utility as instruments for identifying TMD patients with high levels of distress, pain, and disability that can interfere with treatment response and course of Axis I disorders.


The constitutive, glucose-repression-insensitive mutation of the yeast MAL4 locus is an alteration of the MAL43 gene.
Mutations resulting in constitutive production of maltase have been identified at each of the five MAL loci of Saccharomyces yeasts. Here we examine a dominant constitutive, glucose-repression-insensitive allele of the MAL4 locus (MAL4-C). Our results demonstrate that MAL4-C is an alteration in the MAL43 gene, which encodes the positive regulator of the MAL structural genes, and that its product is trans-acting. The MAL43 gene from the MAL4-C strain was cloned and integrated into a series of nonfermenting strains lacking a functional regulatory gene but carrying copies of the maltose permease and maltase structural genes. Expression of the maltase structural gene was both constitutive and insensitive to glucose repression in these transformants. The MAL4-C allele also results in constitutive expression of the unlinked MAL12 gene (encoding maltase) in this strain. In addition, the cloned MAL43 gene was shown to be dominant to the wild-type MAL63 gene. We also show that most of the glucose repression insensitivity of strains carrying the MAL4-C allele results from alteration of MAL43.


Rayleigh scattering of surface plasmons by sub-wavelength holes.
We study the scattering of surface plasmons from sub-wavelength holes and find that it exhibits a stronger wavelength dependence than the traditional λ(-4) scaling found for Rayleigh scattering of light from small particles. This experimental observation is consistent with recent theoretical work and linked to the two-dimensional nature of the surface plasmon and the wavelength dependence of its spatial extent in the third dimension. The scattering cross sections are obtained with a frequency-correlation technique, which compares intensity speckle patterns observed behind various random structures of holes and recorded at different wavelengths. This powerful technique even allows us to distinguish between scattering of surface plasmons into photons and scattering into other surface plasmons.


Nanoscopic imaging of thick heterogeneous soft-matter structures in aqueous solution.
Precise nanometre-scale imaging of soft structures at room temperature poses a major challenge to any type of microscopy because fast thermal fluctuations lead to significant motion blur if the position of the structure is measured with insufficient bandwidth. Moreover, precise localization is also affected by optical heterogeneities, which lead to deformations in the imaged local geometry, the severity depending on the sample and its thickness. Here we introduce quantitative thermal noise imaging, a three-dimensional scanning probe technique, as a method for imaging soft, optically heterogeneous and porous matter with submicroscopic spatial resolution in aqueous solution. By imaging both individual microtubules and collagen fibrils in a network, we demonstrate that structures can be localized with a precision of ∼10 nm and that their local dynamics can be quantified with 50 kHz bandwidth and subnanometre amplitudes. Furthermore, we show how image distortions caused by optically dense structures can be corrected for.


Thiolated human serum albumin cross-linked dextran hydrogels as a macroscale delivery system.
Hydrogels play an important role in macroscale delivery systems by enabling the transport of cells and molecules. Here we present a facile and benign method to prepare a dextran-based hydrogel (Dex-sHSA) using human serum albumin (HSA) as a simultaneous drug carrier and covalent cross-linker. Drug binding affinity of the albumin protein was conserved in the thiolation step using 2-iminothiolane and subsequently, in the in situ gelation step. Oscillation rheometry studies confirmed the formation of a three-dimensional viscoelastic network upon reaction of dextran and the HSA protein. The mechanical properties of Dex-sHSA hydrogel can be tuned by the protein concentration, and the degree of thiolation of sHSA. Sustained release of hydrophobic drugs, such as ibuprofen, paclitaxel and dexamethasone, from the Dex-sHSA network was shown over one week. Hence, this albumin-based dextran hydrogel system demonstrates its potential as a macroscale delivery system of hydrophobic therapeutics for a wide range of biomedical applications.


Inherited neuronal ion channelopathies: new windows on complex neurological diseases.
Studies of genetic forms of epilepsy, chronic pain, and migraine caused by mutations in ion channels have given crucial insights into molecular mechanisms, pathogenesis, and therapeutic approaches to complex neurological disorders. Gain-of-function missense mutations in the brain type-I sodium channel Na(V)1.1 are a primary cause of generalized epilepsy with febrile seizures plus. Loss-of-function mutations in Na(V)1.1 channels cause severe myoclonic epilepsy of infancy, an intractable childhood epilepsy. Studies of a mouse model show that this disease is caused by selective loss of sodium current and excitability of GABAergic inhibitory interneurons, which leads to hyperexcitability, epilepsy, and ataxia. Mutations in the peripheral sodium channel Na(V)1.7 cause familial pain syndromes. Gain-of-function mutations cause erythromelalgia and paroxysmal extreme pain disorder as a result of hyperexcitability of sensory neurons, whereas loss-of-function mutations cause congenital indifference to pain because of attenuation of action potential firing. These experiments have defined correlations between genotype and phenotype in chronic pain diseases and focused attention on Na(V)1.7 as a therapeutic target. Familial hemiplegic migraine is caused by mutations in the calcium channel, Ca(V)2.1, which conducts P/Q-type calcium currents that initiate neurotransmitter release. These mutations increase activation at negative membrane potentials and increase evoked neurotransmitter release at cortical glutamatergic synapses. Studies of a mouse genetic model show that these gain-of-function effects lead to cortical spreading depression, aura, and potentially migraine. Overall, these experiments indicate that imbalance in the activity of excitatory and inhibitory neurons is an important underlying cause of these diseases.


Identification of Stably Expressed lncRNAs as Valid Endogenous Controls for Profiling of Human Glioma.
Recent research indicates that long non-coding RNAs (lncRNA) represent a new family of RNAs that is of fundamental importance for controlling transcription and translation. Thereby, there is increasing evidence that lncRNAs are also important in tumourigenesis. Thereby valid expression profiling using quantitative PCR requires suitable, stably expressed normalisers to achieve reliable and reproducible data. However, no systematic analysis of suitable references in lncRNA studies in human glioma has been performed yet. In this study, we investigated 90 lncRNAs in 30 tissue specimen for the expression stability in human diffuse astrocytoma (WHO-Grade II), anaplastic astrocytoma (WHO-Grade III) and glioblastoma (WHO-Grade IV) both alone as well as in comparison with normal white matter. Our identification procedure included a rigorous bioinformatical selection process that resulted in the inclusion of only highly abundant, equally expressed lncRNAs for further analysis. Additionally, lncRNAs were classified according to their stability value using the NormFinder algorithm. We identified 24 appropriate normalisers suitable for studies in diffuse astrocytoma, 22 for studies in anaplastic astrocytoma and 12 for studies in glioblastoma. Comparing all three glioma entities 7 lncRNAs showed stable expression levels. Addition of normal brain tissue resulted in only 4 suitable lncRNAs. Our findings indicate that 4 lncRNAs (HOXA6as, H19 upstream conserved 1 and 2, Zfhx2as and BC200) are suitable as normalisers in glioma and normal brain. These lncRNAs may thus be regarded as universal references being applicable for the accurate normalisation of lncRNA expression profiling in various glioma (WHO-Grades II-IV) alone and in combination with brain tissue. This enables to perform valid longitudinal studies, e.g. of glioma before and after malignisation to identify changes of lncRNA expressions probably driving malignant transformation.


Thymoquinone, an active constituent of Nigella sativa seeds, binds with bilirubin and protects mice from hyperbilirubinemia and cyclophosphamide-induced hepatotoxicity.
Some reports indicate that thymoquinone (TQ), the main constituent of Nigella sativa seeds, is hepatoprotective. The aim of this study was to determine whether TQ is able to bind directly to bilirubin, and whether TQ or liposomal formulation of TQ (Lip-TQ) can reduce cyclophosphamide (CYP)-induced liver toxicity, serum bilirubin level in mice. The binding of TQ with bilirubin was studied by UV-VIS, fluorescence and Near-UV CD spectroscopy. Inhibition of binding of bilirubin to erythrocytes by TQ was also examined. To increase the in vivo efficacy, Lip-TQ was prepared and used against CYP-induced toxicity. The protective role of TQ or Lip-TQ against CYP-induced toxicity was assessed by determining the liver function parameters, the levels of superoxide dismutase (SOD) and catalase (CAT), and histological studies. It was found that TQ binds to bilirubin and significantly inhibits the binding of bilirubin to erythrocytes. Lip-TQ (10 mg/kg) significantly reduced the levels of aspartate transaminase (AST) from 254 ± 48 to 66 ± 18 IU/L (P < 0.001), alanine transaminase (ALT) from 142 ± 28 to 47.8 ± 16 IU/L (P < 0.05) and serum bilirubin from 2.8 ± 0.50 to 1.24 ± 0.30 mg/dl (P < 0.05). Treatment with Lip-TQ reduced the CYP-induced inflammation and hemorrhage in liver tissues. Moreover, treatment with free or Lip-TQ protected the activity of SOD and CAT in CYP-injected mice. Therefore, TQ can reduce the level of bilirubin in systemic circulation in disease conditions that lead to hyperbilirubinemia and liver toxicity and hence may be used as a supplement in the treatment of liver ailments.


Differential predictive value of depressive versus anxiety symptoms in the prediction of 8-year mortality after acute coronary syndrome.
Both depression and anxiety have been associated with poor prognosis in patients with acute coronary syndrome (ACS). However, certain symptoms and how they are measured may be more important than others. We investigated three different scales to determine their predictive validity. Patients with ACS (N = 598) completed either the Hospital Anxiety and Depression Scales (HADS-A, HADS-D; n = 316) or the Beck Depression Inventory-Fast Screen (n = 282). Their all-cause mortality status was assessed at 8 years. During follow-up, 20% (121/598) of participants died. Cox proportional hazards modeling showed that the HADS-D was predictive of mortality (hazard ratio [HR] = 1.11, 95% confidence interval [CI] = 1.04-1.19), and this association remained significant after adjustment for major clinical/demographic factors, whereas the HADS-A (HR = 0.96, 95% CI = 0.85-1.09) and the Beck Depression Inventory-Fast Screen (HR = 0.99, 95% CI = 0.91-1.08) were not. The following depression items from the HADS-D predicted mortality: "I still enjoy the things I used to enjoy" (HR = 1.38, 95% CI = 1.05-1.82), "I can laugh and see the funny side of things" (HR = 1.48, 95% CI = 1.11-1.96), "I feel as if I am slowed down" (HR = 1.66, 95% CI = 1.24-2.22), and "I look forward with enjoyment to things" (HR = 1.36, 95% CI = 1.08-1.72). Depressive symptoms related to lack of enjoyment or pleasure and physical or cognitive slowing, as measured by the HADS-D, predicted all-cause mortality at 8 years ACS patients, whereas other depressive and anxiety symptoms did not. Whether symptoms of distress predict prognosis in ACS seems to be dependent on the measures and items used.