RJR-2403: a nicotinic agonist with CNS selectivity II. In vivo characterization.
We have evaluated the physiological and behavioral effects of the CNS-selective nicotinic agonist (E)-N-methyl-4-(3-pyridinyl) -3-butene-1-amine (RJR-2403) using a number of different methods, including 1) reversal of pharmacologically induced amnesia in a step-through passive avoidance paradigm, 2) radial arm maze performance in rats with chemically induced brain lesions, 3) changes in HR and blood pressure in rats and 4) changes in body temperature, Y-maze activity, acoustic startle response and respiration in mice. Our results indicate that RJR-2403 is equal to or better than nicotine on measures of CNS function and cognitive enhancement. Specifically, RJR-2403 significantly improved passive avoidance retention after scopolamine-induced amnesia and enhanced both working and reference memory in rats with ibotenic acid lesions of the forebrain cholinergic projection system in an 8-arm radial maze paradigm. By comparison, RJR-2403 was 15 to 30-fold less potent than nicotine in decreasing body temperature, respiration, Y-maze rears and crosses and acoustic startle response. RJR-2403 also demonstrated greatly reduced cardiovascular effects. RJR-2403 was approximately 10-fold less potent than nicotine in increasing HR and 20-fold less potent in increasing blood pressure. These results are consistent with in vitro data indicating this compound's high selectivity for CNS nicotinic ACh receptor subtypes relative to peripheral ganglionic and muscle-type nicotinic ACh receptors. Therefore, RJR-2403 may be a valuable tool for understanding the central and peripheral pharmacology of nicotinic cholinergic systems as well as a potential lead compound for the development of nicotinic therapeutics to treat neurological diseases where cholinergic neurotransmission has been compromised.


Cell-by-cell scanning of whole mitochondrial genomes in aged human heart reveals a significant fraction of myocytes with clonally expanded deletions.
Quantitative information on the cell-to-cell distribution of all possible mitochondrial DNA (mtDNA) mutations in young and aged tissues is needed to assess the relevance of these mutations to the aging process. In the present study, we used PCR amplification of full-length mitochondrial genomes from single cells to scan human cardiomyocytes for all possible large deletions in mtDNA. Analysis of more than 350 individual cells that were derived from three middle-aged and four centenarian donors demonstrates that while most of the cells contain no deletions, in certain cardiomyocytes a significant portion of the mtDNA molecules carried one particular deletion. Different affected cells contained different deletions. Although similar numbers of cells were screened for each donor, these deletion-rich cells were found only in the hearts of old donors, where they occurred at a frequency of up to one in seven cells. These initial observations demonstrate the efficiency of the method and indicate that mitochondrial mutations have the potential to play an important role in human myocardial aging.


Vitamin D receptor controls expression of the anti-aging klotho gene in mouse and human renal cells.
Isoforms of the mammalian klotho protein serve as membrane co-receptors that regulate renal phosphate and calcium reabsorption. Phosphaturic effects of klotho are mediated in cooperation with fibroblast growth factor receptor-1 and its FGF23 ligand. The vitamin D receptor and its 1,25-dihydroxyvitamin D(3) ligand are also crucial for calcium and phosphate regulation at the kidney and participate in a feedback loop with FGF23 signaling. Herein we characterize vitamin D receptor-mediated regulation of klotho mRNA expression, including the identification of vitamin D responsive elements (VDREs) in the vicinity of both the mouse and human klotho genes. In keeping with other recent studies of vitamin D-regulated genes, multiple VDREs control klotho expression, with the most active elements located at some distance (-31 to -46 kb) from the klotho transcriptional start site. We therefore postulate that the mammalian klotho gene is up-regulated by liganded VDR via multiple remote VDREs. The phosphatemic actions of 1,25-dihydroxyvitamin D(3) are thus opposed via the combined phosphaturic effects of FGF23 and klotho, both of which are upregulated by the liganded vitamin D receptor.


Posttransplant complications in patients with marrow failure syndromes: are we improving long-term outcomes?
Inherited bone marrow failure syndromes (IBMFS) encompass a group of rare genetic disorders characterized by bone marrow failure, non-hematologic multisystemic comorbidities, disease defining congenital anomalies, and a susceptibility to myelodysplastic syndrome, acute myeloid leukemia, and in some instances solid tumors. The most common IBMFS include Fanconi anemia, Shwachman-Diamond syndrome, Diamond-Blackfan anemia, and telomere biology disorders/ dyskeratosis congenita. Allogeneic hematopoietic stem cell transplant (HCT) is a well-established curative treatment to correct the hematological manifestations but does not halt or reverse the nonhematological complications and may hasten them. With advances in HCT and in our ability to care for patients with IBMFS, an increasing number of survivors are making it imperative to not only diagnose but also treat late effects from the pre-, peri-, and post-HCT course and complications relating to the natural history of the syndrome. As the field of HCT evolves to allow for the incorporation of alternate graft sources, for expansion of donor options to include unrelated and mismatched donors, and for use of reduced-intensity conditioning or reduced toxicity myeloablative regimens, we have yet to determine if these advances modify the disease-specific course. While long-term outcomes of these patients are often included under one umbrella, this article seeks to address disease-specific post-HCT outcomes within IBMFS.


Precise identification of a human immunodeficiency virus type 1 antigen processing mutant.
Human immunodeficiency virus type 1 (HIV-1) evokes a strong immune response, but the virus persists. Polymorphisms within known antigenic sites result in loss of immune recognition and can be positively selected. Amino acid variation outside known HLA class I restricted epitopes can also enable immune escape by interfering with the processing of the optimal peptide antigen. However, the lack of precise rules dictating epitope generation and the enormous genetic diversity of HIV make prediction of processing mutants very difficult. Polymorphism E169D in HIV-1 reverse transcriptase (RT) is significantly associated with HLA-B*0702 in HIV-1-infected individuals. This polymorphism does not map within a known HLA-B*0702 epitope; instead, it is located five residues downstream of a HLA-B*0702-restricted epitope SPAIFQSSM (SM9). Here we investigate the association between E169D and HLA-B*0702 for immune escape via the SM9 epitope. We show that this single amino acid variation prevents the immune recognition of the flanked SM9 epitope by cytotoxic T cells through lack of generation of the epitope, which is a result of aberrant proteasomal cleavage. The E169D polymorphism also maps within and abrogates the recognition of an HLA-A*03-restricted RT epitope MR9. This study highlights the potential for using known statistical associations as indicators for viral escape but also the complexity involved in interpreting the immunological consequences of amino acid changes in HIV sequences.


Changes in erythrocyte membrane fatty acids during a clinical trial of eicosapentaenoic acid (EPA) supplementation in schizophrenia.
In a previously reported double-blind, placebo-controlled trial of eicosapentaenoic acid (EPA) as supplemental treatment in 40 patients with schizophrenia, we found significant improvement in symptoms as measured by the Positive and Negative Syndrome Scale (PANSS) compared to placebo (Emsley et al. 2002). Here we report changes in fatty acid composition of erythrocyte membranes in the same sample (n = 16 in each group). After 12 weeks of receiving EPA, levels of several saturated and mono-unsaturated fatty acids decreased significantly while levels of n-3 fatty acids increased significantly compared to the placebo group. Increases of n-3 and n-6 fatty acids in the erythrocyte membranes were greater in subjects who improved more than 20% on overall symptoms. Changes in fatty acids correlated significantly with improvement in PANSS sub-scale scores, more so in females than in males. Docosahexaenoic acid (DHA) (22:6n-3) levels increased less than expected, suggesting a possible defect in synthesis or incorporation of DHA into membranes in schizophrenia. Improvement in dyskinesia correlated significantly with an increase in alpha-linolenic acid (18:3n-3; p = 0.03), and a decrease in 20:1n-9 (p = 0.005).


VIPP1 rods engulf membranes containing phosphatidylinositol phosphates.
In cyanobacteria and plants, VIPP1 plays crucial roles in the biogenesis and repair of thylakoid membrane protein complexes and in coping with chloroplast membrane stress. In chloroplasts, VIPP1 localizes in distinct patterns at or close to envelope and thylakoid membranes. In vitro, VIPP1 forms higher-order oligomers of >1 MDa that organize into rings and rods. However, it remains unknown how VIPP1 oligomerization is related to function. Using time-resolved fluorescence anisotropy and sucrose density gradient centrifugation, we show here that Chlamydomonas reinhardtii VIPP1 binds strongly to liposomal membranes containing phosphatidylinositol-4-phosphate (PI4P). Cryo-electron tomography reveals that VIPP1 oligomerizes into rods that can engulf liposomal membranes containing PI4P. These findings place VIPP1 into a group of membrane-shaping proteins including epsin and BAR domain proteins. Moreover, they point to a potential role of phosphatidylinositols in directing the shaping of chloroplast membranes.


Two-colour chewing gum mixing ability test for evaluating masticatory performance in children with mixed dentition: validity and reliability study.
The unappealing taste of the chewing material and the time-consuming repetitive task in masticatory performance tests using artificial foodstuff may discourage children from performing natural chewing movements. Therefore, the aim was to determine the validity and reliability of a two-colour chewing gum mixing ability test for masticatory performance (MP) assessment in mixed dentition children. Masticatory performance was tested in two groups: systemically healthy fully dentate young adults and children in mixed dentition. Median particle size was assessed using a comminution test, and a two-colour chewing gum mixing ability test was applied for MP analysis. Validity was tested with Pearson correlation, and reliability was tested with intra-class correlation coefficient, Pearson correlation and Bland-Altman plots. Both comminution and two-colour chewing gum mixing ability tests revealed statistically significant MP differences between children (n = 25) and adults (n = 27, both P < 0·01). Pearson correlation between comminution and two-colour chewing gum mixing ability tests was positive and significant (r = 0·418, P = 0·002). Correlations for interobserver reliability and test-retest values were significant (r = 0·990, P = 0·0001 and r = 0·995, P = 0·0001). Although both methods could discriminate MP differences, the comminution test detected these differences generally in a wider range compared to two-colour chewing gum mixing ability test. However, considering the high reliability of the results, the two-colour chewing gum mixing ability test can be used to assess masticatory performance in children, especially at non-clinical settings.


Mitral regurgitation following acute myocardial infarction.
Mitral valve regurgitation (MR) is a frequent Doppler echocardiographic finding in patients after acute myocardial infarction (AMI) and an independent predictor of long-term cardiovascular mortality. Reported risk factors include advanced age, prior myocardial infarction, infarct extension, and recurrent ischemia. During the early phase of AMI, transient ischemic MR is common and rarely causes hemodynamic compromise. However, when several chordae tendineae or a papillary muscle ruptures, acute left atrial and ventricular volume overload ensues, leading to abrupt hemodynamic deterioration with cardiogenic shock. Auscultation may be unrevealing due to decreased turbulence. Hence, the importance of a high index of suspicion for acute MR in any patient with acute pulmonary edema in the setting of AMI, especially if left ventricular systolic function is well preserved. Later, ventricular remodeling may lead to MR through annular dilatation or papillary muscle migration with malcoaptation of the leaflets. The widespread availability, ease of use and non-invasive nature of Doppler echocardiography have made it the standard diagnostic tool for detecting MR. Mechanical reperfusion of the infarct-related artery seems to be superior to fibrinolysis in decreasing its incidence acutely and in the long run. Nevertheless, when acute severe MR occurs, unless rapidly diagnosed and treated, this dreaded complication is associated with high morbidity and mortality. Prompt surgical intervention after hemodynamic stabilization is essential to ensure a good short-term and long-term prognosis. This review discusses the incidence, long-term prognosis, associated risk factors, complex pathophysiology, time of occurrence, clinical manifestations, diagnosis, and management of patients with MR after AMI.


Early warning signs of endocrine disruption in adult fish from the ingestion of polyethylene with and without sorbed chemical pollutants from the marine environment.
Plastic debris is associated with several chemical pollutants known to disrupt the functioning of the endocrine system. To determine if the exposure to plastic debris and associated chemicals promotes endocrine-disrupting effects in fish, we conducted a chronic two-month dietary exposure using Japanese medaka (Oryzias latipes) and environmentally relevant concentrations of microplastic (<1mm) and associated chemicals. We exposed fish to three treatments: a no-plastic (i.e. negative control), virgin-plastic (i.e. virgin polyethylene pre-production pellets) and marine-plastic treatment (i.e. polyethylene pellets deployed in San Diego Bay, CA for 3 months). Altered gene expression was observed in male fish exposed to the marine-plastic treatment, whereas altered gene expression was observed in female fish exposed to both the marine- and virgin-plastic treatment. Significant down-regulation of choriogenin (Chg H) gene expression was observed in males and significant down-regulation of vitellogenin (Vtg I), Chg H and the estrogen receptor (ERα) gene expression was observed in females. In addition, histological observation revealed abnormal proliferation of germ cells in one male fish from the marine-plastic treatment. Overall, our study suggests that the ingestion of plastic debris at environmentally relevant concentrations may alter endocrine system function in adult fish and warrants further research.


Wigner function measurement using a lenslet array.
Geometrical-optical arguments have traditionally been used to explain how a lenslet array measures the distribution of light jointly over space and spatial frequency. Here, we rigorously derive the connection between the intensity measured by a lenslet array and wave-optical representations of such light distributions for partially coherent optical beams by using the Wigner distribution function (WDF). It is shown that the action of the lenslet array is to sample a smoothed version of the beam's WDF (SWDF). We consider the effect of lenslet geometry and coherence properties of the beam on this measurement, and we derive an expression for cross-talk between lenslets that corrupts the measurement. Conditions for a high fidelity measurement of the SWDF and the discrepancies between the measured SWDF and the WDF are investigated for a Schell-model beam.


Rifampin pharmacokinetics in children, with and without human immunodeficiency virus infection, hospitalized for the management of severe forms of tuberculosis.
Rifampin is a key drug in antituberculosis chemotherapy because it rapidly kills the majority of bacilli in tuberculosis lesions, prevents relapse and thus enables 6-month short-course chemotherapy. Little is known about the pharmacokinetics of rifampin in children. The objective of this study was to evaluate the pharmacokinetics of rifampin in children with tuberculosis, both human immunodeficiency virus type-1-infected and human immunodeficiency virus-uninfected. Fifty-four children, 21 human immunodeficiency virus-infected and 33 human immunodeficiency virus-uninfected, mean ages 3.73 and 4.05 years (P = 0.68), respectively, admitted to a tuberculosis hospital in Cape Town, South Africa with severe forms of tuberculosis were studied approximately 1 month and 4 months after commencing antituberculosis treatment. Blood specimens for analysis were drawn in the morning, 45 minutes, 1.5, 3.0, 4.0 and 6.0 hours after dosing. Rifampin concentrations were determined by liquid chromatography tandem mass spectrometry. For two sample comparisons of means, the Welch version of the t-test was used; associations between variables were examined by Pearson correlation and by multiple linear regression. The children received a mean rifampin dosage of 9.61 mg/kg (6.47 to 15.58) body weight at 1 month and 9.63 mg/kg (4.63 to 17.8) at 4 months after commencing treatment administered as part of a fixed-dose formulation designed for paediatric use. The mean rifampin area under the curve 0 to 6 hours after dosing was 14.9 and 18.1 microg/hour/ml (P = 0.25) 1 month after starting treatment in human immunodeficiency virus-infected and human immunodeficiency virus-uninfected children, respectively, and 16.52 and 17.94 microg/hour/ml (P = 0.59) after 4 months of treatment. The mean calculated 2-hour rifampin concentrations in these human immunodeficiency virus-infected and human immunodeficiency virus-uninfected children were 3.9 and 4.8 microg/ml (P = 0.20) at 1 month after the start of treatment and 4.0 and 4.6 microg/ml (P = 0.33) after 4 months of treatment. These values are considerably less than the suggested lower limit for 2-hour rifampin concentrations in adults of 8.0 microg/ml and even 4 microg/ml Both human immunodeficiency virus-infected and human immunodeficiency virus-uninfected children with tuberculosis have very low rifampin serum concentrations after receiving standard rifampin dosages similar to those used in adults. Pharmacokinetic studies of higher dosages of rifampin are urgently needed in children to assist in placing the dosage of rifampin used in childhood on a more scientific foundation.


Mosaic Disease of Calla Lily Caused by a New Potyvirus in Taiwan.
In 1998, a new mosaic disease of calla lily (Zantedeschia spp.) was found in Taichung County, Taiwan. Primary symptoms were mosaic and green islands on leaves and discolored spots on flowers. Symptomatic plants were negative in double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA) for Dasheen mosaic virus (DsMV) polyclonal antibody (Agdia Inc., Elkhart, IN), but were positive in indirect ELISA using an anti-potyvirus group monoclonal antibody. Extracts from these plants were infective by mechanical inoculation to seedlings of calla lily and Philodendron selloum, which expressed veinal chlorosis, mosaic, and green island symptoms. Transmission electron microscopic analysis indicated that the virus particles purified from inoculated P. selloum were 695 to 845 nm long. In addition, potyvirus-specific cytoplasmic inclusions were observed in epidermal cells of infected calla lily. The 3'-terminal region of the virus was amplified by reverse-transcription polymerase chain reaction from total RNA or viral RNA using a potyvirus-specific degenerate primer and an oligo(dT) primer. A 1.6-kb amplified fragment was cloned, and three independent clones were sequenced. The sequences included a portion of NIb gene, the coat protein (CP) gene, the 3' untranslated region (3'UTR) and the poly(A) tail. This nucleotide sequence (GenBank Accession No. AF332872) was checked against the international sequence databases using the BLAST program (provided by National Center for Biotechnology Information online at www.ncbi.nlm.nih.gov/blast/ ). The highest identity of the CP amino acid sequences between this unknown virus and other potyviruses is 68%. Amino acid sequence homologies for the CPs between individual potyviruses are 38 to 71%, while those between strains of a potyvirus are more than 90% (1). Therefore, this Zantedeschia-infecting virus is a previously undescribed potyvirus and is herein designated as Zantedeschia mosaic virus (ZaMV). In further analyses, the amino acid sequence identities of the CP gene between ZaMV and 13 other aphid-transmitted potyviruses were 46 to 61% and 9 to 23% for the nucleotide sequence of the 3'UTR. ZaMV and DsMV showed 46% identity in the CP amino acid sequence and 12% identity in the 3'UTR nucleotide sequence, indicating that they are two distinct members of the genus Potyvirus. To our knowledge, this is the first report of natural infection of Zantedeschia spp. by ZaMV, a new potyvirus identified in Taiwan. Reference: (1) D. D. Shukla and C. W. Ward. Arch. Virol. 106:171, 1989.


A new glucose-6-phosphate dehydrogenase variant, G6PD Orissa (44 Ala-->Gly), is the major polymorphic variant in tribal populations in India.
Deficiency of glucose-6-phosphate dehydrogenase (G6PD) is usually found at high frequencies in areas of the world where malaria has been endemic. The frequency and genetic basis of G6PD deficiency have been studied in Africa, around the Mediterranean, and in the Far East, but little such information is available about the situation in India. To determine the extent of heterogeneity of G6PD, we have studied several different Indian populations by screening for G6PD deficiency, followed by molecular analysis of deficient alleles. The frequency of G6PD deficiency varies between 3% and 15% in different tribal and urban groups. Remarkably, a previously unreported deficient variant, G6PD Orissa (44 Ala-->Gly), is responsible for most of the G6PD deficiency in tribal Indian populations but is not found in urban populations, where most of the G6PD deficiency is due to the G6PD Mediterranean (188 Ser-->Phe) variant. The KmNADP of G6PD Orissa is fivefold higher than that of the normal enzyme. This may be due to the fact that the alanine residue that is replaced by glycine is part of a putative coenzyme-binding site.


High-throughput turbidimetric screening for heparin-neutralizing agents and low-molecular-weight heparin mimetics.
Safer heparin-neutralizing agents are currently required to replace protamine, the use of which causes adverse effects such as anaphylaxia. Low-molecular-weight (LMW) heparin mimetics that potentiate antithrombin III (AT) action are also valuable as anti-thrombotics. This paper describes a high-throughput assay for both heparin-neutralizing agents and LMW heparin mimetics without the use of blood preparations. The assay is based on turbidimetric measurement of a solution of collagen, heparin, and a test compound. Native collagen molecules spontaneously form insoluble fibrils when transferred to a physiological buffer, and this process is inhibited by heparin. In the presence of a heparin-neutralizing agent or an LMW heparin mimetic, the inhibitory effect of heparin is canceled and turbidity increase is retrieved. We demonstrated that this assay is effective in detecting potential agents with high reliability (Z' factor=0.9). The screening of a chemical library (34400 compounds) was further performed in a 384-well format, and led to the identification of a novel heparin-neutralizing agent. Since this assay protocol is feasible for an automated high-throughput screening (HTS) system, it could enhance the lead seeking process for drugs related to heparin/heparan sulfate (HS) functions.


Sucrose pretreatment for enucleation: an efficient and non-damage method for removing the spindle of the mouse MII oocyte.
Oocytes enucleated at metaphase II stage can support reprogramming of transferred nucleus and further developing to term. However, the first polar body in mice sometimes migrates away from the original place of expulsion, so the chromosomes of the oocyte will displace from the first polar body. Thus, it is not always possible to successfully enucleate according to the position of the first polar body. Here we use sucrose treatment to visualize metaphase spindle fibers and chromosomes with standard light microscopy. In the manipulation medium containing 3% sucrose, oocytes of poor quality become shrunken, deformed or fragmented, while oocytes of good quality in the same medium would show a swelling around the metaphase chromosomes and a transparent spindle area, shaped like "infinity" and "0". So it is easy to remove the well-distinguished spindle and chromosomes in oocytes of good quality. Re-examined by Hoechst 33342 stain under the UV light, the enucleation rate was 100%. There was no significant difference in IVF and cleavage rates between the sucrose treatment and the control group. In conclusion, this study demonstrated that 3% sucrose pretreatment can give a method for evaluating embryo quality and more importantly, it can, under a common microscope, allow the visualization of the spindle and chromosomes in oocytes of good quality and hence efficiently improve enucleation rate without any harm.


Bangladeshi backyard poultry raisers' perceptions and practices related to zoonotic transmission of avian influenza.
Highly pathogenic avian influenza (H5N1) virus (known as "bird flu") is an important public health concern due to its potential to infect humans and cause a human pandemic. Bangladesh is a high-risk country for an influenza pandemic because of its dense human population, widespread backyard poultry raising, and endemic H5N1 infection in poultry. Understanding poultry raisers' perceived risks and identifying their risk exposures can help to develop interventions to reduce the risk of avian influenza transmission. This paper explores the perception of Bangladeshi backyard poultry raisers regarding poultry sickness and zoonotic disease transmission and relevant practices. We conducted a qualitative study using social mapping (n=2), in-depth interviews (n=40), household mapping (n=40) and observation (n=16), in two backyard poultry-raising communities. The poultry raisers recognized various signs of poultry illness but they did not distinguish among diseases using biomedical classifications. They perceived disease transmission from poultry to poultry, but not from poultry to humans. They usually kept sick poultry under the bed. If the poultry did not recover, they were slaughtered and consumed or sold. The poultry raisers had close contact with sick birds while handling and slaughtering poultry. The poultry raisers are unlikely to follow instructions from health authorities to prevent "bird flu" transmission because many of the instructions ask low-income producers to change their existing practices and require time, money, and financial loss. Villagers are more likely to comply with interventions that help to protect their flocks and address their financial interest.


Inter-disciplinary European guidelines on surgery of severe obesity.
In 2005, for the first time in European history, an extraordinary Expert panel named 'The BSCG' (Bariatric Scientific Collaborative Group), was appointed through joint effort of the major European Scientific Societies which are active in the field of obesity management. Societies that constituted this panel were: IFSO - International Federation for the Surgery of Obesity, IFSO-EC - International Federation for the Surgery of Obesity - European Chapter, EASO - European Association for Study of Obesity, ECOG - European Childhood Obesity Group, together with the IOTF (International Obesity Task Force) which was represented during the completion process by its representative. The BSCG was composed not only of the top officers representing the respective Scientific Societies (four acting presidents, two past presidents, one honorary president, two executive directors), but was balanced with the presence of many other key opinion leaders in the field of obesity. The BSCG composition allowed the coverage of key disciplines in comprehensive obesity management, as well as reflecting European geographical and ethnic diversity. This joint BSCG expert panel convened several meetings which were entirely focused on guidelines creation, during the past two years. There was a specific effort to develop clinical guidelines, which will reflect current knowledge, expertise and evidence based data on morbid obesity treatment.


A comprehensive analysis of the androgen receptor gene and risk of breast cancer: results from the National Cancer Institute Breast and Prostate Cancer Cohort Consortium (BPC3).
Androgens have been hypothesised to influence risk of breast cancer through several possible mechanisms, including their conversion to estradiol or their binding to the oestrogen receptor and/or androgen receptor (AR) in the breast. Here, we report on the results of a large and comprehensive study of the association between genetic variation in the AR gene and risk of breast cancer in the National Cancer Institute Breast and Prostate Cancer Cohort Consortium (BPC3). The underlying genetic variation was determined by first sequencing the coding regions of the AR gene in a panel of 95 advanced breast cancer cases. Second, a dense set of markers from the public database was genotyped in a panel of 349 healthy women. The linkage disequilibrium relationships (blocks) across the gene were then identified, and haplotype-tagging single nucleotide polymorphisms (htSNPs) were selected to capture the common genetic variation across the locus. The htSNPs were then genotyped in the nested breast cancer cases and controls from the Cancer Prevention Study II, European Prospective Investigation into Cancer and Nutrition, Multiethnic Cohort, Nurses' Health Study, and Women's Health Study cohorts (5,603 breast cancer cases and 7,480 controls). We found no association between any genetic variation (SNP, haplotype, or the exon 1 CAG repeat) in the AR gene and risk of breast cancer, nor were any statistical interactions with known breast cancer risk factors observed. Among postmenopausal Caucasian women, common variants of the AR gene are not associated with risk of breast cancer.


Synthesis and characterization of carbazolide-based iridium PNP pincer complexes. Mechanistic and computational investigation of alkene hydrogenation: evidence for an Ir(III)/Ir(V)/Ir(III) catalytic cycle.
New carbazolide-based iridium pincer complexes ((carb)PNP)Ir(C2H4), 3a, and ((carb)PNP)Ir(H)2, 3b, have been prepared and characterized. The dihydride, 3b, reacts with ethylene to yield the cis-dihydride ethylene complex cis-((carb)PNP)Ir(C2H4)(H)2. Under ethylene this complex reacts slowly at 70 °C to yield ethane and the ethylene complex, 3a. Kinetic analysis establishes that the reaction rate is dependent on ethylene concentration and labeling studies show reversible migratory insertion to form an ethyl hydride complex prior to formation of 3a. Exposure of cis-((carb)PNP)Ir(C2H4)(H)2 to hydrogen results in very rapid formation of ethane and dihydride, 3b. DFT analysis suggests that ethane elimination from the ethyl hydride complex is assisted by ethylene through formation of ((carb)PNP)Ir(H)(Et)(C2H4) and by H2 through formation of ((carb)PNP)Ir(H)(Et)(H2). Elimination of ethane from Ir(III) complex ((carb)PNP)Ir(H)(Et)(H2) is calculated to proceed through an Ir(V) complex ((carb)PNP)Ir(H)3(Et) which reductively eliminates ethane with a very low barrier to return to the Ir(III) dihydride, 3b. Under catalytic hydrogenation conditions (C2H4/H2), cis-((carb)PNP)Ir(C2H4)(H)2 is the catalyst resting state, and the catalysis proceeds via an Ir(III)/Ir(V)/Ir(III) cycle. This is in sharp contrast to isoelectronic (PCP)Ir systems in which hydrogenation proceeds through an Ir(III)/Ir(I)/Ir(III) cycle. The basis for this remarkable difference is discussed.


Prevention of 1,2-dimethylhydrazine-induced colon tumorigenesis by HMG-CoA reductase inhibitors, pravastatin and simvastatin, in ICR mice.
3-Hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitors, pravastatin (Pr) and simvastatin (Si), suppressed 1,2-dimethylhydrazine (DMH)-induced colon cancer development in female ICR mice. All mice received an i.p. injection of 10 mg DMH/kg body wt once weekly for 15 weeks. Pr was administered at 0.01, 0.005 and 0.001% levels in drinking water, and Si at 0.01 and 0.002% levels in the diet. All animals had access to Pr or Si throughout the experiments which were terminated at weeks 25 or 30. Histologically most of the tumors were well-differentiated adenocarcinomas. The incidence of colon tumors examined at weeks 25 or 30 was reduced by 67% in the 0.01% Pr group, by 30% in the 0.005% Pr and 0.01% Si groups, and by 24% in the 0.001% Pr and 0.002% Si groups, compared with their respective controls. However, the differences did not reach statistical significance. The number of tumors per mouse was significantly reduced in all groups administered Pr and Si except the 0.001% Pr group as compared to their respective controls. The results from those three independent experiments seem to suggest that HMG-CoA reductase inhibitors may prevent colon tumorigenesis in laboratory animal model.


Genomics in 2011: challenges and opportunities.
As we come to the end of 2011, Genome Biology has asked some members of our Editorial Board for their views on the state of play in genomics. What was their favorite paper of 2011? What are the challenges in their particular research area? Who has had the biggest influence on their careers? What advice would they give to young researchers embarking on a career in research?


Molecular dynamics simulations of the ligand-binding domain of an N-methyl-D-aspartate receptor.
The mechanism of partial agonism at N-methyl-D-aspartate receptors is an unresolved issue, especially with respect to the role of protein dynamics. We have performed multiple molecular dynamics simulations (7 x 20 ns) to examine the behavior of the ligand-binding core of the NR1 subunit with a series of ligands. Our results show that water plays an important role in stabilizing different conformations of the core and how a closed cleft conformation of the protein might be stabilized in the absence of ligands. In the case of ligand-bound simulations with both full and partial agonists, we observed that ligands within the binding cleft may undergo distinct conformational changes, without grossly influencing the degree of cleft closure within the ligand-binding domain. In agreement with recently published crystallographic data, we also observe similar changes in backbone torsions corresponding to the hinge region between the two lobes for the partial agonist, D-cycloserine. This observation rationalizes the classification of D-cycloserine as a partial agonist and should provide a basis with which to predict partial agonism in this class of receptor by analyzing the behavior of these torsions with other potential ligands.


Foraging biology predicts food web complexity.
Food webs, the networks of feeding links between species, are central to our understanding of ecosystem structure, stability, and function. One of the key aspects of food web structure is complexity, or connectance, the number of links expressed as a proportion of the total possible number of links. Connectance (complexity) is linked to the stability of webs and is a key parameter in recent models of other aspects of web structure. However, there is still no fundamental biological explanation for connectance in food webs. Here, we propose that constraints on diet breadth, driven by optimal foraging, provide such an explanation. We show that a simple diet breadth model predicts highly constrained values of connectance as an emergent consequence of individual foraging behavior. When combined with features of real food web data, such as taxonomic and trophic aggregation and cumulative sampling of diets, the model predicts well the levels of connectance and scaling of connectance with species richness, seen in real food webs. This result is a previously undescribed synthesis of foraging theory and food web theory, in which network properties emerge from the behavior of individuals and, as such, provides a mechanistic explanation of connectance currently lacking in food web models.


Biosynthesis of the dimethylbenzene moiety of riboflavin and dimethylbenzimidazole: evidence for the involvement of C-1 of a pentose as a precursor.
The relative incorporations of specially labeled pyruvate, lactate, erythritol, D-erythrose, D-ribose, and D-glucose precursors into the dimethylbenzene carbon atoms of the 5,6-dimethylbenzimidazole unit of vitamin B12 by Propionibacterium shermanii have been determined. The incorporation data provide information regarding the putative four-carbon biosynthetic unit which is involved in the formation of 6,7-dimethyl-8-ribityllumazine and which is the source of the eight dimethylbenzene carbon atoms of both 5,6-dimethylbenzimidazole and riboflavin. The relative incorporations of the labeled lactate and pyruvate precursors are not consistent with either acetoin or 2,3-butanedione functioning as the four-carbon biosynthetic unit. The relative incorporations of the labeled hexose, pentose, and tetrose precursors indicate that the observed incorporation of C-1 of the pentose into the dimethylbenzene carbon atoms does not involve metabolism to a tetrose intermediate, but occurs more directly. It is concluded that the C-1 position of a pentose precursor is involved in the formation of the putative four-carbon biosynthetic unit.


Effect of a 90 g/day low-carbohydrate diet on glycaemic control, small, dense low-density lipoprotein and carotid intima-media thickness in type 2 diabetic patients: An 18-month randomised controlled trial.
This study explored the effect of a moderate (90 g/d) low-carbohydrate diet (LCD) in type 2 diabetes patients over 18 months. Ninety-two poorly controlled type 2 diabetes patients aged 20-80 years with HbA1c ≥7.5% (58 mmol/mol) in the previous three months were randomly assigned to a 90 g/d LCD r traditional diabetic diet (TDD). The primary outcomes were glycaemic control status and change in medication effect score (MES). The secondary outcomes were lipid profiles, small, dense low-density lipoprotein (sdLDL), serum creatinine, microalbuminuria and carotid intima-media thickness (IMT). A total of 85 (92.4%) patients completed 18 months of the trial. At the end of the study, the LCD and TDD group consumed 88.0±29.9 g and 151.1±29.8 g of carbohydrates, respectively (p < 0.05). The 18-month mean change from baseline was statistically significant for the HbA1c (-1.6±0.3 vs. -1.0±0.3%), 2-h glucose (-94.4±20.8 vs. -18.7±25.7 mg/dl), MES (-0.42±0.32 vs. -0.05±0.24), weight (-2.8±1.8 vs. -0.7±0.7 kg), waist circumference (-5.7±2.7 vs. -1.9±1.4 cm), hip circumference (-6.1±1.8 vs. -2.9±1.7 cm) and blood pressure (-8.3±4.6/-5.0±3 vs. 1.6±0.5/2.5±1.6 mmHg) between the LCD and TDD groups (p<0.05). The 18-month mean change from baseline was not significantly different in lipid profiles, sdLDL, serum creatinine, microalbuminuria, alanine aminotransferase (ALT) and carotid IMT between the groups. A moderate (90 g/d) LCD showed better glycaemic control with decreasing MES, lowering blood pressure, decreasing weight, waist and hip circumference without adverse effects on lipid profiles, sdLDL, serum creatinine, microalbuminuria, ALT and carotid IMT than TDD for type 2 diabetic patients.


Characterization of chronic pain and somatosensory function in spinal cord injury subjects.
The pathophysiology of the chronic pain following spinal cord injury (SCI) is unclear. In order to study it's underlying mechanism we characterized the neurological profile of SCI subjects with (SCIP) and without (SCINP) chronic pain. Characterization comprised of thermal threshold testing for warmth, cold and heat pain and tactile sensibility testing of touch, graphesthesia and identification of speed of movement of touch stimuli on the skin. In addition, spontaneously painful areas were mapped in SCIP and evoked pathological pain--allodynia, hyperpathia and wind-up pain evaluated for both groups. Both SCIP and SCINP showed similar reductions in both thermal and tactile sensations. In both groups thermal sensations were significantly more impaired than tactile sensations. Chronic pain was present only in skin areas below the lesion with impaired or absent temperature and heat-pain sensibilities. Conversely, all the thermally impaired skin areas in SCIP were painful while painfree areas in the same subjects were normal. In contrast, chronic pain could be found in skin areas without any impairment in tactile sensibilities. Allodynia could only be elicited in SCIP and a significantly higher incidence of pathologically evoked pain (i.e. hyperpathia and wind-up pain) was seen in the chronic pain areas compared to SCINP. We conclude that damage to the spinothalamic tract (STT) is a necessary condition for the occurrence of chronic pain following SCI. However, STT lesion is not a sufficient condition since it could also be found in SCINP. The abnormal evoked pain seen in SCIP is probably due to neuronal hyperexcitability in these subjects. The fact that apparently identical sensory impairments manifest as chronic pain and hyperexcitability in one subject but not in another implies that either genetic predisposition or subtle differences in the nature of spinal injury determine the emergence of chronic pain following SCI.


Neuronal and intestinal protein kinase d isoforms mediate Na+ (salt taste)-induced learning.
Ubiquitously expressed protein kinase D (PKD) isoforms are poised to disseminate signals carried by diacylglycerol (DAG). However, the in vivo regulation and functions of PKDs are poorly understood. We show that the Caenorhabditis elegans gene, dkf-2, encodes not just DKF-2A, but also a second previously unknown isoform, DKF-2B. Whereas DKF-2A is present mainly in intestine, we show that DKF-2B is found in neurons. Characterization of dkf-2 null mutants and transgenic animals expressing DKF-2B, DKF-2A, or both isoforms revealed that PKDs couple DAG signals to regulation of sodium ion (Na+)-induced learning. EGL-8 (a phospholipase Cbeta4 homolog) and TPA-1 (a protein kinase Cdelta homolog) are upstream regulators of DKF-2 isoforms in vivo. Thus, pathways containing EGL-8-TPA-1-DKF-2 enable learning and behavioral plasticity by receiving, transmitting, and cooperatively integrating environmental signals targeted to both neurons and intestine.


Immune-related genetic enrichment in frontotemporal dementia: An analysis of genome-wide association studies.
Converging evidence suggests that immune-mediated dysfunction plays an important role in the pathogenesis of frontotemporal dementia (FTD). Although genetic studies have shown that immune-associated loci are associated with increased FTD risk, a systematic investigation of genetic overlap between immune-mediated diseases and the spectrum of FTD-related disorders has not been performed. Using large genome-wide association studies (GWASs) (total n = 192,886 cases and controls) and recently developed tools to quantify genetic overlap/pleiotropy, we systematically identified single nucleotide polymorphisms (SNPs) jointly associated with FTD-related disorders-namely, FTD, corticobasal degeneration (CBD), progressive supranuclear palsy (PSP), and amyotrophic lateral sclerosis (ALS)-and 1 or more immune-mediated diseases including Crohn disease, ulcerative colitis (UC), rheumatoid arthritis (RA), type 1 diabetes (T1D), celiac disease (CeD), and psoriasis. We found up to 270-fold genetic enrichment between FTD and RA, up to 160-fold genetic enrichment between FTD and UC, up to 180-fold genetic enrichment between FTD and T1D, and up to 175-fold genetic enrichment between FTD and CeD. In contrast, for CBD and PSP, only 1 of the 6 immune-mediated diseases produced genetic enrichment comparable to that seen for FTD, with up to 150-fold genetic enrichment between CBD and CeD and up to 180-fold enrichment between PSP and RA. Further, we found minimal enrichment between ALS and the immune-mediated diseases tested, with the highest levels of enrichment between ALS and RA (up to 20-fold). For FTD, at a conjunction false discovery rate < 0.05 and after excluding SNPs in linkage disequilibrium, we found that 8 of the 15 identified loci mapped to the human leukocyte antigen (HLA) region on Chromosome (Chr) 6. We also found novel candidate FTD susceptibility loci within LRRK2 (leucine rich repeat kinase 2), TBKBP1 (TBK1 binding protein 1), and PGBD5 (piggyBac transposable element derived 5). Functionally, we found that the expression of FTD-immune pleiotropic genes (particularly within the HLA region) is altered in postmortem brain tissue from patients with FTD and is enriched in microglia/macrophages compared to other central nervous system cell types. The main study limitation is that the results represent only clinically diagnosed individuals. Also, given the complex interconnectedness of the HLA region, we were not able to define the specific gene or genes on Chr 6 responsible for our pleiotropic signal. We show immune-mediated genetic enrichment specifically in FTD, particularly within the HLA region. Our genetic results suggest that for a subset of patients, immune dysfunction may contribute to FTD risk. These findings have potential implications for clinical trials targeting immune dysfunction in patients with FTD.


Blood flow in ductus venosus in early uncomplicated pregnancy.
The ductus venosus is the very important part of fetal venous circulation. It plays a central role in return of venous blood from the placenta. This unique shunt carries well-oxygenated blood from the umbilical vein through the inferior atrial inlet on its way across the foramen ovale. Using Doppler ultrasound, it is possible to assess the blood flow in fetal vessels including ductus venosus. It is observed, in animal and human studies, that the typical waveform for blood flow in ductus venosus in early pregnancy can be different depending on numerous conditions e.g. fetal karyotype. This study is performed to assess the physiologic parameters of blood flow in ductus venosus in uncomplicated early pregnancy. 404 women were examined between 11+0 and 13+6 weeks (+ days) of gestation by ultrasound. Fetal crown-rump length (CRL) was measured to assess the gestational age. The assessment of risk of fetal abnormalities was based on nuchal translucency (NT) measurement. The ductus venosus blood flow with color and spectral Doppler was obtained in all patients. The following features were assessed: pulsatility index (PI), and direction of flow (positive/negative) during atrial contraction (wave A). All cases were followed up to 22 weeks of gestation when the control scan was performed. 30 cases were excluded from the uncomplicated group due to: high risk of fetal abnormalities, fetal loss, confirmed fetal abnormalities and utero-placental pathology. 374 women were considered as uncomplicated pregnancy. In both uncomplicated and complicated groups the mean values for pulsatility index (PI) were established. The mean PI value in uncomplicated pregnancies was: 0.91 (SD +/- 0.32). No significant differences between groups were noticed. In 370 cases of uncomplicated pregnancy the A wave direction was positive but in 1.1% of cases the reverse flow in atrial contraction was observed.


A Kelch repeat protein, Cokel1p, associates with microtubules and is involved in appressorium development in Colletotrichum orbiculare.
Kelch repeat proteins are conserved in diverse organisms and some are known to mediate fundamental cellular functions. We isolated the gene CoKEL1, encoding a novel kelch repeat protein, from Colletotrichum orbiculare. Analysis of a cokel1 mutant indicated that CoKEL1 is involved in proper appressorium development and cell wall synthesis. Appressoria produced by cokel1 disruption mutants showed irregular shape and impairment of turgor generation and the mutant appressoria rarely penetrated to form infection hyphae in host epidermal cells. Accordingly, cokel1 mutants had reduced pathogenicity on host leaves compared with the wild type. Furthermore, the cokel1 mutant was more sensitive to cell-wall-degrading enzymes and showed altered labeling with the cell wall stain Calcofluor white. Cokel1p was localized on cortical and spindle microtubules in vegetative hyphae. These results suggest that Cokel1p is a microtubule-associated protein involved in infection-related morphogenesis and pathogenicity. This is the first report that a kelch repeat protein is required for the pathogenicity of a fungal plant pathogen.


Synapses, axonal and dendritic patterns of GABA-immunoreactive neurons in human cerebral cortex.
Gamma-aminobutyric acid (GABA) containing neurons were characterized in human association cortex by a combination of Golgi impregnation and immunohistochemistry. Neurons were Golgi impregnated, gold toned, drawn and then classified on the basis of their dendritic and axonal arborization in layers I-VI. An antiserum to GABA was used to determine which of the impregnated neurons were immunopositive. Twenty-four GABA-positive cells were Golgi impregnated: 7 were bitufted with their dendrites predominantly radially oriented, and 17 were multipolar stellate cells. Three of the multipolar cells with large somata in the deep layers showed dendritic patterns similar to previously described basket cells. Nine of the multipolar stellate cells in layers III-VI showed characteristics of 'neurogliaform' neurons (Ramón y Cajal, 1899). The somata and the dendritic field of these cells were spherical, with diameters of about 10-15 microns and 200 microns, respectively. Their dendrites were smooth and slightly beaded. The axon collaterals were densely distributed in and around the dendritic field, in a spherical area with a diameter of at least 300 microns. The thin axon collaterals had only occasional 'en passant' swellings. Contacts between the axons of neurogliaform cells and the distal dendrites of Golgi-impregnated pyramidal cells were observed. Electron microscopic immunocytochemistry revealed that GABA immunopositive nerve terminals formed symmetric synaptic contacts with somata, with GABA immunonegative and immunopositive dendritic shafts and with dendritic spines. The results show that GABAergic neurons are heterogeneous with respect to their dendritic and axonal patterns. In addition to the chandelier and basket cells, which have been shown in animal studies to contain GABA, other cell types, most prominently the neurogliaform cells, terminating on the distal parts of neurons, also contain GABA and may have a inhibitory function. Many of the GABAergic terminals make synapses on dendritic spines and shafts in the human cerebral cortex.


MicroRNAs regulate survival in oxygen-deprived environments.
Some animals must endure prolonged periods of oxygen deprivation to survive. One such extreme model is the northern crayfish (Orconectes virilis), that regularly survives year-round hypoxic and anoxic stresses in its warm stagnant summer waters and in its cold, ice-locked winter waters. To elucidate the molecular underpinnings of anoxia resistance in this natural model, we surveyed the expression profiles of 76 highly conserved microRNAs in crayfish hepatopancreas and tail muscle from normoxic, acute 2 h anoxia, and chronic 20 h anoxia exposures. MicroRNAs are known to regulate a diverse array of cellular functions required for environmental stress adaptations, and here we explored their role in anoxia tolerance. The tissue-specific anoxia responses observed herein, with 22 anoxia-responsive microRNAs in the hepatopancreas and only four in muscle, suggest that microRNAs facilitate a reprioritization of resources to preserve crucial organ functions. Bioinformatic microRNA target enrichment analysis predicted that the anoxia-downregulated microRNAs in hepatopancreas targeted Hippo signalling, suggesting that cell proliferation and apoptotic signalling are highly regulated in this liver-like organ during anoxia. Compellingly, miR-125-5p, miR-33-5p and miR-190-5p, all known to target the master regulator of oxygen deprivation responses HIF1 (hypoxia inducible factor-1), were anoxia downregulated in the hepatopancreas. The anoxia-increased transcript levels of the oxygen-dependent subunit HIF1α highlight a potential critical role for miRNA-HIF targeting in facilitating a successful anoxia response. Studying the cytoprotective mechanisms in place to protect against the challenges associated with surviving in oxygen-poor environments is critical to elucidating the vast and substantial role of microRNAs in the regulation of metabolism and stress in aquatic invertebrates.


Postoperative changes of spinal curvature and range of motion in adult patients with cervical spinal cord tumors: analysis of 51 cases and review of the literature.
We sought to determine factors likely to influence postoperative changes in cervical spinal curvature and range of motion (ROM) in patients with cervical spinal cord tumors. Fifty-one patients who underwent tumor excision via a single posterior approach were examined. Surgical methods were classified as follows: group A, expansive open-door laminoplasty (ELAP); group B, conventional laminectomy; and group C, hemilaminectomy (HL). Pre- and postoperative cervical spinal curvature, cervical curvature type, and ROM were evaluated using lateral radiographs. Worsening of cervical curvature type was statistically less frequent in group C than in group A or B (P < 0.05), although no significant difference was evident overall between pre- and postoperative cervical curvature indexes or ROM in any group. Cases of C2 surgery showed more frequent curvature worsening (80%) in group A than in other groups (P < 0.05). Postoperative changes of the cervical curvature in patients with cervical spinal cord tumors were greatly influenced by the surgical method; the cervical curvature was particularly unfavorable when patients had C2 decompression. Careful attention to postoperative changes in the spinal curvature is needed when ELAP is performed, especially if surgery involves C2. HL is the best posterior approach for resection of cervical spinal cord tumors from the viewpoint of postoperative spinal curvature and ROM.


Cellulose Nanofibril Hydrogel Tubes as Sacrificial Templates for Freestanding Tubular Cell Constructs.
The merging of defined nanoscale building blocks with advanced additive manufacturing techniques is of eminent importance for the preparation of multiscale and highly functional materials with de novo designed hierarchical architectures. Here, we demonstrate that hydrogels of cellulose nanofibrils (CNF) can be processed into complex shapes, and used as a sacrificial template to prepare freestanding cell constructs. We showcase our approach for the fabrication of hollow fibers using a controlled extrusion through a circular die into a coagulation bath. The dimensions of the hollow fibers are tunable, and the final tubes combine the nanofibrillar porosity of the CNF hydrogel with a submillimeter wall thickness and centimeter-scale length provided by the additive manufacturing technique. We demonstrate that covalent and supramolecular cross-linking of the CNFs can be used to tailor the mechanical properties of the hydrogel tubes within 1 order of magnitude and in an attractive range for the mechanosensation of cells. The resulting tubes are highly biocompatible and allow for the growth of mouse fibroblasts into confluent cell layers in their inner lumen. A detailed screening of several cellulases enables degradation of the scaffolding, temporary CNF hydrogel tube in a quick and highly cell-friendly way, and allows the isolation of coherent cell tubes. We foresee that the growing capabilities of hydrogel printing techniques in combination with the attractive features of CNFs-sustainable, globally abundant, biocompatible and enzymatically degradable-will allow making plant-based biomaterials with hierarchical structures and on-demand degradation useful, for instance, to engineer complex tissue structures to replace animal models, and for implants.


Effect of royal jelly in ovo injection on embryonic growth, hatchability, and gonadotropin levels of pullet breeder chicks.
The objectives of this study were to compare the following: (1) hatchability, (2) chicks' body and internal organs weights, and (3) plasma gonadotropin levels of hatchlings after in ovo administration of royal jelly (RJ) on Day 7 of incubation. Fertile eggs (n = 270) were injected into the air sac or yolk sac with 0.5 mL normal saline solution consisting of four formulations (normal saline solution with antibiotics, ultrafiltrate RJ, pure RJ, and RJ with antibiotics). The eggs were randomly divided into nine groups of 30 eggs each: (i) C: the control eggs received no injection, (ii) ASA: air sac-injected eggs received normal saline solution with antibiotics, (iii) ARJ: air sac-injected eggs received pure RJ, (iv) ARJA: air sac-injected eggs received RJ with antibiotics, (v) ARJF: air sac-injected eggs received RJ ultrafiltrate solution, (vi) YSA: yolk sac-injected eggs received normal saline solution with antibiotics, and (vii) YRJ: yolk sac-injected eggs received pure RJ, (viii) YRJA: yolk sac-injected eggs received RJ with antibiotics, and (ix) YRJF: yolk sac-injected eggs received ultrafiltrate RJ solution. Hatchability rate was lower in ARJ (46.7%), ARJA (43.3%), ARJF (43.3%), and YRJF (46.7%) groups than in the control (80.0%; P < 0.05). Hatchability rate in ASA (70.0%), YSA (66.7%), YRJ (66.7%), and YRJA (63.3%) groups were comparable to the control (P > 0.05). In ovo injection of RJ and or RJ with antibiotics in both sacs increased chicks' body weight (CWT), heart weight (HWT), and liver weight (LWT) and FSH and LH levels compared with control (P < 0.05). CWT in YRJ (37.02 g), YRJA (37.03 g), ARJ (36.82 g), and ARJA (36.89 g) groups were higher than control (34.9 g; P < 0.05). Similarly, HWT significantly increased in YRJ (0.22 g), YRJA (0.21 g), ARJ (0.20 g), and ARJA (0.20 g) in comparison to control (0.18 g; P < 0.05). In addition, LWT were higher in YRJ (0.83 g), YRJA (0.82g), ARJ (0.81g), and ARJA (0.81g) than control (0.72 g; P < 0.05). Six hours post-hatch, the mean plasma FSH and LH levels in ARJ (1.13 and 2.80 mIu/mL), YRJ (1.32 and 3.36 mIu/mL), ARJA (1.23 and 2.95 mIu/mL), and YRJA (1.31 and 3.28 mIu/mL) groups were higher than in the control (0.56 and 1.48 mIu/mL, P < 0.05). We concluded that in ovo administration of RJ or RJ with antibiotics might be an effective method to increase CWT, chicks' internal organs weights, and LH and FSH secretion rate without deleterious effect on hatchability. However, further research should be conducted to determine the putative endocrine disruptive effects of RJ and its byproducts.


Functional outcomes in high risk ICU patients in Central Australia: a prospective case series.
This case series reports the functional outcomes of a prospective group of patients, thought to be at high risk for future morbidity, admitted to a rural intensive care unit (ICU) for a life-threatening illness. This prospective longitudinal observational study conducted between February and August 2009 in the Alice Springs Hospital ICU included patients considered 'high risk', as evidenced by profound physiological derangement. The participants were prospectively recruited when pre-defined criteria were met. Functional outcomes were measured by performance in the six-minute walk test, and the ability to undertake activities of daily living. Persisting morbidity was crudely measured by hospital re-admission rate. Mortality was measured at 6 months. Eighteen patients consented to take part in the study. Fourteen were Indigenous, and 14 were medical patients. Six-minute walk distance did not improve between ICU discharge and 6 months, and was significantly below that predicted. Almost all patients achieved scores consistent with full independence in basic activities of daily living. Five achieved scores consistent with independence in domestic activities of daily living. Twelve required at least one re-admission, with half the Indigenous subgroup requiring three or more re-admissions. There were four deaths, all Indigenous patients, and three were homeless.< This study demonstrates that follow up in this group at 6 months is both feasible and valuable. There is evidence of persisting morbidity, and increased mortality, particularly among Indigenous patients. Further avenues of research are suggested, including the need for a large multi-centre prospective study.


Growth of confined cancer spheroids: a combined experimental and mathematical modelling approach.
A critical step in the dissemination of ovarian cancer is the formation of multicellular spheroids from cells shed from the primary tumour. The objectives of this study were to apply bioengineered three-dimensional (3D) microenvironments for culturing ovarian cancer spheroids in vitro and simultaneously to build on a mathematical model describing the growth of multicellular spheroids in these biomimetic matrices. Cancer cells derived from human epithelial ovarian carcinoma were embedded within biomimetic hydrogels of varying stiffness and grown for up to 4 weeks. Immunohistochemistry, imaging and growth analyses were used to quantify the dependence of cell proliferation and apoptosis on matrix stiffness, long-term culture and treatment with the anti-cancer drug paclitaxel. The mathematical model was formulated as a free boundary problem in which each spheroid was treated as an incompressible porous medium. The functional forms used to describe the rates of cell proliferation and apoptosis were motivated by the experimental work and predictions of the mathematical model compared with the experimental output. This work aimed to establish whether it is possible to simulate solid tumour growth on the basis of data on spheroid size, cell proliferation and cell death within these spheroids. The mathematical model predictions were in agreement with the experimental data set and simulated how the growth of cancer spheroids was influenced by mechanical and biochemical stimuli including matrix stiffness, culture duration and administration of a chemotherapeutic drug. Our computational model provides new perspectives on experimental results and has informed the design of new 3D studies of chemoresistance of multicellular cancer spheroids.


Rediscovery of the Yesso scallop pathogen Perkinsus qugwadi in Canada, and development of PCR tests.
Perkinsus qugwadi, a pathogenic protozoan parasite of Yesso scallops Patinopecten yessoensis, is found only in cultured populations in British Columbia, Canada. This pathogen was first identified in 1988 and caused significant mortalities at some locations during the early 1990s. Prevalence of infection decreased dramatically following 1995, and the disease was last reported in 1997, leading to speculation that the Yesso scallop stocks in Canada had developed resistance to the disease, or that P. qugwadi had disappeared. However, the present study revealed that infection with P. qugwadi and associated mortality is still occurring in scallops from at least one location in British Columbia. One of the PCR tests developed for P. qugwadi detected the parasite in a 105-fold dilution of DNA extracted from a heavily infected sample and detected 52% more positive scallops than histology; however, the assay also cross-reacted with P. honshuensis and P. olseni. The other PCR test was less sensitive and detected 34% more positives, but did not react to any of the other Perkinsus species tested, suggesting that these PCR tests are powerful tools for screening for the presence of P. qugwadi. Phylogenetic analysis of 1796 bp of SSU rRNA gene sequence clearly indicated that P. qugwadi is positioned basally to other Perkinsus species.


Factors associated with the need of biventricular mechanical circulatory support in children with advanced heart failure.
Postimplantation right ventricular dysfunction is associated with increased morbidity and mortality in ventricular assist device (VAD) recipients. This study aimed to determine the preoperative risk factors for severe right heart failure needing biventricular mechanical circulatory support in children with end-stage heart failure. We reviewed data from 84 children supported with long-term VADs at the German Heart Institute Berlin between January 1999 and October 2010. Right ventricular assist device (RVAD) support was needed for 24 (29%) patients, and the other 60 (71%) were implanted with left ventricular assist devices (LVADs). The median age at implantation was 7 years (12 days-18 years), and the median support time was 41 days (1-432 days). Of the 84 patients, the overall survival to transplantation or recovery of ventricular function was 69%. Compared with children implanted with LVAD, patients receiving biventricular support had significantly higher postoperative mortality (P = 0.04). The multivariate logistic regression indicated that decreased milrinone use was the only preoperative factor independently associated with increased requirement for biventricular support (odds ratio: 0.1, 95% confidence interval: 0.04-0.64, P = 0.01). Children treated with milrinone preoperatively showed improved survival after implantation (P = 0.04). Paediatric patients needing biventricular support had significantly higher postoperative mortality. Preoperative milrinone use might decrease the risk of severe right ventricular failure requiring additional RVAD insertion and improve postimplantation survival in children with advanced heart failure.


Progesterone and interferon tau regulate leukemia inhibitory factor receptor and IL6ST in the ovine uterus during early pregnancy.
The actions of leukemia inhibitory factor (LIF) via LIF receptor (LIFR) and its co-receptor, IL6 signal transducer (IL6ST), are implicated in uterine receptivity to conceptus implantation in a number of species including sheep. The present study determined the effects of the estrous cycle, pregnancy, progesterone (P4), and interferon tau (IFNT) on the expression of LIFR and IL6ST in the ovine uterus. LIFR mRNA and protein were localized to the endometrial luminal (LE) and superficial glandular epithelia (sGE), whereas IL6ST mRNA and protein were localized primarily in the middle to deep GE. Both LIFR and IL6ST mRNAs and protein were more abundant in pregnant than cyclic ewes and increased from days 10 to 20 of pregnancy. Treatment of ovariectomized ewes with P4 and/or infusion of ovine IFNT increased LIFR and IL6ST in endometrial LE/sGE and GE respectively. Co-expression of LIFR and IL6ST as well as phosphorylated STAT3 was observed only in the upper GE of the endometrium as well as in the conceptus trophectoderm on days 18 and 20. In mononuclear trophectoderm and GE cells, LIF elicited an increase in phosphorylated STAT3 and MAPK3/1 MAPK proteins. Collectively, these results suggest that LIFR and IL6ST are both stimulated by IFNT and regulated by P4 in a complex stage- and cell-specific manner, and support the hypothesis that LIF exerts effects on the endometrial GE as well as conceptus trophectoderm during early pregnancy in sheep. Thus, LIF and STAT3 may have biological roles in endometrial function and trophectoderm growth and differentiation.


Resonant absorption and scattering suppression of localized surface plasmons in Ag particles on green LED.
The metallic-structure dependent localized surface plasmons (LSPs) coupling behaviors with InGaN QWs in a green LED epitaxial wafer are investigated by optical transmission, scanning electron microscopy (SEM) and photoluminescence (PL) measurements. Ag nanoparticles (NPs) are formed by thermal annealing Ag layer on the green LED wafer. SEM images show that for higher annealing temperature and/or thicker deposited Ag layer, larger Ag NPs can be produced, leading to the redshift of absorption peaks in the transmission spectra. Time resolved PL (TRPL) measurements indicate when LSP-MQW coupling occurs, PL decay rate is greatly enhanced especially at the resonant wavelength 560 nm. However, the PL intensity is suppressed by 3.5 folds compared to the bare LED. The resonant absorption and PL suppression are simulated by three dimension finite-difference-time-domain (FDTD), which suggests that Ag particle with smaller size and lower height lead to the larger dissipation of LSP.


A novel two-component system PdeK/PdeR regulates c-di-GMP turnover and virulence of Xanthomonas oryzae pv. oryzae.
Two-component systems (TCS) consisting of histidine kinases (HK) and response regulators (RR) play essential roles in bacteria to sense environmental signals and regulate cell functions. One type of RR is involved in metabolism of cyclic diguanylate (c-di-GMP), a ubiquitous bacterial second messenger. Although genomic studies predicted a large number of them existing in different bacteria, only a few have been studied. In this work, we characterized a novel TCS consisting of PdeK(PXO_01018)/PdeR(PXO_ 01019) from Xanthomonas oryzae pv. oryzae, which causes the bacterial leaf blight of rice. PdeR (containing GGDEF, EAL, and REC domains) was shown to have phosphodiesterase (PDE) activity in vitro by colorimetric assays and high-performance liquid chromatography analysis. The PDE activity of full-length PdeR needs to be triggered by HK PdeK. Deletion of pdeK or pdeR in X. oryzae pv. oryzae PXO99(A) had attenuated its virulence on rice. ΔpdeK and ΔpdeR secreted less exopolysaccharide than the wild type but there were no changes in terms of motility or extracellular cellulase activity, suggesting the activity of PdeK/PdeR might be specific.


Noncirrhotic portal hypertension in the adult: case report and review of the literature.
Noncirrhotic portal hypertension results from thrombosis of the extrahepatic portal vein that subsequently is recanalized. Liver function is preserved. In the adult, esophageal variceal hemorrhage is the most common presentation and may occur years after the portal vein thrombosis. We report the case of a 34-year-old man who presented with recurrent esophageal variceal hemorrhage. After ultrasonographic and angiographic evaluation, a diagnosis of idiopathic noncirrhotic portal hypertension was made. Due to recurrent esophageal variceal bleeding, the patient required surgical intervention to control bleeding. The incidence of noncirrhotic portal hypertension is unknown. Multiple etiologies may cause the disorder, although nearly half are idiopathic. The pathogenesis, clinical manifestations, diagnostic evaluation, natural history, prognosis, and management of noncirrhotic portal hypertension are discussed. Endoscopic management of esophageal variceal bleeding is the preferred therapy. However, when endoscopic treatment fails to control variceal hemorrhage, a distal splenorenal shunt is likely to be the most successful operation.


Arxula adeninivorans recombinant guanine deaminase and its application in the production of food with low purine content.
Purines of exogenous and endogenous sources are degraded to uric acid in human beings. Concentrations >6.8 mg uric acid/dl serum cause hyperuricemia and its symptoms. Pharmaceuticals and the reduction of the intake of purine-rich food are used to control uric acid levels. A novel approach to the latter proposition is the enzymatic reduction of the purine content of food by purine-degrading enzymes. Here we describe the production of recombinant guanine deaminase by the yeast Arxula adeninivorans LS3 and its application in food. In media supplemented with nitrogen sources hypoxanthine or adenine, guanine deaminase (AGDA) gene expression is induced and intracellular accumulation of guanine deaminase (Agdap) protein occurs. The characteristics of the guanine deaminase isolated from wild-type strain LS3 and a transgenic strain expressing the AGDA gene under control of the strong constitutive TEF1 promoter were determined and compared. Both enzymes were dimeric and had temperature optima of 55°C with high substrate specificity for guanine and localisation in both the cytoplasm and vacuole of yeast. The enzyme was demonstrated to reduce levels of guanine in food. A mixture of guanine deaminase and other purine degradation enzymes will allow the reduction of purines in purine-rich foods.


Isolation and purification of human biliary vesicles with potent cholesterol-nucleation-promoting activity.
1. Cholesterol nucleation is a critical step in the formation of cholesterol gallstones. This nucleation takes place after aggregation and fusion of cholesterol-rich biliary vesicles, a process probably modulated by biliary proteins. The present study was conducted to identify specific proteins associated with native cholesterol-rich biliary vesicles and to explore their effect on the cholesterol-nucleation time of supersaturated artificial bile. 2. Hepatic bile was obtained from six patients with cholesterol gallstone disease. Biliary vesicles were isolated by ultracentrifugation and were purified by gel filtration chromatography. A small amount of protein (less than 1% by weight) remained associated with the purified cholesterol-rich biliary vesicles. The electrophoretic profile of these proteins was remarkably similar in all six patients, showing the presence of at least six polypeptides (of molecular mass from 52 to 200 kDa), five of them having carbohydrate residues (except the 52 kDa one). The effect of reconstituted biliary vesicle solutions, containing their specific vesicular proteins, on cholesterol-nucleation time was studied by mixing the vesicle solution with artificial supersaturated bile. A potent cholesterol-pronucleating activity, reflected in a 20-70% reduction in nucleation time, was present in the biliary vesicle solutions compared with control solutions having a similar lipid composition. The pronucleating activity disappeared on heating and was not detected in the micellar fraction containing the major proportion of biliary proteins. 3. These results indicate that cholesterol-rich biliary vesicles containing a unique and defined glycoprotein profile can be isolated and purified from human hepatic bile. The potent cholesterol-pronucleating activity of the biliary vesicles from patients with gallstones was unrelated to their lipid composition or cholesterol content.(ABSTRACT TRUNCATED AT 250 WORDS)


Bub1 kinase and Sgo1 modulate pericentric chromatin in response to altered microtubule dynamics.
Tension sensing of bioriented chromosomes is essential for the fidelity of chromosome segregation. The spindle assembly checkpoint (SAC) conveys lack of tension or attachment to the anaphase promoting complex. Components of the SAC (Bub1) phosphorylate histone H2A (S121) and recruit the protector of cohesin, Shugoshin (Sgo1), to the inner centromere. How the chromatin structural modifications of the inner centromere are integrated into the tension sensing mechanisms and the checkpoint are not known. We have identified a Bub1/Sgo1-dependent structural change in the geometry and dynamics of kinetochores and the pericentric chromatin upon reduction of microtubule dynamics. The cluster of inner kinetochores contract, whereas the pericentric chromatin and cohesin that encircle spindle microtubules undergo a radial expansion. Despite its increased spatial distribution, the pericentric chromatin is less dynamic. The change in dynamics is due to histone H2A phosphorylation and Sgo1 recruitment to the pericentric chromatin, rather than microtubule dynamics. Bub1 and Sgo1 act as a rheostat to regulate the chromatin spring and maintain force balance. Through histone H2A S121 phosphorylation and recruitment of Sgo1, Bub1 kinase softens the chromatin spring in response to changes in microtubule dynamics. The geometric alteration of all 16 kinetochores and pericentric chromatin reflect global changes in the pericentromeric region and provide mechanisms for mechanically amplifying damage at a single kinetochore microtubule.


Evaluation of 10 genotypes of sweetpotato for fries.
The potential use of selected genotypes of sweetpotato for fries was evaluated. Moisture, fat, browning index, and beta-carotene content of fries from 10 sweetpotato genotypes, Apomuden, Bohye, Nanungungu, Otoo, CIP442162, Patron, CIP440390, Obare, Dadanyuie, and Tu-Purple, were determined by standard methods. Sensorial properties of the fries were further evaluated by an in-house trained panel of eight members using standard methods. Fat content was highest in CIP442162 and low in Dadanyuie, while beta-carotene content decreased by 44.27% in Bohye and 13.20% in Nanungungu after frying. Browning index was highest in the orange-fleshed and purple genotypes, but this was mostly due to their flesh colors and not the frying conditions. Fries from orange-fleshed genotypes, Apomuden and Nanungungu, were considered to be sweeter than the other genotypes. Detection of caramel and starch (rawness sensation) was very low for all genotypes assessed. Oily mouthcoat, moistness, and sogginess were detected in mostly the orange-fleshed genotypes, particularly Apomuden and Nanungungu. Tu-Purple, Bohye, and CIP440390 produced moderately crunchy fries and had the highest score for desirable attributes compared with the other genotypes. Tu-Purple, Bohye, and CIP440390 could be explored in commercial production of fries for enhanced utilization of developed sweetpotato genotypes.


Effect of amino acids on the stability of spray freeze-dried immunoglobulin G in sugar-based matrices.
The purpose of this study was to prepare spray freeze-dried particles of immunoglobulin G (IgG) using various combinations of trehalose and different amino acids (leucine, phenylalanine, arginine, cysteine, and glycine), and investigate the effect of the amino acids on the stability of IgG during the spray freeze-drying (SFD) process and storage. The morphology and structural integrity of the processed particles were evaluated by physical and spectroscopic techniques. SFD-processed IgG without any excipient resulted in the formation of aggregates corresponding to approximately 14% of IgG. In contrast, IgG formulations stabilized using an optimal level of leucine, phenylalanine, or glycine in the presence of trehalose displayed aggregates <2.2%. In particular, phenylalanine combined with trehalose was most effective in stabilizing IgG against shear, freezing, and dehydration stresses during SFD. Arginine and cysteine were destabilizers displaying aggregation and fragmentation of IgG, respectively. Aggregation and fragmentation were evaluated by dynamic light scattering, ultraviolet spectrophotometry, size-exclusion chromatography, and microchip capillary gel electrophoresis. The IgG formulations prepared with leucine, phenylalanine, or glycine in the presence of trehalose showed good stability after storage at 40 °C and 75% relative humidity for 2 months. Thus, a combination of the excipients trehalose and uncharged, nonpolar amino acids appears effective for production of stable SFD IgG formulations.


Day-to-day fluctuation of point-of-care circulating cathodic antigen test scores and faecal egg counts in children infected with Schistosoma mansoni in Ethiopia.
Determining the variation of circulating cathodic antigen (CCA) in urine and egg counts variation in stool between days in Schistosoma mansoni (S. mansoni) infected individuals is vital to decide whether or not to rely on a single-sample test for diagnosis of Schistosomiasis. In this study, the magnitude of day-to-day variation in urine-CCA test scores and in faecal egg counts was evaluated in school children in Ethiopia. A total of 620 school children (age 8 to 12 years) were examined for S. mansoni infection using double Kato-Katz and single urine-CCA cassette methods (batch 32727) on three consecutive days. The prevalence of S. mansoni infection was 81.1% based on triple urine-CCA-cassette test and 53.1% based on six Kato-Katz thick smears. Among the study participants, 26.3% showed fluctuation in urine CCA and 32.4% showed fluctuation in egg output. Mean egg count as well as number of cases in each class of intensity and intensity of cassette band color varied over the three days of examination. Over 85% of the children that showed day-to-day variations in status of S. mansoni infection from negative to positive or vice versa by the Kato-Katz and the CCA methods had light intensity of infection. The fluctuation in both the CCA test scores and faecal egg count was not associated with age and sex. The current study showed day-to-day variation in CCA and Kato-Katz test results of children infected with S. mansoni. This indicates the necessity of more than one urine or stool samples to be collected on different days for more reliable diagnosis of S. mansoni infection in low endemic areas.


A comparison of methods to measure daily cigarillo consumption among adolescents and young adults.
Cigarillo use is widespread among young people. Accurate assessment of cigarillo consumption is necessary to inform and evaluate tobacco research, but is complicated by product sharing and irregular use. This study compares a conventional approach with a detailed approach for measuring cigarillo consumption. Data are drawn from a cross-sectional, web-based survey of 1089 young (aged 15-28 years) cigarillo smokers. The conventional measure of cigarillo consumption employs two common tobacco use items-the number of days a product was smoked in the past month and the average number of products smoked per day. The detailed measure uses a time line follow-back procedure to assess product use on each of the past 7 days, both in a group and alone. Paired t-tests compare daily cigarillo use estimates from the two methods overall, and are stratified by sample characteristics and behaviours; associations with multiple factors are examined simultaneously using linear regression. Compared with the conventional measure, the detailed measure yields significantly higher daily consumption estimates for moderate and high-level users and for non-daily tobacco users, and significantly lower estimates for those who always share products and daily tobacco users. Differences remain after controlling for demographics and product use behaviours. There are no differences by gender, age, race or multiple product use. The two measurement methods yield significantly different consumption estimates based on sharing behaviour, regularity of use and use level. Improving accuracy in the measurement of tobacco product consumption is important and timely for tobacco control research and policy.


Interferon-λ and interleukin 22 act synergistically for the induction of interferon-stimulated genes and control of rotavirus infection.
The epithelium is the main entry point for many viruses, but the processes that protect barrier surfaces against viral infections are incompletely understood. Here we identified interleukin 22 (IL-22) produced by innate lymphoid cell group 3 (ILC3) as an amplifier of signaling via interferon-λ (IFN-λ), a synergism needed to curtail the replication of rotavirus, the leading cause of childhood gastroenteritis. Cooperation between the receptor for IL-22 and the receptor for IFN-λ, both of which were 'preferentially' expressed by intestinal epithelial cells (IECs), was required for optimal activation of the transcription factor STAT1 and expression of interferon-stimulated genes (ISGs). These data suggested that epithelial cells are protected against viral replication by co-option of two evolutionarily related cytokine networks. These data may inform the design of novel immunotherapy for viral infections that are sensitive to interferons.


A comprehensive mechanistic model of iron metabolism in Saccharomyces cerevisiae.
The ironome of budding yeast (circa 2019) consists of approximately 139 proteins and 5 nonproteinaceous species. These proteins were grouped according to location in the cell, type of iron center(s), and cellular function. The resulting 27 groups were used, along with an additional 13 nonprotein components, to develop a mesoscale mechanistic model that describes the import, trafficking, metallation, and regulation of iron within growing yeast cells. The model was designed to be simultaneously mutually autocatalytic and mutually autoinhibitory - a property called autocatinhibitory that should be most realistic for simulating cellular biochemical processes. The model was assessed at the systems' level. General conclusions are presented, including a new perspective on understanding regulatory mechanisms in cellular systems. Some unsettled issues are described. This model, once fully developed, has the potential to mimic the phenotype (at a coarse-grain level) of all iron-related genetic mutations in this simple and well-studied eukaryote.


Providers' beliefs and behaviors regarding antibiotic prescribing and antibiotic resistance in persons with spinal cord injury or disorder.
Persons with spinal cord injury or disorder (SCI/D) are at increased risk for antibiotic resistance because of recurrent infections and subsequent use of antibiotics. However, there are no studies focused on providers who care for these patients and their perceptions regarding antibiotic use and resistance. To characterize SCI/D provider behavior and attitudes about antibiotic prescribing and resistance. Anonymous internet-based, cross-sectional survey. A total of 314 SCI/D clinicians who prescribe antibiotics (physicians, physician assistants, and nurse practitioners). A total of 118 providers responded (37.6% response rate) including 80 physicians, 20 nurse practitioners, and 18 physician assistants. The majority of respondents agreed with statements regarding the societal impact of antibiotic resistance; only 17.8% agreed that they prescribed antibiotics more than they should, but 61.0% agreed that patient demand was a major reason for prescribing unnecessary antibiotics. The most frequent problematic organisms reported were: methicillin-resistant Staphylococcus aureus (83.1%), multidrug-resistant Pseudomonas (61.0%), and Clostridium difficile (57.6%). The most frequent antibiotics selected for outpatient treatment of community-acquired pneumonia treatment, based on a clinical scenario were azithromycin (36.4%) and respiratory fluoroquinolones (22.9%). These data show that the respondents are aware of and concerned with the problem of antibiotic resistance in their practice. Clinician respondents also endorsed the need to improve their own knowledge and that of their colleagues regarding appropriate antibiotic prescribing. These findings suggest that interventions should focus on provider education, particularly regarding appropriate antibiotic prescribing.


Numerical simulation of micro-particle rotation by the acoustic viscous torque.
We present the first numerical simulation setup for the calculation of the acoustic viscous torque on arbitrarily shaped micro-particles inside general acoustic fields. Under typical experimental conditions, the particle deformation plays a minor role. Therefore, the particle is modeled as a rigid body which is free to perform any time-harmonic and time-averaged translation and rotation. Applying a perturbation approach, the viscoacoustic field around the particle is resolved to obtain the time-averaged driving forces for a subsequent acoustic streaming simulation. For some acoustic fields, the near-boundary streaming around the fluid-suspended particle induces surface forces on the nonrotating particle that integrate into a non-zero acoustic viscous torque. In the equilibrium state, this torque is compensated by an equal and opposite drag torque due to the particle rotation. The rotation-induced flow field is superimposed on the acoustic streaming field to obtain the total fluid motion around the rotating particle. In this work, we only consider cases within the Rayleigh limit even though the presented numerical model is not strictly limited to this regime. After a validation by analytical solutions, the numerical model is applied to challenging experimental cases. For an arbitrary particle density, we consider particle sizes that can be comparable to the viscous boundary layer thickness. This important regime has not been studied before because it lies beyond the validity limits of the available analytical solutions. The detailed numerical analysis in this work predicts nonintuitive phenomena, including an inversion of the rotation direction. Our numerical model opens the door to explore a wide range of experimentally relevant cases, including non-spherical particle rotation. As a step toward application fields such as micro-robotics, the rotation of a prolate ellipsoid is studied.


Diabetic mastopathy, a clinicopathological correlation of 34 cases.
Diabetic mastopathy (DM) is an unusual fibrous disease of the breast. We report 34 cases with clinicopathological correlation and follow-ups. Patients mean age was 57 years and 24 had histories of diabetes mellitus. The DM presented as a palpable breast mass in 85.3% of the cases clinically. Image findings were often as an irregular hypoechoic mass (44.4%) on ultrasound and negative or non-specific density (67.6%) on mammogram. Histologically, we found that the majority of DM occurred in the upper outer/central part of the breast (76%) and the size ranged from 0.5 to 3.7 cm. All cases showed lymphocytic lobulitis and vasculitis, keloidal fibrosis, lobular atrophy and varying degree of epithelioid fibroblasts. Examining the surrounding normal tissue of the excision specimens revealed that the margins of DM were often poorly circumscribed and irregularly demarcated from the sounding breast tissue. Vascular calcifications were identified in 10 of 24 excisional specimens, suggesting a hypothesis that vascular damage and scar/wounding healing process could contribute to the pathogenesis of DM. Twenty-one cases had repeated procedures for uncertain descriptive diagnosis. This is the largest study of diabetic mastopathy to date with clinical and imagining correlation. Recognition of this benign entity is important to better understanding the pathogenesis and to avoid unnecessary procedures.


Heterobifunctional Dyes: Highly Fluorescent Linkers Based on Cyanine Dyes.
Herein, we present a new synthetic route to cyanine-based heterobifunctional dyes and their application as fluorescent linkers between polymers and biomolecules. The synthesized compounds, designed in the visible spectral range, are equipped with two different reactive groups for highly selective conjugation under physiological conditions. By applying indolenine precursors with functionalized benzenes, we achieved water-soluble asymmetric cyanine dyes bearing maleimido and N-hydroxysuccinimidyl functionalities in a three-step synthesis. Spectroscopic characterization revealed good molar absorption coefficients and moderate fluorescence quantum yields. Further reaction with polyethylene glycol yielded dye-polymer conjugates that were subsequently coupled to the antibody cetuximab, often applied in cancer therapy. Successful coupling was confirmed by mass shifts detected by gel electrophoresis. Receptor-binding studies and live-cell imaging revealed that labeling did not alter the biological function. In sum, we provided a successful synthetic pathway to rigid heterobifunctional cyanine dyes that are applicable as fluorescent linkers, for example, for connecting antibodies with macromolecules. Our approach contributes to the field of bioconjugation chemistry, such as antibody-drug conjugates by combining diagnostic and therapeutic approaches.


Early detection of reverse transcriptase activity in plasma of neonates infected with HIV-1: a comparative analysis with RNA-based and DNA-based testing using polymerase chain reaction.
Plasma viral load from 71 HIV-1-infected neonates was measured by using Amp-RT, an ultrasensitive quantitative reverse transcriptase (RT) assay and by nucleic acid sequence-based amplification (NASBA), an RNA-based quantitative assay. Results were then compared with those obtained from detection of proviral DNA in peripheral blood mononuclear cells (PBMCs) by polymerase chain reaction (PCR) using Turnbull analysis. At 5 days of life, 50% of neonates were positive by Amp-RT, 30% were NASBA positive, and 20% were DNA-PCR positive. Through the first 12 days of life, Amp-RT was more sensitive than either NASBA or DNA-PCR in detecting HIV-1 infection. Amp-RT values correlated well with NASBA RNA values, with an overall Pearson's r = 0.63 (95% confidence interval [CI], 0.40-0.78). In proportional hazards analysis of infants aged 14 to 61 days (N = 31), a one-log increase in RNA-based viral load was associated with a > fivefold risk of disease progression when using the U.S. Centers for Disease Control and Prevention (CDC) clinical Category C (CDC-C) or death as an endpoint (p =.014). Kaplan-Meier analysis of these data found that RNA viral loads were able to predict disease progression using CDC-C/death as an endpoint (p = .013). Early quantitative viral load measurements may assist clinicians in diagnosing HIV-1 infection, stratifying risk of disease progression, and implementing a treatment plan using highly active antiretroviral therapy for infants within the first few weeks of life.


Lean body mass and muscle function in head and neck cancer patients and healthy individuals--results from the DAHANCA 25 study.
Loss of lean body mass is common following radiotherapy in patients with head and neck squamous cell carcinoma (HNSCC) and may reduce maximal muscle strength and functional performance. However, the associations between lean body mass, muscle strength and functional performance are unclear and no studies in HNSCC patients have compared the levels of these variables to the levels seen in healthy individuals. The purpose of the present study was to investigate the associations between lean body mass, maximal muscle strength and functional performance in HNSCC patients and to compare the levels of these variables after radiotherapy and after progressive resistance training with the levels in healthy individuals. Lean body mass (dual energy X-ray absorptiometry), maximal muscle strength (isokinetic dynamometry) and functional performance (10 m max gait speed, 30 s chair rise, 30 s arm curl, stair climb) from HNSCC patients from the DAHANCA 25 trials and data from 24 healthy individuals were included. Lean body mass and maximal muscle strength were significantly associated according to the gender and age-adjusted linear regression model (p < 0.0001). In addition, maximal muscle strength were associated with 30 s arm curl performance, 10 m max gait speed and 30 s chair rise (p < 0.0001). Multiple regression analyses showed that HNSCC patients expressed significant lower levels of the investigated variables after radiotherapy than healthy individuals (p < 0.0001), and that all differences were evened out after training. Significant associations were found between lean body mass, maximal muscle strength and functional performance in HNSCC patients. Patients expressed lower levels of these variables compared with healthy individuals, suggesting that lean body mass is a clinically relevant health factor in HNSCC patients.


The effect of non-pharmacological sleep interventions on depression symptoms: A meta-analysis of randomised controlled trials.
Poor sleep is a significant risk factor for depression across the lifespan and sleep problems have been hypothesised to contribute to the onset and maintenance of depression symptoms. However, sleep problems are usually not a direct target of interventions for depression. A range of non-pharmacological treatments can reduce sleep problems but it is unclear whether these interventions also reduce other depression symptoms. The aim of this review was to examine whether non-pharmacological interventions for sleep problems are effective in reducing symptoms of depression. We carried out a systematic search for randomised controlled trials of non-pharmacological sleep interventions that measured depression symptoms as an outcome. Forty-nine trials (n = 5908) were included in a random effects meta-analysis. The pooled standardised mean difference for depression symptoms after treatment for sleep problems was -0.45 (95% CI: -0.55,-0.36). The size of the effect on depression symptoms was moderated by the size of the effect on subjective sleep quality. In studies of participants with mental health problems, sleep interventions had a large effect on depression symptoms (d = -0.81, 95% CI: -1.13,-0.49). The findings indicate that non-pharmacological sleep interventions are effective in reducing the severity of depression, particularly in clinical populations. This suggests that non-pharmacological sleep interventions could be offered as a treatment for depression, potentially improving access to treatment.


Nutrition practices and knowledge of college varsity athletes: a follow-up.
The purpose of this study was to determine nutrition knowledge and behavior of division IA college athletes and to compare such knowledge and behavior with data from a similar survey conducted in 1992. Surveys with distribution instructions and statements of confidentiality were sent randomly to strength and conditioning coordinators (SCCs) at 16 universities. Survey results indicated that women (60.6%) received more nutrition information than men (49.5). SCCs and athletic trainers were the primary nutrition sources for men, whereas university classes and nutritionists were primary for women. Much information was obtained from arguable sources such as magazines, family members, and coaches. Only 3, 11.7, and 29.5% correctly identified recommended percents of total calorie intake for protein, fat, and carbohydrates, respectively. Thirty-seven percent correctly identified the role of vitamins and 54.4% for protein. Creatine and vitamin/mineral supplements were the most common for men and women, respectively. Despite previous recommendations regarding nutrition education of high-level athletes, diminutive changes have occurred in the past 6 years.