Tissue transglutaminase regulates focal adhesion kinase/AKT activation by modulating PTEN expression in pancreatic cancer cells.
Pancreatic ductal adenocarcinoma (PDAC) progresses rapidly and exhibits profound resistance to treatment. We recently reported that a great majority of PDAC tumors and tumor cell lines express elevated levels of tissue transglutaminase (TG2). Here, we provide first evidence that TG2 expression in PDAC cells results in constitutive activation of focal adhesion kinase/AKT by modulating the expression of the tumor suppressor phosphatase PTEN. Using PDAC cell lines, we determined the effect of TG2 overexpression on PTEN stability and functions. We confirmed the correlation between TG2 expression and PTEN levels in a few (n=51) PDAC tumor samples. We observed that expression of TG2 is inversely correlated with PTEN expression in PDAC cells. Ectopic expression of TG2 inhibited PTEN phosphorylation and promoted its degradation by ubiquitin-proteasomal pathway. Conversely, down-regulation of TG2 by small interfering RNA up-regulated PTEN expression. Clinical relevance of these results was evident in an athymic nude mouse model where down-regulation of endogenous TG2 caused a significant retardation in PDAC xenograft growth. Importantly, the analysis of 51 tumor samples from patients with stage II PDAC revealed that overexpression of TG2 was associated with loss of PTEN expression (P=0.023; odds ratio, 4.1). In multivariate analysis, TG2-mediated loss of PTEN was a prognostic factor for overall survival in patients with stage II pancreatic ductal carcinoma independent of tumor stage/lymph node status and tumor differentiation (P=0.01). TG2 expression in PDAC promotes degradation of PTEN by ubiquitin-proteasomal pathway and results in constitutive activation of focal adhesion kinase/AKT cell survival signaling.


Vasoconstriction during volume expansion is independent of central control.
We have previously demonstrated that, in the absence of the rapid acting neural and hormonal controllers of blood pressure, an acute blood volume expansion of only 5% in unanesthetized rats caused an increase in total peripheral resistance (TPR) of 22%. Either whole body autoregulation or the release of a putative ouabainlike factor from the central nervous system (CNS) could have explained these responses. The purpose of the present study was to investigate the possible contribution of a centrally released ouabainlike factor to the vasoconstriction response observed during volume expansion. Because the anteroventral third ventricle (AV3V) region is proposed to be important in the control of this putative factor, we compared the hemodynamic responses to blood volume expansion in rats with AV3V lesions (n = 6), sham lesions (n = 6), and total CNS ablation (n = 6). The results of our studies showed that neither AV3V lesion nor CNS ablation reduced the increases of total peripheral resistance seen with blood volume expansion. We conclude that centrally released factors are not required for vasoconstriction in response to acute volume expansion and that regional autoregulatory mechanisms result in a net increase of systemic vascular resistance (i.e., whole body autoregulation).


Archaeal and Bacterial Communities Associated with the Surface Mucus of Caribbean Corals Differ in Their Degree of Host Specificity and Community Turnover Over Reefs.
Comparative studies on the distribution of archaeal versus bacterial communities associated with the surface mucus layer of corals have rarely taken place. It has therefore remained enigmatic whether mucus-associated archaeal and bacterial communities exhibit a similar specificity towards coral hosts and whether they vary in the same fashion over spatial gradients and between reef locations. We used microbial community profiling (terminal-restriction fragment length polymorphism, T-RFLP) and clone library sequencing of the 16S rRNA gene to compare the diversity and community structure of dominant archaeal and bacterial communities associating with the mucus of three common reef-building coral species (Porites astreoides, Siderastrea siderea and Orbicella annularis) over different spatial scales on a Caribbean fringing reef. Sampling locations included three reef sites, three reef patches within each site and two depths. Reference sediment samples and ambient water were also taken for each of the 18 sampling locations resulting in a total of 239 samples. While only 41% of the bacterial operational taxonomic units (OTUs) characterized by T-RFLP were shared between mucus and the ambient water or sediment, for archaeal OTUs this percentage was 2-fold higher (78%). About half of the mucus-associated OTUs (44% and 58% of bacterial and archaeal OTUs, respectively) were shared between the three coral species. Our multivariate statistical analysis (ANOSIM, PERMANOVA and CCA) showed that while the bacterial community composition was determined by habitat (mucus, sediment or seawater), host coral species, location and spatial distance, the archaeal community composition was solely determined by the habitat. This study highlights that mucus-associated archaeal and bacterial communities differ in their degree of community turnover over reefs and in their host-specificity.


Decreased efficacy of drugs targeting the vascular endothelial growth factor pathway by the epigenetic silencing of FLT1 in renal cancer cells.
The vascular endothelial growth factor (VEGF)-VEGF receptor (VEGFR) signaling pathway is involved in cancer-related biological functions and is a therapeutic target in cancer. However, the influence of epigenetic regulation of VEGF-VEGFR signaling-related genes remains unclear. Here, we evaluated the effects of FLT1 and KDR promoter hypermethylation combined with drugs targeting VEGF-VEGFR signaling on cancer-related phenotypes in renal cancer cells (RCCs) and examined changes in FLT1 and KDR promoter hypermethylation in tissues from patients with renal cancer. In vitro experiments were performed to evaluate the effects of beavacizumab (an anti-VEGF antibody), an anti-FLT1 peptide, an anti-KDR antibody, and the VEGFR tyrosine kinase inhibitors (TKIs) sunitinib and axitinib in 13 RCC lines with different levels of FLT1 and/or KDR promoter methylation and in 2 FLT1 or KDR in vitro knockdown models. The synergistic effects of sunitinib and axitinib treatment were also evaluated in four RCC lines having different levels of FLT1 and/or KDR methylation. In our in vitro experiments, bevacizumab and an anti-KDR antibody did not affect the proliferation of RCCs having FLT1 and/or KDR hypermethylation. In contrast, in RCCs with FLT1 hypermethylation, proliferation inhibition was counteracted by treatment with an anti-FLT1 peptide and both VEGF-TKIs (sunitinib and axitinib). Demethylation with sunitinib or axitinib synergistically increased proliferation inhibition in the RCCs exhibiting FLT1 hypermethylation. Using in vitro FLT1 or KDR knockdown models, decreased proliferation inhibition following anti-FLT1 peptide, sunitinib, and axitinib treatment was observed only in FLT1-knockdown cells. In patients with renal cancer who received sunitinib, FLT1 promoter methylation was higher in renal cancer tissues from eight nonresponders (stable or progressive disease assessed by the Response Evaluation Criteria in Solid Tumors) than in cancer tissues from five responders (complete response or partial response). The present data showed that hypermethylated FLT1 was important for the efficacy of anti-VEGF/VEGFR drugs targeting FLT1 or intracellular VEGFR signaling. FLT1 hypermethylation causing alterations of FLT1 function could serve as a useful biomarker for predicting changes in FLT1 status in RCCs.


A vaccine against Alzheimer`s disease: anything left but faith?
Alzheimer's disease looms as a profound and growing threat to future human health. The disease is thought to be primarily driven by aberrant proteolysis of the amyloid precursor protein (APP) and amyloid beta (Aβ) plaque deposition. We provide an overview of the molecular pathology that leads to an increase in Aβ peptide accumulation, of the mechanism of action for antibody mediated therapies and of the therapeutic vaccines that target Aβ under development. We also discuss the rationale for using vaccines in the early stages of the disease. The major components of β-amyloid plaques are Aβ1-42 and Aβ1-40 peptides derived from the APP. Reducing these plaques by means of passive or active vaccination against Aβ-peptides has been a long-running endeavor but with disappointing results as the impact on disease progression has been minimal. The data gathered to date could suggest that antibodies do not work, mainly because the studies have not been performed in an optimal fashion. The emerging views are that patients should be treated earlier, ideally in the prodromal or symptom free stage, antibody levels have to be high and the correct epitope must be targeted. More clinical trials to fully explore the potential of vaccines are therefore warranted.


Setting the Agenda for Personal Health Records in Low- and Middle-Income Countries.
In the developed world, Personal Health Records (PHRs) have been demonstrated to improve patient adherence, reduce medical errors, improve patient-provider communication, improve chronic disease management, and promote behavior change. PHRs have not been widely adopted in low- and middle-income countries. There is rising use of smartphones, adoption of national-level electronic health systems, and change in historical perceptions on acceptance of technology. It is thus an opportune time to critically examine the potential role and approaches to PHRs in LMICs.


Clinical use of autologous micro-fragmented fat progressively restores pain and function in shoulder osteoarthritis.
Aim: We aim to show that the use of nondigested micro-fragmented adipose tissue (MFat™, Lipogems®) is a viable alternative for treatment of joint pain and inflammation associated dysfunction in shoulder osteoarthritis (OA). Materials & methods: A total of 25 subjects with OA received an injection of MFat™ and were followed at 6, 18 and 52 weeks intervals. Quantitative analysis of pain and function modalities were performed using the visual analog scale and the disabilities of the arm, shoulder and hand, respectively. Results: All study participants reported significant progressive improvement (p < 0.001) from baseline in visual analog scale and disabilities of the arm, shoulder and hand in shoulder OA cases up to a year post. Conclusion: MFat™ therapy improves pain and function in patients with shoulder OA and can provide a long-term alternative to surgical intervention.


Hypothalamic roles of mTOR complex I: integration of nutrient and hormone signals to regulate energy homeostasis.
Mammalian or mechanistic target of rapamycin (mTOR) senses nutrient, energy, and hormone signals to regulate metabolism and energy homeostasis. mTOR activity in the hypothalamus, which is associated with changes in energy status, plays a critical role in the regulation of food intake and body weight. mTOR integrates signals from a variety of "energy balancing" hormones such as leptin, insulin, and ghrelin, although its action varies in response to these distinct hormonal stimuli as well as across different neuronal populations. In this review, we summarize and highlight recent findings regarding the functional roles of mTOR complex 1 (mTORC1) in the hypothalamus specifically in its regulation of body weight, energy expenditure, and glucose/lipid homeostasis. Understanding the role and underlying mechanisms behind mTOR-related signaling in the brain will undoubtedly pave new avenues for future therapeutics and interventions that can combat obesity, insulin resistance, and diabetes.


Ras1 and a putative guanine nucleotide exchange factor perform crucial steps in signaling by the sevenless protein tyrosine kinase.
We have conducted a genetic screen for mutations that decrease the effectiveness of signaling by a protein tyrosine kinase, the product of the Drosophila melanogaster sevenless gene. These mutations define seven genes whose wild-type products may be required for signaling by sevenless. Four of the seven genes also appear to be essential for signaling by a second protein tyrosine kinase, the product of the Ellipse gene. The putative products of two of these seven genes have been identified. One encodes a ras protein. The other locus encodes a protein that is homologous to the S. cerevisiae CDC25 protein, an activator of guanine nucleotide exchange by ras proteins. These results suggest that the stimulation of ras protein activity is a key element in the signaling by sevenless and Ellipse and that this stimulation may be achieved by activating the exchange of GTP for bound GDP by the ras protein.


Adding Postal Follow-Up to a Web-Based Survey of Primary Care and Gastroenterology Clinic Physician Chiefs Improved Response Rates but not Response Quality or Representativeness.
This study assessed whether postal follow-up to a web-based physician survey improves response rates, response quality, and representativeness. We recruited primary care and gastroenterology chiefs at 125 Veterans Affairs medical facilities to complete a 10-min web-based survey on colorectal cancer screening and diagnostic practices in 2010. We compared response rates, response errors, and representativeness in the primary care and gastroenterology samples before and after adding postal follow-up. Adding postal follow-up increased response rates by 20-25 percentage points; markedly greater increases than predicted from a third e-mail reminder. In the gastroenterology sample, the mean number of response errors made by web responders (0.25) was significantly smaller than the mean number made by postal responders (2.18), and web responders provided significantly longer responses to open-ended questions. There were no significant differences in these outcomes in the primary care sample. Adequate representativeness was achieved before postal follow-up in both samples, as indicated by the lack of significant differences between web responders and the recruitment population on facility characteristics. We conclude adding postal follow-up to this web-based physician leader survey improved response rates but not response quality or representativeness.


Variational fractional-spin density-functional theory for diradicals.
Accurate computation of singlet-triplet energy gaps of diradicals remains a challenging problem in density-functional theory (DFT). In this work, we propose a variational extension of our previous work [D. H. Ess, E. R. Johnson, X. Q. Hu, and W. T. Yang, J. Phys. Chem. A 115, 76 (2011)], which applied fractional-spin density-functional theory (FS-DFT) to diradicals. The original FS-DFT approach assumed equal spin-orbital occupancies of 0.5 α-spin and 0.5 β-spin for the two degenerate, or nearly degenerate, frontier orbitals. In contrast, the variational approach (VFS-DFT) optimizes the total energy of a singlet diradical with respect to the frontier-orbital occupation numbers, based on a full configuration-interaction picture. It is found that the optimal occupation numbers are exactly 0.5 α-spin and 0.5 β-spin for diradicals such as O(2), where the frontier orbitals belong to the same multidimensional irreducible representation, and VFS-DFT reduces to FS-DFT for these cases. However, for diradicals where the frontier orbitals do not belong to the same irreducible representation, the optimal occupation numbers can vary between 0 and 1. Furthermore, analysis of CH(2) by VFS-DFT and FS-DFT captures the (1)A(1) and (1)B(1) states, respectively. Finally, because of the static correlation error in commonly used density functional approximations, both VFS-DFT and FS-DFT calculations significantly overestimate the singlet-triplet energy gaps for disjoint diradicals, such as cyclobutadiene, in which the frontier orbitals are confined to separate atomic centers.


Enumerating the economic cost of antimicrobial resistance per antibiotic consumed to inform the evaluation of interventions affecting their use.
Antimicrobial resistance (AMR) poses a colossal threat to global health and incurs high economic costs to society. Economic evaluations of antimicrobials and interventions such as diagnostics and vaccines that affect their consumption rarely include the costs of AMR, resulting in sub-optimal policy recommendations. We estimate the economic cost of AMR per antibiotic consumed, stratified by drug class and national income level. The model is comprised of three components: correlation coefficients between human antibiotic consumption and subsequent resistance; the economic costs of AMR for five key pathogens; and consumption data for antibiotic classes driving resistance in these organisms. These were used to calculate the economic cost of AMR per antibiotic consumed for different drug classes, using data from Thailand and the United States (US) to represent low/middle and high-income countries. The correlation coefficients between consumption of antibiotics that drive resistance in S. aureus, E. coli, K. pneumoniae, A. baumanii, and P. aeruginosa and resistance rates were 0.37, 0.27, 0.35, 0.45, and 0.52, respectively. The total economic cost of AMR due to resistance in these five pathogens was $0.5 billion and $2.9 billion in Thailand and the US, respectively. The cost of AMR associated with the consumption of one standard unit (SU) of antibiotics ranged from $0.1 for macrolides to $0.7 for quinolones, cephalosporins and broad-spectrum penicillins in the Thai context. In the US context, the cost of AMR per SU of antibiotic consumed ranged from $0.1 for carbapenems to $0.6 for quinolones, cephalosporins and broad spectrum penicillins. The economic costs of AMR per antibiotic consumed were considerable, often exceeding their purchase cost. Differences between Thailand and the US were apparent, corresponding with variation in the overall burden of AMR and relative prevalence of different pathogens. Notwithstanding their limitations, use of these estimates in economic evaluations can make better-informed policy recommendations regarding interventions that affect antimicrobial consumption and those aimed specifically at reducing the burden of AMR.


Characterization of a novel ranavirus isolated from grouper Epinephelus tauvina.
A large icosahedral virus was isolated from diseased grouper Epinephelus tauvina. The virus grew well in several cultured fish cell lines, with stable and high infectivity after serial passages in grouper cell line (GP). The virus was sensitive to both acid and heat treatments. Virus replication was inhibited by 5-iodo-2-deoxyuridine (IUDR), indicative of a DNA-containing genome. The virus infectivity was reduced with ether treatment, suggesting that the virus was lipid-enveloped. Electron micrographs showed abundant cytoplasmic icosahedral virons in the virus-infected GP cells. The size of the intracellular nucleocapsid was 154 nm between the opposite sides, or 176 nm between the opposite vertices with an inner electron-dense core of 93 nm. Virus particles were released through budding from plasma membranes with a size of 200 nm in diameter. SDS-PAGE of purified virus revealed 20 structural protein bands and a major capsid protein (MCP) of 49 kDa. A DNA fragment of approximately 500 nucleotides was successfully amplified by polymerase chain reaction (PCR) using the primers from conserved regions of the MCP gene of frog virus 3 (FV3), the type species of Ranavirus. Subsequent multiple alignment and phylogenetic analysis showed that the newly isolated grouper virus was closely related to largemouth bass virus (LMBV), FV3 and Regina ranavirus (RRV). Our data suggests that the virus isolate is a novel member of genus Ranavirus, family Iridoviridae. We tentatively name the virus as Singapore grouper iridovirus (SGIV). SGIV was able to cause serious systemic disease capable of killing 96% of grouper fry.


Tuning of synapse number, structure and function in the cochlea.
Cochlear inner hair cells (IHCs) transmit acoustic information to spiral ganglion neurons through ribbon synapses. Here we have used morphological and physiological techniques to ask whether synaptic mechanisms differ along the tonotopic axis and within IHCs in the mouse cochlea. We show that the number of ribbon synapses per IHC peaks where the cochlea is most sensitive to sound. Exocytosis, measured as membrane capacitance changes, scaled with synapse number when comparing apical and midcochlear IHCs. Synapses were distributed in the subnuclear portion of IHCs. High-resolution imaging of IHC synapses provided insights into presynaptic Ca(2+) channel clusters and Ca(2+) signals, synaptic ribbons and postsynaptic glutamate receptor clusters and revealed subtle differences in their average properties along the tonotopic axis. However, we observed substantial variability for presynaptic Ca(2+) signals, even within individual IHCs, providing a candidate presynaptic mechanism for the divergent dynamics of spiral ganglion neuron spiking.


Tracing immature myelin in acute disseminated encephalomyelitis.
Inherent abnormalities of myelin have been suggested in the pathogenesis of multiple sclerosis (MS). We investigated myelin in acute disseminated encephalomyelitis (ADEM) patients by magnetic resonance spectroscopy (MRS) and cerebrospinal fluid (CSF) analysis for citrulline, a marker of immature myelin. A citrulline peak was observed in the normal appearing white matter of 7/15 patients and of 1/10 age-matched neurological controls (p=0.08). CSF citrulline was elevated in 4/6 patients. Alterations in the composition of myelin might predispose to or follow acute inflammatory disorders of the central nervous system.


Immunogenic Consensus Sequence T helper Epitopes for a Pan-Burkholderia Biodefense Vaccine.
Biodefense vaccines against Category B bioterror agents Burkholderia pseudomallei (BPM) and Burkholderia mallei (BM) are needed, as they are both easily accessible to terrorists and have strong weaponization potential. Burkholderia cepaciae (BC), a related pathogen, causes chronic lung infections in cystic fibrosis patients. Since BPM, BM and BC are all intracellular bacteria, they are excellent targets for T cell-based vaccines. However, the sheer volume of available genomic data requires the aid of immunoinformatics for vaccine design. Using EpiMatrix, ClustiMer and EpiAssembler, a set of immunoinformatic vaccine design tools, we screened the 31 available Burkholderia genomes and performed initial tests of our selections that are candidates for an epitope-based multi-pathogen vaccine against Burkholderia species. Immunoinformatics analysis of 31 Burkholderia genomes yielded 350,004 9-mer candidate vaccine peptides of which 133,469 had perfect conservation across the 10 BM genomes, 175,722 had perfect conservation across the 11 BPM genomes and 40,813 had perfect conservation across the 10 BC genomes. Further screening with EpiMatrix yielded 54,010 high-scoring Class II epitopes; these were assembled into 2,880 longer highly conserved 'immunogenic consensus sequence' T helper epitopes. 100% of the peptides bound to at least one HLA class II allele in vitro, 92.7% bound to at least two alleles, 82.9% to three, and 75.6% of the binding results were consistent with the immunoinformatics analysis. Our results show it is possible to rapidly identify promiscuous T helper epitopes conserved across multiple Burkholderia species and test their binding to HLA ligands in vitro. The next step in our process will be to test the epitopes ex vivo using peripheral leukocytes from BC, BPM infected humans and for immunogenicity in human HLA transgenic mice. We expect that this approach will lead to development of a licensable, pan-Burkholderia biodefense vaccine.


The Cancer Cell Dissemination Machinery as an Immunosuppressive Niche: A New Obstacle Towards the Era of Cancer Immunotherapy.
Although cancer immunotherapy has resulted in unpreceded survival benefits to subsets of oncology patients, accumulating evidence from preclinical animal models suggests that the immunosuppressive tumor microenvironment remains a detrimental factor limiting benefit for many patient subgroups. Recent efforts on lymphocyte-mediated immunotherapies are primarily focused on eliminating cancer foci at primary and metastatic sites, but few studies have investigated the impact of these therapies on the highly complex process of cancer cell dissemination. The metastatic cascade involves the directional streaming of invasive/migratory tumor cells toward specialized blood vessel intravasation gateways, called TMEM doorways, to the peripheral circulation. Importantly, this process occurs under the auspices of a specialized tumor microenvironment, herewith referred to as "Dissemination Trajectory", which is supported by an ample array of tumor-associated macrophages (TAMs), skewed towards an M2-like polarization spectrum, and which is also vital for providing microenvironmental cues for cancer cell invasion, migration and stemness. Based on pre-existing evidence from preclinical animal models, this article outlines the hypothesis that dissemination trajectories do not only support the metastatic cascade, but also embody immunosuppressive niches, capable of providing transient and localized immunosubversion cues to the migratory/invasive cancer cell subpopulation while in the act of departing from a primary tumor. So long as these dissemination trajectories function as "immune deserts", the migratory tumor cell subpopulation remains efficient in evading immunological destruction and seeding metastatic sites, despite administration of cancer immunotherapy and/or other cytotoxic treatments. A deeper understanding of the molecular and cellular composition, as well as the signaling circuitries governing the function of these dissemination trajectories will further our overall understanding on TAM-mediated immunosuppression and will be paramount for the development of new therapeutic strategies for the advancement of optimal cancer chemotherapies, immunotherapies, and targeted therapies.


Health literacy and self-care of patients with heart failure.
Today's complex healthcare system relies heavily on sophisticated self-care regimens. To navigate the system and follow self-care protocols, patients must be able to understand and use health information, which requires health literacy. However, nearly 90 million Americans lack the necessary health literacy skills to adequately care for themselves in the face of a complex healthcare system and self-care regimens. Understanding how to effectively care for one's self is thought to improve heart failure symptoms and patient outcomes, but little is actually known about how health literacy influences self-care in patients with heart failure. The purpose of this pilot study was to examine the relationship between health literacy and self-care of patients with heart failure. Patients with a diagnosis of heart failure were recruited from a variety of community settings. Participants completed the Short-Form Test of Functional Health Literacy (measured health literacy), the Self-care Index of Heart Failure (measured self-care maintenance, management, and confidence), and a demographic questionnaire. Spearman ρ correlations were used to assess the strength of the relationship between health literacy level and self-care scores. Among the 49 participants recruited, health literacy was positively related to self-care maintenance (Rs = 0.357, P = .006). Health literacy had a negative relationship with self-care management (Rs = -0.573, P = .001). There was no association between health literacy and self-care confidence (Rs = 0.201, P = .083). This project provides preliminary data regarding the association between health literacy and self-care in heart failure, showing support for higher health-literate patients performing more self-care maintenance, which has been shown to improve patient outcomes in heart failure. Patients with higher health literacy trended toward having greater self-care confidence, which can increase the likelihood of performing self-care, but this finding was not statistically significant. It was unexpected to find that lower health-literate patients performed more self-care management.


Molecular Mechanism for Azeotrope Formation in Ethanol/Benzene Binary Mixtures through Gibbs Ensemble Monte Carlo Simulation.
Azeotropes have been studied for decades due to the challenges they impose on separation processes but fundamental understanding at the molecular level remains limited. Although molecular simulation has demonstrated its capability of predicting mixture vapor-liquid equilibrium (VLE) behaviors, including azeotropes, its potential for mechanistic investigation has not been fully exploited. In this study, we use the united atom transferable potentials for phase equilibria (TraPPE-UA) force field to model the ethanol/benzene mixture, which displays a positive azeotrope. Gibbs ensemble Monte Carlo (GEMC) simulation is performed to predict the VLE phase diagram, including an azeotrope point. The results accurately agree with experimental measurements. We argue that the molecular mechanism of azeotrope formation cannot be fully understood by studying the mixture liquid-state stability at the azeotrope point alone. Rather, azeotrope occurrence is only a reflection of the changing relative volatility between the two components over a much wider composition range. A thermodynamic criterion is thus proposed on the basis of the comparison of partial excess Gibbs energy between the components. In the ethanol/benzene system, molecular energetics shows that with increasing ethanol mole fraction, its volatility initially decreases but later plateaus, while benzene volatility is initially nearly constant and only starts to decrease when its mole fraction is low. Analysis of the mixture liquid structure, including a detailed investigation of ethanol hydrogen-bonding configurations at different composition levels, reveals the underlying molecular mechanism for the changing volatilities responsible for the azeotrope.


Variation in susceptibility to Bacillus thuringiensis toxins among unselected strains of Plutella xylostella.
So far, the only insect that has evolved resistance in the field to Bacillus thuringiensis toxins is the diamondback moth (Plutella xylostella). Documentation and analysis of resistant strains rely on comparisons with laboratory strains that have not been exposed to B. thuringiensis toxins. Previously published reports show considerable variation among laboratories in responses of unselected laboratory strains to B. thuringiensis toxins. Because different laboratories have used different unselected strains, such variation could be caused by differences in bioassay methods among laboratories, genetic differences among unselected strains, or both. Here we tested three unselected strains against five B. thuringiensis toxins (Cry1Aa, Cry1Ab, Cry1Ac, Cry1Ca, and Cry1Da) using two bioassay methods. Tests of the LAB-V strain from The Netherlands in different laboratories using different bioassay methods yielded only minor differences in results. In contrast, side-by-side comparisons revealed major genetic differences in susceptibility between strains. Compared with the LAB-V strain, the ROTH strain from England was 17- to 170-fold more susceptible to Cry1Aa and Cry1Ac, respectively, whereas the LAB-PS strain from Hawaii was 8-fold more susceptible to Cry1Ab and 13-fold more susceptible to Cry1Da and did not differ significantly from the LAB-V strain in response to Cry1Aa, Cry1Ac, or Cry1Ca. The relative potencies of toxins were similar among LAB-V, ROTH, and LAB-PS, with Cry1Ab and Cry1Ac being most toxic and Cry1Da being least toxic. Therefore, before choosing a standard reference strain upon which to base comparisons, it is highly advisable to perform an analysis of variation in susceptibility among field and laboratory populations.


Cbl-mediated degradation of Lyn and Fyn induced by constitutive fibroblast growth factor receptor-2 activation supports osteoblast differentiation.
Fibroblast growth factors (FGFs) play an important regulatory role in skeletal development and bone formation. However, the FGF signaling mechanisms controlling osteoblast function are poorly understood. Here, we identified a role for the Src family members Lyn and Fyn in osteoblast differentiation promoted by constitutive activation of FGF receptor-2 (FGFR2). We show that the overactive FGFR2 S252W mutation induced decreased Src family kinase tyrosine phosphorylation and activity associated with decreased Lyn and Fyn protein expression in human osteoblasts. Pharmacological stimulation of Src family kinases or transfection with Lyn or Fyn vectors repressed alkaline phosphatase (ALP) up-regulation induced by overactive FGFR2. Inhibition of proteasome activity restored normal Lyn and Fyn expression and ALP activity in FGFR2 mutant osteoblasts. Immunoprecipitation studies showed that Lyn, Fyn, and FGFR2 interacted with the ubiquitin ligase c-Cbl and ubiquitin. Transfection with c-Cbl in which the RING finger was disrupted or with c-Cbl with a point mutation that abolishes the binding ability of the Cbl phosphotyrosine-binding domain restored Src kinase activity and Lyn, Fyn, and FGFR2 levels and reduced ALP up-regulation in mutant osteoblasts. Thus, constitutive FGFR2 activation induces c-Cbl-dependent Lyn and Fyn proteasome degradation, resulting in reduced Lyn and Fyn kinase activity, increased ALP expression, and FGFR2 down-regulation. This reveals a common Cbl-mediated negative feedback mechanism controlling Lyn, Fyn, and FGFR2 degradation in response to overactive FGFR2 and indicates a role for Cbl-dependent down-regulation of Lyn and Fyn in osteoblast differentiation induced by constitutive FGFR2 activation.


Seasonality of global and Arctic black carbon processes in the Arctic Monitoring and Assessment Programme models.
This study quantifies black carbon (BC) processes in three global climate models and one chemistry transport model, with focus on the seasonality of BC transport, emissions, wet and dry deposition in the Arctic. In the models, transport of BC to the Arctic from lower latitudes is the major BC source for this region. Arctic emissions are very small. All models simulated a similar annual cycle of BC transport from lower latitudes to the Arctic, with maximum transport occurring in July. Substantial differences were found in simulated BC burdens and vertical distributions, with Canadian Atmospheric Global Climate Model (CanAM) (Norwegian Earth System Model, NorESM) producing the strongest (weakest) seasonal cycle. CanAM also has the shortest annual mean residence time for BC in the Arctic followed by Swedish Meteorological and Hydrological Institute Multiscale Atmospheric Transport and Chemistry model, Community Earth System Model, and NorESM. Overall, considerable differences in wet deposition efficiencies in the models exist and are a leading cause of differences in simulated BC burdens. Results from model sensitivity experiments indicate that convective scavenging outside the Arctic reduces the mean altitude of BC residing in the Arctic, making it more susceptible to scavenging by stratiform (layer) clouds in the Arctic. Consequently, scavenging of BC in convective clouds outside the Arctic acts to substantially increase the overall efficiency of BC wet deposition in the Arctic, which leads to low BC burdens and a more pronounced seasonal cycle compared to simulations without convective BC scavenging. In contrast, the simulated seasonality of BC concentrations in the upper troposphere is only weakly influenced by wet deposition in stratiform clouds, whereas lower tropospheric concentrations are highly sensitive.


TBCD links centriologenesis, spindle microtubule dynamics, and midbody abscission in human cells.
Microtubule-organizing centers recruit alpha- and beta-tubulin polypeptides for microtubule nucleation. Tubulin synthesis is complex, requiring five specific cofactors, designated tubulin cofactors (TBCs) A-E, which contribute to various aspects of microtubule dynamics in vivo. Here, we show that tubulin cofactor D (TBCD) is concentrated at the centrosome and midbody, where it participates in centriologenesis, spindle organization, and cell abscission. TBCD exhibits a cell-cycle-specific pattern, localizing on the daughter centriole at G1 and on procentrioles by S, and disappearing from older centrioles at telophase as the protein is recruited to the midbody. Our data show that TBCD overexpression results in microtubule release from the centrosome and G1 arrest, whereas its depletion produces mitotic aberrations and incomplete microtubule retraction at the midbody during cytokinesis. TBCD is recruited to the centriole replication site at the onset of the centrosome duplication cycle. A role in centriologenesis is further supported in differentiating ciliated cells, where TBCD is organized into "centriolar rosettes". These data suggest that TBCD participates in both canonical and de novo centriolar assembly pathways.


Severe neonatal complication of transverse lie after preterm premature rupture of membranes.
Both transverse lie and preterm premature rupture of membranes (PPROM) are associated with neonatal morbidity and mortality. We present a neonate born at 29 weeks gestation with severe birth trauma after PPROM and transverse lie. The patient had extensive swelling and areas of desquamated and necrotic skin of the right lower limb. Neonatal compartment syndrome (NCS) was suspected. Perfusion of the limb improved after decompressing subcutaneous incisions. A fetus in transverse lie may be mechanically damaged in the case of PPROM, especially at an early gestational age. Early recognition is of great interest in the management and prognosis of NCS.


Dexamethasone destabilizes cyclooxygenase 2 mRNA by inhibiting mitogen-activated protein kinase p38.
The stability of cyclooxygenase 2 (Cox-2) mRNA is regulated positively by proinflammatory stimuli acting through mitogen-activated protein kinase (MAPK) p38 and negatively by anti-inflammatory glucocorticoids such as dexamethasone. A tetracycline-regulated reporter system was used to investigate mechanisms of regulation of Cox-2 mRNA stability. Dexamethasone was found to destabilize beta-globin-Cox-2 reporter mRNAs by inhibiting p38. This inhibition occurred at the level of p38 itself: stabilization of reporter mRNA by a kinase upstream of p38 was blocked by dexamethasone, while stabilization by a kinase downstream of p38 was insensitive to dexamethasone. Inhibition of p38 activity by dexamethasone was observed in a variety of cell types treated with different activating stimuli. Furthermore, inhibition of p38 was antagonized by the anti-glucocorticoid RU486 and was delayed and actinomycin D sensitive, suggesting that ongoing glucocorticoid receptor-dependent transcription is required.


Lean management-the journey from toyota to healthcare.
The evolution of production systems is tightly linked to the story of Toyota Motor Company (TMC) that has its roots around 1918. The term "lean" was coined in 1990 following the exploration of the Toyota model that led to the "transference" thesis sustaining the concept that manufacturing problems and technologies are universal problems faced by management and that these concepts can be emulated in non-Japanese enterprises. Lean is a multi-faceted concept and requires organizations to exert effort along several dimensions simultaneously; some consider a successful implementation either achieving major strategic components of lean, implementing practices to support operational aspects, or providing evidence that the improvements are sustainable in the long term. The article explores challenges and opportunities faced by organizations that intend incorporating lean management principles and presents the specific context of the healthcare industry. Finally, the concepts of "essential few" and customer value are illustrated through a simple example of process change following lean principles, which was implemented in a dental school in the United States.


Community Acquired Burkholderia cepacia Bacteraemia Presenting as MODS in an Immunocompetent Individual: An Unusual Case.
Burkholderia cepacia has been recognized as a group of highly virulent organisms known as Burkholderia cepacia complex (Bcc). Bcc are ubiquitous in nature and most commonly found in moist environment, on plant roots and soil. Because of its high intrinsic antibiotic resistance, Bcc is a major cause of morbidity and mortality in hospitalized patients. It is reported most commonly in immunocompromised patients especially in patients with cystic fibrosis. Here, we report a rare case report of bacteraemia by Burkholderia cepacia in an immunocompetent male, who presented with fever and Multiple Organ Dysfunction Syndrome (MODS). Burkholderia cepacia was isolated from his blood culture, which he acquired from his work place. He was treated successfully and discharged after negative blood culture.


Survival from breast, colon, lung, ovarian and rectal cancer by geographical remoteness in New South Wales, Australia, 2000-2008.
This study aims to compare survival from breast, colon, lung, ovarian and rectal cancer by geographical remoteness in New South Wales (NSW). Retrospective population-wide registry study. NSW, Australia. A total of 107 060 NSW residents, who were diagnosed with any of the five cancers between 01 January 2000 and 31 December 2008. Kaplan-Meier survival curves and proportional hazards regression were used to compare survival by geographical remoteness of residence at diagnosis, controlling for gender, age and extent of disease at diagnosis. Remoteness was classified using standard definitions: major city, inner regional (InnReg), outer regional (OutReg) and remote (including very remote). Significant differences in survival (likelihood of death) were identified in all five cancers: breast (adjusted hazard ratio(HR) = 1.22 (95% confidence interval (CI), 1.001-1.48) in regionalised and HR = 1.30 (1.02-1.64) in metastatic disease for OutReg areas); colon (HR = 1.14 (1.01-1.29) for OutReg areas in metastatic disease); lung (HR range = 1.08-1.35 (1.01-1.48) for most non-metropolitan areas in all stages of disease excepting regionalised); ovarian (HR = 1.32 (1.06-1.65) for OutReg areas in metastatic disease, HR = 1.40 (1.04-1.90) for InnReg areas and HR = 1.68 (1.02-2.77) for OutReg areas in unknown stage of disease) and rectal (HR = 1.37 (1.05-1.78) for OutReg areas in localised and HR = 1.14 (1.002-1.30) for InnReg areas in regionalised disease). Where significant differences were found, major cities tended to show the best survival, whereas OutReg areas tended to show the worst. Although no definitive interpretation could be made regarding remote areas due to small patient numbers, their survival appeared relatively favourable. Reasons that contribute to the differences observed and the disparate results between cancer types need to be further explored in order to facilitate targeted solutions in reducing survival inequality between NSW regions.


Objective and subjective measures of sedentary behavior and physical activity.
To examine the convergent validity of the ActiGraph and activPAL accelerometers with the Bouchard Activity Record (BAR) in adults. Sedentary behavior and walking were evaluated in all instruments; standing and moderate-to-vigorous physical activity (MVPA) was evaluated only in those that detected such variables. Thirty-two participants wore the accelerometers and completed the BAR concurrently for 1 d. Descriptive statistics and delta values were reported for all instruments. Summary time spent in sedentary behavior and walking was compared between all instruments using repeated-measures ANOVA. Dependent t-tests were used to analyze summary time in 1) standing between activPAL and BAR and 2) MVPA between ActiGraph and BAR. Bland-Altman plots were interpreted for systematic bias. On a detailed level, concurrent time interval data were compared using mean percent agreement and κ statistics. There was a significant difference found in summary time spent in sedentary behavior apparent between ActiGraph and activPAL as well as between ActiGraph and BAR. There was also a significant difference detected in time spent in walking, apparent between ActiGraph and activPAL, and between ActiGraph and BAR. In the time interval analysis, mean percent agreement ranged from 54.0% (for walking detected by ActiGraph and activPAL) to 86.7% (for MVPA by ActiGraph and BAR). κ values ranged from 0.25 (for walking by ActiGraph and activPAL) to 0.70 (for sedentary behavior between activPAL and BAR). Differences were also found in standing and MVPA. The activPAL and BAR showed convergence on both summary and concurrent time interval levels in both sedentary behavior and walking. The comparative discordance between activPAL and BAR with ActiGraph was likely a function of different approaches used to distinguish sedentary behavior from walking.


C-terminal domain of insulin-like growth factor (IGF) binding protein-6: structure and interaction with IGF-II.
IGFs are important mediators of growth. IGF binding proteins (IGFBPs) 1-6 regulate IGF actions and have IGF-independent actions. The C-terminal domains of IGFBPs contribute to high-affinity IGF binding and modulation of IGF actions and confer some IGF-independent properties, but understanding how they achieve this has been constrained by the lack of a three-dimensional structure. We therefore determined the solution structure of the C-domain of IGFBP-6 using nuclear magnetic resonance (NMR). The domain consists of a thyroglobulin type 1 fold comprising an alpha-helix followed by a loop, a three-stranded antiparallel beta-sheet incorporating a second loop, and finally a disulfide-bonded flexible third loop. The IGF-II binding site on the C-domain was identified by examining NMR spectral changes upon complex formation. It consists of a largely hydrophobic surface patch involving the alpha-helix, the first beta-strand, and the first and second loops. The site was confirmed by mutagenesis of several residues, which resulted in decreased IGF binding affinity. The IGF-II binding site lies adjacent to surfaces likely to be involved in glycosaminoglycan binding of IGFBPs, which might explain their decreased IGF affinity when bound to glycosaminoglycans, and nuclear localization. Our structure provides a framework for understanding the roles of IGFBP C-domains in modulating IGF actions and conferring IGF-independent actions, as well as ultimately for the development of therapeutic IGF inhibitors for diseases including cancer.


Multi- and Single-Joint Resistance Exercises Promote Similar Plantar Flexor Activation in Resistance Trained Men.
The present study aimed to compare soleus, lateral, and medial gastrocnemius muscles activation during leg press and calf raise exercises in trained men. The study involved 22 trained men (27.1 ± 3.6 years, 82.7 ± 6.6 kg, 177.5 ± 5.2 cm, 3.6 ± 1.4 experience years) who performed one set of each exercise using a 10-repetition maximum (10RM) load in a counterbalanced randomized order and separated by 10 min of rest. The electromyographic signal was measured for the three major plantar flexors: soleus, medial, and lateral gastrocnemius. A comparison between exercises showed that the mean adjusted by peak values during the leg press were 49.20% for the gastrocnemius lateralis, 51.31% for the gastrocnemius medialis, and 50.76% for the soleus. Values for calf raise were 50.70%, 52.19%, and 51.34% for the lateral, medial gastrocnemius, and soleus, respectively. There were no significant differences between exercises for any muscle (lateral gastrocnemius (p = 0.230), medial gastrocnemius (p = 0.668), and soleus (p = 0.535)). The present findings suggest that both leg press and calf raises can be used with the purpose to recruit triceps surae muscles. This bring the suggestion that one can chose between exercises based on personal preferences and practical aspects, without any negative impact on muscle activation.


Matching Inhaler Devices with Patients: The Role of the Primary Care Physician.
Poor inhaler technique and nonadherence impair the efficacy of medications for asthma and chronic obstructive pulmonary disease (COPD). A range of factors, including age, dexterity, inspiratory capacity, cognitive ability, health literacy, and ethnicity, can impact a patient's ability and intention to use their device. Treatment success can also be influenced by patient preferences and perceptions. Therefore, it is important that healthcare professionals effectively match inhaler devices to individual patients' needs and abilities and empower patients by including them in treatment decisions. Physicians must, therefore, fully understand the characteristics of each device, as well as their patients' demographic characteristics and comorbidities. Following device selection, patient training and education, including a physical demonstration of the device, are key to eliminate any critical errors that may impact on health outcomes. Inhaler technique should be frequently rechecked. This review will examine the important role of primary care providers in the selection of appropriate inhaler devices and provision of training for patients with COPD and asthma to optimize correct inhaler use and adherence. An overview of the key features of available devices and of the factors to consider when selecting devices will be provided in the context of current asthma and COPD guidelines.


A role for chromatin remodeling in transcriptional termination by RNA polymerase II.
Chromatin remodeling can facilitate the recruitment of RNA polymerase II (Pol II) to targeted promoters, as well as enhancing the level of transcription. Here, we describe a further key role for chromatin remodeling in transcriptional termination. Using a genetic screen in S. pombe, we identified the CHD-Mi2 class chromatin remodeling ATPase, Hrp1, as a termination factor. In S. cerevisiae, we show that transcriptional termination and chromatin structure at the 3' ends of three genes all depend on the activity of the Hrp1 homolog, Chd1p, either alone or redundantly with the ISWI ATPases, Isw1p, and Isw2p. We suggest that chromatin remodeling of termination regions is a necessary prelude to efficient Pol II termination.


Remnant cholesterol, low-density lipoprotein cholesterol, and blood pressure as mediators from obesity to ischemic heart disease.
Obesity leads to increased ischemic heart disease (IHD) risk, but the risk is thought to be mediated through intermediate variables and may not be caused by increased weight per se. To test the hypothesis that the increased IHD risk because of obesity is mediated through lipoproteins, blood pressure, glucose, and C-reactive protein. Approximately 90 000 participants from Copenhagen were included in a Mendelian randomization design with mediation analyses. Associations were examined using conventional measurements of body mass index and intermediate variables and using genetic variants associated with these. During ≤22 years of follow-up 13 945 participants developed IHD. The increased IHD risk caused by obesity was partly mediated through elevated levels of nonfasting remnant cholesterol and low-density lipoprotein cholesterol, through elevated blood pressure, and possibly also through elevated nonfasting glucose levels; however, reduced high-density lipoprotein cholesterol and elevated C-reactive protein levels were not mediators in genetic analyses. The 3 intermediate variables that explained the highest excess risk of IHD from genetically determined obesity were low-density lipoprotein cholesterol with 8%, systolic blood pressure with 7%, and remnant cholesterol with 7% excess risk of IHD. Corresponding observational excess risks using conventional body mass index were 21%, 11%, and 20%, respectively. The increased IHD risk because of obesity was partly mediated through elevated levels of nonfasting remnant and low-density lipoprotein cholesterol and through elevated blood pressure. Our results suggest that there may be benefit to gain by reducing levels of these risk factors in obese individuals not able to achieve sustained weight loss.


Oropharyngeal group A streptococcal colonization disrupts latent Epstein-Barr virus infection.
Epstein-Barr virus (EBV) infects >90% of the human population within the first 2 decades of life and establishes reversible latent infection in B cells. The stimuli that lead to switching from latent to lytic EBV infection in vivo are still elusive. Group A streptococci (GAS) are a common cause of bacterial pharyngotonsillitis in children and adolescents and colonize the tonsils and pharynx of up to 20% of healthy children. Thus, concomitant presence of EBV and GAS in the same individual is frequent. Here, we show that EBV carriers who are colonized with GAS shed EBV particles in higher numbers in their saliva, compared with EBV carriers not colonized with GAS. Messenger RNA levels of the master lytic regulatory EBV gene BZLF1 were more frequently detected in tonsils from EBV carriers colonized with GAS than from EBV carriers not colonized. Heat-killed GAS, potentially mimicking GAS colonization, elicited lytic EBV in latently infected lymphoblastoid cell lines (LCLs) partially via Toll-like receptor 2 triggering, as did purified GAS peptidoglycan. Thus, colonization by GAS might benefit EBV by increasing the EBV load in saliva and thereby enhancing the likelihood of EBV spread to other hosts.


Low infiltration of tumor-associated macrophages in high c-Myb-expressing breast tumors.
Tumor-associated macrophages (TAMs) are prominent components of tumor stroma that promotes tumorigenesis. Many soluble factors participate in the deleterious cross-talk between TAMs and transformed cells; however mechanisms how tumors orchestrate their production remain relatively unexplored. c-Myb is a transcription factor recently described as a negative regulator of a specific immune signature involved in breast cancer (BC) metastasis. Here we studied whether c-Myb expression is associated with an increased presence of TAMs in human breast tumors. Tumors with high frequency of c-Myb-positive cells have lower density of CD68-positive macrophages. The negative association is reflected by inverse correlation between MYB and CD68/CD163 markers at the mRNA levels in evaluated cohorts of BC patients from public databases, which was found also within the molecular subtypes. In addition, we identified potential MYB-regulated TAMs recruiting factors that in combination with MYB and CD163 provided a valuable clinical multigene predictor for BC relapse. We propose that identified transcription program running in tumor cells with high MYB expression and preventing macrophage accumulation may open new venues towards TAMs targeting and BC therapy.


Implementation of an algorithm based on the Runge-Kutta-Fehlberg technique and the potential energy as a reaction coordinate to locate intrinsic reaction paths.
The intrinsic reaction coordinate (IRC) curve is used widely as a representation of the Reaction Path and can be parameterized taking the potential energy as a reaction coordinate (Aguilar-Mogas et al., J Chem Phys 2008, 128, 104102). Taking this parameterization and its variational nature, an algorithm is proposed that permits to locate this type of curve joining two points from an arbitrary curve that joints the same initial and final points. The initial and final points are minima of the potential energy surface associated with the geometry of reactants and products of the reaction whose mechanism is under study. The arbitrary curves are moved toward the IRC curve by a Runge-Kutta-Fehlberg technique. This technique integrates a set of differential equations resulting from the minimization until value zero of the line integral over the Weierstrass E-function. The Weierstrass E-function is related with the second variation in the theory of calculus of variations. The algorithm has been proved in real chemical systems.


Floral isolation is the major reproductive barrier between a pair of rewarding orchid sister species.
The crucial role of reproductive isolation in speciation has long been recognized; however, a limited number of studies quantify different isolation barriers and embed reproductive isolation in a phylogenetic context. In this study, we investigate reproductive isolation between the often sympatrically occurring orchid species, Gymnadenia conopsea and G. odoratissima. We examine the phylogenetic relationship between the two species and analyse floral isolation, fruit set and seed viability from interspecies crosses, as well as the ploidy level. Additionally, we quantify interspecies differences in floral signals and morphology. The results suggest that the two species have a sister-species relationship. In terms of reproductive isolation, we found complete floral isolation between the two species, but little to no post-pollination isolation; the species also mostly had the same ploidy level in the studied populations. We also show clear distinctions in floral signals, as well as in floral size and spur length. We propose that respective adaptation to short- vs. long-tongued pollinators was the driver of speciation in the here studied Gymnadenia species. Our study supports the key role of floral isolation in orchid speciation and shows that floral isolation is not restricted to highly specialized pollination systems, but can also occur between species with less specialized pollination.


New Model for Predicting Adsorption of Polar Molecules in Metal-Organic Frameworks with Unsaturated Metal Sites.
Conventional molecular models fail to correctly describe interactions of adsorbates with coordinatively unsaturated sites (CUS) present in a large number of metal-organic frameworks (MOFs). Here, we confirm the failure of these models for a prototypical polar adsorbate, carbon monoxide, and show that simply adjusting their parameters leads to poor agreement with experimental isotherms when outside the fitting conditions. We propose a new approach that combines quantum mechanical density functional theory (DFT) with Monte Carlo simulations to rigorously account for specific interactions at the CUS. By explicitly including electrostatic interactions and employing accurate DFT functionals that describe dispersion interactions, our modeling approach becomes generally applicable to both polar and nonpolar molecules. We demonstrate that this CUS model leads to substantial improvement in carbon monoxide adsorption isotherm predictions, and correctly captures the coordination binding mechanism. This paper represents a major stepping stone in the development of a robust, transferable and generally applicable approach to describe the complex interactions between gas molecules and CUS, with great potential for use in large-scale screening studies.


Local vascularized flaps for augmentation of Reinke's space.
The purpose of this study is to describe and test a novel surgical strategy for augmentation of Reinke's space using vascularized flaps: a thyroid ala perichondrium flap (TAP) and a composite thyroid ala perichondrium flap (CTAP) from the anterior larynx. We hypothesized that these specially designed vascularized flaps would remain viable once inset into the lamina propria, and that they would not disrupt rheologic, biomechanical, and histologic properties of the native vocal fold. Experimental. In vivo canine model. The length and volume of test flaps harvested in six adult human cadaveric larynges were analyzed to determine suitability for use in augmentation in the lamina propria. Also, 12 beagles randomly underwent unilateral placement of either TAP or CTAP, which were designed in accordance with the human adult cadaveric experiments. Flap perfusion was measured before and after harvest with laser Doppler. After 1 month, the beagles were humanely sacrificed and their larynges subjected to aerodynamic and acoustic evaluation using an excised larynx apparatus. The vocal fold lamina propria of four larynges--two TAP and two CTAP--underwent rheologic evaluation using a simple-shear rheometer. The remaining eight larynges underwent quantitative histologic and immunohistochemical evaluation. The survival and complication (swallowing, airway, local wound) rates of all dogs were noted. Initial studies with adult human cadaveric larynges established that TAP and CTAP possessed length and volume greater than native lamina propria. In the canine experiments, the perfusion change in the flaps was similar between flap groups. The damping ratio (ζ), dynamic viscosity (η'), elastic shear modulus (G'), and viscous shear modulus (G″) of treated and untreated native vocal folds were not statistically different. The glottic function measures of vocal efficiency, laryngeal resistance, jitter, shimmer, and harmonics-to-noise ratio (HNR) of treated and normal larynges were not statistically different. Similarly, the values for collagen, elastin, and glycosaminoglycans (GAGs) in treated and untreated vocal folds were not statistically different. Also, neither neochrondrogenesis nor neoosteogenesis was detected in any treated vocal fold. The values for vascular and cellular proliferation in treated and untreated vocal folds were not statistically different. All test dogs survived and had no complications related to swallowing, airway distress, or the local wound. The test flaps described and tested in this study appear to have conceptually attractive features for augmentation of Reinke's space. When placed in an in vivo setting TAP and CTAP did not reveal unfavorable vascular, rheologic, aerodynamic, acoustic, or histologic characteristics. There was no unanticipated morbidity or mortality to the test animals. Long-term viability of these flaps is unknown. TAP and CTAP may open novel pathways for correction of glottic defects and may offer crossover opportunities with tissue engineering techniques.


Engineering induction of singular neural rosette emergence within hPSC-derived tissues.
Human pluripotent stem cell (hPSC)-derived neural organoids display unprecedented emergent properties. Yet in contrast to the singular neuroepithelial tube from which the entire central nervous system (CNS) develops in vivo, current organoid protocols yield tissues with multiple neuroepithelial units, a.k.a. neural rosettes, each acting as independent morphogenesis centers and thereby confounding coordinated, reproducible tissue development. Here, we discover that controlling initial tissue morphology can effectively (>80%) induce single neural rosette emergence within hPSC-derived forebrain and spinal tissues. Notably, the optimal tissue morphology for observing singular rosette emergence was distinct for forebrain versus spinal tissues due to previously unknown differences in ROCK-mediated cell contractility. Following release of geometric confinement, the tissues displayed radial outgrowth with maintenance of a singular neuroepithelium and peripheral neuronal differentiation. Thus, we have identified neural tissue morphology as a critical biophysical parameter for controlling in vitro neural tissue morphogenesis furthering advancement towards biomanufacture of CNS tissues with biomimetic anatomy and physiology.


Temporal Evolution of Generalization during Learning in Linear Networks.
We study generalization in a simple framework of feedforward linear networks with n inputs and n outputs, trained from examples by gradient descent on the usual quadratic error function. We derive analytical results on the behavior of the validation function corresponding to the LMS error function calculated on a set of validation patterns. We show that the behavior of the validation function depends critically on the initial conditions and on the characteristics of the noise. Under certain simple assumptions, if the initial weights are sufficiently small, the validation function has a unique minimum corresponding to an optimal stopping time for training for which simple bounds can be calculated. There exists also situations where the validation function can have more complicated and somewhat unexpected behavior such as multiple local minima (at most n) of variable depth and long but finite plateau effects. Additional results and possible extensions are briefly discussed.


Seasonal infection rates of Batrachochytrium dendrobatidis in populations of northern green frog Lithobates clamitans melanota tadpoles.
Few studies have documented seasonal variation of Batrachochytrium dendrobatidis (Bd) infection rates in larval amphibians. We identified 4 natural populations of northern green frogs Lithobates clamitans melanota in Pennsylvania (USA) that contained Bd-infected tadpoles during post-wintering collections in May and June, after hibernating tadpoles had overwintered in wetlands. However, we failed to detect infected tadpoles at those wetlands when pre-wintering collections were made in late July through early September. We observed 2 cohorts of tadpoles that appeared to lack Bd-infected individuals in pre-wintering collections, yet contained Bd-infected individuals the following spring. We also observed 4 cohorts of pre-wintering tadpoles that were Bd-free, even though post-wintering tadpoles collected earlier in the year were infected with Bd. Our results suggest that tadpoles either reduce Bd infections during the summer months, and/or infections proliferate sometime prior to (or shortly after) tadpoles emerge from hibernation. It is unlikely that pre-wintering tadpoles were too small to detect Bd zoospores because (1) there was no correlation between Bd zoospore levels and tadpole size or stage, and (2) size was not a significant predictor of infection status. These results suggest that, while sampling larvae can be an effective means of collecting large sample sizes, investigators in our Mid-Atlantic region should conduct sampling by early summer to maximize the chances of detecting Bd. Further research is warranted to determine whether wetland topography and warm, shallow microhabitats within wetlands contribute to a population's ability to drastically reduce Bd prevalence prior to overwintering at ponds.


Phosphoaspirin (MDC-43), a novel benzyl ester of aspirin, inhibits the growth of human cancer cell lines more potently than aspirin: a redox-dependent effect.
Aspirin is chemopreventive against colon and probably other cancers, but this effect is relatively weak and its chronic administration to humans is associated with significant side effects. Because of these limitations, extensive effort has been exerted to improve the pharmacological properties of aspirin. We have determined the anticancer activity and mechanisms of action of the novel para positional isomer of phosphoaspirin [P-ASA; MDC-43; 4-((diethoxyphosphoryloxy)methyl)phenyl 2-acetoxybenzoate]. P-ASA inhibited the growth of 10 human cancer cell lines originating from colon, lung, liver, pancreas and breast, at least 18- to 144-fold more potently than conventional aspirin. P-ASA achieved this effect by modulating cell kinetics; compared with controls, P-ASA reduced cell proliferation by up to 68%, increased apoptosis 5.5-fold and blocked cell cycle progression in the G(2)/M phase. P-ASA increased intracellular levels of reactive oxygen species (ROS), depleted glutathione levels and modulated cell signaling predominantly through the mitogen-activated protein kinase (p38 and c-jun N-terminal kinase), cyclooxygenase (COX) and nuclear factor-kappa B pathways. P-ASA targeted the mitochondria, increasing mitochondrial superoxide anion levels; this effect on ROS led to collapsed mitochondrial membrane potential and triggered the intrinsic apoptotic pathway. The antioxidant N-acetyl cysteine abrogated the cell growth inhibitory and signaling effects of P-ASA, underscoring the centrality of ROS in its mechanism of action. Our results, establishing P-ASA as a potent inhibitor of the growth of several human cancer cell lines, suggest that it may possess broad anticancer properties. We conclude that the novel P-ASA is a promising anticancer agent, which merits further evaluation.


Comparative effects of dexmedetomidine, propofol, sevoflurane, and S-ketamine on regional cerebral glucose metabolism in humans: a positron emission tomography study.
The highly selective α2-agonist dexmedetomidine has become a popular sedative for neurointensive care patients. However, earlier studies have raised concern that dexmedetomidine might reduce cerebral blood flow without a concomitant decrease in metabolism. Here, we compared the effects of dexmedetomidine on the regional cerebral metabolic rate of glucose (CMRglu) with three commonly used anaesthetic drugs at equi-sedative doses. One hundred and sixty healthy male subjects were randomised to EC50 for verbal command of dexmedetomidine (1.5 ng ml-1; n=40), propofol (1.7 μg ml-1; n=40), sevoflurane (0.9% end-tidal; n=40) or S-ketamine (0.75 μg ml-1; n=20) or placebo (n=20). Anaesthetics were administered using target-controlled infusion or vapouriser with end-tidal monitoring. 18F-labelled fluorodeoxyglucose was administered 20 min after commencement of anaesthetic administration, and high-resolution positron emission tomography with arterial blood activity samples was used to quantify absolute CMRglu for whole brain and 15 brain regions. At the time of [F18]fluorodeoxyglucose injection, 55% of dexmedetomidine, 45% of propofol, 85% of sevoflurane, 45% of S-ketamine, and 0% of placebo subjects were unresponsive. Whole brain CMRglu was 63%, 71%, 71%, and 96% of placebo in the dexmedetomidine, propofol, sevoflurane, and S-ketamine groups, respectively (P<0.001 between the groups). The lowest CMRglu was observed in nearly all brain regions with dexmedetomidine (P<0.05 compared with all other groups). With S-ketamine, CMRglu did not differ from placebo. At equi-sedative doses in humans, potency in reducing CMRglu was dexmedetomidine>propofol>ketamine=placebo. These findings alleviate concerns for dexmedetomidine-induced vasoconstriction and cerebral ischaemia. NCT02624401.


Genomic and biological characterization of exon 4 KRAS mutations in human cancer.
Mutations in RAS proteins occur widely in human cancer. Prompted by the confirmation of KRAS mutation as a predictive biomarker of response to epidermal growth factor receptor (EGFR)-targeted therapies, limited clinical testing for RAS pathway mutations has recently been adopted. We performed a multiplatform genomic analysis to characterize, in a nonbiased manner, the biological, biochemical, and prognostic significance of Ras pathway alterations in colorectal tumors and other solid tumor malignancies. Mutations in exon 4 of KRAS were found to occur commonly and to predict for a more favorable clinical outcome in patients with colorectal cancer. Exon 4 KRAS mutations, all of which were identified at amino acid residues K117 and A146, were associated with lower levels of GTP-bound RAS in isogenic models. These same mutations were also often accompanied by conversion to homozygosity and increased gene copy number, in human tumors and tumor cell lines. Models harboring exon 4 KRAS mutations exhibited mitogen-activated protein/extracellular signal-regulated kinase kinase dependence and resistance to EGFR-targeted agents. Our findings suggest that RAS mutation is not a binary variable in tumors, and that the diversity in mutant alleles and variability in gene copy number may also contribute to the heterogeneity of clinical outcomes observed in cancer patients. These results also provide a rationale for broader KRAS testing beyond the most common hotspot alleles in exons 2 and 3.


Development of Nationwide Recommendations to Support Prenatal Counseling in Extreme Prematurity.
To develop a nationwide, evidence-based framework to support prenatal counseling in extreme prematurity, focusing on organization, decision-making, content, and style aspects. A nationwide multicenter RAND-modified Delphi method study was performed between November 2016 and December 2017 in the Netherlands. Firstly, recommendations were extracted from literature and previous studies. Secondly, an expert panel (n = 21) with experienced parents, obstetricians, and neonatologists rated the recommendations on importance for inclusion in the framework. Thirdly, ratings were discussed in a consensus meeting. The final set of recommendations was approved and transformed into a framework. A total of 101 recommendations on organization, decision-making, content, and style were included in the framework, including tools to support personalization. The most important recommendations regarding organization were to have both parents involved in the counseling with both the neonatologist and obstetrician. The shared decision-making model was recommended for deciding between active support and comfort care. Main recommendations regarding content of conversation were explanation of treatment options, information on survival, risk of permanent consequences, impossibility to predict an individual course, possibility for multiple future decision moments, and a discussion on parental values and standards. It was considered important to avoid jargon, check understanding, and provide a summary. The expert panel, patient organization, and national professional associations (gynecology and pediatrics) approved the framework. A nationwide, evidence-based framework for prenatal counseling in extreme prematurity was developed. It contains recommendations and tools for personalization in the domains of organization, decision-making, content, and style of prenatal counseling.


Degarelix for Treating Advanced Hormone-Dependent Prostate Cancer: An Evidence Review Group Perspective of a NICE Single Technology Appraisal.
As part of its Single Technology Appraisal Process, the National Institute for Health and Care Excellence (NICE) invited the manufacturer of degarelix (Ferring Pharmaceuticals) to submit evidence for the clinical and cost effectiveness of degarelix for the treatment of advanced hormone-dependent prostate cancer. The School of Health and Related Research Technology Appraisal Group at the University of Sheffield was commissioned to act as the independent Evidence Review Group (ERG). The ERG produced a critical review of the evidence contained within the company's submission to NICE. The evidence, which included a randomised controlled trial (RCT) of degarelix versus leuprorelin, found that degarelix was non-inferior to leuprorelin for reduction of testosterone levels and that degarelix achieved a more rapid suppression of prostate-specific antigen levels and subsequently decreased incidences of testosterone flare associated with luteinising hormone releasing-hormone (LHRH) agonists. However, protection against testosterone flare for the comparators in the clinical trials was not employed in line with UK clinical practice. Further claims surrounding overall survival, cardiovascular adverse events and clinical equivalence of the comparator drugs from six RCTs of degarelix should be regarded with caution because of flaws and inconsistencies in the pooling of trial data to draw conclusions. The cost-effectiveness evidence included a de novo economic model. Based on the ERG's preferred base case, the deterministic incremental cost-effectiveness analysis (ICER) for degarelix versus 3-monthly triptorelin was £14,798 per quality-adjusted life-year (QALY) gained. Additional scenario analyses undertaken by the ERG resulted in ICERs for degarelix versus 3-monthly triptorelin ranging from £17,067 to £35,589 per QALY gained. Subgroup analyses undertaken using the Appraisal Committee's preferred assumptions suggested that degarelix was not cost effective for the subgroup with metastatic disease but could be cost effective for the subgroup with spinal metastases. The company submitted further evidence to NICE following an initial negative Appraisal Committee decision. Further analyses from the Decision Support Unit found that that, whilst some evidence indicated that degarelix could be cost effective for a small subgroup of people with spinal cord compression (SCC), data on the potential size of this subgroup and the rate of SCC were insufficient to estimate an ICER based on the evidence submitted by the company and a separately commissioned systematic review. NICE recommended degarelix as an option for treating advanced hormone-dependent prostate cancer in people with spinal metastases, only if the commissioner can achieve at least the same discounted drug cost as that available to the UK NHS in June 2016.


High cholesterol diet activates ER stress mediated apoptosis in testes tissue: Role of α-tocopherol.
The seminiferous tubules where spermatogenesis occurs are enveloped and protected by the Sertoli cells to support germ cells undergoing meiosis to produce haploid gametes. Clearly, induction of apoptosis in seminiferous tubules leads to abnormalities in spermatogenesis and male infertility. Studies demonstrated that increased hyperlipidemia impairs male infertility and spermatogenesis by enhancing seminiferous tubules apoptosis. However, molecular mechanisms underlying high-cholesterol-mediated testicular damage remain poorly elucidated. In this scope, we established a rabbit model and investigated the role of endoplasmic reticulum (ER) stress on high cholesterol diet induced seminiferous tubule apoptosis. Histopatological examinations revealed increased seminifer tubule apoptosis in testes of rabbits fed high cholesterol diet. In addition, phosphorylated forms of IRE1 and PERK, two well-identified markers of ER stress, were significantly induced in accordance with high cholesterol diet. High cholesterol diet also exhibited CHOP induction in testes, indicating increased ER stress related apoptosis. Supplementation of α-tocopherol significantly attenuated cholesterol mediated ER stress, and restored seminiferous tubules apoptosis. Taken together, our findings suggest that α-tocopherol might be capable to reduce testicular damage via ameliorating histopatological features and inhibiting seminiferous tubules apoptosis in hypercholesterolemic rabbits.


Effect of Reduced Phosphoric Acid Pre-etching Times on Enamel Surface Characteristics and Shear Fatigue Strength Using Universal Adhesives.
To examine the effect of reduced phosphoric acid pre-etching times on enamel fatigue bond strength of universal adhesives and surface characteristics by using atomic force microscopy (AFM). Three universal adhesives were used in this study (Clearfil Universal Bond [C], G-Premio Bond [GP], Scotchbond Universal Adhesive [SU]). Four pre-etching groups were employed: enamel pre-etched with phosphoric acid and immediately rinsed with an air-water spray, and enamel pre-etched with phosphoric acid for 5, 10, or 15 s. Ground enamel was used as the control group. For the initial bond strength test, 15 specimens per etching group for each adhesive were used. For the shear fatigue test, 20 specimens per etching group for each adhesive were loaded using a sine wave at a frequency of 20 Hz for 50,000 cycles or until failure occurred. Initial shear bond strengths and fatigue shear strengths of composite adhesively bonded to ground and pre-etched enamel were determined. AFM observations of ground and pre-etched enamel were also conducted, and surface roughness as well as surface area were evaluated. The initial shear bond strengths and fatigue shear strengths of the universal adhesives in the pre-etched groups were significantly higher than those of the control group, and were not influenced by the pre-etching time. Significantly higher surface roughness and surface area of enamel surfaces in pre-etched groups were observed compared with those in the control group. While the surface area was not significantly influenced by etching time, surface roughness of the enamel surfaces in the pre-etched groups significantly increased with pre-etching time. The results of this in vitro study suggest that reduced phosphoric acid pre-etching times do not impair the fatigue bond strength of universal adhesives. Although fatigue bond strength and surface area were not influenced by phosphoric-acid etching times, surface roughness increased with increasing etching time.


Aberrant expression of interleukin-1β and inflammasome activation in human malignant gliomas.
Glioblastoma is the most frequent and malignant form of primary brain tumor with grave prognosis. Mounting evidence supports that chronic inflammation (such as chronic overactivation of IL-1 system) is a crucial event in carcinogenesis and tumor progression. IL-1 also is an important cytokine with species-dependent regulations and roles in CNS cell activation. While much attention is paid to specific anti-tumor immunity, little is known about the role of chronic inflammation/innate immunity in glioma pathogenesis. In this study, we examined whether human astrocytic cells (including malignant gliomas) can produce IL-1 and its role in glioma progression. We used a combination of cell culture, real-time PCR, ELISA, western blot, immunocytochemistry, siRNA and plasmid transfection, micro-RNA analysis, angiogenesis (tube formation) assay, and neurotoxicity assay. Glioblastoma cells produced large quantities of IL-1 when activated, resembling macrophages/microglia. The activation signal was provided by IL-1 but not the pathogenic components LPS or poly IC. Glioblastoma cells were highly sensitive to IL-1 stimulation, suggesting its relevance in vivo. In human astrocytes, IL-1β mRNA was not translated to protein. Plasmid transfection also failed to produce IL-1 protein, suggesting active repression. Suppression of microRNAs that can target IL-1α/β did not induce IL-1 protein. Glioblastoma IL-1β processing occurred by the NLRP3 inflammasome, and ATP and nigericin increased IL-1β processing by upregulating NLRP3 expression, similar to macrophages. RNAi of annexin A2, a protein strongly implicated in glioma progression, prevented IL-1 induction, demonstrating its new role in innate immune activation. IL-1 also activated Stat3, a transcription factor crucial in glioma progression. IL-1 activated glioblastoma-conditioned media enhanced angiogenesis and neurotoxicity. Our results demonstrate unique, species-dependent immune activation mechanisms involving human astrocytes and astrogliomas. Specifically, the ability to produce IL-1 by glioblastoma cells may confer them a mesenchymal phenotype including increased migratory capacity, unique gene signature and proinflammatory signaling.


Association of Sugar-Sweetened Beverage Intake during Infancy with Dental Caries in 6-year-olds.
To examine whether sugar-sweetened beverage (SSB) intake during infancy is associated with dental caries by age 6, a longitudinal analysis of 1,274 U.S. children was conducted using data from the 2005-2007 Infant Feeding Practices Study II and the 2012 Follow-up Study at 6 years of age. The exposure variables were maternal-reported SSB intakes during infancy (i.e., any SSB intake during infancy, age at SSB introduction during infancy, and average frequency of SSB intake during 10-12 months of age). The outcome variable was maternal-reported dental caries of their 6-year-old in his/her lifetime. Multivariable logistic regression analyses were used to calculate adjusted odds ratios (aOR) for associations of SSB intake during infancy with having dental caries among 6-year-olds after controlling for baseline characteristics of children and mothers and child's tooth brushing habits and sweet food intake at follow-up. Based on maternal recall, almost 40% of 6-year-olds had dental caries in their lifetime. Adjusted odds of having dental caries was significantly associated with higher frequency of SSB intake during 10-12 months (aOR=1.83 for ≥3 times/week, vs. none). Any SSB intake during infancy and age at SSB introduction during infancy were not associated with dental caries. In conclusion, frequent SSB intake during 10-12 months of age significantly increased the likelihood of having dental caries among 6-year-olds. Late infancy may be an important time for mothers to establish healthy beverage practices for their children. These findings can be used to inform efforts to reduce dental caries among children.


The Efficacy, Safety and Applications of Medical Hypnosis.
The efficacy and safety of hypnotic techniques in somatic medicine, known as medical hypnosis, have not been supported to date by adequate scientific evidence. We systematically reviewed meta-analyses of randomized controlled trials (RCTs) of medical hypnosis. Relevant publications (January 2005 to June 2015) were sought in the Cochrane databases CDSR and DARE, and in PubMed. Meta-analyses involving at least 400 patients were included in the present analysis. Their methodological quality was assessed with AMSTAR (A Measurement Tool to Assess Systematic Reviews). An additional search was carried out in the CENTRAL and PubMed databases for RCTs of waking suggestion (therapeutic suggestion without formal trance induction) in somatic medicine. Out of the 391 publications retrieved, five were reports of metaanalyses that met our inclusion criteria. One of these meta-analyses was of high methodological quality; three were of moderate quality, and one was of poor quality. Hypnosis was superior to controls with respect to the reduction of pain and emotional stress during medical interventions (34 RCTs, 2597 patients) as well as the reduction of irritable bowel symptoms (8 RCTs, 464 patients). Two meta-analyses revealed no differences between hypnosis and control treatment with respect to the side effects and safety of treatment. The effect size of hypnosis on emotional stress during medical interventions was low in one meta-analysis, moderate in one, and high in one. The effect size on pain during medical interventions was low. Five RCTs indicated that waking suggestion is effective in medical procedures. Medical hypnosis is a safe and effective complementary technique for use in medical procedures and in the treatment of irritable bowel syndrome. Waking suggestions can be a component of effective doctor-patient communication in routine clinical situations.


The Cys allele of the DRD2 Ser311Cys polymorphism has a dominant effect on risk for schizophrenia: evidence from fixed- and random-effects meta-analyses.
Previously we derived independent estimates of the effect of the dopamine D2 receptor (DRD2) Ser311Cys polymorphism on risk for schizophrenia using fixed- and random-effects meta-analyses. Both analyses identified a significant association between the Cys allele and schizophrenia, but neither included all available data. Furthermore, genotype data were not evaluated in either analysis, thus precluding any determination of the mode of inheritance. The present study was conducted to resolve discrepancies between the existing meta-analyses, and provide more comprehensive and accurate estimates of the nature and magnitude of the influence of the Ser311Cys polymorphism on risk for schizophrenia. All discrepancies between the two sets of previously meta-analyzed studies were identified and resolved to the mutual satisfaction of the authors, and the final dataset was analyzed independently by fixed- and random-effects meta-analyses. A total of 27 samples, comprising 3,707 schizophrenia patients and 5,363 control subjects, were included in the analyses of allelic association, while smaller numbers of studies and subjects were included in each of the genotypic association analyses. A significant effect of the Cys allele was observed under both fixed-effects (odds ratio [OR] = 1.4; P = 0.002) and random-effects (OR = 1.4; P = 0.007) models. Cys/Ser heterozygotes were at elevated risk for schizophrenia when compared to Ser/Ser homozygotes (fixed- and random-effects OR = 1.4, p(s) <or= 0.005), but Cys/Cys homozygotes were at no elevated risk relative to heterozygotes (OR = 1.0, p(s) >or= 0.948). There was no evidence of heterogeneity, excessive influence of any single study, or publication bias in any of the analyses, suggesting that the effect of this DRD2 polymorphism on schizophrenia risk is reliable and uniform across populations, and our estimates of its magnitude are robust and accurate.


Osmotic regulation of vasopressin and renin in spontaneously hypertensive rats.
Abnormalities in the vasopressin and renin systems have been reported in spontaneously hypertensive rats (SHR). Therefore, studies were performed to evaluate the responsiveness of these systems to changes in plasma osmolality and sodium concentration. These variables were manipulated in vivo by intraperitoneal administration of distilled water, isotonic saline, or hypertonic saline to 8- and 18-week-old SHR and normotensive Wistar-Kyoto rats (WKY). Animals were decapitated 30 minutes later, and trunk blood was collected. The hypertonic saline injections resulted in an increase in plasma osmolality and serum sodium at both ages (p less than 0.001). Serum vasopressin was higher in all groups of animals receiving hypertonic saline (1200 mosm/kg H2O; p less than 0.05), but the magnitude of increase was not significantly different in the SHR and WKY at either age. Serum renin activity was lower in SHR than in WKY following acute decreases in serum sodium at 8 weeks, but it was the same for both strains at 18 weeks. Both kidney renin content and concentration were lower in SHR than in WKY at 18 weeks but not at 8 weeks. Therefore, the suppressed renin response to acute osmotic challenge in 8-week-old SHR is not the consequence of reduced kidney renin content. The vasopressin response to osmotic stimulation also was evaluated in vitro using hypothalamoneurohypophyseal explants obtained from 5-, 8-, and 18-week-old SHR and WKY.(ABSTRACT TRUNCATED AT 250 WORDS)


Stress reactivity and attachment security.
Seventy-three 18-month-olds were tested in the Ainsworth Strange Situation. These children were a subset of 83 infants tested at 2, 4, 6, and 15 months during their well-baby examinations with inoculations. Salivary cortisol, behavioral distress, and maternal responsiveness measures obtained during these clinic visits were examined in relation to attachment classifications. In addition, parental report measures of the children's social fearfulness in the 2nd year of life were used to classify the children into high-fearful versus average- to low-fearful groups. In the 2nd year, the combination of high fearfulness and insecure versus secure attachment was associated with higher cortisol responses to both the clinic exam-inoculation situation and the Strange Situation. Thus, attachment security moderates the physiological consequences of fearful, inhibited temperament. Regarding the 2-, 4-, and 6-month data, later attachment security was related to greater maternal responsiveness and lower cortisol baselines. Neither cortisol nor behavioral reactivity to the inoculations predicted later attachment classifications. There was some suggestion, however, that at their 2-month checkup, infants who would later be classified as insecurely attached exhibited larger dissociations between the magnitude of their behavioral and hormonal response to the inoculations. Greater differences between internal (hormonal) and external (crying) responses were also negatively correlated with maternal responsiveness and positively correlated with pretest cortisol levels during these early months of life.


Recent advances in management of transient ischaemic attacks and minor ischaemic strokes.
The risk of recurrent stroke during the first few days after a transient ischaemic attack or minor stroke is much higher than previously estimated. However, there is substantial variation worldwide in how patients with suspected transient ischaemic attack or minor stroke are investigated and treated in the acute phase: some health-care systems provide immediate emergency inpatient care and others provide non-emergency outpatient clinical assessment. This review considers what is known about the early prognosis after transient ischaemic attack and minor ischaemic stroke, what factors identify individuals at particularly high early risk of stroke, and what evidence there is that urgent preventive treatment is likely to be effective in reducing the early risk of stroke.


Vitalism in contemporary chiropractic: a help or a hinderance?
Chiropractic emerged in 1895 and was promoted as a viable health care substitute in direct competition with the medical profession. This was an era when there was a belief that one cause and one cure for all disease would be discovered. The chiropractic version was a theory that most diseases were caused by subluxated (slightly displaced) vertebrae interfering with "nerve vibrations" (a supernatural, vital force) and could be cured by adjusting (repositioning) vertebrae, thereby removing the interference with the body's inherent capacity to heal. DD Palmer, the originator of chiropractic, established chiropractic based on vitalistic principles. Anecdotally, the authors have observed that many chiropractors who overtly claim to be "vitalists" cannot define the term. Therefore, we sought the origins of vitalism and to examine its effects on chiropractic today. Vitalism arose out of human curiosity around the biggest questions: Where do we come from? What is life? For some, life was derived from an unknown and unknowable vital force. For others, a vital force was a placeholder, a piece of knowledge not yet grasped but attainable. Developments in science have demonstrated there is no longer a need to invoke vitalistic entities as either explanations or hypotheses for biological phenomena. Nevertheless, vitalism remains within chiropractic. In this examination of vitalism within chiropractic we explore the history of vitalism, vitalism within chiropractic and whether a vitalistic ideology is compatible with the legal and ethical requirements for registered health care professionals such as chiropractors. Vitalism has had many meanings throughout the centuries of recorded history. Though only vaguely defined by chiropractors, vitalism, as a representation of supernatural force and therefore an untestable hypothesis, sits at the heart of the divisions within chiropractic and acts as an impediment to chiropractic legitimacy, cultural authority and integration into mainstream health care.


Fruit Wines Inhibitory Activity Against α-Glucosidase.
Fruit wines are well known for their profound health-promoting properties including both enzyme activations and inhibitions. They may act preventive in regard to diabetes melitus and other chronic diseases. Potential α-glucosidase inhibitory activity of fruit wines made from blueberry, black chokeberry, blackberry, raspberry and sour cherry was the subject of this study. In order to increase the alcohol content due to enriched extraction of total phenolics, sugar was added in the fruit pomace of the half of the examined fruit wine samples. Compared with acarbose used as a positive control (IC50 = 73.78 µg/mL), all fruit wine samples exhibited higher α-glucosidase inhibitory activity. Indeed, blueberry wine samples stood out, both prepared with IC50 = 24.14 µg/mL, lyophilised extract yield 3.23% and without IC50 = 46.39 µg/mL, lyophilised extract yield 2.89% and with addition of sugar before fermentation. Chlorogenic acid predominantly contributed to α-glucosidase inhibitory activity of the blueberry, black chokeberry and sour cherry wine samples. However, ellagic acid, a potent α-glucosidase inhibitor possessing a planar structure, only slightly affected the activity of the blueberry wine samples, due to the lower concentration. In addition to this, molecular docking study of chlorogenic acid pointed out the importance of binding energy (-8.5 kcal/mol) for the inhibition of the enzyme. In summary, fruit wines made from blueberry should be primarily taken into consideration as a medicinal food targeting diabetes mellitus type 2 in the early stage, if additional studies would confirm their therapeutic potential for the control of postprandial hyperglycemia.


USP10 regulates p53 localization and stability by deubiquitinating p53.
Stability and localization of p53 is essential for its tumor suppressor function. Ubiquitination by the E3 ubiquitin ligase Mdm2 is the major regulatory mechanism of p53, which induces p53 nuclear export and degradation. However, it is unclear whether ubiquitinated cytoplasmic p53 can be recycled. Here, we report that USP10, a cytoplasmic ubiquitin-specific protease, deubiquitinates p53, reversing Mdm2-induced p53 nuclear export and degradation. After DNA damage, USP10 is stabilized, and a fraction of USP10 translocates to the nucleus to activate p53. The translocation and stabilization of USP10 is regulated by ATM -mediated phosphorylation of USP10 at Thr42 and Ser337. Finally, USP10 suppresses tumor cell growth in cells with wild-type p53, with USP10 expression downregulated in a high percentage of clear cell carcinomas, known to have few p53 mutations. These findings reveal USP10 to be a novel regulator of p53, providing an alternative mechanism of p53 inhibition in cancers with wild-type p53.


NMR structure and dynamics of Q4D059, a kinetoplastid-specific and conserved protein from Trypanosoma cruzi.
Q4D059 (UniProt accession number), is an 86-residue protein from Trypanosoma cruzi, conserved in the related kinetoplastid parasites Trypanosoma brucei and Leishmania major. These pathogens are the causal agents of the neglected diseases: Chagas, sleeping sickness and leishmaniases respectively and had recently their genomes sequenced. Q4D059 shows low sequence similarity with mammal proteins and because of its essentiality demonstrated in T. brucei, it is a potential target for anti-parasitic drugs. The 11 hypothetical proteins homologous to Q4D059 are all uncharacterized proteins of unknown function. Here, the solution structure of Q4D059 was solved by NMR and its backbone dynamics was characterized by (15)N relaxation parameters. The structure is composed by a parallel/anti-parallel three-stranded β-sheet packed against four helical regions. The structure is well defined by ca. 9 NOEs per residue and a backbone rmsd of 0.50±0.05 Å for the representative ensemble of 20 lowest-energy structures. The structure is overall rigid except for N-terminal residues A(9) to D(11) at the beginning of β1, K(38), V(39) at the end of helix H3 with rapid motion in the ps-ns timescale and G(25) (helix H2), I(68) (β2) and V(78) (loop 3) undergoing internal motion in the μs-ms timescale. Limited structural similarities were found in protein structures deposited in the PDB, therefore functional inferences based on protein structure information are not clear. Q4D059 adopts a α/β fold that is slightly similar to the ATPase sub-domain IIB of the heat-shock protein 70 (HSP70) and to the N-terminal domain of the ribosomal protein L11.