α-Conotoxin Decontamination Protocol Evaluation: What Works and What Doesn’t

Nine publically available biosafety protocols for safely handling conotoxin peptides were tested to evaluate their decontamination efficacy. Circular dichroism (CD) spectroscopy and mass spectrometry (MS) were used to assess the effect of each chemical treatment on the secondary and primary structure of α-CTx MII (L10V, E11A). Of the nine decontamination methods tested, treatment with 1% (m/v) solution of the enzymatic detergent Contrex™ EZ resulted in a 76.8% decrease in α-helical content as assessed by the mean residue ellipticity at 222 nm, and partial peptide digestion was demonstrated using high performance liquid chromatography mass spectrometry (HPLC-MS). Additionally, treatment with 6% sodium hypochlorite (m/v) resulted in 80.5% decrease in α-helical content and complete digestion of the peptide. The Contrex™ EZ treatment was repeated with three additional α-conotoxins (α-CTxs), α-CTxs LvIA, ImI and PeIA, which verified the decontamination method was reasonably robust. These results support the use of either 1% Contrex™ EZ solution or 6% sodium hypochlorite in biosafety protocols for the decontamination of α-CTxs in research laboratories

CyberKnife Boost for Patients with Cervical Cancer Unable to Undergo Brachytherapy

Standard radiation therapy for patients undergoing primary chemosensitized radiation for carcinomas of the cervix usually consists of external beam radiation followed by an intracavitary brachytherapy boost. On occasion, the brachytherapy boost cannot be performed due to unfavorable anatomy or because of coexisting medical conditions. We examined the safety and efficacy of using CyberKnife stereotactic body radiotherapy (SBRT) as a boost to the cervix after external beam radiation in those patients unable to have brachytherapy to give a more effective dose to the cervix than with conventional external beam radiation alone. Six consecutive patients with anatomic or medical conditions precluding a tandem and ovoid boost were treated with combined external beam radiation and CyberKnife boost to the cervix. Five patients received 45 Gy to the pelvis with serial intensity-modulated radiation therapy boost to the uterus and cervix to a dose of 61.2 Gy. These five patients received an SBRT boost to the cervix to a dose of 20 Gy in five fractions of 4 Gy each. One patient was treated to the pelvis to a dose of 45 Gy with an external beam boost to the uterus and cervix to a dose of 50.4 Gy. This patient received an SBRT boost to the cervix to a dose of 19.5 Gy in three fractions of 6.5 Gy. Five percent volumes of the bladder and rectum were kept to ≤75 Gy in all patients (i.e., V75 Gy ≤ 5%). All of the patients remain locally controlled with no evidence of disease following treatment. Grade 1 diarrhea occurred in 4/6 patients during the conventional external beam radiation. There has been no grade 3 or 4 rectal or bladder toxicity. There were no toxicities observed following SBRT boost. At a median follow-up of 14 months, CyberKnife radiosurgical boost is well tolerated and efficacious in providing a boost to patients with cervix cancer who are unable to undergo brachytherapy boost. Further follow-up is required to see if these results remain durable

N-Functionalised TsDPEN catalysts for asymmetric transfer hydrogenation; synthesis and applications

A series of Ru(II)/arene complexes containing N-alkylated derivatives of TsDPEN were prepared and tested in the asymmetric transfer hydrogenation (ATH) of ketones. The results demonstrated that a wide variety of functionality were tolerated on the basic amine of the TsDPEN ligand, without significantly disrupting the ability of the catalyst to catalyse hydrogen transfer reactions

Expression and Purification of a Cleavable Recombinant Fortilin from \u3ci\u3eEscherichia coli\u3c/i\u3e for Structure Activity Studies

Complications related to atherosclerosis account for approximately 1 in 4 deaths in the United States and treatment has focused on lowering serum LDL-cholesterol levels with statins. However, approximately 50% of those diagnosed with atherosclerosis have blood cholesterol levels within normal parameters. Human fortilin is an anti-apoptotic protein and a factor in macrophage-mediated atherosclerosis and is hypothesized to protect inflammatory macrophages from apoptosis, leading to subsequent cardiac pathogenesis. Fortilin is unique because it provides a novel drug target for atherosclerosis that goes beyond lowering cholesterol and utilization of a solution nuclear magnetic resonance (NMR) spectroscopy, structure-based drug discovery approach requires milligram quantities of pure, bioactive, recombinant fortilin. Here, we designed expression constructs with different affinity tags and protease cleavage sites to find optimal conditions to obtain the quantity and purity of protein necessary for structure activity relationship studies. Plasmids encoding fortilin with maltose binding protein (MBP), 6-histidine (6His) and glutathione-S-transferase (GST), N- terminal affinity tags were expressed and purified from Escherichia coli (E. coli). Cleavage sites with tobacco etch virus (TEV) protease and human rhinovirus (HRV) 3C protease were assessed. Despite high levels of expression of soluble protein, the fusion constructs were resistant to proteinases without the inclusion of amino acids between the cleavage site and N-terminus. We surveyed constructs with increasing lengths of glycine/serine (GGS) linkers between the cleavage site and fortilin and found that inclusion of at least one GGS insert led to successful protease cleavage and pure fortilin with conserved binding to calcium as measured by NMR

Differential physiological responses to environmental change promote woody shrub expansion

© The Author(s), 2013. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in Ecology and Evolution 3 (2013): 1149–1162, doi:10.1002/ece3.525.Direct and indirect effects of warming are increasingly modifying the carbon-rich vegetation and soils of the Arctic tundra, with important implications for the terrestrial carbon cycle. Understanding the biological and environmental influences on the processes that regulate foliar carbon cycling in tundra species is essential for predicting the future terrestrial carbon balance in this region. To determine the effect of climate change impacts on gas exchange in tundra, we quantified foliar photosynthesis (Anet), respiration in the dark and light (RD and RL, determined using the Kok method), photorespiration (PR), carbon gain efficiency (CGE, the ratio of photosynthetic CO2 uptake to total CO2 exchange of photosynthesis, PR, and respiration), and leaf traits of three dominant species – Betula nana, a woody shrub; Eriophorum vaginatum, a graminoid; and Rubus chamaemorus, a forb – grown under long-term warming and fertilization treatments since 1989 at Toolik Lake, Alaska. Under warming, B. nana exhibited the highest rates of Anet and strongest light inhibition of respiration, increasing CGE nearly 50% compared with leaves grown in ambient conditions, which corresponded to a 52% increase in relative abundance. Gas exchange did not shift under fertilization in B. nana despite increases in leaf N and P and near-complete dominance at the community scale, suggesting a morphological rather than physiological response. Rubus chamaemorus, exhibited minimal shifts in foliar gas exchange, and responded similarly to B. nana under treatment conditions. By contrast, E. vaginatum, did not significantly alter its gas exchange physiology under treatments and exhibited dramatic decreases in relative cover (warming: −19.7%; fertilization: −79.7%; warming with fertilization: −91.1%). Our findings suggest a foliar physiological advantage in the woody shrub B. nana that is further mediated by warming and increased soil nutrient availability, which may facilitate shrub expansion and in turn alter the terrestrial carbon cycle in future tundra environments.This study was supported by the National Science Foundation #0732664; Australian Research Council DP0986823; and Marsden Fund of the Royal Society of New Zealand

Parsing the Behavioral and Brain Mechanisms of Third-Party Punishment.

UnlabelledThe evolved capacity for third-party punishment is considered crucial to the emergence and maintenance of elaborate human social organization and is central to the modern provision of fairness and justice within society. Although it is well established that the mental state of the offender and the severity of the harm he caused are the two primary predictors of punishment decisions, the precise cognitive and brain mechanisms by which these distinct components are evaluated and integrated into a punishment decision are poorly understood. Using fMRI, here we implement a novel experimental design to functionally dissociate the mechanisms underlying evaluation, integration, and decision that were conflated in previous studies of third-party punishment. Behaviorally, the punishment decision is primarily defined by a superadditive interaction between harm and mental state, with subjects weighing the interaction factor more than the single factors of harm and mental state. On a neural level, evaluation of harms engaged brain areas associated with affective and somatosensory processing, whereas mental state evaluation primarily recruited circuitry involved in mentalization. Harm and mental state evaluations are integrated in medial prefrontal and posterior cingulate structures, with the amygdala acting as a pivotal hub of the interaction between harm and mental state. This integrated information is used by the right dorsolateral prefrontal cortex at the time of the decision to assign an appropriate punishment through a distributed coding system. Together, these findings provide a blueprint of the brain mechanisms by which neutral third parties render punishment decisions.Significance statementPunishment undergirds large-scale cooperation and helps dispense criminal justice. Yet it is currently unknown precisely how people assess the mental states of offenders, evaluate the harms they caused, and integrate those two components into a single punishment decision. Using a new design, we isolated these three processes, identifying the distinct brain systems and activities that enable each. Additional findings suggest that the amygdala plays a crucial role in mediating the interaction of mental state and harm information, whereas the dorsolateral prefrontal cortex plays a crucial, final-stage role, both in integrating mental state and harm information and in selecting a suitable punishment amount. These findings deepen our understanding of how punishment decisions are made, which may someday help to improve them

Medical consequences of pathogenic CNVs in adults: Analysis of the UK Biobank

Background: Genomic CNVs increase the risk for early-onset neurodevelopmental disorders, but their impact on medical outcomes in later life is still poorly understood. The UK Biobank allows us to study the medical consequences of CNVs in middle and old age in half a million well-phenotyped adults. Methods: We analysed all Biobank participants for the presence of 54 CNVs associated with genomic disorders or clinical phenotypes, including their reciprocal deletions or duplications. After array quality control and exclusion of first-degree relatives, we compared 381 452 participants of white British or Irish origin who carried no CNVs with carriers of each of the 54 CNVs (ranging from 5 to 2843 persons). We used logistic regression analysis to estimate the risk of developing 58 common medical phenotypes (3132 comparisons). Results and conclusions: Many of the CNVs have profound effects on medical health and mortality, even in people who have largely escaped early neurodevelopmental outcomes. Forty-six CNV–phenotype associations were significant at a false discovery rate threshold of 0.1, all in the direction of increased risk. Known medical consequences of CNVs were confirmed, but most identified associations are novel. Deletions at 16p11.2 and 16p12.1 had the largest numbers of significantly associated phenotypes (seven each). Diabetes, hypertension, obesity and renal failure were affected by the highest numbers of CNVs. Our work should inform clinicians in planning and managing the medical care of CNV carriers

Elementary effects for models with dimensional inputs of arbitrary type and range: Scaling and trajectory generation

The Elementary Effects method is a global sensitivity analysis approach for identifying (un)important parameters in a model. However, it has almost exclusively been used where inputs are dimensionless and take values on [0, 1]. Here, we consider models with dimensional inputs, inputs taking values on arbitrary intervals or discrete inputs. In such cases scaling effects by a function of the input range is essential for correct ranking results. We propose two alternative dimensionless sensitivity indices by normalizing the scaled mean or median of absolute effects. Testing these indices with 9 trajectory generation methods on 4 test functions (including the Penman-Monteith equation for evapotranspiration) reveals that: i) scaled elementary effects are necessary to obtain correct parameter importance rankings; ii) small step-size methods typically produce more accurate rankings; iii) it is beneficial to compute and compare both sensitivity indices; and iv) spread and discrepancy of the simulation points are poor proxies for trajectory generation method performance

Cancer stem cells: Mediators of tumorigenesis and metastasis in head and neck squamous cell carcinoma

BackgroundCancer stem cells (CSCs) represent a subpopulation of cells responsible for tumor growth. Their role in head and neck squamous cell carcinoma (HNSCC) tumorigenesis and metastasis remains uncertain.MethodsWound healing and an orthotopic animal model were used to study cells expressing the CSC phenotype (CD44high and aldehyde dehydrogenase [ALDH]+) and assess mobility, tumorigenesis, and metastasis. A prospective collection of 40 patient‐derived primary HNSCC specimens were analyzed for CSC‐proportion compared to clinical variables.ResultsCSCs exhibited significantly faster wound closure and greater tumorigenesis and regional metastasis in vivo than non‐CSCs. In primary patient tumors, size and advanced stage were correlated with elevated proportion of CSCs, however, not with survival.ConclusionHNSCC stem cells mediate tumorigenesis and regional metastasis in vivo. In primary patient tumors, CSC‐proportion was associated with tumor size and stage, but not with metastatic spread or survival. CSC burden alone may only represent a minor variable in understanding CSCs and metastasis. © 2014 Wiley Periodicals, Inc. Head Neck 37: 317–326, 2015Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/110728/1/hed23600.pd