The origin and future of offshore medical schools in the Caribbean.
The Flexner Report of 1910 concluded that there were too many medical schools in the United States of America (U.S.A.). In the wake of this conclusion, Flexner recommended higher admission and graduation standards for medical schools, the expected impact being fewer medical schools and graduates. One unintended consequence of this recommendation was a male-only policy by some universities to accommodate the smaller numbers of males. There were privately owned medical schools, unaffiliated to any college or universities which were run primarily for profit. Degrees were usually obtained after only two years of study and laboratory and dissection exercises were not part of the training. The adoption of Flexner's recommendation resulted in medical education becoming more expensive; it also created opportunities for persons without the financial means or the matriculation requirements to find alternative training programmes among those "medical schools" which did not adopt Flexner's recommendations. This paper traces the further development of these "medical schools" to facilitate the need for more doctors in the context of global maldistribution of doctors which has resulted in many medically underserved areas, more so in developing countries (1).


Identity, morals, and taboos: beliefs as assets.
We develop a theory of moral behavior, individual and collective, based on a general model of identity in which people care about "who they are" and infer their own values from past choices. The model sheds light on many empirical puzzles inconsistent with earlier approaches. Identity investments respond nonmonotonically to acts or threats, and taboos on mere thoughts arise to protect beliefs about the "priceless" value of certain social assets. High endowments trigger escalating commitment and a treadmill effect, while competing identities can cause dysfunctional capital destruction. Social interactions induce both social and antisocial norms of contribution, sustained by respectively shunning free riders or do-gooders.


Cavity length resonances in a nanosecond singly resonant optical parametric oscillator.
Resonant output energy enhancement in a singly resonant nondegenerate type-I optical parametric oscillator with a volume Bragg grating output coupler is demonstrated. The resonances occur when the pump laser and parametric oscillator cavity length ratio is an integer or a fraction of small integers. Although the length resonances are similar to those observed in doubly resonant optical parametric oscillators, the physical mechanism is distinctly different. The resonances in the singly resonant oscillator are caused by correlation of the instantaneous power between the quasi-periodic multimode pump laser beam and the OPO signal.


Which growth parameters can affect mortality in cerebral palsy?
The objective of this study was to identify growth parameters that can affect mortality of cerebral palsy (CP). This was a birth cohort study based on the National Health Screening Program for Infants and Children database along with the National Health Insurance Service, which were linked using a personal identifier number. The birth cohort consisted of 2 191 956 subjects, representing 93.5% of live births from 2007-2011, with maximal 10-year follow-up (range, 5-10 years) until October 2016. Subjects with CP were identified. Growth parameters in terms of birth weight, underweight (weight-for-age below the 3rd percentile), rate of body weight gain were collected, along with all-cause mortality after the age of 1 year. Prevalence of CP was 2.0 per 1000 live births (95% CI, 1.94-2.06). All-cause mortality after the age of 1 year was 0.09 deaths/1000 person-years (95% CI, 0.08-0.09) in the general population (GP) and 2.85 deaths/1000 person-years (95% CI, 2.32-3.50) in subjects with CP during the follow-up. Therefore, the incidence rate ratio for all-cause mortality was 32.15 (95% CI, 25.72-39.76) in subjects with CP compared to GP. Presence of underweight was significantly associated with higher mortality in both subjects with CP and GP, where the adjusted hazard ratio of death was 2.60 (95% CI, 1.93-3.50) at the age of 18-24 months, 3.12 at 30-36 months, 4.37 at 42-48 months, 5.12 at 54-60 months, and 4.17 at 66-71 months. Birth weight did not affect mortality in both subjects with CP and GP after the age of 1 year (p > 0.05). While subjects with CP shows higher mortality, underweight is an important growth parameter that affects all-cause mortality of both subjects with CP and GP. This study urges increased awareness that subjects with CP who are underweight require special care.


'AND'-based fluorescence scaffold for the detection of ROS/RNS and a second analyte.
Traditionally, fluorescence probes have focused on the detection of a single biomarker for a specific process. In this work, we set out to develop a number of fluorescence probes that enable the detection of a chosen analyte in the presence of reactive oxygen/nitrogen species (ROS/RNS). These fluorescence probes when activated result in the formation of the highly fluorescent pink dye, resorufin. Therefore, we have labelled these fluorescent probes as 'Pinkments'. Our first 'Pinkment' was shown to detect biologically relevant concentrations of ONOO- and have an excellent selectivity against other ROS/RNS. Pinkment-OH was developed to provide a core unit which could be easily functionalised to produce a range of 'AND' based fluorescence probes for the detection of ROS/RNS and a second analyte. For proof of concept, we synthesised Pinkment-OTBS and Pinkment-OAc. These 'AND'-based probes were successfully shown to detect ROS/RNS and F- or esterase, respectively.


Response of Nitrobacter spp. ribosomal gene and transcript abundance following nitrite starvation and exposure to mechanistically distinct inhibitors.
The Nitrobacter spp. rRNA gene (rDNA) and relative rRNA transcript abundance (rRNAt/rDNA ratio) were evaluated in response to sudden changes in the nitrite oxidation rate. The rDNA abundance poorly indicated sudden transitions in the rate, whereas the relative rRNAt abundance usually varied quickly and significantly. In response to changes in nitrite concentration, 8 h were required for the rRNAt/rDNA ratio to transition from a minimum value at nitrite starvation (approximately 0.07) to a maximum value with excess nitrite present (approximately 4), and 5 h were required for this metric to return to the minimum value after nitrite starvation re-ensued. Generally, the relative rRNAt abundance dropped significantly after 4.5 h of exposure to three different inhibitors. A sharp decline in the rRNAt/rDNA ratio occurred during exposure to 3,5-DCP (from 4 down to 0.2) even as the fractional inhibition level remained low (< 0.10); the minimum ratio value was observed when nitrite oxidation was completely inhibited. The ratio decreased significantly during exposure to azide (from 4 to 0.5) and H+ (from 2 to 0.2), but only when the fractional inhibition levels were high (> 0.8). Interestingly, when the pH was suddenly changed to 4.5, inhibiting nitrite oxidation completely, the rRNAt/rDNA metric did not decline suggesting that rRNAt processing was inhibited. This effect was not observed during severe inhibition with 3,5-DCP and azide. Overall, the findings indicate the relative rRNAt abundance can be used to closely track in situ Nitrobacter spp. activity and in most instances will reveal inhibition events with the potential to impact treatment performance in reactors where Nitrobacter spp. are dominant.


High throughput methods to measure the propensity of compounds to form intramolecular hydrogen bonding.
Implementation of IMHB considerations in drug discovery needs robust and validated descriptors to experimentally verify the propensity of compounds to exhibit IMHBs. The first part of the paper presents an overview of the most common techniques to measure the propensity of compounds to form IMHBs. Then we review and discuss recently proposed high throughput (HT) physicochemical descriptors (i.e. Δlog Poct-tol, EPSA and log k'80 PLRP-S) which provide the same information. Analysis of the available data enabled us to extract guidelines for the application of these descriptors in drug discovery programs.


Effectiveness of biological surrogates for predicting patterns of marine biodiversity: a global meta-analysis.
The use of biological surrogates as proxies for biodiversity patterns is gaining popularity, particularly in marine systems where field surveys can be expensive and species richness high. Yet, uncertainty regarding their applicability remains because of inconsistency of definitions, a lack of standard methods for estimating effectiveness, and variable spatial scales considered. We present a Bayesian meta-analysis of the effectiveness of biological surrogates in marine ecosystems. Surrogate effectiveness was defined both as the proportion of surrogacy tests where predictions based on surrogates were better than random (i.e., low probability of making a Type I error; P) and as the predictability of targets using surrogates (R(2)). A total of 264 published surrogacy tests combined with prior probabilities elicited from eight international experts demonstrated that the habitat, spatial scale, type of surrogate and statistical method used all influenced surrogate effectiveness, at least according to either P or R(2). The type of surrogate used (higher-taxa, cross-taxa or subset taxa) was the best predictor of P, with the higher-taxa surrogates outperforming all others. The marine habitat was the best predictor of R(2), with particularly low predictability in tropical reefs. Surrogate effectiveness was greatest for higher-taxa surrogates at a <10-km spatial scale, in low-complexity marine habitats such as soft bottoms, and using multivariate-based methods. Comparisons with terrestrial studies in terms of the methods used to study surrogates revealed that marine applications still ignore some problems with several widely used statistical approaches to surrogacy. Our study provides a benchmark for the reliable use of biological surrogates in marine ecosystems, and highlights directions for future development of biological surrogates in predicting biodiversity.


Synthesis, characterization, and photoinduced antibacterial activity of porphyrin-type photosensitizers conjugated to the antimicrobial peptide apidaecin 1b.
Antimicrobial photodynamic therapy (aPDT) is an emerging treatment for bacterial infections that is becoming increasingly more attractive because of its effectiveness against multi-antibiotic-resistant strains and unlikelihood of inducing bacterial resistance. Among the strategies to enhance the efficacy of PDT against Gram-negative bacteria, the binding to a cationic antimicrobial peptide offers the attractive prospect for improving both the water solubilty and the localization of the photoactive drug in bacteria. In this work we have compared a number of free and apidaecin-conjugated photosensitizers (PSs) differing in structure and charge. Our results indicate that the conjugation of per se ineffective highly hydrophobic PSs to a cationic peptide produces a photosensitizing agent effective against Gram-negative bacteria. Apidaecin cannot improve the phototoxic activity of cationic PSs, which mainly depends on a very high yield of singlet oxygen production in the surroundings of the bacterial outer membrane. Apidaecin-PS conjugates appear most promising for treatment protocols requiring repeated washing after sensitizer delivery.


Learning neuroendoscopy with an exoscope system (video telescopic operating monitor): Early clinical results.
Steep learning curve is found initially in pure endoscopic procedures. Video telescopic operating monitor (VITOM) is an advance in rigid-lens telescope systems provides an alternative method for learning basics of neuroendoscopy with the help of the familiar principle of microneurosurgery. The aim was to evaluate the clinical utility of VITOM as a learning tool for neuroendoscopy. Video telescopic operating monitor was used 39 cranial and spinal procedures and its utility as a tool for minimally invasive neurosurgery and neuroendoscopy for initial learning curve was studied. Video telescopic operating monitor was used in 25 cranial and 14 spinal procedures. Image quality is comparable to endoscope and microscope. Surgeons comfort improved with VITOM. Frequent repositioning of scope holder and lack of stereopsis is initial limiting factor was compensated for with repeated procedures. Video telescopic operating monitor is found useful to reduce initial learning curve of neuroendoscopy.


The interacting roles of climate, soils, and plant production on soil microbial communities at a continental scale.
Soil microbial communities control critical ecosystem processes such as decomposition, nutrient cycling, and soil organic matter formation. Continental scale patterns in the composition and functioning of microbial communities are related to climatic, biotic, and edaphic factors such as temperature and precipitation, plant community composition, and soil carbon, nitrogen, and pH. Although these relationships have been well explored individually, the examination of the factors that may act directly on microbial communities vs. those that may act indirectly through other ecosystem properties has not been well developed. To further such understanding, we utilized structural equation modeling (SEM) to evaluate a set of hypotheses about the direct and indirect effects of climatic, biotic, and edaphic variables on microbial communities across the continental United States. The primary goals of this work were to test our current understanding of the interactions among climate, soils, and plants in affecting microbial community composition, and to examine whether variation in the composition of the microbial community affects potential rates of soil enzymatic activities. A model of interacting factors created through SEM shows several expected patterns. Distal factors such as climate had indirect effects on microbial communities by influencing plant productivity, soil mineralogy, and soil pH, but factors related to soil organic matter chemistry had the most direct influence on community composition. We observed that both plant productivity and soil mineral composition were important indirect influences on community composition at the continental scale, both interacting to affect organic matter content and microbial biomass and ultimately community composition. Although soil hydrolytic enzymes were related to the moisture regime and soil carbon, oxidative enzymes were also affected by community composition, reflected in the abundance of soil fungi. These results highlight that soil microbial communities can be modeled within the context of multiple interacting ecosystem properties acting both directly and indirectly on their composition and function, and this provides a rich and informative context with which to examine communities. This work also highlights that variation in climate, microbial biomass, and microbial community composition can affect maximum rates of soil enzyme activities, potentially influencing rates of decomposition and nutrient mineralization in soils.


Arabidopsis thaliana plants expressing Rift Valley fever virus antigens: Mice exhibit systemic immune responses as the result of oral administration of the transgenic plants.
The zoonotic Rift Valley fever virus affects livestock and humans in Africa and on the Arabian Peninsula. The economic impact of this pathogen due to livestock losses, as well as its relevance to public health, underscores the importance of developing effective and easily distributed vaccines. Vaccines that can be delivered orally are of particular interest. Here, we report the expression in transformed plants (Arabidopsis thaliana) of Rift Valley fever virus antigens. The antigens used in this study were the N protein and a deletion mutant of the Gn glycoprotein. Transformed lines were analysed for specific mRNA and protein content by RT-PCR and Western blotting, respectively. Furthermore, the plant-expressed antigens were evaluated for their immunogenicity in mice fed the transgenic plants. After oral intake of fresh transgenic plant material, a proportion of the mice elicited specific IgG antibody responses, as compared to the control animals that were fed wild-type plants and of which none sero-converted. Thus, we show that transgenic plants can be readily used to express and produce Rift Valley Fever virus proteins, and that the plants are immunogenic when given orally to mice. These are promising findings and provide a basis for further studies on edible plant vaccines against the Rift Valley fever virus.


A Genome-wide Association Study of Nonsyndromic Cleft Palate Identifies an Etiologic Missense Variant in GRHL3.
Cleft palate (CP) is a common birth defect occurring in 1 in 2,500 live births. Approximately half of infants with CP have a syndromic form, exhibiting other physical and cognitive disabilities. The other half have nonsyndromic CP, and to date, few genes associated with risk for nonsyndromic CP have been characterized. To identify such risk factors, we performed a genome-wide association study of this disorder. We discovered a genome-wide significant association with a missense variant in GRHL3 (p.Thr454Met [c.1361C>T]; rs41268753; p = 4.08 × 10(-9)) and replicated the result in an independent sample of case and control subjects. In both the discovery and replication samples, rs41268753 conferred increased risk for CP (OR = 8.3, 95% CI 4.1-16.8; OR = 2.16, 95% CI 1.43-3.27, respectively). In luciferase transactivation assays, p.Thr454Met had about one-third of the activity of wild-type GRHL3, and in zebrafish embryos, perturbed periderm development. We conclude that this mutation is an etiologic variant for nonsyndromic CP and is one of few functional variants identified to date for nonsyndromic orofacial clefting. This finding advances our understanding of the genetic basis of craniofacial development and might ultimately lead to improvements in recurrence risk prediction, treatment, and prognosis.


Nuclear import of exogenous FGF1 requires the ER-protein LRRC59 and the importins Kpnα1 and Kpnβ1.
Fibroblast growth factor 1 (FGF1) taken up by cells into endocytic vesicles can be translocated across vesicular membranes into the cytosol and the nucleus where it has a growth regulatory activity. Previously, leucine-rich repeat containing 59 (LRRC59) was identified as an intracellular binding partner of FGF1, but its biological role remained unknown. Here, we show that LRRC59 is strictly required for nuclear import of exogenous FGF1. siRNA-mediated depletion of LRRC59 did not inhibit the translocation of FGF1 into cytosol, but blocked the nuclear import of FGF1. We also found that an nuclear localization sequence (NLS) in FGF1, Ran GTPase, karyopherin-α1 (Kpnα1), and Kpnβ1 were required for nuclear import of FGF1. Nuclear import of exogenous FGF2, which depends on CEP57/Translokin, was independent of LRRC59, but was dependent on Kpnα1 and Kpnβ1, while the nuclear import of FGF1 was independent of CEP57. LRRC59 is a membrane-anchored protein that localizes to the endoplasmic reticulum (ER) and the nuclear envelope (NE). We found that LRRC59 possesses NLS-like sequences in its cytosolic part that can mediate nuclear import of soluble LRRC59 variants, and that the localization of LRRC59 to the NE depends on Kpnβ1. We propose that LRRC59 facilitates transport of cytosolic FGF1 through nuclear pores by interaction with Kpns and movement of LRRC59 along the ER and NE membranes.


Vascular risk status as a predictor of later-life depressive symptoms: a cohort study.
Common etiology of vascular diseases and later-life depression may provide important synergies for prevention. We examined whether standard clinical risk profiles developed for vascular diseases also predict depressive symptoms in older adults. Data were drawn from the Whitehall II study with baseline examination in 1991; follow-up screenings in 1997, 2003, and 2008; and additional disease ascertainment from hospital data and registry linkage on 5318 participants (mean age 54.8 years, 31% women) without depressive symptoms at baseline. Vascular risk was assessed with the Framingham Cardiovascular, Coronary Heart Disease, and Stroke Risk Scores. New depressive symptoms at each follow-up screening were identified by General Health Questionnaire caseness, a Center for Epidemiologic Studies Depression Scale score ≥16, and use of antidepressant medication. Diagnosed vascular disease (that is, coronary heart disease or stroke) was associated with an increased risk for depressive symptoms, age- and sex-adjusted odds ratios from 1.5 (95% confidence interval 1.0-2.2) to 2.0 (1.4-3.0), depending on the indicator of depressive symptoms. Among participants without manifest vascular disease, the Stroke Risk Score was associated with Center for Epidemiologic Studies Depression Scale depressive symptoms before age 65 (age- and sex-adjusted odds ratio per 10% absolute change in the score = 3.1 [1.5-6.5]), but none of the risk scores predicted new-onset depressive symptoms in those aged ≥65 (odds ratios from .8 to 1.2). These data suggest that public health measures to improve vascular risk status will influence the incidence of later-life depressive symptoms via reduced rates of manifest vascular disease.


Avocado Intake, and Longitudinal Weight and Body Mass Index Changes in an Adult Cohort.
Avocados contain nutrients and bioactive compounds that may help reduce the risk of becoming overweight/obese. We prospectively examined the effect of habitual avocado intake on changes in weight and body mass index (BMI). In the Adventist Health Study (AHS-2), a longitudinal cohort (~55,407; mean age ~56 years; U.S. and Canada), avocado intake (standard serving size 32 g/day) was assessed by a food frequency questionnaire (FFQ). Self-reported height and weight were collected at baseline. Self-reported follow-up weight was collected with follow-up questionnaires between four and 11 years after baseline. Using the generalized least squares (GLS) approach, we analyzed repeated measures of weight in relation to avocado intake. Marginal logistic regression analyses were used to calculate the odds of becoming overweight/obese, comparing low (>0 to <32 g/day) and high (≥32 g/day) avocado intake to non-consumers (reference). Avocado consumers who were normal weight at baseline, gained significantly less weight than non-consumers. The odds (OR (95% CI)) of becoming overweight/obese between baseline and follow-up was 0.93 (0.85, 1.01), and 0.85 (0.60, 1.19) for low and high avocado consumers, respectively. Habitual consumption of avocados may reduce adult weight gain, but odds of overweight/obesity are attenuated by differences in initial BMI values.


The 5-CNL Front-of-Pack Nutrition Label Appears an Effective Tool to Achieve Food Substitutions towards Healthier Diets across Dietary Profiles.
Front-of-pack (FOP) nutrition labels are considered helpful tools to help consumers making healthier food choices, thus improving their diet. In France, the implementation of a FOP nutrition label-the 5-Colour Nutrition Label (5-CNL)-is currently under consideration. Our objective was to investigate dietary profiles in a French adult population using the 5-CNL, and to assess its potential impact in improving the diet through substitution of foods. Subjects included in the NutriNet-Santé cohort, who had completed three 24-h dietary records were included in this cross-sectional analysis. Mutually exclusive clusters of individuals were identified using the percentage of energy derived from foods of each of the 5-CNL colours as input variables. Three scenarios of substitution of foods for healthier alternative using the 5-CNL were tested. Food group and dietary intakes, socio-demographic and lifestyle data were compared across clusters using ANOVAs or Chi-square tests, as appropriate. We identified three mutually exclusive dietary profiles: 'Healthy' (N = 28 095, 29.3% of the sample), with high consumption of fruit, vegetables, whole cereals and fish; 'Western' (N = 33 386, 34.8%); with high consumption of sweetened beverages, breakfast cereal, cheese, fatty and sugary foods; 'Traditional' (N = 34 461, 35.1%), with high consumption of potatoes, bread, meat and dairy desserts. Overall, substitutions strategies led to an increase in the number of subjects reaching the recommended intakes in energy, macro and micronutrients. Increases were particularly high in the 'Western' pattern for lipids and saturates intakes: from 16.2% reaching the recommended amount for lipids (13.5% for saturates) to 60.6% and 85.7% respectively. The use of the 5-CNL as an indicator of food choice meaningfully characterizes clusters of dietary habits and appears as an effective tool to help improving the nutritional quality of the diet.


Deferasirox for managing iron overload in people with myelodysplastic syndrome.
The myelodysplastic syndrome (MDS) comprises a diverse group of haematopoietic stem cell disorders. Due to symptomatic anaemia, most people with MDS require supportive therapy including repeated red blood cell (RBC) transfusions. In combination with increased iron absorption, this contributes to the accumulation of iron resulting in secondary iron overload and the risk of organ dysfunction and reduced life expectancy. Since the human body has no natural means of removing excess iron, iron chelation therapy, i.e. the pharmacological treatment of iron overload, is usually recommended. However, it is unclear whether or not the newer oral chelator deferasirox leads to relevant benefit. To evaluate the effectiveness and safety of oral deferasirox for managing iron overload in people with myelodysplastic syndrome (MDS). We searched the following databases up to 03 April 2014: MEDLINE, EMBASE, The Cochrane Library, Biosis Previews, Web of Science, Derwent Drug File and four trial registries: Current Controlled Trials (www.controlled-trials.com), ClinicalTrials.gov (www.clinicaltrials.gov), ICTRP (www.who.int./ictrp/en/), and German Clinical Trial Register (www.drks.de). Randomised controlled trials (RCTs) comparing deferasirox with no therapy, placebo or with another iron-chelating treatment schedule. We did not identify any trials eligible for inclusion in this review. No trials met our inclusion criteria. However, we identified three ongoing and one completed trial (published as an abstract only and in insufficient detail to permit us to decide on inclusion) comparing deferasirox with deferoxamine, placebo or no treatment. We planned to report evidence from RCTs that evaluated the effectiveness of deferasirox compared to either placebo, no treatment or other chelating regimens, such as deferoxamine, in people with MDS. However, we did not identify any completed RCTs addressing this question.We found three ongoing and one completed RCT (published as an abstract only and in insufficient detail) comparing deferasirox with deferoxamine, placebo or no treatment and data will hopefully be available soon. These results will be important to inform physicians and patients on the advantages and disadvantages of this treatment option.


Microscale mapping of extracellular matrix elasticity of mouse joint cartilage: an approach to extracting bulk elasticity of soft matter with surface roughness.
Cartilage is composed of cells and an extracellular matrix, the latter being a composite of a collagen mesh interpenetrated by proteoglycans responsible for tissue osmotic swelling. The matrix composition and structure vary through the tissue depth. Mapping such variability requires tissue sectioning to gain access. The resulting surface roughness, and concomitant proteoglycan loss contribute to large uncertainties in elastic modulus estimates. To extract elasticity values for the bulk matrix which are not obfuscated by the indeterminate surface layer, we developed a novel experimental and data analysis methodology. We analyzed the surface roughness to optimize the probe size, and performed high-resolution (1 μm) elasticity mapping on thin (∼12 μm), epiphyseal newborn mouse cartilage sections cut parallel to the bone longitudinal axis or normal to the articular surface. Mild fixation prevented the major proteoglycan loss observed in unfixed specimens but not the stress release that resulted in thickness changes in the sectioned matrix. Our novel data analysis method introduces a virtual contact point as a fitting parameter for the Hertz model, to minimize the effects of surface roughness and corrects for the finite section thickness. Our estimates of cartilage elasticity converge with increasing indentation depth and, unlike previous data interpretations, are consistent with linearly elastic material. A high cell density that leaves narrow matrix septa between cells may cause the underestimation of elastic moduli, whereas fixation probably causes an overestimation. The proposed methodology has broader relevance to nano- and micro-indentation of soft materials with multiple length scales of organization and whenever surface effects (including roughness, electrostatics, van der Waals forces, etc.) become significant.


A Phase I Study of the Safety, Pharmacokinetics, and Pharmacodynamics of Combination Therapy with Refametinib plus Sorafenib in Patients with Advanced Cancer.
To assess the safety and tolerability of the small-molecule allosteric MEK inhibitor refametinib combined with sorafenib, in patients with advanced solid malignancies. This phase I dose-escalation study included an expansion phase at the maximum tolerated dose (MTD). Patients received refametinib/sorafenib twice daily for 28 days, from a dose of refametinib 5 mg plus sorafenib 200 mg to a dose of refametinib 50 mg plus sorafenib 400 mg. Plasma levels of refametinib, refametinib metabolite M17, and sorafenib were measured for pharmacokinetic assessments. Tumors were biopsied at the MTD for analysis of MEK pathway mutations and ERK phosphorylation. Thirty-two patients were enrolled in the dose-escalation cohort. The MTD was refametinib 50 mg twice daily plus sorafenib 400 mg twice daily. The most common treatment-related toxicities were diarrhea and fatigue. Refametinib was readily absorbed following oral administration (plasma half-life of ∼16 hours at the MTD), and pharmacokinetic parameters displayed near-dose proportionality, with less than 2-fold accumulation after multiple dosing. Another 30 patients were enrolled in the MTD cohort; 19 had hepatocellular carcinoma. The combination was associated with significantly reduced ERK phosphorylation in 5 out of 6 patients biopsied, with the greatest reductions in those with KRAS or BRAF mutations. Disease was stabilized in approximately half of patients, and 1 patient with colorectal cancer achieved a partial response at the MTD lasting approximately 1 year. In this phase I study, refametinib plus sorafenib was well tolerated, with good oral absorption, near-dose proportionality, and target inhibition in a range of tumor types. Clin Cancer Res; 22(10); 2368-76. ©2015 AACR.


Binding of diamine oxidase activity to rat and guinea pig microvascular endothelial cells. Comparisons with lipoprotein lipase binding.
Microvascular endothelial cells from rat and guinea pig fat pads were shown to bind diamine oxidase (DAO) activity when incubated with soluble extracts of placenta (33 DAO U/mg of placenta) and a purified placental enzyme preparation (94 U/micrograms of protein). The extent of binding was dependent on the concentration of enzyme activity and tissue. Saturation of binding sites with 5,000 U of DAO/ml resulted in levels of bound activity (up to 11-13 U/mg of endothelial cells) in excess of that observed in all tissues except placenta. Scatchard plots suggested that there were at least two DAO binding sites (apparent Km 92 and 2,450 U/ml). Although the same cell preparations bound 125I-labeled lipoprotein lipase (LPL), the presence of LPL on the endothelial cell surface did not interfere with the binding of DAO activity except when cells were exposed to high concentrations of LPL. Alternatively, bound DAO activity was partially displaced (up to 33%) only with high concentrations (30 micrograms/ml) of LPL. DAO activity may thus be bound to at least two populations of sites, one of which may bind LPL. Both enzymes, however, were displaced by heparin (0.05-5 U/ml) and DAO binding was impaired by prior treatment of cells with proteolytic and glycosaminoglycandegrading enzymes. The demonstration of DAO binding to vascular endothelial cells provides a further example of the ability of these cells to bind enzymes at their surface and thereby act on biologically active substances in the circulation.


Pathway analysis of transcriptomic data shows immunometabolic effects of vitamin D.
Unbiased genomic screening analyses have highlighted novel immunomodulatory properties of the active form of vitamin D, 1,25-dihydroxyvitamin D (1,25(OH)2D). However, clearer interpretation of the resulting gene expression data is limited by cell model specificity. The aim of the current study was to provide a broader perspective on common gene regulatory pathways associated with innate immune responses to 1,25(OH)2D, through systematic re-interrogation of existing gene expression databases from multiple related monocyte models (the THP-1 monocytic cell line (THP-1), monocyte-derived dendritic cells (DCs) and monocytes). Vitamin D receptor (VDR) expression is common to multiple immune cell types, and thus, pathway analysis of gene expression using data from multiple related models provides an inclusive perspective on the immunomodulatory impact of vitamin D. A bioinformatic workflow incorporating pathway analysis using PathVisio and WikiPathways was utilized to compare each set of gene expression data based on pathway-level context. Using this strategy, pathways related to the TCA cycle, oxidative phosphorylation and ATP synthesis and metabolism were shown to be significantly regulated by 1,25(OH)2D in each of the repository models (Z-scores 3.52-8.22). Common regulation by 1,25(OH)2D was also observed for pathways associated with apoptosis and the regulation of apoptosis (Z-scores 2.49-3.81). In contrast to the primary culture DC and monocyte models, the THP-1 myelomonocytic cell line showed strong regulation of pathways associated with cell proliferation and DNA replication (Z-scores 6.1-12.6). In short, data presented here support a fundamental role for active 1,25(OH)2D as a pivotal regulator of immunometabolism.


Validation of a prototype DiodeAir for small field dosimetry.
Standard commercial diode detectors over-respond within small radiation fields, an effect largely attributable to the relatively high mass-density of silicon. However, Monte Carlo studies can be used to optimise dosimeter designs and have demonstrated that 'mass-density compensation'-for example, introducing a low-density air-gap upstream of a diode's high-density silicon volume-can substantially improve instrument response. In this work we used egs_chamber Monte Carlo simulations to predict the ideal air-gap thickness for a PTW 60017 unshielded diode detector. We then developed a prototype instrument incorporating that air-gap and, for a 6 MV linac, tested it experimentally against EBT3 film. We also tested a further three prototypes with different air-gap thicknesses. Our results demonstrate that for a 10 × 10 cm(2) reference field the DiodeAir, a PTW 60017 diode with a built-in air-gap of 1 mm, has on-axis correction factors near unity. Laterally the DiodeAir performs very well off-axis and reports FWHM and penumbra values consistent with those measured using EBT3. For PDD measurement, the performance of the DiodeAir matches that of the original PTW 60017. The experimental focus of this work was 6 MV but we also simulated the on-axis response of the DiodeAir within 15 MV beams and found that our modification proved robust to this substantial increase in beam energy. However, the original diode 60017 does exhibit low energy scatter dependencies and may over-respond to high linac dose-rates such that applying the mass-density compensation method to an alternative instrument (particularly a diamond detector) could ultimately take us even closer to the small-field ideal.


T cell immunity as a tool for studying epigenetic regulation of cellular differentiation.
Cellular differentiation is regulated by the strict spatial and temporal control of gene expression. This is achieved, in part, by regulating changes in histone post-translational modifications (PTMs) and DNA methylation that in turn, impact transcriptional activity. Further, histone PTMs and DNA methylation are often propagated faithfully at cell division (termed epigenetic propagation), and thus contribute to maintaining cellular identity in the absence of signals driving differentiation. Cardinal features of adaptive T cell immunity include the ability to differentiate in response to infection, resulting in acquisition of immune functions required for pathogen clearance; and the ability to maintain this functional capacity in the long-term, allowing more rapid and effective pathogen elimination following re-infection. These characteristics underpin vaccination strategies by effectively establishing a long-lived T cell population that contributes to an immunologically protective state (termed immunological memory). As we discuss in this review, epigenetic mechanisms provide attractive and powerful explanations for key aspects of T cell-mediated immunity - most obviously and notably, immunological memory, because of the capacity of epigenetic circuits to perpetuate cellular identities in the absence of the initial signals that drive differentiation. Indeed, T cell responses to infection are an ideal model system for studying how epigenetic factors shape cellular differentiation and development generally. This review will examine how epigenetic mechanisms regulate T cell function and differentiation, and how these model systems are providing general insights into the epigenetic regulation of gene transcription during cellular differentiation.


Frequency and Phase Synchronization in Distributed (Implantable-Transcutaneous) Neural Interfaces.
Synchronized oscillations are a ubiquitous feature of neuronal circuits and can modulate online information transfer and plasticity between brain areas. The disruption of these oscillatory processes is associated with the symptoms of several brain disorders. While conventional therapeutic high-frequency deep brain stimulation can perturb neuronal oscillations, manipulating the timing of oscillatory activity between areas more precisely could provide a more efficient and effective method of modulating these activities. Here we describe a prototype circuit for synchronizing the clocks between an active implantable and an external sensing and stimulation system that could be used to achieve this goal. Our specific focus is on synchronizing the systems for paired-associative stimulation. The ability to repetitively drive two brain regions with a fixed latency has specific implications for neural plasticity. Furthermore, the general concept can be applied for many potential applications involving distributed neural interfaces.


Bevacizumab beyond first progression is associated with prolonged overall survival in metastatic colorectal cancer: results from a large observational cohort study (BRiTE).
Bevacizumab provides a survival benefit in first- and second-line metastatic colorectal cancer (mCRC). In a large, observational, bevacizumab treatment study (Bevacizumab Regimens: Investigation of Treatment Effects and Safety [BRiTE]) in patients who had mCRC, a longer-than-expected overall survival (OS) of 25.1 months was reported. The association between various pre- and post-treatment factors (including the use of bevacizumab beyond first progression [BBP]) and survival was examined. The 1,445 of 1,953 previously untreated patients with mCRC who were enrolled in BRiTE and who experienced disease progression (PD) were classified into three groups: no post-PD treatment (n = 253), post-PD treatment without bevacizumab (no BBP; n = 531), and BBP (n = 642). Relevant baseline and on-study variables, including BBP, were analyzed with a Cox model with respect to their independent effect on survival beyond first PD. Median OS was 25.1 months (95% CI, 23.4 to 27.5 months), and median progression-free survival was 10.0 months in the overall BRiTE population. Baseline and postbaseline factors were well balanced between the BBP and no-BBP groups. Median OS rates were 12.6, 19.9, and 31.8 months in the no post-PD treatment, no-BBP, and BBP groups, respectively. In multivariate analyses, compared with no BBP, BBP was strongly and independently associated with improved survival (HR, 0.48; P < .001). Hypertension that required medication was the only bevacizumab-related safety event that occurred more frequently in the BBP group (24.6% v 19.2%). These results from a large, prospective, observational study suggest that continued vascular endothelial growth factor inhibition with bevacizumab beyond initial PD could play an important role improving the overall success of therapy for patients who have mCRC.


Technical aspects of osteoid osteoma ablation in children using MR-guided high intensity focussed ultrasound.
Osteoid osteoma (OO) is a painful bone tumour occurring in children and young adults. Magnetic resonance imaging-guided high intensity focussed ultrasound (MR-HIFU) allows non-invasive treatment without ionising radiation exposure, in contrast to the current standard of care treatment with radiofrequency ablation (RFA). This report describes technical aspects of MR-HIFU ablation in the first 8 paediatric OO patients treated in a safety and feasibility clinical trial (total enrolment of up to 12 patients). OO lesions and adjacent periosteum were treated with MR-HIFU ablation in 5-20 sonications (sonication duration = 16-48 s, frequency = 1.2 MHz, acoustic power = 20-160 W). Detailed treatment workflow, patient positioning and coupling strategies, as well as temperature and tissue perfusion changes were summarised and correlated. MR-HIFU ablation was feasible in all eight cases. Ultrasound standoff pads were shaped to conform to extremity contours providing acoustic coupling and aided patient positioning. The energy delivered was 10 ± 7 kJ per treatment, raising maximum temperature to 83 ± 3 °C. Post ablation contrast-enhanced MRI showed ablated volumes ranging 0.46-19.4 cm3 extending further into bone (7 ± 4 mm) than into soft tissue (4 ± 6 mm, p = 0.01, Mann-Whitney). Treatment time ranged 30-86 min for sonication and 160 ± 40 min for anaesthesia. No serious treatment-related adverse events were observed. Complete pain relief with no medication occurred in 7/8 patients within 28 days following treatment. MR-HIFU ablation of painful OO appears technically feasible in children and it may become a non-invasive and radiation-free alternative for painful OO. Therapy success, efficiency, and applicability may be improved through specialised equipment designed more specifically for extremity bone ablation.


Identification of SLC41A3 as a novel player in magnesium homeostasis.
Regulation of the body Mg(2+) balance takes place in the distal convoluted tubule (DCT), where transcellular reabsorption determines the final urinary Mg(2+) excretion. The basolateral Mg(2+) extrusion mechanism in the DCT is still unknown, but recent findings suggest that SLC41 proteins contribute to Mg(2+) extrusion. The aim of this study was, therefore, to characterize the functional role of SLC41A3 in Mg(2+) homeostasis using the Slc41a3 knockout (Slc41a3(-/-)) mouse. By quantitative PCR analysis it was shown that Slc41a3 is the only SLC41 isoform with enriched expression in the DCT. Interestingly, serum and urine electrolyte determinations demonstrated that Slc41a3(-/-) mice suffer from hypomagnesemia. The intestinal Mg(2+) absorption capacity was measured using the stable (25)Mg(2+) isotope in mice fed a low Mg(2+) diet. (25)Mg(2+) uptake was similar in wildtype (Slc41a3(+/+)) and Slc41a3(-/-) mice, although Slc41a3(-/-) animals exhibited increased intestinal mRNA expression of Mg(2+) transporters Trpm6 and Slc41a1. Remarkably, some of the Slc41a3(-/-) mice developed severe unilateral hydronephrosis. In conclusion, SLC41A3 was established as a new factor for Mg(2+) handling.


Hypertension as a risk factor for renal disease. Review of clinical and epidemiological evidence.
Renal failure is a well-recognized complication of accelerated and malignant hypertension that can be prevented by appropriate antihypertensive therapy. The risk of renal disease in patients with milder forms of hypertension is less certain. Death certificate, insurance industry, and community-based observational studies provide little information with respect to the risk of kidney damage in the hypertensive patient. Randomized controlled trials of antihypertensive drug therapy provide equivocal results. The strongest evidence in support of the hypothesis that mild hypertension increases the risk of subsequent renal disease comes from analyses of the Hypertension Detection and Follow-up Program and the Medicare End-Stage Renal Disease (ESRD) Program. The risk of blood pressure-related ESRD appears to be especially high in blacks, independent of age, sex, and prevalence of hypertension. Although these results need to be confirmed, the data raise the possibility that the effects of high blood pressure on the kidney are different in blacks than in whites. Based on current trends, it is projected that the number of beneficiaries in the Medicare ESRD Program will continue to increase during the next 30-40 years and that the diagnosis of hypertensive ESRD will become increasingly common. Additional studies to characterize the relation between mild hypertension and subsequent risk of renal disease and to confirm the corresponding benefits of antihypertensive therapy are urgently needed.


Modelling the genital herpes epidemic.
Mathematical models are useful tools for summarizing and testing current knowledge about a system and predicting trends. Models have shown that medical and behavioural changes can substantially affect herpes simplex virus type 2 (HSV-2) transmission and can be used to develop rational epidemic control policies. The spread of the genital herpes epidemic and the potential impact of HSV antiviral treatment in the immunocompetent population have been addressed by four models. HSV drug resistance to antiviral drugs is predicted to be minimal. Assuming that drug-resistant mutants are attenuated both in infectivity and reactivity, one model predicted that even after 25 years, only 5 in 10,000 individuals will shed drug-resistant virus, even if rates of usage of antivirals are high. The models show that increased usage of episodic antiviral therapy will be beneficial in reducing the herpes epidemic. Results also show that the transmission rate can be reduced by preventing infection (safer sex), reduced time spent in non-monogamous relationships or the advent of effective therapeutic HSV vaccines. One model has indicated that suppressive therapy will have only a minimal impact on HSV prevalence; however, the results of this modelling study are limited as it assumed that suppressive therapy would only be given to incident infections. More recent research using a model based upon virological core groups (and treating both incident and prevalent infections) shows that suppressive therapy could cause a substantial reduction in HSV-2 incidence rates. Current modelling is also focused on modelling how HSV-2 antiviral treatment will impact the HIV epidemic.


Wild-type and central DNA flap defective HIV-1 lentiviral vector genomes: intracellular visualization at ultrastructural resolution levels.
HIV-1 and other lentiviruses have the unique ability among retroviruses to efficiently replicate in non-dividing cells as a result of the active nuclear import of their DNA genome across an interphasic nuclear membrane. Previous work has shown that a three-stranded DNA structure synthesized during HIV-1 reverse transcription, called the central DNA flap, acts as a cis-determinant of HIV-1 genome nuclear import. Concordantly, DNA Flap re-insertion in lentiviral-derived gene therapy vectors stimulates gene transfer efficiencies and complements the level of nuclear import to wild-type levels quantitatively indistinguishable from wild-type virus in all cell types and tissues examined so far. In order to define the precise nature of the replicative defect of DNA flap mutant viruses, we carried out in situ DNA hybridization experiments with electron microscopy to determine the subcellular localization of DNA flap mutant and wild-type HIV-1 genomes. We found that Flap defective DNA genomes accumulate at the cytoplasmic face of the nuclear membrane with no overlap across the nuclear membrane, whereas wild-type genomes localize throughout the nuclear compartment. These data provide an unequivocal confirmation of the role of the DNA flap in HIV-1 nuclear import and further establish that the DNA flap controls a step that immediately precedes translocation through the nuclear pore. Further, the widespread distribution of wild-type genomes within the open chromatin confirms the recent genome-wide mapping of HIV-1 cDNA integration sites and points to an as-yet poorly understood step of intranuclear transport of HIV-1 pre-integration complexes.


Drosophila heat shock response requires the JNK pathway and phosphorylation of mixed lineage kinase at a conserved serine-proline motif.
Defining context specific requirements for proteins and pathways is a major challenge in the study of signal transduction. For example, the stress-activated protein kinase (SAPK) pathways are comprised of families of closely related transducers that are activated in a variety of tissues and contexts during development and organismal homeostasis. Consequently, redundant and pleiotropic effects have hampered a complete understanding of the individual contributions of transducers in distinct contexts. Here, we report on the function of a context-specific regulatory phosphorylation site, PXSP, in the Drosophila mixed lineage kinase protein, Slpr, a mitogen-activated protein kinase kinase kinase (MAP3K) in the Jun Kinase (JNK) pathway. Genetic analysis of the function of non-phosphorylatable (PXAP) and phosphomimetic mutant (PXEP) Slpr transgenes in several distinct contexts revealed minimal effects in JNK-dependent tissue closure processes but differential requirements in heat stress response. In particular, PXAP expression resulted in sensitivity of adults to sustained heat shock, like p38 and JNK pathway mutants. In contrast, PXEP overexpression conferred some resistance. Indeed, phosphorylation of the PXSP motif is enriched under heat shock conditions and requires in part, the p38 kinases for the enrichment. These data suggest that coordination of signaling between p38 and Slpr serves to maintain JNK signaling during heat stress. In sum, we demonstrate a novel role for JNK signaling in the heat shock response in flies and identify a posttranslational modification on Slpr, at a conserved site among MAP3K mixed lineage kinase family members, which bolsters stress resistance with negligible effects on JNK-dependent developmental processes.


Educational attainment and nutrient consumption patterns: the Minnesota Heart Survey.
To explore whether dietary patterns differ according to educational attainment, we examined the associations of nutrient intake with educational level in a sample of 825 male and 893 female Twin Cities (Minneapolis and St. Paul) residents aged 25 to 74 years who were surveyed between 1980 and 1982. Of particular interest was the pattern of macronutrient consumption, since the proportion of fat and type of fat in the diet is thought to be related to the development of chronic disease. Estimates of nutrient intake were based on 24-hour dietary recalls, while information on education was obtained through interview. Educational attainment was positively associated with nutrient consumption patterns that may decrease risk of chronic disease for both men and women. For women, the proportion of dietary fat was less (p = .03) and carbohydrate greater (p = .004) with increasing education. For men, the Keys dietary score decreased as education increased (p = .05) (high scores indicate greater blood cholesterol-raising effect of the diet). Inverse associations of average total serum cholesterol level with education in both men (p = .02) and women (p = .06) support these dietary associations. These results suggest that educational attainment is related positively to eating patterns that may carry a decreased risk of cardiovascular and other chronic disease. Whether or not this association is causal, it would appear to point to a need for more effective nutrition education strategies for those with less education.


The bimusical brain is not two monomusical brains in one: evidence from musical affective processing.
Complex auditory exposures in ambient environments include systems of not only linguistic but also musical sounds. Because musical exposure is often passive, consisting of listening rather than performing, examining listeners without formal musical training allows for the investigation of the effects of passive exposure on our nervous system without active use. Additionally, studying listeners who have exposure to more than one musical system allows for an evaluation of how the brain acquires multiple symbolic and communicative systems. In the present fMRI study, listeners who had been exposed to Western-only (monomusicals) and both Indian and Western musical systems (bimusicals) since childhood and did not have significant formal musical training made tension judgments on Western and Indian music. Significant group by music interactions in temporal and limbic regions were found, with effects predominantly driven by between-music differences in temporal regions in the monomusicals and by between-music differences in limbic regions in the bimusicals. Effective connectivity analysis of this network via structural equation modeling (SEM) showed significant path differences across groups and music conditions, most notably a higher degree of connectivity and larger differentiation between the music conditions within the bimusicals. SEM was also used to examine the relationships among the degree of music exposure, affective responses, and activation in various brain regions. Results revealed a more complex behavioral-neural relationship in the bimusicals, suggesting that affective responses in this group are shaped by multiple behavioral and neural factors. These three lines of evidence suggest a clear differentiation of the effects of the exposure of one versus multiple musical systems.


Associations of alcohol, nicotine, cannabis, and drug use/dependence with educational attainment: evidence from cotwin-control analyses.
Although substance use is associated with reduced educational attainment, this association may be owing to common risk factors such as socioeconomic disadvantage. We tested whether alcohol, nicotine, and illicit drug use and dependence were associated with lifetime educational attainment after controlling for familial background characteristics. Data were from a 1987 questionnaire and a 1992 telephone diagnostic interview of 6,242 male twins (n = 3,121 pairs; mean age = 41.9 years in 1992) who served in the U.S. military during the Vietnam era and therefore, were eligible for educational benefits after military service. Reduced educational attainment (<16 years) was examined in twin pairs discordant for substance use history. Substance use and dependence risk factors assessed were early alcohol and cannabis use, daily nicotine use, lifetime cannabis use, and alcohol, nicotine, cannabis, and any illicit drug dependence. Three significant differences were observed between at-risk twins and their cotwins: Compared to their low-risk cotwins, likelihood of completing <16 years of education was significantly increased for the following: (i) twins who used alcohol before age 18 (adjusted OR = 1.44; 95% CI: 1.02 to 2.05), (ii) twins with a lifetime alcohol dependence diagnosis (adjusted OR = 1.76; 95% CI: 1.27 to 2.44), and (iii) twins who had used nicotine daily for 30 or more days (adjusted OR = 2.54, 95% CI: 1.55 to 4.17). However, no differences in education were observed among twin pairs discordant for cannabis initiation, early cannabis use, or cannabis, nicotine, or any illicit drug dependence. Even in a veteran population with access to military educational benefits, early alcohol use, alcohol dependence, and daily nicotine use remained significantly associated with years of education after controlling for shared familial contributions to educational attainment. The association between other substances and educational attainment was explained by familial factors common to these substance use phenotypes and adult educational attainment.


Aire-expressing thymic medullary epithelial cells originate from β5t-expressing progenitor cells.
The thymus provides multiple microenvironments that are essential for the development and repertoire selection of T lymphocytes. The thymic cortex induces the generation and positive selection of T lymphocytes, whereas the thymic medulla establishes self-tolerance among the positively selected T lymphocytes. Cortical thymic epithelial cells (cTECs) and medullary TECs (mTECs) constitute the major stromal cells that structurally form and functionally characterize the cortex and the medulla, respectively. cTECs and mTECs are both derived from the endodermal epithelium of the third pharyngeal pouch. However, the molecular and cellular characteristics of the progenitor cells for the distinct TEC lineages are unclear. Here we report the preparation and characterization of mice that express the recombinase Cre instead of β5t, a proteasome subunit that is abundant in cTECs and not detected in other cell types, including mTECs. By crossing β5t-Cre knock-in mice with loxP-dependent GFP reporter mice, we found that β5t-Cre-mediated recombination occurs specifically in TECs but not in any other cell types in the mouse. Surprisingly, in addition to cTECs, β5t-Cre-loxP-mediated GFP expression was detected in almost all mTECs. These results indicate that the majority of mTECs, including autoimmune regulator-expressing mTECs, are derived from β5t-expressing progenitor cells.


AA16, a new lytic polysaccharide monooxygenase family identified in fungal secretomes.
Lignocellulosic biomass is considered as a promising alternative to fossil resources for the production of fuels, materials and chemicals. Efficient enzymatic systems are needed to degrade the plant cell wall and overcome its recalcitrance. A widely used producer of cellulolytic cocktails is the ascomycete Trichoderma reesei, but this organism secretes a limited set of enzymes. To improve the saccharification yields, one strategy is to upgrade the T. reesei enzyme cocktail with enzymes produced by other biomass-degrading filamentous fungi isolated from biodiversity. In this study, the enzymatic cocktails secreted by five strains from the genus Aspergillus (Aspergillus japonicus strains BRFM 405, 1487, 1489, 1490 and Aspergillus niger strain BRFM 430) were tested for their ability to boost a T. reesei reference cocktail for the saccharification of pretreated biomass. Proteomic analysis of fungal secretomes that significantly improved biomass degradation showed that the presence of proteins belonging to a putative LPMO family previously identified by genome analysis and awaiting experimental demonstration of activity. Members of this novel LPMO family, named AA16, are encountered in fungi and oomycetes with life styles oriented toward interactions with plant biomass. One AA16 protein from Aspergillus aculeatus (AaAA16) was produced to high level in Pichia pastoris. LPMO-type enzyme activity was demonstrated on cellulose with oxidative cleavage at the C1 position of the glucose unit. AaAA16 LPMO was found to significantly improve the activity of T. reesei CBHI on cellulosic substrates. Although Aspergillus spp. has been investigated for decades for their CAZymes diversity, we identified members of a new fungal LPMO family using secretomics and functional assays. Properties of the founding member of the AA16 family characterized herein could be of interest for use in biorefineries.


Contribution of lumbrical muscle activity to the paradoxical extension phenomenon induced by injuries to the finger flexor tendons.
The "Extensor habitus" phenomenon occurs in finger flexor tendon injuries and consists of a paradoxical extension of the interphalangeal joints after an attempt to flex the finger. The mechanism of extension is considered to be a contraction of the flexor digitorum profundus that is then transmitted via the lumbrical muscle structure to the extensor expansion. Using electromyography, we recorded the lumbrical muscle activity during the paradoxical extension phenomenon to determine whether the lumbrical muscle contributed to this event. Two patterns of electromyographical activity of the lumbrical muscle were observed. Group I (6 fingers) displayed electrical activities in the lumbrical muscle during flexion tasks, while group II (12 fingers) did not. In group I, the lesions were mainly located in zone V, and the response to range of motion exercises was satisfactory. In group II, nearly all of the lesion were located in zone II, and half of the cases required additional surgical interventions. Group II appeared to exhibit the "Extensor habitus" phenomenon, while group I exhibited an "Extensor habitus-like phenomenon." To distinguish between these two phenomena, an electromyographical examination of the lumbrical muscle must be performed.


Dihydrochalcones: Implication in resistance to oxidative stress and bioactivities against advanced glycation end-products and vasoconstriction.
Flavonoids are a group of polyphenol compounds with known antioxidant activities. Among them, dihydrochalcones are mainly found in apple leaves (Malus domestica). Glycosylated dihydrochalcones were previously found in large amounts in leaves of two genotypes of Malus with contrasting resistance to fire blight, a bacterial disease caused by Erwinia amylovora. In the present study we demonstrate that soluble polyphenol patterns comprised phloridzin alone or in combination with two additional dihydrochalcones, identified as sieboldin and trilobatin. Presence of sieboldin in young leaves correlated well with a high 1,1-diphenyl-2-picrylhydrazyl (DPPH) free radical scavenging activity. Moreover, these leaves displayed enhanced tolerance to paraquat, a photooxidative-stress generating herbicide. Interestingly, phloridzin had a high activity in the oxygen radical absorbance capacity (ORAC) assay, but its presence alone in leaves did not correlate with tolerance to paraquat. In order to further characterise the activity of these compounds, we tested their ability to prevent oxidative-dependent formation of advanced glycation end-products (AGEs) and phenylephrine-induced contraction of isolated rat mesenteric arteries. The antioxidant capacity of sieboldin was clearly demonstrated by showing that this compound (i) prevented vasoconstriction and (ii) inhibited AGEs formation. Both assays provided interesting information concerning a potential use of sieboldin as a therapeutic. Hence, our results strongly argue for a bioactivity of dihydrochalcones as functional antioxidants in the resistance of Malus leaves to oxidative stress. In addition, we demonstrate for the first time that sieboldin is a powerful multipotent antioxidant, effective in preventing physiopathological processes. Further work should aim at demonstrating the potential use of this compound as a therapeutic in treating free radical-involving diseases.


Monitoring tissue oxygen heterogeneities and their influence on optical glucose measurements in an animal model.
The purpose of this study was to characterize the heterogeneity of oxygen partial pressure in different adipose tissue zones and to assess the possibility of compensating these heterogeneities during optical glucose measurements. In this proof of concept study, the heterogeneity of oxygen partial pressure was determined in the adipose tissue of a pig by using 48 oxygen sensors in 3 zones of the abdominal region at two different blood oxygen levels. Sensor oxygen values correlated well with reference blood oxygen values and we identified heterogeneities in oxygen partial pressure among the defined zones of the abdominal region. Significant differences in the mean oxygen partial pressure were found when comparing the three abdominal zones but no significant differences were found when comparing two sensors located in close proximity (on one cannula). The low heterogeneity on one cannula allows the compensation of physiological oxygen variations for optical glucose measurements by using an additional oxygen sensor in close proximity to the glucose sensor. In addition, this setup can be used to continuously monitor tissue oxygenation e.g. in patients with adipose tissue dysfunction or serve limb ischemia.


Enhanced reactivity and selectivity of asymmetric oxa-Michael addition of 2'-hydroxychalcones in carbon confined spaces.
Carbon nanotubes (CNTs) are employed as nanoscale reaction vessels for the asymmetric oxa-Michael addition of 2'-hydroxychalcones. A systematic comparison of the catalytic activities of chiral phosphoric acid treated ferrite nanoparticles (chiral ferrites) has been studied for the synthesis of flavanones. Higher reactivity and selectivity with switching of enantiomers were observed when these chiral ferrites are inside the CNT channel.


Use of activity-based probes to develop high throughput screening assays that can be performed in complex cell extracts.
High throughput screening (HTS) is one of the primary tools used to identify novel enzyme inhibitors. However, its applicability is generally restricted to targets that can either be expressed recombinantly or purified in large quantities. Here, we described a method to use activity-based probes (ABPs) to identify substrates that are sufficiently selective to allow HTS in complex biological samples. Because ABPs label their target enzymes through the formation of a permanent covalent bond, we can correlate labeling of target enzymes in a complex mixture with inhibition of turnover of a substrate in that same mixture. Thus, substrate specificity can be determined and substrates with sufficiently high selectivity for HTS can be identified. In this study, we demonstrate this method by using an ABP for dipeptidyl aminopeptidases to identify (Pro-Arg)2-Rhodamine as a specific substrate for DPAP1 in Plasmodium falciparum lysates and Cathepsin C in rat liver extracts. We then used this substrate to develop highly sensitive HTS assays (Z'>0.8) that are suitable for use in screening large collections of small molecules (i.e >300,000) for inhibitors of these proteases. Finally, we demonstrate that it is possible to use broad-spectrum ABPs to identify target-specific substrates. We believe that this approach will have value for many enzymatic systems where access to large amounts of active enzyme is problematic.


Computational analysis of candidate disease genes and variants for salt-sensitive hypertension in indigenous Southern Africans.
Multiple factors underlie susceptibility to essential hypertension, including a significant genetic and ethnic component, and environmental effects. Blood pressure response of hypertensive individuals to salt is heterogeneous, but salt sensitivity appears more prevalent in people of indigenous African origin. The underlying genetics of salt-sensitive hypertension, however, are poorly understood. In this study, computational methods including text- and data-mining have been used to select and prioritize candidate aetiological genes for salt-sensitive hypertension. Additionally, we have compared allele frequencies and copy number variation for single nucleotide polymorphisms in candidate genes between indigenous Southern African and Caucasian populations, with the aim of identifying candidate genes with significant variability between the population groups: identifying genetic variability between population groups can exploit ethnic differences in disease prevalence to aid with prioritisation of good candidate genes. Our top-ranking candidate genes include parathyroid hormone precursor (PTH) and type-1 angiotensin II receptor (AGTR1). We propose that the candidate genes identified in this study warrant further investigation as potential aetiological genes for salt-sensitive hypertension.


Low voltage tunable liquid crystal lens.
In this Letter, we report a method to prepare a liquid crystal (LC) lens based on a variable pretilt angle of a photoalignment (PA) layer. With precise control of the irradiance, such PA provides good control on the pretilt angle that can be tuned in the range of 1°-89°. Therefore, the precise control of the irradiance generates a spatially varying pretilt angle with any desired pretilt angle profile. Thus, the tunable LC lenses have been fabricated and characterized. With low voltage operations, easy fabrication, and relatively high switching speed, such lenses can be applied in many modern optical and photonic devices.


Prospects for a new, safer and more effective TB vaccine.
Tuberculosis in infants and young children remains an all too common cause of morbidity and mortality in high burden countries, despite the fact that the majority of these children receive vaccination with BCG in infancy. BCG confers incomplete and variable protection against pulmonary tuberculosis [PTB] and is unsafe in HIV positive persons. Newer TB vaccines, which, it is hoped, will either replace or complement BCG are being developed and a number of these have reached the stage of clinical trials, with two booster vaccines going into Phase IIB trials in 2009. Prospects for at least one new licensed TB vaccine within the next 5-10 years appear reasonable. This article explores some of the issues around the development of new vaccines against TB and details the leading candidates.


A randomised controlled trial of the effect of music therapy and verbal relaxation on chemotherapy-induced anxiety.
To determine the effect of music therapy and verbal relaxation on state anxiety and anxiety-induced physiological manifestations among patients with cancer before and after chemotherapy. Cancer and its treatment provoke a series of changes in the emotional sphere of the patient's anxiety. Music therapy and verbal relaxation had reported the anxiety reduction effect on patients with cancer receiving chemotherapy. Few studies have been undertaken comparing music therapy and verbal relaxation in differentiating high-normal state anxiety subsample. A randomised controlled trial and permuted block design were used. Outpatient chemotherapy clinic operated by a University medical centre in southern Taiwan. Ninety-eight patients were randomised into three groups: the music therapy group received one-hour single music session; the verbal relaxation group received 30 minutes of guided relaxation; the control group received usual care. Spielberger State-Trait Anxiety Instrument, Emotional Visual Analog Scale, three biobehavioural indicators: skin temperature, heart rate and consciousness level were measured during and after chemotherapy. Music therapy had a greater positive effect on postchemotherapy anxiety than verbal relaxation and control groups and a significantly increase in skin temperature. Patients with high state anxiety receiving music therapy had a greater drop in postchemotherapy anxiety than did the normal state anxiety subsample. Both music and verbal relaxation therapy are effective in reducing chemotherapy-induced anxiety. Thirty minutes of intervention initiates anxiety reduction. Patients with high state anxiety receiving chemotherapy obtain the most benefit from music or verbal relaxation. Prior to chemotherapy, patients with high state anxiety must be sorted from all patients as they are more responsive to interventions. Oncology nurses can offer music and verbal relaxation as adjuvant interventions to reduce chemotherapy-induced anxiety and enhance the quality of care.


Integrating the internal and external training loads in soccer.
This study aimed to assess the relationships of fitness in soccer players with a novel integration of internal and external training load (TL). Ten amateur soccer players performed a lactate threshold (LT) test followed by a soccer simulation (Ball-Sport Endurance and Sprint Test [BEAST90mod]). The results from the LT test were used to determine velocity at lactate threshold (vLT), velocity at onset of blood lactate accumulation (vOBLA), maximal oxygen uptake (VO2max), and the heart rate-blood lactate profile for calculation of internal TL (individualized training impulse, or iTRIMP). The total distance (TD) and high intensity distance (HID) covered during the BEAST90mod were measured using GPS technology that allowed measurement of performance and external TL. The internal TL was divided by the external TL to form TD:iTRIMP and HID:iTRIMP ratios. Correlation analyses assessed the relationships between fitness measures and the ratios to performance in the BEAST90mod. vLT, vOBLA, and VO2max showed no significant relationship to TD or HID. HID:iTRIMP significantly correlated with vOBLA (r = .65, P = .04; large), and TD:iTRIMP showed a significant correlation with vLT (r = .69, P = .03; large). The results suggest that the integrated use of ratios may help in the assessment of fitness, as performance alone showed no significant relationships with fitness.


Polarisation effects on the solvation properties of alcohols.
Alcohol solvents are significantly more polar than expected based on the measured H-bonding properties of monomeric alcohols in dilute solution. Self-association of alcohols leads to formation of cyclic aggregates and linear polymeric chains that have a different polarity from the alcohol monomer. Cyclic aggregates are less polar than the monomer, and the chain ends of linear polymers are more polar. The solvation properties of alcohols therefore depend on the interplay of these self-association equilibria and the equilibria involving interactions with solutes. Twenty-one different molecular recognition probes of varying polarity were used to probe the solvation properties of alkane-alcohol mixtures across a wide range of different solvent compositions. The results allow dissection of the complex equilibria present in these systems. Formation of a H-bond between two alcohol molecules leads to polarisation of the hydroxyl groups, resulting in an increase in binding affinity for subsequent interactions with the unbound donor and acceptor sites. The H-bond donor parameter (α) for these sites increases from 2.7 to 3.5, and the H-bond acceptor parameter (β) increases from 5.3 to 6.9. Polarisation is a short range effect limited to the first H-bond in a chain, and formation of subsequent H-bonds in longer chains does not further increase the polarity of chain ends. H-bond donor sites involved in a H-bond are unavailable for further interactions, because the formation of a bifurcated three-centre H-bond is three orders of magnitude less favourable than formation of a conventional two-centre H-bond. These findings are reproduced by quantum chemical calculations of the molecular electrostatic potential surfaces of alcohol aggregates. Thus, the overall solvation properties of alcohols depend on the speciation of different aggregates, the polarities of these species and the polarities of the solutes. At low alcohol concentrations, polar solutes are solvated by alcohol monomers, and at higher alcohol concentrations, solutes are solvated by the more polar chain ends of linear polymers. The less polar cyclic aggregates are less important for interactions with solutes. Similar behavior was found for ten different alcohol solvents. Tertiary alcohols are marginally less polar solvents than primary alcohols, due to steric interactions that destabilises the formation of polymeric aggregates leading to lower concentrations of polar chain ends. One alcohol with an electron-withdrawing substituent was studied, and this solvent showed slightly different behavior, because the H-bond donor and acceptor properties are different.


TRMT5 Mutations Cause a Defect in Post-transcriptional Modification of Mitochondrial tRNA Associated with Multiple Respiratory-Chain Deficiencies.
Deficiencies in respiratory-chain complexes lead to a variety of clinical phenotypes resulting from inadequate energy production by the mitochondrial oxidative phosphorylation system. Defective expression of mtDNA-encoded genes, caused by mutations in either the mitochondrial or nuclear genome, represents a rapidly growing group of human disorders. By whole-exome sequencing, we identified two unrelated individuals carrying compound heterozygous variants in TRMT5 (tRNA methyltransferase 5). TRMT5 encodes a mitochondrial protein with strong homology to members of the class I-like methyltransferase superfamily. Both affected individuals presented with lactic acidosis and evidence of multiple mitochondrial respiratory-chain-complex deficiencies in skeletal muscle, although the clinical presentation of the two affected subjects was remarkably different; one presented in childhood with failure to thrive and hypertrophic cardiomyopathy, and the other was an adult with a life-long history of exercise intolerance. Mutations in TRMT5 were associated with the hypomodification of a guanosine residue at position 37 (G37) of mitochondrial tRNA; this hypomodification was particularly prominent in skeletal muscle. Deficiency of the G37 modification was also detected in human cells subjected to TRMT5 RNAi. The pathogenicity of the detected variants was further confirmed in a heterologous yeast model and by the rescue of the molecular phenotype after re-expression of wild-type TRMT5 cDNA in cells derived from the affected individuals. Our study highlights the importance of post-transcriptional modification of mitochondrial tRNAs for faithful mitochondrial function.


Furanosic forms of sugars: conformational equilibrium of methyl β-d-ribofuranoside.
The investigation of an isolated ribofuranose unit in the gas phase reveals the intrinsic conformational landscape of the biologically active sugar form. We report the rotational spectra of two conformers of methyl β-d-ribofuranoside in a supersonic jet expansion. Both conformers adopt a near twisted ((3)T2) ring conformation with the methoxy and hydroxymethyl substituents involved in various intramolecular hydrogen bonds.


Modular, Bioorthogonal Strategy for the Controlled Loading of Cargo into a Protein Nanocage.
Virus capsids, i.e., viruses devoid of their genetic material, are suitable nanocarriers for biomedical applications such as drug delivery and diagnostic imaging. For this purpose, the reliable encapsulation of cargo in such a protein nanocage is crucial, which can be accomplished by the covalent attachment of the compounds of interest to the protein domains positioned at the interior of the cage. This approach is particularly valid for the capsid proteins of the cowpea chlorotic mottle virus (CCMV), which have their N-termini located at the inside of the capsid structure. Here, we examined several site-selective modification methods for covalent attachment and encapsulation of cargo at the N-terminus of the CCMV protein. Initially, we explored approaches to introduce an N-terminal azide functionality, which would allow the subsequent bioorthogonal modification with a strained alkyne to attach the desired cargo. As these methods showed compatibility issues with the CCMV capsid proteins, a strategy based on 2-pyridinecarboxaldehydes for site-specific N-terminal protein modification was employed. This method allowed the successful modification of the proteins, and was applied for the introduction of a bioorthogonal vinylboronic acid moiety. In a subsequent reaction, the proteins could be modified further with a fluorophore using the tetrazine ligation. The application of capsid assembly conditions on the functionalized proteins led to successful particle formation, showing the potential of this covalent encapsulation strategy.


Pharmacokinetic interactions between primaquine and chloroquine.
Chloroquine combined with primaquine has been the standard radical curative regimen for Plasmodium vivax and Plasmodium ovale malaria for over half a century. In an open-label crossover pharmacokinetic study, 16 healthy volunteers (4 males and 12 females) aged 20 to 47 years were randomized into two groups of three sequential hospital admissions to receive a single oral dose of 30 mg (base) primaquine, 600 mg (base) chloroquine, and the two drugs together. The coadministration of the two drugs did not affect chloroquine or desethylchloroquine pharmacokinetics but increased plasma primaquine concentrations significantly (P ≤ 0.005); the geometric mean (90% confidence interval [CI]) increases were 63% (47 to 81%) in maximum concentration and 24% (13 to 35%) in total exposure. There were also corresponding increases in plasma carboxyprimaquine concentrations (P ≤ 0.020). There were no significant electrocardiographic changes following primaquine administration, but there was slight corrected QT (QTc) (Fridericia) interval lengthening following chloroquine administration (median [range] = 6.32 [-1.45 to 12.3] ms; P < 0.001), which was not affected by the addition of primaquine (5.58 [1.74 to 11.4] ms; P = 0.642). This pharmacokinetic interaction may explain previous observations of synergy in preventing P. vivax relapse. This trial was registered at ClinicalTrials.gov under reference number NCT01218932.


Antigen-specific immunosuppression in human malaria due to Plasmodium falciparum.
Proliferative responses of T lymphocytes to antigens specific and not specific for malaria were investigated in 32 adult patients in eastern Thailand during acute infection with Plasmodium falciparum malaria and during their convalescence. Immune unresponsiveness to malarial antigen, which persisted for more than four weeks in 37.5% of the individuals, was present in all patients, irrespective of parasitemia or severity of clinical illness. Suppression of responses to nonspecific antigens was less profound and observed only in patients with moderately severe or cerebral malaria. The depressed functional responses were associated with a loss of T lymphocytes--both helper and suppressor subsets--from the peripheral blood; these responses were recovered once parasites were cleared. These results indicate that blood-stage plasmodial infections may suppress responses important for immunity to malaria and so allow the parasite to survive. They further suggest that patients acutely or even recently infected with P. falciparum may not respond as well to a malaria vaccine as would uninfected individuals.


Cathepsin B, L, and S cleave and inactivate secretory leucoprotease inhibitor.
A number of serine proteases, matrix metalloproteases, and cysteine proteases were evaluated for their ability to cleave and inactivate the antiprotease, secretory leucoprotease inhibitor (SLPI). None of the serine proteases or the matrix metalloproteases examined cleaved the SLPI protein. However, incubation with cathepsins B, L, and S resulted in the cleavage and inactivation of SLPI. All three cathepsins initially cleaved SLPI between residues Thr(67) and Tyr(68). The proteolytic cleavage of SLPI by all three cathepsins resulted in the loss of the active site of SLPI and the inactivation of SLPI anti-neutrophil elastase capacity. Cleavage and inactivation were catalytic with respect to the cathepsins, so that the majority of a 400-fold excess of SLPI was inactivated within 15 min by cathepsins L and S. Analysis of epithelial lining fluid samples from individuals with emphysema indicated the presence of cleaved SLPI in these samples whereas only intact SLPI was observed in control epithelial lining fluid samples. Active cathepsin L was shown to be present in emphysema epithelial lining fluid and inhibition of this protease prevented the cleavage of recombinant SLPI added to emphysema epithelial lining fluid. Taken together with previous data that demonstrates that cathepsin L inactivates alpha(1)-antitrypsin, these findings indicate the involvement of cathepsins in the diminution of the lung antiprotease screen possibly leading to lung destruction in emphysema.


Shedding of Viable Virus in Asymptomatic SARS-CoV-2 Carriers.
Information regarding the infectivity of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in asymptomatic carriers is scarce. In order to determine the duration of infectivity and its correlation with reverse transcription-PCR (RT-PCR) results and time since initial positive PCR test in this population, we evaluated SARS-CoV-2 cell infectivity in nasopharyngeal samples longitudinally obtained from asymptomatic carriers who disembarked from a cruise ship during a COVID-19 outbreak. Of 166 nasopharyngeal samples collected from 39 asymptomatic carriers every 48 h until two consecutive negative PCR test results were obtained, SARS-CoV-2 was successfully isolated from 9 PCR-positive samples which were obtained from 7 persons (18%; 7/39). Viable viruses were isolated predominantly within 7 days after the initial positive PCR test, except for one person who shed viable virus until day 15. The median crossing point (Cp) value of RT-PCR of culture-positive samples was 24.6 (interquartile range [IQR], 20.4 to 25.8; range, 17.9 to 30.3), and Cp values were significantly associated with isolation of viable virus (odds ratio, 0.496; 95% confidence interval [CI], 0.329 to 0.747; P value, 0.001), which was consistent with existing data for symptomatic patients. Genome sequence analysis of SARS-CoV-2 samples consecutively obtained from a person who shed viable virus for 15 days identified the emergence of two novel single nucleotide variants (C8626T transition and C18452T transition) in the sample collected on day 15, with the latter corresponding to an amino acid substitution in nonstructural protein 14. The impact of these mutations on prolonged viable-virus shedding is unclear. These findings underscore the potential role of asymptomatic carriers in transmission.IMPORTANCE A growing number of studies suggest the potential role of asymptomatic SARS-CoV-2 carriers as a major driver of the COVID-19 pandemic; however, virological assessment of asymptomatic infection has largely been limited to reverse transcription-PCR (RT-PCR), which can be persistently positive without necessarily indicating the presence of viable virus (e.g., replication-competent virus). Here, we evaluated the infectivity of asymptomatic SARS-CoV-2 carriers by detecting SARS-CoV-2-induced cytopathic effects on Vero cells using longitudinally obtained nasopharyngeal samples from asymptomatic carriers. We show that asymptomatic carriers can shed viable virus until 7 days after the initial positive PCR test, with one outlier shedding until day 15. The crossing point (Cp) value of RT-PCR was the leading predictive factor for virus viability. These findings provide additional insights into the role of asymptomatic carriers as a source of transmission and highlight the importance of universal source control measures, along with isolation policy for asymptomatic carriers.


Psychopathology and academic performance, social well-being, and social preference at school: the TRAILS study.
Psychopathology during adolescence has been associated with poor academic performance, low social well-being, and low social preference by peers at school. However, previous research has not accounted for comorbid psychopathology, informant-specific associations between psychopathology and functioning, and gender and age differences. This study addresses these limitations by examining adolescents' psychopathology and functioning at school, reported by child, parent, teacher, and peers during primary and secondary school in a large Dutch longitudinal cohort study (N = 2230). Teacher reports of psychopathology, especially regarding attention problems and withdrawn/depressed problems, followed by parent reports regarding hyperactivity, were most strongly associated with academic performance. The same held for social preference which was associated with teacher and parent ratings of withdrawn/depressed problems and hyperactivity. In contrast, social well-being was best predicted by child reports (at primary school) of affective problems. In girls, the association between ADHD problems and poor academic performance was stronger than in boys and conduct problems were more often associated with poor school functioning in general. These findings can help identify adolescents at risk for poor functioning and design interventions that effectively reduce or prevent poor school functioning.


Antihyperglycemic effect of polyphenols from Acerola (Malpighia emarginata DC.) fruit.
A crude acerola polyphenol fraction (C-AP) was prepared by subjecting an acerola extract to a C18 cartridge column, and eluting the adsorbed fraction with ethanol containing 10% of acetic acid. C-AP appeared in a previous study to have an inhibitory effect on alpha-glucosidase and particularly on maltase activities. To elucidate the antihyperglycemic effect of C-AP further, we examined the regulation by C-AP of glucose uptake in Caco-2 cell; this resulted in the inhibition of glucose uptake. We next conducted single administration tests of glucose and maltose to ICR mice to investigate whether C-AP really controlled the intestinal glucose absorption in an animal body. The results showed that C-AP significantly suppressed the plasma glucose level after administering both glucose and maltose, suggesting that C-AP had a preventive effect on hyperglycemia in the postprandial state. The mechanism for this effect is considered to have been both suppression of the intestinal glucose transport and the inhibition of alpha-glucosidase. Despite such a preventive effect, the therapeutic effect of C-AP on hyperglycemia appeared to be low from the experiment with KKAy mice.


Biochemical, biophysical, and mutational analyses of subunit interactions of the human cytomegalovirus nuclear egress complex.
Nuclear egress, the trafficking of herpesvirus nucleocapsids from the nucleus to the cytoplasm, involves two conserved viral proteins that form a complex at the nuclear envelope, referred to as the nuclear egress complex. In human cytomegalovirus, these two proteins are called UL50 and UL53. To study UL50 and UL53 in molecular detail, these proteins were expressed in bacteria and purified. To obtain highly expressed, pure proteins, it was necessary to truncate both constructs based on sequence conservation and predicted secondary structural elements. Size exclusion chromatography and analytical ultracentrifugation studies indicated that the truncated form of UL50 is a monomer in solution, that the truncated form of UL53 is a homodimer, and that, when mixed, the two proteins form a heterodimer. To identify residues of UL53 crucial for homodimerization and for heterodimerization with UL50, we constructed and expressed mutant forms of UL53 containing alanine substitutions in a predicted helix. Isothermal titration calorimetry was used to measure the binding affinities of the UL53 mutants to UL50. UL53 residues, the replacement of which reduced binding to UL50, form a surface on one face of the predicted helix. Moreover, most of the substitutions that reduce UL53-UL50 interactions also reduced homodimerization. Substitutions that reduced the interaction between UL50 and UL53 in vitro also reduced colocalization of full-length UL50 and UL53 at the nuclear rim in transfected cells. These results demonstrate direct protein-protein interactions between these proteins that are likely to be mediated by a helix, and they have implications for understanding nuclear egress and for drug discovery.


Temperance.
The kind of temperance discussed is the virtue of moderation concerning the pleasures of eating and drinking. A person lacks this moderation if either his desire for such pleasures causes conflicts with his own standards of appropriate behaviour or the standards themselves attach too much importance to the pleasures concerned. Opinions vary about the proper status of these pleasures, but people surely value them too highly if they place them above their health or the moral duty not to cause unnecessary suffering. Temperance concerns medicine because of its connection with health, ethics because it involves not only the good life for the individual but also the welfare of other creatures.


Long-term solid fuel use and risks of major eye diseases in China: A population-based cohort study of 486,532 adults.
Over 3.5 billion individuals worldwide are exposed to household air pollution from solid fuel use. There is limited evidence from cohort studies on associations of solid fuel use with risks of major eye diseases, which cause substantial disease and economic burden globally. The China Kadoorie Biobank recruited 512,715 adults aged 30 to 79 years from 10 areas across China during 2004 to 2008. Cooking frequency and primary fuel types in the 3 most recent residences were assessed by a questionnaire. During median (IQR) 10.1 (9.2 to 11.1) years of follow-up, electronic linkages to national health insurance databases identified 4,877 incident conjunctiva disorders, 13,408 cataracts, 1,583 disorders of sclera, cornea, iris, and ciliary body (DSCIC), and 1,534 cases of glaucoma. Logistic regression yielded odds ratios (ORs) for each disease associated with long-term use of solid fuels (i.e., coal or wood) compared to clean fuels (i.e., gas or electricity) for cooking, with adjustment for age at baseline, birth cohort, sex, study area, education, occupation, alcohol intake, smoking, environmental tobacco smoke, cookstove ventilation, heating fuel exposure, body mass index, prevalent diabetes, self-reported general health, and length of recall period. After excluding participants with missing or unreliable exposure data, 486,532 participants (mean baseline age 52.0 [SD 10.7] years; 59.1% women) were analysed. Overall, 71% of participants cooked regularly throughout the recall period, of whom 48% used solid fuels consistently. Compared with clean fuel users, solid fuel users had adjusted ORs of 1.32 (1.07 to 1.37, p < 0.001) for conjunctiva disorders, 1.17 (1.08 to 1.26, p < 0.001) for cataracts, 1.35 (1.10 to 1.66, p = 0.0046) for DSCIC, and 0.95 (0.76 to 1.18, p = 0.62) for glaucoma. Switching from solid to clean fuels was associated with smaller elevated risks (over long-term clean fuel users) than nonswitching, with adjusted ORs of 1.21 (1.07 to 1.37, p < 0.001), 1.05 (0.98 to 1.12, p = 0.17), and 1.21 (0.97 to 1.50, p = 0.088) for conjunctiva disorders, cataracts, and DSCIC, respectively. The adjusted ORs for the eye diseases were broadly similar in solid fuel users regardless of ventilation status. The main limitations of this study include the lack of baseline eye disease assessment, the use of self-reported cooking frequency and fuel types for exposure assessment, the risk of bias from delayed diagnosis (particularly for cataracts), and potential residual confounding from unmeasured factors (e.g., sunlight exposure). Among Chinese adults, long-term solid fuel use for cooking was associated with higher risks of not only conjunctiva disorders but also cataracts and other more severe eye diseases. Switching to clean fuels appeared to mitigate the risks, underscoring the global health importance of promoting universal access to clean fuels.


Amplification of the synovial inflammatory response through activation of mitogen-activated protein kinases and nuclear factor kappaB using ligation of CD40 on CD14+ synovial cells from patients with rheumatoid arthritis.
To determine the signal transduction pathways in CD14+ synovial cells from patients with rheumatoid arthritis (RA) after CD40 ligation, and to examine their role in amplifying synovial inflammation in affected joints. Expression of messenger RNA was analyzed using quantitative reverse transcription-polymerase chain reaction. Cytokines and chemokines were measured using enzyme-linked immunosorbent assay. Activation of kinases was detected using Western blotting. Nuclear translocation of NF-kappaB was examined using immunohistochemistry. CD14+ synovial cells were enriched using magnetic cell sorting. Fibroblast-like synoviocytes (FLS) were obtained by passaging primary synovial cell culture. Stimulation of CD14+ synovial cells from RA patients by recombinant soluble CD154 (rsCD154) significantly induced expression of tumor necrosis factor alpha (TNFalpha), interleukin-1alpha (IL-1alpha), and IL-1beta. CD14+ RA synovial cells stimulated with rsCD154 plus interferon-gamma (IFNgamma) induced significantly higher production of IL-6, IL-8, and monocyte chemoattractant protein 1 by FLS compared with unstimulated CD14+ synovial cells, through TNFalpha-, IL-1alpha-, and IL-1beta-mediated pathways. Stimulation with rsCD154 plus IFNgamma induced the activation of ERK-1/2, p38 MAPK, and NF-kappaB. Specific inhibitors of MAPK/ERK-1/2 kinases and p38 MAPK significantly reduced the production of TNFalpha and IL-1beta by rsCD154 plus IFNgamma-stimulated CD14+ synovial cells, and also inhibited production of these cytokines by freshly isolated synovial cells from RA patients. These data indicate that the CD40-CD154 interaction activates the ERK, p38, and NF-kappaB pathways in CD14+ synovial cells from RA patients to produce TNFalpha, IL-1alpha, and IL-1beta, which in turn amplifies inflammatory responses by stimulating FLS. Inhibition of the CD40-CD154 interaction or its signal transduction pathways would be a strong and efficient strategy for the management of synovial inflammation in RA.