PI3KC2α, a class II PI3K, is required for dynamin-independent internalization pathways.
Increasing evidence indicates that cellular uptake of several molecules can occur independently of functional dynamin, but the molecular players that regulate dynamin-independent endocytosis and the subsequent trafficking steps are still largely unknown. A survival-based short-hairpin (sh) RNA screen using a cell line expressing a diphtheria toxin receptor (DTR, officially known as HBEGF) anchored to GPI (DTR-GPI), which internalizes diphtheria toxin (DT, officially known as DTX) in a dynamin-independent manner, identified PI3KC2α, a class II phosphoinositide 3-kinase (PI3K), as a specific regulator of dynamin-independent DT internalization. We found that the internalization of several proteins that enter the cell through dynamin-independent pathways led to a relocalization of PI3KC2α to cargo-positive vesicles. Furthermore, downregulation of PI3KC2α impaired internalization of CD59 as well as fluid-phase endocytosis. Our data suggest a general role for PI3KC2α in regulating physiologically relevant dynamin-independent internalization pathways by recruiting early endosome antigen 1 (EEA1) to vesicular compartments, a step required for the intracellular trafficking of vesicles generated by dynamin-independent endocytic pathways.


Short-chain fatty acid-initiated cell cycle arrest and apoptosis of colonic epithelial cells is linked to mitochondrial function.
Butyrate, a short-chain fatty acid produced during microbial fermentation of fiber, induces growth arrest, differentiation, and apoptosis of colonic epithelial cells in vitro, and our prior work has shown that this induction is tightly linked to mitochondrial activity. Here we demonstrate that 12 h following induction, SW620 human colonic carcinoma cells accumulate simultaneously in G0-G1 and G2-M of the cell cycle. Four h later, during this G0-G1 to G2-M arrest, cells begin to undergo apoptosis. Using a series of unrelated agents that modulate mitochondrial functions, we demonstrate that mitochondrial electron transport and membrane potential are critical in initiation of this butyrate-mediated growth arrest and apoptosis. Colonic tumorigenesis is characterized by abnormalities in proliferation, apoptosis, and mitochondrial activities. Thus, butyrate may reduce risk for colon cancer by inducing a pathway that enhances mitochondrial function, ultimately resulting in initiation of growth arrest and apoptosis of colonic epithelial cells.


Cancer mortality rates in Appalachia: descriptive epidemiology and an approach to explaining differences in outcomes.
Cancer is a leading cause of death in the Appalachian region of the United States. Existing studies compare regional mortality rates to those of the entire nation. We compare cancer mortality rates in Appalachia to those of the nation, with additional comparisons of Appalachian and non-Appalachian counties within the 13 states that contain the Appalachian region. Lung/bronchus, colorectal, female breast and cervical cancers, as well as all cancers combined, are included in analysis. Linear regression is used to identify independent associations between ecological socioeconomic and demographic variables and county-level cancer mortality outcomes. There is a pattern of high cancer mortality rates in the 13 states containing Appalachia compared to the rest of the United States. Mortality rate differences exist between Appalachian and non-Appalachian counties within the 13 states, but these are not consistent. Lung cancer is a major problem in Appalachia; most Appalachian counties within the 13 states have significantly higher mortality rates than in-state, non-Appalachian counterparts. Mortality rates from all cancers combined also appear to be worse overall within Appalachia, but part of this disparity is likely driven by lung cancer. Education and income are generally associated with cancer mortality, but differences in the strength and direction of these associations exist depending on location and cancer type. Improving high school graduation rates in Appalachia could result in a meaningful long term reduction in lung cancer mortality. The relative importance of household income level to cancer outcomes may be greater outside the Appalachian regions within these states.


Anomalous heat transfer modes of nanofluids: a review based on statistical analysis.
This paper contains the results of a concise statistical review analysis of a large amount of publications regarding the anomalous heat transfer modes of nanofluids. The application of nanofluids as coolants is a novel practise with no established physical foundations explaining the observed anomalous heat transfer. As a consequence, traditional methods of performing a literature review may not be adequate in presenting objectively the results representing the bulk of the available literature. The current literature review analysis aims to resolve the problems faced by researchers in the past by employing an unbiased statistical analysis to present and reveal the current trends and general belief of the scientific community regarding the anomalous heat transfer modes of nanofluids. The thermal performance analysis indicated that statistically there exists a variable enhancement for conduction, convection/mixed heat transfer, pool boiling heat transfer and critical heat flux modes. The most popular proposed mechanisms in the literature to explain heat transfer in nanofluids are revealed, as well as possible trends between nanofluid properties and thermal performance. The review also suggests future experimentation to provide more conclusive answers to the control mechanisms and influential parameters of heat transfer in nanofluids.


Is there an excess of significant findings in published studies of psychotherapy for depression?
Many studies have examined the efficacy of psychotherapy for major depressive disorder (MDD) but publication bias against null results may exist in this literature. However, to date, the presence of an excess of significant findings in this literature has not been explicitly tested. We used a database of 1344 articles on the psychological treatment of depression, identified through systematic search in PubMed, PsycINFO, EMBASE and the Cochrane database of randomized trials. From these we identified 149 studies eligible for inclusion that provided 212 comparisons. We tested for an excess of significant findings using the method developed by Ioannidis and Trikalinos (2007), and compared the distribution of p values in this literature with the distribution in the antidepressant literature, where publication bias is known to be operating. The average statistical power to detect the effect size indicated by the meta-analysis was 49%. A total of 123 comparisons (58%) reported a statistically significant difference between treatment and control groups, but on the basis of the average power observed, we would only have expected 104 (i.e. 49%) to do so. There was therefore evidence of an excess of significance in this literature (p = 0.010). Similar results were obtained when these analyses were restricted to studies including a cognitive behavioural therapy (CBT) arm. Finally, the distribution of p values for psychotherapy studies resembled that for published antidepressant studies, where publication bias against null results has already been established. The small average size of individual psychotherapy studies is only sufficient to detect large effects. Our results indicate an excess of significant findings relative to what would be expected, given the average statistical power of studies of psychotherapy for major depression.


Size of the amino acid side chain at position 158 of cytochrome b is critical for an active cytochrome bc1 complex and for photosynthetic growth of Rhodobacter capsulatus.
The nonphotosynthetic mutant R126 of Rhodobacter capsulatus has a cytochrome (cyt) bc1 complex (EC 188.8.131.52) with a defective quinol oxidation Qz(o,p) site but a functional quinone reduction Qc(i,n) site. Genetic analyses of this mutant have indicated that a single-base-pair change has replaced Gly-158 of cyt b with Asp. In this work, Gly-158 was changed by oligonucleotide-mediated mutagenesis to several other amino acids to define its role on quinol oxidation catalyzed by the cyt bc1 complex. The effects of the mutations were analyzed by measuring the photosynthetic growth rate of mutants and the activity of their cyt bc1 complexes. The mutants overproduced the cyt bc1 complex, assembled its subunits, and incorporated its prosthetic groups as shown by immunoblotting and optical difference spectroscopy. Of 14 amino acid residues tested at position 158 of cyt b all but alanine and serine resulted in a marked decrease of cyt bc1 activity and failed to support photosynthetic growth of R. capsulatus. The photosynthesis-competent mutants, Gly-158----Ala and Gly-158----Ser, had lower cyt bc1 complex activities that were resistant to myxothiazol, but not to stigmatellin, quinol oxidation inhibitors. These findings indicated that the specific role of Gly-158 of cyt b on quinol oxidation and myxothiazol binding may be related to the small size of its side chain and are discussed in terms of the structure and function of the quinol oxidation site of the cyt bc1 complex.


Investigation of N Deficiency in Cherry Trees Using Visible and Near-Infrared Spectra Part of the Spectrum in Field Condition.
This objective of the study was to develop a model for the determination of N deficiency in cherry trees using a combination of visible near infrared methods and spectro-radiometric measurement. In our experimental design, cherry seedlings were grown under various N deficiency conditions in nutrient-controlled containers. The reflectance values of plant leaves were measured using a spectro-radiometer. Plant leaves samples were simultaneously collected. Their nutrient contents were determined in the laboratory. Afterwards, we performed a statistical comparison of the reflectance values. Sample analysis results established the significant wavelengths. Moreover, we received accurate regression models for predicting N deficiency in cherry leaves that were grown in nutrient solutions. Next, we verified the model validity by measuring the reflectance of the leaves collected from cherry orchards at various locations using a spectroradiometer. Nutrient deficiencies were calculated using the developed model, and then, the predicted and measured data were compared to evaluate model validity. From these results, we determined the wavelengths that yielded the most accurate results for N prediction, selected from the blue and green regions of the spectrum. We established that for N prediction in cherry trees, the simplest model can be created using 560 and 570 nm wavelengths. However, the evaluated model can be applicable only under certain conditions. We concluded that in order to develop a prediction method with sufficient application capacity, as well as the ability to assess nutritional and physiological characteristics, the ecology condition of the plant should be properly considered based on the model.


Patient perspective: reasons and methods for measuring fatigue in rheumatoid arthritis.
The experience of fatigue has been reported by a large proportion of people with rheumatoid arthritis (RA), and it is often the most important problem for individual patients. A systematic analysis of patient focus group discussions revealed 3 overarching themes: fatigue is overwhelming and different from normal tiredness; it permeates every sphere of life; and self-management is variable, but professional support is rare. A systematic search for articles measuring fatigue discovered 23 scales, 6 of which have sufficient evidence of validity to pass the OMERACT filter. Some preliminary data indicate that fatigue measurement is sensitive to change induced by some interventions in RA. This issue, and the question of whether measuring fatigue adds additional information to measurements made using the current core set of measures, will be addressed at OMERACT 8.


Results of metal-on-metal hybrid hip resurfacing for Crowe type-I and II developmental dysplasia.
Modern hip resurfacing implants may increase stability and preserve more bone than conventional total hip arthroplasty. The purpose of this retrospective study was to analyze the mid-term results in a consecutive series of middle-aged patients with developmental dysplasia of the hip treated with hybrid resurfacing joint arthroplasty. Metal-on-metal hip resurfacing was performed in fifty-one patients (fifty-nine hips), forty-two of whom were female and nine of whom were male. The average age at the time of surgery was 43.7 years. Radiographic and clinical data were collected at six weeks, at three months, and at yearly follow-up visits. Seven hips had Crowe type-II developmental dysplasia of the hip and fifty-two had type-I. The follow-up period ranged from 4.2 to 9.5 years (average, 6.0 years). Initial stability was achieved in all but three hips. The clinical outcomes, as rated with the University of California at Los Angeles (UCLA) hip score, improved significantly compared with the preoperative ratings. On the average, the pain rating improved from 3.2 to 9.3 points; the score for walking, from 6.0 to 9.7 points; the score for function, from 5.7 to 9.6 points; and the score for activity, from 4.6 to 7.3 points (all p = 0.0001). The mean Short Form-12 (SF-12) mental score increased from 46.6 to 53.5 points, and the mean SF-12 physical score increased from 31.7 to 51.4 points (both p < 0.0001). The mean postoperative Harris hip score was 92.5 points. On the average, the range of flexion improved from 106 degrees to 129.6 degrees ; the abduction-adduction arc, from 41.9 degrees to 76.9 degrees ; and the rotation arc in extension, from 32.1 degrees to 84.8 degrees (all p = 0.0001). Four patients delivered a total of six healthy babies since the time of implantation of the prosthesis. Radiographic analysis showed a decrease in the mean body weight lever arm from 118.5 mm preoperatively to 103.9 mm postoperatively (p = 0.007). There were five femoral failures requiring conversion to a total hip arthroplasty. One hip showed a radiolucency around the metaphyseal femoral stem. There were no complete acetabular radiolucencies, and all sockets remained well fixed. The mid-term results of metal-on-metal resurfacing in patients with Crowe type-I or II developmental dysplasia of the hip were disappointing with respect to the durability of the femoral component. However, the fixation of the porous-coated acetabular components without adjuvant fixation was excellent despite incomplete lateral acetabular coverage of the socket. More rigorous patient selection and especially meticulous bone preparation are essential to minimize femoral neck fractures and loosening after this procedure.


Calcium peroxide from ambient to high pressures.
Structures of calcium peroxide (CaO2) are investigated in the pressure range 0-200 GPa using the ab initio random structure searching (AIRSS) method and density functional theory (DFT) calculations. At 0 GPa, there are several CaO2 structures very close in enthalpy, with the ground-state structure dependent on the choice of exchange-correlation functional. Further stable structures for CaO2 with C2/c, I4/mcm and P21/c symmetries emerge at pressures below 40 GPa. These phases are thermodynamically stable against decomposition into CaO and O2. The stability of CaO2 with respect to decomposition increases with pressure, with peak stability occurring at the CaO B1-B2 phase transition at 65 GPa. Phonon calculations using the quasiharmonic approximation show that CaO2 is a stable oxide of calcium at mantle temperatures and pressures, highlighting a possible role for CaO2 in planetary geochemistry. We sketch the phase diagram for CaO2, and find at least five new stable phases in the pressure-temperature ranges 0 ≤ P ≤ 60 GPa, 0 ≤ T ≤ 600 K, including two new candidates for the zero-pressure ground state structure.


The HML mating-type cassette of Saccharomyces cerevisiae is regulated by two separate but functionally equivalent silencers.
Mating-type genes resident in the silent cassette HML at the left arm of chromosome III are repressed by the action of four SIR gene products, most likely mediated through two cis-acting sites located on opposite sides of the locus. We showed that deletion of either of these two cis-acting sites from the chromosome did not yield any detectable derepression of HML, while deletion of both sites yielded full expression of the locus. In addition, each of these sites was capable of exerting repression of heterologous genes inserted in their vicinity. Thus, HML expression is regulated by two independent silencers, each fully competent for maintaining repression. This situation was distinct from the organization of the other silent locus, HMR, at which a single silencer served as the predominant repressor of expression. Examination of identifiable domains and binding sites within the HML silencers suggested that silencing activity can be achieved by a variety of combinations of various functional domains.


Serological profiling of the EBV immune response in Chronic Fatigue Syndrome using a peptide microarray.
Epstein-Barr-Virus (EBV) plays an important role as trigger or cofactor for various autoimmune diseases. In a subset of patients with Chronic Fatigue Syndrome (CFS) disease starts with infectious mononucleosis as late primary EBV-infection, whereby altered levels of EBV-specific antibodies can be observed in another subset of patients. We performed a comprehensive mapping of the IgG response against EBV comparing 50 healthy controls with 92 CFS patients using a microarray platform. Patients with multiple sclerosis (MS), systemic lupus erythematosus (SLE) and cancer-related fatigue served as controls. 3054 overlapping peptides were synthesised as 15-mers from 14 different EBV proteins. Array data was validated by ELISA for selected peptides. Prevalence of EBV serotypes was determined by qPCR from throat washing samples. EBV type 1 infections were found in patients and controls. EBV seroarray profiles between healthy controls and CFS were less divergent than that observed for MS or SLE. We found significantly enhanced IgG responses to several EBNA-6 peptides containing a repeat sequence in CFS patients compared to controls. EBNA-6 peptide IgG responses correlated well with EBNA-6 protein responses. The EBNA-6 repeat region showed sequence homologies to various human proteins. Patients with CFS had a quite similar EBV IgG antibody response pattern as healthy controls. Enhanced IgG reactivity against an EBNA-6 repeat sequence and against EBNA-6 protein is found in CFS patients. Homologous sequences of various human proteins with this EBNA-6 repeat sequence might be potential targets for antigenic mimicry.


Cost-effectiveness of a genetic test for breast cancer risk.
Genetic testing of seven single-nucleotide polymorphisms (7SNP) can improve estimates of risk of breast cancer relative to the Gail risk test alone, for the purpose of recommending MRI screening for women at high risk. A simulation of breast cancer and health care processes was used to conduct a virtual trial comparing the use of the 7SNP test with the Gail risk test to categorize patients by risk. Average-risk patients received annual mammogram, whereas high-risk patients received annual MRI. Cancer incidence was based on Surveillance, Epidemiology, and End Results data and validated to Cancer Prevention Study II Nutrition Cohort data. Risk factor values were drawn from National Health and Nutrition Examination Survey (NHANES-4) and Prostate, Lung, Colorectal, and Ovarian Cancer Screening Trial data. Mammogram characteristics were derived from Breast Cancer Surveillance Consortium data. The test was most cost-effective when given to patients at an intermediate lifetime risk of breast cancer. For patients with a risk of 16% to 28%, it resulted in a 1.91% reduction in cancer deaths, saving 0.005 quality-adjusted life years per person at a cost of $163,264 per QALY. These results were sensitive to the age at which the test is given, the discount rate, and the costs of the genetic test and MRI. The cost effectiveness of using the 7SNP test for patients with intermediate Gail risk is similar to that of other recommended strategies, including annual MRI for patients with a lifetime risk greater than 20% or BRCA1/2 mutations.


A hybrid qPCR/SNP array approach allows cost efficient assessment of KIR gene copy numbers in large samples.
Killer Immunoglobulin-like Receptors (KIRs) are surface receptors of natural killer cells that bind to their corresponding Human Leukocyte Antigen (HLA) class I ligands, making them interesting candidate genes for HLA-associated autoimmune diseases, including type 1 diabetes (T1D). However, allelic and copy number variation in the KIR region effectively mask it from standard genome-wide association studies: single nucleotide polymorphism (SNP) probes targeting the region are often discarded by standard genotype callers since they exhibit variable cluster numbers. Quantitative Polymerase Chain Reaction (qPCR) assays address this issue. However, their cost is prohibitive at the sample sizes required for detecting effects typically observed in complex genetic diseases. We propose a more powerful and cost-effective alternative, which combines signals from SNPs with more than three clusters found in existing datasets, with qPCR on a subset of samples. First, we showed that noise and batch effects in multiplexed qPCR assays are addressed through normalisation and simultaneous copy number calling of multiple genes. Then, we used supervised classification to impute copy numbers of specific KIR genes from SNP signals. We applied this method to assess copy number variation in two KIR genes, KIR3DL1 and KIR3DS1, which are suitable candidates for T1D susceptibility since they encode the only KIR molecules known to bind with HLA-Bw4 epitopes. We find no association between KIR3DL1/3DS1 copy number and T1D in 6744 cases and 5362 controls; a sample size twenty-fold larger than in any previous KIR association study. Due to our sample size, we can exclude odds ratios larger than 1.1 for the common KIR3DL1/3DS1 copy number groups at the 5% significance level. We found no evidence of association of KIR3DL1/3DS1 copy number with T1D, either overall or dependent on HLA-Bw4 epitope. Five other KIR genes, KIR2DS4, KIR2DL3, KIR2DL5, KIR2DS5 and KIR2DS1, in high linkage disequilibrium with KIR3DL1 and KIR3DS1, are also unlikely to be significantly associated. Our approach could potentially be applied to other KIR genes to allow cost effective assaying of gene copy number in large samples.


Speed and eccentricity tuning reveal a central role for the velocity-based cue to 3D visual motion.
Two binocular cues are thought to underlie the visual perception of three-dimensional (3D) motion: a disparity-based cue, which relies on changes in disparity over time, and a velocity-based cue, which relies on interocular velocity differences. The respective building blocks of these cues, instantaneous disparity and retinal motion, exhibit very distinct spatial and temporal signatures. Although these two cues are synchronous in naturally moving objects, disparity-based and velocity-based mechanisms can be dissociated experimentally. We therefore investigated how the relative contributions of these two cues change across a range of viewing conditions. We measured direction-discrimination sensitivity for motion though depth across a wide range of eccentricities and speeds for disparity-based stimuli, velocity-based stimuli, and "full cue" stimuli containing both changing disparities and interocular velocity differences. Surprisingly, the pattern of sensitivity for velocity-based stimuli was nearly identical to that for full cue stimuli across the entire extent of the measured spatiotemporal surface and both were clearly distinct from those for the disparity-based stimuli. These results suggest that for direction discrimination outside the fovea, 3D motion perception primarily relies on the velocity-based cue with little, if any, contribution from the disparity-based cue.


Estimation and prediction for a mechanistic model of measles transmission using particle filtering and maximum likelihood estimation.
Disease incidence reported directly within health systems frequently reflects a partial observation relative to the true incidence in the population. State-space models present a general framework for inferring both the dynamics of infectious disease processes and the unobserved burden of disease in the population. Here, we present a state-space model of measles transmission and vaccine-based interventions at the country-level and a particle filter-based estimation procedure. Our dynamic transmission model builds on previous work by incorporating population age-structure to allow explicit representation of age-targeted vaccine interventions. We illustrate the performance of estimators of model parameters and predictions of unobserved states on simulated data from two dynamic models: one on the annual time-scale of observations and one on the biweekly time-scale of the epidemiological dynamics. We show that our model results in approximately unbiased estimates of unobserved burden and the underreporting rate. We further illustrate the performance of the fitted model for prediction of future disease burden in the next one to 15 years.


Ethylene Signaling Is Required for Fully Functional Tension Wood in Hybrid Aspen.
Tension wood (TW) in hybrid aspen trees forms on the upper side of displaced stems to generate a strain that leads to uplifting of the stem. TW is characterized by increased cambial growth, reduced vessel frequency and diameter, and the presence of gelatinous, cellulose-rich (G-)fibers with its microfibrils oriented parallel to the fiber cell axis. Knowledge remains limited about the molecular regulators required for the development of this special xylem tissue with its characteristic morphological, anatomical, and chemical features. In this study, we use transgenic, ethylene-insensitive (ETI) hybrid aspen trees together with time-lapse imaging to show that functional ethylene signaling is required for full uplifting of inclined stems. X-ray diffraction and Raman microspectroscopy of TW in ETI trees indicate that, although G-fibers form, the cellulose microfibril angle in the G-fiber S-layer is decreased, and the chemical composition of S- and G-layers is altered than in wild-type TW. The characteristic asymmetric growth and reduction of vessel density is suppressed during TW formation in ETI trees. A genome-wide transcriptome profiling reveals ethylene-dependent genes in TW, related to cell division, cell wall composition, vessel differentiation, microtubule orientation, and hormone crosstalk. Our results demonstrate that ethylene regulates transcriptional responses related to the amount of G-fiber formation and their properties (chemistry and cellulose microfibril angle) during TW formation. The quantitative and qualitative changes in G-fibers are likely to contribute to uplifting of stems that are displaced from their original position.


Green synthesis of imine-based covalent organic frameworks in water.
Dynamic covalent bonds have been advantageously used to direct the synthesis of crystalline porous covalent organic frameworks (COFs). Unlike the standard synthetic protocols that involve harsh conditions, this work provides a high-yield "one-pot" green synthesis of imine-based COFs in water. Additionally, this aqueous synthesis can be performed under microwave conditions, considerably reducing the reaction time.


Coronary Artery Calcium From Early Adulthood to Middle Age and Left Ventricular Structure and Function.
Background The relationship of coronary artery calcium (CAC) with adverse cardiac remodeling is not well established. We aimed to study the association of CAC in middle age and change in CAC from early adulthood to middle age with left ventricular (LV) function. Methods CAC score was measured by computed tomography at CARDIA study (Coronary Artery Risk Development in Young Adults) year-15 examination and at year-25 examination (Y25) in 3043 and 3189 participants, respectively. CAC score was assessed as a continuous variable and log-transformed to account for nonlinearity. Change in CAC from year-15 examination to Y25 was evaluated as the absolute difference of log-transformed CAC from year-15 examination to Y25. LV structure and function were evaluated by echocardiography at Y25. Results At Y25, mean age was 50.1±3.6 years, 56.6% women, 52.4% black. In the multivariable analysis at Y25, higher CAC was related to higher LV mass (β=1.218; adjusted P=0.007), higher LV end-diastolic volume (β=0.811; adjusted P=0.007), higher LV end-systolic volume (β=0.350; adjusted P=0.048), higher left atrial volume (β=0.214; adjusted P=0.009), and higher E/e' ratio (β=0.059; adjusted P=0.014). CAC was measured at both year-15 examination and Y25 in 2449 individuals. Higher change in CAC score during follow-up was independently related to higher LV mass index in blacks (β=4.789; adjusted P<0.001), but not in whites (β=1.051; adjusted P=0.283). Conclusions Higher CAC in middle age is associated with higher LV mass and volumes and worse LV diastolic function. Being free of CAC from young adulthood to middle age correlates to better LV function at middle age. Higher change in CAC score during follow-up is independently related to higher LV mass index in blacks.


Synchronization of HeLa cell cultures by inhibition of DNA polymerase alpha with aphidicolin.
Both the inhibitory effect of aphidicolin on the replicative alpha-polymerase and the reversibility of its action in vivo (Pedrali-Noy & Spadari, 1979, Biochem. Biophys. Res. Commun. 88, 1194-2002) allow the synchronization of cells in culture. Aphidicolin prevents G1 cells from entering the DNA synthetic period, blocks cells in "S" phase, allows G2, M and G1 cells to continue the cell cycle and to accumulate at the G1/S border. Aphidicolin is a more useful reagent than hydroxyurea and thymidine because it does not affect cell viability or "S" phase duration and does not interfere with the synthesis of dNTPs or DNA polymerases. In fact cells exposed to the drug continue to synthesize all three DNA polymerases alpha, beta and gamma as well as all dNTPs which, when the block is removed, are present at levels optimal for DNA initiation and replication. The technique is simple and can be applied to cells growing in suspension or monolayers and allows one to harvest large quantities of synchronized cells.


The role of cystoscopy in the diagnosis of Hunner's ulcer disease.
Interstitial cystitis can be classified into Hunner's ulcer and nonulcer disease, which are easily distinguished by cystoscopic examination. Although therapeutic options may differ between these 2 groups, the diagnosis of interstitial cystitis is currently established by many clinicians in the absence of cystoscopy. We determined whether clinical parameters alone without cystoscopic evaluation could reliably distinguish the patient with Hunner's ulcer vs nonHunner's ulcer disease. Data were collected on 184 women and 39 men who met National Institute of Diabetes, Digestive and Kidney Diseases criteria for interstitial cystitis and who were evaluated at our institution between 1990 and 2005. A total of 86 patients with Hunner's ulcer were consecutively identified. Of patients with nonHunner's ulcer disease seen during that period a cohort of 137 who were consecutively identified were selected as a comparison group. Clinical data on each patient were collected. The groups were compared by the 2-sample t test, the Mann-Whitney test when appropriate and the chi-square test. No significant differences in clinical parameters were found between women and men in either group. The female-to-male ratio was 6:1 and 3:1 in the nonHunner's and Hunner's ulcer groups, respectively. The mean age of patients with Hunner's ulcer was significantly higher than that of patients with nonHunner's ulcer disease (60 vs 47 years, t test p <0.001). No significant differences in symptom duration, history of gross hematuria, history of comorbid disease or visual analog pain scores were found between the 2 groups. Microscopic hematuria was present in 27 (31%) and 29 patients (21%) with Hunner's ulcer and nonHunner's ulcer disease, respectively (chi-square test p <0.086). Although there have been recently published methods and markers by which to differentiate Hunner's ulcer vs nonHunner's ulcer interstitial cystitis, our data demonstrate that standard clinical evaluation cannot reliably distinguish these groups. These findings suggest that cystoscopy is needed to accurately identify patients with Hunner's ulcer.


Factors affecting outcome in frameless non-isocentric stereotactic radiosurgery for trigeminal neuralgia: a multicentric cohort study.
Stereotactic radiosurgery (SRS) is an effective treatment for trigeminal neuralgia (TN). Nevertheless, a proportion of patients will experience recurrence and treatment-related sensory disturbances. In order to evaluate the predictors of efficacy and safety of image-guided non-isocentric radiosurgery, we analyzed the impact of trigeminal nerve volume and the nerve dose/volume relationship, together with relevant clinical characteristics. Two-hundred and ninety-six procedures were performed on 262 patients at three centers. In 17 patients the TN was secondary to multiple sclerosis (MS). Trigeminal pain and sensory disturbances were classified according to the Barrow Neurological Institute (BNI) scale. Pain-free-intervals were investigated using Kaplan Meier analyses. Univariate and multivariate Cox regression analyses were performed to identify predictors. The median follow-up period was 38 months, median maximal dose 72.4 Gy, median target nerve volume 25 mm3, and median prescription dose 60 Gy. Pain control rate (BNI I-III) at 6, 12, 24, 36, 48, and 60 months were 96.8, 90.9, 84.2, 81.4, 74.2, and 71.2%, respectively. Overall, 18% of patients developed sensory disturbances. Patients with volume ≥ 30 mm3 were more likely to maintain pain relief (p = 0.031), and low integral dose (< 1.4 mJ) tended to be associated with more pain recurrence than intermediate (1.4-2.7 mJ) or high integral dose (> 2.7 mJ; low vs. intermediate: log-rank test, χ2 = 5.02, p = 0.019; low vs. high: log-rank test, χ2 = 6.026, p = 0.014). MS, integral dose, and mean dose were the factors associated with pain recurrence, while re-irradiation and MS were predictors for sensory disturbance in the multivariate analysis. The dose to nerve volume ratio is predictive of pain recurrence in TN, and re-irradiation has a major impact on the development of sensory disturbances after non-isocentric SRS. Interestingly, the integral dose may differ significantly in treatments using apparently similar dose and volume constraints.


The influence of the α/β ratio on treatment time iso-effect relationships in the central nervous system.
Purpose: To investigate the influence of changes in α/β ratio (range 1.5-3 Gy) on iso-effective doses, with varying treatment time, in spinal cord and central nervous system tissues with comparable radio-sensitivity. It is important to establish if an α/β ratio of 2 Gy, the accepted norm for neuro-oncology iso-effect estimations, can be used.Methods: The rat spinal cord irradiation data of Pop et al. provided ED50 values for radiation myelopathy for treatment times that varied from minutes to ∼6 days. Analysis using biphasic repair kinetics, allowing for variable dose-rates, provided the best fit with repair half-times of 0.19 and 2.16 hr, each providing ∼50% of overall repair; with an α/β ratio 2.47 Gy (CI 1.5-3.95 Gy). Using the above data set, graphical methods were used to investigate changes in the repair parameters for differing fixed α/β ratios between 1.5 and 3.0 Gy. Two different intermittent dose delivery equations were used to evaluate the implications in a radiosurgery setting.Results: Changes in the α/β ratio (1.5-3.0 Gy) have a minor effect on equivalent doses for radiation myelopathy for treatment durations of a few hours. Changing the α/β value from 2 Gy to 2.47 Gy, modified equivalent single doses by < 1% when overall treatment times ranged from 0.1 to 5.0 hr. Significant changes were only found for treatment times longer than 5-10 hr. These two α/β ratios were also compared in a practical radiosurgery situation, using two different models for estimating BED, again there was no significant loss of accuracy.Conclusions: It is reasonable to use an α/β ratio of 2 Gy for CNS tissue, with the same repair half-times as published in the original publication by Pop et al., in situations where the assessment of the BED in radiosurgery is used with other form of radiotherapy. In radiosurgery, the variation in BED with treatment duration (for a fixed physical dose) is very similar, but absolute BED values depend on the α/β value. In radiosurgery, clinical recommendations obtained using BED calculations using the originally proposed α/β ratio of 2.47 Gy are still appropriate. For calculations involving a combination of radiosurgery and other modalities, such as fractionated radiotherapy, it would be appropriate in all cases to apply a value of 2 Gy, the accepted norm in neuro-oncology, without significant loss of accuracy in the radio-surgical component. This may have important applications in retreatment situations.


Influence of protein corona and caveolae-mediated endocytosis on nanoparticle uptake and transcytosis.
Transcytosis of nanoparticles (NPs) is emerging as an attractive alternative to the paracellular route in cancer drug delivery with studies suggesting targeting caveolae-mediated endocytosis to maximize NP transcytosis. However, there are limited studies on transcytosis of NPs, especially for corona-coated NPs. Most studies focused on cellular uptake as an indirect measure of the NP's transcellular permeability (Pd). Here, we probed the effect of protein corona on the uptake and transcytosis of 20, 40, 100, and 200 nm polystyrene nanoparticles (pNP-PC) across HUVECs in a microfluidic channel that modelled the microvasculature. We observed increased cell uptake with size of pNP-PC although it was the smallest 20 nm pNP-PC that exhibited the highest transcellular Pd. In the absence of corona however, cell uptake decreased with size, and the largest 200 nm pNP-PEG exhibited the lowest transcellular Pd. By inhibiting caveolae-mediated endocytosis in HUVECs, smaller pNPs had a larger drop in cell uptake than larger pNPs, regardless of surface coating. However, only the smallest (20 nm) and largest (200 nm) pNP-PC had a decrease in Pd following inhibition with MβCD. Our findings showed that the protein corona affected the transcytosis of NPs, and their uptake by caveolae-mediated endocytosis did not necessarily lead to transcytosis.


Measurement of triplet optical densities of organic compounds in the presence of photodecomposition.
We have developed a method to measure triplet optical densities OD(r)(lambda) when considerable photodecomposition is present. The method is demonstrated on Coumarin 120. By focusing an UV cw laser beam on a sample with the aid of a lens, high triplet optical densities OD(T)(lambda) can be recorded. However, the excitation of organic compounds with light causes varying degrees of photodecomposition. In order to take photodecomposition into account, one records triplet optical-density values as well as the accumulation of absorbing photoproducts as a function of time. Turning off the cw laser excitation, one records the accumulation of photoproducts only. Separating the two processes, one can determine how the triplet optical density OD(T) declines as a function of time t. The obtained curve can be expressed with an equation that appears to decline exponentially with time. This allows one to extrapolate back to t = 0 and recover OD(T) when there was no photodecomposition. The extrapolated OD(T) values are used to obtain triplet-extinction coefficients (T)(lambda) by McClure's method of Coumarin 120.


Moth olfactory receptor neurons adjust their encoding efficiency to temporal statistics of pheromone fluctuations.
The efficient coding hypothesis predicts that sensory neurons adjust their coding resources to optimally represent the stimulus statistics of their environment. To test this prediction in the moth olfactory system, we have developed a stimulation protocol that mimics the natural temporal structure within a turbulent pheromone plume. We report that responses of antennal olfactory receptor neurons to pheromone encounters follow the temporal fluctuations in such a way that the most frequent stimulus timescales are encoded with maximum accuracy. We also observe that the average coding precision of the neurons adjusted to the stimulus-timescale statistics at a given distance from the pheromone source is higher than if the same encoding model is applied at a shorter, non-matching, distance. Finally, the coding accuracy profile and the stimulus-timescale distribution are related in the manner predicted by the information theory for the many-to-one convergence scenario of the moth peripheral sensory system.


Evaluation of Efficacy of a Skin Care Regimen Containing Methyl Estradiolpropanoate (MEP) for Treating Estrogen Deficient Skin
BACKGROUND: Aging is a complex process due to the interplay of intrinsic factors (such as genetics and hormones) and extrinsic factors (including ultraviolet radiation and pollution). A significant cause of intrinsic aging in women is the loss of estrogen as a result of the onset of menopause. OBJECTIVE: A single site experience trial to assess the efficacy of Emepelle (Biopelle, Ferndale Pharma Group, Ferndale, MI), a skin care regimen containing Methyl Estradiolpropanoate (MEP)®, for the treatment of Estrogen Deficient Skin (EDS). The secondary objective was to assess patient tolerability and satisfaction. METHODS: Fourteen female subjects aged 53-68 years who were amenorrheic for 1-10 years (mean, 5 years), with at least a Grade II in Wrinkling (fine to moderate-depth wrinkles, moderate number of lines) and score of at least 5 (of 9; moderate-to-severe) in elastosis on the clinician-assessed Fitzpatrick-Goldman Classification of Wrinkling and Elastosis Scale, and a 3 or greater on the Investigator Facial Skin Hydration Scale, were included in the study. The subjects were instructed to apply the product Emepelle Serum in the morning, and the product Emepelle Night Cream in the evening to the entire freshly washed and dried face. Follow up visits were performed at 8 weeks, 14 weeks, and 20 weeks to evaluate efficacy and safety. Canfield Visia Complexion Analysis and standard photography was performed at baseline and at each follow up visit. RESULTS: On a 0-4 Facial Hydration Scale, 100% of study participants by week 20 showed at least one-grade improvement and 93% saw two grades or more improvement in hydration. 100% of study participants showed aesthetic improvements per investigator-assessed Global Aesthetic Improvement Scale (GAIS) at week 14. By week 20, 93% of study participants responded that the combination of Emepelle Serum and Night Cream regimen helped improve wrinkles, texture, and color, and 86% of study participants responded that Emepelle helped improve sun-damage, thickness, and integrity. In the Quality of Life questionnaire, 86% responded that Emepelle helped alleviate some or all of the skin issues they developed since entering menopause. Investigator clinical assessment scored patients with a 53% improvement in texture, 21% improvement in keratoses, and 15% improvement in laxity on the Alexiades-Armenakas Comprehensive Grading Scale for Assessment of Skin Aging and Photodamage by the end of the study at week 20. CONCLUSIONS: Patients in the study indicated satisfaction with the formulations of Emepelle Serum and Night Cream. Younger patients showed significant improvement by about 8 weeks. For patients who have been in menopause longer, significant improvement was seen by week 20, suggesting MEP's potential ability to reactivate dormant estrogen receptors. J Drugs Dermatol. 2019;18(12):1226-1230.


Aortic homograft endocarditis caused by Cardiobacterium hominis and complicated by agranulocytosis due to ceftriaxone.
The present report describes a very rare case of an aortic homograft valve endocarditis caused by Cardiobacterium hominis. The case was complicated by an agranulocytosis after 3 weeks of antibiotic treatment induced by ceftriaxone. Alternative oral treatment with ciprofloxacin and rifampicin was successful, no surgical intervention was needed and homograft function could be preserved.


Endovascular laser therapy for varicose veins: an evidence-based analysis.
The objective of the MAS evidence review was to conduct a systematic review of the available evidence on the safety, effectiveness, durability and cost-effectiveness of endovascular laser therapy (ELT) for the treatment of primary symptomatic varicose veins (VV). The Ontario Health Technology Advisory Committee (OHTAC) met on November 27, 2009 to review the safety, effectiveness, durability and cost-effectiveness of ELT for the treatment of primary VV based on an evidence-based review by the Medical Advisory Secretariat (MAS). CLINICAL CONDITION: VV are tortuous, twisted, or elongated veins. This can be due to existing (inherited) valve dysfunction or decreased vein elasticity (primary venous reflux) or valve damage from prior thrombotic events (secondary venous reflux). The end result is pooling of blood in the veins, increased venous pressure and subsequent vein enlargement. As a result of high venous pressure, branch vessels balloon out leading to varicosities (varicose veins). SYMPTOMS TYPICALLY AFFECT THE LOWER EXTREMITIES AND INCLUDE (BUT ARE NOT LIMITED TO): aching, swelling, throbbing, night cramps, restless legs, leg fatigue, itching and burning. Left untreated, venous reflux tends to be progressive, often leading to chronic venous insufficiency (CVI). A NUMBER OF COMPLICATIONS ARE ASSOCIATED WITH UNTREATED VENOUS REFLUX: including superficial thrombophlebitis as well as variceal rupture and haemorrhage. CVI often results in chronic skin changes referred to as stasis dermatitis. Stasis dermatitis is comprised of a spectrum of cutaneous abnormalities including edema, hyperpigmentation, eczema, lipodermatosclerosis and stasis ulceration. Ulceration represents the disease end point for severe CVI. CVI is associated with a reduced quality of life particularly in relation to pain, physical function and mobility. In severe cases, VV with ulcers, QOL has been rated to be as bad or worse as other chronic diseases such as back pain and arthritis. Lower limb VV is a common disease affecting adults and estimated to be the seventh most common reason for physician referral in the US. There is a strong familial predisposition to VV with the risk in offspring being 90% if both parents affected, 20% when neither is affected, and 45% (25% boys, 62% girls) if one parent is affected. Globally, the prevalence of VV ranges from 5% to 15% among men and 3% to 29% among women varying by the age, gender and ethnicity of the study population, survey methods and disease definition and measurement. The annual incidence of VV estimated from the Framingham Study was reported to be 2.6% among women and 1.9% among men and did not vary within the age range (40-89 years) studied. Approximately 1% of the adult population has a stasis ulcer of venous origin at any one time with 4% at risk. The majority of leg ulcer patients are elderly with simple superficial vein reflux. Stasis ulcers are often lengthy medical problems and can last for several years and, despite effective compression therapy and multilayer bandaging are associated with high recurrence rates. Recent trials involving surgical treatment of superficial vein reflux have resulted in healing and significantly reduced recurrence rates. ENDOVASCULAR LASER THERAPY FOR VV: ELT is an image-guided, minimally invasive treatment alternative to surgical stripping of superficial venous reflux. It does not require an operating room or general anesthesia and has been performed in outpatient settings by a variety of medical specialties including surgeons (vascular or general), interventional radiologists and phlebologists. Rather than surgically removing the vein, ELT works by destroying, cauterizing or ablating the refluxing vein segment using heat energy delivered via laser fibre. Prior to ELT, colour-flow Doppler ultrasonography is used to confirm and map all areas of venous reflux to devise a safe and effective treatment plan. The ELT procedure involves the introduction of a guide wire into the target vein under ultrasound guidance followed by the insertion of an introducer sheath through which an optical fibre carrying the laser energy is advanced. A tumescent anesthetic solution is injected into the soft tissue surrounding the target vein along its entire length. This serves to anaesthetize the vein so that the patient feels no discomfort during the procedure. It also serves to insulate the heat from damaging adjacent structures, including nerves and skin. Once satisfactory positioning has been confirmed with ultrasound, the laser is activated. Both the laser fibre and the sheath are simultaneously, slowly and continuously pulled back along the length of the target vessel. At the end of the procedure, homeostasis is then achieved by applying pressure to the entry point. Adequate and proper compression stockings and bandages are applied after the procedure to reduce the risk of venous thromboembolism, and to reduce postoperative bruising and tenderness. Patients are encouraged to walk immediately after the procedure and most patients return to work or usual activity within a few days. Follow-up protocols vary, with most patients returning 1-3 weeks later for an initial follow-up visit. At this point, the initial clinical result is assessed and occlusion of the treated vessels is confirmed with ultrasound. Patients often have a second follow-up visit 1-3 months following ELT at which time clinical evaluation and ultrasound are repeated. If required, sclerotherapy may be performed during the ELT procedure or at any follow-up visits. Endovascular laser for the treatment of VV was approved by Health Canada as a class 3 device in 2002. The treatment has been an insured service in Saskatchewan since 2007 and is the only province to insure ELT. Although the treatment is not an insured service in Ontario, it has been provided by various medical specialties since 2002 in over 20 private clinics. The MAS evidence-based review was performed as an update to the 2007 health technology review performed by the Australian Medical Services Committee (MSAC) to support public financing decisions. The literature search was performed on August 18, 2009 using standard bibliographic databases for studies published from January 1, 2007 to August 15, 2009. Search alerts were generated and reviewed for additional relevant literature up until October 1, 2009. English language full-reports and human studiesOriginal reports with defined study methodologyReports including standardized measurements on outcome events such as technical success, safety, effectiveness, durability, quality of life or patient satisfactionReports involving ELT for VV (great or small saphenous veins)Randomized controlled trials (RCTs), systematic reviews and meta-analysesCohort and controlled clinical studies involving > 1 month ultrasound imaging follow-up Non systematic reviews, letters, comments and editorialsReports not involving outcome events such as safety, effectiveness, durability, or patient satisfaction following an intervention with ELTReports not involving interventions with ELT for VVPilot studies or studies with small samples ( < 50 subjects) The MAS evidence search identified 14 systematic reviews, 29 cohort studies on safety and effectiveness, four cost studies and 12 randomized controlled trials involving ELT, six of these comparing endovascular laser with surgical ligation and saphenous vein stripping. Since 2007, 22 cohort studies involving 10,883 patients undergoing ELT of the great saphenous vein (GSV) have been published. Imaging defined treatment effectiveness of mean vein closure rates were reported to be greater than 90% (range 93%- 99%) at short term follow-up. Longer than one year follow-up was reported in five studies with life table analysis performed in four but the follow up was still limited at three and four years. The overall pooled major adverse event rate, including DVT, PE, skin burns or nerve damage events extracted from these studies, was 0.63% (69/10,883). The overall level of evidence of randomized trials comparing ELT with surgical ligation and vein stripping (n= 6) was graded as moderate to high. Recovery after treatment was significantly quicker after ELT (return to work median number of days, 4 vs. 17; p= .005). Major adverse events occurring after surgery were higher [(1.8% (n=4) vs. 0.4% (n = 1) 1 but not significantly. Treatment effectiveness as measured by imaging vein absence or closure, symptom relief or quality of life similar in the two treatment groups and both treatments resulted in statistically significantly improvements in these outcomes. Recurrence was low after both treatments at follow up but neovascularization (growth of new vessels, a key predictor of long term recurrence was significantly more common (18% vs. 1%; p = .001) after surgery. Although patient satisfaction was reported to be high (>80%) with both treatments, patient preferences evaluated through recruitment process, physician reports and consumer groups were strongly in favour of ELT. For patients minimal complications, quick recovery and dependability of outpatient scheduling were key considerations. As clinical effectiveness of the two treatments was similar, a cost-analysis was performed to compare differences in resources and costs between the two procedures. A budget impact analysis for introducing ELT as an insured service was also performed. The average case cost (based on Ontario hospital costs and medical resources) for surgical vein stripping was estimated to be $1,799. Because of the uncertainties with resources associated with ELT, in addition to the device related costs, hospital costs were varied and assumed to be the same as or less than (40%) those for surgery resulting in an average ELT case cost of $2,025 or $1,602. (ABSTRACT TRUNCATED)


Controlled nanometric fibers of self-assembled designed protein scaffolds.
The use of biological molecules as platforms for templating and nanofabrication is an emerging field. Here, we use designed protein building blocks based on small repetitive units (consensus tetratricopeptide repeat - CTPR) to generate fibrillar linear nanostructures by controlling the self-assembly properties of the units. We fully characterize the kinetics and thermodynamics of the assembly and describe the polymerization process by a simple model that captures the features of the structures formed under defined conditions. This work, together with previously established functionalization potential, sets up the basis for the application of these blocks in the fabrication and templating of complex hybrid nanostructures.


A new family of Ty1-copia-like retrotransposons originated in the tomato genome by a recent horizontal transfer event.
Rider is a novel and recently active Ty1-copia-like retrotransposon isolated from the T3238fer mutant of tomato. Structurally, it is delimited by a duplication of target sites and contains two long terminal direct repeats and an internal open reading frame, which encodes a Ty1-copia-type polyprotein with characteristic protein domains required for retrotransposition. The family of Rider elements has an intermediate copy number and is scattered in the chromosomes of tomato. Rider family members in the tomato genome share high sequence similarity, but different structural groups were identified (full-size elements, deletion derivatives, and solo LTRs). Southern blot analysis in Solanaceae species showed that Rider was a Lycopersicon-specific element. Sequence analysis revealed that among other plants, two Arabidopsis elements (named as Rider-like 1 and Rider-like 2) are most similar to Rider in both the coding and noncoding regions. RT-PCR analysis indicates that Rider is constitutively expressed in tomato plants. The phylogeny-based parsimony analysis and the sequence substitution analyses of these data suggest that these Rider-like elements originated from a recent introgression of Rider into the tomato genome by horizontal transfer 1-6 million years ago. Considering its transcriptional activity and the recent insertion of the element into at least two genes, Rider is a recently active retrotransposon in the tomato genome.


A Synthetic Biology Approach to Engineering Living Photovoltaics.
The ability to electronically interface living cells with electron accepting scaffolds is crucial for the development of next-generation biophotovoltaic technologies. Although recent studies have focused on engineering synthetic interfaces that can maximize electronic communication between the cell and scaffold, the efficiency of such devices is limited by the low conductivity of the cell membrane. This review provides a materials science perspective on applying a complementary, synthetic biology approach to engineering membrane-electrode interfaces. It focuses on the technical challenges behind the introduction of foreign extracellular electron transfer pathways in bacterial host cells and the past and future efforts to engineer photosynthetic organisms with artificial electron-export capabilities for biophotovoltaic applications. The article highlights advances in engineering protein-based, electron-exporting conduits in a model host organism, E. coli, before reviewing state-of-the-art biophotovoltaic technologies that use both unmodified and bioengineered photosynthetic bacteria with improved electron transport capabilities. A thermodynamic analysis is used to propose an energetically feasible pathway for extracellular electron transport in engineered cyanobacteria and identify metabolic bottlenecks amenable to protein engineering techniques. Based on this analysis, an engineered photosynthetic organism expressing a foreign, protein-based electron conduit yields a maximum theoretical solar conversion efficiency of 6-10% without accounting for additional bioengineering optimizations for light-harvesting.


Investigation of Astaxanthin Effect on Cisplatin Ototoxicity in Rats by Using Otoacoustic Emission, Total Antioxidant Capacity, and Histopathological Methods.
Cisplatin-induced ototoxicity is related to oxidative stress. Astaxanthin is one of the most powerful antioxidants in nature. To investigate the protective effect of astaxanthin on cisplatin-induced ototoxicity. Thirty-five Sprague Dawley female rats were divided into 5 groups: control, cisplatin, and cisplatin with 10, 20, and 40 mg/kg astaxanthin groups. Cisplatin group received a single intraperitoneal injection of 14 mg/kg cisplatin. While saline was administered in the control group, in the other 3 groups, 10, 20, and 40 mg/kg daily doses of astaxanthin were administered through orogastric cannula before administration of cisplatin. Baseline and 10th day otoacoustic emission tests were administered. An intracardiac blood sample was taken to measure total antioxidant capacity (TAC), and the cochleas of the animals were investigated histopathologically. Hearing level of astaxanthin 40 mg/kg + cisplatin group was higher at 24 kHz and 32 kHz frequencies compared to the cisplatin group. The TAC value of the cisplatin group was lower than both the control and astaxanthin + cisplatin groups (P < .05). On histopathological examination, the other groups were deformed compared to the control group, but no statistically significant difference was observed between the astaxanthin + cisplatin and cisplatin groups. Astaxanthin showed protective effect at high frequencies when it was administered at high dose. Thus, astaxanthin may have protective effect against cisplatin-induced ototoxicity.


No Association between Circulating Levels and Genetic Variants of IL-6 and TNF-α and Colon Adenoma.
Interleukin 6 (IL-6) and tumor necrosis factor alpha (TNF-α), two important inflammatory cytokines, have been inconsistently associated with risk of colon neoplasia in epidemiological studies. However, research to date has not adequately assessed whether race-specific differences may exist in associations between biomarkers and genetic variants of these cytokines and colorectal adenoma - the precursor lesions of colorectal cancer. We sought to determine whether circulating levels of IL-6 and TNF-α, or genetic polymorphisms in IL-6and TNF-α were associated with colon adenoma and if so, whether that association differed by race. We analyzed the associations of circulating levels and single nucleotide polymorphisms (SNPs) of IL-6 and TNF-α with risk of colon adenomas in a colonoscopy -based case-control study of 401 incident adenoma cases and 1,050 controls. We used multivariate unconditional logistic regression models to estimate the odds ratios (OR) and 95% confidence intervals (95% CI) for levels or genotypes (log additive models) of IL-6 and TNF-α. Compared to the bottom tertile of IL-6, the adjusted ORs were: 1.06 (0.75-1.44), 1.01 (0.72-1.40), respectively for the 2nd and 3rd tertiles (ptrend = 0.10); the corresponding ORs for TNF-α were: 0.85 (0.63-1.15) and 1.01 (0.75-1.36), respectively (ptrend = 0.39). Race-stratified analyses did not reveal any significant association. There were also no statistically significant associations between IL-6 and TNF-α SNPs and colon adenoma. Our results do not support pre-diagnostic levels of IL-6, TNF-α or their genetic variants as significant risk factors for the development of colon adenoma.


Pharmacogenetics in American Indian populations: analysis of CYP2D6, CYP3A4, CYP3A5, and CYP2C9 in the Confederated Salish and Kootenai Tribes.
Cytochrome P450 enzymes play a dominant role in drug elimination and variation in these genes is a major source of interindividual differences in drug response. Little is known, however, about pharmacogenetic variation in American Indian and Alaska Native (AI/AN) populations. We have developed a partnership with the Confederated Salish and Kootenai Tribes (CSKT) in northwestern Montana to address this knowledge gap. We resequenced CYP2D6 in 187 CSKT individuals and CYP3A4, CYP3A5, and CYP2C9 in 94 CSKT individuals. We identified 67 variants in CYP2D6, 15 in CYP3A4, 10 in CYP3A5, and 41 in CYP2C9. The most common CYP2D6 alleles were CYP2D6*4 and *41 (20.86 and 11.23%, respectively). CYP2D6*3, *5, *6, *9, *10, *17, *28, *33, *35, *49, *1xN, *2xN, and *4xN frequencies were less than 2%. CYP3A5*3, CYP3A4*1G, and *1B were detected with frequencies of 92.47, 26.81, and 2.20%, respectively. Allelic variation in CYP2C9 was low: CYP2C9*2 (5.17%) and *3 (2.69%). In general, allele frequencies in CYP2D6, CYP2C9, and CYP3A5 were similar to those observed in European Americans. There was, however, a marked divergence in CYP3A4 for the CYP3A4*1G allele. We also observed low levels of linkage between CYP3A4*1G and CYP3A5*1 in the CSKT. The combination of nonfunctional CYP3A5*3 and putative reduced function CYP3A4*1G alleles may predict diminished clearance of CYP3A substrates. These results highlight the importance of carrying out pharmacogenomic research in AI/AN populations and show that extrapolation from other populations is not appropriate. This information could help optimize drug therapy for the CSKT population.


International Prevalence and Mechanisms of SARS-CoV-2 in Childhood Arterial Ischemic Stroke During the COVID-19 Pandemic.
Data from the early pandemic revealed that 0.62% of children hospitalized with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) had an acute arterial ischemic stroke (AIS). In a larger cohort from June 2020 to December 2020, we sought to determine whether our initial point estimate was stable as the pandemic continued and to understand radiographic and laboratory data that may clarify mechanisms of pediatric AIS in the setting of SARS-CoV-2. We surveyed international sites with pediatric stroke expertise to determine numbers of hospitalized SARS-CoV-2 patients <18 years, numbers of incident AIS cases among children (29 days to <18 years), frequency of SARS-CoV-2 testing for children with AIS, and numbers of childhood AIS cases positive for SARS-CoV-2 June 1 to December 31, 2020. Two stroke neurologists with 1 neuroradiologist determined whether SARS-CoV-2 was the main stroke risk factor, contributory, or incidental. Sixty-one centers from 21 countries provided AIS data. Forty-eight centers (78.7%) provided SARS-CoV-2 hospitalization data. SARS-CoV-2 testing was performed in 335/373 acute AIS cases (89.8%) compared with 99/166 (59.6%) in March to May 2020, P<0.0001. Twenty-three of 335 AIS cases tested (6.9%) were positive for SARS-CoV-2 compared with 6/99 tested (6.1%) in March to May 2020, P=0.78. Of the 22 of 23 AIS cases with SARS-CoV-2 in whom we could collect additional data, SARS-CoV-2 was the main stroke risk factor in 6 (3 with arteritis/vasculitis, 3 with focal cerebral arteriopathy), a contributory factor in 13, and incidental in 3. Elevated inflammatory markers were common, occurring in 17 (77.3%). From centers with SARS-CoV-2 hospitalization data, of 7231 pediatric patients hospitalized with SARS-CoV-2, 23 had AIS (0.32%) compared with 6/971 (0.62%) from March to May 2020, P=0.14. The risk of AIS among children hospitalized with SARS-CoV-2 appeared stable compared with our earlier estimate. Among children in whom SARS-CoV-2 was considered the main stroke risk factor, inflammatory arteriopathies were the stroke mechanism.


Isolation of Methyl Gallate from Toxicodendron sylvestre and Its Effect on Tomato Bacterial Wilt.
We determined that the methanol extract of Toxicodendron sylvestre could significantly inhibit Ralstonia solanacearum in vitro and in planta. One compound responsible for the antibacterial activity was isolated from the ethyl ether extract and identified as methyl gallate (MG) based on its 1H and 13C nuclear magnetic resonance data as well as mass spectroscopy. MG displayed broad-spectrum activity against plant-pathogenic bacteria, and strong inhibitory effects on the growth of plant-pathogenic oomycetes. The half inhibition concentration of MG on R. solanacearum was 8.3 mg/liter, which was tested by the agar dilution method. Disease control trials in planta showed that both natural and synthetic MG could effectively reduce the incidence of tomato bacterial wilt, and there was no significant difference between them in control efficacy. This is the first report on the use of MG for the control of a plant bacterial disease. Because of its broad-spectrum antimicrobial activity and significant control efficacy on tomato bacterial wilt in planta, MG shows potential to be used as a bactericide to control plant bacterial wilt.


Reproducibility of interictal spike propagation in children with refractory epilepsy.
Interictal spikes are a characteristic feature of invasive electroencephalography (EEG) recordings in children with refractory epilepsy. Spikes frequently co-occur across multiple brain regions with discernable latencies, suggesting that spikes can propagate through distributed neural networks. The purpose of this study was to examine the long-term reproducibility of spike propagation patterns over hours to days of interictal recording. Twelve children (mean age 13.1 years) were retrospectively studied. A mean ± standard deviation (SD) of 47.2 ± 40.1 hours of interictal EEG recordings were examined per patient (range 17.5-166.5 hours). Interictal recordings were divided into 30-minute segments. Networks were extracted based on the frequency of spike coactivation between pairs of electrodes. For each 30-minute segment, electrodes were assigned a "Degree Preference (DP)" based on the tendency to appear upstream or downstream within propagation sequences. The consistency of DPs across segments ("DP-Stability") was quantified using the Spearman rank correlation. Regions exhibited highly stable preferences to appear upstream, intermediate, or downstream in spike propagation sequences. Across networks, the mean ± SD DP-Stability was 0.88 ± 0.07, indicating that propagation patterns observed in 30-minute segments were representative of the patterns observed in the full interictal window. At the group level, regions involved in seizure generation appeared more upstream in spike propagation sequences. Interictal spike propagation is a highly reproducible output of epileptic networks. These findings shed new light on the spatiotemporal dynamics that may constrain the network mechanisms of refractory epilepsy.


Effects of electrospray mechanisms and structural relaxation on polylactide ion conformations in the gas phase: insights from ion mobility spectrometry and molecular dynamics simulations.
Recent advances in molecular dynamics (MD) simulations have made it possible to examine the behavior of large charged droplets that contain analytes such as proteins or polymers, thereby providing insights into electrospray ionization (ESI) mechanisms. In the present study, we use this approach to investigate the release of polylactide (PLA) ions from water/acetonitrile ESI droplets. We found that cationized gaseous PLA ions can be formed via various competing pathways. Some MD runs showed extrusion and subsequent separation of polymer chains from the droplet, as envisioned by the chain ejection model (CEM). On other occasions the PLA chains remained inside the droplets and were released after solvent evaporation to dryness, consistent with the charge residue model (CRM). Following their release from ESI droplets, the nascent gaseous PLA ions were subjected to structural relaxation for several μs in vacuo. The MD conformations generated in this way for various PLA charge states compared favorably to experimental results obtained by ion mobility spectrometry-mass spectrometry (IMS-MS). The structures of all PLA ions evolved during relaxation in the gas phase. However, some macroion species retained features that resembled their nascent structures. For this subset of ions, the IMS-MS response appears to be strongly correlated with the ESI release mechanism (CEM vs. CRM). The former favored extended structures, whereas the latter preferentially generated compact conformers.


High frequency plasmid recombination mediated by 28 bp direct repeats.
The stability in Escherichia coli of a mammalian expression vector (pCIneo) and its derivative candidate DNA vaccine (pGPV-PV) is described. These multicopy pMB1-type plasmids were found to recombine in several recA E. coli strains due to the presence of two 28 bp direct repeats flanking intervening sequences of 1.6 kb (pCIneo) and 3.2 kb (pGPV-PV). In this recombination event, one of the direct repeats and the intervening sequence were deleted or duplicated, originating monomeric or/and hetero-dimeric plasmid forms, respectively. Additionally, the plasmid rearrangement led to the acquisition of a kanamycin resistance phenotype. Recombination frequencies between 7.8 x 10(-7) and 3.1 x 10(-5) were determined for DH5alpha and JM109(DE3) strains, respectively. Higher recombination frequencies were found in cells previously grown up to stationary growth phase being the monomeric plasmid form the prevalent one. Real-time PCR quantification revealed the presence of approximately 1.5 x 10(4) recombined molecules per 2 x 10(5 )cells pre-kanamycin exposure. Under selective pressure of this antibiotic, the number of recombined molecules increased approximately 2,000-fold in a 48-h period replacing the original plasmid form. The high frequency, at which deletion-duplication occurred in the absence of kanamycin selective pressure, should be regarded as a safety concern. This work highlights the impact of mutational hot spots on expression and cloning plasmid vectors and the need to carefully design plasmid vectors.


Insights into the Activity and Substrate Binding of Xylella fastidiosa Polygalacturonase by Modification of a Unique QMK Amino Acid Motif Using Protein Chimeras.
Polygalacturonases (EC 188.8.131.52) catalyze the random hydrolysis of 1, 4-alpha-D-galactosiduronic linkages in pectate and other galacturonans. Xylella fastidiosa possesses a single polygalacturonase gene, pglA (PD1485), and X. fastidiosa mutants deficient in the production of polygalacturonase are non-pathogenic and show a compromised ability to systemically infect grapevines. These results suggested that grapevines expressing sufficient amounts of an inhibitor of X. fastidiosa polygalacturonase might be protected from disease. Previous work in our laboratory and others have tried without success to produce soluble active X. fastidiosa polygalacturonase for use in inhibition assays. In this study, we created two enzymatically active X. fastidiosa / A. vitis polygalacturonase chimeras, AX1A and AX2A to explore the functionality of X. fastidiosa polygalacturonase in vitro. The AX1A chimera was constructed to specifically test if recombinant chimeric protein, produced in Escherichia coli, is soluble and if the X. fastidiosa polygalacturonase catalytic amino acids are able to hydrolyze polygalacturonic acid. The AX2A chimera was constructed to evaluate the ability of a unique QMK motif of X. fastidiosa polygalacturonase, most polygalacturonases have a R(I/L)K motif, to bind to and allow the hydrolysis of polygalacturonic acid. Furthermore, the AX2A chimera was also used to explore what effect modification of the QMK motif of X. fastidiosa polygalacturonase to a conserved RIK motif has on enzymatic activity. These experiments showed that both the AX1A and AX2A polygalacturonase chimeras were soluble and able to hydrolyze the polygalacturonic acid substrate. Additionally, the modification of the QMK motif to the conserved RIK motif eliminated hydrolytic activity, suggesting that the QMK motif is important for the activity of X. fastidiosa polygalacturonase. This result suggests X. fastidiosa polygalacturonase may preferentially hydrolyze a different pectic substrate or, alternatively, it has a different mechanism of substrate binding than other polygalacturonases characterized to date.


Mechanisms of Yersinia YopO kinase substrate specificity.
Yersinia bacteria cause a range of human diseases, including yersiniosis, Far East scarlet-like fever and the plague. Yersiniae modulate and evade host immune defences through injection of Yersinia outer proteins (Yops) into phagocytic cells. One of the Yops, YopO (also known as YpkA) obstructs phagocytosis through disrupting actin filament regulation processes - inhibiting polymerization-promoting signaling through sequestration of Rac/Rho family GTPases and by using monomeric actin as bait to recruit and phosphorylate host actin-regulating proteins. Here we set out to identify mechanisms of specificity in protein phosphorylation by YopO that would clarify its effects on cytoskeleton disruption. We report the MgADP structure of Yersinia enterocolitica YopO in complex with actin, which reveals its active site architecture. Using a proteome-wide kinase-interacting substrate screening (KISS) method, we identified that YopO phosphorylates a wide range of actin-modulating proteins and located their phosphorylation sites by mass spectrometry. Using artificial substrates we clarified YopO's substrate length requirements and its phosphorylation consensus sequence. These findings provide fresh insight into the mechanism of the YopO kinase and demonstrate that YopO executes a specific strategy targeting actin-modulating proteins, across multiple functionalities, to compete for control of their native phospho-signaling, thus hampering the cytoskeletal processes required for macrophage phagocytosis.


Effect of Helicobacter pylori eradication in the treatment of Japanese patients with chronic idiopathic urticaria.
Infection with Helicobacter pylori has been associated with chronic idiopathic urticaria (CIU). The aim of this study was to investigate the efficacy of H. pylori eradication in the treatment of patients with CIU. Fifty patients with CIU (16 men and 34 women; age 25-75 years) and 100 sex- and age-matched control subjects were enrolled in the study. Presence of IgG antibody to H. pylori was examined by serology. H. pylori-seropositive patients with CIU received endoscopy to confirm H. pylori infection. Patients infected with H. pylori received eradication therapy comprising lansoprazole, amoxicillin, and either clarithromycin or metronidazole. At least 2 months after finishing the eradication therapy, a (13)C-urea breath test was performed, and the effect of eradication therapy on the CIU was scored, using a three-point scale, as complete remission, partial remission, or no improvement. In the 50 patients with CIU, 26 (52%) were H. pylori-seropositive, while 48% (48/100) of the control subjects were seropositive (statistically not significant). Nineteen out of the 26 patients with CIU infected with H. pylori received eradication therapy, and eradication was successful in 17 patients. In the 17 H. pylori-eradicated patients, 6 (35%) had complete remission and 11 (65%) had complete remission or partial remission. On the other hand, in the 9 patients without H. pylori eradication, only 2 (22%) showed partial remission and 7 (78%) had no improvement. Eradication of H. pylori would be a valid choice for patients with CIU, although the prevalence of H. pylori infection is not higher in patients with CIU than it is in controls.


Plasma mediated collagen-I-coating of metal implant materials to improve biocompatibility.
This study describes the collagen-I coating of titanium and steel implants via cold low-pressure gas plasma treatment. To analyze the coatings in terms of biocompatibility osteoblast-like osteosarcoma cells and human leukocytes were cultivated on the metal surfaces. Two different implant materials were assessed (Ti6Al4V, X2CrNiMo18) and four different surface properties were evaluated: (a) plasma pretreated and collagen-I coated implant materials; (b) collagen-I dip-coated without plasma pretreatment; (c) plasma treated but not collagen-I coated; (d) standard implant materials served as control. The different coating characteristics were analyzed by scanning electron microscopy (SEM). For adhesion and viability tests calcein-AM staining of the cells and Alamar blue assays were performed. The quantitative analysis was conducted by computer assisted microfluorophotography and spectrometer measurements. SEM analysis revealed that stable collagen-I coatings could not be achieved on the dip-coated steel and titanium alloys. Only due to pretreatment with low-pressure gas plasma a robust deposition of collagen I on the surface could be achieved. The cell viability and cell attachment rate on the plasma pretreated, collagen coated surfaces was significantly (p < 0.017) increased compared to the non coated surfaces. Gas plasma treatment is a feasible method for the deposition of proteins on metal implant materials resulting in an improved biocompatibility in vitro. (c) 2010 Wiley Periodicals, Inc. J Biomed Mater Res, 2010.


Exposure to high-pH medium increases the incidence and extent of dye coupling between rat hippocampal CA1 pyramidal neurons in vitro.
Previous studies have demonstrated that dye coupling between neurons can be reduced by maneuvers that reduce intracellular pH. However, it is apparent that marked extracellular alkaline- as well as acid-going pH shifts can occur in the mammalian CNS. In light of the fact that an applied change in extracellular pH may produce a damped change in intracellular pH in the same direction, in this study we have examined the effects of exposure to high extracellular pH (achieved by raising [HCO3-] at a constant PCO2) on the incidence of Lucifer yellow dye coupling between CA1 pyramidal neurons in the rat hippocampal slice. Under standard conditions (pH 7.4), 44% of CA1 pyramidal neurons were dye-coupled, and the mean number of neurons stained per injection was 1.62. A marked increase in the incidence (88%) and extent (mean number of neurons stained per injection, 3.25) of dye coupling was observed during exposure to high-pH medium (pH 7.9). Under both standard and high-pH conditions, dye coupling was associated with the ability of CA1 pyramidal neurons to generate bursts of action potentials in response to intracellularly applied depolarizing current pulses. The results provide additional evidence that dye coupling between hippocampal pyramidal neurons may be modulated dynamically and may have implications for the genesis of synchronized epileptiform activity under alkalotic conditions.


Maternal gestational diabetes mellitus and the risk of subsequent pediatric cardiovascular diseases of the offspring: a population-based cohort study with up to 18 years of follow up.
The prevalence of gestational diabetes mellitus (GDM) has been increasing worldwide. We aimed to study the effect of GDM on the risk for childhood cardiovascular morbidity of the offspring. A population-based cohort analysis was performed comparing total and different subtypes of cardiovascular related pediatric diagnoses among offspring of mothers with GDM vs. offspring of mothers with no diabetes. The analysis included all singletons born between the years 1991-2014. Cardiovascular related morbidities included hospitalizations involving a pre-defined set of ICD-9 codes. Mothers with pregestational diabetes, Infants with congenital malformations, multiple gestations, and perinatal deaths were excluded from the analysis. A Cox proportional hazards model was constructed to adjust for confounders. The study population included 216197 newborns which met the inclusion criteria; among them 4.4% (9460) were born to mothers with GDM controlled by diet and exercise (GDM A1) and 0.3% (724) were born to mothers with GDM requiring oral treatment or insulin (GDM A2). A significant association was noted between GDM and the rate of cardiovascular related hospitalizations (0.97 for GDM A2 vs. 0.57 for GDM A1 vs. 0.33 for no GDM, respectively; p < 0.001). The association remained significant and independent for GDM A1 only while adjusting for relevant confounders [adjusted HR = 1.6 (1.2-2.2); p value 0.001]. A significant association is noted between the GDM and the rate of cardiovascular hospitalizations of the offspring. However, in our population GDM A1 is an independent risk factor for pediatric cardiovascular morbidity of the offspring.


Origin and evolution of a new gene expressed in the Drosophila sperm axoneme.
Sdic is a new gene that evolved recently in the lineage of Drosophila melanogaster. It was formed from a duplication and fusion of the gene AnnX, which encodes annexin X, and Cdic, which encodes the intermediate polypeptide chain of the cytoplasmic dynein. The fusion joins AnnX exon 4 with Cdic intron 3, which brings together three putative promoter elements for testes-specific expression of Sdic: the distal conserved element (DCE) and testes-specific element (TSE) are derived from AnnX, and the proximal conserved element (PCE) from Cdic intron 3. Sdic transcription initiates within the PCE, and translation is initiated within the sequence derived from Cdic intron 3, continuing through a 10 base pair insertion that creates a new splice donor site that enables the new coding sequence derived from intron 3 to be joined with the coding sequence of Cdic exon 4. A novel protein is created lacking 100 residues at the amino end that contain sequence motifs essential for the function of cytoplasmic dynein intermediate chains. Instead, the amino end is a hydrophobic region of 16 residues that resembles the amino end of axonemal dynein intermediate chains from other organisms. The downstream portion of Sdic features large deletions eliminating Cdic exons v2 and v3, as well as multiple frameshift deletions or insertions. The new protein becomes incorporated into the tail of the mature sperm and may function as an axonemal dynein intermediate chain. The new Sdic gene is present in about 10 tandem repeats between the wildtype Cdic and AnnX genes located near the base of the X chromosome. The implications of these findings are discussed relative to the origin of new gene functions and the process of speciation.


Mode-dependent loss and gain: statistics and effect on mode-division multiplexing.
In multimode fiber transmission systems, mode-dependent loss and gain (collectively referred to as MDL) pose fundamental performance limitations. In the regime of strong mode coupling, the statistics of MDL (expressed in decibels or log power gain units) can be described by the eigenvalue distribution of zero-trace Gaussian unitary ensemble in the small-MDL region that is expected to be of interest for practical long-haul transmission. Information-theoretic channel capacities of mode-division-multiplexed systems in the presence of MDL are studied, including average and outage capacities, with and without channel state information.


Crystal structure of oxygen-evolving photosystem II from Thermosynechococcus vulcanus at 3.7-A resolution.
Photosystem II (PSII) is a multisubunit membrane protein complex performing light-induced electron transfer and water-splitting reactions, leading to the formation of molecular oxygen. The first crystal structure of PSII from a thermophilic cyanobacterium Thermosynechococcus elongatus was reported recently [Zouni, A., Witt, H. T., Kern, J., Fromme, P., Krauss, N., Saenger, W. & Orth, P. (2001) Nature 409, 739-743)] at 3.8-A resolution. To analyze the PSII structure in more detail, we have obtained the crystal structure of PSII from another thermophilic cyanobacterium, Thermosynechococcus vulcanus, at 3.7-A resolution. The present structure was built on the basis of the sequences of PSII large subunits D1, D2, CP47, and CP43; extrinsic 33- and 12-kDa proteins and cytochrome c550; and several low molecular mass subunits, among which the structure of the 12-kDa protein was not reported previously. This yielded much information concerning the molecular interactions within this large protein complex. We also show the arrangement of chlorophylls and cofactors, including two beta-carotenes recently identified in a region close to the reaction center, which provided important clues to the secondary electron transfer pathways around the reaction center. Furthermore, possible ligands for the Mn-cluster were determined. In particular, the C terminus of D1 polypeptide was shown to be connected to the Mn cluster directly. The structural information obtained here provides important insights into the mechanism of PSII reactions.


Metabolic phenotype in the brothers of women with polycystic ovary syndrome.
Hyperandrogenemia, insulin resistance, and dyslipidemia demonstrate familial aggregation in the female first-degree relatives of women with polycystic ovary syndrome (PCOS), suggesting that these defects are heritable. Hyperandrogenemia also appears to be the male reproductive phenotype. We performed this study to test the hypothesis that brothers of women with PCOS have metabolic defects similar to those of their proband sisters. This was a prospective case-control study performed at four academic medical centers in the U.S. Fasting blood was obtained from 196 non-Hispanic white brothers of women with PCOS and 169 control men of age, BMI, and ethnicity comparable to those of brothers. A separate analysis was performed by study site to assess potential regional variations in metabolic parameters. Overall, brothers of women with PCOS had significantly higher total (P = 0.001) and LDL cholesterol (P = 0.01) as well as triglyceride levels (P = 0.01) compared with control men, although there were regional variations in these differences. There were significant positive correlations between brothers and their sisters with PCOS for total (rho = 0.2, P = 0.009) and LDL cholesterol (rho = 0.3, P = 0.001) and triglyceride (rho = 0.2, P = 0.05) levels. Brothers also had significantly higher fasting insulin levels and homeostatic index of insulin resistance (P = 0.02 for both comparisons) compared with control men. Brothers of women with PCOS have dyslipidemia as well as evidence for insulin resistance similar to that of their proband sisters with PCOS. These findings are consistent with the hypothesis that some metabolic abnormalities in PCOS are heritable and are not sex specific.


Measuring the functional responses of farmland birds: an example for a declining seed-feeding bunting.
1. Many farmland bird species have undergone significant declines. It is important to predict the effect of agricultural change on these birds and their response to conservation measures. This requirement could be met by mechanistic models that predict population size from the optimal foraging behaviour and fates of individuals within populations. A key component of these models is the functional response, the relationship between food and competitor density and feeding rate. 2. This paper describes a method for measuring functional responses of farmland birds, and applies this method to a declining farmland bird, the corn bunting Miliaria calandra L. We derive five alternative models to predict the functional responses of farmland birds and parameterize these for corn bunting. We also assess the minimum sample sizes required to predict accurately the functional response. 3. We show that the functional response of corn bunting can be predicted accurately from a few behavioural parameters (searching rate, handling time, vigilance time) that are straightforward to measure in the field. These parameters can be measured more quickly than the alternative of measuring the functional response directly. 4. While corn bunting violated some of the assumptions of Holling's disk equation (model 1 in our study), it still provided the most accurate fit to the observed feeding rates while remaining the most statistically simple model tested. Our other models may be more applicable to other species, or corn bunting feeding in other locations. 5. Although further tests are required, our study shows how functional responses can be predicted, simplifying the development of mechanistic models of farmland bird populations.


Formation of hydroxyl radical from San Joaquin Valley particles extracted in a cell-free surrogate lung fluid.
Previous studies have suggested that the adverse health effects from ambient particulate matter (PM) are linked to the formation of reactive oxygen species (ROS) by PM in cardiopulmonary tissues. While hydroxyl radical ((•)OH) is the most reactive of the ROS species, there are few quantitative studies of (•)OH generation from PM. Here we report on (•)OH formation from PM collected at an urban (Fresno) and rural (Westside) site in the San Joaquin Valley (SJV) of California. We quantified (•)OH in PM extracts using a cell-free, phosphate-buffered saline (PBS) solution with or without 50μM ascorbate (Asc). The results show that generally the urban Fresno PM generates much more (•)OH than the rural Westside PM. The presence of Asc at a physiologically relevant concentration in the extraction solution greatly enhances (•)OH formation from all the samples. Fine PM (PM(2.5)) generally makes more (•)OH than the corresponding coarse PM (PM(cf), i.e. with diameters of 2.5 to 10 μm) normalized by air volume collected, while the coarse PM typically generates more (•)OH normalized by PM mass. (•)OH production by SJV PM is reduced on average by (97±6)% when the transition metal chelator desferoxamine (DSF) is added to the extraction solution, indicating a dominant role of transition metals. By measuring calibration curves of (•)OH generation from copper and iron, and quantifying copper and iron concentrations in our particle extracts, we find that PBS-soluble copper is primarily responsible for (•)OH production by the SJV PM, while iron often makes a significant contribution. Extrapolating our results to expected burdens of PM-derived (•)OH in human lung lining fluid suggests that typical daily PM exposures in the San Joaquin Valley are unlikely to result in a high amount of pulmonary (•)OH, although high PM events could produce much higher levels of (•)OH, which might lead to cytotoxicity.


Relationship between the forces acting on the horse's back and the movements of rider and horse while walking on a treadmill.
The exact relationship between the saddle pressure pattern during one stride cycle and the movements of horse and rider at the walk are poorly understood and have never been investigated in detail. The movements of rider and horse account for the force distribution pattern under the saddle. Vertical ground reaction forces (GRF), kinematics of horse and rider as well as saddle forces (FS) were measured synchronously in 7 high level dressage horses while being ridden on an instrumented treadmill at walk. Discrete values of the total saddle forces (FStot) were determined for each stride and related to kinematics and GRF. The pressure sensitive mat was divided into halves and sixths to assess the force distribution over the horse's back in more detail. Differences were tested using a one sample t test (P < 0.05). FStot of all the horses showed 3 peaks (P1-P3) and 3 minima (M1-M3) in each half-cycle, which were systematically related to the footfall sequence of the walk. Looking at the halves of the mat, force curves were 50% phase-shifted. The analysis of the FS of the 6 sections showed a clear association to the rider's and horse's movements. The saddle force distribution during an entire stride cycle has a distinct pattern although the force fluctuations of the FStot are small. The forces in the front thirds were clearly related to the movement of the front limbs, those in the mid part to the lateral flexion of the horse's spine and the loading of the hind part was mainly influenced by the axial rotation and lateral bending of the back. These data can be used as a reference for comparing different types of saddle fit.


Genome sequence analysis of in vitro and in vivo phenotypes of Bunyamwera and Ngari virus isolates from northern Kenya.
Biological phenotypes of tri-segmented arboviruses display characteristics that map to mutation/s in the S, M or L segments of the genome. Plaque variants have been characterized for other viruses displaying varied phenotypes including attenuation in growth and/or pathogenesis. In order to characterize variants of Bunyamwera and Ngari viruses, we isolated individual plaque size variants; small plaque (SP) and large plaque (LP) and determined in vitro growth properties and in vivo pathogenesis in suckling mice. We performed gene sequencing to identify mutations that may be responsible for the observed phenotype. The LP generally replicated faster than the SP and the difference in growth rate was more pronounced in Bunyamwera virus isolates. Ngari virus isolates were more conserved with few point mutations compared to Bunyamwera virus isolates which displayed mutations in all three genome segments but majority were silent mutations. Contrary to expectation, the SP of Bunyamwera virus killed suckling mice significantly earlier than the LP. The LP attenuation may probably be due to a non-synonymous substitution (T858I) that mapped within the active site of the L protein. In this study, we identify natural mutations whose exact role in growth and pathogenesis need to be determined through site directed mutagenesis studies.


An assessment of human gastric fluid composition as a function of PPI usage.
The standard of care for chronic gastro-esophageal reflux disease (GERD), which affects up to 40% of the population, is the use of drugs such as proton pump inhibitors (PPIs) that block the production of stomach acid. Despite widespread use, the effects of PPIs on gastric fluid remain poorly characterized. In this study, gastric fluid was collected from patients undergoing cardiac surgery who were not (n = 40) or were (n = 25) actively taking PPIs. Various enzymatic and immunoassays as well as mass spectrometry were utilized to analyze the concentrations of bile, gastricsin, trypsin, and pepsin in the gastric fluid. Proteomic analyses by mass spectrometry suggested that degradation of trypsin at low pH might account, at least in part, for the observation that patients taking PPIs have a greater likelihood of having high concentrations of trypsin in their gastric fluid. In general, the concentrations of all analytes evaluated varied over several orders of magnitude, covering a minimum of a 2000-fold range (gastricsin) and a maximum of a 1 × 10(6) -fold range (trypsin). Furthermore, the concentrations of various analytes were poorly correlated with one another in the samples. For example, trypsin and bile concentrations showed a significant (P < 0.0001) but not strong correlation (r = 0.54). Finally, direct assessment of bacterial concentrations by flow cytometry revealed that PPIs did not cause a profound increase in microbial load in the gastric fluid. These results further delineate the profound effects that PPI usage has on the physiology of the stomach.