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Find video protocols related to scientific articles indexed in Pubmed.
Zygomatic arch fracture in a 40-year-old woman.
Adv Emerg Nurs J
PUBLISHED: 10-31-2014
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This case describes an injury to a 40-year-old woman, employed as a softball team coach, who presented to an emergency department (ED) after sustaining a line drive hit to the right side of the face with a softball during practice. Upon arrival to the ED, the patient complained of moderate pain, swelling, and bruising over the right side of her midface area. After evaluation in the ED, the patient received a diagnosis of a zygomatic arch fracture that was further managed by an occupational medicine nurse practitioner, a plastic surgeon, and an ear, nose, and throat (ENT) physician specialist.
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Cx46 Gap Junctions Provide a Pathway for the Delivery of Glutathione to the Lens Nucleus.
J. Biol. Chem.
PUBLISHED: 10-07-2014
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Maintenance of adequate levels of glutathione (GSH) in the lens nucleus is critical for protection of lens proteins from the effects of oxidative stress and for lens transparency. How GSH is transported to the nucleus is unknown. We show that GSH diffuses to nucleus from the outer cortex, where a high concentration of the anti-oxidant is established by synthesis or uptake, via the network of gap junctions. Using electrophysiological measurements, we found that channels formed by Cx46 and Cx50, the two connexin isoforms expressed in the lens, were moderately cation-selective (PNa:PCl ~ 5:1 for Cx46 and ~ 3:1 for Cx50). Single channel permeation of the larger GSH anion was low but detectable (PNa:PGSH ~ 12:1 for Cx46 and ~ 8:1 for Cx50) whereas permeation of divalent anion glutathione disulfide (GSSG) was undetectable. Measurement of GSH levels in the lenses from connexin knockout (KO) mice indicated Cx46, and not Cx50, is necessary for transport of GSH to the core. Levels of GSH in the nucleus were markedly reduced in Cx46 KO, whereas they were unaffected by Cx50 KO. We also show that GSH delivery to the nucleus is not dependent on the lens microcirculation, which is believed to be responsible for extracellular transport of other nutrients to membrane transporters in the core. These results indicate that glutathione diffuses from cortical fiber cells to the nucleus via gap junction channels formed by Cx46. We present a model of GSH diffusion from outer cells to inner fiber cells through gap junctions.
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Unheard voices: outcomes of tertiary care for treatment-refractory psychosis.
Psychiatr Bull (2014)
PUBLISHED: 09-20-2014
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Aims and method In up to a quarter of patients, schizophrenia is resistant to standard treatments. We undertook a naturalistic study of 153 patients treated in the tertiary referral in-patient unit of the National Psychosis Service based at the Maudsley Hospital in London. A retrospective analysis of symptoms on admission and discharge was undertaken using the OPCRIT tool, along with preliminary economic modelling of potential costs related to changes in accommodation. Results In-patient treatment demonstrated statistically significant improvements in all symptom categories in patients already identified as having schizophrenia refractory to standard secondary care. The preliminary cost analysis showed net savings to referring authorities due to changes from pre- to post-discharge accommodation. Clinical implications Despite the enormous clinical, personal and societal burden of refractory psychotic illnesses, there is insufficient information on the outcomes of specialised tertiary-level care. Our pilot data support its utility in all domains measured.
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Spatiotemporally consistent genomic signatures of reproductive isolation in a moving hybrid zone.
Evolution
PUBLISHED: 09-19-2014
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Studies of hybrid zone dynamics often investigate a single sampling period and draw conclusions from that temporal snapshot. Stochasticity can, however, result in loci with spurious outlier patterns, which is exacerbated by limited temporal or geographic sampling. Comparing admixed populations from different geographic regions is one way to detect repeatedly divergent genomic regions potentially involved in reproductive isolation. Temporal comparisons also allow us to control partially for the role of stochasticity, but the power of temporal sampling has not yet been adequately explored. In North America, black-capped (Poecile atricapillus) and Carolina (P. carolinensis) chickadees hybridize in a contact zone extending from New Jersey to Kansas. The hybrid zone is likely maintained by strong intrinsic selection against hybrids, and it is moving north. We used a reduced representation genomic approach and temporally spaced sampling-two samples of ?80 individuals separated by a decade-to determine the pattern and consistency of selection and genomic introgression in the chickadee hybrid zone. We report consistently low introgression for highly divergent loci between P. atricapillus and P. carolinensis in this moving hybrid zone. This is strong evidence that these loci may be linked to genomic regions involved in reproductive isolation between chickadees.
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Is Sequestrectomy a Viable Alternative to Microdiscectomy? A Systematic Review of the Literature.
Clin. Orthop. Relat. Res.
PUBLISHED: 09-03-2014
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Traditionally, lumbar discectomy involves removal of the free disc fragment followed by aggressive or conservative excision of the intervertebral disc. In selected patients, however, it is possible to remove only the free fragment or sequester without clearing the intervertebral disc space. However, there is some controversy about whether that approach is sufficient to prevent recurrent symptoms and to provide adequate pain relief.
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Visualizing a protein quake with time-resolved X-ray scattering at a free-electron laser.
Nat. Methods
PUBLISHED: 08-10-2014
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We describe a method to measure ultrafast protein structural changes using time-resolved wide-angle X-ray scattering at an X-ray free-electron laser. We demonstrated this approach using multiphoton excitation of the Blastochloris viridis photosynthetic reaction center, observing an ultrafast global conformational change that arises within picoseconds and precedes the propagation of heat through the protein. This provides direct structural evidence for a 'protein quake': the hypothesis that proteins rapidly dissipate energy through quake-like structural motions.
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Eluding the illusion? Schizophrenia, dopamine and the McGurk effect.
Front Hum Neurosci
PUBLISHED: 08-05-2014
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Perceptions are inherently probabilistic; and can be potentially manipulated to induce illusory experience by the presentation of ambiguous or improbable evidence under selective (spatio-temporal) constraints. Accordingly, perception of the McGurk effect, by which individuals misperceive specific incongruent visual and auditory vocal cues, rests upon effective probabilistic inference. Here, we report findings from a behavioral investigation of illusory perception and related metacognitive evaluation during audiovisual integration, conducted in individuals with schizophrenia (n = 30) and control subjects (n = 24) matched in terms of age, sex, handedness and parental occupation. Controls additionally performed the task after an oral dose of amisulpride (400 mg). Individuals with schizophrenia were observed to exhibit illusory perception less frequently than controls, despite non-significant differences in perceptual performance during control conditions. Furthermore, older individuals with schizophrenia exhibited reduced rates of illusory perception. Subsequent analysis revealed a robust inverse relationship between illness chronicity and the illusory perception rate in this group. Controls demonstrated non-significant modulation of perception by amisulpride; amisulpride was, however, found to elicit increases in subjective confidence in perceptual performance. Overall, these findings are consistent with the idea that impairments in probabilistic inference are exhibited in schizophrenia and exacerbated by illness chronicity. The latter suggests that associated processes are a potentially worthwhile target for therapeutic intervention.
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Examining educational attainment, prepregnancy smoking rate, and delay discounting as predictors of spontaneous quitting among pregnant smokers.
Exp Clin Psychopharmacol
PUBLISHED: 07-28-2014
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We investigated three potential predictors (educational attainment, prepregnancy smoking rate, and delay discounting [DD]) of spontaneous quitting among pregnant smokers. These predictors were examined alone and in combination with other potential predictors using study-intake assessments from controlled clinical trials examining the efficacy of financial incentives for smoking cessation and relapse prevention. Data from 349 pregnant women (231 continuing smokers and 118 spontaneous quitters) recruited from the greater Burlington, VT, area contributed to this secondary analysis, including psychiatric/sociodemographic characteristics, smoking characteristics, and performance on a computerized DD task. Educational attainment, smoking rate, and DD values were each significant predictors of spontaneous quitting in univariate analyses. A model examining those three predictors together retained educational attainment as a main effect and revealed a significant interaction of DD and smoking rate (i.e., DD was a significant predictor at lower but not higher smoking rates). A final model considering all potential predictors, included education, the interaction of DD and smoking rate, and five additional predictors (i.e., stress ratings, the belief that smoking during pregnancy will "greatly harm my baby," age of smoking initiation, marital status, and prior quit attempts during pregnancy). The study presented here contributes new knowledge on predictors of spontaneous quitting among pregnant smokers with substantive practical implications for reducing smoking during pregnancy.
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The female urinary microbiome: a comparison of women with and without urgency urinary incontinence.
MBio
PUBLISHED: 07-10-2014
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Bacterial DNA and live bacteria have been detected in human urine in the absence of clinical infection, challenging the prevailing dogma that urine is normally sterile. Urgency urinary incontinence (UUI) is a poorly understood urinary condition characterized by symptoms that overlap urinary infection, including urinary urgency and increased frequency with urinary incontinence. The recent discovery of the urinary microbiome warrants investigation into whether bacteria contribute to UUI. In this study, we used 16S rRNA gene sequencing to classify bacterial DNA and expanded quantitative urine culture (EQUC) techniques to isolate live bacteria in urine collected by using a transurethral catheter from women with UUI and, in comparison, a cohort without UUI. For these cohorts, we demonstrated that the UUI and non-UUI urinary microbiomes differ by group based on both sequence and culture evidences. Compared to the non-UUI microbiome, sequencing experiments revealed that the UUI microbiome was composed of increased Gardnerella and decreased Lactobacillus. Nine genera (Actinobaculum, Actinomyces, Aerococcus, Arthrobacter, Corynebacterium, Gardnerella, Oligella, Staphylococcus, and Streptococcus) were more frequently cultured from the UUI cohort. Although Lactobacillus was isolated from both cohorts, distinctions existed at the species level, with Lactobacillus gasseri detected more frequently in the UUI cohort and Lactobacillus crispatus most frequently detected in controls. Combined, these data suggest that potentially important differences exist in the urinary microbiomes of women with and without UUI, which have strong implications in prevention, diagnosis, or treatment of UUI. Importance: New evidence indicates that the human urinary tract contains microbial communities; however, the role of these communities in urinary health remains to be elucidated. Urgency urinary incontinence (UUI) is a highly prevalent yet poorly understood urinary condition characterized by urgency, frequency, and urinary incontinence. Given the significant overlap of UUI symptoms with those of urinary tract infections, it is possible that UUI may have a microbial component. We compared the urinary microbiomes of women affected by UUI to those of a comparison group without UUI, using both high-throughput sequencing and extended culture techniques. We identified statistically significant differences in the frequency and abundance of bacteria present. These differences suggest a potential role for the urinary microbiome in female urinary health.
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Room-temperature macromolecular serial crystallography using synchrotron radiation.
IUCrJ
PUBLISHED: 07-01-2014
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A new approach for collecting data from many hundreds of thousands of microcrystals using X-ray pulses from a free-electron laser has recently been developed. Referred to as serial crystallography, diffraction patterns are recorded at a constant rate as a suspension of protein crystals flows across the path of an X-ray beam. Events that by chance contain single-crystal diffraction patterns are retained, then indexed and merged to form a three-dimensional set of reflection intensities for structure determination. This approach relies upon several innovations: an intense X-ray beam; a fast detector system; a means to rapidly flow a suspension of crystals across the X-ray beam; and the computational infrastructure to process the large volume of data. Originally conceived for radiation-damage-free measurements with ultrafast X-ray pulses, the same methods can be employed with synchrotron radiation. As in powder diffraction, the averaging of thousands of observations per Bragg peak may improve the ratio of signal to noise of low-dose exposures. Here, it is shown that this paradigm can be implemented for room-temperature data collection using synchrotron radiation and exposure times of less than 3?ms. Using lysozyme microcrystals as a model system, over 40?000 single-crystal diffraction patterns were obtained and merged to produce a structural model that could be refined to 2.1?Å resolution. The resulting electron density is in excellent agreement with that obtained using standard X-ray data collection techniques. With further improvements the method is well suited for even shorter exposures at future and upgraded synchrotron radiation facilities that may deliver beams with 1000 times higher brightness than they currently produce.
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Examining vulnerability to smokeless tobacco use among adolescents and adults meeting diagnostic criteria for major depressive disorder.
Exp Clin Psychopharmacol
PUBLISHED: 06-30-2014
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Smoking prevalence is unevenly distributed in the U.S. population, with those with mental illness, other substance use disorders, and lower socioeconomic status being especially vulnerable. Less research has been conducted on the association between these same vulnerabilities and smokeless tobacco (ST) use. The present study examined cigarette and ST use among adolescents and adults who met diagnostic criteria for major depressive disorder in the National Survey on Drug Use and Health (NSDUH). Utilizing the most recent (2011) NSDUH, we compared odds for current cigarette smoking and ST use among adolescents and adults meeting criteria for past-year major depressive disorder to the general population, after adjusting for potential confounding influences of sociodemographic and other substance use characteristics. Analyses were conducted to examine sex as a moderator of the relation between major depressive disorder and tobacco use. Odds for current cigarette smoking among those classified with major depressive disorder were increased among adolescents (OR = 1.33, 95% CI [1.05, 1.69], p = .021) and adults (OR = 1.70, 95% CI [1.47, 1.97], p < .0005), and odds for current ST use did not differ among adolescents (OR = 0.90, 95% CI [0.54, 1.49], p = .678) and were lower among adults (OR = 0.68, 95% CI [0.51, 0.91], p = .010). Sex was not a significant moderator in adolescents or adults. Major depressive disorder is associated with increased risk for smoking but not ST use among adolescents and adults further demonstrating heterogeneity in predictors of vulnerability to use of different tobacco products.
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Mapping the continuous reciprocal space intensity distribution of X-ray serial crystallography.
Philos. Trans. R. Soc. Lond., B, Biol. Sci.
PUBLISHED: 06-11-2014
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Serial crystallography using X-ray free-electron lasers enables the collection of tens of thousands of measurements from an equal number of individual crystals, each of which can be smaller than 1 µm in size. This manuscript describes an alternative way of handling diffraction data recorded by serial femtosecond crystallography, by mapping the diffracted intensities into three-dimensional reciprocal space rather than integrating each image in two dimensions as in the classical approach. We call this procedure 'three-dimensional merging'. This procedure retains information about asymmetry in Bragg peaks and diffracted intensities between Bragg spots. This intensity distribution can be used to extract reflection intensities for structure determination and opens up novel avenues for post-refinement, while observed intensity between Bragg peaks and peak asymmetry are of potential use in novel direct phasing strategies.
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Phasing coherently illuminated nanocrystals bounded by partial unit cells.
Philos. Trans. R. Soc. Lond., B, Biol. Sci.
PUBLISHED: 06-11-2014
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With the use of highly coherent femtosecond X-ray pulses from a free-electron laser, it is possible to record protein nanocrystal diffraction patterns with far more information than is present in conventional crystallographic diffraction data. It has been suggested that diffraction phases may be retrieved from such data via iterative algorithms, without the use of a priori information and without restrictions on resolution. Here, we investigate the extension of this approach to nanocrystals with edge terminations that produce partial unit cells, and hence cannot be described by a common repeating unit cell. In this situation, the phase problem described in previous work must be reformulated. We demonstrate an approximate solution to this phase problem for crystals with random edge terminations.
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Post-refinement method for snapshot serial crystallography.
Philos. Trans. R. Soc. Lond., B, Biol. Sci.
PUBLISHED: 06-11-2014
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A post-refinement procedure has been devised for 'snapshot' diffraction data consisting entirely of partially recorded reflections, each diffraction pattern from a crystal in an orientation unrelated to the others. Initial estimates of the diffraction geometry are used to calculate initial partialities, which are then used to scale the entire dataset together to produce initial estimates of the fully integrated intensities. The geometrical parameters for each pattern are then refined to maximize the agreement between these estimates and the calculated intensities in each pattern, and the procedure repeated iteratively. The performance of the procedure was investigated using simulated data and found to yield a significant improvement in the data quality.
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Medial scapular winging associated with rib fractures and plating corrected with pectoralis major transfer.
Int J Surg Case Rep
PUBLISHED: 06-09-2014
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Rib plating is becoming increasingly common as a method for stabilizing a flail chest resulting from multiple rib fractures. Recent guidelines recommend surgical stabilization of a flail chest based on consistent evidence of its efficacy and lack of major safety concerns. But complications of this procedure can occur and are wide ranging.
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Cheetah: software for high-throughput reduction and analysis of serial femtosecond X-ray diffraction data.
J Appl Crystallogr
PUBLISHED: 06-01-2014
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The emerging technique of serial X-ray diffraction, in which diffraction data are collected from samples flowing across a pulsed X-ray source at repetition rates of 100?Hz or higher, has necessitated the development of new software in order to handle the large data volumes produced. Sorting of data according to different criteria and rapid filtering of events to retain only diffraction patterns of interest results in significant reductions in data volume, thereby simplifying subsequent data analysis and management tasks. Meanwhile the generation of reduced data in the form of virtual powder patterns, radial stacks, histograms and other meta data creates data set summaries for analysis and overall experiment evaluation. Rapid data reduction early in the analysis pipeline is proving to be an essential first step in serial imaging experiments, prompting the authors to make the tool described in this article available to the general community. Originally developed for experiments at X-ray free-electron lasers, the software is based on a modular facility-independent library to promote portability between different experiments and is available under version 3 or later of the GNU General Public License.
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Aberrant Connexin26 Hemichannels Underlying Keratitis-Ichthyosis-Deafness Syndrome Are Potently Inhibited by Mefloquine.
J. Invest. Dermatol.
PUBLISHED: 05-29-2014
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Keratitis-ichthyosis-deafness (KID) syndrome is an ectodermal dysplasia caused by dominant mutations of connexin26 (Cx26). Loss of Cx26 function causes nonsyndromic sensorineural deafness, without consequence in the epidermis. Functional analyses have revealed that a majority of KID-causing mutations confer a novel expansion of hemichannel activity, mediated by connexin channels in a nonjunctional configuration. Inappropriate Cx26 hemichannel opening is hypothesized to compromise keratinocyte integrity and epidermal homeostasis. Pharmacological modulators of Cx26 are needed to assess the pathomechanistic involvement of hemichannels in the development of hyperkeratosis in KID syndrome. We have used electrophysiological assays to evaluate small-molecule analogs of quinine for suppressive effects on aberrant hemichannel currents elicited by KID mutations. Here, we show that mefloquine (MFQ) inhibits several mutant hemichannel forms implicated in KID syndrome when expressed in Xenopus laevis oocytes (IC50?16??M), using an extracellular divalent cation, zinc (Zn(++)), as a nonspecific positive control for comparison (IC50?3??M). Furthermore, we used freshly isolated transgenic keratinocytes to show that micromolar concentrations of MFQ attenuated increased macroscopic membrane currents in primary mouse keratinocytes expressing human Cx26-G45E, a mutation that causes a lethal form of KID syndrome.Journal of Investigative Dermatology advance online publication, 16 October 2014; doi:10.1038/jid.2014.408.
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Signal design and courtship presentation coincide for highly biased delivery of an iridescent butterfly mating signal.
Evolution
PUBLISHED: 05-27-2014
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Sensory drive theory contends that signalling systems should evolve to optimize transmission between senders and intended receivers, while minimising visibility to eavesdroppers where possible. In visual communication systems, the high directionality afforded by iridescent colouration presents underappreciated avenues for mediating this trade-off. This hypothesis predicts functional links between signal design and presentation such that visual conspicuousness is maximised only under ecologically relevant settings and/or to select audiences. We addressed this prediction using Hypolimnas bolina, a butterfly in which males possess ultraviolet markings on their dorsal wing surfaces with a narrow angular reflectance function. Males bearing brighter dorsal markings are increasingly attractive to females, but also likely more conspicuous to predators. Our data indicate that, during courtship (and given the ritualized wingbeat dynamics at these times), males position themselves relative to females in such a way as to simultaneously maximise three components of known or putative signal conspicuousness: brightness, area, and iridescent flash. This suggests that male signal design and display have coevolved for the delivery of an optimally conspicuous signal to courted females. More broadly, these findings imply a potential signalling role for iridescence itself, and pose a novel example for how signal design may coevolve with the behavioural context of display. This article is protected by copyright. All rights reserved.
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Gene flow and the maintenance of species boundaries.
Mol. Ecol.
PUBLISHED: 05-06-2014
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Hybrid zones are regions where individuals from genetically differentiated populations meet and mate, resulting in at least some offspring of mixed ancestry. Patterns of gene flow (introgression) in hybrid zones vary across the genome, allowing assessment of the role of individual genes or genome regions in reproductive isolation. Here, we document patterns of introgression between two recently diverged species of field crickets. We sampled at a very fine spatial scale and genotyped crickets for 110 highly differentiated single nucleotide polymorphisms (SNPs) identified through transcriptome scans. Using both genomic and geographic cline analysis, we document remarkably abrupt transitions (<100 m) in allele frequencies for 50 loci, despite high levels of gene flow at other loci. These are among the steepest clines documented for any hybridizing taxa. Furthermore, the cricket hybrid zone provides one of the clearest examples of the semi-permeability of species boundaries. Comparisons between data from the fine-scale transect and data (for the same set of markers) from sampling a much larger area in a different region of the cricket hybrid zone reveal consistent patterns of introgression for individual loci. The consistency in patterns of introgression between these two distant and distinct regions of the hybrid zone suggests that strong selection is acting to maintain abrupt discontinuities within the hybrid zone and that genomic regions with restricted introgression likely include genes that contribute to nonecological prezygotic barriers.
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Serial time-resolved crystallography of photosystem II using a femtosecond X-ray laser.
Nature
PUBLISHED: 05-04-2014
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Photosynthesis, a process catalysed by plants, algae and cyanobacteria converts sunlight to energy thus sustaining all higher life on Earth. Two large membrane protein complexes, photosystem I and II (PSI and PSII), act in series to catalyse the light-driven reactions in photosynthesis. PSII catalyses the light-driven water splitting process, which maintains the Earth's oxygenic atmosphere. In this process, the oxygen-evolving complex (OEC) of PSII cycles through five states, S0 to S4, in which four electrons are sequentially extracted from the OEC in four light-driven charge-separation events. Here we describe time resolved experiments on PSII nano/microcrystals from Thermosynechococcus elongatus performed with the recently developed technique of serial femtosecond crystallography. Structures have been determined from PSII in the dark S1 state and after double laser excitation (putative S3 state) at 5 and 5.5 Å resolution, respectively. The results provide evidence that PSII undergoes significant conformational changes at the electron acceptor side and at the Mn4CaO5 core of the OEC. These include an elongation of the metal cluster, accompanied by changes in the protein environment, which could allow for binding of the second substrate water molecule between the more distant protruding Mn (referred to as the 'dangler' Mn) and the Mn3CaOx cubane in the S2 to S3 transition, as predicted by spectroscopic and computational studies. This work shows the great potential for time-resolved serial femtosecond crystallography for investigation of catalytic processes in biomolecules.
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Sex differences in COMT polymorphism effects on prefrontal inhibitory control in adolescence.
Neuropsychopharmacology
PUBLISHED: 04-03-2014
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Catecholamine-0-methyl-transferase (COMT) gene variation effects on prefrontal blood oxygenation-level-dependent (BOLD) activation are robust; however, despite observations that COMT is estrogenically catabolized, sex differences in its prefrontal repercussions remain unclear. Here, in a large sample of healthy adolescents stratified by sex and Val(158)Met genotype (n=1133), we examine BOLD responses during performance of the stop-signal task in right-hemispheric prefrontal regions fundamental to inhibitory control. A significant sex-by-genotype interaction was observed in pre-SMA during successful-inhibition trials and in both pre-SMA and inferior frontal cortex during failed-inhibition trials with Val homozygotes displaying elevated activation compared with other genotypes in males but not in females. BOLD activation in the same regions significantly mediated the relationship between COMT genotype and inhibitory proficiency as indexed by stop-signal reaction time in males alone. These sexually dimorphic effects of COMT on inhibitory brain activation have important implications for our understanding of the contrasting patterns of prefrontally governed psychopathology observed in males and females.
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Ketamine as the prototype glutamatergic antidepressant: pharmacodynamic actions, and a systematic review and meta-analysis of efficacy.
Ther Adv Psychopharmacol
PUBLISHED: 04-02-2014
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The burden of depressive disorders and the frequent inadequacy of their current pharmacological treatments are well established. The anaesthetic and hallucinogenic drug ketamine has provoked much interest over the past decade or so as an extremely rapidly acting antidepressant that does not modify 'classical' monoaminergic receptors. Current evidence has shown several ways through which it might exert therapeutic antidepressant actions: blockade of glutamatergic NMDA receptors and relative upregulation of ?-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) subtypes may alter cortical connectivity patterns; through intracellular changes in protein expression, including the proteins mammalian target of rapamycin (mTOR) and brain-derived neurotrophic factor (BDNF); and alteration of intracellular signalling cascades. The clinical evidence demonstrates rapid improvements in mood and suicidal thinking in most participants, although study numbers have generally been small and many trials are unblinded and methodologically weak. There is a small body of work to suggest ketamine might also augment electroconvulsive therapy and potentially have a role as a surgical anaesthetic in depressed patients. A major problem is that the effects of ketamine appear temporary, disappearing after days to weeks (although longer benefits have been sustained in some), and attempts to circumvent this through pharmacological augmentation have been disappointing thus far. These exciting data are providing new insights into neurobiological models of depression, and potentially opening up a new class of antidepressants, but there are significant practical and ethical issues about any future mainstream clinical role it might have.
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Functional magnetic resonance imaging of impaired sensory prediction in schizophrenia.
JAMA Psychiatry
PUBLISHED: 03-04-2014
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Forward models predict the sensory consequences of planned actions and permit discrimination of self- and non-self-elicited sensation; their impairment in schizophrenia is implied by an abnormality in behavioral force-matching and the flawed agency judgments characteristic of positive symptoms, including auditory hallucinations and delusions of control.
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Liver-specific GH receptor gene-disrupted (LiGHRKO) mice have decreased endocrine IGF-I, increased local IGF-I, and altered body size, body composition, and adipokine profiles.
Endocrinology
PUBLISHED: 02-11-2014
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GH is an important regulator of body growth and composition as well as numerous other metabolic processes. In particular, liver plays a key role in the GH/IGF-I axis, because the majority of circulating "endocrine" IGF-I results from GH-stimulated liver IGF-I production. To develop a better understanding of the role of liver in the overall function of GH, we generated a strain of mice with liver-specific GH receptor (GHR) gene knockout (LiGHRKO mice). LiGHRKO mice had a 90% decrease in circulating IGF-I levels, a 300% increase in circulating GH, and significant changes in IGF binding protein (IGFBP)-1, IGFBP-2, IGFBP-3, IGFBP-5, and IGFBP-7. LiGHRKO mice were smaller than controls, with body length and body weight being significantly decreased in both sexes. Analysis of body composition over time revealed a pattern similar to those found in GH transgenic mice; that is, LiGHRKO mice had a higher percentage of body fat at early ages followed by lower percentage of body fat in adulthood. Local IGF-I mRNA levels were significantly increased in skeletal muscle and select adipose tissue depots. Grip strength was increased in LiGHRKO mice. Finally, circulating levels of leptin, resistin, and adiponectin were increased in LiGHRKO mice. In conclusion, LiGHRKO mice are smaller despite increased local mRNA expression of IGF-I in several tissues, suggesting that liver-derived IGF-I is indeed important for normal body growth. Furthermore, our data suggest that novel GH-dependent cross talk between liver and adipose is important for regulation of adipokines in vivo.
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Physiologic and metabolic safety of butyrylcholinesterase gene therapy in mice.
Vaccine
PUBLISHED: 02-06-2014
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In continuing efforts to develop gene transfer of human butyrylcholinesterase (BChE) as therapy for cocaine addiction, we conducted wide-ranging studies of physiological and metabolic safety. For that purpose, mice were given injections of adeno-associated virus (AAV) vector or helper-dependent adenoviral (hdAD) vector encoding human or mouse BChE mutated for optimal cocaine hydrolysis. Age-matched controls received saline or AAV-luciferase control vector. At times when transduced BChE was abundant, physiologic and metabolic parameters in conscious animals were evaluated by non-invasive Echo-MRI and an automated "Comprehensive Laboratory Animal Monitoring System" (CLAMS). Despite high vector doses (up to 10(13) particles per mouse) and high levels of transgene protein in the plasma (?1500-fold above baseline), the CLAMS apparatus revealed no adverse physiologic or metabolic effects. Likewise, body composition determined by Echo-MRI, and glucose tolerance remained normal. A CLAMS study of vector-treated mice given 40 mg/kg cocaine showed none of the physiologic and metabolic fluctuations exhibited in controls. We conclude that neither the tested vectors nor great excesses of circulating BChE affect general physiology directly, while they protect mice from disturbance by cocaine. Hence, viral gene transfer of BChE appears benign and worth exploring as a therapy for cocaine abuse and possibly other disorders as well.
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Genomic and morphological analysis of a semipermeable avian hybrid zone suggests asymmetrical introgression of a sexual signal.
Evolution
PUBLISHED: 01-31-2014
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Hybrid zones are geographic regions where differentiated taxa meet and potentially exchange genes. Increasingly, genomic analyses have demonstrated that many hybrid zones are semipermeable boundaries across which introgression is highly variable. In some cases, certain alleles penetrate across the hybrid zone in only one direction, recombining into the alternate genome. We investigated this phenomenon using genomic (genotyping-by-sequencing) and morphological (plumage reflectance spectrophotometry) analyses of the hybrid zone between two subspecies of the red-backed fairy-wren (Malurus melanocephalus) that differ conspicuously in a sexual signal, male back plumage color. Geographic cline analyses revealed a highly variable pattern of differential introgression, with many narrow coincident clines combined with several significantly wider clines, suggesting that the hybrid zone is a semipermeable tension zone. The plumage cline was shifted significantly into the genomic background of the orange subspecies, consistent with sexual selection driving asymmetrical introgression of red plumage alleles across the hybrid zone. This interpretation is supported by previous experimental work demonstrating an extra-pair mating advantage for red males, but the role of genetic dominance in driving this pattern remains unclear. This study highlights the potential for sexual selection to erode taxonomic boundaries and promote gene flow, particularly at an intermediate stage of divergence.
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Climate-mediated movement of an avian hybrid zone.
Curr. Biol.
PUBLISHED: 01-27-2014
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The interaction between sibling species that share a zone of contact is a multifaceted relationship affected by climate change [1, 2]. Between sibling species, interactions may occur at whole-organism (direct or indirect competition) or genomic (hybridization and introgression) levels [3-5]. Tracking hybrid zone movements can provide insights about influences of environmental change on species interactions [1]. Here, we explore the extent and mechanism of movement of the contact zone between black-capped chickadees (Poecile atricapillus) and Carolina chickadees (Poecile carolinensis) at whole-organism and genomic levels. We find strong evidence that winter temperatures limit the northern extent of P. carolinensis by demonstrating a current-day association between the range limit of this species and minimum winter temperatures. We further show that this temperature limitation has been consistent over time because we are able to accurately hindcast the previous northern range limit under earlier climate conditions. Using genomic data, we confirm northward movement of this contact zone over the past decade and highlight temporally consistent differential-but limited-geographic introgression of alleles. Our results provide an informative example of the influence of climate change on a contact zone between sibling species.
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Lipidic cubic phase injector facilitates membrane protein serial femtosecond crystallography.
Nat Commun
PUBLISHED: 01-24-2014
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Lipidic cubic phase (LCP) crystallization has proven successful for high-resolution structure determination of challenging membrane proteins. Here we present a technique for extruding gel-like LCP with embedded membrane protein microcrystals, providing a continuously renewed source of material for serial femtosecond crystallography. Data collected from sub-10-?m-sized crystals produced with less than 0.5 mg of purified protein yield structural insights regarding cyclopamine binding to the Smoothened receptor.
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Myostatin and sarcopenia: opportunities and challenges - a mini-review.
Gerontology
PUBLISHED: 01-17-2014
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The progressive loss of skeletal muscle mass, strength and/or function with advancing age, termed sarcopenia, poses a major threat to independence and quality of life. Therefore, there is significant merit in better understanding the biology of sarcopenia and developing therapeutic interventions to prevent, slow or reverse its progression. Since the discovery of myostatin, a potent negative regulator of growth that is highly enriched in skeletal muscle, there has been great interest in it as a potential mediator of sarcopenia as well as a therapeutic target. The complex biology of myostatin, the promise of myostatin inhibition as an effective means to counter sarcopenia, and the challenges facing its clinical translation are reviewed herein.
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Uncertainty and confidence from the triple-network perspective: voxel-based meta-analyses.
Brain Cogn
PUBLISHED: 01-11-2014
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Our subjective confidence about particular events is related to but independent from the objective certainty of the stimuli we encounter. Surprisingly, previous investigations of the neurophysiological correlates of confidence and uncertainty have largely been carried out separately. After systematically reviewing the blood oxygenation-level dependent functional magnetic resonance imaging (BOLD fMRI) literature, and splitting studies on the basis of their task requirements, a voxel-based meta-analysis was performed to identify: (i) those regions which are replicably modulated by the uncertainty of environmental conditions; (ii) those regions whose activity is robustly affected by our subjective confidence; and (iii) those regions differentially activated at these contrasting times. In further meta-analyses the consistency of activation between these judgement types was assessed. Increased activation was consistently observed in the salience (anterior cingulate cortex and insula) and central executive network (dorsolateral prefrontal and posterior parietal cortices) in conditions of increased uncertainty; by contrast, default mode network (midline cortical and medial temporal lobe) regions robustly exhibited a positive relationship with subjective confidence. Regions including right parahippocampal gyrus were positively modulated by magnitude across both certainty and confidence judgements. This region was also shown to be more significantly modulated by confidence magnitude as compared with degree of environmental certainty. The functional and methodological implications of these findings are discussed with a view to improving future investigation of the neural basis of metacognitive judgement.
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Long-term patient outcomes after surgical stabilization of rib fractures.
Am. J. Surg.
PUBLISHED: 01-04-2014
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Rib fractures are common, and can be disabling. Recently, there has been increased interest in surgical stabilization of rib fractures (SSRF). It is difficult to define long-term benefits of the procedure. This is a descriptive study of patient outcomes after SSRF.
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Invading and expanding: range dynamics and ecological consequences of the greater white-toothed shrew (Crocidura russula) invasion in Ireland.
PLoS ONE
PUBLISHED: 01-01-2014
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Establishing how invasive species impact upon pre-existing species is a fundamental question in ecology and conservation biology. The greater white-toothed shrew (Crocidura russula) is an invasive species in Ireland that was first recorded in 2007 and which, according to initial data, may be limiting the abundance/distribution of the pygmy shrew (Sorex minutus), previously Ireland's only shrew species. Because of these concerns, we undertook an intensive live-trapping survey (and used other data from live-trapping, sightings and bird of prey pellets/nest inspections collected between 2006 and 2013) to model the distribution and expansion of C. russula in Ireland and its impacts on Ireland's small mammal community. The main distribution range of C. russula was found to be approximately 7,600 km2 in 2013, with established outlier populations suggesting that the species is dispersing with human assistance within the island. The species is expanding rapidly for a small mammal, with a radial expansion rate of 5.5 km/yr overall (2008-2013), and independent estimates from live-trapping in 2012-2013 showing rates of 2.4-14.1 km/yr, 0.5-7.1 km/yr and 0-5.6 km/yr depending on the landscape features present. S. minutus is negatively associated with C. russula. S. minutus is completely absent at sites where C. russula is established and is only present at sites at the edge of and beyond the invasion range of C. russula. The speed of this invasion and the homogenous nature of the Irish landscape may mean that S. minutus has not had sufficient time to adapt to the sudden appearance of C. russula. This may mean the continued decline/disappearance of S. minutus as C. russula spreads throughout the island.
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Dysconnectivity of neurocognitive networks at rest in very-preterm born adults.
Neuroimage Clin
PUBLISHED: 01-01-2014
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Advances in neonatal medicine have resulted in a larger proportion of preterm-born individuals reaching adulthood. Their increased liability to psychiatric illness and impairments of cognition and behaviour intimate lasting cerebral consequences; however, the central physiological disturbances remain unclear. Of fundamental importance to efficient brain function is the coordination and contextually-relevant recruitment of neural networks. Large-scale distributed networks emerge perinatally and increase in hierarchical complexity through development. Preterm-born individuals exhibit systematic reductions in correlation strength within these networks during infancy. Here, we investigate resting-state functional connectivity in functional magnetic resonance imaging data from 29 very-preterm (VPT)-born adults and 23 term-born controls. Neurocognitive networks were identified with spatial independent component analysis conducted using the Infomax algorithm and employing Icasso procedures to enhance component robustness. Network spatial focus and spectral power were not generally significantly affected by preterm birth. By contrast, Granger-causality analysis of the time courses of network activity revealed widespread reductions in between-network connectivity in the preterm group, particularly along paths including salience-network features. The potential clinical relevance of these Granger-causal measurements was suggested by linear discriminant analysis of topological representations of connection strength, which classified individuals by group with a maximal accuracy of 86%. Functional connections from the striatal salience network to the posterior default mode network informed this classification most powerfully. In the VPT-born group it was additionally found that perinatal factors significantly moderated the relationship between executive function (which was reduced in the VPT-born as compared with the term-born group) and generalised partial directed coherence. Together these findings show that resting-state functional connectivity of preterm-born individuals remains compromised in adulthood; and present consistent evidence that the striatal salience network is preferentially affected. Therapeutic practices directed at strengthening within-network cohesion and fine-tuning between-network inter-relations may have the potential to mitigate the cognitive, behavioural and psychiatric repercussions of preterm birth.
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Serial femtosecond crystallography of G protein-coupled receptors.
Science
PUBLISHED: 12-21-2013
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X-ray crystallography of G protein-coupled receptors and other membrane proteins is hampered by difficulties associated with growing sufficiently large crystals that withstand radiation damage and yield high-resolution data at synchrotron sources. We used an x-ray free-electron laser (XFEL) with individual 50-femtosecond-duration x-ray pulses to minimize radiation damage and obtained a high-resolution room-temperature structure of a human serotonin receptor using sub-10-micrometer microcrystals grown in a membrane mimetic matrix known as lipidic cubic phase. Compared with the structure solved by using traditional microcrystallography from cryo-cooled crystals of about two orders of magnitude larger volume, the room-temperature XFEL structure displays a distinct distribution of thermal motions and conformations of residues that likely more accurately represent the receptor structure and dynamics in a cellular environment.
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Vulnerability to Smokeless Tobacco Use Among Those Dependent on Alcohol or Illicit Drugs.
Nicotine Tob. Res.
PUBLISHED: 09-30-2013
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Individuals dependent on alcohol or illicit drugs are vulnerable to cigarette smoking and related adverse health outcomes. Less research has been conducted regarding whether these same groups are vulnerable to smokeless tobacco (ST) use. The goal of this study is to examine vulnerability to ST use among individuals with other drug dependence.
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The effects of age on lens transport.
Invest. Ophthalmol. Vis. Sci.
PUBLISHED: 09-26-2013
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Age-related nuclear cataracts involve denaturation and aggregation of intracellular proteins. We have documented age-dependent changes in membrane transport in the mouse lens to see what might initiate changes in the intracellular milieu.
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Functional Effects of Cx50 Mutations Associated with Congenital Cataracts.
Am. J. Physiol., Cell Physiol.
PUBLISHED: 09-04-2013
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Mutations in Cx50 cause dominant cataracts in both humans and mice. The exact mechanisms by which mutations cause these variable phenotypes are poorly understood. We examined the functional properties of gap junctions made by three Cx50 mutations, V44E, D47N and V79L, expressed in mammalian cell lines. V44E trafficked to the plasma membrane properly and formed gap junctional plaques. However, the mutant did not form functional gap junctions when expressed alone, or with wildtype (WT) Cx46 and Cx50, indicating that V44E is a dominant negative inhibitor of WT connexin function. In contrast, D47N subunits did not localize to junctional plaques or form functional homotypic gap junctions; however, mixed expression of D47N and WT subunits of either Cx50 or Cx46 resulted in functional intercellular channels, with high levels of coupling. Single channel studies indicated that D47N formed heteromeric channels with WT Cx46 with unique properties. Unlike either V44E or D47N, V79L formed functional homotypic intercellular channels. However, the mutation caused an alteration in voltage gating and a reduction in the open probability, resulting in much lower levels of conductance in cells expressing V79L alone, or together with WT connexin subunits. Thus, each mutation produced distinct changes in the properties of junctional coupling. V44E failed to form intercellular channels in any configuration, D47N formed only heteromeric channels with WT connexins and V79L formed homotypic and heteromeric channels with altered properties. These results suggest that unique interactions between mutant and wild-type lens connexins might underlie the development of various cataract phenotypes in humans.
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Preclinical Studies on Neurobehavioral and Neuromuscular Effects of Cocaine Hydrolase Gene Therapy in Mice.
J. Mol. Neurosci.
PUBLISHED: 08-30-2013
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Cocaine hydrolase gene transfer of mutated human butyrylcholinesterase (BChE) is evolving as a promising therapy for cocaine addiction. BChE levels after gene transfer can be 1,500-fold above those in untreated mice, making this enzyme the second most abundant plasma protein. Because mutated BChE is approximately 70 % as efficient in hydrolyzing acetylcholine as wild-type enzyme, it is important to examine the impact on cholinergic function. Here, we focused on memory and cognition (Stone T-maze), basic neuromuscular function (treadmill endurance and grip strength), and coordination (Rotarod). BALB/c mice were given adeno-associated virus vector or helper-dependent adenoviral vector encoding mouse or human BChE optimized for cocaine. Age-matched controls received saline or luciferase vector. Despite high doses (up to 10(13) particles per mouse) and high transgene expression (1,000-fold above baseline), no deleterious effects of vector treatment were seen in neurobehavioral functions. The vector-treated mice performed as saline-treated and luciferase controls in maze studies and strength tests, and their Rotarod and treadmill performance decreased less with age. Thus, neither the viral vectors nor the large excess of BChE caused observable toxic effects on the motor and cognitive systems investigated. This outcome justifies further steps toward an eventual clinical trial of vector-based gene transfer for cocaine abuse.
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The critical importance of defined media conditions in Daphnia magna nanotoxicity studies.
Toxicol. Lett.
PUBLISHED: 08-07-2013
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Due to the widespread use of silver nanoparticles (AgNPs), the likelihood of them entering the environment has increased and they are known to be potentially toxic. Currently, there is little information on the dynamic changes of AgNPs in ecotoxicity exposure media and how this may affect toxicity. Here, the colloidal stability of three different sizes of citrate-stabilized AgNPs was assessed in standard strength OECD ISO exposure media, and in 2-fold (media2) and 10-fold (media10) dilutions by transmission electron microscopy (TEM) and atomic force microscopy (AFM) and these characteristics were related to their toxicity towards Daphnia magna. Aggregation in undiluted media (media1) was rapid, and after diluting the medium by a factor of 2 or 10, aggregation was reduced, with minimal aggregation over 24h occurring in media10. Acute toxicity measurements were performed using 7nm diameter particles in media1 and media10. In media10 the EC50 of the 7nm particles for D. magna neonates was calculated to be 7.46?gL(-1) with upper and lower 95% confidence intervals of 6.84?gL(-1) and 8.13?gL(-1) respectively. For media1, an EC50 could not be calculated, the lowest observed adverse effect concentration (LOAEC) of 11.25?gL(-1) indicating a significant reduction in toxicity compared to that in media10. The data suggest the increased dispersion of nanoparticles leads to enhanced toxicity, emphasising the importance of appropriate media composition to fully assess nanoparticle toxicity in aquatic ecotoxicity tests.
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Do you see what I see? Sex differences in the discrimination of facial emotions during adolescence.
Emotion
PUBLISHED: 08-05-2013
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During adolescence social relationships become increasingly important. Establishing and maintaining these relationships requires understanding of emotional stimuli, such as facial emotions. A failure to adequately interpret emotional facial expressions has previously been associated with various mental disorders that emerge during adolescence. The current study examined sex differences in emotional face processing during adolescence. Participants were adolescents (n = 1951) with a target age of 14, who completed a forced-choice emotion discrimination task. The stimuli used comprised morphed faces that contained a blend of two emotions in varying intensities (11 stimuli per set of emotions). Adolescent girls showed faster and more sensitive perception of facial emotions than boys. However, both adolescent boys and girls were most sensitive to variations in emotion intensity in faces combining happiness and sadness, and least sensitive to changes in faces comprising fear and anger. Furthermore, both sexes overidentified happiness and anger. However, the overidentification of happiness was stronger in boys. These findings were not influenced by individual differences in the level of pubertal maturation. These results indicate that male and female adolescents differ in their ability to identify emotions in morphed faces containing emotional blends. The findings provide information for clinical studies examining whether sex differences in emotional processing are related to sex differences in the prevalence of psychiatric disorders within this age group. (PsycINFO Database Record (c) 2013 APA, all rights reserved).
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Structure of a photosynthetic reaction centre determined by serial femtosecond crystallography.
Nat Commun
PUBLISHED: 07-19-2013
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Serial femtosecond crystallography is an X-ray free-electron-laser-based method with considerable potential to have an impact on challenging problems in structural biology. Here we present X-ray diffraction data recorded from microcrystals of the Blastochloris viridis photosynthetic reaction centre to 2.8?Å resolution and determine its serial femtosecond crystallography structure to 3.5?Å resolution. Although every microcrystal is exposed to a dose of 33?MGy, no signs of X-ray-induced radiation damage are visible in this integral membrane protein structure.
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Neural primacy of the salience processing system in schizophrenia.
Neuron
PUBLISHED: 06-20-2013
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For effective information processing, two large-scale distributed neural networks appear to be critical: a multimodal executive system anchored on the dorsolateral prefrontal cortex (DLPFC) and a salience system anchored on the anterior insula. Aberrant interaction among distributed networks is a feature of psychiatric disorders such as schizophrenia. We used whole-brain Granger causal modeling using resting fMRI and observed a significant failure of both the feedforward and reciprocal influence between the insula and the DLPFC in schizophrenia. Further, a significant failure of directed influence from bilateral visual cortices to the insula was also seen in patients. These findings provide compelling evidence for a breakdown of the salience-execution loop in the clinical expression of psychosis. In addition, this offers a parsimonious explanation for the often-observed "frontal inefficiency," the failure to recruit prefrontal system when salient or novel information becomes available in patients with schizophrenia.
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Sensing the wavefront of x-ray free-electron lasers using aerosol spheres.
Opt Express
PUBLISHED: 06-06-2013
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Characterizing intense, focused x-ray free electron laser (FEL) pulses is crucial for their use in diffractive imaging. We describe how the distribution of average phase tilts and intensities on hard x-ray pulses with peak intensities of 10(21) W/m(2) can be retrieved from an ensemble of diffraction patterns produced by 70 nm-radius polystyrene spheres, in a manner that mimics wavefront sensors. Besides showing that an adaptive geometric correction may be necessary for diffraction data from randomly injected sample sources, our paper demonstrates the possibility of collecting statistics on structured pulses using only the diffraction patterns they generate and highlights the imperative to study its impact on single-particle diffractive imaging.
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Application of Partial Directed Coherence to the Analysis of Resting-State EEG-fMRI Data.
Brain Connect
PUBLISHED: 06-04-2013
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Abstract The simultaneous acquisition of electroencephalography (EEG) and functional magnetic resonance imaging (fMRI) data potentially allows measurement of brain signals with both high spatial and temporal resolution. Partial directed coherence (PDC) is a Granger causality measure in the frequency domain, which is often used to infer the intensity of information flow over the brain from EEG data. In the current study, we propose a new approach to investigate functional connectivity in resting-state (RS) EEG-fMRI data by combining time-varying PDC with the analysis of blood oxygenation level-dependent (BOLD) signal fluctuations. Basically, we aim to identify brain circuits that are more active when the information flow is increased between distinct remote neuronal modules. The usefulness of the proposed method is illustrated by application to simultaneously recorded EEG-fMRI data from healthy subjects at rest. Using this approach, we decomposed the nodes of RS networks in fMRI data according to the frequency band and directed flow of information provided from EEG. This approach therefore has the potential to inform our understanding of the regional characteristics of oscillatory processes in the human brain.
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Crystallographic data processing for free-electron laser sources.
Acta Crystallogr. D Biol. Crystallogr.
PUBLISHED: 05-17-2013
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A processing pipeline for diffraction data acquired using the `serial crystallography methodology with a free-electron laser source is described with reference to the crystallographic analysis suite CrystFEL and the pre-processing program Cheetah. A detailed analysis of the nature and impact of indexing ambiguities is presented. Simulations of the Monte Carlo integration scheme, which accounts for the partially recorded nature of the diffraction intensities, are presented and show that the integration of partial reflections could be made to converge more quickly if the bandwidth of the X-rays were to be increased by a small amount or if a slight convergence angle were introduced into the incident beam.
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Alcohol injection for Mortons neuroma: a five-year follow-up.
Foot Ankle Int
PUBLISHED: 05-13-2013
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Although many treatment modalities are available for Mortons neuroma (MN), studies looking at the long-term effectiveness of most forms of treatment are scarce. The injection of MN with alcohol has gained popularity over the past 10 years with widespread media coverage. Many surgeons have anecdotally questioned the long-term effectiveness of this treatment. We reviewed a cohort of patients at an average 5-year follow-up to assess the medium-term results of alcohol injection.
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AKT activation promotes PTEN hamartoma tumor syndrome-associated cataract development.
J. Clin. Invest.
PUBLISHED: 04-11-2013
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Mutations in the human phosphatase and tensin homolog (PTEN) gene cause PTEN hamartoma tumor syndrome (PHTS), which includes cataract development among its diverse clinical pathologies. Currently, it is not known whether cataract formation in PHTS patients is secondary to other systemic problems, or the result of the loss of a critical function of PTEN within the lens. We generated a mouse line with a lens-specific deletion of Pten (PTEN KO) and identified a regulatory function for PTEN in lens ion transport. Specific loss of PTEN in the lens resulted in cataract. PTEN KO lenses exhibited a progressive age-related increase in intracellular hydrostatic pressure, along with, increased intracellular sodium concentrations, and reduced Na+/K+-ATPase activity. Collectively, these defects lead to lens swelling, opacities and ultimately organ rupture. Activation of AKT was highly elevated in PTEN KO lenses compared to WT mice. Additionally, pharmacological inhibition of AKT restored normal Na+/K+-ATPase activity in primary cultured lens cells and reduced lens pressure in intact lenses from PTEN KO animals. These findings identify a direct role for PTEN in the regulation of lens ion transport through an AKT-dependent modulation of Na+/K+-ATPase activity, and provide a new animal model to investigate cataract development in PHTS patients.
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Adaptive evolution during an ongoing range expansion: the invasive bank vole (Myodes glareolus) in Ireland.
Mol. Ecol.
PUBLISHED: 04-03-2013
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Range expansions are extremely common, but have only recently begun to attract attention in terms of their genetic consequences. As populations expand, demes at the wave front experience strong genetic drift, which is expected to reduce genetic diversity and potentially cause allele surfing, where alleles may become fixed over a wide geographical area even if their effects are deleterious. Previous simulation models show that range expansions can generate very strong selective gradients on dispersal, reproduction, competition and immunity. To investigate the effects of range expansion on genetic diversity and adaptation, we studied the population genomics of the bank vole (Myodes glareolus) in Ireland. The bank vole was likely introduced in the late 1920s and is expanding its range at a rate of ~2.5 km/year. Using genotyping-by-sequencing, we genotyped 281 bank voles at 5979 SNP loci. Fourteen sample sites were arranged in three transects running from the introduction site to the wave front of the expansion. We found significant declines in genetic diversity along all three transects. However, there was no evidence that sites at the wave front had accumulated more deleterious mutations. We looked for outlier loci with strong correlations between allele frequency and distance from the introduction site, where the direction of correlation was the same in all three transects. Amongst these outliers, we found significant enrichment for genic SNPs, suggesting the action of selection. Candidates for selection included several genes with immunological functions and several genes that could influence behaviour.
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The effect of size and species on lens intracellular hydrostatic pressure.
Invest. Ophthalmol. Vis. Sci.
PUBLISHED: 03-06-2013
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Previous experiments showed that mouse lenses have an intracellular hydrostatic pressure that varied from 335 mm Hg in central fibers to 0 mm Hg in surface cells. Model calculations predicted that in larger lenses, all else equal, pressure should increase as the lens radius squared. To test this prediction, lenses of different radii from different species were studied.
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Understanding the basis of diminished gene flow between hybridizing chromosome races of the house mouse.
Evolution
PUBLISHED: 03-01-2013
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Speciation may be promoted in hybrid zones if there is an interruption to gene flow between the hybridizing forms. For hybridizing chromosome races of the house mouse in Valtellina (Italy), distinguished by whole-arm chromosomal rearrangements, previous studies have shown that there is greater interruption to gene flow at the centromeres of chromosomes that differ between the races than at distal regions of the same chromosome or at the centromeres of other chromosomes. Here, by increasing the number of markers along race-specific chromosomes, we reveal a decay in between-race genetic differentiation from the centromere to the distal telomere. For the first time, we use simulation models to investigate the possible role of recombination suppression and hybrid breakdown in generating this pattern. We also consider epistasis and selective sweeps as explanations for isolated chromosomal regions away from the centromere showing differentiation between the races. Hybrid breakdown alone is the simplest explanation for the decay in genetic differentiation with distance from the centromere. Robertsonian fusions/whole-arm reciprocal translocations are common chromosomal rearrangements characterizing both closely related species and races within species, and this fine-scale empirical analysis suggests that the unfitness associated with these rearrangements in the heterozygous state may contribute to the speciation process.
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The human Cx26-D50A and Cx26-A88V mutations causing keratitis-ichthyosis-deafness syndrome display increased hemichannel activity.
Am. J. Physiol., Cell Physiol.
PUBLISHED: 02-27-2013
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Mutations in the human gene encoding connexin 26 (Cx26 or GJB2) cause either nonsyndromic deafness or syndromic deafness associated with skin diseases. That distinct clinical disorders can be caused by different mutations within the same gene suggests that different channel activities influence the ear and skin. Here we use three different expression systems to examine the functional characteristics of two Cx26 mutations causing either mild (Cx26-D50A) or lethal (Cx26-A88V) keratitis-ichthyosis-deafness (KID) syndrome. In either cRNA-injected Xenopus oocytes, transfected HeLa cells, or transfected primary human keratinocytes, we show that both Cx26-D50A and Cx26-A88V form active hemichannels that significantly increase membrane current flow compared with wild-type Cx26. This increased membrane current accelerated cell death in low extracellular calcium solutions and was not due to increased mutant protein expression. Elevated mutant hemichannel currents could be blocked by increased extracellular calcium concentration. These results show that these two mutations exhibit a shared gain of functional activity and support the hypothesis that increased hemichannel activity is a common feature of human Cx26 mutations responsible for KID syndrome.
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Dysregulated but not decreased salience network activity in schizophrenia.
Front Hum Neurosci
PUBLISHED: 02-18-2013
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Effective estimation of the salience of environmental stimuli underlies adaptive behavior, while related aberrance is believed to undermine rational thought processes in schizophrenia. A network including bilateral frontoinsular cortex (FIC) and dorsal anterior cingulate cortex (dACC) has been observed to respond to salient stimuli using functional magnetic resonance imaging (fMRI). To test the hypothesis that activity in this salience network (SN) is less discriminately modulated by contextually-relevant stimuli in schizophrenia than in healthy individuals, fMRI data were collected in 20 individuals with schizophrenia and 13 matched controls during performance of a modified monetary incentive delay (MID) task. After quantitatively identifying spatial components representative of the FIC and dACC features of the SN, two principal analyses were conducted. In the first, modulation of SN activity by salience was assessed by measuring response to trial outcome. First-level general linear models were applied to individual-specific time-courses of SN activity identified using spatial independent component analysis (ICA). This analysis revealed a significant salience-by-performance-by-group interaction on the best-fit FIC components activity at trial outcome, whereby healthy individuals but not individuals with schizophrenia exhibited greater distinction between the response to hits and misses in high salience trials than in low salience trials. The second analysis aimed to ascertain whether SN component amplitude differed between the study groups over the duration of the experiment. Independent-samples T-tests on back-projected, percent-signal-change scaled SN component images importantly showed that the groups did not differ in the overall amplitude of SN expression over the entire dataset. These findings of dysregulated but not decreased SN activity in schizophrenia provide physiological support for mechanistic conceptual frameworks of delusional thought formation.
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Phenotypic variation across chromosomal hybrid zones of the common shrew (Sorex araneus) indicates reduced gene flow.
PLoS ONE
PUBLISHED: 01-01-2013
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Sorex araneus, the Common shrew, is a species with more than 70 karyotypic races, many of which form parapatric hybrid zones, making it a model for studying chromosomal speciation. Hybrids between races have reduced fitness, but microsatellite markers have demonstrated considerable gene flow between them, calling into question whether the chromosomal barriers actually do contribute to genetic divergence. We studied phenotypic clines across two hybrid zones with especially complex heterozygotes. Hybrids between the Novosibirsk and Tomsk races produce chains of nine and three chromosomes at meiosis, and hybrids between the Moscow and Seliger races produce chains of eleven. Our goal was to determine whether phenotypes show evidence of reduced gene flow at hybrid zones. We used maximum likelihood to fit tanh cline models to geometric shape data and found that phenotypic clines in skulls and mandibles across these zones had similar centers and widths as chromosomal clines. The amount of phenotypic differentiation across the zones is greater than expected if it were dissipating due to unrestricted gene flow given the amount of time since contact, but it is less than expected to have accumulated from drift during allopatric separation in glacial refugia. Only if heritability is very low, Ne very high, and the time spent in allopatry very short, will the differences we observe be large enough to match the expectation of drift. Our results therefore suggest that phenotypic differentiation has been lost through gene flow since post-glacial secondary contact, but not as quickly as would be expected if there was free gene flow across the hybrid zones. The chromosomal tension zones are confirmed to be partial barriers that prevent differentiated races from becoming phenotypically homogenous.
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Radiation damage in protein serial femtosecond crystallography using an x-ray free-electron laser.
Phys. Rev. B Condens. Matter Mater. Phys.
PUBLISHED: 12-01-2011
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X-ray free-electron lasers deliver intense femtosecond pulses that promise to yield high resolution diffraction data of nanocrystals before the destruction of the sample by radiation damage. Diffraction intensities of lysozyme nanocrystals collected at the Linac Coherent Light Source using 2 keV photons were used for structure determination by molecular replacement and analyzed for radiation damage as a function of pulse length and fluence. Signatures of radiation damage are observed for pulses as short as 70 fs. Parametric scaling used in conventional crystallography does not account for the observed effects.
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Varying molecular interactions by coverage in supramolecular surface chemistry.
Chem. Commun. (Camb.)
PUBLISHED: 11-09-2011
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The possibility of modifying the intermolecular interactions of absorbed benzene-carboxylic acids from coordination to hydrogen bonding by changing their surface coverage is demonstrated through a combination of scanning tunnelling microscopy, X-ray photoemission spectroscopy and density functional theory calculations.
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The Cx26-G45E mutation displays increased hemichannel activity in a mouse model of the lethal form of keratitis-ichthyosis-deafness syndrome.
Mol. Biol. Cell
PUBLISHED: 10-26-2011
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Mutations in the GJB2 gene (Cx26) cause deafness in humans. Most are loss-of-function mutations and cause nonsyndromic deafness. Some mutations produce a gain of function and cause syndromic deafness associated with skin disorders, such as keratitis-ichthyosis-deafness syndrome (KIDS). Cx26-G45E is a lethal mutation linked to KIDS that forms constitutively active connexin hemichannels. The pathomechanism(s) by which mutant Cx26 hemichannels perturb normal epidermal cornification are poorly understood. We created an animal model for KIDS by generating an inducible transgenic mouse expressing Cx26-G45E in keratinocytes. Cx26-G45E mice displayed reduced viability, hyperkeratosis, scaling, skin folds, and hair loss. Histopathology included hyperplasia, acanthosis, papillomatosis, increased cell size, and osteal plugging. These abnormalities correlated with human KIDS pathology and were associated with increased hemichannel currents in transgenic keratinocytes. These results confirm the pathogenic nature of the G45E mutation and provide a new model for studying the role of aberrant connexin hemichannels in epidermal differentiation and inherited connexin disorders.
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Long-term impact of web-based tools, leadership feedback, and policies on inpatient antipsychotic polypharmacy.
Psychiatr Serv
PUBLISHED: 10-05-2011
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This column describes a series of interventions to decrease antipsychotic polypharmacy in the New York State Office of Mental Health (NYSOMH) network of psychiatric hospitals. Phase 1 consisted of implementation of the Psychiatric Services Clinical Knowledge Enhancement System (PSYCKES), a Web-based application supporting clinical decision making and quality improvement, and a policy requiring approval by NYSOMHs medical director to prescribe more than two antipsychotics per patient. In phase 2 hospital leaders received feedback from the office of the medical director identifying specific patients on polypharmacy. In phase 3, access to PSYCKES continued, but the prior-approval policy and feedback were discontinued. Polypharmacy decreased significantly during phase 1, from 16.9 to 9.7 inpatients per 1,000, and decreased further in phase 2, to 3.9 inpatients per 1,000. In phase 3 the prevalence of antipsychotic polypharmacy remained low at six-month follow-up (3.1 inpatients per 1,000), despite the ending of state-level oversight. On long-term follow-up, polypharmacy increased, eventually rising to 9.2 inpatients per 1,000 after 36 months, but remained well below baseline levels.
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Exome sequencing identifies a spectrum of mutation frequencies in advanced and lethal prostate cancers.
Proc. Natl. Acad. Sci. U.S.A.
PUBLISHED: 09-26-2011
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To catalog protein-altering mutations that may drive the development of prostate cancers and their progression to metastatic disease systematically, we performed whole-exome sequencing of 23 prostate cancers derived from 16 different lethal metastatic tumors and three high-grade primary carcinomas. All tumors were propagated in mice as xenografts, designated the LuCaP series, to model phenotypic variation, such as responses to cancer-directed therapeutics. Although corresponding normal tissue was not available for most tumors, we were able to take advantage of increasingly deep catalogs of human genetic variation to remove most germline variants. On average, each tumor genome contained ~200 novel nonsynonymous variants, of which the vast majority was specific to individual carcinomas. A subset of genes was recurrently altered across tumors derived from different individuals, including TP53, DLK2, GPC6, and SDF4. Unexpectedly, three prostate cancer genomes exhibited substantially higher mutation frequencies, with 2,000-4,000 novel coding variants per exome. A comparison of castration-resistant and castration-sensitive pairs of tumor lines derived from the same prostate cancer highlights mutations in the Wnt pathway as potentially contributing to the development of castration resistance. Collectively, our results indicate that point mutations arising in coding regions of advanced prostate cancers are common but, with notable exceptions, very few genes are mutated in a substantial fraction of tumors. We also report a previously undescribed subtype of prostate cancers exhibiting "hypermutated" genomes, with potential implications for resistance to cancer therapeutics. Our results also suggest that increasingly deep catalogs of human germline variation may challenge the necessity of sequencing matched tumor-normal pairs.
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Unsupervised classification of single-particle X-ray diffraction snapshots by spectral clustering.
Opt Express
PUBLISHED: 09-22-2011
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Single-particle experiments using X-ray Free Electron Lasers produce more than 10(5) snapshots per hour, consisting of an admixture of blank shots (no particle intercepted), and exposures of one or more particles. Experimental data sets also often contain unintentional contamination with different species. We present an unsupervised method able to sort experimental snapshots without recourse to templates, specific noise models, or user-directed learning. The results show 90% agreement with manual classification.
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Genetic testing for early detection of individuals at risk of coronary heart disease and monitoring response to therapy: challenges and promises.
Curr Atheroscler Rep
PUBLISHED: 08-11-2011
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Coronary heart disease (CHD) often presents suddenly with little warning. Traditional risk factors are inadequate to identify the asymptomatic high-risk individuals. Early identification of patients with subclinical coronary artery disease using noninvasive imaging modalities would allow the early adoption of aggressive preventative interventions. Currently, it is impractical to screen the entire population with noninvasive coronary imaging tools. The use of relatively simple and inexpensive genetic markers of increased CHD risk can identify a population subgroup in which benefit of atherosclerotic imaging modalities would be increased despite nominal cost and radiation exposure. Additionally, genetic markers are fixed and need only be measured once in a patients lifetime, can help guide therapy selection, and may be of utility in family counseling.
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Lens intracellular hydrostatic pressure is generated by the circulation of sodium and modulated by gap junction coupling.
J. Gen. Physiol.
PUBLISHED: 06-01-2011
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We recently modeled fluid flow through gap junction channels coupling the pigmented and nonpigmented layers of the ciliary body. The model suggested the channels could transport the secretion of aqueous humor, but flow would be driven by hydrostatic pressure rather than osmosis. The pressure required to drive fluid through a single layer of gap junctions might be just a few mmHg and difficult to measure. In the lens, however, there is a circulation of Na(+) that may be coupled to intracellular fluid flow. Based on this hypothesis, the fluid would cross hundreds of layers of gap junctions, and this might require a large hydrostatic gradient. Therefore, we measured hydrostatic pressure as a function of distance from the center of the lens using an intracellular microelectrode-based pressure-sensing system. In wild-type mouse lenses, intracellular pressure varied from ?330 mmHg at the center to zero at the surface. We have several knockout/knock-in mouse models with differing levels of expression of gap junction channels coupling lens fiber cells. Intracellular hydrostatic pressure in lenses from these mouse models varied inversely with the number of channels. When the lens circulation of Na(+) was either blocked or reduced, intracellular hydrostatic pressure in central fiber cells was either eliminated or reduced proportionally. These data are consistent with our hypotheses: fluid circulates through the lens; the intracellular leg of fluid circulation is through gap junction channels and is driven by hydrostatic pressure; and the fluid flow is generated by membrane transport of sodium.
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Pathological hemichannels associated with human Cx26 mutations causing Keratitis-Ichthyosis-Deafness syndrome.
Biochim. Biophys. Acta
PUBLISHED: 05-23-2011
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Connexin (Cx) proteins form intercellular gap junction channels by first assembling into single membrane hemichannels that then dock to connect the cytoplasm of two adjacent cells. Gap junctions are highly specialized structures that allow the direct passage of small molecules between cells to maintain tissue homeostasis. Functional activity of nonjunctional hemichannels has now been shown in several experimental systems. Hemichannels may constitute an important diffusional exchange pathway with the extracellular space, but the extent of their normal physiological role is currently unknown. Aberrant hemichannel activity has been linked to mutations of connexin proteins involved in genetic diseases. Here, we review a proposed role for hemichannels in the pathogenesis of Keratitis-Ichthyosis-Deafness (KID) syndrome associated with connexin26 (Cx26) mutations. Continued functional evaluation of mutated hemichannels linked to human hereditary disorders may provide additional insights into the mechanisms governing their regulation in normal physiology and dysregulation in disease. This article is part of a Special Issue entitled: The Communicating junctions, composition, structure and characteristics.
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Experimental observation of evanescent modes at the interface to slow-light photonic crystal waveguides.
Opt Lett
PUBLISHED: 04-12-2011
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We experimentally study the fields close to an interface between two photonic crystal waveguides that have different dispersion properties. After the transition from a waveguide in which the group velocity of light is v(g) ~ c/10 to a waveguide in which it is v(g) ~ c/100, we observe a gradual increase in the field intensity and the lateral spreading of the mode. We attribute this evolution to the existence of a weakly evanescent mode that exponentially decays away from the interface. We compare this to the situation where the transition between the waveguides only leads to a minor change in group velocity and show that, in that case, the evolution is absent. Furthermore, we apply novel numerical mode extraction techniques to confirm experimental results.
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Aggregation and dispersion of silver nanoparticles in exposure media for aquatic toxicity tests.
J Chromatogr A
PUBLISHED: 03-14-2011
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Silver nanoparticles (AgNPs) are currently being very widely used in industry, mainly because of their anti-bacterial properties, with applications in many areas. Once released into the environment, the mobility, bioavailability, and toxicity of AgNPs in any ecosystem are dominated by colloidal stability. There have been studies on the stability or the aggregation of various nanoparticles (NPs) under a range of environmental conditions, but there is little information on fully characterised AgNPs in media used in (eco)toxicity studies. In this study, monodisperse 7, 10 and 20 nm citrate-stabilised AgNPs were synthesised, characterised and then fractionated and sized by flow field-flow fractionation (FFF) and measured with dynamic light scattering (DLS) in different dilutions of the media recommended by OECD for Daphnia magna (water flea) toxicity testing. Stability of NPs was assessed over 24 h, and less so over 21 days, similar time periods to the OECD acute and chronic toxicity tests for D. magna. All particles aggregated quickly in the media with high ionic strength (media1), resulting in a loss of colour from the solution. The size of particles could be measured by DLS in most cases after 24h, although a fractogram by FFF could not be obtained due to aggregation and polydispersity of the sample. After diluting the media by a factor of 2, 5 or 10, aggregation was reduced, although the smallest NPs were unstable under all media conditions. Media diluted up to 10-fold in the absence of AgNPs did not induce any loss of mobility or fecundity in D. magna. These results confirm that standard OECD media causes aggregation of AgNPs, which result in changes in organism exposure levels and the nature of the exposed particles compared to exposure to fully dispersed particles. Setting aside questions of dose metrics, significant and substantial reduction in concentration over exposure period suggests that literature data are in the main improperly interpreted and nanoparticles are likely to have far greater biological effects than suggested thus far by poorly controlled exposures. We recommend that the standard OECD media is diluted by a factor of ca. 10 for use with these NPs and this test media, which reduces AgNP aggregation without affecting the viability of the text organism.
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Phasing of coherent femtosecond X-ray diffraction from size-varying nanocrystals.
Opt Express
PUBLISHED: 03-04-2011
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The scattering between Bragg reflections from nanocrystals is used to aid solution of the phase problem. We describe a method for reconstructing the charge density of a typical molecule within a single unit cell, if sufficiently finely-sampled "snap-shot" diffraction data (as provided a free-electron X-ray laser) are available from many nanocrystals of different sizes lying in random orientations. By using information on the particle-size distribution within the patterns, this digital method succeeds, using all the data, without knowledge of the distribution of particle size or requiring atomic-resolution data.
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Motion-related artefacts in EEG predict neuronally plausible patterns of activation in fMRI data.
Neuroimage
PUBLISHED: 02-23-2011
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The simultaneous acquisition and subsequent analysis of EEG and fMRI data is challenging owing to increased noise levels in the EEG data. A common method to integrate data from these two modalities is to use aspects of the EEG data, such as the amplitudes of event-related potentials (ERP) or oscillatory EEG activity, to predict fluctuations in the fMRI data. However, this relies on the acquisition of high quality datasets to ensure that only the correlates of neuronal activity are being studied. In this study, we investigate the effects of head-motion-related artefacts in the EEG signal on the predicted T2*-weighted signal variation. We apply our analyses to two independent datasets: 1) four participants were asked to move their feet in the scanner to generate small head movements, and 2) four participants performed an episodic memory task. We created T2*-weighted signal predictors from indicators of abrupt head motion using derivatives of the realignment parameters, from visually detected artefacts in the EEG as well as from three EEG frequency bands (theta, alpha and beta). In both datasets, we found little correlation between the T2*-weighted signal and EEG predictors that were not convolved with the canonical haemodynamic response function (cHRF). However, all convolved EEG predictors strongly correlated with the T2*-weighted signal variation in various regions including the bilateral superior temporal cortex, supplementary motor area, medial parietal cortex and cerebellum. The finding that movement onset spikes in the EEG predict T2*-weighted signal intensity only when the time course of movements is convolved with the cHRF, suggests that the correlated signal might reflect a BOLD response to neural activity associated with head movement. Furthermore, the observation that broad-spectral EEG spikes tend to occur at the same time as abrupt head movements, together with the finding that abrupt movements and EEG spikes show similar correlations with the T2*-weighted signal, indicates that the EEG spikes are produced by abrupt movement and that continuous regressors of EEG oscillations contain motion-related noise even after stringent correction of the EEG data. If not properly removed, these artefacts complicate the use of EEG data as a predictor of T2*-weighted signal variation.
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Structure-factor analysis of femtosecond microdiffraction patterns from protein nanocrystals.
Acta Crystallogr., A, Found. Crystallogr.
PUBLISHED: 02-16-2011
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A complete set of structure factors has been extracted from hundreds of thousands of femtosecond single-shot X-ray microdiffraction patterns taken from randomly oriented nanocrystals. The method of Monte Carlo integration over crystallite size and orientation was applied to experimental data from Photosystem I nanocrystals. This arrives at structure factors from many partial reflections without prior knowledge of the particle-size distribution. The data were collected at the Linac Coherent Light Source (the first hard-X-ray laser user facility), to which was fitted a hydrated protein nanocrystal injector jet, according to the method of serial crystallography. The data are single still diffraction snapshots, each from a different nanocrystal with sizes ranging between 100 nm and 2 µm, so the angular width of Bragg peaks was dominated by crystal-size effects. These results were compared with single-crystal data recorded from large crystals of Photosystem I at the Advanced Light Source and the quality of the data was found to be similar. The implications for improving the efficiency of data collection by allowing the use of very small crystals, for radiation-damage reduction and for time-resolved diffraction studies at room temperature are discussed.
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Single mimivirus particles intercepted and imaged with an X-ray laser.
Nature
PUBLISHED: 02-05-2011
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X-ray lasers offer new capabilities in understanding the structure of biological systems, complex materials and matter under extreme conditions. Very short and extremely bright, coherent X-ray pulses can be used to outrun key damage processes and obtain a single diffraction pattern from a large macromolecule, a virus or a cell before the sample explodes and turns into plasma. The continuous diffraction pattern of non-crystalline objects permits oversampling and direct phase retrieval. Here we show that high-quality diffraction data can be obtained with a single X-ray pulse from a non-crystalline biological sample, a single mimivirus particle, which was injected into the pulsed beam of a hard-X-ray free-electron laser, the Linac Coherent Light Source. Calculations indicate that the energy deposited into the virus by the pulse heated the particle to over 100,000?K after the pulse had left the sample. The reconstructed exit wavefront (image) yielded 32-nm full-period resolution in a single exposure and showed no measurable damage. The reconstruction indicates inhomogeneous arrangement of dense material inside the virion. We expect that significantly higher resolutions will be achieved in such experiments with shorter and brighter photon pulses focused to a smaller area. The resolution in such experiments can be further extended for samples available in multiple identical copies.
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Femtosecond X-ray protein nanocrystallography.
Nature
PUBLISHED: 02-05-2011
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X-ray crystallography provides the vast majority of macromolecular structures, but the success of the method relies on growing crystals of sufficient size. In conventional measurements, the necessary increase in X-ray dose to record data from crystals that are too small leads to extensive damage before a diffraction signal can be recorded. It is particularly challenging to obtain large, well-diffracting crystals of membrane proteins, for which fewer than 300 unique structures have been determined despite their importance in all living cells. Here we present a method for structure determination where single-crystal X-ray diffraction snapshots are collected from a fully hydrated stream of nanocrystals using femtosecond pulses from a hard-X-ray free-electron laser, the Linac Coherent Light Source. We prove this concept with nanocrystals of photosystem I, one of the largest membrane protein complexes. More than 3,000,000 diffraction patterns were collected in this study, and a three-dimensional data set was assembled from individual photosystem I nanocrystals (?200?nm to 2??m in size). We mitigate the problem of radiation damage in crystallography by using pulses briefer than the timescale of most damage processes. This offers a new approach to structure determination of macromolecules that do not yield crystals of sufficient size for studies using conventional radiation sources or are particularly sensitive to radiation damage.
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JoVE Visualize is a tool created to match the last 5 years of PubMed publications to methods in JoVE's video library.

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In developing our video relationships, we compare around 5 million PubMed articles to our library of over 4,500 methods videos. In some cases the language used in the PubMed abstracts makes matching that content to a JoVE video difficult. In other cases, there happens not to be any content in our video library that is relevant to the topic of a given abstract. In these cases, our algorithms are trying their best to display videos with relevant content, which can sometimes result in matched videos with only a slight relation.