High-frequency oscillatory ventilation versus conventional mechanical ventilation for very-low-birth-weight infants.
The efficacy and safety of early high-frequency oscillatory ventilation as compared with conventional synchronized intermittent mandatory ventilation for the treatment of infants with very low birth weight have not been established. We conducted a randomized, multicenter clinical trial to determine whether infants treated with early high-frequency oscillatory ventilation were more likely than infants treated with synchronized intermittent mandatory ventilation to be alive without requiring supplemental oxygen at 36 weeks of postmenstrual age. Eligible infants weighed 601 to 1200 g at birth, were less than four hours of age, had received one dose of surfactant, and required ventilation with a mean airway pressure of at least 6 cm of water and a fraction of inspired oxygen of at least 0.25. Infants were stratified according to birth weight and exposure to prenatal corticosteroids and then randomly assigned to high-frequency oscillatory ventilation or synchronized intermittent mandatory ventilation. Ventilation was managed according to protocols designed to optimize lung inflation and blood gas values. Five hundred infants were enrolled in the study. Infants randomly assigned to high-frequency oscillatory ventilation were successfully extubated earlier than infants assigned to synchronized intermittent mandatory ventilation (P<0.001). Of infants assigned to high-frequency oscillatory ventilation, 56 percent were alive without a need for supplemental oxygen at 36 weeks of postmenstrual age, as compared with 47 percent of those receiving synchronized intermittent mandatory ventilation (P=0.046). There was no difference between the groups in the risk of intracranial hemorrhage, cystic periventricular leukomalacia, or other complications. There was a small but significant benefit of high-frequency oscillatory ventilation in terms of the pulmonary outcome for very-low-birth-weight infants without an increase in the occurrence of other complications of premature birth.


Assembly of bio-nanoparticles for double controlled drug release.
A critical limiting factor of chemotherapy is the unacceptably high toxicity. The use of nanoparticle based drug carriers has significantly reduced the side effects and facilitated the delivery of drugs. Source of the remaining side effect includes (1) the broad final in vivo distribution of the administrated nanoparticles, and (2) strong basal drug release from nanoparticles before they could reach the tumor. Despite the advances in pH-triggered release, undesirable basal drug release has been a constant challenge under in vivo conditions. In this study, functionalized single walled carbon nanohorn supported immunoliposomes were assembled for paclitaxel delivery. The immunoliposomes were formulated with polyethylene glycol, thermal stable and pH sensitive phospholipids. Each nanohorn was found to be encapsulated within one immunoliposome. Results showed a highly pH dependent release of paclitaxel in the presence of serum at body temperature with minimal basal release under physiological conditions. Upon acidification, paclitaxel was released at a steady rate over 30 days with a cumulative release of 90% of the loaded drug. The drug release results proved our hypothesized double controlled release mechanism from the nanoparticles. Other results showed the nanoparticles have doubled loading capacity compared to that of traditional liposomes and higher affinity to breast cancer cells overexpressing Her2 receptors. Internalized nanoparticles were found in lysosomes.


The high burden of infant deaths in rural Burkina Faso: a prospective community-based cohort study.
Infant mortality rates (IMR) remain high in many sub-Saharan African countries, especially in rural settings where access to health services may be limited. Studies in such communities can provide relevant data on the burden of and risk factors for infant death. We measured IMR and explored risk factors for infant death in a cohort of children born in Banfora Health District, a rural area in South-West Burkina Faso. A prospective community-based cohort study was nested within the PROMISE-EBF trial (NCT00397150) in 24 villages of the study area. Maternal and infant baseline characteristics were collected at recruitment and after birth, respectively. Home visits were conducted at weeks 3, 6, 12, 24 and 52 after birth. Descriptive statistics were calculated using robust standard errors to account for cluster sampling. Cox multivariable regression was used to investigate potential risk factors for infant death. Among the 866 live born children included in the study there were 98 infant deaths, yielding an IMR of 113 per 1000 live births (95% CI: 89-143). Over 75% of infant deaths had occurred by 6 months of age and the post neonatal infant mortality rate was 67 per 1000 live births (95% CI: 51-88). Infections (35%) and preterm births complications (23%) were the most common probable causes of death by 6 months. Multivariable analyses identified maternal history of child death, polygyny, twin births and poor anthropometric z-scores at week-3 as factors associated with increased risk of infant death. We observed a very high IMR in a rural area of Burkina Faso, a country where 75% of the population lives in rural settings. Community-based health interventions targeting mothers and children at high risk are urgently needed to reduce the high burden of infant deaths in these areas.


Cortrak-Assisted Feeding Tube Insertion: A Comprehensive Review of Adverse Events in the MAUDE Database.
Electromagnetic devices to guide feeding tube placement such as the CORTRAK Enteral Access System have shown promising results; however, researchers in recent studies have expressed concern that a higher level of user expertise may be required for safe use. To review adverse events related to CORTRAK-assisted feeding tube insertion reported in the Food and Drug Administration's Manufacturer and User Facility Device Experience (MAUDE) database. A retrospective, secondary analysis of the MAUDE database was performed to evaluate adverse events (ie, injury or death of patient) related to CORTRAK. Fifty-four adverse events between January 1, 2006 and February 29, 2016 were identified and reviewed. Most events (98%) involved feeding tube placement in the lungs (37%, left lung; 46%, right lung; 15%, not specified). Lung complications included pneumothorax (77%) and pneumonitis (21%). Death occurred in 17% of lung placements. Clinicians failed to recognize placement in 89% of CORTRAK insertion tracings reviewed. Lung placement is not unique to CORTRAK and is an inherent risk of all feeding tube insertions. In known or suspected lung placement, feeding tubes should be removed and radiography performed to assess for pneumothorax. Clinicians must observe closely for lung placement and discriminate lung from gastric placement on insertion tracings. Clinicians require specialized training and experience to develop competency in using the CORTRAK device, although the exact amount of experience needed is unknown.


Changing spatial patterns of DNA replication in the noise-damaged chick cochlea.
The purpose of the present study was to examine the spatio-temporal pattern of cell proliferation in the chick cochlea in response to the sensory hair cell loss induced by a 1.5 kHz pure tone at 120 dB SPL (1 dB = 20 muPa) for 48 h. DNA replication was evaluated with the bromodeoxyuridine (BrdU) pulse-fix technique. One group of birds was given multiple injections of BrdU (50 mg/kg) over a period of 8 h at various starting times during or after the exposure. Afterwards, their cochleas were removed and processed as whole mounts for BrdU immunohistochemistry. The cochleas of a second group of acoustically traumatized chicks were evaluated by scanning electron microscopy in order to determine the spatio-temporal pattern of hair cell loss. Hair cell loss was first observed 12 h after the start of the exposure and DNA replication started near the inferior edge of the hair cell lesion 24-32 h after the start of the exposure, i.e. 12-20 h after the first sign of hair cell loss. The site of hair cell loss and DNA replication shifted toward the superior edge of the basilar papilla as the exposure continued. The rate of DNA replication accelerated and reached its peak near the end of the 48 h exposure. The estimated latency of cell proliferation after hair cell loss was faster and the duration of DNA replication shorter than that observed in other sensory systems. The spatio-temporal pattern of DNA replication follows the spatio-temporal gradient of hair cell loss, suggesting that cell proliferation is triggered by hair cell loss itself rather than by intrinsic positional cues or gradients.


Activation of the PI3K/AKT pathway in Merkel cell carcinoma.
Merkel cell carcinoma (MCC) is a highly aggressive skin cancer with an increasing incidence. The understanding of the molecular carcinogenesis of MCC is limited. Here, we scrutinized the PI3K/AKT pathway, one of the major pathways activated in human cancer, in MCC. Immunohistochemical analysis of 41 tumor tissues and 9 MCC cell lines revealed high levels of AKT phosphorylation at threonine 308 in 88% of samples. Notably, the AKT phosphorylation was not correlated with the presence or absence of the Merkel cell polyoma virus (MCV). Accordingly, knock-down of the large and small T antigen by shRNA in MCV positive MCC cells did not affect phosphorylation of AKT. We also analyzed 46 MCC samples for activating PIK3CA and AKT1 mutations. Oncogenic PIK3CA mutations were found in 2/46 (4%) MCCs whereas mutations in exon 4 of AKT1 were absent. MCC cell lines demonstrated a high sensitivity towards the PI3K inhibitor LY-294002. This finding together with our observation that the PI3K/AKT pathway is activated in the majority of human MCCs identifies PI3K/AKT as a potential new therapeutic target for MCC patients.


Highly sensitive SERS analysis of the cyclic Arg-Gly-Asp peptide ligands of cells using nanogap antennas.
The cyclic RGD (cRGD) peptide ligands of cells have become widely used for treating several cancers. We report a highly sensitive analysis of c(RGDfC) using surface enhanced Raman spectroscopy (SERS) using single dimer nanogap antennas in aqueous environment. Good agreement between characteristic peaks of the SERS and the Raman spectra of bulk c(RGDfC) with its peptide's constituents were observed. The exhibited blinking of the SERS spectra and synchronization of intensity fluctuations, suggest that the SERS spectra acquired from single dimer nanogap antennas was dominated by the spectrum of single to a few molecules. SERS spectra of c(RGDfC) could be used to detect at the nanoscale, the cells' transmembrane proteins binding to its ligand. SERS of cyclic RGD on nanogap antenna.


Differences across health care systems in outcome and cost-utility of surgical and conservative treatment of chronic low back pain: a study protocol.
There is little evidence on differences across health care systems in choice and outcome of the treatment of chronic low back pain (CLBP) with spinal surgery and conservative treatment as the main options. At least six randomised controlled trials comparing these two options have been performed; they show conflicting results without clear-cut evidence for superior effectiveness of any of the evaluated interventions and could not address whether treatment effect varied across patient subgroups. Cost-utility analyses display inconsistent results when comparing surgical and conservative treatment of CLBP. Due to its higher feasibility, we chose to conduct a prospective observational cohort study. This study aims to examine if1. Differences across health care systems result in different treatment outcomes of surgical and conservative treatment of CLBP2. Patient characteristics (work-related, psychological factors, etc.) and co-interventions (physiotherapy, cognitive behavioural therapy, return-to-work programs, etc.) modify the outcome of treatment for CLBP3. Cost-utility in terms of quality-adjusted life years differs between surgical and conservative treatment of CLBP. This study will recruit 1000 patients from orthopaedic spine units, rehabilitation centres, and pain clinics in Switzerland and New Zealand. Effectiveness will be measured by the Oswestry Disability Index (ODI) at baseline and after six months. The change in ODI will be the primary endpoint of this study. Multiple linear regression models will be used, with the change in ODI from baseline to six months as the dependent variable and the type of health care system, type of treatment, patient characteristics, and co-interventions as independent variables. Interactions will be incorporated between type of treatment and different co-interventions and patient characteristics. Cost-utility will be measured with an index based on EQol-5D in combination with cost data. This study will provide evidence if differences across health care systems in the outcome of treatment of CLBP exist. It will classify patients with CLBP into different clinical subgroups and help to identify specific target groups who might benefit from specific surgical or conservative interventions. Furthermore, cost-utility differences will be identified for different groups of patients with CLBP. Main results of this study should be replicated in future studies on CLBP.


Can electric fields explain inter-individual variability in transcranial direct current stimulation of the motor cortex?
The effects of transcranial direct current stimulation (tDCS) on motor cortical excitability are highly variable between individuals. Inter-individual differences in the electric fields generated in the brain by tDCS might play a role in the variability. Here, we explored whether these fields are related to excitability changes following anodal tDCS of the primary motor cortex (M1). Motor evoked potentials (MEPs) were measured in 28 healthy subjects before and after 20 min sham or 1 mA anodal tDCS of right M1 in a double-blind crossover design. The electric fields were individually modelled based on magnetic resonance images. Statistical analysis indicated that the variability in the MEPs could be partly explained by the electric fields, subjects with the weakest and strongest fields tending to produce opposite changes in excitability. To explain the findings, we hypothesized that the likely locus of action was in the hand area of M1, and the effective electric field component was that in the direction normal to the cortical surface. Our results demonstrate that a large part of inter-individual variability in tDCS may be due to differences in the electric fields. If this is the case, electric field dosimetry could be useful for controlling the neuroplastic effects of tDCS.


Polarised epithelial monolayers of the gastric mucosa reveal insights into mucosal homeostasis and defence against infection.
Helicobacter pylori causes life-long colonisation of the gastric mucosa, leading to chronic inflammation with increased risk of gastric cancer. Research on the pathogenesis of this infection would strongly benefit from an authentic human in vitro model. Antrum-derived gastric glands from surgery specimens served to establish polarised epithelial monolayers via a transient air-liquid interface culture stage to study cross-talk with H. pylori and the adjacent stroma. The resulting 'mucosoid cultures', so named because they recapitulate key characteristics of the gastric mucosa, represent normal stem cell-driven cultures that can be passaged for months. These highly polarised columnar epithelial layers encompass the various gastric antral cell types and secrete mucus at the apical surface. By default, they differentiate towards a foveolar, MUC5AC-producing phenotype, whereas Wnt signalling stimulates proliferation of MUC6-producing cells and preserves stemness-reminiscent of the gland base. Stromal cells from the lamina propria secrete Wnt inhibitors, antagonising stem-cell niche signalling and inducing differentiation. On infection with H. pylori, a strong inflammatory response is induced preferentially in the undifferentiated basal cell phenotype. Infection of cultures for several weeks produces foci of viable bacteria and a persistent inflammatory condition, while the secreted mucus establishes a barrier that only few bacteria manage to overcome. Gastric mucosoid cultures faithfully reproduce the features of normal human gastric epithelium, enabling new approaches for investigating the interaction of H. pylori with the epithelial surface and the cross-talk with the basolateral stromal compartment. Our observations provide striking insights in the regulatory circuits of inflammation and defence.


Gas mixing apparatus for determining cardiac output by CO2 rebreathing.
The carbon dioxide rebreathing technique is widely used for determination of cardiac output during exercise. The equilibration method of Collier et at. (J. Appl. Physiol. 9:25, 1956) is generally preferred over the exponential method of Defares (J. Appl. Physiol. 13:159, 1968). However, the equilibration method requires the volume and initial CO2 percentage in the rebreathing bag to be adjusted according to the work rate. A device for mixing two gases (100% O2 and 20% CO2/80% O2) was constructed for this purpose. Multistage regulators are attached to the gas tanks and connected to a medical gas mixer via high-pressure air hoses. A variable time-delay switch causes a solenoid valve to open for 1.0-10s to deliver a predetermined gas volume. The device was found to accurately deliver a preset volume and concentration of gas to the rebreathing bag. A gas mixing apparatus simplifies the equilibration CO2 rebreathing technique by allowing the investigator to easily select the initial volume and percentage of CO2.


Fasting for weight loss: an effective strategy or latest dieting trend?
With the increasing obesity epidemic comes the search for effective dietary approaches for calorie restriction and weight loss. Here I examine whether fasting is the latest 'fad diet' as portrayed in popular media and discuss whether it is a safe and effective approach or whether it is an idiosyncratic diet trend that promotes short-term weight loss, with no concern for long-term weight maintenance. Fasting has long been used under historical and experimental conditions and has recently been popularised by 'intermittent fasting' or 'modified fasting' regimes, in which a very low-calorie allowance is allowed, on alternate days (ADF) or 2 days a week (5:2 diet), where 'normal' eating is resumed on non-diet days. It is a simple concept, which makes it easy to follow with no difficult calorie counting every other day. This approach does seem to promote weight loss, but is linked to hunger, which can be a limiting factor for maintaining food restriction. The potential health benefits of fasting can be related to both the acute food restriction and chronic influence of weight loss; the long-term effect of chronic food restriction in humans is not yet clear, but may be a potentially interesting future dietary strategy for longevity, particularly given the overweight epidemic. One approach does not fit all in the quest to achieve body weight control, but this could be a dietary strategy for consideration. With the obesity epidemic comes the search for dietary strategies to (i) prevent weight gain, (ii) promote weight loss and (iii) prevent weight regain. With over half of the population of the United Kingdom and other developed countries being collectively overweight or obese, there is considerable pressure to achieve these goals, from both a public health and a clinical perspective. Certainly not one dietary approach will solve these complex problems. Although there is some long-term success with gastric surgical options for morbid obesity, there is still a requirement for dietary approaches for weight management for the overweight and obese population, particularly as invasive interventions carry post-operative risk of death due to complications. Effective dietary interventions are required that promote long-term adherence and sustained beneficial effects on metabolic and disease markers. In general, such interventions need to be palatable and satiating, meet minimal nutritional requirements, promote loss of fat and preserve lean body mass, ensure long-term safety, be simple to administer and monitor and have widespread public health utility. Intermittent fasting or alternate day fasting may be an option for achieving weight loss and maintenance.


The time course of changes during motor sequence learning: a whole-brain fMRI study.
There is a discrepancy between the results of imaging studies in which subjects learn motor sequences. Some experiments have shown decreases in the activation of some areas as learning increased, whereas others have reported learning-related increases as learning progressed. We have exploited fMRI to measure changes in blood oxygen leve-dependent (BOLD) signal throughout the course of learning. T2*-weighted echo-planar images were acquired over the whole brain for 40 min while the subjects learned a sequence eight moves long by trial and error. The movements were visually paced every 3.2 s and visual feedback was provided to the subjects. A baseline period followed each activation period. The effect due to the experimental conditions was modeled using a square-wave function, time locked to their occurrence. Changes over time in the difference between activation and baseline signal were modeled using a set of polynomial basis functions. This allowed us to take into account linear as well as nonlinear changes over time. Low-frequency changes over time common to both activation and baseline conditions (and thus not learning related) were modeled and removed. Linear and nonlinear changes of BOLD signal over time were found in prefrontal, premotor, and parietal cortex and in neostriatal and cerebellar areas. Single-unit recordings in nonhuman primates during the learning of motor tasks have clearly shown increased activity early in learning, followed by a decrease as learning progressed. Both phenomena can be observed at the population level in the present study.


Temporal integration of auxin information for the regulation of patterning.
Positional information is essential for coordinating the development of multicellular organisms. In plants, positional information provided by the hormone auxin regulates rhythmic organ production at the shoot apex, but the spatio-temporal dynamics of auxin gradients is unknown. We used quantitative imaging to demonstrate that auxin carries high-definition graded information not only in space but also in time. We show that, during organogenesis, temporal patterns of auxin arise from rhythmic centrifugal waves of high auxin travelling through the tissue faster than growth. We further demonstrate that temporal integration of auxin concentration is required to trigger the auxin-dependent transcription associated with organogenesis. This provides a mechanism to temporally differentiate sites of organ initiation and exemplifies how spatio-temporal positional information can be used to create rhythmicity.


Effects of baked and raw salmon fillet on lipids and n-3 PUFAs in serum and tissues in Zucker fa/fa rats.
Knowledge of the health impact of consuming heat-treated versus raw fish fillet is limited. To investigate effects of baked or raw salmon fillet intake on lipids and n-3 PUFAs in serum and tissues, obese Zucker fa/fa rats were fed diets containing 25% of protein from baked or raw salmon fillet and 75% of protein from casein, or casein as the sole protein source (control group) for four weeks. Salmon diets had similar composition of amino and fatty acids. Growth and energy intake were similar in all groups. Amounts of lipids and n-3 PUFAs in serum, liver and skeletal muscle were similar between rats fed baked or raw salmon fillet. When compared to the control group, rats fed baked salmon had lower serum total and LDL cholesterol and higher serum triacylglycerol levels. Both raw and baked salmon groups had lower HDL cholesterol level when compared to control rats. In conclusion, baking as a preparation method does not alter protein and fat qualities of salmon fillets, and intake of baked and raw salmon fillets gave similar effects on lipids and n-3 PUFAs in serum and tissues from rats.


Focus on TILs: prognostic significance of tumor infiltrating lymphocytes in human colorectal cancer.
Human colorectal cancer tissues are infiltrated by various immune/inflammatory cells, usually along the invasive margin. These responses tend be regarded as "non-specific". However, it is now clear that these cellular responses, particularly lymphocytic reactions, are independent prognostic factors for a better survival. Immunohistochemical subset analyses have generally disclosed that the number of T-lymphocytes is important. The effects of these T cells tend to be more manifest when the observation periods are longer. These data suggest that some degrees of anti-tumor immunity exist in human colorectal carcinomas. However, human tumors are generally composed of various histologic subtypes, which sometimes complicates these analyses and simple explanations may be misleading. Considered with precaution, these morphologic analyses on immune cell subtypes are important tools to understand the immune responses in human cancers.


Replacement of starch from corn with nonforage fiber from distillers grains and soyhulls in diets of lactating dairy cows.
Forty Holstein cows were used in a completely randomized design with a 2-wk covariate period followed by a 6-wk experimental period to evaluate incremental substitution of nonforage fiber provided by dried distillers grains with solubles (DDGS) and soyhulls (SH) for starch provided by corn in the diet. Diets provided decreasing concentrations of starch: 29% starch with 0% DDGS; 26% starch with 7% DDGS; 23% starch with 14% DDGS; and 20% starch with 21% DDGS. Diets contained 27% corn silage, 22% alfalfa hay, and 51% concentrate mix and were formulated to be 17% crude protein, 4.7% fat, and 23% neutral detergent fiber from forage. Total neutral detergent fiber increased as DDGS and SH were included in the diet. Soyhulls were included in a linear fashion along with DDGS to replace soybean meal and expeller soybean meal, thereby maintaining a similar crude protein content across diets. Dry matter intake decreased linearly; consequently, feed efficiency tended to increase linearly as starch was replaced by nonforage fiber. There was no effect of diet on milk production or milk fat and protein percentage or yield. Milk fatty acid profiles were similar across diets. Other response variables, including 4% fat-corrected milk, total solids, and milk urea nitrogen, were unaffected by dietary treatments. Ruminal volatile fatty acid concentration did not differ between diets. Concentrations of blood glucose and beta-hydroxybutyrate were similar across diets. Results from this research suggest that nonforage fiber from DDGS can partially substitute for starch from corn in dairy cow diets without affecting milk production and milk composition. Economic analysis of the diets showed that feeding DDGS and SH in substitution of corn was cost-effective. Results from this experiment indicate that DDGS and SH can replace corn as an energy source to decrease feed costs.


Active Inference and Epistemic Value in Graphical Models.
The Free Energy Principle (FEP) postulates that biological agents perceive and interact with their environment in order to minimize a Variational Free Energy (VFE) with respect to a generative model of their environment. The inference of a policy (future control sequence) according to the FEP is known as Active Inference (AIF). The AIF literature describes multiple VFE objectives for policy planning that lead to epistemic (information-seeking) behavior. However, most objectives have limited modeling flexibility. This paper approaches epistemic behavior from a constrained Bethe Free Energy (CBFE) perspective. Crucially, variational optimization of the CBFE can be expressed in terms of message passing on free-form generative models. The key intuition behind the CBFE is that we impose a point-mass constraint on predicted outcomes, which explicitly encodes the assumption that the agent will make observations in the future. We interpret the CBFE objective in terms of its constituent behavioral drives. We then illustrate resulting behavior of the CBFE by planning and interacting with a simulated T-maze environment. Simulations for the T-maze task illustrate how the CBFE agent exhibits an epistemic drive, and actively plans ahead to account for the impact of predicted outcomes. Compared to an EFE agent, the CBFE agent incurs expected reward in significantly more environmental scenarios. We conclude that CBFE optimization by message passing suggests a general mechanism for epistemic-aware AIF in free-form generative models.


Directional transneuronal infection by pseudorabies virus is dependent on an acidic internalization motif in the Us9 cytoplasmic tail.
The Us9 gene is conserved among most alphaherpesviruses. In pseudorabies virus (PRV), the Us9 protein is a 98-amino-acid, type II membrane protein found in the virion envelope. It localizes to the trans-Golgi network (TGN) region in infected and transfected cells and is maintained in this compartment by endocytosis from the plasma membrane. Viruses with Us9 deleted have no observable defects in tissue culture yet have reduced virulence and restricted spread to retinorecipient neurons in the rodent brain. In this report, we demonstrate that Us9-promoted transneuronal spread in vivo is dependent on a conserved acidic motif previously shown to be essential for the maintenance of Us9 in the TGN region and recycling from the plasma membrane. Mutant viruses with the acidic motif deleted have an anterograde spread defect indistinguishable from that of Us9 null viruses. Transneuronal spread, however, is not dependent on a dileucine endocytosis motif in the Us9 cytoplasmic tail. Through alanine scanning mutagenesis of the acidic motif, we have identified two conserved tyrosine residues that are essential for Us9-mediated spread as well as two serine residues, comprising putative consensus casein kinase II sites, that modulate the rate of PRV transneuronal spread in vivo.


Towards understanding the first genome sequence of a crenarchaeon by genome annotation using clusters of orthologous groups of proteins (COGs).
Standard archival sequence databases have not been designed as tools for genome annotation and are far from being optimal for this purpose. We used the database of Clusters of Orthologous Groups of proteins (COGs) to reannotate the genomes of two archaea, Aeropyrum pernix, the first member of the Crenarchaea to be sequenced, and Pyrococcus abyssi. A. pernix and P. abyssi proteins were assigned to COGs using the COGNITOR program; the results were verified on a case-by-case basis and augmented by additional database searches using the PSI-BLAST and TBLASTN programs. Functions were predicted for over 300 proteins from A. pernix, which could not be assigned a function using conventional methods with a conservative sequence similarity threshold, an approximately 50% increase compared to the original annotation. A. pernix shares most of the conserved core of proteins that were previously identified in the Euryarchaeota. Cluster analysis or distance matrix tree construction based on the co-occurrence of genomes in COGs showed that A. pernix forms a distinct group within the archaea, although grouping with the two species of Pyrococci, indicative of similar repertoires of conserved genes, was observed. No indication of a specific relationship between Crenarchaeota and eukaryotes was obtained in these analyses. Several proteins that are conserved in Euryarchaeota and most bacteria are unexpectedly missing in A. pernix, including the entire set of de novo purine biosynthesis enzymes, the GTPase FtsZ (a key component of the bacterial and euryarchaeal cell-division machinery), and the tRNA-specific pseudouridine synthase, previously considered universal. A. pernix is represented in 48 COGs that do not contain any euryarchaeal members. Many of these proteins are TCA cycle and electron transport chain enzymes, reflecting the aerobic lifestyle of A. pernix. Special-purpose databases organized on the basis of phylogenetic analysis and carefully curated with respect to known and predicted protein functions provide for a significant improvement in genome annotation. A differential genome display approach helps in a systematic investigation of common and distinct features of gene repertoires and in some cases reveals unexpected connections that may be indicative of functional similarities between phylogenetically distant organisms and of lateral gene exchange.


Isolation and partial characterization of the human homologue of Thy-1.
A glycoprotein of 25,000 daltons was isolated from a human T lymphoblastoid cell line (MOLT-3). A monovalent antiserum raised against this antigen precipitated mouse Thy-1.1 antigen as well as a 25,000-dalton antigen from human blood T cells. The unabsorbed serum was also cytotoxic for both mouse and human T cells. By multiple criteria, this human T antigen appears to be homologous with mouse Thy-1.


Design and operation of the wide angular-range chopper spectrometer ARCS at the Spallation Neutron Source.
The wide angular-range chopper spectrometer ARCS at the Spallation Neutron Source (SNS) is optimized to provide a high neutron flux at the sample position with a large solid angle of detector coverage. The instrument incorporates modern neutron instrumentation, such as an elliptically focused neutron guide, high speed magnetic bearing choppers, and a massive array of (3)He linear position sensitive detectors. Novel features of the spectrometer include the use of a large gate valve between the sample and detector vacuum chambers and the placement of the detectors within the vacuum, both of which provide a window-free final flight path to minimize background scattering while allowing rapid changing of the sample and sample environment equipment. ARCS views the SNS decoupled ambient temperature water moderator, using neutrons with incident energy typically in the range from 15 to 1500 meV. This range, coupled with the large detector coverage, allows a wide variety of studies of excitations in condensed matter, such as lattice dynamics and magnetism, in both powder and single-crystal samples. Comparisons of early results to both analytical and Monte Carlo simulation of the instrument performance demonstrate that the instrument is operating as expected and its neutronic performance is understood. ARCS is currently in the SNS user program and continues to improve its scientific productivity by incorporating new instrumentation to increase the range of science covered and improve its effectiveness in data collection.


Direct visualization of the microtubule lattice seam both in vitro and in vivo.
Microtubules are constructed from alpha- and beta-tubulin heterodimers that are arranged into protofilaments. Most commonly there are 13 or 14 protofilaments. A series of structural investigations using both electron microscopy and x-ray diffraction have indicated that there are two potential lattices (A and B) in which the tubulin subunits can be arranged. Electron microscopy has shown that kinesin heads, which bind only to beta-tubulin, follow a helical path with a 12-nm pitch in which subunits repeat every 8-nm axially, implying a primarily B-type lattice. However, these helical symmetry parameters are not consistent with a closed lattice and imply that there must be a discontinuity or "seam" along the microtubule. We have used quick-freeze deep-etch electron microscopy to obtain the first direct evidence for the presence of this seam in microtubules formed either in vivo or in vitro. In addition to a conventional single seam, we have also rarely found microtubules in which there is more than one seam. Overall our data indicates that microtubules have a predominantly B lattice, but that A lattice bonds between tubulin subunits are found at the seam. The cytoplasmic microtubules in mouse nerve cells also have predominantly B lattice structure and A lattice bonds at the seam. These observations have important implications for the interaction of microtubules with MAPs and with motor proteins, and for example, suggest that kinesin motors may follow a single protofilament track.


Coordinated movement, neuromuscular synaptogenesis and trans-synaptic signaling defects in Drosophila galactosemia models.
The multiple galactosemia disease states manifest long-term neurological symptoms. Galactosemia I results from loss of galactose-1-phosphate uridyltransferase (GALT), which converts galactose-1-phosphate + UDP-glucose to glucose-1-phosphate + UDP-galactose. Galactosemia II results from loss of galactokinase (GALK), phosphorylating galactose to galactose-1-phosphate. Galactosemia III results from the loss of UDP-galactose 4'-epimerase (GALE), which interconverts UDP-galactose and UDP-glucose, as well as UDP-N-acetylgalactosamine and UDP-N-acetylglucosamine. UDP-glucose pyrophosphorylase (UGP) alternatively makes UDP-galactose from uridine triphosphate and galactose-1-phosphate. All four UDP-sugars are essential donors for glycoprotein biosynthesis with critical roles at the developing neuromuscular synapse. Drosophila galactosemia I (dGALT) and II (dGALK) disease models genetically interact; manifesting deficits in coordinated movement, neuromuscular junction (NMJ) development, synaptic glycosylation, and Wnt trans-synaptic signalling. Similarly, dGALE and dUGP mutants display striking locomotor and NMJ formation defects, including expanded synaptic arbours, glycosylation losses, and differential changes in Wnt trans-synaptic signalling. In combination with dGALT loss, both dGALE and dUGP mutants compromise the synaptomatrix glycan environment that regulates Wnt trans-synaptic signalling that drives 1) presynaptic Futsch/MAP1b microtubule dynamics and 2) postsynaptic Frizzled nuclear import (FNI). Taken together, these findings indicate UDP-sugar balance is a key modifier of neurological outcomes in all three interacting galactosemia disease models, suggest that Futsch homolog MAP1B and the Wnt Frizzled receptor may be disease-relevant targets in epimerase and transferase galactosemias, and identify UGP as promising new potential therapeutic target for galactosemia neuropathology.


Peptide-dependent expression of HLA-B7 on antigen processing-deficient T2 cells.
Class I MHC Ag presentation and cell surface expression largely depend on peptide transport into the ER/cis-Golgi by TAP, the transporter associated with Ag processing. Despite this dependency, class I MHC molecules are expressed at low levels on the surface of TAP-deficient T2 cells. We studied the peptide dependency of HLA-B7 expression in transfected T2 cells. HLA-B7 expression was affected by mutations at 19 out of 23 peptide-binding groove residues, but not by nine mutations outside of the peptide-binding groove. T2 cell surface HLA-A2, -B7, and -B51 had similar stabilities, and approximately half of these class I molecules had a long t1/2 consistent with tight peptide binding. Using metabolically labeled T2 cells, HLA-A2-bound peptide eluted as five prominent peaks, but HLA-B7-bound peptide was not detected. In contrast, HLA-B7-eluted peptides were detected spectrophotometrically. These data suggest that HLA-A2 and HLA-B7 molecules utilize distinct TAP-independent peptide supply mechanisms to different degrees. Equivalent amounts of HLA-B7 from TAP- and TAP+ cells yielded similar amounts of peptide, which had the characteristic HLA-B7 peptide-binding motif. The dependency of HLA-B7 cell surface expression on peptide-binding groove residues, the stability of cell surface class I molecules, and the ability to detect HLA-B7-bound peptide indicate that the low level expression on T2 cells is largely peptide dependent. TAP-independent peptide Ag presentation may allow immune recognition of intracellular pathogens that interfere with TAP-dependent peptide transport.


The segment-specific gene Krox-20 encodes a transcription factor with binding sites in the promoter region of the Hox-1.4 gene.
Krox-20 is a mouse zinc finger gene expressed in a segment-specific manner in the early central nervous system, which makes it a potential developmental control gene. In this report, we show that the Krox-20 protein binds in vitro to two specific DNA sites located upstream from the homeobox containing gene Hox-1.4. The nucleotide sequence recognized by Krox-20 is closely related to the Sp1 target sequence, which is consistent with the similarity existing between the zinc fingers of the two proteins. In co-transfection experiments in cultured cells, Krox-20 dramatically activates transcription from the herpes simplex virus thymidine kinase promoter when an oligomer of its binding site is present in cis close to the promoter. Analysis of mutated binding sites demonstrates that the level of activation by Krox-20 correlates with the affinity of the protein for the mutant sequence. These data indicate that Krox-20 constitutes a sequence-specific DNA-binding transcription factor. Parallel analysis of the expression of Krox-20 and Hox-1.4 in the neural tube by in situ hybridization revealed no overlap, arguing against direct interactions between these two genes. The possible involvement of Krox-20 in the regulation of the transcription of other homeobox genes is discussed in view of their respective patterns of expression.


Pilot study protocol to inform a future longitudinal study of ageing using linked administrative data: Healthy AGeing in Scotland (HAGIS).
Population ageing is a welcome testament to improvements in the social, economic and health circumstances over the life course. However, these successes necessitate that we understand more about the pathways of ageing to plan and cost our health and social care services, to support our ageing population to live healthier for longer and to make adequate provisions for retirement. Longitudinal studies of ageing facilitate such understanding in many countries around the world. Scotland presently does not have a longitudinal study of ageing, despite dramatic increases to its ageing population and its poor health record. Healthy AGeing in Scotland (HAGIS) constitutes the launch of Scotland's first comprehensive longitudinal study of ageing. A sample of 1000 people aged 50+ years will be invited to take part in a household social survey. The innovative sampling procedure used administrative data to identify eligible households. Anonymised survey responses will be linked to administrative data. Ethics approval was obtained from the host institution for the study design and from the Public Benefits and Privacy Panel for administrative data linkage. Anonymised survey data will be deposited with the UK Data Service. A subset of survey data, harmonised with other global ageing studies, will be available via the Gateway to Global Aging platform. These data will enable powerful cross-country comparisons across the social, economic and health domains that will be relevant for national and international research.Research publications from the HAGIS team will be disseminated through journal articles and national and international conferences. The findings will support current and future research and policy debate on ageing populations.


ID1 and ID3 are Negative Regulators of TGFβ2-Induced Ocular Hypertension and Compromised Aqueous Humor Outflow Facility in Mice.
In POAG, elevated IOP remains the major risk factor in irreversible vision loss. Increased TGFβ2 expression in POAG aqueous humor and in the trabecular meshwork (TM) amplifies extracellular matrix (ECM) deposition and reduces ECM turnover in the TM, leading to a decreased aqueous humor (AH) outflow facility and increased IOP. Inhibitor of DNA binding proteins (ID1 and ID3) inhibit TGFβ2-induced fibronectin and PAI-1 production in TM cells. We examined the effects of ID1 and ID3 gene expression on TGFβ2-induced ocular hypertension and decreased AH outflow facility in living mouse eyes. IOP and AH outflow facility changes were determined using a mouse model of Ad5-hTGFβ2C226S/C288S-induced ocular hypertension. The physiological function of ID1 and ID3 genes were evaluated using Ad5 viral vectors to enhance or knockdown ID1/ID3 gene expression in the TM of BALB/cJ mice. IOP was measured in conscious mice using a Tonolab impact tonometer. AH outflow facilities were determined by constant flow infusion in live mice. Over-expressing ID1 and ID3 significantly blocked TGFβ2-induced ocular hypertension (P < 0.0001). Although AH outflow facility was significantly decreased in TGFβ2-transduced eyes (P < 0.04), normal outflow facility was preserved in eyes injected concurrently with ID1 or ID3 along with TGFβ2. Knockdown of ID1 or ID3 expression exacerbated TGFβ2-induced ocular hypertension. Increased expression of ID1 and ID3 suppressed both TGFβ2-elevated IOP and decreased AH outflow facility. ID1 and/or ID3 proteins thus may show promise as future candidates as IOP-lowering targets in POAG.


Maximizing the opening of eye diagrams for slow-light systems.
We present a data-fidelity metric for quantifying distortion in slow-light optical pulse delay devices. We demonstrate the utility of this metric by applying it to the performance optimization of gain-based slow-light delay systems for Gaussian and super-Gaussian pulses. Symmetric Lorentzian double-line and triple-line gain systems are optimized and achieve maximum delay of 1.5 and 1.7 times the single-line gain system delay, respectively. The resulting double-line gain system design is qualitatively similar to the double-line gain system designed with a previous metric, but is tuned specifically to constrain data fidelity.


Microbiological laboratory hazard of bearded men.
An investigation was conducted to evaluate the hypothesis that a bearded man subjects his family and friends to risk of infection if his beard is contaminated by infectious microorganisms while he is working in a microbiological laboratory. Bearded and unbearded men were tested with Serratia marcescens and Bacillus subtilis var. niger. Contact aerosol transmission from a contaminated beard on a mannequin to a suitable host was evaluated with both Newcastle disease virus and Clostridium botulinum toxin, type A. The experiments showed that beards retained microorganisms and toxin despite washing with soap and water. Although washing reduced the amount of virus or toxin, a sufficient amount remained to produce disease upon contact with a suitable host.


Versatile design and synthesis of nano-barcodes.
Encoded nano-structures/particles have been used for barcoding and are in great demand for the simultaneous analysis of multiple targets. Due to their nanoscale dimension(s), nano-barcodes have been implemented favourably for bioimaging, in addition to their security and multiplex bioassay application. In designing nano-barcodes for a specific application, encoding techniques, synthesis strategies, and decoding techniques need to be considered. The encoding techniques to generate unique multiple codes for nano-barcodes are based on certain encoding elements including optical (fluorescent and non-fluorescent), graphical, magnetic, and phase change properties of nanoparticles or their different shapes and sizes. These encoding elements can generally be embedded inside, decorated on the surface of nanostructures or self-assembled to prepare the nano-barcodes. The decoding techniques for each encoding technique are different and need to be suitable for the desired applications. This review will provide a thorough discussion on designing nano-barcodes, focusing on the encoding techniques, synthesis methods, and decoding for applications including bio-detection, imaging, and anti-counterfeiting. Additionally, associated challenges in the field and potential solutions will also be discussed. We believe that a comprehensive understanding on this topic could significantly contribute towards the advancement of nano-barcodes for a broad spectrum of applications.


Growth of well-arrayed ZnO nanorods on thinned silica fiber and application for humidity sensing.
Thinned silica fibers were fabricated by drawing conventional single mode silica fiber through flame heated method and well-arrayed ZnO nanorods were grown on the thinned silica fibers by a hydrothermal method. The structure enables efficient light coupling between the fiber and the nanorods. With the unique property of high surface to volume ratio of one-dimensional ZnO nanorods, light coupled to nanorods array enhances the optical interaction between the device and the ambient environment. Sensitive humidity sensor was demonstrated by launching laser into ZnO nanorod-covered fibers. Theoretical and experimental results are presented.


International aspects of the AIDS/HIV epidemic.
This review provides the reader with pertinent information on the epidemiology, prevention, and new technologies of the ongoing HIV pandemic. These aspects are key to international policy discussions surrounding the public health response to the international spread of HIV. Our understanding of the impact of AIDS on other diseases is evolving, as is our insight into the demographic and economic effects of the epidemic on the global community. Observations on the success of certain prevention strategies allow rational allocation of resources in newly affected epidemic areas. Information on the origin and nature of HIV transmission exemplifies the phenomenon of global emerging infections. As world populations are brought closer together through transportation, communication, trade, and commerce, insight into emerging infections of epidemic potential becomes increasingly important to the practitioner of public health. Although important, legal and social aspects of the epidemic will not be emphasized here. The epidemics of HIV/AIDS in the United States and Europe are not reviewed here. The global pandemic has recently been described in an overview in this publication to which the reader is also referred.


On the relationship between the "default mode network" and the "social brain".
The default mode network (DMN) of the brain consists of areas that are typically more active during rest than during active task performance. Recently however, this network has been shown to be activated by certain types of tasks. Social cognition, particularly higher-order tasks such as attributing mental states to others, has been suggested to activate a network of areas at least partly overlapping with the DMN. Here, we explore this claim, drawing on evidence from meta-analyses of functional MRI data and recent studies investigating the structural and functional connectivity of the social brain. In addition, we discuss recent evidence for the existence of a DMN in non-human primates. We conclude by discussing some of the implications of these observations.


Bone marrow transplantation for chronic myeloid leukemia with volunteer unrelated donors using ex vivo or in vivo T-cell depletion: major prognostic impact of HLA class I identity between donor and recipient.
Between August 1985 and July 1994, we performed 115 volunteer unrelated donor (VUD) bone marrow transplants (BMT) for first chronic phase (n = 86) or advanced phase (n = 29) chronic myeloid leukemia (CML). Standard serologic HLA typing of potential donors and recipients was supplemented with one-dimensional isoelectric focusing (IEF) for class I proteins, allogenotyping for DR and DQ alleles using DNA restriction fragment length polymorphism (RFLP) analysis, and the measurement of antirecipient major histocompatibility complex (MHC) cytotoxic T-lymphocyte precursor cells in the donors' blood (CTLp assay). Recipients were conditioned for transplantation with a combination of high-dose chemotherapy and total body irradiation (n = 103) or high-dose chemotherapy alone (n = 12). Twenty eight recipients received ex vivo T-cell-depleted marrow, and 84 underwent some form of in vivo T-cell depletion. The probability of severe (grades III or IV) acute graft-versus-host disease (aGVHD) was 24%, and that of extensive chronic graft-versus-host disease (cGVHD), 38%. Proportional hazards regression analysis showed an association between low frequency CTLp and a reduced incidence of severe aGVHD (relative risk [RR], 0.28; P = .0035). The probability of relapse at 3 years was 23%, with first chronic phase disease being independently associated with a lower risk of relapse (RR, 0.71; P = .01). The overall leukemia-free survival (LFS) at 3 years was 37%; the LFS for the first chronic phase and advanced phase recipients was 41% and 26%, respectively. First chronic phase disease (RR, 0.56; P = .063) and the combination of recipient cytomegalovirus (CMV) seronegativity and an IEF-matched donor (RR, 0.48; P = .011) were both associated with improved LFS. The probabilities of survival and LFS for patients under 40 years of age transplanted in first chronic phase from an IEF-matched donor were 73% and 50%, respectively. We conclude that VUD BMT is a reasonable option for patients with CML; when using ex vivo or in vivo T-cell depletion, optimal results are achieved in patients transplanted in chronic phase with marrow from donors without demonstrable class I HLA mismatch and a low CTLp frequency.


Factors associated with inflammation markers, a cross-sectional analysis.
Epidemiological studies have reported associations between circulating inflammation markers and risk of chronic diseases. It is of interest to examine whether risk factors for these diseases are associated with inflammation. We conducted a cross-sectional analysis to evaluate whether reproductive and lifestyle factors and circulating vitamin D were associated with inflammation markers, including C-reactive protein, cytokines (IL-1β, IL-2, IL-4, IL-5, IL-6, IL-10, IL-12p40, IL-12p70, IL-13, TNFα), and cytokine modulators (IL-1RA, sIL-1RII, sIL-2Ra, sIL-4R, sIL-6R, sTNF-R1/R2), among 616 healthy women. We confirmed associations of several inflammation markers with age and BMI. We also observed significantly higher levels of certain inflammation markers in postmenopausal vs. premenopausal women (TNFα, sIL-1RII, sIL-2Ra), with increasing parity (IL-12p40), and with higher circulating 25(OH) vitamin D (IL-13) and lower levels among current users of non-steroidal anti-inflammatory drugs (NSAIDs) (IL-1β, IL-2, IL-10, IL-12p70, and IL-12p40), current smokers (IL-4, IL-13, IL-12p40), and women with a family history of breast or ovarian cancer (IL-4, IL-10, IL-13). Our findings suggest that risk factors for chronic diseases (age, BMI, menopausal status, parity, NSAID use, family history of breast and ovarian cancer, and smoking) are associated with inflammation markers in healthy women.


Neurothekeoma palpebrae in association with multiple superficial angiomyxomas: Tegumental Angiomyxoma-Neurothekeoma syndrome (TAN syndrome).
We report a case of 10-year-old Indian girl with history of multiple superficial angiomyxoma, presented with three months history of painless right upper lid swelling. There were no visual dysfunctions. Previously, the patient had multiple superficial angiomyxoma (left pinna, left upper cheek, left upper limb, chest, right axilla, hard palate) and epidermal cyst (chin). The histopathological specimens were negative to S-100 protein antibody. Systemic review and family history was unremarkable. Excision biopsy and upper lid reconstruction were performed. Intraoperatively the tumor was multilobulated, firm, well encapsulated and did not invade the underlying tarsal plate. Histopathological features of the upperlid tumor were consistent with nerves sheath myxoma (neurothekeoma). To the best of the authors' knowledge, this is the first reported case of neurothekeoma in association with multiple superficial angiomyxoma.


Scaling up proven public health interventions through a locally owned and sustained leadership development programme in rural Upper Egypt.
In 2002, the Egypt Ministry of Health and Population faced the challenge of improving access to and quality of services in rural Upper Egypt in the face of low morale among health workers and managers.From 1992 to 2000, the Ministry, with donor support, had succeeded in reducing the nationwide maternal mortality rate by 52%. Nevertheless, a gap remained between urban and rural areas. In 2002, the Ministry, with funding from the United States Agency for International Development and assistance from Management Sciences for Health, introduced a Leadership Development Programme (LDP) in Aswan Governorate. The programme aimed to improve health services in three districts by increasing managers' ability to create high performing teams and lead them to achieve results.The programme introduced leadership and management practices and a methodology for identifying and addressing service delivery challenges. Ten teams of health workers participated. In 2003, after participation in the LDP, the districts of Aswan, Daraw and Kom Ombo increased the number of new family planning visits by 36%, 68% and 20%, respectively. The number of prenatal and postpartum visits also rose.After the United States funding ended, local doctors and nurses scaled up the programme to 184 health care facilities (training more than 1000 health workers). From 2005 to 2007, the Leadership Development Programme participants in Aswan Governorate focused on reducing the maternal mortality rate as their annual goal. They reduced it from 85.0 per 100,000 live births to 35.5 per 100,000. The reduction in maternal mortality rate was much greater than in similar governorates in Egypt. Managers and teams across Aswan demonstrated their ability to scale up effective public health interventions though their increased commitment and ownership of service challenges. When teams learn and apply empowering leadership and management practices, they can transform the way they work together and develop their own solutions to complex public health challenges. Committed health teams can use local resources to scale up effective public health interventions.


Mutation Screening of mtDNA Combined Targeted Exon Sequencing in a Cohort With Suspected Hereditary Optic Neuropathy.
Leber hereditary optic neuropathy (LHON) and autosomal dominant optic atrophy (ADOA) are the two commonest forms of hereditary optic neuropathy. The aim of this study was to comprehensively investigate the incidence and spectrum of mutations in patients with suspected hereditary optic neuropathy by combining mitochondrial DNA (mtDNA) genome-wide and targeted exon sequencing. A cohort of 1101 subjects were recruited to participate in the study, comprising 177 families (177 probands and their family members, a total of 537 subjects, including 254 patients) and 164 sporadic cases with suspected hereditary optic neuropathy, and 400 unrelated control subjects for genetic analysis: all subjects (including control subjects) underwent a comprehensive ophthalmologic examination and were subjected to sequencing analysis of mtDNA genome-wide and targeted exon. Overall, targeted exon sequencing was used to screen 792 genes associated with common hereditary eye diseases, and the mtDNA genome-wide were screened by next-generation sequencing. We found variants detected in 168 (40.2%, 168/418) of the 418 patients screened. Among these, 132 cases (78.6%, 132/168) were detected with known LHON disease-causing mtDNA variants; 40 cases (23.8%, 40/168) were detected with nuclear DNA (ntDNA) variants, which included 36 cases (21.4%, 36/168) with detected OPA1 mutations, 4 patients (2.4%, 4/168) with detected OPA3 mutations, and 2 patients (1.2%, 2/168) with detected TMEM126A homozygous mutation. Coexistence variation (mtDNA/mtDNA [n = 16], ntDNA/ntDNA [n = 4], mtDNA/ntDNA [n = 7]) was found in 27 patients (16.4%, 27/165), including mtDNA/ntDNA coexistence variation that was detected in seven patients. Among these ntDNA mutations, 38 distinct disease-causing variants, including autosomal recessive heterozygous mutations, were detected, which included 22 novel variants and two de novo variants. Total haplogroup distribution showed that 34.5% (29/84) and 28.6% (24/84) of the affected subjects with m.11778G>A belonged to haplogroup D and M, with a high frequency of subhaplogroups D4, D5, and M7. The LHON-mtDNA mutations are the commonest genetic defects in this Chinese cohort, followed by the OPA1 mutations. To our knowledge, this is the first comprehensive study of LHON, ADOA, and autosomal recessive optic atrophy combined with mtDNA genome-wide and targeted exon sequencing, as well as haplogroup analysis, in a large cohort of Chinese patients with suspected hereditary optic neuropathy. Our findings provide a powerful basis for genetic counseling in patients with suspected hereditary optic neuropathy. We applied mtDNA genome-wide sequencing combined with panel-based targeted exon sequencing to explore the pathogenic variation spectrum and genetic characteristics of patients with suspected hereditary optic neuropathy, providing a comprehensive research strategy for clinical assistant diagnosis, treatment, and genetic counseling.


Detection of steroid 21-hydroxylase alleles using gene-specific PCR and a multiplexed ligation detection reaction.
Steroid 21-hydroxylase deficiency is the most common cause of congenital adrenal hyperplasia, an inherited inability to synthesize cortisol that occurs in 1 in 10,000-15,000 births. Affected females are born with ambiguous genitalia, a condition that can be ameliorated by administering dexamethasone to the mother for most of gestation. Prenatal diagnosis is required for accurate treatment of affected females as well as for genetic counseling purposes. Approximately 95% of mutations causing this disorder result from recombinations between the gene encoding the 21-hydroxylase enzyme (CYP21) and a linked, highly homologous pseudogene (CYP21P). Approximately 20% of these mutations are gene deletions, and the remainder are gene conversions that transfer any of nine deleterious mutations from the CYP21P pseudogene to CYP21. We describe a methodology for genetic diagnosis of 21-hydroxylase deficiency that utilizes gene-specific PCR amplification in conjunction with thermostable DNA ligase to discriminate single nucleotide variations in a multiplexed ligation detection assay. The assay has been designed to be used with either fluorescent or radioactive detection of ligation products by electrophoresis on denaturing acrylamide gels and is readily adaptable for use in other disease systems.


Influences of Dilute Organic Adsorbates on the Hydration of Low-Surface-Area Silicates.
Competitive adsorption of dilute quantities of certain organic molecules and water at silicate surfaces strongly influence the rates of silicate dissolution, hydration, and crystallization. Here, we determine the molecular-level structures, compositions, and site-specific interactions of adsorbed organic molecules at low absolute bulk concentrations on heterogeneous silicate particle surfaces at early stages of hydration. Specifically, dilute quantities (∼0.1% by weight of solids) of the disaccharide sucrose or industrially important phosphonic acid species slow dramatically the hydration of low-surface-area (∼1 m(2)/g) silicate particles. Here, the physicochemically distinct adsorption interactions of these organic species are established by using dynamic nuclear polarization (DNP) surface-enhanced solid-state NMR techniques. These measurements provide significantly improved signal sensitivity for near-surface species that is crucial for the detection and analysis of dilute adsorbed organic molecules and silicate species on low-surface-area particles, which until now have been infeasible to characterize. DNP-enhanced 2D (29)Si{(1)H}, (13)C{(1)H}, and (31)P{(1)H} heteronuclear correlation and 1D (29)Si{(13)C} rotational-echo double-resonance NMR measurements establish hydrogen-bond-mediated adsorption of sucrose at distinct nonhydrated and hydrated silicate surface sites and electrostatic interactions with surface Ca(2+) cations. By comparison, phosphonic acid molecules are found to adsorb electrostatically at or near cationic calcium surface sites to form Ca(2+)-phosphonate complexes. Although dilute quantities of both types of organic molecules effectively inhibit hydration, they do so by adsorbing in distinct ways that depend on their specific architectures and physicochemical interactions. The results demonstrate the feasibility of using DNP-enhanced NMR techniques to measure and assess dilute adsorbed molecules and their molecular interactions on low-surface-area materials, notably for compositions that are industrially relevant.


Noise-resistant fitting for spherical harmonics.
Spherical harmonic (SH) basis functions have been widely used for representing spherical functions in modeling various illumination properties. They can compactly represent low-frequency spherical functions. However, when the unconstrained least square method is used for estimating the SH coefficients of a hemispherical function, the magnitude of these SH coefficients could be very large. Hence, the rendering result is very sensitive to quantization noise (introduced by modern texture compression like S3TC, IEEE half float data type on GPU, or other lossy compression methods) in these SH coefficients. Our experiments show that, as the precision of SH coefficients is reduced, the rendered images may exhibit annoying visual artifacts. To reduce the noise sensitivity of the SH coefficients, this paper first discusses how the magnitude of SH coefficients affects the rendering result when there is quantization noise. Then, two fast fitting methods for estimating the noise-resistant SH coefficients are proposed. They can effectively control the magnitude of the estimated SH coefficients and, hence, suppress the rendering artifacts. Both statistical and visual results confirm our theory.


The spectrum of autosomal dominant polycystic kidney disease in children.
The natural history of autosomal dominant polycystic kidney disease (ADPKD) has not been well described in children, and it is not known whether a relationship exists between renal structural abnormalities and function in children as has been seen in adults. Therefore, 140 children from 67 ADPKD families were studied in a prospective study. Only 22 children came with a previous diagnosis of ADPKD. In 44% of all children, at least one cyst was found on ultrasound at a mean age of 8.7 yr. Of these, 60% were classified as having moderate disease on the basis of a total cyst number of 1 to 10 cysts, whereas 40% were considered to have severe disease with a total of more than 10 cysts. There was a significant relationship between the severity of the renal structural involvement and the frequency of flank and back pain, hypertension, and impaired renal concentrating capacity. However, GFR were not reduced in children with ADPKD and did not relate to structural severity. Thirty-nine children were seen for a follow-up visit 2 to 5 yr after the initial visit. No child had progressed from nonaffected to affected with ADPKD, but three of four children with only one cyst at the time of the initial study had progressed to bilateral cysts. Among the 22 ADPKD children who had a follow-up study, there was progression of the disease, reflected by an increase in cyst number and an increase in the frequency of pain and hypertension. However, GFR remained stable in all children.(ABSTRACT TRUNCATED AT 250 WORDS)


On the complexity of computing and learning with multiplicative neural networks.
In a great variety of neuron models, neural inputs are combined using the summing operation. We introduce the concept of multiplicative neural networks that contain units that multiply their inputs instead of summing them and thus allow inputs to interact nonlinearly. The class of multiplicative neural networks comprises such widely known and well-studied network types as higher-order networks and product unit networks. We investigate the complexity of computing and learning for multiplicative neural networks. In particular, we derive upper and lower bounds on the Vapnik-Chervonenkis (VC) dimension and the pseudo-dimension for various types of networks with multiplicative units. As the most general case, we consider feedforward networks consisting of product and sigmoidal units, showing that their pseudo-dimension is bounded from above by a polynomial with the same order of magnitude as the currently best-known bound for purely sigmoidal networks. Moreover, we show that this bound holds even when the unit type, product or sigmoidal, may be learned. Crucial for these results are calculations of solution set components bounds for new network classes. As to lower bounds, we construct product unit networks of fixed depth with super-linear VC dimension. For sigmoidal networks of higher order, we establish polynomial bounds that, in contrast to previous results, do not involve any restriction of the network order. We further consider various classes of higher-order units, also known as sigma-pi units, that are characterized by connectivity constraints. In terms of these, we derive some asymptotically tight bounds. Multiplication plays an important role in both neural modeling of biological behavior and computing and learning with artificial neural networks. We briefly survey research in biology and in applications where multiplication is considered an essential computational element. The results we present here provide new tools for assessing the impact of multiplication on the computational power and the learning capabilities of neural networks.


The acoustic signature of decaying resonant phospholipid microbubbles.
Sub-capillary sized microbubbles offer improved techniques for diagnosis and therapy of vascular related disease using ultrasound. Their physical interaction with ultrasound remains an active research field that aims to optimize techniques. The aim of this study is to investigate whether controlled microbubble disruption upon exposure to consecutive ultrasound exposures can be achieved. Single lipid-shelled microbubble scattered echoes have been measured in response to two consecutive imaging pulses, using a calibrated micro-acoustic system. The nonlinear evolution of microbubble echoes provides an exact signature above and below primary and secondary resonance, which has been identified using theoretical results based on the Mooney-Rivlin strain softening shell model. Decaying microbubbles follow an irreversible trajectory through the resonance peak, causing the evolution of specific microbubble spectral signatures. The characteristics of the microbubble motion causes varying amounts of shell material to be lost during microbubble decay. Incident ultrasound field parameters can thus accurately manipulate the regulated shedding of shell material, which has applications for both imaging applications and localized drug delivery strategies.


Unrestricted hepatocyte transduction with adeno-associated virus serotype 8 vectors in mice.
Recombinant adeno-associated virus (rAAV) vectors can mediate long-term stable transduction in various target tissues. However, with rAAV serotype 2 (rAAV2) vectors, liver transduction is confined to only a small portion of hepatocytes even after administration of extremely high vector doses. In order to investigate whether rAAV vectors of other serotypes exhibit similar restricted liver transduction, we performed a dose-response study by injecting mice with beta-galactosidase-expressing rAAV1 and rAAV8 vectors via the portal vein. The rAAV1 vector showed a blunted dose-response similar to that of rAAV2 at high doses, while the rAAV8 vector dose-response remained unchanged at any dose and ultimately could transduce all the hepatocytes at a dose of 7.2 x 10(12) vector genomes/mouse without toxicity. This indicates that all hepatocytes have the ability to process incoming single-stranded vector genomes into duplex DNA. A single tail vein injection of the rAAV8 vector was as efficient as portal vein injection at any dose. In addition, intravascular administration of the rAAV8 vector at a high dose transduced all the skeletal muscles throughout the body, including the diaphragm, the entire cardiac muscle, and substantial numbers of cells in the pancreas, smooth muscles, and brain. Thus, rAAV8 is a robust vector for gene transfer to the liver and provides a promising research tool for delivering genes to various target organs. In addition, the rAAV8 vector may offer a potential therapeutic agent for various diseases affecting nonhepatic tissues, but great caution is required for vector spillover and tight control of tissue-specific gene expression.


Effects of naltrexone plus topiramate on ethanol self-administration and tyrosine hydroxylase gene expression changes.
The purpose of this study was to evaluate the effects of naltrexone (0.7 mg/kg) and/or topiramate (25 mg/kg) on ethanol consumption and the motivation to drink in an oral-operant conditioning paradigm in C57BL/6 mice. Subsequent real-time polymerase chain reaction (PCR) experiments were performed to analyze gene expression changes in tyrosine hydroxylase (TH) in the ventral tegmental area (VTA). The administration of naltrexone significantly reduced ethanol consumption and the motivation to drink during the different stages of the experiment, whereas the treatment with topiramate resulted in a much lower effect. Interestingly, the administration of naltrexone plus topiramate reduced ethanol consumption markedly compared with single-drug treatment. The water self-administration paradigm was also performed using the same drugs and no differences were found between treatment groups. Real-time PCR analyses revealed that naltrexone significantly normalized the increase of TH gene expression in the VTA induced by ethanol, whereas the administration of topiramate did not produce any significant effect. In the ethanol self-administration procedure, the combination of both drugs further reduced TH gene expression, reaching statistical significance compared with the vehicle, naltrexone or topiramate groups. Taken together, these findings indicate that the administration of naltrexone plus topiramate further reduced ethanol consumption and the motivation to drink in comparison with single-drug treatment. This action may be due, at least in part, to a greater decrease in TH gene expression in the VTA. These results suggest that the combination of both drugs deserves further exploration for the treatment of problems related to alcohol consumption.


Traumatic subluxation/dislocation of the peroneal tendons.
Traumatic subluxation/dislocation of the peroneal tendons has been reported following a variety of sports-related activities. The peroneal musculature contracts reflexively during the injury and overcomes the restraining soft tissue. The tendons can then dislocate anteriorly from behind the distal fibula. Some patients have anatomical variations of the posterolateral ankle that predispose them to injury. If the physician is unaware of the injury, the diagnosis may be missed in the acute setting and can develop into a chronic, disabling condition. Treatment of the acute injury is controversial because advocates exist for both conservative and surgical therapies. The chronic, painful lesion should be treated operatively. A tremendous number of surgical procedures have been described for the treatment of chronic tendon dislocations. Choice of a surgical procedure depends upon the anatomy of the peroneal groove and the retinaculum, and the nature of the damage to the area.


Maternal stress-induced reduction in birth weight as a marker for adult affective state.
It is known that adverse events experienced by a pregnant woman may be reflected upon the developing fetus and adversely affect its mental wellbeing in later life. In a recent study by our group, prenatal stress was associated with a clear increase in anxiety- and depression-related behavior in male, but not female Sprague-Dawley offspring. Since birth weight data were recorded we were able to determine whether birth weight, as an important outcome measure of fetal distress, may be used as a predictive indicator for adult performance. For this purpose, a correlation analysis was performed, aimed at studying the possible link between stress-induced fetal growth restriction and adult affective state. Male birth weight correlated positively to depression-related behavior in the forced swim test. Furthermore, it weight was correlated negatively to basal, and positively to stress-induced, plasma corticosterone levels in adulthood. Female birth weight did not correlate to any of the studied outcome measures. These data suggest that male birth weight may represent a valuable indicative marker for variations in adult affective state with a developmental origin.


Aggressive and impulsive behaviors in Frontotemporal dementia and Alzheimer's disease.
Differentiating FTD and AD is of great clinical significance, due to the very different efficacy of cholinesterase inhibitors in the two disorders. Previous studies have pointed to behavioral differences in FTD and AD, but less attention has been paid to comparing aggressive and impulsive behaviors in the AD and the FTD. Nursing home residents diagnosed with AD (NINCDS/ADRDA criteria) or the behavioral variant of FTD (Frontotemporal Dementia and Pick's Disease Working Group criteria) were included in the research group. Individuals diagnosed with AD who showed atrophy of the frontal lobes in neuroimaging were excluded from the study, as were patients matching the clinical criteria for bvFTD, but with lesions in the temporal lobes. Agitation and aggressive behaviors were assessed using the Cohen-Mansfield Agitation Inventory (CMAI), whereas behavioral and psychological symptoms were assessed with the Neuropsychiatric Inventory - Nursing Home Version (NPI-NH). The overall NPI-NH score, including with some partial scores (apathy/indifference and disinhibition), together with the intensity of aberrant motor behavior differentiated individuals with AD from those with bvFTD. On the CMAI scale, both the overall score and physical aggressive behaviors differentiated the two study groups. Our research confirms previous results indicating a greater intensity of behavioral pathology in FTD. A detailed assessment of aggressive behaviors revealed further differences between AD and FTD in this respect.


Impact of diabetes on healthcare costs in a population-based cohort: a cost analysis.
To estimate the healthcare costs attributable to diabetes in Ontario, Canada using a propensity-matched control design and health administrative data from the perspective of a single-payer healthcare system. Incident diabetes cases among adults in Ontario were identified from the Ontario Diabetes Database between 2004 and 2012 and matched 1:3 to control subjects without diabetes identified in health administrative databases on the basis of sociodemographics and propensity score. Using a comprehensive source of administrative databases, direct per-person costs (Canadian dollars 2012) were calculated. A cost analysis was performed to calculate the attributable costs of diabetes; i.e. the difference of costs between patients with diabetes and control subjects without diabetes. The study sample included 699 042 incident diabetes cases. The costs attributable to diabetes were greatest in the year after diagnosis [C$3,785 (95% CI 3708, 3862) per person for women and C$3,826 (95% CI 3751, 3901) for men], increasing substantially for older age groups and patients who died during follow-up. After accounting for baseline comorbidities, attributable costs were primarily incurred through inpatient acute hospitalizations, physician visits and prescription medications and assistive devices. The excess healthcare costs attributable to diabetes are substantial and pose a significant clinical and public health challenge. This burden is an important consideration for decision-makers, particularly given increasing concern over the sustainability of the healthcare system, aging population structure and increasing prevalence of diabetic risk factors, such as obesity.


Further refinement of COL4A1 and COL4A2 related cortical malformations.
Mutations in COL4A1 have been reported in schizencephaly and porencephaly combined with microbleeds or calcifications, often associated with ocular and renal abnormalities, myopathy, elevated creatine kinase levels and haemolytic anaemia. In this study, we aimed to clarify the phenotypic spectrum of COL4A1/A2 mutations in the context of cortical malformations that include schizencephaly, polymicrogyria and/or heterotopia. We screened for COL4A1/A2 mutations in 9 patients with schizencephaly and/or polymicrogyria suspected to be caused by vascular disruption and leading to a cerebral haemorrhagic ischaemic event. These included 6 cases with asymmetrical or unilateral schizencephaly and/or polymicrogyria and 3 cases with bilateral schizencephaly. One de novo missense COL4A1 mutation (c.3715 G > A, p.(Gly1239Arg)) and two COL4A2 mutations were found, respectively in one familial case (c.4129G > A, p.(Gly1377Arg)) and one sporadic patient (c.1776+1G > A). In three other cases, COL4A1 variants of unknown significance were identified. None of our patients demonstrated neuromuscular or hematological anomalies. Brain malformations included a combination of schizencephaly, mainly asymmetrical, with porencephaly or ventriculomegaly (3/3 mutated patients). We did not observe microbleeds or microcalcifications in any of our cases, hence we do not believe that they represent a distinctive feature of COL4A1/A2 mutations. Our study further emphasizes the need to search for both COL4A1 and COL4A2 mutations in children presenting with uni- or bilateral polymicrogyria with schizencephaly, even in the absence of intracranial microbleeds, calcification or associated systemic features.


Long-term outcomes of living donor liver transplantation in patients with a prior history of nonhepatic malignancy.
Posttransplant malignancy has become a significant cause of mortality. Data on the long-term outcomes of patients with pretransplant nonhepatic malignancy (PTM) after living donor liver transplantation (LDLT) are scarce, although the recipients of other organs with PTM have been reported to have a poor survival. Fifteen patients with PTM (4.4%) among the 342 adult recipients were identified in our LDLT programs. The outcomes of the patients with PTM after LDLT were compared to those of patients without PTM in terms of the all-cause mortality and cancer-specific mortality (defined as mortality related to malignancy expect for hepatocellular carcinoma, cholangiocarcinoma, or neuroendocrine tumor). The sites of PTM included the breast in six, stomach in two, and colon, lung, kidney, uterine, thyroid, larynx, and acute myelogenous leukemia in one each. The median interval from the PTM treatment to LDLT was 57 months (range, 2-298). The patients who received the curative treatment for PTM were selected as the recipients. No patients with PTM had recurrence during the follow-up period. The 1-, 5-, and 10-year patient survival rates were 100%, 92.9%, and 92.9% in the PTM group and 86.2%, 76.7%, and 68.5% in the non-PTM group, respectively (p = 0.142). Likewise, there was no significant difference between the two groups in the cancer-specific mortality. In conclusion, the patients with PTM had comparable outcomes with regard to mortality and cancer-specific mortality compared with those without PTM. This study showed that the patients with PTM can obtain an acceptable outcome after LDLT when carefully selected.


Unspecific metabolic blood parameters as used in clinical routine may differentiate malignant from benign cerebral tumors.
The investigation of rather insensitive metabolic parameters (protein, fibrinogen, blood urea nitrogen (BUN), blood glucose) reveals significant differences between tumor-bearing and tumor-free patients as well as benign and malignant neoplasms. Whereas metastases and glioblastomas (GBM) show significantly elevated BUN levels (21.9 +/- 1.7; 8 +/- 2.2 mg/dl) compared to benign tumors (meningioma WHO I, astrocytoma I, II) (16 +/- 0.9 mg/dl) and tumor-free matched controls (e.g. 13.9 +/- 1.4 mg/dl) only metastases depict higher glucose (141.7 +/- 11mg/dl) counts. Fibrinogen, significantly elevated in malignancy (395 +/- 25.2; 397.2 +/- 25.9 mg/dl) is without difference between meningioma, astrocytoma (253.2 +/- 16.6; 271.5 +/- 16.5 mg/dl) and controls (e.g. 270.1 +/- 10.8 mg/dl). Correlating BUN with total protein reveals a metabolic mismatch to nearly all tumor patients, regardless of dignity, as compared to tumor-free patients. Neuroendocrinoimmunological changes are the most likely reason for these overt as well as occult findings, making investigation of more sensitive metabolic parameters a rewarding task.


Nonspecific abdominal pain: A follow-up survey.
Little data have been published on the outcome of patients discharged from the emergency department (ED) after being diagnosed with nonspecific abdominal pain (NSAP). This study aimed to investigate short-term follow-up of patients discharged with a diagnosis of NSAP from the ED. This prospective, observational study was conducted in the University-based ED and enrolled all consecutive adult patients who were diagnosed as NSAP out of patients presented with abdominal pain (AP). The main outcome measure was the presence of recurrent AP resulting in referral to the ED and specific diagnoses within the first 3- and 90-day postdischarge. On the 3rd and 90th days, all patients discharged with NSAP from the ED were asked questions, and their response entered into a questionnaire. A total of 684 patients presented with AP, of which 299 (46%) had a diagnosis of NSAP within the 4-month period. Fifty cases (16%) could not be included due to inability to access. Eighty-one out of 249 patients (32.5%) complained of recurrent AP within the first 3 days. Twenty-two cases (8.8%) were readmitted to ED once again in the meantime, and ten received specific diagnoses including three with acute abdomen. Within 90 days, additional nine patients out of 20 (45%) with recurrent AP received specific diagnoses including two with acute abdomen. Certain specific underlying entities can be missed in patients considered to have NSAP and discharged from the ED. Adherence to timely follow-up and repeated examinations are of vital importance in these patients.


Effects of Rituximab and Infliximab Treatment on Carboxypeptidase B and Its Substrates in RA Synovium.
We evaluated the synovial effects of 2 potent biologic rheumatoid arthritis (RA) therapies, focusing on their effect on the expression level of carboxypeptidase B (CPB) and its substrates. Patients with RA receiving infliximab (IFX; n = 9) or rituximab (RTX; n = 5) had an arthroscopic synovial biopsy at baseline and 16 weeks posttherapy. Expression of CPB, C5a, osteopontin (OPN), CD3, CD20, CD55, and CD68 was assessed by immunohistochemistry and image analysis, and compared with OA synovium. RA disease activity score was assessed at multiple timepoints. Serial serum samples were analyzed for soluble CPB and C5a levels. The baseline clinical characteristics of patients receiving IFX and RTX were similar. At the time of the second biopsy, 50% of patients had achieved a European League Against Rheumatism good or moderate response. At baseline, expression of CPB, C5a, and OPN was markedly higher in RA compared with OA synovium and correlated with mononuclear cell infiltration. There was an overall reduction in synovial expression of CPB, C5a, and OPN paralleling a reduction in mononuclear cell infiltration, but these changes were not associated with clinical response. After an early reduction in serum C5a levels, these returned to baseline levels at later timepoints. In response to IFX and RTX treatment, RA synovial expression of CPB, C5a, and OPN decrease independently of the clinical response, reflecting the complex proinflammatory and antiinflammatory effects of this pathway.


Multiscale 3D-printing of microfluidic AFM cantilevers.
Microfluidic atomic force microscopy (AFM) cantilever probes have all the functionalities of a standard AFM cantilever along with fluid pipetting. They have a channel inside the cantilever and an aperture at the tip. Such probes are useful for precise fluid manipulation at a desired location, for example near or inside cells. They are typically made by complex microfabrication process steps, resulting in expensive probes. Here, we used two different 3D additive manufacturing techniques, stereolithography and two-photon polymerization, to directly print ready-to-use microfluidic AFM cantilever probes. This approach has considerably reduced the fabrication time and increased the design freedom. One of the probes, 564 μm long, 30 μm wide, 30 μm high, with a 25 μm diameter channel and 2.5 μm wall thickness had a spring constant of 3.7 N m-1 and the polymer fabrication material had an elastic modulus of 4.2 GPa. Using these 3D printed probes, AFM imaging of a surface, puncturing of the cell membrane, and aspiration at the single cell level have been demonstrated.


Intravenous Topiramate: Pharmacokinetics in Dogs with Naturally Occurring Epilepsy.
Barriers to developing treatments for human status epilepticus include the inadequacy of experimental animal models. In contrast, naturally occurring canine epilepsy is similar to the human condition and can serve as a platform to translate research from rodents to humans. The objectives of this study were to characterize the pharmacokinetics of an intravenous (IV) dose of topiramate (TPM) in dogs with epilepsy and evaluate its effect on intracranial electroencephalographic (iEEG) features. Five dogs with naturally occurring epilepsy were used for this study. Three were getting at least one antiseizure drug as maintenance therapy including phenobarbital (PB). Four (ID 1-4) were used for the 10 mg/kg IV TPM + PO TPM study, and three (ID 3-5) were used for the 20 mg/kg IV TPM study. IV TPM was infused over 5 min at both doses. The animals were observed for vomiting, diarrhea, ataxia, and lethargy. Blood samples were collected at scheduled pre- and post-dose times. Plasma concentrations were measured using a validated high-performance liquid chromatography-mass spectrometry method. Non-compartmental and population compartmental modeling were performed (Phoenix WinNonLin and NLME) using plasma concentrations from all dogs in the study. iEEG was acquired in one dog. The difference between averaged iEEG energy levels at 15 min pre- and post-dose was assessed using a Kruskal-Wallis test. No adverse events were noted. TPM concentration-time profiles were best fit by a two compartment model. PB co-administration was associated with a 5.6-fold greater clearance and a ~4-fold shorter elimination half-life. iEEG data showed that TPM produced a significant energy increase at frequencies >4 Hz across all 16 electrodes within 15 min of dosing. Simulations suggested that dogs on an enzyme inducer would require 25 mg/kg, while dogs on non-inducing drugs would need 20 mg/kg to attain the target concentration (20-30 μg/mL) at 30 min post-dose. This study shows that IV TPM has a relatively rapid onset of action, loading doses appear safe, and the presence of PB necessitates a higher dose to attain targeted concentrations. Consequently, it is a good candidate for further evaluation for treatment of seizure emergencies in dogs and people.


Low temperature flow lithography.
Flow lithography (FL) is a microfluidic technique distinguished for its ability to produce hydrogel microparticles of various geometrical and chemical designs. While FL is typically performed in room temperature, this paper reports a new technique called low temperature flow lithography that uses low synthesis temperature to increase the degree of polymerization of microparticles without compromising other aspects of flow lithography. We suggest that decreased oxygen diffusivity in low temperature is responsible for the increase in polymerization. Microparticles that exhibit a higher degree of polymerization display a more developed polymer network, ultimately resulting in a more defined morphology, higher incorporation of materials of interest, and improved functional performance. This work demonstrates the increase in the degree of polymerization by examining the temperature effect on both the physical and chemical structures of particles. We show applications of this technique in synthesizing thin microparticles and enhancing microparticle-based detection of microRNA. Low temperature FL offers a simple and easy method of improving the degree of polymerization, which can be implemented in a wide range of FL applications.


Neuropilin 1 deficiency on CD4+Foxp3+ regulatory T cells impairs mouse melanoma growth.
Infiltration of Foxp3(+) regulatory T (T reg) cells is considered to be a critical step during tumor development and progression. T reg cells supposedly suppress locally an effective anti-tumor immune response within tumor tissues, although the precise mechanism by which T reg cells infiltrate the tumor is still unclear. We provide evidence that Neuropilin 1 (Nrp-1), highly expressed by Foxp3(+) T reg cells, regulates the immunological anti-tumor control by guiding T reg cells into the tumor in response to tumor-derived vascular endothelial growth factor (VEGF). We demonstrate for the first time that T cell-specific ablation of Nrp-1 expression results in a significant breakdown in tumor immune escape in various transplantation models and in a spontaneous, endogenously driven melanoma model associated with strongly reduced tumor growth and prolonged tumor-free survival. Strikingly, numbers of tumor-infiltrating Foxp3(+) T reg cells were significantly reduced accompanied by enhanced activation of CD8(+) T cells within tumors of T cell-specific Nrp-1-deficient mice. This phenotype can be reversed by adoptive transfer of Nrp-1(+) T reg cells from wild-type mice. Thus, our data strongly suggest that Nrp-1 acts as a key mediator of Foxp3(+) T reg cell infiltration into the tumor site resulting in a dampened anti-tumor immune response and enhanced tumor progression.


Evolution of chromosome organization driven by selection for reduced gene expression noise.
The distribution of genes on eukaryotic chromosomes is nonrandom, but the reasons behind this are not well understood. The commonly observed clustering of essential genes is a case in point. Here we model and test a new hypothesis. Essential proteins are unusual in that random fluctuations in abundance (noise) can be highly deleterious. We hypothesize that persistently open chromatin domains are sinks for essential genes, as they enable reduced noise by avoidance of transcriptional bursting associated with chromatin remodeling. Simulation of the model captures clustering and correctly predicts that (i) essential gene clusters are associated with low nucleosome occupancy (ii) noise-sensitive nonessential genes cluster with essential genes (iii) nonessential genes of similar knockout fitness are physically linked (iv) genes in domains rich in essential genes have low noise (v) essential genes are rare subtelomerically and (vi) essential gene clusters are preferentially conserved. We conclude that different noise characteristics of different genomic domains favors nonrandom gene positioning. This has implications for gene therapy and understanding transgenic phenotypes.


Elevational gradients in fish diversity in the Himalaya: water discharge is the key driver of distribution patterns.
Studying diversity and distribution patterns of species along elevational gradients and understanding drivers behind these patterns is central to macroecology and conservation biology. A number of studies on biogeographic gradients are available for terrestrial ecosystems, but freshwater ecosystems remain largely neglected. In particular, we know very little about the species richness gradients and their drivers in the Himalaya, a global biodiversity hotspot. We collated taxonomic and distribution data of fish species from 16 freshwater Himalayan rivers and carried out empirical studies on environmental drivers and fish diversity and distribution in the Teesta river (Eastern Himalaya). We examined patterns of fish species richness along the Himalayan elevational gradients (50-3800 m) and sought to understand the drivers behind the emerging patterns. We used generalized linear models (GLM) and generalized additive models (GAM) to examine the richness patterns; GLM was used to investigate relationship between fish species richness and various environmental variables. Regression modelling involved stepwise procedures, including elimination of collinear variables, best model selection, based on the least Akaike's information criterion (AIC) and the highest percentage of deviance explained (D(2)). This maiden study on the Himalayan fishes revealed that total and non-endemic fish species richness monotonously decrease with increasing elevation, while endemics peaked around mid elevations (700-1500 m). The best explanatory model (synthetic model) indicated that water discharge is the best predictor of fish species richness patterns in the Himalayan rivers. This study, carried out along one of the longest bioclimatic elevation gradients of the world, lends support to Rapoport's elevational rule as opposed to mid domain effect hypothesis. We propose a species-discharge model and contradict species-area model in predicting fish species richness. We suggest that drivers of richness gradients in terrestrial and aquatic ecosystems are likely to be different. These studies are crucial in context of the impacts of unprecedented on-going river regulation on fish diversity and distribution in the Himalaya.


The history of the Coast Salish "woolly dogs".
DNA and traditional knowledge reveal the history of an extinct dog bred for its wool.