Long-term safety and efficacy of factor IX gene therapy in hemophilia B.
In patients with severe hemophilia B, gene therapy that is mediated by a novel self-complementary adeno-associated virus serotype 8 (AAV8) vector has been shown to raise factor IX levels for periods of up to 16 months. We wanted to determine the durability of transgene expression, the vector dose-response relationship, and the level of persistent or late toxicity. We evaluated the stability of transgene expression and long-term safety in 10 patients with severe hemophilia B: 6 patients who had been enrolled in an initial phase 1 dose-escalation trial, with 2 patients each receiving a low, intermediate, or high dose, and 4 additional patients who received the high dose (2×10(12) vector genomes per kilogram of body weight). The patients subsequently underwent extensive clinical and laboratory monitoring. A single intravenous infusion of vector in all 10 patients with severe hemophilia B resulted in a dose-dependent increase in circulating factor IX to a level that was 1 to 6% of the normal value over a median period of 3.2 years, with observation ongoing. In the high-dose group, a consistent increase in the factor IX level to a mean (±SD) of 5.1±1.7% was observed in all 6 patients, which resulted in a reduction of more than 90% in both bleeding episodes and the use of prophylactic factor IX concentrate. A transient increase in the mean alanine aminotransferase level to 86 IU per liter (range, 36 to 202) occurred between week 7 and week 10 in 4 of the 6 patients in the high-dose group but resolved over a median of 5 days (range, 2 to 35) after prednisolone treatment. In 10 patients with severe hemophilia B, the infusion of a single dose of AAV8 vector resulted in long-term therapeutic factor IX expression associated with clinical improvement. With a follow-up period of up to 3 years, no late toxic effects from the therapy were reported. (Funded by the National Heart, Lung, and Blood Institute and others; ClinicalTrials.gov number, NCT00979238.).


Clonazepam treatment of lysergic acid diethylamide-induced hallucinogen persisting perception disorder with anxiety features.
An unique and intriguing characteristic of lysergic acid diethylamide (LSD) and LSD-like substances is the recurrence of some of the symptoms which appear during the intoxication, in the absence of recent intake of hallucinogens. Hallucinogen persisting perception disorder (HPPD) is a condition in which the re-experiencing of one or more perceptual symptoms causes significant distress or impairment in social, occupational or other important areas of functioning and may be extremely debilitating. Benzodiazepines are one of the recommended agents for the treatment of HPPD but it is unclear which of them may be more helpful. The goal of our investigation was to assess the efficacy of clonazepam in the treatment of LSD-induced HPPD. Sixteen patients fulfilled entrance criteria. All complained of HPPD with anxiety features for at least 3 months and were drug free at least 3 months. They received clonazepam 2 mg/day for 2 months. Follow-up was continued for 6 months. They were weekly evaluated during the 2 months of clonazepam administration and monthly during the follow-up period using the Clinical Global Impression Scale, a Self-report Scale and Hamilton Anxiety Rating Scale. Patients reported a significant relief and the presence of only mild symptomatology during the clonazepam administration. This improvement was clearly sustained and persisted during a 6-month follow-up period. This study suggests that high potency benzodiazepines like clonazepam, which has serotonergic properties, may be more effective than low-potency benzodiazepines in the treatment of some patients with LSD-induced HPPD.


Evidence for biphasic uncoating during HIV-1 infection from a novel imaging assay.
Uncoating of the HIV-1 core plays a critical role during early post-fusion stages of infection but is poorly understood. Microscopy-based assays are unable to easily distinguish between intact and partially uncoated viral cores. In this study, we used 5-ethynyl uridine (EU) to label viral-associated RNA during HIV production. At early time points after infection with EU-labeled virions, the viral-associated RNA was stained with an EU-specific dye and was detected by confocal microscopy together with viral proteins. We observed that detection of the viral-associated RNA was specific for EU-labeled virions, was detected only after viral fusion with target cells, and occurred after an initial opening of the core. In vitro staining of cores showed that the opening of the core allowed the small molecule dye, but not RNase A or antibodies, inside. Also, staining of the viral-associated RNA, which is co-localized with nucleocapsid, decays over time after viral infection. The decay rate of RNA staining is dependent on capsid (CA) stability, which was altered by CA mutations or a small molecule inducer of HIV-1 uncoating. While the staining of EU-labeled RNA was not affected by inhibition of reverse transcription, the kinetics of core opening of different CA mutants correlated with initiation of reverse transcription. Analysis of the E45A CA mutant suggests that initial core opening is independent of complete capsid disassembly. Taken together, our results establish a novel RNA accessibility-based assay that detects an early event in HIV-1 uncoating and can be used to further define this process.


Target for improvement: a cluster randomised trial of public involvement in quality-indicator prioritisation (intervention development and study protocol).
Public priorities for improvement often differ from those of clinicians and managers. Public involvement has been proposed as a way to bridge the gap between professional and public clinical care priorities but has not been studied in the context of quality-indicator choice. Our objective is to assess the feasibility and impact of public involvement on quality-indicator choice and agreement with public priorities. We will conduct a cluster randomised controlled trial comparing quality-indicator prioritisation with and without public involvement. In preparation for the trial, we developed a 'menu' of quality indicators, based on a systematic review of existing validated indicator sets. Participants (public representatives, clinicians, and managers) will be recruited from six participating sites. In intervention sites, public representatives will be involved through direct participation (public representatives, clinicians, and managers will deliberate together to agree on quality-indicator choice and use) and consultation (individual public recommendations for improvement will be collected and presented to decision makers). In control sites, only clinicians and managers will take part in the prioritisation process. Data on quality-indicator choice and intended use will be collected. Our primary outcome will compare quality-indicator choice and agreement with public priorities between intervention and control groups. A process evaluation based on direct observation, videorecording, and participants' assessment will be conducted to help explain the study's results. The marginal cost of public involvement will also be assessed. We identified 801 quality indicators that met our inclusion criteria. An expert panel agreed on a final set of 37 items containing validated quality indicators relevant for chronic disease prevention and management in primary care. We pilot tested our public-involvement intervention with 27 participants (11 public representatives and 16 clinicians and managers) and our study instruments with an additional 21 participants, which demonstrated the feasibility of the intervention and generated important insights and adaptations to engage public representatives more effectively. To our knowledge, this study is the first trial of public involvement in quality-indicator prioritisation, and its results could foster more effective upstream engagement of patients and the public in clinical practice improvement. NTR2496 (Netherlands National Trial Register, http://www.trialregister.nl).


Design of composite measure schemes for comparative severity assessment in animal-based neuroscience research: A case study focussed on rat epilepsy models.
Comparative severity assessment of animal models and experimental interventions is of utmost relevance for harm-benefit analysis during ethical evaluation, an animal welfare-based model prioritization as well as the validation of refinement measures. Unfortunately, there is a lack of evidence-based approaches to grade an animal's burden in a sensitive, robust, precise, and objective manner. Particular challenges need to be considered in the context of animal-based neuroscientific research because models of neurological disorders can be characterized by relevant changes in the affective state of an animal. Here, we report about an approach for parameter selection and development of a composite measure scheme designed for precise analysis of the distress of animals in a specific model category. Data sets from the analysis of several behavioral and biochemical parameters in three different epilepsy models were subjected to a principal component analysis to select the most informative parameters. The top-ranking parameters included burrowing, open field locomotion, social interaction, and saccharin preference. These were combined to create a composite measure scheme (CMS). CMS data were subjected to cluster analysis enabling the allocation of severity levels to individual animals. The results provided information for a direct comparison between models indicating a comparable severity of the electrical and chemical post-status epilepticus models, and a lower severity of the kindling model. The new CMS can be directly applied for comparison of other rat models with seizure activity or for assessment of novel refinement approaches in the respective research field. The respective online tool for direct application of the CMS or for creating a new CMS based on other parameters from different models is available at https://github.com/mytalbot/cms. However, the robustness and generalizability needs to be further assessed in future studies. More importantly, our concept of parameter selection can serve as a practice example providing the basis for comparable approaches applicable to the development and validation of CMS for all kinds of disease models or interventions.


An Epidemiological Study of Child Marriages in a Rural Community of Gujarat.
India has the maximum number of child marriages (CMs; < 18 years) because of the size of its population, and in 47% of all marriages the bride is a child. Children who are married at young age are exposed to multiple risks pertaining to their physical, mental, and social health. (i) To estimate the prevalence of CM in rural population. (ii) To study the determinants and health effects of CM. (iii) To assess the awareness among the married women regarding the health implications of CM. Community-based cross-sectional study conducted in Ardi village of Anand district. All the married women of the village were surveyed to find out the prevalence of CM. For collection of other relevant information, only those women having a married life of less than 10years were interviewed using semicoded and pretested questionnaire. Data collected were analyzed using Statistical Package for Social Sciences (SPSS) 17.0 software. Proportions, ratios, χ(2) test, and Fisher's exact test. The prevalence of CM was found to be 71.5%. Caste and spouse's education were revealed as important determinants for CM. CM was found to be significantly associated with mother's age at birth of first child, delayed antenatal care (ANC), spontaneous abortion, preterm delivery, low birth weight (LBW), health problems in new born baby, faulty feeding practices, lack of knowledge regarding family welfare methods, and health implications of CM. Exceptionally high prevalence of CM in rural community and its serious health consequences warrant stricter enforcement of legislation, better educational opportunities for girls, and easy access to quality health services.


Direct Observation of Incoherent Cherenkov Diffraction Radiation in the Visible Range.
We report on the observation of incoherent Cherenkov radiation emitted by a 5.3 GeV positron beam circulating in the Cornell electron-positron storage ring as the beam passes in the close vicinity of the surface of a fused silica radiator (i.e., at a distance larger than 0.8 mm). The shape of the radiator was designed in order to send the Cherenkov photons towards the detector, consisting of a compact optical system equipped with an intensified camera. The optical system allows both the measurements of 2D images and angular distribution including polarization study. The corresponding light intensity has been measured as a function of the distance between the beam and the surface of the radiator and has shown a good agreement with theoretical predictions. For highly relativistic particles, a large amount of incoherent radiation is produced in a wide spectral range. A light yield of 0.8×10^{-3} photon per particle per turn has been measured at a wavelength of 600±10 nm in a 2 cm long radiator and for an impact parameter of 1 mm. This will find applications in accelerators as noninvasive beam diagnostics for both leptons and hadrons.


Conservative amino acid substitution in the myelin proteolipid protein of jimpymsd mice.
The capacity for synthesizing and maintaining a compact myelin sheath is destroyed in a number of inborn errors of myelin metabolism. One class of hypomyelinating mutations, which displays an X-linked pattern of inheritance, is distinguished by marked disturbances in oligodendrocyte differentiation. We have defined the molecular defect in one such mutant that lacks mature oligodendrocytes, the X-linked jimpy myelin synthesis deficient (jpmsd) trait in mice. The structure of the gene encoding the most abundant myelin protein, proteolipid protein (PLP), was determined by mapping and partially sequencing genomic clones from jpmsd and wild-type mice. Jpmsd mice have a single base change in PLP, a C----T transition in exon 6 that would substitute a valine for alanine in both PLP and its alternatively spliced isoform, DM20. The mutation was confirmed by polymerase chain reaction-amplifying exon 6 from genomic DNA and then either sequencing the amplified DNA or directly probing exon 6 with oligonucleotides designed to detect a single base mismatch. The conservative amino acid replacement in PLP/DM20 of jpmsd mice results in a pleiotropic phenotype similar to that observed for the allelic mutation jimpy, in which a splicing defect has radically altered the PLP/DM20 protein. The accelerated turnover of oligodendrocytes in both mouse mutants suggests a function for PLP/DM20 in oligodendrocyte differentiation distinct from the role of these proteolipid proteins as structural components of the myelin sheath.


The satiating hormone amylin enhances neurogenesis in the area postrema of adult rats.
Adult neurogenesis in the subgranular zone and subventricular zone is generally accepted, but its existence in other brain areas is still controversial. Circumventricular organs, such as the area postrema (AP) have recently been described as potential neurogenic niches in the adult brain. The AP is the major site of action of the satiating hormone amylin. Amylin has been shown to promote the formation of neuronal projections originating from the AP in neonatal rodents but the role of amylin in adult neurogenesis remains unknown. To test this, we first performed an RNA-sequencing of the AP of adult rats acutely injected with either amylin (20 μg/kg), amylin plus the amylin receptor antagonist AC187 (500 μg/kg) or vehicle. Second, animals were subcutaneously equipped with minipumps releasing either amylin (50 μg/kg/day) or vehicle for 3 weeks to assess cell proliferation and differentiation with the 5'-bromo-2-deoxyuridine (BrdU) technique. Acute amylin injections affected genes involved in pathways and processes that control adult neurogenesis. Amylin consistently upregulated NeuroD1 transcript and protein in the adult AP, and this effect was blocked by the co-administration of AC187. Further, chronic amylin treatment increased the number of newly proliferated AP-cells and significantly promoted their differentiation into neurons rather than astrocytes. Our findings revealed a novel role of the satiating hormone amylin in promoting neurogenesis in the AP of adult rats.


Cellular and molecular regulation of skeletal muscle side population cells.
Muscle progenitor cells (satellite cells) function in the maintenance and repair of adult skeletal muscle. Side population (SP) cells are enriched in repopulating activity and also reside in adult skeletal muscle. In this study, we observed that Abcg2 is a determinant of the SP cell phenotype. Using reverse transcription polymerase chain reaction and immunohistochemical techniques, we localized Abcg2-expressing cells in the interstitium and in close approximation to the vasculature of adult skeletal muscle. Muscle SP cells are able to differentiate into myotubes and increase in number after cardiotoxin-induced muscle injury. Similar to myogenic progenitor cells, muscle SP cells express Foxk1 and are decreased in number in Foxk1 mutant skeletal muscle. Using emerging technologies, we examine the molecular signature of muscle SP cells from normal, injured, and Foxk1 mutant skeletal muscle to define common and distinct molecular programs. We propose that muscle SP cells are progenitor cells that participate in repair and regeneration of adult skeletal muscle.


Presurgical Nasoalveolar Molding of Bilateral Cleft Lip and Palate Infants: An Orthodontist's Point of View.
Nonsyndromic complete cleft lip and palate deformity is primarily functional, then esthetic, and finally but not least importantly, a dental challenge. Feeding and facial appearance are important during the first years of a newborn. Nutrition is universally provided by passive feeding plates. If the Cleft Team prefers to use active plates, alveolar molding combined with nasal approaches in infants is the best method to improve esthetics to date. Orthodontists are predominantly responsible for achieving both the goals. After those difficulties have been met in early days of the life, dentists are mainly responsible for the treatment thereafter. If the infants have a dentoalveolar unity without any fistulas and correctly aligned maxillary deciduous teeth, this is a real success. Therefore, this article is an overview of presurgical infant orthopedics and its contribution to subsequent dental practice.


Third-Party Testing Nutritional Supplement Knowledge, Attitudes, and Use Among an NCAA I Collegiate Student-Athlete Population.
Dietary supplements, sports foods, and ergogenic supplements are consumed to increase performance, recovery, and health, but risk contamination with illegal substances. Third-party testing programs may assist in regulating the purity and safety of supplements, yet athletes' attitudes and use of such programs are not widely reported. This study examined nutritional supplement knowledge, attitudes, and use, as well as the purchase of third-party tested supplements among university student-athletes (N = 138). Knowledge of nutritional supplements yielded a median and (IQR) score of 25% (17 to 42%). Sixteen percent of student-athletes said they were knowledgeable about supplements and their effects, p < 0.001. All athletes stated they used a dietary supplement or sports food at least once within the last 12 months, and 77% consumed at least one "claimed to be" ergogenic supplement. Sixty-six percent of student-athletes purchased nutritional supplements not provided by the athletic department. Females athletes were more likely to consume a combination of vitamins and single minerals, a larger variety of sports foods, exotic berries, herbs, maca root powder, ribose, ephedra, colostrum, and hydroxy-methyl-buterate (HMB) than males. Over 90% believed it was essential to know if a supplement was third-party tested. However, only 57% stated the supplements bought were third-party tested. No sex differences were found for nutritional supplement knowledge, attitudes, and use of third-party testing programs. Our results indicate a need to improve student-athletes' attitudes toward and knowledge of nutritional supplements, and the initiation of programs to assist in the choosing and consuming of third-party tested supplements.


Tolerogenicity is not an absolute property of a dendritic cell.
Pharmacological modulation is known to temper the immune capacity of DC, enhancing the notion that modulated Ag-bearing DC might be used therapeutically to induce tolerance. We have investigated phenotypic features shared by such DC, and queried their potential to tolerize in different settings. Immature, IL-10, TGF-beta and 1alpha,25-dihydroxyvitamin D(3)-modulated BMDC all induced tolerance to male skin in female TCR transgenic A1.RAG mice, and the modulated DC also tolerized after exposure to the TLR4-ligand LPS. Transcript profiling revealed that this was achieved despite retaining much of the normal LPS-maturation response. No shared tolerance-associated transcripts could be identified. Equivalent BMDC could not tolerize in Marilyn TCR-transgenic mice. Simultaneous presentation of both A1.RAG and Marilyn peptide-Ag (Dby-H2E(k) and Dby-H2A(b)) on immature (C57BL/6JxCBA/Ca) F1 BMDC also only achieved tolerance in A1.RAG mice. Both strains registered Ag, but Foxp3(+) Treg were only induced in A1.RAG mice. In contrast, Marilyn T cells showed greater proliferation and an inflammatory bias, in response to Ag presented by immature F1 BMDC in vitro. In summary, while pharmacological agents can skew DC to reinforce their immature tolerogenic phenotype, the outcome of presentation is ultimately an integrated response including T-cell-intrinsic components that can over-ride for immune activation.


Outcomes of treatment of multiple ligament knee injuries.
Knee dislocations are rare and potentially devastating injuries. Significant displacement of the tibia and femur commonly disrupts multiple knee ligaments and also often results in profound disruption to the surrounding soft tissue envelope. Open wounds and neurologic and vascular insult can put the involved limb in jeopardy. Following reduction, the optimal management of the dislocated knee is unknown. Surgery to repair and/or reconstruct torn structures likely affords superior long-term function over nonoperative immobilization strategies. The role of early versus delayed surgery, repair versus reconstruction, and autograft versus allograft tissue for reconstruction remain topics of debate. High-quality research efforts to investigate these controversies are hampered by the heterogeneous nature of the injuries themselves and the many treatment strategies available.


iPiG: integrating peptide spectrum matches into genome browser visualizations.
Proteogenomic approaches have gained increasing popularity, however it is still difficult to integrate mass spectrometry identifications with genomic data due to differing data formats. To address this difficulty, we introduce iPiG as a tool for the integration of peptide identifications from mass spectrometry experiments into existing genome browser visualizations. Thereby, the concurrent analysis of proteomic and genomic data is simplified and proteomic results can directly be compared to genomic data. iPiG is freely available from https://sourceforge.net/projects/ipig/. It is implemented in Java and can be run as a stand-alone tool with a graphical user-interface or integrated into existing workflows. Supplementary data are available at PLOS ONE online.


Key elements of bioanalytical method validation for small molecules.
Method validation is a process that demonstrates that a method will successfully meet or exceed the minimum standards recommended in the Food and Drug Administration (FDA) guidance for accuracy, precision, selectivity, sensitivity, reproducibility, and stability. This article discusses the validation of bioanalytical methods for small molecules with emphasis on chromatographic techniques. We present current thinking on validation requirements as described in the current FDA Guidance and subsequent 2006 Bioanalytical Methods Validation Workshop white paper.


Specific interaction of human Tamm-Horsfall gylcoprotein with leucoagglutinin, a lectin from Phaseolus vulgaris (red kidney bean).
Human Tamm-Horsfall glycoprotein inhibits lymphocyte transformation induced by leucoagglutinin and haemagglutinin from Phaseolus vulgaris (red kidney bean). The glycoprotein interacts with the two lectins, giving insoluble precipitates. The interaction with leucoagglutinin is highly specific, and the shape of the precipitin curve is that of an antigen-antibody reaction; precipitation is specifically inhibited by N-acetyl-D-galactosamine. Results are discussed, and it is suggested that inhibition of lymphocyte transformation is due to competition between human Tamm-Horsfall glycoprotein and carbohydrate receptors on lymphocytes for the two lectins. The interaction between human Tamm-Horsfall glycoprotein and Phaseolus vulgaris lectins has been used to develop a one-step procedure for the separation of the two lectins by affinity chromatography on (human Tamm-Horsfall-glycoprotein)-Sepharose.


Nativity is associated with sugar-sweetened beverage and fast-food meal consumption among Mexican-origin women in Texas border colonias.
Trends of increasing obesity are especially pronounced among Mexican-origin women. There is little understanding of dietary patterns among U.S.- and Mexico-born Mexican-origin individuals residing in new-destination immigrant communities in the United States, especially behaviors related to obesity, such as consumption of sugar-sweetened beverages (SSB) and fast-food meals (FFM). The study used survey data of 599 adult Mexican-origin women from the 610 women who completed the 2009 Colonia Household and Community Food Resource Assessment (C-HCFRA), which was completed in person by trained promotora-researchers in 44 colonias near the Texas border towns of Progreso and La Feria. Data included demographic characteristics (age, education, nativity or country of birth, household income, household composition, and employment status), access to transportation, self-reported height and weight, food and nutrition assistance program participation, and consumption of SSB and FFM. Descriptive statistics were calculated by nativity (U.S.-born vs. Mexico-born); multivariable linear regression models were estimated for correlates of consumption of SSB and FFM. There are three major findings related to nativity. First, U.S.-born women consumed more SSB and FFM than Mexican-born counterparts in the same areas of colonias. Second, in the combined sample and controlling for other population characteristics, being born in Mexico was independently associated with FFM (fewer FFM), but not with SSB. Third, in analyses stratified by nativity, FFM and SSB were associated with each other among both nativity groups. Among Mexico-born women only, age, presence of a child, or being a lone parent was significantly associated with SSB; full-time employment, being a lone parent, and SSB consumption were each independently associated with increased frequency of FFM. Our analyses revealed differences in prevalence and correlates of SSB and FFM based on country of birth. Nativity, as a proxy for acculturation, may indicate the extent that immigrants have adopted behaviors from their new environment. However, nativity could also indicate limited accessibility to resources such as food/nutrition assistance programs, transportation, and proper documentation. Additionally, future research should focus on expanding our understanding of the meaning of nativity among individuals who share common contextual factors, but may have different life course experiences and resources needed to transition into a new place. Additional measures should be considered such as educational and occupational background, migration history, documentation status, and dietary acculturation, which may better explain heterogeneity within Hispanic subgroups.


Dynamic Monitoring Reveals Motor Task Characteristics in Prehistoric Technical Gestures.
Reconstructing ancient technical gestures associated with simple tool actions is crucial for understanding the co-evolution of the human forelimb and its associated control-related cognitive functions on the one hand, and of the human technological arsenal on the other hand. Although the topic of gesture is an old one in Paleolithic archaeology and in anthropology in general, very few studies have taken advantage of the new technologies from the science of kinematics in order to improve replicative experimental protocols. Recent work in paleoanthropology has shown the potential of monitored replicative experiments to reconstruct tool-use-related motions through the study of fossil bones, but so far comparatively little has been done to examine the dynamics of the tool itself. In this paper, we demonstrate that we can statistically differentiate gestures used in a simple scraping task through dynamic monitoring. Dynamics combines kinematics (position, orientation, and speed) with contact mechanical parameters (force and torque). Taken together, these parameters are important because they play a role in the formation of a visible archaeological signature, use-wear. We present our new affordable, yet precise methodology for measuring the dynamics of a simple hide-scraping task, carried out using a pull-to (PT) and a push-away (PA) gesture. A strain gage force sensor combined with a visual tag tracking system records force, torque, as well as position and orientation of hafted flint stone tools. The set-up allows switching between two tool configurations, one with distal and the other one with perpendicular hafting of the scrapers, to allow for ethnographically plausible reconstructions. The data show statistically significant differences between the two gestures: scraping away from the body (PA) generates higher shearing forces, but requires greater hand torque. Moreover, most benchmarks associated with the PA gesture are more highly variable than in the PT gesture. These results demonstrate that different gestures used in 'common' prehistoric tasks can be distinguished quantitatively based on their dynamic parameters. Future research needs to assess our ability to reconstruct these parameters from observed use-wear patterns.


Neuroethical considerations: cognitive liberty and converging technologies for improving human cognition.
Developers of NBIC (Nano-Bio-Info-Cogno) technologies face a multitude of obstacles, not the least of which is navigating the public ethics of their applied research. Biotechnologies have received widespread media attention and spawned heated interest in their perceived social implications. Now, in view of the rapidly expanding purview of neuroscience and the growing array of technologic developments capable of affecting or monitoring cognition, the emerging field of neuroethics calls for a consideration of the social and ethical implications of neuroscientific discoveries and trends. To negotiate the complex ethical issues at stake in new and emerging kinds of technologies for improving human cognition, we need to overcome political, disciplinary, and religious sectarianism. We need analytical models that protect values of personhood at the heart of a functional democracy-values that allow, as much as possible, for individual decision-making, despite transformations in our understanding and ability to manipulate cognitive processes. Addressing cognitive enhancement from the legal and ethical notion of "cognitive liberty" provides a powerful tool for assessing and encouraging NBIC developments.


HTO - A complementary ergonomics approach.
The field of human factors and ergonomics constitutes a strong potential in systems analysis, design and improvement. However, it is difficult to communicate its potential value. This paper addresses how the human-technology-organization (HTO) concept can be defined and supports the understanding, communication and development of the systems' character and potential of human factors and ergonomics. Empirical examples from the authors' experiences of working with the HTO concept in R&D and teaching are illustrated, including its usefulness as: 1) a conceptual model; 2) an analysis framework; 3) a meta methodology; 4) a pedagogical tool; and 5) a design tool. The use of HTO provides guidance on how the system can be designed to better support health, individual and systems performance. It is further suggested that there is a strong potential for developing the theory, applications and methodological aspects of HTO.


Adipose triglyceride lipase (ATGL) expression in human skeletal muscle is type I (oxidative) fiber specific.
Accumulation of triacylglycerol (TAG) and lipid intermediates in skeletal muscle plays an important role in the etiology of insulin resistance and type 2 diabetes mellitus. Disturbances in skeletal muscle lipid turnover and lipolysis may contribute significantly to this. So far, knowledge on the regulation of muscle lipolysis is limited. Recently the identification of a new lipase was reported: adipose triglyceride lipase (ATGL). ATGL deficient animals show significant lipid accumulation in skeletal muscle, which may indicate that ATGL plays a pivotal role in skeletal muscle lipolysis. However, until now, it is still unknown whether ATGL protein is expressed in human skeletal muscle. Therefore, the aim of the present study was to investigate whether ATGL is expressed at the protein level in human skeletal muscle, and to examine whether its expression is fiber-type specific. To accomplish this, we established an imunohistochemical and immunofluorescent staining procedure to study ATGL protein expression in relation to fiber type in human vastus lateralis muscle of eight male subjects (BMI range: 21.0-34.5 kg/m2 and age: 38-59 years). In the present paper we report for the first time that ATGL protein is indeed expressed in human skeletal muscle. Moreover, ATGL is exclusively expressed in type I (oxidative) muscle fibers, suggesting a pivotal role for ATGL in intramuscular fatty acid handling, lipid storage and breakdown.


Detecting and classifying three different hand movement types through electroencephalography recordings for neurorehabilitation.
Brain-computer interfaces can be used for motor substitution and recovery; therefore, detection and classification of movement intention are crucial for optimal control. In this study, palmar, lateral and pinch grasps were differentiated from the idle state and classified from single-trial EEG using only information prior to the movement onset. Fourteen healthy subjects performed the three grasps 100 times, while EEG was recorded from 25 electrodes. Temporal and spectral features were extracted from each electrode, and feature reduction was performed using sequential forward selection (SFS) and principal component analysis (PCA). The detection problem was investigated as the ability to discriminate between movement preparation and the idle state. Furthermore, all task pairs and the three movements together were classified. The best detection performance across movements (79 ± 8 %) was obtained by combining temporal and spectral features. The best movement-movement discrimination was obtained using spectral features: 76 ± 9 % (2-class) and 63 ± 10 % (3-class). For movement detection and discrimination, the performance was similar across grasp types and task pairs; SFS outperformed PCA. The results show it is feasible to detect different grasps and classify the distinct movements using only information prior to the movement onset, which may enable brain-computer interface-based neurorehabilitation of upper limb function through Hebbian learning mechanisms.


Treatment of patients with MYC rearrangement positive large B-cell lymphoma with R-CHOP plus lenalidomide: results of a multicenter HOVON phase II trial.
Patients with MYC-rearrangement positive large B-cell lymphoma (MYC+ LBCL) have an inferior prognosis following standard first-line therapy with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone (R-CHOP) as compared to patients without MYC rearrangement. Although intensive chemotherapy regimens yield higher remission rates, toxicity remains a concern. Lenalidomide is an oral immunomodulatory drug which downregulates MYC and its target genes thereby providing support using lenalidomide as additional therapeutic option for MYC+ LBCL. A phase II trial was conducted evaluating the efficacy of lenalidomide (15 mg day 1-14) in combination with R-CHOP (R2CHOP) in newly diagnosed MYC+ LBCL patients identified through a nationwide MYC-FISH screening program. The primary endpoint was complete metabolic response (CMR) on centrally reviewed 18F-fluorodeoxyglucose (18F-FDG) positron emission tomography (PET)-computer tomography (CT)-scan at end-of-treatment. Secondary endpoints were overall survival (OS), disease-free survival (DFS) and event-free survival (EFS). Eighty-two patients with stage II-IV MYC+ LBCL were treated with 6 cycles of R2CHOP. At EOT, 67% (confidence interval (CI) 58-75%) of the patients reached CMR. With a median follow-up of 25.4 months, 2-year estimates (95% CI) for OS, DFS, EFS were 73% (62-82%), 75% (63-84%) and 63% (52-73%) respectively. In this prospective trial for newly diagnosed MYC+ LBCL patients, we found that administering R2CHOP was safe, and yields comparable CMR and survival rates as in studies applying more intensive chemotherapy regimens. Hence, these findings offer new prospects for MYC+ LBCL patients and warrant comparison in prospective randomized clinical trials. This trial was registered at www.clinicaltrialsregister.eu (#2014-002654-39).


Zygotic Venlafaxine Exposure Impacts Behavioral Programming by Disrupting Brain Serotonin in Zebrafish.
The antidepressant venlafaxine, a selective serotonin and norepinephrine reuptake inhibitor, is present in surface waters downstream of wastewater treatment plants. We previously showed that zygotic venlafaxine deposition alters larval behavior in zebrafish (Danio rerio), but the mechanisms were unknown. Here we tested the hypothesis that venlafaxine disrupts central serotonergic development, leading to impaired behavioral responses in zebrafish larvae. This was tested by microinjecting embryos with venlafaxine immediately after fertilization and performing spatial distribution of serotonin immunoreactivity, as well as characterizing target genes involved in serotonin turnover in the zebrafish brain. We provide evidence that venlafaxine exposure reduces serotonin immunoreactivity and tyrosine hydroxylase-positive cell populations in specific larval brain regions, and this corresponded with reduced larval activity observed in the drug-exposed group. Lowered serotonin was not due to either reduced synthesis or increased breakdown capacity. However, co-injection of serotonin alongside venlafaxine in embryos recovered brain serotonin immunoreactivity, tyrosine hydroxylase-positive cell populations, and rescued venlafaxine-mediated behavioral changes. Overall, our results demonstrate for the first time that early life exposure to venlafaxine perturbs brain development, which may be due to reduced serotonin, leading to altered larval behavior in zebrafish.


Effects of a refugee elective on medical student perceptions.
There are growing numbers of refugees throughout the world. Refugee health is a relatively unstudied and rarely taught component of medical education. In response to this need, a Refugee Health Elective was begun. Medical student perceptions toward cultural aspects of medicine and refugee health before and after participation in the elective were measured. Preliminary questionnaires were given to all preclinical students at the academic year commencement with follow-up questionnaires at the refugee elective's conclusion. Both questionnaires examined students' comfort in interacting with patients and familiarity with refugee medical issues, alternative medical practices, and social hindrances to medical care. The preliminary answers served as a control and follow-up questionnaire data were separated into participant/non-participant categories. All preclinical medical students at two Midwestern medical schools were provided the opportunity to participate in the Refugee Health Elective and surveys. The 3 data groups were compared using unadjusted and adjusted analysis techniques with the Kruskall-Wallis, Bonferroni and ANCOVA adjustment. P-values < 0.05 were considered significant. 408 and 403 students filled out the preliminary and follow-up questionnaires, respectfully, 42 of whom participated in the elective. Students considering themselves minorities or multilingual were more likely to participate. Elective participants were more likely to be able to recognize the medical/mental health issues common to refugees, to feel comfortable interacting with foreign-born patients, and to identify cultural differences in understanding medical/mental health conditions, after adjusting for minority or multilingual status. As medical schools integrate a more multicultural curriculum, a Refugee Health Elective for preclinical students can enhance awareness and promote change in attitude toward medical/mental health issues common to refugees. This elective format offers tangible and effective avenues for these topics to be addressed.


Increasing imputation and prediction accuracy for Chinese Holsteins using joint Chinese-Nordic reference population.
This study investigated the effect of including Nordic Holsteins in the reference population on the imputation accuracy and prediction accuracy for Chinese Holsteins. The data used in this study include 85 Chinese Holstein bulls genotyped with both 54K chip and 777K (HD) chip, 2862 Chinese cows genotyped with 54K chip, 510 Nordic Holstein bulls genotyped with HD chip, and 4398 Nordic Holstein bulls genotyped with 54K chip and with deregressed proofs for five milk production traits. Based on these data, the accuracy of imputation from 54K to HD marker data and the accuracy of genomic predictions in Chinese Holstein were assessed. The allele correct rate increased around 2.7 and 1.7% in imputation from the 54K to the HD marker data for Chinese Holstein bulls and cows, respectively, when the Nordic HD-genotyped bulls were included in the reference data for imputation. However, the prediction accuracy was improved slightly when using the marker data imputed based on the combined HD reference data, compared with using the marker data imputed based on the Chinese HD reference data only. On the other hand, when using the combined reference population including 4398 Nordic Holstein bulls, the accuracy of genomic predictions increased 6.5 percentage points together with a reduction of prediction bias. The HD markers did not outperform the 54K markers in genomic prediction based on the present data. The results indicate that for Chinese Holsteins, it is necessary to genotype more individuals with 54K chip to increase reference population rather than increasing marker density.


Within-Mother Estimates of the Effects of WIC on Birth Outcomes in New York City.
There is a large literature suggesting that "WIC works" to improve birth outcomes. However, methodological limitations related to selection into the WIC program have left room for doubt about this conclusion. This paper uses birth records from New York City to address some limitations of the previous literature. We estimate models with mother fixed effects to control for fixed characteristics of mothers and we directly investigate the way that time-varying characteristics of mothers affect selection into the WIC program. We find that WIC is associated with reductions in low birth weight, even among full term infants, and with reductions in the probability that a child is "small for dates." These improvements are associated with a reduction in the probability that the mother gained too little weight during pregnancy. Improvements tend to be largest for first born children. We also find that women on WIC are more likely to be diagnosed with chronic conditions, and receive more intensive medical services, a finding that may reflect improved access to medical care.


β4 Integrin signaling induces expansion of prostate tumor progenitors.
The contextual signals that regulate the expansion of prostate tumor progenitor cells are poorly defined. We found that a significant fraction of advanced human prostate cancers and castration-resistant metastases express high levels of the β4 integrin, which binds to laminin-5. Targeted deletion of the signaling domain of β4 inhibited prostate tumor growth and progression in response to loss of p53 and Rb function in a mouse model of prostate cancer (PB-TAg mice). Additionally, it suppressed Pten loss-driven prostate tumorigenesis in tissue recombination experiments. We traced this defect back to an inability of signaling-defective β4 to sustain self-renewal of putative cancer stem cells in vitro and proliferation of transit-amplifying cells in vivo. Mechanistic studies indicated that mutant β4 fails to promote transactivation of ErbB2 and c-Met in prostate tumor progenitor cells and human cancer cell lines. Pharmacological inhibition of ErbB2 and c-Met reduced the ability of prostate tumor progenitor cells to undergo self-renewal in vitro. Finally, we found that β4 is often coexpressed with c-Met and ErbB2 in human prostate cancers and that combined pharmacological inhibition of these receptor tyrosine kinases exerts antitumor activity in a mouse xenograft model. These findings indicate that the β4 integrin promotes prostate tumorigenesis by amplifying ErbB2 and c-Met signaling in tumor progenitor cells.


Percutaneous irreversible electroporation for treatment of locally advanced pancreatic cancer following chemotherapy or radiochemotherapy.
Irreversible electroporation (IRE) is a non-thermal based tumor ablation method used close to vessels and ducts and has the potential of treating locally advanced pancreatic cancer (LAPC). The aim of this study was to evaluate the efficacy and safety of IRE in patients with LAPC after chemo- and/or radio-chemotherapy. Twenty-four patients with biopsy proven LAPC and who had received chemo- and/or radio-chemotherapy with no signs of metastases were included and treated with ultrasound guided percutaneous IRE under general anesthesia. The median overall survival from diagnosis of LAPC was 17.9 months; this included 7.0 months after IRE. Median time from IRE was 6.1 months to local progression and 2.7 months to observation of metastases. Local control was observed in nine patients. IRE related complications were observed in 11 patients, three of which were serious complications. There was no IRE related mortality. Percutaneous IRE is reasonably safe in LAPC after chemo-/radio-chemotherapy and with promising results regarding efficacy.


[AIDS. New challenges for social and medical prevention].
Anti-retroviral therapy (ART) produces spectacular improvements in life expectancy and quality of life for people infected with HIV, and contributes to primary prevention in the wider population by reducing the viral load. Many people infected with HIV begin therapy later than indicated, while, despite ongoing prevention efforts, the number of new HIV diagnoses is increasing, along with the incidence of other STIs and, in identifiable sub-groups, of hepatitis C, above all among men who have sex with men (MSM). The prevention consequences of this complex situation are discussed in the context of the alternative between control and containment (Suchstrategie) and inclusion and cooperation (Lernstrategie), arguing for HIV prevention to be integrated in the broader paradigm of sexual health and sub-group-specific efforts to increase the willingness to undergo testing both through community-based campaigns and in the health care context on the basis of informed consent and counseling. Above all ethical considerations mitigate against an undifferentiated test-and-treat approach. The contribution identifies research gaps and institutional obstacles that stand in the way of achievable advances and productive linkage of social and medical prevention.


Bufalin suppresses the proliferation and metastasis of renal cell carcinoma by inhibiting the PI3K/Akt/mTOR signaling pathway.
Bufalin, one of the active ingredients of the Chinese drug Chan su, exhibits significant antitumor activity against various cancer types. However, the role of bufalin in renal cell carcinoma (RCC) remains unclear. In the present study, it was demonstrated that bufalin inhibited cell proliferation, blocked the cell cycle in the G2/M phase, and reduced the metastasis of human RCC ACHN cells via the upregulation of p21waf/cip1 and E-cadherin and the downregulation of cyclin dependent kinase 1, cyclin B1, N-cadherin, and hypoxia-inducible factor-1α (HIF-1α). Further mechanistic study revealed that bufalin reduced the expression of phosphorylated (phospho)-Akt and phospho-mammalian target of rapamycin (mTOR). Moreover, HIF-1α expression may be regulated through the inhibition of the phosphoinositide 3-kinase (PI3K)/protein kinase B (Akt)/mTOR signaling pathway. Thus, the present results suggest that bufalin induces cell cycle arrest and suppresses metastasis; this process may be associated with the PI3K/Akt/mTOR signaling pathway. Accordingly, it is suggested that bufalin is a therapeutic agent for RCC.


Assessment of inpatient paediatric care in first referral level hospitals in 13 districts in Kenya.
The district hospital is considered essential for delivering basic, cost-effective health care to children in resource poor countries. We aimed to investigate the performance of these facilities in Kenya. Government hospitals providing first referral level care were prospectively sampled from 13 Kenyan districts. Workload statistics and data documenting the management and care of admitted children were obtained by specially trained health workers. Data from 14 hospitals were surveyed with routine statistics showing considerable variation in inpatient paediatric mortality (range 4-15%) and specific case fatality rates (eg, anaemia 3-46%). The value of these routine data is seriously undermined by missing data, apparent avoidance of a diagnosis of HIV/AIDS, and absence of standard definitions. Case management practices are often not in line with national or international guidelines. For malaria, signs defining severity such as the level of consciousness and degree of respiratory distress are often not documented (range per hospital 0-100% and 9-77%, respectively), loading doses of quinine are rarely given (3% of cases) and dose errors are not uncommon. Resource constraints such as a lack of nutritional supplements for malnourished children also restrict the provision of basic, effective care. Even crude performance measures suggest there is a great need to improve care and data quality, and to identify and tackle key health system constraints at the first referral level in Kenya. Appropriate intervention might lead to more effective use of health workers' efforts in such hospitals.


Ribosome profiling: a Hi-Def monitor for protein synthesis at the genome-wide scale.
Ribosome profiling or ribo-seq is a new technique that provides genome-wide information on protein synthesis (GWIPS) in vivo. It is based on the deep sequencing of ribosome protected mRNA fragments allowing the measurement of ribosome density along all RNA molecules present in the cell. At the same time, the high resolution of this technique allows detailed analysis of ribosome density on individual RNAs. Since its invention, the ribosome profiling technique has been utilized in a range of studies in both prokaryotic and eukaryotic organisms. Several studies have adapted and refined the original ribosome profiling protocol for studying specific aspects of translation. Ribosome profiling of initiating ribosomes has been used to map sites of translation initiation. These studies revealed the surprisingly complex organization of translation initiation sites in eukaryotes. Multiple initiation sites are responsible for the generation of N-terminally extended and truncated isoforms of known proteins as well as for the translation of numerous open reading frames (ORFs), upstream of protein coding ORFs. Ribosome profiling of elongating ribosomes has been used for measuring differential gene expression at the level of translation, the identification of novel protein coding genes and ribosome pausing. It has also provided data for developing quantitative models of translation. Although only a dozen or so ribosome profiling datasets have been published so far, they have already dramatically changed our understanding of translational control and have led to new hypotheses regarding the origin of protein coding genes.


Differentiating between monozygotic twins through DNA methylation-specific high-resolution melt curve analysis.
Although short tandem repeat profiling is extremely powerful in identifying individuals from crime scene stains, it is unable to differentiate between monozygotic (MZ) twins. Efforts to address this include mutation analysis through whole genome sequencing and through DNA methylation studies. Methylation of DNA is affected by environmental factors; thus, as MZ twins age, their DNA methylation patterns change. This can be characterized by bisulfite treatment followed by pyrosequencing. However, this can be time-consuming and expensive; thus, it is unlikely to be widely used by investigators. If the sequences are different, then in theory the melting temperature should be different. Thus, the aim of this study was to assess whether high-resolution melt curve analysis can be used to differentiate between MZ twins. Five sets of MZ twins provided buccal swabs that underwent extraction, quantification, bisulfite treatment, polymerase chain reaction amplification and high-resolution melting curve analysis targeting two markers, Alu-E2F3 and Alu-SP. Significant differences were observed between all MZ twins targeting Alu-E2F3 and in four of five MZ twins targeting Alu-SP (P<0.05). Thus, it has been demonstrated that bisulfite treatment followed by high-resolution melting curve analysis could be used to differentiate between MZ twins.


Disentangling nutritional factors and household characteristics related to child stunting and maternal overweight in Guatemala.
The aim of this study was to identify nutritional factors and households characteristics associated with child stunting, maternal overweight and the familial coexistence of both types of malnutrition. In Guatemala, 2000, with nationally representative data, we selected 2261 households with at least one child aged 12-60 months and his/her mother. Nutritional status was assessed in children (e.g., stunting as height-for-age Z-score<-2) and mothers (e.g., overweight as body mass index > or =25 kg/m(2)) and identified the presence of both, child stunting and maternal overweight in the same household (SCOM). With logistic regression models we assessed the association of the malnutrition indicators with individual and household socio-economic and health characteristics. SCOM was identified in 18% of households. Socio-economic status (SES) of SCOM households was significantly lower than SES of households with non-stunted children. SCOM households, compared to those with normal-stature children and normal weight mothers, were more likely to have mothers of short stature (adjusted odds ratio-OR+/-95% CI=3.1 (2.1-4.7)), higher parity (1.2 (1.1-1.3)), currently working (1.7 (1.1-2.6), and self-identified as indigenous (2.0 (1.3-3.1)). Factors associated with stunting in children such as poverty, maternal short stature and indigenousness, were predictors of SCOM. These findings support the notion that SCOM is an extension of the malnutrition spectrum in the most disadvantaged population groups in countries that are in the middle of their nutrition transitions such as Guatemala. At the same time it revealed that these populations are already in the stage of chronic, nutrition related diseases associated with less physical activity and more access to highly processed foods of low cost, high dietary energy and low nutrient density in important population groups. The challenge for the decision makers and service deliverers is to guide SCOM households to deal equally with both extremes of the malnutrition continuum.


Epigenetic Control of Skeletal Development by the Histone Methyltransferase Ezh2.
Epigenetic control of gene expression is critical for normal fetal development. However, chromatin-related mechanisms that activate bone-specific programs during osteogenesis have remained underexplored. Therefore, we investigated the expression profiles of a large cohort of epigenetic regulators (>300) during osteogenic differentiation of human mesenchymal cells derived from the stromal vascular fraction of adipose tissue (AMSCs). Molecular analyses establish that the polycomb group protein EZH2 (enhancer of zeste homolog 2) is down-regulated during osteoblastic differentiation of AMSCs. Chemical inhibitor and siRNA knockdown studies show that EZH2, a histone methyltransferase that catalyzes trimethylation of histone 3 lysine 27 (H3K27me3), suppresses osteogenic differentiation. Blocking EZH2 activity promotes osteoblast differentiation and suppresses adipogenic differentiation of AMSCs. High throughput RNA sequence (mRNASeq) analysis reveals that EZH2 inhibition stimulates cell cycle inhibitory proteins and enhances the production of extracellular matrix proteins. Conditional genetic loss of Ezh2 in uncommitted mesenchymal cells (Prrx1-Cre) results in multiple defects in skeletal patterning and bone formation, including shortened forelimbs, craniosynostosis, and clinodactyly. Histological analysis and mRNASeq profiling suggest that these effects are attributable to growth plate abnormalities and premature cranial suture closure because of precocious maturation of osteoblasts. We conclude that the epigenetic activity of EZH2 is required for skeletal patterning and development, but EZH2 expression declines during terminal osteoblast differentiation and matrix production.


Lactation and maternal risk of type 2 diabetes: a population-based study.
Lactation has been associated with improvements in maternal glucose metabolism. We explored the relationships between lactation and risk of type 2 diabetes in a well-characterized, population-representative cohort of women, aged 40-78 years, who were members of a large integrated health care delivery organization in California and enrolled in the Reproductive Risk factors for Incontinence Study at Kaiser (RRISK), between 2003 and 2008. Multivariable logistic regression was used to control for age, parity, race, education, hysterectomy, physical activity, tobacco and alcohol use, family history of diabetes, and body mass index while examining the impact of duration, exclusivity, and consistency of lactation on risk of having developed type 2 diabetes. Of 2233 women studied, 1828 were mothers; 56% had breastfed an infant for > or =1 month. In fully adjusted models, the risk of type 2 diabetes among women who consistently breastfed all of their children for > or =1 month remained similar to that of women who had never given birth (odds ratio [OR] 1.01; 95% confidence interval [CI], 0.56-1.81). In contrast, mothers who had never breastfed an infant were more likely to have developed type 2 diabetes than nulliparous women (OR 1.93; 95% CI, 1.14-3.27) [corrected]. Mothers who never exclusively breastfed were more likely to have developed type 2 diabetes than mothers who exclusively breastfed for 1-3 months (OR 1.52; 95% CI, 1.11-2.10). Risk of type 2 diabetes increases when term pregnancy is followed by <1 month of lactation, independent of physical activity and body mass index in later life. Mothers should be encouraged to exclusively breastfeed all of their infants for at least 1 month.


Determinants of quantitative spectral electroencephalography in early Alzheimer's disease: cognitive function, regional cerebral blood flow, and computed tomography.
Electroencephalography (EEG) bands may have different clinical or physiological correlates at initial diagnosis of Alzheimer's disease (AD). We studied 163 consecutive patients with probable (n = 105) and possible (n = 58) AD with measurements of cognitive function (CAMCOG), regional cerebral blood flow (rCBF) with single photon emission computed tomography using technetium-99m-labeled hexamethylpropylene amine oxime, and computed tomography (CT). Lower CAMCOG scores were significantly and most strongly associated with lower parieto-occipital and fronto-central alpha power. In a separate analysis of cognitive domains, disturbances in language, praxis, attention, and abstraction were also significantly and most consistently related to decrease in alpha power. Presence of cortical atrophy as measured on CT showed some statistically significant relations with EEG bands, but these associations were not consistent. Lower temporal and parietal rCBF were significantly related to lower parieto-occipital alpha activity. Presence of leukoaraiosis was significantly associated with lower beta values, but also with higher absolute theta and delta activity. The results suggest that alpha on EEG is most closely linked to cognitive function and rCBF, while beta and theta activity more likely reflect lower cortical or subcortical changes. Our study thus provides evidence that the EEG bands reflect differential pathophysiologic changes in AD.


Influence of oriented nanofibrous PCL scaffolds on quantitative gene expression during neural differentiation of mouse embryonic stem cells.
Neural differentiation of mouse embryonic stem cells in combination with three-dimensional electrospun nanofibers as an artificial extracellular matrix can be utilized to reconstruct a spinal cord defect. In this study, random and parallel-aligned nanofibrous poly ɛ-caprolactone was fabricated using electrospinning. Its hydrophobicity was modified by O2 plasma treatment to facilitate enhanced cell attachment. Embryoid bodies (EBs), which contain all three embryonic germ layers, were cultured on poly ɛ-caprolactone scaffolds to study the effect of fiber orientation on cell morphology and differentiation. Cell morphology and neuron-specific gene and protein expressions were, respectively, evaluated by scanning electron microscopy, real-time polymerase chain reaction, and immunocytochemistry. Although two types of nanofibrous scaffolds showed neural marker expression at the protein level, cells on randomly oriented scaffolds showed short-range topographical guidance and stretched across multiple directions, whereas cells on the parallel scaffolds exhibited long extension with enhanced neuron outgrowth along the fiber, producing oriented extracellular matrix, leading to direct cell migration and nerve regeneration. Quantitative real-time polymerase chain reaction showed that both aligned and random electrospun nanofibers downregulated the precursor neural marker Nestin compared with that in the control group, a gelatin-coated tissue culture plate (T). Analysis also showed higher expression of dorso-ventral neural markers (Isl1/2 and Lim1/2) than motor neuron progenitor markers (Pax6, Nkx6.1, and olig2) in aligned nanofibers than in the T group. Moreover, aligned nanofibers showed higher expression of mature neural specific markers such as β-tub and Map2 than those in the randomly oriented scaffolds. Therefore, we conclude that nanofibers with different orientations can support the neural lineage, but aligned nanofibrous scaffolds are superior candidates to promote the advancement of neural precursors to achieve maturity during the differentiation process.


Selection of muscle and nerve-cuff electrodes for neuroprostheses using customizable musculoskeletal model.
Neuroprosthetic systems based on functional electrical stimulation aim to restore motor function to individuals with paralysis following spinal cord injury. Identifying the optimal electrode set for the neuroprosthesis is complicated because it depends on the characteristics of the individual (such as injury level), the force capacities of the muscles, the movements the system aims to restore, and the hardware limitations (number and type of electrodes available). An electrode-selection method has been developed that uses a customized musculoskeletal model. Candidate electrode sets are created based on desired functional outcomes and the hard ware limitations of the proposed system. Inverse-dynamic simulations are performed to determine the proportion of target movements that can be accomplished with each set; the set that allows the most movements to be performed is chosen as the optimal set. The technique is demonstrated here for a system recently developed by our research group to restore whole-arm movement to individuals with high-level tetraplegia. The optimal set included selective nerve-cuff electrodes for the radial and musculocutaneous nerves; single-channel cuffs for the axillary, suprascapular, upper subscapular, and long-thoracic nerves; and muscle-based electrodes for the remaining channels. The importance of functional goals, hardware limitations, muscle and nerve anatomy, and surgical feasibility are highlighted.


The PML-associated protein DEK regulates the balance of H3.3 loading on chromatin and is important for telomere integrity.
Histone variant H3.3 is deposited in chromatin at active sites, telomeres, and pericentric heterochromatin by distinct chaperones, but the mechanisms of regulation and coordination of chaperone-mediated H3.3 loading remain largely unknown. We show here that the chromatin-associated oncoprotein DEK regulates differential HIRA- and DAAX/ATRX-dependent distribution of H3.3 on chromosomes in somatic cells and embryonic stem cells. Live cell imaging studies show that nonnucleosomal H3.3 normally destined to PML nuclear bodies is re-routed to chromatin after depletion of DEK. This results in HIRA-dependent widespread chromatin deposition of H3.3 and H3.3 incorporation in the foci of heterochromatin in a process requiring the DAXX/ATRX complex. In embryonic stem cells, loss of DEK leads to displacement of PML bodies and ATRX from telomeres, redistribution of H3.3 from telomeres to chromosome arms and pericentric heterochromatin, induction of a fragile telomere phenotype, and telomere dysfunction. Our results indicate that DEK is required for proper loading of ATRX and H3.3 on telomeres and for telomeric chromatin architecture. We propose that DEK acts as a "gatekeeper" of chromatin, controlling chromatin integrity by restricting broad access to H3.3 by dedicated chaperones. Our results also suggest that telomere stability relies on mechanisms ensuring proper histone supply and routing.


[Effect of acupuncture stimulation of Zusanli (ST 36) on cerebral regional homogeneity in volunteer subjects with different constitutions: a resting-state fMRI study].
To observe the synchronism difference of brain region activities in response to acupuncture stimulation of Zusanli (ST 36) in healthy volunteer subjects with different acupuncture analgesia sensitivity, so as to study the central factors influencing acupuncture intervention outcomes. Forty-five healthy volunteer subjects with different constitutions (different sensitivities in response to needling stimulation) were divided into insensitive group, normal group and sensitive group (n = 15). The pressure pain threshold (PPT) of the Zusanli (ST 36) region before and after acupuncture stimulation of ST36 was assessed using visual analog scale (VAS). Two weeks later after acupuncture stimulation of ST 36, resting-state fMRI images were acquired by using a nuclear magnetic resonance imaging system and analyzed by using DPARSFV 2.1 software package, software SPM 8 and REST 1.7. The cerebral regional homogeneity (ReHo) of the subjects was then calculated by Resting-State fMRI Data Analysis Toolkit (REST). Compared with pre-acupuncture, PPT levels of the normal and sensitive groups were significantly increased after acupuncture of ST 36 (P < 0.05), and that of the insensitive group had no significant change (P > 0.05). Following acupuncture stimulation of ST 36, the insensitive group only showed a significant decreased ReHo in the left fusiform gyrus, left inferior temporal gyrus, bilateral postcentral gyrus, and left anterior central gyrus. In the normal group, a significantly increased ReHo was found in left brainstem, the right cerebellum posterior lobe, right parahippocampa gyrus, right fusiform gyrus, left angular gyrus, temporal lobe and the left frontal lobe; and a significantly decreased ReHo in the occipital lobes and the right superior temporal gyrus after acupuncture stimulation of ST 36. In the sensitive group, a markedly increased ReHo was found in the left brainstem, bilateral cerebellum posterior lobes, left inferior temporal gyrus, basal ganglia, the left insular lobe, anterior cingutate, frontal lobe, inferior parietal lobule, and the right supplementary motor area, and an obviously decreased ReHo found in the bilateral occipital lobes, fusiform gyrus, posterior central gyrus, the right posterior cingutate, the left temporal lobe and the left paracentral lobule, etc. after acupuncture of ST 36. Constitution-associated needling sensation may be an important influential factor for acupuncture analgesia in normal subjects. The change of ReHo in different cerebral areas is probably responsible for the difference of acupuncture analgesia in different constitution people.


Two phase 3 trials of bapineuzumab in mild-to-moderate Alzheimer's disease.
Bapineuzumab, a humanized anti-amyloid-beta monoclonal antibody, is in clinical development for the treatment of Alzheimer's disease. We conducted two double-blind, randomized, placebo-controlled, phase 3 trials involving patients with mild-to-moderate Alzheimer's disease--one involving 1121 carriers of the apolipoprotein E (APOE) ε4 allele and the other involving 1331 noncarriers. Bapineuzumab or placebo, with doses varying by study, was administered by intravenous infusion every 13 weeks for 78 weeks. The primary outcome measures were scores on the 11-item cognitive subscale of the Alzheimer's Disease Assessment Scale (ADAS-cog11, with scores ranging from 0 to 70 and higher scores indicating greater impairment) and the Disability Assessment for Dementia (DAD, with scores ranging from 0 to 100 and higher scores indicating less impairment). A total of 1090 carriers and 1114 noncarriers were included in the efficacy analysis. Secondary outcome measures included findings on positron-emission tomographic amyloid imaging with the use of Pittsburgh compound B (PIB-PET) and cerebrospinal fluid phosphorylated tau (phospho-tau) concentrations. There were no significant between-group differences in the primary outcomes. At week 78, the between-group differences in the change from baseline in the ADAS-cog11 and DAD scores (bapineuzumab group minus placebo group) were -0.2 (P=0.80) and -1.2 (P=0.34), respectively, in the carrier study; the corresponding differences in the noncarrier study were -0.3 (P=0.64) and 2.8 (P=0.07) with the 0.5-mg-per-kilogram dose of bapineuzumab and 0.4 (P=0.62) and 0.9 (P=0.55) with the 1.0-mg-per-kilogram dose. The major safety finding was amyloid-related imaging abnormalities with edema among patients receiving bapineuzumab, which increased with bapineuzumab dose and APOE ε4 allele number and which led to discontinuation of the 2.0-mg-per-kilogram dose. Between-group differences were observed with respect to PIB-PET and cerebrospinal fluid phospho-tau concentrations in APOE ε4 allele carriers but not in noncarriers. Bapineuzumab did not improve clinical outcomes in patients with Alzheimer's disease, despite treatment differences in biomarkers observed in APOE ε4 carriers. (Funded by Janssen Alzheimer Immunotherapy and Pfizer; Bapineuzumab 301 and 302 ClinicalTrials.gov numbers, NCT00575055 and NCT00574132, and EudraCT number, 2009-012748-17.).


Efficient synthesis of enzymatically active calf chymosin in Saccharomyces cerevisiae.
We have constructed a high-efficiency expression vector to direct the synthesis of heterologous polypeptides in yeast. The vector is termed a sandwich expression vector as the heterologous gene is inserted between the 5' and 3' control regions of the efficiently expressed yeast PGK gene. We have used this vector to direct the expression of three derivatives of the calf chymosin cDNA gene; preprochymosin, prochymosin and chymosin. Prochymosin is synthesised to at least 5% of total yeast-cell protein and furthermore, it can be readily activated to produce an enzyme which has milk-clotting activity.


Synthesis of the Novel Covalent Cysteine Proteases Inhibitor with Iodoacetic Functional Group.
This work presents the synthesis of the novel covalent inhibitor of cysteine proteases where epoxide has been replaced by the iodoacetyl functional group. The molecule, similar in action to E-64 and DCG-04, the commonly applied inhibitors, is additionally biotinylated and contains tyrosyl iodination sites. The Fmoc solid phase synthesis has been applied. Conjugation of iodoacetic acid with the peptide was optimized by testing different conjugation agents. The purity of the final product was verified by mass spectrometry and its bioactivity was tested by incubation with a model cysteine protease-staphopain C. Finally, it was shown that the synthesized inhibitor binds to the protein at the ratio of 1:1. More detailed analysis by means of tandem mass spectrometry proved that the inhibitor binds to the cysteine present in the active site of the enzyme.


Competing demands in the office visit: what influences mammography recommendations?
The multiple competing demands of the busy office visit have been shown to interfere with delivery of preventive services. In this study we used physician recommendations for screening mammography to examine the relative importance of physician, patient, and visit characteristics in determining on which patient visits this preventive service will be provided. Physicians in the Ambulatory Sentinel Practice Network (ASPN) completed a questionnaire describing their knowledge, attitudes, and beliefs about screening mammography. They also described the content of a series of nonacute care visits with women aged 40 to 75 years with regard to making a recommendation when the patient was due for screening mammography. The data were linked, and univariate and multivariate logistic regression methods were used to examine the relative importance of physician, patient, and visit characteristics on making a recommendation for mammography. Ninety-three physicians reported making a recommendation for screening mammography on 53.1% of nonacute visits. When modeling physician, patient, and visit characteristics separately, 70% of the variability in the model is explained by physician characteristics only, 63% by patient characteristics only, and 73% by visit characteristics only. A combined model using all characteristics explained 85% of the variability. Although characteristics of physicians and patients can predict frequency of recommendations for mammography, the specific characteristics of the visit are equally important. Efforts to improve delivery of preventive services in primary care that emphasize physician education and performance feedback are unlikely to increase rates of mammography recommendation. Effective strategies must consider the multiple competing demands faced by patients and physicians during each office visit and seek ways for assisting them in setting rational priorities for services.


Molecular epidemiology of malaria in Cameroon. XVIII. Polymorphisms of the Plasmodium falciparum merozoite surface antigen-2 gene in isolates from symptomatic patients.
Merozoite surface antigen-2 (MSA-2) is a polymorphic genetic marker that is highly discriminatory for characterizing Plasmodium falciparum field isolates. Genetic diversity of isolates obtained from symptomatic patients residing in Yaounde, Cameroon was analyzed by an allele-specific polymerase chain reaction and direct sequencing of amplification products. Of 137 isolates, 25 (18%) had only FC27-type alleles, 40 (29%) had only 3D7-type alleles, and 72 (53%) had multiple parasite populations with both alleles. Of 295 fragments, 145 (49.2%) and 150 (50.8%) belonged to FC27 and 3D7 alleles, respectively. There were 23 different MSA-2 alleles (10 FC27-type and 13 3D7-type that yielded 44 different combinations in multiple infections). DNA sequencing showed distinct individual sequences. Sequences belonging to the FC27 allelic family were relatively conserved, with most of the polymorphism arising from differences in the number of repeat units. In contrast, the sequences within the GSA-rich region in 3D7 allelic family were less conserved, but many of the sequences in Cameroonian isolates have been identified in other isolates from geographically distant origins. Our results show an extensive diversity of the central region of MSA-2 in size, allelic family, combinations of these two features in multiple infections, and sequence variations underlying the complex population structure of P. falciparum clinical isolates in Yaounde, Cameroon.


Hypometabolism as the ultimate defence in stress response: how the comparative approach helps understanding of medically relevant questions.
First conceptualized from breath-hold diving mammals, later recognized as the ultimate cell autonomous survival strategy in anoxia-tolerant vertebrates and burrowing or hibernating rodents, hypometabolism is typically recruited by resilient organisms to withstand and recover from otherwise life-threatening hazards. Through the coordinated down-regulation of biosynthetic, proliferative and electrogenic expenditures at times when little ATP can be generated, a metabolism turned 'down to the pilot light' allows the re-balancing of energy demand with supply at a greatly suppressed level in response to noxious exogenous stimuli or seasonal endogenous cues. A unifying hallmark of stress-tolerant organisms, the adaptation effectively prevents lethal depletion of ATP, thus delineating a marked contrast with susceptible species. Along with disengaged macromolecular syntheses, attenuated transmembrane ion shuttling and PO2 -conforming respiration rates, the metabolic slowdown in tolerant species usually culminates in a non-cycling, quiescent phenotype. However, such a reprogramming also occurs in leading human pathophysiologies. Ranging from microbial infections through ischaemia-driven infarcts to solid malignancies, cells involved in these disorders may again invoke hypometabolism to endure conditions non-permissive for growth. At the same time, their reduced activities underlie the frequent development of a general resistance to therapeutic interventions. On the other hand, a controlled induction of hypometabolic and/or hypothermic states by pharmacological means has recently stimulated intense research aimed at improved organ preservation and patient survival in situations requiring acutely administered critical care. The current review article therefore presents an up-to-date survey of concepts and applications of a coordinated and reversibly down-regulated metabolic rate as the ultimate defence in stress responses.


Anxious and non-anxious major depressive disorder in the World Health Organization World Mental Health Surveys.
To examine cross-national patterns and correlates of lifetime and 12-month comorbid DSM-IV anxiety disorders among people with lifetime and 12-month DSM-IV major depressive disorder (MDD). Nationally or regionally representative epidemiological interviews were administered to 74 045 adults in 27 surveys across 24 countries in the WHO World Mental Health (WMH) Surveys. DSM-IV MDD, a wide range of comorbid DSM-IV anxiety disorders, and a number of correlates were assessed with the WHO Composite International Diagnostic Interview (CIDI). 45.7% of respondents with lifetime MDD (32.0-46.5% inter-quartile range (IQR) across surveys) had one of more lifetime anxiety disorders. A slightly higher proportion of respondents with 12-month MDD had lifetime anxiety disorders (51.7%, 37.8-54.0% IQR) and only slightly lower proportions of respondents with 12-month MDD had 12-month anxiety disorders (41.6%, 29.9-47.2% IQR). Two-thirds (68%) of respondents with lifetime comorbid anxiety disorders and MDD reported an earlier age-of-onset (AOO) of their first anxiety disorder than their MDD, while 13.5% reported an earlier AOO of MDD and the remaining 18.5% reported the same AOO of both disorders. Women and previously married people had consistently elevated rates of lifetime and 12-month MDD as well as comorbid anxiety disorders. Consistently higher proportions of respondents with 12-month anxious than non-anxious MDD reported severe role impairment (64.4 v. 46.0%; χ 2 1 = 187.0, p < 0.001) and suicide ideation (19.5 v. 8.9%; χ 2 1 = 71.6, p < 0.001). Significantly more respondents with 12-month anxious than non-anxious MDD received treatment for their depression in the 12 months before interview, but this difference was more pronounced in high-income countries (68.8 v. 45.4%; χ 2 1 = 108.8, p < 0.001) than low/middle-income countries (30.3 v. 20.6%; χ 2 1 = 11.7, p < 0.001). Patterns and correlates of comorbid DSM-IV anxiety disorders among people with DSM-IV MDD are similar across WMH countries. The narrow IQR of the proportion of respondents with temporally prior AOO of anxiety disorders than comorbid MDD (69.6-74.7%) is especially noteworthy. However, the fact that these proportions are not higher among respondents with 12-month than lifetime comorbidity means that temporal priority between lifetime anxiety disorders and MDD is not related to MDD persistence among people with anxious MDD. This, in turn, raises complex questions about the relative importance of temporally primary anxiety disorders as risk markers v. causal risk factors for subsequent MDD onset and persistence, including the possibility that anxiety disorders might primarily be risk markers for MDD onset and causal risk factors for MDD persistence.


Solving a 50 year mystery of a missing OPA1 mutation: more insights from the first family diagnosed with autosomal dominant optic atrophy.
Up to the 1950s, there was an ongoing debate about the diversity of hereditary optic neuropathies, in particular as to whether all inherited optic atrophies can be ascribed to Leber's hereditary optic neuropathy (LHON) or represent different disease entities. In 1954 W. Jaeger published a detailed clinical and genealogical investigation of a large family with explicit autosomal dominant segregation of optic atrophy thus proving the existence of a discrete disease different from LHON, which is nowadays known as autosomal dominant optic atrophy (ADOA). Since the year 2000 ADOA is associated with genomic mutations in the OPA1 gene, which codes for a protein that is imported into mitochondria where it is required for mitochondrial fusion. Interestingly enough, the underlying mutation in this family has not been identified since then. We have reinvestigated this family with the aim to identify the mutation and to further clarify the underlying pathomechanism. Patients showed a classical non-syndromic ADOA. The long term deterioration in vision in the two teenagers examined 50 years later is of particular note 5/20 to 6/120. Multiplex ligation probe amplification revealed a duplication of the OPA1 exons 7-9 which was confirmed by long distance PCR and cDNA analysis, resulting in an in-frame duplication of 102 amino acids. Segregation was verified in 53 available members of the updated pedigree and a penetrance of 88% was calculated. Fibroblast cultures from skin biopsies were established to assess the mitochondrial network integrity and to qualitatively and quantitatively study the consequences of the mutation on transcript and protein level. Fibroblast cultures demonstrated a fragmented mitochondrial network. Processing of the OPA1 protein was altered. There was no correlation of the OPA1 transcript levels and the OPA1 protein levels in the fibroblasts. Intriguingly an overall decrease of mitochondrial proteins was observed in patients' fibroblasts, while the OPA1 transcript levels were elevated. The thorough study of this family provides a detailed clinical picture accompanied by a molecular investigation of patients' fibroblasts. Our data show a classic OPA1-associated non-syndromic ADOA segregating in this family. Cell biological findings suggest that OPA1 is regulated by post-translational mechanisms and we would like to hypothesize that loss of OPA1 function might lead to impaired mitochondrial quality control. With the clinical, genetic and cell biological characterisation of a family described already more than 50 years ago, we span more than half a century of research in optic neuropathies.


Cerebral Artery Remodeling in Rodent Models of Subarachnoid Hemorrhage.
Vasospasm is known to contribute to delayed cerebral ischemia following subarachnoid hemorrhage (SAH). We hypothesized that vasospasm initiates structural changes within the vessel wall, possibly aggravating ischemia and leading to resistance to vasodilator treatment. We therefore investigated the effect of blood on cerebral arteries with respect to contractile activation and vascular remodeling. In vitro experiments on rodent basilar and middle cerebral arteries showed a gradual contraction in response to overnight exposure to blood. After incubation with blood, a clear inward remodeling was found, reducing the caliber of the passive vessel. The transglutaminase inhibitor L682.777 fully prevented this remodeling. Translation of the in vitro findings to an in vivo SAH model was attempted in rats, using both a single prechiasmatic blood injection model and a double cisterna magna injection model, and in mice, using a single prechiasmatic blood injection. However, we found no substantial changes in active or passive biomechanical properties in vivo. We conclude that extravascular blood can induce matrix remodeling in cerebral arteries, which reduces vascular caliber. This remodeling depends on transglutaminase activity. However, the current rodent SAH models do not permit in vivo confirmation of this mechanism.


Snakebite is Under Appreciated: Appraisal of Burden from West Africa.
Snakebite envenoming (SBE) is a major problem in rural areas of West Africa (WA). Compared to other Neglected Tropical Diseases (NTD), the public health burden of SBE has not been well characterized. We estimated the impact of snakebite mortality and morbidity using the Disability Adjusted Life Years (DALYs) metrics for 16 countries in WA. We used the reported annual number of SB deaths and mean age at time of SB and converted these into years of life lost (YLL). Similarly, the years of life lived with disability (YLD) were estimated by multiplying the number of amputations by the respective disability weight of 0.13. In WA, the annual cases of SB mortality and amputations ranged from 24 (95% Confidence Interval: 19-29) and 28 (17-48) respectively in Guinea-Bissau with the highest estimates of 1927 (1529-2333) and 2368 (1506-4043) respectively in Nigeria. We calculated that the annual DALYs associated with a SB death ranged from 1550 DALYs (95%CI: 1227-1873 DALYs) in Guinea Bissau to 124,484 DALYs (95%CI: 98,773-150,712 DALYs) in Nigeria. The annual DALYs associated with amputation for the two countries were 149 DALYs (95%CI: 91-256 DALYs) and 12,621 DALYs (95%CI: 8027-21,549 DALYs) respectively. The total burden of SBE was estimated at 319,874 DALYs (95% CI: 248,357-402,654 DALYs) in the 16 countries in WA. These estimates are similar, and in some instances even higher, than for other NTDs encountered in WA (e.g., Buruli ulcer, Echinococcosis, Intestinal Nematode Infections, Leishmaniasis, Onchocerchiasis, Trachoma and Trypanosomiasis) as reported in the Global Burden of Diseases 2010 (GBD). The public health burden of SBE in WA is very substantial and similar to other more widely recognized NTDs. Efforts and funding commensurate with its burden should be made available for the control of snakebite in the sub-region.


Thyroid Function in 509 Premature Newborns Below 31 Weeks of Gestational Age: Evaluation and Follow-up
Preterm and low birth weight (LBW) neonates may present with thyroid dysfunction during a critical period for neurodevelopment. These alterations can be missed on routine congenital hypothyroidism (CH) screening which only measures thyroid stimulating hormone (TSH). The objective of this study was to evaluate a protocol for thyroid function screening (TFS) six years after national implementation. Serum TSH and free thyroxine (fT4) were measured during the second week of life in neonates below 31 weeks. Patients with abnormal TFS (fT4 <0.8 ng/dL and/or TSH >5 mU/L) were followed up with repeated tests until normal levels were reported. Patients who were still on levothyroxine (LT4) at three years of age were re-evaluated. Five-hundred and nine neonates were included. Thyroid dysfunction was detected in 170 neonates (33%); CH n=20 (3.9%) including typical CH n=1; delayed TSH elevation CH n=19; hypothyroxinemia of prematurity (HOP) n=15 (2.9%); and transient hyperthyrotropinemia n=135 (26.5%). Twenty-one neonates (4.1%) were treated (20 for CH and 1 for HOP). At 3-year follow-up only three patients were diagnosed with permanent CH and still need treatment. LBW infants tended to have TSH levels higher than those with adequate weight. This protocol was able to detect thyroid dysfunction in preterm neonates who were not identified by the current program based on TSH determination in whole-blood. This thyroid dysfunction seems to resolve spontaneously in a few months in the great majority of neonates, but in some cases LT4 could be needed. There is a critical need for specific guidelines regarding the follow-up and re-evaluation of transient CH in preterm neonates.


Administration of the non-steroidal anti-inflammatory drug ibuprofen increases macrophage concentrations but reduces necrosis during modified muscle use.
To test the hypothesis that ibuprofen administration during modified muscle use reduces muscle necrosis and invasion by select myeloid cell populations. Rats were subjected to hindlimb unloading for 10 days, after which they experienced muscle reloading by normal weight-bearing to induce muscle inflammation and necrosis. Some animals received ibuprofen by intraperitoneal injection 8 h prior to the onset of muscle reloading, and then again at 8 and 16 h following the onset of reloading. Other animals received buffer injection at 8 h prior to reloading and then ibuprofen at 8 and 16 h following the onset of reloading. Control animals received buffer only at each time point. Quantitative immunohistochemical analysis was used to assess the presence of necrotic muscle fibers, total inflammatory infiltrate, neutrophils, ED1+ macrophages and ED2+ macrophages at 24 h following the onset of reloading. Administration of ibuprofen beginning 8 h prior to reloading caused significant reduction in the concentration of necrotic fibers, but increased the concentration of inflammatory cells in muscle. The increase in inflammatory cells was attributable to a 2.6-fold increase in the concentration of ED2+ macrophages. Animals treated with ibuprofen 8 h following the onset of reloading showed no decrease in muscle necrosis or increase in ED2+ macrophage concentrations. Administration of ibuprofen prior to increased muscle loading reduces muscle damage, but increases the concentration of macrophages that express the ED2 antigen. The increase in ED2+ macrophage concentration and decrease in necrosis may be mechanistically related because ED2+ macrophages have been associated with muscle regeneration and repair.


Spectral Properties and Excitation Relaxation of Novel Fucoxanthin Chlorophyll a/c-Binding Protein Complexes.
Fucoxanthin chlorophyll a/c-binding proteins (FCPs) are unique light harvesters for some photosynthetic organisms. There were several reports for the alterations of FCPs in response to light conditions. Here, we present the spectral profiles and excitation dynamics of novel FCP complexes isolated from the diatom Chaetoceros gracilis. Under a red-light condition, C. gracilis cells expressed three types of FCP complexes, two of which are very similar to FCP complexes found in the white-light grown cells, and one of which is the novel FCP complex. The combination of steady-state absorption and fluorescence spectra and time-resolved fluorescence spectra revealed that, compared to other types of FCP complexes, the novel FCP complexes exhibited red-shifted absorption and fluorescence spectra and fast decay of excitation. This finding will provide new insights into not only the light-harvesting strategies of diatoms but also the diversity of light adaptation machinery for photosynthetic organisms.


PCR-restriction fragment length polymorphism (RFLP) analyses reveal both extensive clonality and local genetic differences in Candida albicans.
Using a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method to obtain genotypes for the diploid pathogenic yeast, Candida albicans, we analysed 204 C. albicans isolates from three populations of the Duke University community: two from clinical sources [one from patients infected with human immunodeficiency virus (HIV) and the other from patients without HIV infection], and the third from healthy student volunteers. The results indicated: (i) extensive evidence for clonality within and between populations of C. albicans; and (ii) greater genotypic and gene diversities in the nonclinical population than those derived from clinical specimens, regardless of HIV status. The two clinical populations were genetically more similar to each other than either was to the population consisting of isolates from healthy people. Within each population sample there was a general lack of heterozygotes, and random associations of alleles within and between loci were found in less than 50% of the loci or pairs of loci. These findings were consistent between the two sets of samples analysed: those including all isolates and those including only clone-corrected isolates. Possible mechanisms are presented to explain the observed patterns of genetic variation within and between C. albicans populations.


Identification of hydroxyxanthones as Na/K-ATPase ligands.
We have screened a chemical library and identified several novel structures of Na/K-ATPase inhibitors. One group of these inhibitors belongs to polyphenolic xanthone derivatives. Functional characterization reveals the following properties of this group of inhibitors. First, like ouabain, they are potent inhibitors of the purified Na/K-ATPase. Second, their effects on the Na/K-ATPase depend on the number and position of phenolic groups. Methylation of these phenolic groups reduces the inhibitory effect. Third, further characterization of the most potent xanthone derivative, MB7 (3,4,5,6-tetrahydroxyxanthone), reveals that it does not change either Na(+) or ATP affinity of the enzyme. Finally, unlike that of ouabain, the inhibitory effect of MB7 on Na/K-ATPase is not antagonized by K(+). Moreover, MB7 does not activate the receptor Na/K-ATPase/Src complex and fails to stimulate protein kinase cascades in cultured cells. Thus, we have identified a group of novel Na/K-ATPase ligands that can inhibit the pumping function without stimulating the signaling function of Na/K-ATPase.


Molecularly imprinted polymer grafted graphene for simultaneous electrochemical sensing of 4, 4-methylene diphenylamine and aniline by differential pulse voltammetry.
A sensitive and selective electrochemical sensor based on molecular imprinting polymers grafted graphene (MIPs-G) was developed for simultaneous measurement of 4, 4-methylene diphenylamine (MDA) and aniline by differential pulse voltammetry (DPV). MIPs-G was synthesized via free radical polymerization reaction using MDA and 4-vinyl pyridine as template molecule and functional monomer respectively. The obtained nanocomposites were characterized by transmission electron microscope, scanning electron microscope, Fourier transform infrared spectrometry, Raman spectroscopy and thermal gravimetric analysis. After removal template molecules, MIPs-G was used as electrode material for aromatic amines measurements. Due to the high binding affinity towards MDA and the π-π interaction with aniline, the MIPs-G based electrochemical sensor can be used for simultaneous detection of MDA and aniline by DPV. Under the optimized condition, the oxidation potentials for MDA and aniline were observed at 0.62 and 0.72 V, respectively. Moreover, MIPs-G modified glassy carbon electrode shows linear response towards MDA and aniline simultaneously over the range of 1.0 to 15 μM. The MIPs-G based electrochemical sensor has been applied for simultaneous measurement of MDA and aniline in plastic tableware.


Desaturation during exercise is not a sufficient mechanism for prediction of osteoporosis in non-cystic fibrosis bronchiectasis.
Recent research has proposed an association between desaturation during a six minute walking test (6MWT) and osteoporosis in an elderly group of individuals with non-cystic fibrosis bronchiectasis. A causative pathway through activation of hypoxia-inducible factor 1-alpha (HIF-1α) has been proposed. Queries regarding the statistical approaches used are identified and discussed within this correspondence. These predominate around the use of linear regression models to predict osteoporosis in a group that is already osteoporotic, presenting with extreme values for bone mineral density (BMD). Further queries are raised regarding the HIF-1α pathway, and physical activity (PA) is proposed as an upstream mechanism for both reduced exercise tolerance and low BMD. It is suggested that osteoporosis cannot be predicted in a group that is already osteoporotic, and that PA is likely to be the causative mechanism between desaturation in the 6MWT and low BMD in non-cystic fibrosis bronchiectasis.