Sleep fragmentation impairs ventilatory long-term facilitation via adenosine A1 receptors.
Sleep fragmentation (SF), a primary feature of obstructive sleep apnoea (OSA), impairs hippocampal long-term potentiation and causes cognitive/attention deficits. However, its influence upon respiratory control has hardly been studied. This study examined the effect of SF on ventilatory long-term facilitation (LTF, a persistent augmentation of respiratory activity after episodic hypoxia) and the hypoxic ventilatory response (HVR), and investigated the role of adenosine A1 receptors in these SF effects in conscious adult male Sprague-Dawley rats. SF, confirmed by sleep architecture recordings, was achieved by periodic, forced locomotion in a rotating drum (30 s rotation/90 s stop for 24 h). LTF, elicited by five episodes of 5 min poikilocapnic hypoxia (10% O2) with 5 min intervals, was measured by plethysmography. Resting ventilation and metabolic rate were unchanged, HVR was reduced (150.6 +/- 3.5% versus 110.4 +/- 12.3%) and LTF was eliminated (22.6 +/- 0.5% versus -0.1 +/- 1.3%) shortly after 24 h SF. The SF-induced impairments were SF duration dependent, and completely reversible as HVR (< 24 h) and LTF (< 48 h) returned spontaneously to their pre-SF values. The SF-impaired HVR was improved (130.3 +/- 4.2%) and SF-eliminated LTF was restored (19.6 +/- 0.9%) by systemic injection of the adenosine A1 receptor antagonist 8-CPT (2.5 mg kg(-1)) approximately 30 min before LTF elicitation. Both HVR and LTF were also similarly impaired by 24 h total sleep deprivation or 24 h repeated cage tapping-induced SF, but not by a 24 h locomotion control protocol for SF. Collectively, these data suggest that: (1) 24 h SF impairs LTF and poikilocapnic HVR; (2) these impairments require A1 receptors; and (3) SF of OSA may exacerbate OSA via impaired ventilatory control mechanisms.


Contribution of N- and C-terminal Kv4.2 channel domains to KChIP interaction [corrected].
Association of Shal gene-related voltage-gated potassium (Kv4) channels with cytoplasmic Kv channel interacting proteins (KChIPs) influences inactivation gating and surface expression. We investigated both functional and biochemical consequences of mutations in cytoplasmic N and C-terminal Kv4.2 domains to characterize structural determinants for KChIP interaction. We performed a lysine-scanning mutagenesis within the proximal 40 amino acid portion and a structure-based mutagenesis in the tetramerization 1 (T1) domain of Kv4.2. In addition, the cytoplasmic Kv4.2 C-terminus was truncated at various positions. Wild-type and mutant Kv4.2 channels were coexpressed with KChIP2 isoforms in mammalian cell lines. The KChIP2-induced modulation of Kv4.2 currents was studied with whole-cell patch clamp and the binding of KChIP2 isoforms to Kv4.2 channels with coimmunoprecipitation experiments. Our results define one major interaction site for KChIPs, including amino acids in the proximal N-terminus between residues 11 and 23, where binding and functional modulation are essentially equivalent. A further interaction site includes residues in the T1 domain. Notably, C-terminal deletions also had marked effects on KChIP2-dependent gating modulation and KChIP2 binding, revealing a previously unknown involvement of domains within the cytoplasmic Kv4.2 C-terminus in KChIP interaction. Less coincidence of binding and functional modulation indicates a more loose 'anchoring' at T1- and C-terminal interaction sites. Our results refine and extend previously proposed structural models for Kv4.2/KChIP complex formation.


Inhibition of glioblastoma malignancy by Lgl1.
lethal giant larvae (lgl) was first identified as a tumor suppressor in Drosophila, where its loss repressed the differentiation and promoted the invasion of neuroblasts, the Drosophila equivalent of the neural stem cell. Recently we have shown that a human homolog of Lgl, Lgl1 (LLGL1), is constitutively phosphorylated and inactivated in glioblastoma cells; this occurs as a downstream consequence of PTEN loss, one of the most frequent genetic events in glioblastoma. Here we have investigated the consequences of this loss of functional Lgl1 in glioblastoma in vivo. We used a doxycycline-inducible system to express a non-phosphorylatable, constitutively active version of Lgl1 (Lgl3SA) in either a glioblastoma cell line or primary glioblastoma cells isolated under neural stem cell culture conditions from patients. In both types of cells, expression of Lgl3SA, but not wild type Lgl1, inhibited cell motility in vitro. Induction of Lgl3SA in intracerebral xenografts markedly reduced the in vivo invasion of primary glioblastoma cells. Lgl3SA expression also induced the differentiation of glioblastoma cells in vitro and in vivo along the neuronal lineage. Thus the central features of Lgl function as a tumor suppressor in Drosophila are conserved in human glioblastoma.


Immunohistochemical analysis of angiotensin converting enzyme in Sardinian pterygium.
Pterygium is a common ocular surface disorder characterized by excessive cell proliferation, inflammation, fibrosis, angiogenesis and extracellular matrix remodeling. The Angiotensin converting enzyme (ACE or ACE I) is the major component of the Renin-angiotensin system (RAS) converting the inactive decapeptide Angiotensin I (Ang I) to the active octapeptide Angiotensin II (Ang II). Besides this 'classical role', it can act as transcriptional regulator in response to external stimuli that may lead to cell damage and tissue remodeling. Due to this role, it can be internalized into the nuclear compartment to act as transcriptional factor for proteins involved in the inflammatory response. The aim of the present study was to determine ACE expression and localization in pterygium and culture pterygium cells by immunohistochemistry. Our results are the first to demonstrate nuclear immunolocalization of ACE, more so in pterygium compared to conjunctiva epithelial cells in histological sections. ACE was not detected in the nuclei of subcultivated pterygium epithelial cells. The nuclear localization of ACE may be correlated with an anti-inflammatory path mediated by activation of its transcriptional role.


Arthroscopic bankart suture-anchor repair: radiological and clinical outcome at minimum 10 years of follow-up.
There are not many reports in the literature about the long-term outcomes in terms of recurrence and degenerative changes after arthroscopic capsulolabral reconstruction for anterior shoulder instability. The aim of this study was to evaluate long-term follow-up (minimum 10 years) of arthroscopic suture-anchor repair for traumatic unidirectional anterior instability, with special emphasis on the radiological evidence of arthritis and clinical outcome. Case series; Level of evidence, 4. Forty-two patients (43 shoulders) treated at our institute from 1995 to 1997 were included in the study. Thirty patients (31 shoulders) were available for clinical and radiological examination (71%). The mean follow-up was 10.9 years (range, 9.8-14.3 years). Patients were evaluated preoperatively and after surgery using the University of California, Los Angeles (UCLA), Simple Shoulder Test (SST), and Rowe score. Patient satisfaction was determined by asking the patients if they would do this operation again. Radiological outcome was used to evaluate the incidence and grade of arthritis according to the Samilson-Prieto classification. At the final follow-up examination, 5 patients (16%) reported an atraumatic recurrent instability, while 2 recurrences (7%) occurred after a major injury. Three of the 7 recurrences occurred 6 years after surgery. All of the patients in the recurrence group except 1 were contact or overhead athletes. Twenty-six patients were satisfied (84%) with the outcome. The SST showed an improvement of shoulder function in 23 cases, the UCLA score improved from 21.8 to 32.1, and the Rowe score showed excellent or good results in 77.3% of cases. Twenty-two patients (71%) were able to return to their preoperative sports level. Radiographic findings showed 9 cases with mild arthritis (29%) and 3 cases with moderate arthritis (10%). The recurrence rate deteriorated with time. Involvement in contact sports and overhead activities appears to be a risk factor for recurrence of instability, although this could not be proved statistically with the numbers available, whereas age, gender, and number of preoperative dislocations did not reveal any correlation with recurrence. Degenerative changes of the glenohumeral joint were noted but had no significant effect on the clinical outcomes.


Adaptive beamforming for photoacoustic imaging.
An adaptive photoacoustic image reconstruction technique that combines coherence factor (CF) weighting and the minimum variance (MV) method is introduced. The backprojection method is widely used to reconstruct photoacoustic tomography images. Owing to the scattering of light, the quality of the photoacoustic imaging can be degraded. CF, an adaptive weighting technique, is known to improve the lateral resolution of photoacoustic images. In addition, an MV adaptive beamforming method can further improve the image quality by suppressing signals from off-axis directions. Experimental studies are performed to quantify the spatial resolution and contrast of the adaptive photoacoustic beamforming methods.


Ambient Particulate Matter Air Pollution Exposure and Mortality in the NIH-AARP Diet and Health Cohort.
Outdoor fine particulate matter (≤ 2.5 μm; PM2.5) has been identified as a global health threat, but the number of large U.S. prospective cohort studies with individual participant data remains limited, especially at lower recent exposures. We aimed to test the relationship between long-term exposure PM2.5 and death risk from all nonaccidental causes, cardiovascular (CVD), and respiratory diseases in 517,041 men and women enrolled in the National Institutes of Health-AARP cohort. Individual participant data were linked with residence PM2.5 exposure estimates across the continental United States for a 2000-2009 follow-up period when matching census tract-level PM2.5 exposure data were available. Participants enrolled ranged from 50 to 71 years of age, residing in six U.S. states and two cities. Cox proportional hazard models yielded hazard ratio (HR) estimates per 10 μg/m3 of PM2.5 exposure. PM2.5 exposure was significantly associated with total mortality (HR = 1.03; 95% CI: 1.00, 1.05) and CVD mortality (HR = 1.10; 95% CI: 1.05, 1.15), but the association with respiratory mortality was not statistically significant (HR = 1.05; 95% CI: 0.98, 1.13). A significant association was found with respiratory mortality only among never smokers (HR = 1.27; 95% CI: 1.03, 1.56). Associations with 10-μg/m3 PM2.5 exposures in yearly participant residential annual mean, or in metropolitan area-wide mean, were consistent with baseline exposure model results. Associations with PM2.5 were similar when adjusted for ozone exposures. Analyses of California residents alone also yielded statistically significant PM2.5 mortality HRs for total and CVD mortality. Long-term exposure to PM2.5 air pollution was associated with an increased risk of total and CVD mortality, providing an independent test of the PM2.5-mortality relationship in a new large U.S. prospective cohort experiencing lower post-2000 PM2.5 exposure levels. Thurston GD, Ahn J, Cromar KR, Shao Y, Reynolds HR, Jerrett M, Lim CC, Shanley R, Park Y, Hayes RB. 2016. Ambient particulate matter air pollution exposure and mortality in the NIH-AARP Diet and Health cohort. Environ Health Perspect 124:484-490; http://dx.doi.org/10.1289/ehp.1509676.


Spatial language facilitates spatial cognition: evidence from children who lack language input.
Does spatial language influence how people think about space? To address this question, we observed children who did not know a conventional language, and tested their performance on nonlinguistic spatial tasks. We studied deaf children living in Istanbul whose hearing losses prevented them from acquiring speech and whose hearing parents had not exposed them to sign. Lacking a conventional language, the children used gestures, called homesigns, to communicate. In Study 1, we asked whether homesigners used gesture to convey spatial relations, and found that they did not. In Study 2, we tested a new group of homesigners on a Spatial Mapping Task, and found that they performed significantly worse than hearing Turkish children who were matched to the deaf children on another cognitive task. The absence of spatial language thus went hand-in-hand with poor performance on the nonlinguistic spatial task, pointing to the importance of spatial language in thinking about space.


The early use of Antibiotics for at Risk CHildren with InfluEnza-like illness (ARCHIE): a double-blind randomised placebo-controlled trial.
The UK government stockpiles co-amoxiclav to treat bacterial complications during influenza pandemics. This pragmatic trial examines whether early co-amoxiclav use reduces reconsultation due to clinical deterioration in "at risk" children presenting with influenza-like illness (ILI) in primary or ambulatory care. "At risk" children aged from 6 months to 12 years presenting within 5 days of ILI onset were randomly assigned to oral co-amoxiclav 400/57 or a placebo twice daily for 5 days (dosing based on age±weight). "At risk" groups included children with respiratory, cardiac and neurological conditions. Randomisation was stratified by region and used a non-deterministic minimisation algorithm to balance age and current seasonal influenza vaccination status. Our target sample size was 650 children which would have allowed us to detect a reduction in the proportion of children reconsulting due to clinical deterioration from 40% to 26%, with 90% power and 5% two-tailed alpha error (including allowance for 25% loss to follow-up and an inflation factor of 1.041). Participants, caregivers and investigators were blinded to treatment allocation. Intention-to-treat analysis included all randomised participants with primary outcome data on reconsultation due to clinical deterioration within 28 days. Safety analysis included all randomised participants. ISRCTN 70714783. EudraCT 2013-002822-21. We recruited 271 children between February 11, 2015 and April 20, 2018. Primary outcome data were available for 265 children. Only 61 out of 265 children (23.0%) reconsulted due to clinical deterioration. No evidence of a treatment effect was observed for reconsultation due to clinical deterioration (33 out of 133 for co-amoxiclav (24.8%) and 28 out of 132 (21.2%) for placebo; adjusted risk ratio (RR) 1.16, 95% confidence interval (CI) 0.75-1.80). There was also no evidence of a difference between groups in the proportion of children for whom one or more adverse events (AEs) were reported (32 out of 136 (23.5%) for co-amoxiclav and 22 out of 135 (16.3%) for placebo; adjusted RR 1.45, 95% CI 0.90-2.34). In total, 66 AEs were reported (co-amoxiclav, n=37; placebo, n=29). Nine serious AEs were reported per group, although none were considered related to study medication. Our trial did not find evidence that treatment with co-amoxiclav reduces risk of reconsultation due to clinical deterioration in "at risk" children who present early with ILI during influenza season. Our findings therefore do not support early co-amoxiclav use in children with seasonal ILI.


Restoration of intracortical and thalamocortical circuits after transplantation of bone marrow mesenchymal stem cells into the ischemic brain of mice.
Transplantation of bone marrow mesenchymal stem cells (BMSCs) provides a promising regenerative medicine for stroke. Whether BMSC therapy could repair ischemia-damaged neuronal circuits and recover electrophysiological activity has largely been unknown. To address this issue, BMSCs were implanted into the ischemic barrel cortex of adult mice 1 and 7 days after focal barrel cortex stroke. Two days after the first transplantation (3 days after stroke), the infarct volume determined by TTC staining was significantly smaller in BMSC-treated compared to vehicle-treated stroke mice. The behavioral corner test showed better long-term recovery of sensorimotor function in BMSC-treated mice. Six weeks poststroke, thalamocortical slices were prepared and neuronal circuit activity in the peri-infarct region of the barrel cortex was determined by extracellular recordings of evoked field potentials. In BMSC-transplanted brain slices, the ischemia-disrupted intracortical activity from layer 4 to layer 2/3 was noticeably recovered, and the thalamocortical circuit connection was also partially restored. In contrast, much less and slower recovery was seen in control animals of barrel cortex stroke. Immunohistochemical staining disclosed that the density of neurons, axons, and blood vessels in the peri-infarct region was significantly higher in BMSC-treated mice, accompanied with enhanced local blood flow recovery. Western blotting showed that BMSC treatment increased the expression of stromal cell-derived factor-1 (SDF-1), vascular endothelial growth factor (VEGF), and brain-derived neurotrophic factor (BDNF) in the peri-infarct region. Moreover, the expression of the axonal growth associated protein-43 (GAP-43) was markedly increased, whereas the axonal growth inhibiting proteins ROCK II and NG2 were suppressed in the BMSC-treated brains. BMSC transplantation also promoted directional migration and survival of doublecortin (DCX)-positive neuroblasts in the peri-infarct region. The present investigation thus provides novel evidence that BMSC transplantation has the potential to repair the ischemia-damaged neural networks and restore lost neuronal connections. The recovered circuit activity likely contributes to the improved sensorimotor function after focal ischemic stroke and BMSC transplantation.


Comprehensive analysis of hormone and genetic variation in 36 genes related to steroid hormone metabolism in pre- and postmenopausal women from the breast and prostate cancer cohort consortium (BPC3).
Sex steroids play a central role in breast cancer development. This study aimed to relate polymorphic variants in 36 candidate genes in the sex steroid pathway to serum concentrations of sex steroid hormones and SHBG. Data on 700 genetic polymorphisms were combined with existing hormone assays and data on breast cancer incidence, within the European Prospective Investigation into Cancer and Nutrition (EPIC) and the Nurses' Health Study (NHS) cohorts; significant findings were reanalyzed in the Multiethnic Cohort (MEC). We analyzed data from a pooled sample of 3852 pre- and postmenopausal Caucasian women from EPIC and NHS and 454 postmenopausal women from MEC. Outcome measures were SHBG, testosterone, dehydroepiandrosterone (DHEAS), androstenedione, estrone (E1), and estradiol (E2) as well as breast cancer risk. Globally significant associations were found among pre- and postmenopausal women combined between levels of SHBG and the SHBG gene and between DHEAS and the FSHR and AKR1C3 genes. Among postmenopausal women, serum E1 and E2 were significantly associated with the genes CYP19 and FSHR, and E1 was associated with ESR1. None of the variants related to serum hormone levels showed any significant association with breast cancer risk. We confirmed associations between serum levels of SHBG and the SHBG gene and of E1 and E2 and the CYP19 and ESR1 genes. Novel associations were observed between FSHR and DHEAS, E1, and E2 and between AKR1C3 and DHEAS.


Cell penetrating peptides for in vivo molecular imaging applications.
Cell penetrating peptides (CPPs) are a relatively new class of peptides that have the promising capability to cross cell membranes. While details remain to be resolved, various non-receptor-mediated endocytic pathways likely contribute most to the cell penetrating properties of these peptides. CPPs have been used to deliver many different cargos - ranging from radionuclides and other peptides to antibodies and nanoparticles - into cells. Besides many different drug delivery applications, CPPs have also seen a limited use in molecular imaging. Molecular imaging of intracellular and intranuclear targets, by techniques such as SPECT, PET, optical imaging, and MRI, relies heavily on the delivery of contrast agents to the cytoplasm and/or nuclei of the target tissue. Therefore, the number of applications in molecular imaging of intracellular targets has remained relatively low, because of the effective barrier presented by the cell membrane. One of the key strategies to overcome this challenge is the introduction of membrane-transducing peptides in the design of new contrast agents. This review presents an overview of the literature on CPPs, focusing on their use for molecular imaging. Applications using proteins and peptides, DNA/RNA, and CPP-loaded cells as the imaging agents will be looked at. Moreover, the difficulties and pitfalls regarding the use of CPPs in molecular imaging will be discussed.


Developmental regulation of polyglutamylated alpha- and beta-tubulin in mouse brain neurons.
Polyglutamylation is an important posttranslational modification of tubulin that is very active in nerve cells, where it accounts for the main factor responsible for tubulin heterogeneity. In the present work, we have analyzed quantitative and qualitative changes in glutamylated alpha- and beta-tubulin occurring during neuronal differentiation in culture. Glutamylated alpha- and beta-tubulin both markedly accumulate during this process with a time course remarkably similar to that observed in vivo during brain development. However, the characteristics of the glutamylation of the two subunits are not exactly the same. Glutamylated alpha-tubulin is already abundant in very young neurons and displays, at this stage, a wide range of its degree of glutamylation (1 to 6 glutamyl units present in the lateral polyglutamyl chain), which remains unchanged during the entire period of the culture. Glutamylated beta-tubulin is present at very low levels in young neurons and its accumulation during differentiation is accompanied by a progressive increase in its degree of glutamylation from 2 to 6 glutamyl units. Posttranslational incorporation of [3H]glutamate into alpha- and beta-tubulin decreases during differentiation, as well as the rate of the reverse deglutamylation reaction, suggesting that accumulation of glutamylated tubulin is accompanied by a decrease in the turnover of glutamyl units onto tubulin. Neuronal differentiation is also accompanied by an increase of other posttranslationally modified forms of tubulin, including acetylated and non-tyrosinatable alpha-tubulin, which can occur in combination with polyglutamylation and contributes to increase the complexity of tubulin in mature neurons.


Contrasting benefits of different artemisinin combination therapies as first-line malaria treatments using model-based cost-effectiveness analysis.
There are currently several recommended drug regimens for uncomplicated falciparum malaria in Africa. Each has different properties that determine its impact on disease burden. Two major antimalarial policy options are artemether-lumefantrine (AL) and dihydroartemisinin-piperaquine (DHA-PQP). Clinical trial data show that DHA-PQP provides longer protection against reinfection, while AL is better at reducing patient infectiousness. Here we incorporate pharmacokinetic-pharmacodynamic factors, transmission-reducing effects and cost into a mathematical model and simulate malaria transmission and treatment in Africa, using geographically explicit data on transmission intensity and seasonality, population density, treatment access and outpatient costs. DHA-PQP has a modestly higher estimated impact than AL in 64% of the population at risk. Given current higher cost estimates for DHA-PQP, there is a slightly greater cost per case averted, except in areas with high, seasonally varying transmission where the impact is particularly large. We find that a locally optimized treatment policy can be highly cost effective for reducing clinical malaria burden.


[Analysis of 15 cases of avian influenza virus (H7N9) infection].
Objective: To describe the clinical, chest imaging, pathological manifestations and therapeutic experience of human infection with A/H7N9 virus. Methods: The features of 15 laboratory-confirmed cases of human infection with A/H7N9 virus in Taizhou, Jiangsu Province were retrospectively analyzed. Results: The 15 patients with confirmed viral pneumonia included 12 males and 3 females, with a median age of 61 years(ranging from 33 to 81 years). Twelve patients had a history of exposure to the poultry trading places, or direct contact with ill/dead avian, while 3 patients denied exposure or contact. The most common initial symptoms were fever, coughing, and respiratory distress. The illness progressed rapidly to acute respiratory distress syndrome (ARDS). Lab tests showed normal (8 cases) or decreased (7 cases)white blood cell count , decreased (13 cases) lymphocyte count and proportion , increased creatine kinase (CK, 12 cases) and lactate dehydrogenase (LDH, 15 cases), and respiratory failure (13 cases). Chest radiographic examination showed that the most common features were inflammatory infiltration in the lung, with partial consolidation. The average time of the diagnosis with influenza viral nucleic acid and onset of an oral anti-influenza drug were 7.1 days and 6.5 days. All patients were treated by antiviral drugs (oral oseltamivir 150 mg q12 h and/or intravenous paramivir 600 mg qd), with mechanical ventilation in 9 cases, glucocorticoid therapy in 5 cases (intravenous methylprednisolone in 3 and dexamethasone in 2 patients), extracorporeal membrane oxygenation (ECMO) therapy in 2 cases, continuous renal replacement therapy (CRRT) in 6 cases, and artificial liver therapy in 1 case. The pulmonary pathology was observed from post-mortem biopsy for 2 fatal cases. Patient 1 had diffuse alveolar damage with inflammatory exudation, hyaline membrane formation, and cellular infiltration. Patient 2 had widened alveolar septum, lymphocyte and monocyte cell infiltration in the alveolar septa, and interstitial fibrous proliferation. Nine patients were discharged, and 6 died. Conclusions: Patients with influenza A/H7N9 virus mostly presented with fever, cough, and were prone to progression to viral pneumonia. Once acute respiratory distress and important organ dysfunction occurred, the fatality rate was higher. Early diagnosis and rational treatment were critical for better outcomes.


High-power femtosecond-terahertz pulse induces a wound response in mouse skin.
Terahertz (THz) technology has emerged for biomedical applications such as scanning, molecular spectroscopy, and medical imaging. Although a thorough assessment to predict potential concerns has to precede before practical utilization of THz source, the biological effect of THz radiation is not yet fully understood with scant related investigations. Here, we applied a femtosecond-terahertz (fs-THz) pulse to mouse skin to evaluate non-thermal effects of THz radiation. Analysis of the genome-wide expression profile in fs-THz-irradiated skin indicated that wound responses were predominantly mediated by transforming growth factor-beta (TGF-β) signaling pathways. We validated NFκB1- and Smad3/4-mediated transcriptional activation in fs-THz-irradiated skin by chromatin immunoprecipitation assay. Repeated fs-THz radiation delayed the closure of mouse skin punch wounds due to up-regulation of TGF-β. These findings suggest that fs-THz radiation initiate a wound-like signal in skin with increased expression of TGF-β and activation of its downstream target genes, which perturbs the wound healing process in vivo.


Pro: Can neuropathology really confirm the exact diagnosis?
Recent advances in the clinical diagnostic instruments for diagnosing Alzheimer's disease (AD) and in neuroimaging may cast doubt in the minds of some practitioners about the continued need for neuropathology to provide the ultimate diagnosis. Certainly the majority of cases of AD can be clinically correctly diagnosed by experienced clinicians but many cases are given this label by less experienced practitioners. Even after the most thorough work-up, a few cases of confidently diagnosed AD turn out to be something else when microscopy of the brain is undertaken. Even for neuropathologists, however, it can be difficult to correctly assign cognitive decline to the various pathological processes that can be found together in an older brain. We need further clinicopathogical study to enlighten us about, for example, the contribution of commonly found cerebrovascular disease to dementia. Human studies are also needed to explore the changes in pathology that new treatments for AD may produce.


Hepatocyte death: a clear and present danger.
The hepatocyte is especially vulnerable to injury due to its central role in xenobiotic metabolism including drugs and alcohol, participation in lipid and fatty acid metabolism, its unique role in the enterohepatic circulation of bile acids, the widespread prevalence of hepatotropic viruses, and its existence within a milieu of innate immune responding cells. Apoptosis and necrosis are the most widely recognized forms of hepatocyte cell death. The hepatocyte displays many unique features regarding cell death by apoptosis. It is quite susceptible to death receptor-mediated injury, and its death receptor signaling pathways involve the mitochondrial pathway for efficient cell killing. Also, death receptors can trigger lysosomal disruption in hepatocytes which further promote cell and tissue injury. Interestingly, hepatocytes are protected from cell death by only two anti-apoptotic proteins, Bcl-x(L) and Mcl-1, which have nonredundant functions. Endoplasmic reticulum stress or the unfolded protein response contributes to hepatocyte cell death during alterations of lipid and fatty acid metabolism. Finally, the current information implicating RIP kinases in necrosis provides an approach to more fully address this mode of cell death in hepatocyte injury. All of these processes contributing to hepatocyte injury are discussed in the context of potential therapeutic strategies.


The Role of Drug Transporters in the Pharmacokinetics of Antibiotics.
Various transporters, including efflux transporters and uptake transporters, play an important role in the pharmacokinetics of drugs. Currently, studies suggest that several antibiotics also serve as substrates for transporters. In addition, these antibiotics are usually combined with other drugs to treat diseases, more effectively. Therefore, it is necessary to focus on the role of transporters in pharmacokinetics and drug-drug interactions of antibiotics. This review summarizes the findings of recent studies as well as information retrieved from several databases (until June 2015): ISI Web of KnowledgeSM (ISI WoK), SciFinder (Caplus, Medline, Registry, Casreact, Chrmlist, and Chemcasts) and PubMed (indexed for Medline). The present review provides useful information for the study of transporters in the pharmacokinetics and drug-drug interactions of antibiotics, and should assist researchers investigating these topics. The drug transporters mediate intestinal absorption, hepatic uptake, and kidney or biliary excretion. It is necessary to focus on drug-drug interactions when these antibiotics are combined with other chemical substances that are also the substrates for transporters.


The economic burden of major adult visual disorders in the United States.
To estimate the societal economic burden and the governmental budgetary impact of the following visual disorders among US adults aged 40 years and older: visual impairment, blindness, refractive error, age-related macular degeneration, cataracts, diabetic retinopathy, and primary open-angle glaucoma. We estimated 3 components of economic burden: direct medical costs, other direct costs, and productivity losses. We used private insurance and Medicare claims data to estimate direct medical costs; epidemiologic evidence from multiple published sources to estimate other direct costs, such as nursing home costs; and data from the Survey of Income and Program Participation to estimate productivity losses. We used budgetary documents and our direct medical and other direct cost estimates to approximate the governmental budgetary impact. We estimated that the annual total financial burden of major adult visual disorders is $35.4 billion ($16.2 billion in direct medical costs, $11.1 billion in other direct costs, and $8 billion in productivity losses) and that the annual governmental budgetary impact is $13.7 billion. Major visual disorders among Americans older than 40 years result in substantial economic costs for the US economy. Well-designed public health programs may have the ability to reduce this burden in the future.


Associations between parental rules, style of communication and children's screen time.
Research suggests an inverse association between parental rules and screen time in pre-adolescents, and that parents' style of communication with their children is related to the children's time spent watching TV. The aims of this study were to examine associations of parental rules and parental style of communication with children's screen time and perceived excessive screen time in five European countries. UP4FUN was a multi-centre, cluster randomised controlled trial with pre- and post-test measurements in each of five countries; Belgium, Germany, Greece, Hungary and Norway. Questionnaires were completed by the children at school and the parent questionnaire was brought home. Three structural equation models were tested based on measures of screen time and parental style of communication from the pre-test questionnaires. Of the 152 schools invited, 62 (41 %) schools agreed to participate. In total 3325 children (average age 11.2 years and 51 % girls) and 3038 parents (81 % mothers) completed the pre-test questionnaire. The average TV/DVD times across the countries were between 1.5 and 1.8 h/day, while less time was used for computer/games console (0.9-1.4 h/day). The children's perceived parental style of communication was quite consistent for TV/DVD and computer/games console. The presence of rules was significantly associated with less time watching TV/DVD and use of computer/games console time. Moreover, the use of an autonomy-supportive style was negatively related to both time watching TV/DVD and use of computer/games console time. The use of a controlling style was related positively to perceived excessive time used on TV/DVD and excessive time used on computer/games console. With a few exceptions, results were similar across the five countries. This study suggests that an autonomy-supportive style of communicating rules for TV/DVD or computer/ games console use is negatively related to children's time watching TV/DVD and use of computer/games console time. In contrast, a controlling style is associated with more screen time and with more perceived excessive screen time in particular. Longitudinal research is needed to further examine effects of parental style of communication on children's screen time as well as possible reciprocal effects. International Standard Randomized Controlled Trial Number Register, registration number: ISRCTN34562078 . Date applied29/07/2011, Date assigned11/10/2011.


Steroids in the amniotic fluid of the rhesus monkey (Macaca mulatta).
Representative C21, C19 and C18 steroids have been measured in the amniotic fluid and maternal femoral plasma of rhesus monkeys during the last 50 days of pregnancy. The concentrations of cortisol, progesterone, androstenedione, oestrone sulphate all increased in amniotic fluid before spontaneous parturition. In contrast, amniotic fluid oestradiol levels remained low and unchanged, and were always less than the concentrations in the maternal plasma. The changes in progesterone, cortisol and oestrone concentrations in amniotic fluid occurred independently of significant changes in the concentration of these steroids in the maternal plasma. Betamethasone administered to monkeys after day 150 of pregnancy suppressed amniotic fluid and maternal plasma cortisol. This treatment markedly suppressed the concentration of progesterone in amniotic fluid, but did not alter its concentration in maternal plasma. It is concluded that during late pregnancy in the rhesus monkey there is an increase in the concentration of some steroids in amniotic fluid independent of alterations in their concentrations in maternal plasma. These changes could reflect an increase in foetal adrenal steroid secretion.


Probiotics (Lactobacillus rhamnosus R0011 and acidophilus R0052) reduce the expression of toll-like receptor 4 in mice with alcoholic liver disease.
The role of lipopolysaccharide (LPS) and toll-like receptor 4 (TLR 4) in the pathogenesis of alcoholic liver disease (ALD) has been widely established. We evaluated the biological effects of probiotics (Lactobacillus rhamnosus R0011 and acidophilus R0052), KRG (Korea red ginseng), and urushiol (Rhus verniciflua Stokes) on ALD, including their effects on normal and high-fat diet in mice. One hundred C57BL/6 mice were classified into normal (N) and high-fat diet (H) groups. Each group was divided into 5 sub-groups: control, alcohol, alcohol+probiotics, alcohol+KRG, and alcohol+urushiol. A liver function test, histology, electron-microscopy, interleukin (IL)-1β, tumor necrosis factor (TNF)-α, IL-6, and IL-10, and TLR 4 were evaluated and compared. In the N group, probiotics, KRG, and urushiol significantly reduced levels of TNF-α (12.3±5.1, 13.4±3.9, and 12.1±4.3 vs. 27.9±15.2 pg/mL) and IL-1β (108.4±39.4, 75.0±51.0, and 101.1±26.8 vs. 162.4±37.5 pg/mL), which were increased by alcohol. Alcohol-induced TLR 4 expression was reduced by probiotics and urushiol (0.7±0.2, and 0.8±0.1 vs. 1.0±0.3, p<0.001). In the H group, IL-10 was significantly increased by probiotics and KRG, compared with alcohol (25.3±15.6 and 20.4±6.2 vs. 7.6±5.6 pg/mL) and TLR 4 expression was reduced by probiotics (0.8±0.2 vs. 1.0±0.3, p = 0.007). Alcohol-induced TLR 4 expression was down-regulated by probiotics in the normal and high-fat diet groups. Probiotics, KRG, and urushiol might be effective in the treatment of ALD by regulating the gut-liver axis.


Long-Term Engraftment of Human Cardiomyocytes Combined with Biodegradable Microparticles Induces Heart Repair.
Cardiomyocytes derived from human induced pluripotent stem cells (hiPSC-CMs) are a promising cell source for cardiac repair after myocardial infarction (MI) because they offer several advantages such as potential to remuscularize infarcted tissue, integration in the host myocardium, and paracrine therapeutic effects. However, cell delivery issues have limited their potential application in clinical practice, showing poor survival and engraftment after transplantation. In this work, we hypothesized that the combination of hiPSC-CMs with microparticles (MPs) could enhance long-term cell survival and retention in the heart and consequently improve cardiac repair. CMs were obtained by differentiation of hiPSCs by small-molecule manipulation of the Wnt pathway and adhered to biomimetic poly(lactic-co-glycolic acid) MPs covered with collagen and poly(d-lysine). The potential of the system to support cell survival was analyzed in vitro, demonstrating a 1.70-fold and 1.99-fold increase in cell survival after 1 and 4 days, respectively. The efficacy of the system was tested in a mouse MI model. Interestingly, 2 months after administration, transplanted hiPSC-CMs could be detected in the peri-infarct area. These cells not only maintained the cardiac phenotype but also showed in vivo maturation and signs of electrical coupling. Importantly, cardiac function was significantly improved, which could be attributed to a paracrine effect of cells. These findings suggest that MPs represent an excellent platform for cell delivery in the field of cardiac repair, which could also be translated into an enhancement of the potential of cell-based therapies in other medical applications.


Does social support in addition to ART make a difference? Comparison of households with TASO and MOH PLWHA in Central Uganda.
Social support in addition to antiretroviral therapy (ART) has been indicated to be beneficial to person living with HIV/AIDS (PLWHA) and their families, but very few ART service providers go beyond ART. This study investigates whether receipt of social support in addition to ART for PLWHA makes the households that they reside in better off than households that have PLWHA but are without social support. The analysis uses data comprising of 450 households, which is a sub-sample from the 2010/2011 Centre for Health Economics Ugandan HIV Survey, a cross-sectional survey of 596 households that was undertaken in Uganda. Data were collected from households of clients that obtained ART from two major ART service providers in Central Uganda; The AIDS Support Organisation (TASO) and Ministry of Health (MOH), Uganda. Probit models and ordinary least squares regressions are employed to compare outcomes for individuals from households with a TASO or MOH client. Outcomes for individuals in households with a TASO PLWHA are hypothesised to be superior to those from households with an MOH PLWHA given that the benefits from social support accrue not only to the PLWHA but also to the household and communities they belong to. The results confirm that individuals from a household with a TASO PLWHA are better off in terms of physical health outcomes including better productivity as non-wage labour hours and having more cash in hand and having savings. The findings highlight the importance of additional support to HIV/AIDS clients and have implications for supplementation of ART service provision with other services to maximise the benefits from ART in resource constrained countries like Uganda.


Preclinical Evaluation of the Pan-FGFR Inhibitor LY2874455 in FRS2-Amplified Liposarcoma.
Background: FGFR inhibition has been proposed as treatment for dedifferentiated liposarcoma (DDLPS) with amplified FRS2, but we previously only demonstrated transient cytostatic effects when treating FRS2-amplified DDLPS cells with NVP-BGJ398. Methods: Effects of the more potent FGFR inhibitor LY2874455 were investigated in three DDLPS cell lines by measuring effects on cell growth and apoptosis in vitro and also testing efficacy in vivo. Genome, transcriptome and protein analyses were performed to characterize the signaling components in the FGFR pathway. Results: LY2874455 induced a stronger, longer-lasting growth inhibitory effect and moderate level of apoptosis for two cell lines. The third cell line, did not respond to FGFR inhibition, suggesting that FRS2 amplification alone is not sufficient to predict response. Importantly, efficacy of LY2874455 was confirmed in vivo, using an independent FRS2-amplified DDLPS xenograft model. Expression of FRS2 was similar in the responding and non-responding cell lines and we could not find any major difference in downstream FGFR signaling. The only FGF expressed by unstimulated non-responding cells was the intracellular ligand FGF11, whereas the responding cell lines expressed extracellular ligand FGF2. Conclusion: Our study supports LY2874455 as a better therapy than NVP-BGJ398 for FRS2-amplified liposarcoma, and a clinical trial is warranted.


Heterozygosity-fitness correlations and associative overdominance: new detection method and proof of principle in the Iberian wild boar.
Heterozygosity-fitness correlations (HFC) may result from a genome-wide process--inbreeding--or local effects within the genome. The majority of empirical studies reporting HFCs have attributed correlations to inbreeding depression. However, HFCs are unlikely to be caused by inbreeding depression because heterozygosity measured at a small number of neutral markers is unlikely to accurately capture a genome-wide pattern. Testing the strengths of localized effects caused by associative overdominance has proven challenging. In their current paper, Amos and Acevedo-Whitehouse present a novel test for local HFCs. Using stochastic simulations, they determine the conditions under which single-locus HFCs arise, before testing the strength of the correlation between the neutral marker and a linked gene under selection in their simulations. They used insights gained from simulation to statistically investigate the likely cause of correlations between heterozygosity and disease status using data on bovine tuberculosis infections in a wild boar population. They discover that a single microsatellite marker is an excellent predictor of tuberculosis progression in infected individuals. The results are relevant for wild boar management but, more generally, they demonstrate how single-locus HFCs could be used to identify coding loci under selection in free-living populations.


Comparing thousands of circular genomes using the CGView Comparison Tool.
Continued sequencing efforts coupled with advances in sequencing technology will lead to the completion of a vast number of small genomes. Whole-genome comparisons represent an important part of the analysis of any new genome sequence, as they can provide a better understanding of the biology and evolution of the source organism. Visualization of the results is important, as it allows information from a variety of sources to be integrated and interpreted. However, existing graphical comparison tools lack features needed for efficiently comparing a new genome to hundreds or thousands of existing sequences. Moreover, existing tools are limited in terms of the types of comparisons that can be performed, the extent to which the output can be customized, and the ease with which the entire process can be automated. The CGView Comparison Tool (CCT) is a package for visually comparing bacterial, plasmid, chloroplast, or mitochondrial sequences of interest to existing genomes or sequence collections. The comparisons are conducted using BLAST, and the BLAST results are presented in the form of graphical maps that can also show sequence features, gene and protein names, COG (Clusters of Orthologous Groups of proteins) category assignments, and sequence composition characteristics. CCT can generate maps in a variety of sizes, including 400 Megapixel maps suitable for posters. Comparisons can be conducted within a particular species or genus, or all available genomes can be used. The entire map creation process, from downloading sequences to redrawing zoomed maps, can be completed easily using scripts included with the CCT. User-defined features or analysis results can be included on maps, and maps can be extensively customized. To simplify program setup, a CCT virtual machine that includes all dependencies preinstalled is available. Detailed tutorials illustrating the use of CCT are included with the CCT documentation. CCT can be used to visually compare a reference sequence to thousands of existing genomes or sequence collections (next-generation sequencing reads for example) on a standard desktop computer. It provides analysis and visualization functionality not available in any existing circular genome visualization tool. By visually presenting sequence conservation information along with functional classifications and sequence composition characteristics, CCT can be a useful tool for identifying rapidly evolving or novel sequences, horizontally transferred sequences, or unusual functional properties in newly sequenced genomes. CCT is freely available for download at http://stothard.afns.ualberta.ca/downloads/CCT/.


Human airway musculature on a chip: an in vitro model of allergic asthmatic bronchoconstriction and bronchodilation.
Many potential new asthma therapies that show promise in the pre-clinical stage of drug development do not demonstrate efficacy during clinical trials. One factor contributing to this problem is the lack of human-relevant models of the airway that recapitulate the tissue-level structural and functional phenotypes of asthma. Hence, we sought to build a model of a human airway musculature on a chip that simulates healthy and asthmatic bronchoconstriction and bronchodilation in vitro by engineering anisotropic, laminar bronchial smooth muscle tissue on elastomeric thin films. In response to a cholinergic agonist, the muscle layer contracts and induces thin film bending, which serves as an in vitro analogue for bronchoconstriction. To mimic asthmatic inflammation, we exposed the engineered tissues to interleukin-13, which resulted in hypercontractility and altered relaxation in response to cholinergic challenge, similar to responses observed clinically in asthmatic patients as well as in studies with animal tissue. Moreover, we reversed asthmatic hypercontraction using a muscarinic antagonist and a β-agonist which are used clinically to relax constricted airways. Importantly, we demonstrated that targeting RhoA-mediated contraction using HA1077 decreased basal tone, prevented hypercontraction, and improved relaxation of the engineered tissues exposed to IL-13. These data suggest that we can recapitulate the structural and functional hallmarks of human asthmatic musculature on a chip, including responses to drug treatments for evaluation of safety and efficacy of new drugs. Further, our airway musculature on a chip provides an important tool for enabling mechanism-based search for new therapeutic targets through the ability to evaluate engineered muscle at the levels of protein expression, tissue structure, and tissue function.


Discourse, syntax, and prosody: the brain reveals an immediate interaction.
Speech is structured into parts by syntactic and prosodic breaks. In locally syntactic ambiguous sentences, the detection of a syntactic break necessarily follows detection of a corresponding prosodic break, making an investigation of the immediate interplay of syntactic and prosodic information impossible when studying sentences in isolation. This problem can be solved, however, by embedding sentences in a discourse context that induces the expectation of either the presence or the absence of a syntactic break right at a prosodic break. Event-related potentials (ERPs) were compared to acoustically identical sentences in these different contexts. We found in two experiments that the closure positive shift, an ERP component known to be elicited by prosodic breaks, was reduced in size when a prosodic break was aligned with a syntactic break. These results establish that the brain matches prosodic information against syntactic information immediately.


Innate immunity, epigenetics and autoimmunity in rheumatoid arthritis.
Rheumatoid arthritis (RA) is a chronic inflammatory autoimmune disease characterized by the progressive and irreversible destruction of joints. RA remains an incurable condition, although a new class of drugs, biologicals, have made a major breakthrough in targeting and/or eliminating the immune cells, including T cells, B cells and monocytes/macrophages from the joints. That we cannot (yet?) cure the disease is most likely due to the lack of therapeutic targeting the endogenously activated RA synovial fibroblasts (RASF). Most interestingly, RASF express Toll-like receptors (TLRs) 1-6 rendering them prone to activation by exogenous and endogenous TLR ligands and resulting in the production of numerous powerful chemokines and cytokines. These factors are responsible for the repopulation of immune cells in the joints after ceasing cell depleting therapies. To characterize the molecular mechanisms of synovial activation, a new approach studying the epigenetic characteristics of RASF has been recently undertaken. Thereby, the pattern of histone acetylation, DNA methylation and gene expression regulating microRNA are being explored. Since auto-antibodies have the most predictive and diagnostic value for RA, it is challenging to study more comprehensively the contribution of auto-antibodies to the disease. A new screening technique, serological analysis of recombinant human cDNA expression library (SEREX), adapted from cancer research allowed for the identification of novel auto-antibodies in RA, including anti-serpin E2 auto-antibodies. The serpin E2 auto-antibodies were found to inhibit the activity of serpin E2 and have potentially a functional role in the disease. The recent findings in the field of innate immunity, epigenetics and autoimmunity related to the pathogenesis of RA are in the scope of this review.


Closed-loop real-time simulation model of hemodynamics and oxygen transport in the cardiovascular system.
Computer technology enables realistic simulation of cardiovascular physiology. The increasing number of clinical surgical and medical treatment options imposes a need for better understanding of patient-specific pathology and outcome prediction. A distributed lumped parameter real-time closed-loop model with 26 vascular segments, cardiac modelling with time-varying elastance functions and gradually opening and closing valves, the pericardium, intrathoracic pressure, the atrial and ventricular septum, various pathological states and including oxygen transport has been developed. Model output is pressure, volume, flow and oxygen saturation from every cardiac and vascular compartment. The model produces relevant clinical output and validation of quantitative data in normal physiology and qualitative directions in simulation of pathological states show good agreement with published data. The results show that it is possible to build a clinically relevant real-time computer simulation model of the normal adult cardiovascular system. It is suggested that understanding qualitative interaction between physiological parameters in health and disease may be improved by using the model, although further model development and validation is needed for quantitative patient-specific outcome prediction.


Benefits and Harms in Pivotal Trials of Oral Centrally Acting Antiobesity Medicines: A Systematic Review and Meta-Analysis.
To evaluate the benefits and harms of oral centrally acting antiobesity medicinal products in pivotal trials. The European Medicines Agency and Federal Drug Administration websites, PubMed, and ClinicalTrials.gov were searched to identify pivotal trials used to gain marketing authorizations. Pivotal phase III trials on which marketing authorizations were based were included. The data were analyzed by using Cochrane Review Manager (RevMan), and quality assessments for each outcome were performed by using the Grading of Recommendation, Assessment, Development, and Evaluation (GRADE). Five products (16 trials with 24,555 participants) were included. Significantly more participants who took the antiobesity products achieved ≥ 5% reduction in body weight (risk ratio [RR] 2.39; 95% CI: 2.09-2.74; GRADE = low). However, the products significantly increased the risk of adverse events (RR 1.12; 95% CI: 1.07-1.17; GRADE = very low) and the risk of discontinuation because of adverse events (RR 1.52; 95% CI: 1.33-1.74; GRADE = low). There were no significant differences for most outcomes between currently approved and withdrawn products. Although oral centrally acting antiobesity products generate modest weight losses, they also increase the risks of adverse events and discontinuations because of adverse events. The premarketing benefit-to-harm profiles of currently available products and products that were later withdrawn because of harms are similar. Targeted study designs, better outcomes reporting, and improved postmarketing monitoring of harms are needed.


Overdrainage shunt complications in idiopathic normal-pressure hydrocephalus and lumbar puncture opening pressure.
Management of idiopathic normal-pressure hydrocephalus (iNPH) is hard because the diagnosis is difficult and shunt surgery has high complication rates. An important complication is overdrainage, which often can be treated with adjustable-shunt valve manipulations but also may result in the need for subdural hematoma evacuation. The authors evaluated shunt surgery overdrainage complications in iNPH and their relationship to lumbar puncture opening pressure (LPOP). The authors reviewed the charts of 164 consecutive patients with iNPH who underwent shunt surgery at their institution from 2005 to 2011. They noted age, sex, presenting symptoms, symptom duration, hypertension, body mass index (BMI), imaging findings of atrophy, white matter changes, entrapped sulci, LPOP, valve opening pressure (VOP) setting, number of valve adjustments, serious overdrainage (subdural hematoma requiring surgery), radiological overdrainage (subdural hematomas or hygroma seen on postoperative imaging), clinical overdrainage (sustained or postural headache), other complications, and improvements in gait, urine control, and memory. Eight patients (5%) developed subdural hematomas requiring surgery. All had an LPOP of greater than 160 mm H2O and an LPOP-VOP of greater than 40 mm H2O. Radiological overdrainage was more common in those with an LPOP of greater than 160 mm H2O than in those with an LPOP of less than 160 mm H2O (38% vs. 21%, respectively; p = 0.024). The BMI was also significantly higher in those with an LPOP of greater than 160 mm H2O (median 30.2 vs. 27.0, respectively; p = 0.005). Serious overdrainage that caused subdural hematomas and also required surgery after shunting was related to LPOP and LPOP-VOP, which in turn were related to BMI. If this can be replicated, individuals with a high LPOP should have their VOP set close to the LPOP, or even higher. In doing this, perhaps overdrainage complications can be reduced.


N-acetylcysteine increases the frequency of bone marrow pro-B/pre-B cells, but does not reverse cigarette smoking-induced loss of this subset.
We previously showed that mice exposed to cigarette smoke for three weeks exhibit loss of bone marrow B cells at the Pro-B-to-pre-B cell transition, but the reason for this is unclear. The antioxidant N-acetylcysteine (NAC), a glutathione precursor, has been used as a chemopreventive agent to reduce adverse effects of cigarette smoke exposure on lung function. Here we determined whether smoke exposure impairs B cell development by inducing cell cycle arrest or apoptosis, and whether NAC treatment prevents smoking-induced loss of developing B cells. Groups of normal mice were either exposed to filtered room air or cigarette smoke with or without concomitant NAC treatment for 5 days/week for three weeks. Bone marrow B cell developmental subsets were enumerated, and sorted pro-B (B220(+)CD43(+)) and pre-B (B220(+)CD43(-)) cell fractions were analyzed for cell cycle status and the percentage of apoptotic cells. We find that, compared to sham controls, smoke-exposed mice have ∼60% fewer pro-B/pre-B cells, regardless of NAC treatment. Interestingly, NAC-treated mice show a 21-38% increase in total bone marrow cellularity and lymphocyte frequency and about a 2-fold increase in the pro-B/pre-B cell subset, compared to sham-treated controls. No significant smoking- or NAC-dependent differences were detected in frequency of apoptotic cells or the percentage cells in the G1, S, or G2 phases of the cycle. The failure of NAC treatment to prevent smoking-induced loss of bone marrow pre-B cells suggests that oxidative stress is not directly responsible for this loss. The unexpected expansion of the pro-B/pre-B cell subset in response to NAC treatment suggests oxidative stress normally contributes to cell loss at this developmental stage, and also reveals a potential side effect of therapeutic administration of NAC to prevent smoking-induced loss of lung function.


Hirschsprung disease is linked to defects in neural crest stem cell function.
Genes associated with Hirschsprung disease, a failure to form enteric ganglia in the hindgut, were highly up-regulated in gut neural crest stem cells relative to whole-fetus RNA. One of these genes, the glial cell line-derived neurotrophic factor (GDNF) receptor Ret, was necessary for neural crest stem cell migration in the gut. GDNF promoted the migration of neural crest stem cells in culture but did not affect their survival or proliferation. Gene expression profiling, combined with reverse genetics and analyses of stem cell function, suggests that Hirschsprung disease is caused by defects in neural crest stem cell function.


Climate change and adaptational impacts in coastal systems: the case of sea defences.
We briefly review how coastal ecosystems are responding to and being impacted by climate change, one of the greatest challenges facing society today. In adapting to rising and stormier seas associated with climate change, coastal defence structures are proliferating and becoming dominant coastal features, particularly in urbanised areas. Whilst the primary function of these structures is to protect coastal property and infrastructure, they inevitably have a significant secondary impact on the local environment and ecosystems. In this review we outline some of the negative and positive effects of these structures on physical processes, impacts on marine species, and the novel engineering approaches that have been employed to improve the ecological value of these structures in recent years. Finally we outline guidelines for an environmentally sensitive approach to design of such structures in the marine environment.


Near-infrared-emitting heteroleptic cationic iridium complexes derived from 2,3-diphenylbenzo[g]quinoxaline as in vitro theranostic photodynamic therapy agents.
Five heteroleptic cationic iridium complexes with a π-expansive cyclometalating 2,3-diphenylbenzo[g]quinoxaline (dpbq) ligand (C^N ligand) and different diimine ligands (N^N ligands) (i.e. 2,2'-bipyridine (bpy, 1), phenanthroline (phen, 2), 2-(2-pyridinyl)quinoline (pqu, 3), 2,2'-bisquinoline (bqu, 4), and 2-(quinolin-2-yl)quinoxaline (quqo, 5)) were synthesized and characterized. The lowest-energy singlet electronic transitions (S1 states) were mainly dpbq ligand-centred 1ILCT (intraligand charge transfer)/1MLCT (metal to ligand charge transfer) transitions mixed with some 1π,π* transitions for complexes 1-4 with increased contributions from 1LLCT (ligand to ligand charge transfer) in 3 and 4. For complex 5, the S1 state was switched to the 1LLCT/1MLCT transitions. All five complexes displayed weak near-infrared (NIR) phosphorescence, with maximal emission output spanning 700-1400 nm and quantum yields being on the order of 10-3. The triplet state absorptions of 1-4 all resembled that of the [Ir(dpbq)2Cl]2 dimer with lifetimes of ca. 400 ns, while the TA spectrum of 5 possessed the characteristics of both the quqo ligand and the [Ir(dpbq)2Cl]2 dimer with a bi-exponential decay of ca. 5 μs and 400 ns. While the photophysics of these complexes differ slightly, their theranostic photodynamic therapy (PDT) effects varied drastically. All of the complexes were biologically active toward melanoma cells. Complexes 2 and 3 were the most cytotoxic, with 230-340 nM activity and selectivity factors for melanoma cells over normal skin fibroblasts of 34 to 40 fold. Complexes 2, 3, and 5 became very potent cytotoxins with light activation, with EC50 values as low as 12-18 nM. This potent nanomolar light-triggered activity combined with a lower dark toxicity resulted in 5 having a phototherapeutic index (PI) margin of almost 275. The bpy coligand led to the least amount of dark toxicity of 1, while phen and pqu produced cytotoxic but selective complexes 2 and 3. The quqo coligand produced the most potent complex 5 for in vitro PDT, both in terms of photocytotoxicity and PI. All Ir(iii) complexes exhibited very bright NIR phosphorescence in melanoma cells. The wide range of cytotoxicity and photocytotoxicity effects within a relatively small class of complexes highlights the importance of the identity of the coligand in the biological activity of the π-expansive biscyclometalated Ir(iii) complexes, and their bright NIR emission in live cells demonstrates their potential as theranostic PDT agents.


Prolonged exposure to bacterial toxins downregulated expression of toll-like receptors in mesenchymal stromal cell-derived osteoprogenitors.
Human mesenchymal stromal cells (MSCs, also known as mesenchymal stem cells) are multipotent cells with potential therapeutic value. Owing to their osteogenic capability, MSCs may be clinically applied for facilitating osseointegration in dental implants or orthopedic repair of bony defect. However, whether wound infection or oral microflora may interfere with the growth and osteogenic differentiation of human MSCs remains unknown. This study investigated whether proliferation and osteogenic differentiation of MSCs would be affected by potent gram-positive and gram-negative derived bacterial toxins commonly found in human settings. We selected lipopolysaccharide (LPS) from Escherichia coli and lipoteichoic acid (LTA) from Streptococcus pyogenes as our toxins of choice. Our findings showed both LPS and LTA did not affect MSC proliferation, but prolonged LPS challenge upregulated the osteogenic differentiation of MSCs, as assessed by alkaline phosphatase activity and calcium deposition. Because toll-like receptors (TLRs), in particularly TLR4 and TLR2, are important for the cellular responsiveness to LPS and LTA respectively, we evaluated their expression profiles serially from MSCs to osteoblasts by quantitative PCR. We found that during osteogenic differentiation, MSC-derived osteoprogenitors gradually expressed TLR2 and TLR4 by Day 12. But under prolonged incubation with LPS, MSC-derived osteoprogenitors had reduced TLR2 and TLR4 gene expression. This peculiar response to LPS suggests a possible adaptive mechanism when MSCs are subjected to continuous exposure with bacteria. In conclusion, our findings support the potential of using human MSCs as a biological graft, even under a bacterial toxin-rich environment.


Applicability of Trichoscopy in Scalp Seborrheic Dermatitis.
Seborrheic dermatitis (SD) is a chronic-recidive inflammatory skin disorder with predilection in areas rich of sebaceous gland. The most common clinical manifestations are pruritus and scales. Although SD can be diagnosed without special tools, other examinations may be needed to determine additional specific therapy. Trichoscopy is one of the noninvasive tools which can help to diagnose SD as it can provide the microstructure view of the scalp. This descriptive study was conducted to explore the trichoscopic features of SD and its characteristics. There were 96 SD patients enrolled in this study. The scalp was divided into four areas, and each area was scored based on Seborrheic Area Severity Index, comprising erythema, desquamation, number of papules, and percentage of lesion area. The most severe area was examined with a trichoscopy to observe the characteristics of hair and scalp. The association between trichoscopic findings and SD severity was analyzed with Fisher's exact test. Overall, the participants were 36% males and 64% females with the mean age of 30 (13-70) years old. Based on the trichoscopic examination, the most common findings were thick hair shafts (72%), white scales (69%), arborizing thin vessels (38%), yellowish area (36%), and structureless red area (19%). These findings were not significantly different between mild and moderate SD (P > 0.05). Considering the merits and demerits of trichoscopic examination, it can be helpful to aid the diagnosis of SD. Further studies in Asian population with greater sample size are needed to demonstrate more significant result.


A shift from adaptive to innate immunity: a potential mechanism of disease progression in multiple sclerosis.
Multiple sclerosis is postulated to be a T cell-mediated autoimmune disease characterised by a relapsing-remitting stage followed by a secondary progressive phase. The relapsing remitting phase may involve waves of proinflammatory Th1 and Th17 cells that infiltrate the nervous system, provoking a clinical attack. The activity of these cells is modulated by other populations of regulatory T cells and the balance between the pro-inflammatory and regulatory T cells is critical for determining disease activity. Promoting the activity of regulatory cells is a potentially beneficial therapeutic strategy, and probably contributes to the action of glatiramer acetate. The progressive phase of multiple sclerosis is believed to be secondary to neurodegenerative changes triggered by inflammation. The status of the innate immune system and its relationship to the stages of multiple sclerosis has been poorly defined until recently. However, recent data suggest that these results demonstrate abnormalities of dendritic cell activation or maturation may underlie the transition to the progressive phase of the disease. Preventing this transition, perhaps by acting at the level of the innate immune system, is an important treatment goal. The identification of biomarkers to predict disease course and treatment response is a major challenge in multiple sclerosis research. Studies using antigen arrays have identified antibody patterns related to CNS antigens and heat-shock proteins that are associated with different disease stages and with response to therapy. In the future, such antibody repertoires could be used as biomarkers for the diagnosis and evaluation of patients with multiple sclerosis, for matching treatments to individual patients and, potentially, to identify healthy individuals at risk for this autoimmune disease.


Differences in Sexual Practices, Sexual Behavior and HIV Risk Profile between Adolescents and Young Persons in Rural and Urban Nigeria.
We aimed to determine differences in sexual practices, HIV sexual risk behaviors, and HIV risk profile of adolescents and young persons' in rural and urban Nigeria. We recruited 772 participants 15 to 24 years old from urban and rural townships in Nigeria through a household survey. Information on participants' socio-demographic profile (age sex, residential area, number of meals taken per day), sexual practices (vagina, oral and anal sex; heterosexual and homosexual sex; sex with spouse, casual acquaintances, boy/girlfriend and commercial sex workers), sexual behavior (age of sexual debut, use of condom, multiple sex partners, transactional sex and age of sexual partner), and other HIV risk factors (use of alcohol and psychoactive substances, reason for sexual debut, knowledge of HIV prevention and HIV transmission, report of STI symptoms) were collected through an interviewer administered questionnaire. Differences in sexual behavior and sexual practices of adolescents and HIV risk profile of adolescents and young persons resident in urban and rural areas were determined. More than half (53.5%) of the respondents were sexually active, with more residing in the rural than urban areas (64.9% vs 44.1%; p<0.001) and more resident in the rural area reporting having more than one sexual partner (29.5% vs 20.4%; p = 0.04). Also, 97.3% of sexually active respondents reported having vaginal sex, 8.7% reported oral sex and 1.9% reported anal sex. More male than female respondents in the urban area used condoms during the last vaginal sexual intercourse (69.1% vs 51.9%; p = 0.02), and reported sex with casual partners (7.0% vs 15.3%; p = 0.007). More female than male respondents residing in the rural area engaged in transactional sex (1.0% vs 6.7%; p = 0.005). More females than males in both rural (3.6% vs 10.2%; p = 0.04) and urban (4.7% vs 26.6%; p<0.001) areas self-reported a history of discharge. More females than males in both rural (1.4% vs 17.0%; p = 0.04) and urban (15.0% vs 29.1%; p<0.001) areas self-reported a history of itching. There are differences in the sexual behavior and practices of adolescents and young persons' residing in the urban and rural area with implication for HIV prevention programming.


Tunable light and drug induced depletion of target proteins.
Biological processes in development and disease are controlled by the abundance, localization and modification of cellular proteins. We have developed versatile tools based on recombinant E3 ubiquitin ligases that are controlled by light or drug induced heterodimerization for nanobody or DARPin targeted depletion of endogenous proteins in cells and organisms. We use this rapid, tunable and reversible protein depletion for functional studies of essential proteins like PCNA in DNA repair and to investigate the role of CED-3 in apoptosis during Caenorhabditis elegans development. These independent tools can be combined for spatial and temporal depletion of different sets of proteins, can help to distinguish immediate cellular responses from long-term adaptation effects and can facilitate the exploration of complex networks.


Intracranial vascular calcifications, glioblastoma multiforme, and lead poisoning.
A 72-year-man with previous lead poisoning presented with raised intracranial pressure and localizing neurologic signs. CT scans showed a high-grade glioma and extensive intracranial calcifications, which proved to be vascular in distribution on postmortem examination. The latter findings support the concept of dystrophic calcification following lead-induced cerebrovascular injury. Lead poisoning should be considered in the differential diagnosis of unexplained intracranial calcifications. There is also evidence from previous studies to suggest a causative relationship between lead poisoning and development of glioma.


Vulnerability of rehabilitated agricultural production systems to invasion by nontarget plant species.
Vast areas of arable land have been retired from crop production and "rehabilitated" to improved system states through landowner incentive programs in the United States (e.g., Conservation and Wetland Reserve Programs), as well as Europe (i.e., Agri-Environment Schemes). Our review of studies conducted on invasion of rehabilitated agricultural production systems by nontarget species elucidates several factors that may increase the vulnerability of these systems to invasion. These systems often exist in highly fragmented and agriculturally dominated landscapes, where propagule sources of target species for colonization may be limited, and are established under conditions where legacies of past disturbance persist and prevent target species from persisting. Furthermore, rehabilitation approaches often do not include or successfully attain all target species or historical ecological processes (e.g., hydrology, grazing, and/or fire cycles) key to resisting invasion. Uncertainty surrounds ways in which nontarget species may compromise long term goals of improving biodiversity and ecosystem services through rehabilitation efforts on former agricultural production lands. This review demonstrates that more studies are needed on the extent and ecological impacts of nontarget species as related to the goals of rehabilitation efforts to secure current and future environmental benefits arising from this widespread conservation practice.


Enrichment of glycoproteins using nanoscale chelating concanavalin A monolithic capillary chromatography.
Immobilized lectin chromatography can be employed for glycoprotein enrichment, but commonly used columns have limitations of yield and resolution. To improve efficiency and to make the technique applicable to minimal sample material, we have developed a nanoscale chelating Concanavalin A (Con A) monolithic capillary prepared using GMA-EDMA (glycidyl methacrylate-co-ethylene dimethacrylate) as polymeric support. Con A was immobilized on Cu(II)-charged iminodiacetic acid (IDA) regenerable sorbents by forming a IDA:Cu(II):Con A sandwich affinity structure that has high column capacity, as well as stability. When compared with conventional Con A lectin chromatography, the monolithic capillary enabled the better reproducible detection of over double the number of unique N-glycoproteins in human urine samples. Utility for analysis of minimal biological samples was confirmed by the successful elucidation of glycoprotein profiles in mouse urine samples at the microliter scale. The improved efficiency of the nanoscale monolithic capillary will impact the analysis of glycoproteins in complex biological samples, especially where only limited material may be available.


Myoendothelial coupling through Cx40 contributes to EDH-induced vasodilation in murine renal arteries: evidence from experiments and modelling.
Regulation of renal vascular resistance plays a major role in controlling arterial blood pressure. The endothelium participates in this regulation as endothelial derived hyperpolarization plays a significant role in smaller renal arteries and arterioles, but the exact mechanisms are still unknown. To investigate the role of vascular gap junctions and potassium channels in the renal endothelial derived hyperpolarization. In interlobar arteries from wild-type and connexin40 knockout mice, we assessed the role of calcium-activated small (SK) and intermediate (IK) conductance potassium channels. The role of inward rectifier potassium channels (Kir) and Na+ /K+ -ATPases was evaluated as was the contribution from gap junctions. Mathematical models estimating diffusion of ions and electrical coupling in myoendothelial gap junctions were used to interpret the results. Lack of connexin40 significantly reduces renal endothelial hyperpolarization. Inhibition of SK and IK channels significantly attenuated renal EDH to a similar degree in wild-type and knockout mice. Inhibition of Kir and Na+ /K+ -ATPases affected the response in wild-type and knockout mice but at different levels of stimulation. The model confirms that activation of endothelial SK and IK channels generates a hyperpolarizing current that enters the vascular smooth muscle cells. Also, extracellular potassium increases sufficiently to activate Kir and Na+ /K+ -ATPases. Renal endothelial hyperpolarization is mainly initiated by activation of IK and SK channels. The model shows that hyperpolarization can spread through myoendothelial gap junctions but enough potassium is released to activate Kir and Na+ /K+ -ATPases. Reduced coupling seems to shift the signalling pathway towards release of potassium. However, an alternative pathway also exists and needs to be investigated.


Acute consumption of flavanol-rich cocoa and the reversal of endothelial dysfunction in smokers.
This study was designed to assess the effect of flavanol-rich food on the circulating pool of bioactive nitric oxide (NO) and endothelial dysfunction in smokers. Studies suggest that smoking-related vascular disease is caused by impaired NO synthesis and that diets rich in flavanols can increase bioactive NO in plasma. In smokers (n = 11), the effects of flavanol-rich cocoa on circulating NO species in plasma (RXNO) measured by reductive gas-phase chemiluminescence and endothelial function as assessed by flow-mediated dilation (FMD) were characterized in a dose-finding study orally administering cocoa containing 88 to 370 mg flavanols and in a randomized double-blind crossover study using 100 ml cocoa drink with high (176 to 185 mg) or low (<11 mg) flavanol content on two separate days. In addition to cocoa drink, ascorbic acid and NO-synthase inhibitor L-NMMA (n = 4) were applied. There were significant increases in RXNO (21 +/- 3 nmol/l to 29 +/- 5 nmol/l) and FMD (4.5 +/- 0.8% to 6.9 +/- 0.9%, each p < 0.05) at 2 h after ingestion of 176 to 185 mg flavanols, a dose potentially exerting maximal effects. These changes correlated with increases in flavanol metabolites. Cocoa-associated increases in RXNO and FMD were reversed by L-NMMA. Ascorbic acid had no effect. The circulating pool of bioactive NO and endothelium-dependent vasodilation is acutely increased in smokers following the oral ingestion of a flavanol-rich cocoa drink. The increase in circulating NO pool may contribute to beneficial vascular health effects of flavanol-rich food.


"Passage to Paradise" ethics and end-of-life decisions in children.
We describe our personal practice in the care of the dying child. There is no on right model of care, and the multidisciplinary team must be sensitive to individual needs. It is important to get the balance of palliation and active treatment right (important at all stages of the illness). Key is careful planning by the whole multidisciplinary team and unhurried and frequent communication with the parents by a few key people. It is important to keep re-checking levels of understanding of the family; at times of stress, little information may be retained. Care should extend to the wider family and friends, and it is important also to consider the needs of hospital staff, both the immediate core team and beyond. Respect for religious and cultural issues is essential, but should not be allowed to interfere with optimum treatment of the child. The validity of different patterns of grieving should be acknowledged. Regular review is essential. There need to be open discussions about capping care, so that only appropriate interventions are given. This may be more difficult if the dying child is awaiting transplantation. The place of death (home, hospice, hospital) will be a matter of individual choice. After the death of the child, we offer follow up care for both the family and the staff.


A High-Throughput Regeneration and Transformation Platform for Production of Genetically Modified Banana.
Banana (Musa spp.) is an important staple food as well as cash crop in tropical and subtropical countries. Various bacterial, fungal, and viral diseases and pests such as nematodes are major constraints in its production and are currently destabilizing the banana production in sub-Saharan Africa. Genetic engineering is a complementary option used for incorporating useful traits in banana to bypass the long generation time, polyploidy, and sterility of most of the cultivated varieties. A robust transformation protocol for farmer preferred varieties is crucial for banana genomics and improvement. A robust and reproducible system for genetic transformation of banana using embryogenic cell suspensions (ECS) has been developed in this study. Two different types of explants (immature male flowers and multiple buds) were tested for their ability to develop ECS in several varieties of banana locally grown in Africa. ECS of banana varieties "Cavendish Williams" and "Gros Michel" were developed using multiple buds, whereas ECS of "Sukali Ndiizi" was developed using immature male flowers. Regeneration efficiency of ECS was about 20,000-50,000 plantlets per ml of settled cell volume (SCV) depending on variety. ECS of three different varieties were transformed through Agrobacterium-mediated transformation using gusA reporter gene and 20-70 independent transgenic events per ml SCV of ECS were regenerated on selective medium. The presence and integration of gusA gene in transgenic plants was confirmed by PCR, dot blot, and Southern blot analysis and expression by histochemical GUS assays. The robust transformation platform was successfully used to generate hundreds of transgenic lines with disease resistance. Such a platform will facilitate the transfer of technologies to national agricultural research systems (NARS) in Africa.


Mercury species in seawater and sediment of Xiamen western sea area adjacent to a coal-fired power plant.
The purpose of this study was to investigate the concentrations and spatial distributions of mercury (Hg) species in seawater (including dissolved, particle, and total Hg) and sediment (including total and methyl Hg) of the Xiamen western sea area adjacent to a coal-fired power plant. The influence of the wastewater discharged from the seawater desulphurization (De-SO2) system of the power plant is discussed. Concentration of the three species of mercury in seawater from 18 sampling sites varied: dissolved ranged from 0.70 to 4.65 ng/L (mean 1.47 ng/L; median 1.12 ng/L); particulate ranged from not detected to 90.52 ng/L (mean 10.47 ng/L; median 1.26 ng/L); and total was 1.51 to 92.88 ng/L (mean 11.94 ng/L; median 2.84 ng/L). High concentrations of total Hg and particulate Hg, more than 70 ng/L, were observed in the area adjacent to the outfalls of the power plant. The Hg from the waste seawater of the power plant might be re-emitted to the atmosphere because it exists in seawater mainly in particulate attached form. The sediment total Hg concentrations ranged from 0.055 to 0.201 microg/g, with a mean of 0.126 microg/g and a median of 0.125 microg/g. Low methyl Hg concentrations in the sediment were observed in the study area, ranging from 0.017 to 0.256 ng/g as Hg, with a mean of 0.087 ng/g and a median of 0.081 ng/g. The ratios of methyl Hg to total Hg in the sediment were low, with a mean and median of 0.069%. Both total and methyl Hg were significantly linked to the sediment organic carbon (SOC).


Utilization of cardiac troponin assays in adult and pediatric populations: Guideline recommendations vs. reality.
We hypothesized significant gaps remained for cardiac troponin (cTn) utilization in the United States, despite an emerging evidence base and guideline recommendations. We tested this hypothesis and investigated differences and trends between the use of cTn in adult versus pediatric hospitals. Individuals were identified by a targeted distribution through personal contact and professional society email lists. Participants completed an online survey (Qualtrics) from 07/15/13 to 07/26/13. The 31-item questionnaire used skipped logic and collected data about the respondent and their institutional cTn clinical practices. Data tabulation and analysis were conducted using Qualtrics and Microsoft Excel. A total of 159 unique laboratories responded to the survey, representing primarily adult (81%) versus pediatric (19%) institutions. 59% of laboratories utilize the guideline recommended 99th percentile as the upper reference limit (URL) for cTn, with large variability in reporting practices among users of the same assay. 73% of laboratories reported simultaneous ordering of other cardiac biomarkers with cTn, a majority which included CK-MB. Interpretive comments were used with cTn in 71 laboratories with a significant amount of heterogeneity. Pediatric hospitals reported a lower frequency of cTn orders and were less likely to consider elevated cTn a critical value. Gaps in current utilization and reporting of cTn exist, along with practices inconsistent with clinical guideline recommendations. Implementation of the 99th percentile and serial sampling protocols will be critical to adoption of high-sensitivity cTn assays. Differences in chest pain etiology are the most likely reason for the notable differences between cTn use in adults versus pediatric hospitals.


Genome Modification in Enterococcus faecalis OG1RF Assessed by Bisulfite Sequencing and Single-Molecule Real-Time Sequencing.
Enterococcus faecalis is a Gram-positive bacterium that natively colonizes the human gastrointestinal tract and opportunistically causes life-threatening infections. Multidrug-resistant (MDR) E. faecalis strains have emerged, reducing treatment options for these infections. MDR E. faecalis strains have large genomes containing mobile genetic elements (MGEs) that harbor genes for antibiotic resistance and virulence determinants. Bacteria commonly possess genome defense mechanisms to block MGE acquisition, and we hypothesize that these mechanisms have been compromised in MDR E. faecalis. In restriction-modification (R-M) defense, the bacterial genome is methylated at cytosine (C) or adenine (A) residues by a methyltransferase (MTase), such that nonself DNA can be distinguished from self DNA. A cognate restriction endonuclease digests improperly modified nonself DNA. Little is known about R-M in E. faecalis. Here, we use genome resequencing to identify DNA modifications occurring in the oral isolate OG1RF. OG1RF has one of the smallest E. faecalis genomes sequenced to date and possesses few MGEs. Single-molecule real-time (SMRT) and bisulfite sequencing revealed that OG1RF has global 5-methylcytosine (m5C) methylation at 5'-GCWGC-3' motifs. A type II R-M system confers the m5C modification, and disruption of this system impacts OG1RF electrotransformability and conjugative transfer of an antibiotic resistance plasmid. A second DNA MTase was poorly expressed under laboratory conditions but conferred global N(4)-methylcytosine (m4C) methylation at 5'-CCGG-3' motifs when expressed in Escherichia coli. Based on our results, we conclude that R-M can act as a barrier to MGE acquisition and likely influences antibiotic resistance gene dissemination in the E. faecalis species. The horizontal transfer of antibiotic resistance genes among bacteria is a critical public health concern. Enterococcus faecalis is an opportunistic pathogen that causes life-threatening infections in humans. Multidrug resistance acquired by horizontal gene transfer limits treatment options for these infections. In this study, we used innovative DNA sequencing methodologies to investigate how a model strain of E. faecalis discriminates its own DNA from foreign DNA, i.e., self versus nonself discrimination. We also assess the role of an E. faecalis genome modification system in modulating conjugative transfer of an antibiotic resistance plasmid. These results are significant because they demonstrate that differential genome modification impacts horizontal gene transfer frequencies in E. faecalis.


Wetting kinetics of oil mixtures on fluorinated model cellulose surfaces.
The wetting of two different model cellulose surfaces has been studied; a regenerated cellulose (RG) surface prepared by spin-coating, and a novel multilayer film of poly(ethyleneimine) and a carboxymethylated microfibrillated cellulose (MFC). The cellulose films were characterized in detail using atomic force microscopy (AFM) and X-ray photoelectron spectroscopy (XPS). AFM indicates smooth and continuous films on a nanometer scale and the RMS roughness of the RG cellulose and MFC surfaces was determined to be 3 and 6 nm, respectively. The cellulose films were modified by coating with various amounts of an anionic fluorosurfactant, perfluorooctadecanoic acid, or covalently modified with pentadecafluorooctanyl chloride. The fluorinated cellulose films were used to follow the spreading mechanisms of three different oil mixtures. The viscosity and surface tension of the oils were found to be essential parameters governing the spreading kinetics on these surfaces. XPS and dispersive surface energy measurements were made on the cellulose films coated with perfluorooctadecanoic acid. A strong correlation was found between the surface concentration of fluorine, the dispersive surface energy and the contact angle of castor oil on the surface. A dispersive surface energy less than 18 mN/m was required in order for the cellulose surface to be non-wetting (theta e>90 degrees ) by castor oil.


Weight stigma in frequent exercisers: Overt, demeaning and condescending.
The aim of this study was to qualitatively examine weight stigma in individuals who exercise frequently. In total, six focus groups, comprising 30 participants aged 18-25 years, were conducted using convenience sampling. All participants were frequent exercisers. Five themes emerged in the data with participants discussing bullying, the consequences of obesity, causes of obesity, lack of willpower and interventions to reduce obesity. This study is the first qualitative examination of weight stigmatisation in frequent exercisers, where the beliefs reported by focus group participants suggest that frequent exercisers stigmatise, discriminate and dehumanise obese people. Future research to examine the impact of weight stigma on exercise motivation and behaviour of obese people appears warranted.


Genetic heterogeneity in autism: From single gene to a pathway perspective.
The extreme genetic heterogeneity of autism spectrum disorder (ASD) represents a major challenge. Recent advances in genetic screening and systems biology approaches have extended our knowledge of the genetic etiology of ASD. In this review, we discuss the paradigm shift from a single gene causation model to pathway perturbation model as a guide to better understand the pathophysiology of ASD. We discuss recent genetic findings obtained through next-generation sequencing (NGS) and examine various integrative analyses using systems biology and complex networks approaches that identify convergent patterns of genetic elements associated with ASD.


An improved method for quantitative ChIP studies of nuclear receptor function.
Chromatin immunoprecipitation (ChIP) is a valuable tool for the endocrine researcher, providing a means to measure the recruitment of hormone-activated nuclear receptors, for example. However, the technique can be challenging to perform and has multiple experimental steps, risking introduction of error at each. The data produced can be challenging to interpret; several different methods are commonly used for normalising data and thus comparing between conditions. Absolute, sensitive quantification of protein-bound DNA is important for correct interpretation of the data. In addition, such quantification can help the investigator in troubleshooting experiments. Here, we outline a ChIP strategy combining droplet digital PCR for accurate quantification with an internal spike-in control for normalisation. This combination strengthens the reliability of ChIP data and allows the operator to optimise their protocol with greater confidence.


Origins and unconventional behavior of neutrophils in developing zebrafish.
The first leukocytes that arise in the development of vertebrate embryos are the primitive macrophages, which differentiate in the yolk sac and then quickly invade embryonic tissues. These macrophages have been considered to constitute a separate lineage, giving rise to no other cell type. Using an in vivo photoactivatable cell tracer in the transparent zebrafish (Danio rerio) embryo, we demonstrated that this lineage also gave rise to an equal or higher number of neutrophilic granulocytes. We were surprised to find that the differentiation of these primitive neutrophils occurs only after primitive myeloid progenitors have dispersed in the tissues. By 2 days after fertilization, these neutrophils have become the major leukocyte type found wandering in the epidermis and mesenchyme. Like the primitive macrophages, all primitive and larval neutrophils express PU.1 and L-plastin and they are highly attracted to local infections, yet only a small fraction of them phagocytose microbes, and to a much lesser extent per cell than the macrophages. They are also attracted to variously stressed or malformed tissues, suggesting a wider role than antimicrobial defense.


Fate of premalignant clones during the asymptomatic phase preceding lymphoid malignancy.
Almost all cancers are preceded by a prolonged period of clinical latency during which a combination of cellular events helps move carcinogen-exposed cells towards a malignant phenotype. Hitherto, investigating the fate of premalignant cells in vivo remained strongly hampered by the fact that these cells are usually indistinguishable from their normal counterparts. Here, for the first time, we have designed a strategy able to reconstitute the replicative history of the bona fide premalignant clone in an animal model, the sheep experimentally infected with the lymphotropic bovine leukemia virus. We have shown that premalignant clones are early and clearly distinguished from other virus-exposed cells on the basis of their degree of clonal expansion and genetic instability. Detectable as early as 0.5 month after the beginning of virus exposure, premalignant cells displayed a two-step pattern of extensive clonal expansion together with a mutation load approximately 6 times higher than that of other virus-exposed cells that remained untransformed during the life span of investigated animals. There was no fixation of somatic mutations over time, suggesting that they regularly lead to cellular death, partly contributing to maintain a normal lymphocyte count during the prolonged premalignant stage. This equilibrium was finally broken after a period of 18.5 to 60 months of clinical latency, when a dramatic decrease in the genetic instability of premalignant cells coincided with a rapid increase in lymphocyte count and lymphoma onset.


Cholinergic drugs for Alzheimer's disease enhance in vitro dopamine release.
Alzheimer's disease is a neurodegenerative disorder associated with a decline in cognitive abilities. Patients also frequently have noncognitive symptoms, such as anxiety, depression, apathy, and psychosis, that impair daily living. The most commonly prescribed treatments for Alzheimer's disease are acetylcholinesterase inhibitors, such as donepezil and galantamine. Enhanced cholinergic functions caused by these compounds are believed to underlie improvements in learning, memory, and attention. The noncognitive aspects of dementia, however, are usually linked to serotonin and dopamine rather than acetylcholine because those neurotransmitter systems most directly influence mood, emotional balance, and psychosis. Fast-scan cyclic voltammetry applied to mouse striatal brain slices was used to measure the real-time release of dopamine arising from spontaneous activity or from single electrical stimulations. At concentrations that include their prescribed dosage ranges, donepezil (1-1000 nM) and galantamine (50-1000 nM) increase action potential-dependent dopamine release. Consistent with previous literature, the data support slightly different modes of action for donepezil and galantamine. The ability of these commonly prescribed drugs to alter catecholamine release may underlie their influence over noncognitive symptoms of dementia. Furthermore, these results suggest that acting via nicotinic receptors, these drugs may serve presently untapped therapeutic roles by altering dopamine release in other disorders involving dopaminergic systems.