Neurons, astrocytes, and oligodendrocytes of the rat cerebral cortex originate from separate progenitor cells: an ultrastructural analysis of clonally related cells.
The diverse array of neurons and glia in the mammalian cerebral cortex arises from proliferating cells of the ventricular zone that surrounds the lateral ventricles of the developing brain. A fundamental but unresolved question is whether the individual cells of the ventricular zone are committed to producing progeny of only one particular phenotype or whether they generate progeny of more than one phenotype. We have begun to address this question by asking if individual cells of the ventricular zone generate exclusively neurons or glia at the onset of cortical neurogenesis in the rat. To assess the phenotypes of cells derived from a common progenitor cell, retroviral-mediated gene transfer was used to introduce the reporter gene, Escherichia coli beta-galactosidase, into ventricular zone cells at embryonic day 15 or 16. We used histochemistry to reveal beta-galactosidase-expressing cells in the mature rat cerebral cortex. Isolated clusters of beta-galactosidase-expressing cells, presumably clones, were identified in serial sections. Since the histochemical reaction product is electron dense, each cell could be examined at the ultrastructural level and assigned definitively to one of the major classes of cells in the cerebral cortex on the basis of well-established morphological criteria. This approach overcomes the problems of cell type identification encountered with light microscopy, where it is not always possible to distinguish between different cell phenotypes. We found that virtually all clones contained cells of exclusively one type: either all astrocytes, all oligodendrocytes, or all neurons. Furthermore, each particular cell type exhibited a different pattern and intensity of staining. The neuronal clones, with one exception, were composed of either all pyramidal cells (projection neurons), or all nonpyramidal cells (interneurons). The size and composition of neuronal clones did not seem related to their position in the cerebral cortex. Collectively, our observations indicate that separate progenitor cells exist for pyramidal neurons, nonpyramidal neurons, astrocytes, and oligodendrocytes. The striking phenotypic homogeneity in the clones arising from individual progenitor cells suggests that by the onset of cortical neurogenesis, at least some lineage restrictions have already occurred among the precursor cell population. Thus, our results suggest that lineage may play a pivotal role in determining some of the functionally important phenotypic attributes of cells in the cerebral cortex.


Validation of the ThinPrep Papanicolaou Test for Cervical Cancer Diagnosis.
Although the accuracy of the Thin-Prep Papanicolaou (Pap) Test has been demonstrated in the detection of precursor lesions to cervical cancer, its performance in identifying invasive cancer has not been adequately evaluated. These studies were designed to determine the effectiveness of the ThinPrep method in diagnosing invasive cervical cancer. In four clinical studies, 47 cases of cervical cancer were processed by both conventional and ThinPrep techniques, and the cytological diagnoses and detection rates were compared. Additional studies evaluated the potential of cell loss during the ThinPrep filtration process. A diagnosis of cancer was made in 45 cases (95.7%) by the ThinPrep method and in 44 cases by conventional Pap smear (93.6%). In the two cases not diagnosed by the ThinPrep method, one was diagnosed as atypical squamous cells of undetermined significance and one as atypical glandular cells of undetermined significance. In the conventional smears, one of the three cases not identified as cancer was atypical squamous cells of undetermined significance, and the other two cases were unsatisfactory. In addition, the ability of the ThinPrep method to demonstrate tumor diathesis and to retain diagnostic cells by the filtration process was confirmed. These studies indicate that the ThinPrep method is at least equivalent to the conventional Pap smear method for the demonstration of squarnous cell carcinoma, adenocarcinoma, and other cervical malignancies.


Eya protein phosphatase activity regulates Six1-Dach-Eya transcriptional effects in mammalian organogenesis.
The precise mechanistic relationship between gene activation and repression events is a central question in mammalian organogenesis, as exemplified by the evolutionarily conserved sine oculis (Six), eyes absent (Eya) and dachshund (Dach) network of genetically interacting proteins. Here, we report that Six1 is required for the development of murine kidney, muscle and inner ear, and that it exhibits synergistic genetic interactions with Eya factors. We demonstrate that the Eya family has a protein phosphatase function, and that its enzymatic activity is required for regulating genes encoding growth control and signalling molecules, modulating precursor cell proliferation. The phosphatase function of Eya switches the function of Six1-Dach from repression to activation, causing transcriptional activation through recruitment of co-activators. The gene-specific recruitment of a co-activator with intrinsic phosphatase activity provides a molecular mechanism for activation of specific gene targets, including those regulating precursor cell proliferation and survival in mammalian organogenesis.


Selecting informative data for developing peptide-MHC binding predictors using a query by committee approach.
Strategies for selecting informative data points for training prediction algorithms are important, particularly when data points are difficult and costly to obtain. A Query by Committee (QBC) training strategy for selecting new data points uses the disagreement between a committee of different algorithms to suggest new data points, which most rationally complement existing data, that is, they are the most informative data points. In order to evaluate this QBC approach on a real-world problem, we compared strategies for selecting new data points. We trained neural network algorithms to obtain methods to predict the binding affinity of peptides binding to the MHC class I molecule, HLA-A2. We show that the QBC strategy leads to a higher performance than a baseline strategy where new data points are selected at random from a pool of available data. Most peptides bind HLA-A2 with a low affinity, and as expected using a strategy of selecting peptides that are predicted to have high binding affinities also lead to more accurate predictors than the base line strategy. The QBC value is shown to correlate with the measured binding affinity. This demonstrates that the different predictors can easily learn if a peptide will fail to bind, but often conflict in predicting if a peptide binds. Using a carefully constructed computational setup, we demonstrate that selecting peptides with a high QBC performs better than low QBC peptides independently from binding affinity. When predictors are trained on a very limited set of data they cannot be expected to disagree in a meaningful way and we find a data limit below which the QBC strategy fails. Finally, it should be noted that data selection strategies similar to those used here might be of use in other settings in which generation of more data is a costly process.


Photoprotective properties of Prunella vulgaris and rosmarinic acid on human keratinocytes.
UVA radiation provokes the generation of reactive oxygen species (ROS), which induce oxidative stress in the exposed cells leading to extensive cellular damage and cell death either by apoptosis or necrosis. One approach to protecting human skin against the harmful effects of UV radiation is by using herbal compounds as photoprotectants. This study evaluated the protective effects of Prunella vulgaris L. (Labiatae) and its main phenolic acid component, rosmarinic acid (RA), against UVA-induced changes in a human keratinocyte cell line (HaCaT). Human keratinocytes exposed to UVA (10-30 J/cm(2)) were treated with an extract of P. vulgaris (1-75 mg/l) or RA (0.9-18 mg/l) for 4h. P. vulgaris and RA exhibited ability to reduce the UVA-caused decrease in a cell viability monitored by neutral red retention and by LDH release into medium. The P. vulgaris extract and RA significantly suppressed UVA-induced ROS production, which manifests as a decrease in intracellular lipid peroxidation, elevation of ATP and reduced glutathione. Post-treatment with P. vulgaris extract and RA also significantly reduced DNA damage. In addition, UVA-induced activation of caspase-3 was inhibited by treatment with P. vulgaris and RA. The P. vulgaris extract and RA demonstrated a concentration-dependent photoprotection (maximum at 25-50 mg/l and 9 mg/l, respectively). These results suggest that P. vulgaris and RA, used in skin care cosmetics, may offer protection against UVA-induced oxidative stress and may be beneficial as a supplement in photoprotective dermatological preparations.


Anti-Müllerian hormone and risk of ovarian cancer in nine cohorts.
Animal and experimental data suggest that anti-Müllerian hormone (AMH) serves as a marker of ovarian reserve and inhibits the growth of ovarian tumors. However, few epidemiologic studies have examined the association between AMH and ovarian cancer risk. We conducted a nested case-control study of 302 ovarian cancer cases and 336 matched controls from nine cohorts. Prediagnostic blood samples of premenopausal women were assayed for AMH using a picoAMH enzyme-linked immunosorbent assay. Odds ratios (ORs) and 95% confidence intervals (CIs) were calculated using multivariable-adjusted conditional logistic regression. AMH concentration was not associated with overall ovarian cancer risk. The multivariable-adjusted OR (95% CI), comparing the highest to the lowest quartile of AMH, was 0.99 (0.59-1.67) (Ptrend : 0.91). The association did not differ by age at blood draw or oral contraceptive use (all Pheterogeneity : ≥0.26). There also was no evidence for heterogeneity of risk for tumors defined by histologic developmental pathway, stage, and grade, and by age at diagnosis and time between blood draw and diagnosis (all Pheterogeneity : ≥0.39). In conclusion, this analysis of mostly late premenopausal women from nine cohorts does not support the hypothesized inverse association between prediagnostic circulating levels of AMH and risk of ovarian cancer.


Detection of a K-ras mutation in urine of patients with colorectal cancer.
We previously demonstrated that human urine contains small, 150 to 250 nucleotide-sized, soluble DNA derived from the circulation, which may be useful in the detection of colorectal cancer. In this report we have determined the stability of DNA in urine and have found that the half-life time interval of this small, fragmented DNA is at least 4 hours post collection. We further compared, in a blinded study, the frequency of detecting mutated K-ras sequence in DNA isolated from plasma and urine derived from individuals who have either a colorectal carcinoma (CRC), or adenomatous polyps that contain a mutation in codon 12 of the K-ras proto-oncogene. There was an 83% concurrence of mutated DNA detected in urine and its corresponding disease tissue from the same individuals, when paired urine and tissue sections from 20 patients with either CRC or adenomatous polyps were analyzed for K-ras mutation. However, only a 56% concurrence was observed when the matched plasma specimens were tested from these 20 patients. These results suggest that urine might be a better resource for detecting K-ras mutation in circulating DNA.


Antimicrobial resistance in large clostridial toxin-negative, binary toxin-positive Clostridium difficile ribotypes.
Antimicrobial resistance (AMR) is commonly found in Clostridium difficile strains and plays a major role in strain evolution. We have previously reported the isolation of large clostridial toxin-negative, binary toxin-producing (A-B-CDT+) C. difficile strains from colonised (and in some instances diarrhoeic) food animals, as well as from patients with diarrhoea. To further characterise these strains, we investigated the phenotypic and genotypic AMR profiles of a diverse collection of A-B-CDT+C. difficile strains. The in vitro activities of 10 antimicrobial agents were determined for 148 A-B-CDT+C. difficile strains using an agar dilution methodology. Whole-genome sequencing and in silico genotyping was performed on 53 isolates to identify AMR genes. All strains were susceptible to vancomycin, metronidazole and fidaxomicin, antimicrobials currently considered first-line treatments for C. difficile infection (CDI). Differences in antimicrobial phenotypes between PCR ribotypes (RTs) were observed but were minimal. Phenotypic resistance was observed in 13 isolates to tetracycline (TetR, MIC = 16 mg/L), moxifloxacin (MxfR, MIC = 16 mg/L), erythromycin (EryR, MIC ≥128 mg/L) and clindamycin (CliR, MIC = 8 mg/L). The MxfR strain (RT033) possessed mutations in gyrA/B, while the TetR (RT033) strain contained a tetM gene carried on the conjugative transposon Tn6190. All EryR and CliR strains (RT033, QX521) were negative for the erythromycin ribosomal methylase gene ermB, suggesting a possible alternative mechanism of resistance. This work describes the presence of multiple AMR genes in A-B-CDT+C. difficile strains and provides the first comprehensive analysis of the AMR repertoire in these lineages isolated from human, animal, food and environmental sources.


Commitment to cyst formation in Giardia.
Giardia trophozoites differentiate into infectious cysts (encystment) in response to physiological stimuli; encystment is crucial for Giardia's transmission, survival and pathogenesis. In vitro, Giardia encysts when bile sequesters lipids necessary for this lipid auxotroph, and in vivo they encyst to infect new hosts. In this study, we investigated, for the first time, commitment to encystment in Giardia using both molecular and cellular techniques. We show that after 3-6 h in inducing conditions, encysting trophozoites continue to encyst regardless of whether the inducing stimulus remains. We propose that a trophozoite's inability to revert to a growing or dividing trophozoite represents a commitment to encystment. The onset of commitment correlated with the appearance of encystment specific vesicles (ESVs) and encystment specific protein synthesis. These observations suggest the involvement of regulatory pathways with the ability to 'remember' a transient signal long after its removal; a property that enables encysting trophozoites to complete the encystment process should the unfavourable triggering condition(s) change. The ability to form cysts in response to transient signals or, as we have highlighted in this paper, the ability of a small percentage of the population to form cysts without an inducer is vital for the maintenance of infection within populations.


Long-term persistence of human donor alveolar macrophages in lung transplant recipients.
Alveolar macrophages (AMFs) are critical regulators of lung function, and may participate in graft rejection following lung transplantation. Recent studies in experimental animals suggest that most AMFs are self-maintaining cells of embryonic origin, but knowledge about the ontogeny and life span of human AMFs is scarce. To follow the origin and longevity of AMFs in patients with lung transplantation for more than 100 weeks, we obtained transbronchial biopsies from 10 gender-mismatched patients with lung transplantation. These were subjected to combined in situ hybridisation for X/Y chromosomes and immunofluorescence staining for macrophage markers. Moreover, development of AMFs in humanised mice reconstituted with CD34+ umbilical cord-derived cells was assessed. The number of donor-derived AMFs was unchanged during the 2 year post-transplantation period. A fraction of the AMFs proliferated locally, demonstrating that at least a subset of human AMFs have the capacity to self-renew. Lungs of humanised mice were found to abundantly contain populations of human AMFs expressing markers compatible with a monocyte origin. Moreover, in patients with lung transplantation we found that recipient monocytes seeded the alveoli early after transplantation, and showed subsequent phenotypical changes consistent with differentiation into proliferating mature AMFs. This resulted in a stable mixed chimerism between donor and recipient AMFs throughout the 2-year period. The finding that human AMFs are maintained in the lung parenchyma for several years indicates that pulmonary macrophage transplantation can be a feasible therapeutic option for patients with diseases caused by dysfunctional AMFs. Moreover, in a lung transplantation setting, long-term persistence of donor AMFs may be important for the development of chronic graft rejection.


Plants control the seasonal dynamics of microbial N cycling in a beech forest soil by belowground C allocation.
Soil microbes in temperate forest ecosystems are able to cycle several hundreds of kilograms of N per hectare per year and are therefore of paramount importance for N retention. Belowground C allocation by trees is an important driver of seasonal microbial dynamics and may thus directly affect N transformation processes over the course of the year. Our study aimed at unraveling plant controls on soil N cycling in a temperate beech forest at a high temporal resolution over a time period of two years, by investigating the effects of tree girdling on microbial N turnover. In both years of the experiment, we discovered (1) a summer N mineralization phase (between July and August) and (2) a winter N immobilization phase (November-February). The summer mineralization phase was characterized by a high N mineralization activity, low microbial N uptake, and a subsequent high N availability in the soil. During the autumn/winter N immobilization phase, gross N mineralization rates were low, and microbial N uptake exceeded microbial N mineralization, which led to high levels of N in the microbial biomass and low N availability in the soil. The observed immobilization phase during the winter may play a crucial role for ecosystem functioning, since it could protect dissolved N that is produced by autumn litter degradation from being lost from the ecosystem during the phase when plants are mostly inactive. The difference between microbial biomass N levels in winter and spring equals 38 kg N/ha and may thus account for almost one-third of the annual plant N demand. Tree girdling strongly affected annual N cycling: the winter N immobilization phase disappeared in girdled plots (microbial N uptake and microbial biomass N were significantly reduced, while the amount of available N in the soil solution was enhanced). This was correlated to a reduced fungal abundance in autumn in girdled plots. By releasing recently fixed photosynthates to the soil, plants may thus actively control the annual microbial N cycle. Tree belowground C allocation increases N accumulation in microorganisms during the winter which may ultimately feed back on plant N availability in the following growing season.


Flexible cerebral connectivity patterns subserve contextual modulations of pain.
The perception of pain can be significantly modulated by the behavioral context. Here, we investigated how contextual modulations of pain are subserved in the human brain. We independently modulated the attentional and emotional context of painful stimuli and recorded brain activity by using functional magnetic resonance imaging. Our results confirm that attention to pain and a negative emotional context increases pain perception and this is concomitantly associated with increased neural activity in the anterior insular cortex. Connectivity analyses further reveal that during attentional and emotional modulations of pain, the anterior insula selectively and flexibly connects to attentional and emotional brain networks in frontoparietal and medial temporal lobe areas, respectively. We conclude that the flexible functional connectivity of the anterior insula to other functional systems of the brain, for example, attentional and emotional brain networks, subserves the extraordinary sensitivity of the pain experience to contextual modulations.


Role of Physical Activity in Parkinson's Disease.
Parkinson's disease (PD) is common, age-dependent neurodegenerative disorder caused by a severe loss of the nigrostriatal dopaminergic neurons. Given the projected increase in the number of people with PD over the coming decades, interventions aimed at minimizing morbidity and improve quality of life are crucial. There is currently no fully proven pharmacological therapy that can modify or slow the disease progression. Physical activity (PA) can complement pharmacological therapy to manage the inherent decline associated with the disease. The evidence indicates that upregulation of neurotrophins and nerve growth factors are potentially critical mediators of the beneficial effects associated with PA. Accumulating evidence suggests that patients with PD might benefit from PA in a number of ways, from general improvements in health to disease-specific effects and potentially, disease-modifying effects. Various forms of PA that have shown beneficial effects in PD include - aerobic exercises, treadmill training, dancing, traditional Chinese exercise, yoga, and resistance training. In this review, we explored available research that addresses the impact of exercise and PA on PD. The original articles with randomized control trials, prospective cohort studies, longitudinal studies, meta-analysis, and relevant review articles from 2005 to 2017 were selected for the present review. Many gaps remain in our understanding of the most effective exercise intervention for PD symptoms, the mechanisms underlying exercise-induced changes and the best way to monitor response to therapy. However, available research suggests that exercise is a promising, cost-effective, and low-risk intervention to improve both motor and nonmotor symptoms in patients with PD. Thus, PA should be prescribed and encouraged in all PD patients.


14-3-3s regulate fructose-2,6-bisphosphate levels by binding to PKB-phosphorylated cardiac fructose-2,6-bisphosphate kinase/phosphatase.
The cardiac isoform of 6-phosphofructo-2-kinase/ fructose-2,6-bisphosphatase (PFK-2), regulator of the glycolysis-stimulating fructose-2,6-bisphosphate, was among human HeLa cell proteins that were eluted from a 14-3-3 affinity column using the phosphopeptide ARAApSAPA. Tryptic mass fingerprinting and phospho-specific antibodies showed that Ser466 and Ser483 of 14-3-3-affinity-purified PFK-2 were phosphorylated. 14-3-3 binding was abolished by selectively dephosphorylating Ser483, and 14-3-3 binding was restored when both Ser466 and Ser483 were phosphorylated with PKB, but not when Ser466 alone was phosphorylated by AMPK. Furthermore, the phosphopeptide RNYpS(483)VGS blocked binding of PFK-2 to 14-3-3s. These data indicate that 14-3-3s bind to phosphorylated Ser483. When HeLa cells expressing HA-tagged PFK-2 were co-transfected with active PKB or stimulated with IGF-1, HA-PFK-2 was phosphorylated and bound to 14-3-3s. The response to IGF-1 was abolished by PI 3-kinase inhibitors. In addition, IGF-1 promoted the binding of endogenous PFK-2 to 14-3-3s. When cells were transduced with penetratin-linked AARAApSAPA, we found that this reagent bound specifically to 14-3-3s, blocked the IGF-1-induced binding of HA-PFK-2 to 14-3-3s, and completely inhibited the IGF-1-induced increase in cellular fructose-2,6-bisphosphate. These findings suggest that PKB-dependent binding of 14-3-3s to phospho-Ser483 of cardiac PFK-2 mediates the stimulation of glycolysis by growth factor.


Perturbation of physiological systems by nanoparticles.
Nanotechnology is having a tremendous impact on our society. However, societal concerns about human safety under nanoparticle exposure may derail the broad application of this promising technology. Nanoparticles may enter the human body via various routes, including respiratory pathways, the digestive tract, skin contact, intravenous injection, and implantation. After absorption, nanoparticles are carried to distal organs by the bloodstream and the lymphatic system. During this process, they interact with biological molecules and perturb physiological systems. Although some ingested or absorbed nanoparticles are eliminated, others remain in the body for a long time. The human body is composed of multiple systems that work together to maintain physiological homeostasis. The unexpected invasion of these systems by nanoparticles disturbs normal cell signaling, impairs cell and organ functions, and may even cause pathological disorders. This review examines the comprehensive health risks of exposure to nanoparticles by discussing how nanoparticles perturb various physiological systems as revealed by animal studies. The potential toxicity of nanoparticles to each physiological system and the implications of disrupting the balance among systems are emphasized.


Ecophysiology of Zigadenus nuttallii, a toxic spring ephemeral in a warm season grassland: effect of defoliation and fire.
Zigadenus nuttallii, a highly toxic spring ephemeral in tallgrass prairie, was studied in 1985 to ascertain: 1) several ecophysiological characteristics of the species, 2) seasonal patterns of biomass accumulation, and 3) its response to defoliation and fire. The maximum photosynthetic rate of Z. nuttallii measured in unburned prairie was 13.2 μmoles CO2 m-2 s-1 which occurred at 24-28° C and an incident quantum flux of 0.8-1.0 mmoles m-2 s-1. Maximum stomatal conductance measured was 5.4 mm s-1. Early in the season, belowground storage organs (bulbs) decreased in mass and supplied much of the energy for growth of leaves, even though CO2 uptake was possible. Buld mass did not increase until about 6 weeks after shoot emergence implying that, at this time, leaves had become a source rather than a sink for carbohydrates. The result of a single, severe defoliation event was a decrease in biomass of bulbs, leaves and reproductive structures in Z. nuttallii. Intrinsic compensatory mechanisms were not detected. In contrast, fire, which also defoliated plants, did not result in any biomass decrease at the end of the season. Improved post-fire microclimate and increased nutrient supply (extrinsic factors) may have contributed to higher photosynthetic rates and led to biomass compensation in burned prairie. These data support arguments that intrinsic compensatory mechanisms have evolved in response to chronic herbivory.


Species-scanning mutagenesis of the serotonin transporter reveals residues essential in selective, high-affinity recognition of antidepressants.
The serotonin transporter (SERT) is a high-affinity sodium/chloride-dependent neurotransmitter transporter responsible for reuptake of serotonin from the extracellular space. SERT is a selective target of several clinically important antidepressants. In a cross-species analysis comparing human and bovine SERTs, the kinetic parameters for serotonin uptake were found to be similar, however, the pharmacological profiles of the two transporters differ. Following transient expression in COS-1 cells, IC(50) values were determined for several antidepressants and psychostimulants. The potencies of the antidepressants citalopram, fluoxetine, paroxetine and imipramine were several-fold higher at hSERT compared with bSERT. No species selectivity was observed for the antidepressants fluvoxamine, and sertraline or for the psychostimulants cocaine, the cocaine analogue beta-carbomethoxy-3beta-(4-iodophenyl)tropane, or for 3,4-methylenedioxymethamphetamine (MDMA). Analysis of six hSERT/bSERT chimeras and subsequent species-scanning mutagenesis of each isoform revealed methionine-180, tyrosine-495, and phenylalanine-513 to be responsible for the increase in citalopram and paroxetine potencies at hSERT and methionine-180 and phenylalanine-513 to confer species selectivity at hSERT for fluoxetine and imipramine. Results were obtained by doing the forward, bovine to human, mutations and confirmed by doing the reverse mutations. Citalopram analogues were used to define the roles of methionine-180, tyrosine-495, and phenylalanine-513 and to reveal molecular interactions with individual functional groups of citalopram. We suggest that methionine-180 interacts with the heterocyclic nucleus of citalopram or stabilizes the binding pocket and phenylalanine-513 to be a steric blocker of antidepressant recognition.


GWAS meta-analysis of over 29,000 people with epilepsy identifies 26 risk loci and subtype-specific genetic architecture.
Epilepsy is a highly heritable disorder affecting over 50 million people worldwide, of which about one-third are resistant to current treatments. Here we report a multi-ancestry genome-wide association study including 29,944 cases, stratified into three broad categories and seven subtypes of epilepsy, and 52,538 controls. We identify 26 genome-wide significant loci, 19 of which are specific to genetic generalized epilepsy (GGE). We implicate 29 likely causal genes underlying these 26 loci. SNP-based heritability analyses show that common variants explain between 39.6% and 90% of genetic risk for GGE and its subtypes. Subtype analysis revealed markedly different genetic architectures between focal and generalized epilepsies. Gene-set analyses of GGE signals implicate synaptic processes in both excitatory and inhibitory neurons in the brain. Prioritized candidate genes overlap with monogenic epilepsy genes and with targets of current antiseizure medications. Finally, we leverage our results to identify alternate drugs with predicted efficacy if repurposed for epilepsy treatment.


Proper autophagy is indispensable for angiogenesis during chick embryo development.
People have known that autophagy plays a very important role in many physiological and pathological events. But the role of autophagy on embryonic angiogenesis still remains obscure. In this study, we demonstrated that Atg7, Atg8 and Beclin1 were expressed in the plexus vessels of angiogenesis at chick yolk sac membrane and chorioallantoic membrane. Interfering in autophagy with autophagy inducer or inhibitor could restrict the angiogenesis in vivo, which might be driven by the disorder of angiogenesis-related gene expressions, and also lead to embryonic hemorrhage, which was due to imperfection cell junctions in endothelial cells including abnormal expressions of tight junction, adheren junction and desmosome genes. Using HUVECs, we revealed that cell viability and migration ability changed with the alteration of cell autophagy exposed to RAPA or 3-MA. Interestingly, tube formation assay showed that HUVECs ability of tube formation altered with the change of Atg5, Atg7 and Atg8 manipulated by the transfection of their corresponding siRNA or plasmids. Moreover, the lost cell polarity labeled by F-actin and the absenced β-catenin in RAPA-treated and 3-MA-treated cell membrane implied intracellular cytoskeleton alteration was induced by the activation and depression of autophagy. Taken together, our current experimental data reveal that autophagy is really involved in regulating angiogenesis during embryo development.


Genomic Dissection and Expression Profiling Revealed Functional Divergence in Triticum aestivum Leucine Rich Repeat Receptor Like Kinases (TaLRRKs).
The leucine rich repeat receptor like kinases (LRRK) constitute the largest subfamily of receptor like kinases (RLK), which play critical roles in plant development and stress responses. Herein, we identified 531 TaLRRK genes in Triticum aestivum (bread wheat), which were distributed throughout the A, B, and D sub-genomes and chromosomes. These were clustered into 233 homologous groups, which were mostly located on either homeologous chromosomes from various sub-genomes or in proximity on the same chromosome. A total of 255 paralogous genes were predicted which depicted the role of duplication events in expansion of this gene family. Majority of TaLRRKs consisted of trans-membrane region and localized on plasma-membrane. The TaLRRKs were further categorized into eight phylogenetic groups with numerous subgroups on the basis of sequence homology. The gene and protein structure in terms of exon/intron ratio, domains, and motifs organization were found to be variably conserved across the different phylogenetic groups/subgroups, which indicated a potential divergence and neofunctionalization during evolution. High-throughput transcriptome data and quantitative real time PCR analyses in various developmental stages, and biotic and abiotic (heat, drought, and salt) stresses provided insight into modus operandi of TaLRRKs during these conditions. Distinct expression of majority of stress responsive TaLRRKs homologous genes suggested their specified role in a particular condition. These results provided a comprehensive analysis of various characteristic features including functional divergence, which may provide the way for future functional characterization of this important gene family in bread wheat.


A model for predicting the GEARS score from virtual reality surgical simulator metrics.
Surgical education relies heavily upon simulation. Assessment tools include robotic simulator assessments and Global Evaluative Assessment of Robotic Skills (GEARS) metrics, which have been validated. Training programs use GEARS for proficiency testing; however, it requires a trained human evaluator. Due to limited time, learners are reliant on surgical simulator feedback to improve their skills. GEARS and simulator scores have been shown to be correlated but in what capacity is unknown. Our goal is to develop a model for predicting GEARS score using simulator metrics. Linear and multivariate logistic regressions were used on previously reported data by this group. Subjects performed simple (Ring and Rail 1) and complex (Suture Sponge 1) tasks on simulators, the dV-Trainer (dVT) and the da Vinci Skills Simulator (dVSS). They were scored via simulator metrics and GEARS. A linear model for each simulator and exercise showed a positive linear correlation. Equations were developed for predicting GEARS Total Score from simulator Overall Score. Next, the effects of each individual simulator metric on the GEARS Total Score for each simulator and exercise were examined. On the dVSS, Excessive Instrument Force was significant for Ring and Rail 1 and Instrument Collision was significant for Suture Sponge 1. On the dVT, Time to Complete was significant for both exercises. Once the significant variables were identified, multivariate models were generated. Comparing the predicted GEARS Total Score from the linear model (using only simulator Overall Score) to that using the multivariate model (using the significant variables for each simulator and exercise), the results were similar. Our results suggest that trainees can use simulator Overall Score to predict GEARS Total Score using our linear regression equations. This can improve the training process for those preparing for high-stakes assessments.


Antimicrobial susceptibility testing results from 13 hospitals in Viet Nam: VINARES 2016-2017.
To analyse data from 2016-17 from a hospital-based antimicrobial resistance surveillance with national coverage in a network of hospitals Viet Nam. We analysed data from 13 hospitals, 3 less than the dataset from the 2012-13 period. Identification and antimicrobial susceptibility testing data from the clinical microbiology laboratories from samples sent in for routine diagnostics were used. Clinical and Laboratory Standards Institute 2018 guidelines were used for antimicrobial susceptibility testing interpretation. WHONET was used for data entry, management and analysis. 42,553 deduplicated isolates were included in this analysis; including 30,222 (71%) Gram-negative and 12,331 (29%) Gram-positive bacteria. 8,793 (21%) were from ICUs and 7,439 (18%) isolates were from invasive infections. Escherichia coli and Staphylococcus aureus were the most frequently detected species with 9,092 (21%) and 4,833 isolates (11%), respectively; followed by Klebsiella pneumoniae (3,858 isolates - 9.1%) and Acinetobacter baumannii (3,870 isolates - 9%). Bacteria were mainly isolated from sputum (8,798 isolates - 21%), blood (7,118 isolates - 17%) and urine (5,202 isolates - 12%). Among Gram-positives 3,302/4,515 isolates (73%) of S. aureus were MRSA; 99/290 (34%) of Enterococcus faecium were resistant to vancomycin; and 58% (663/1,136) of Streptococcus pneumoniae proportion were reduced susceptible to penicillin. Among Gram-negatives 59% (4,085/6,953) and 40% (1,186/2,958) of E. coli and K. pneumoniae produced ESBL and 29% (376/1,298) and 11% (961/8,830) were resistant to carbapenems, respectively. 79% (2855/3622) and 45% (1,514/3,376) of Acinetobacter spp. and Pseudomonas aeruginosa were carbapenem resistant, respectively. 88% (804/911) of Haemophilus influenzae were ampicillin resistant and 18/253 (7%) of Salmonella spp. and 7/46 (15%) of Shigella spp. were resistant to fluoroquinolones. The number of isolates from which data were submitted in the 2016-2017 period was twice as high as in 2012-2013. AMR proportions were higher in 2016-2017 for most pathogen-antimicrobial combinations of interest including imipenem-resistant A. baumannii, P. aeruginosa and Enterobacterales. The data show alarmingly high and increasing resistant proportions among important organisms in Viet Nam. AMR proportions varied across hospital types and should be interpreted with caution because existing sampling bias and missing information on whether isolates were community or hospital acquired. Affordable and scalable ways to adopt a sample- or case-based approach across the network should be explored and clinical data should be integrated to help provide more accurate inferences of the surveillance data.


Inactivation of Ku-mediated end joining suppresses mec1Delta lethality by depleting the ribonucleotide reductase inhibitor Sml1 through a pathway controlled by Tel1 kinase and the Mre11 complex.
RAD53 and MEC1 are essential Saccharomyces cerevisiae genes required for the DNA replication and DNA damage checkpoint responses. Their lethality can be suppressed by increasing the intracellular pool of deoxynucleotide triphosphates. We report that deletion of YKU70 or YKU80 suppresses mec1Delta, but not rad53Delta, lethality. We show that suppression of mec1Delta lethality is not due to Ku--associated telomeric defects but rather results from the inability of Ku- cells to efficiently repair DNA double strand breaks by nonhomologous end joining. Consistent with these results, mec1Delta lethality is also suppressed by lif1Delta, which like yku70Delta and yku80Delta, prevents nonhomologous end joining. The viability of yku70Delta mec1Delta and yku80Delta mec1Delta cells depends on the ATM-related Tel1 kinase, the Mre11-Rad50-Xrs2 complex, and the DNA damage checkpoint protein Rad9. We further report that this Mec1-independent pathway converges with the Rad53/Dun1-regulated checkpoint kinase cascade and leads to the degradation of the ribonucleotide reductase inhibitor Sml1.


Preclinical evaluation of engineered oncolytic herpes simplex virus for the treatment of pediatric solid tumors.
Recently, investigators showed that mice with syngeneic murine gliomas that were treated with a neuroattenuated oncolytic herpes simplex virus-1 (oHSV), M002, had a significant increase in survival. M002 has deletions in both copies of the γ134.5 gene, enabling replication in tumor cells but precluding infection of normal cells. Previous studies have shown antitumor effects of other oHSV against a number of adult tumors including hepatocellular carcinoma and renal cell carcinoma. The purpose of the current study was to investigate the oncolytic potential of M002 against difficult to treat pediatric liver and kidney tumors. We showed that the oHSV, M002, infected, replicated, and decreased cell survival in hepatoblastoma, malignant rhabdoid kidney tumor, and renal sarcoma cell lines. In addition, we showed that in murine xenografts, treatment with M002 significantly increased survival and decreased tumor growth. Finally, these studies showed that the primary entry protein for oHSV, CD111 (nectin-1) was present in human hepatoblastoma and malignant rhabdoid kidney tumor specimens. We concluded that M002 effectively targeted these rare aggressive tumor types and that M002 may have potential for use in children with unresponsive or relapsed pediatric solid tumors.


[General principles and first clinical trials of therapeutic vaccines against cancer].
In this brief review we describe the general principles of the antigenicity of human tumor cells, which can be recognized by T lymphocytes and particularly by cytolytic T lymphocytes. This antigenicity of tumor cells lead to the development of therapeutic anticancer vaccines that should induce tumor regressions or prevent the development of metastases in the vaccinated patients. We provide some information on a set of small scale clinical trials that we have carried out with metastatic melanoma patients, and which have provided encouraging though limited clinical results. Detailed immunological analyses with some of these vaccinated patients showed strong anti-tumor T cell responses and suggested that the main limiting factor for clinical efficacy is a phenomenon of resistance of the tumor to T lymphocyte attack. Current research projects aim at elucidating the mechanisms of this resistance and to develop new vaccination strategies that circumvent this roadblock.


Newport Green, a fluorescent sensor of weakly bound cellular Zn(2+): competition with proteome for Zn(2).
Newport Green (NPG) is a recognized sensor of cellular Zn(2+) that displays fluorescence enhancement upon binding to Zn(2+). Because of its modest affinity for Zn(2+), the extent of its capacity to bind cellular Zn(2+) is unclear. The present study investigated the range of reactivity of NPG(ESTER) with cells, isolated (Zn)-proteome, and model Zn-proteins. The sensor accumulated in pig kidney LLC-PK1 cells and was slowly (>40 min) hydrolyzed to the fluorescent, acid form, NPG(ACID). The powerful, cell permeant Zn(2+) chelator, N,N,N',N'-tetrakis(2-pyridylmethyl)-ethane-1,2-diamine (TPEN) failed to quench the growing fluorescence emission, indicating that Zn-NPG(ACID) had not formed and NPG-Zn-protein adduct species probably were not present. Furthermore, NPG(ACID) did not bind to Zn-carbonic anhydrase or Zn-alcohol dehydrogenase, two proteins that form adducts with some other sensors. Strikingly, most of the NPG(ACID) that had been converted from NPG(ESTER) was detected in the extracellular medium not the cells. As a result, after cells were incubated with NPG(ESTER) and then Zn-pyrithione to raise the internal concentration of mobile Zn(2+), Zn-NPG(ACID) was only observed in the external medium. Residual cellular NPG(ACID) was unable to bind extra intracellular Zn(2+) delivered by pyrithione. Proteome isolated from the sonicated cell supernatant was also unreactive with NPG(ACID). Titration of proteome or glutathione with Zn(2+) in the presence of NPG(ACID) revealed that NPG(ACID) only weakly competes for mobile Zn(2+) in the presence of these cellular components. In addition, when proteomic Zn(2+) was released by a nitric oxide donor or N-ethyl-maleimide, little Zn(2+) was detected by NPG(ACID). However, exposure to nitric oxide independently enhanced the fluorescence properties of NPG(ACID). Thus, the biochemical properties of NPG related to cellular Zn(2+) chelation deepen the question of how it functions as a Zn(2+) sensor.


In vivo Cigarette Smoke Exposure Decreases CCL20, SLPI, and BD-1 Secretion by Human Primary Nasal Epithelial Cells.
Smokers and individuals exposed to second-hand cigarette smoke have a higher risk of developing chronic sinus and bronchial infections. This suggests that cigarette smoke (CS) has adverse effects on immune defenses against pathogens. Epithelial cells are important in airway innate immunity and are the first line of defense against infection. Airway epithelial cells not only form a physical barrier but also respond to the presence of microbes by secreting antimicrobials, cytokines, and chemokines. These molecules can lyse infectious microorganisms and/or provide signals critical to the initiation of adaptive immune responses. We examined the effects of CS on antimicrobial secretions of primary human nasal epithelial cells (PHNECs). Compared to non-CS-exposed individuals, PHNEC from in vivo CS-exposed individuals secreted less chemokine ligand (C-C motif) 20 (CCL20), Beta-defensin 1 (BD-1), and SLPI apically, less BD-1 and SLPI basolaterally, and more CCL20 basolaterally. Cigarette smoke extract (CSE) exposure in vitro decreased the apical secretion of CCL20 and beta-defensin 1 by PHNEC from non-CS-exposed individuals. Exposing PHNEC from non-CS exposed to CSE also significantly decreased the levels of many mRNA transcripts that are involved in immune signaling. Our results show that in vivo or in vitro exposure to CS alters the secretion of key antimicrobial peptides from PHNEC, but that in vivo CS exposure is a much more important modifier of antimicrobial peptide secretion. Based on the gene expression data, it appears that CSE disrupts multiple immune signaling pathways in PHNEC. Our results provide mechanistic insight into how CS exposure alters the innate immune response and increases an individual's susceptibility to pathogen infection.


Eosinophilic meningitis caused by Angiostrongylus cantonensis: report of 17 cases.
To describe two outbreaks of Angiostrongylus cantonensis infection that occurred in Kaohsiung, Taiwan, during 1998 and 1999, and to characterize the source of the outbreaks and the clinical manifestations of the disease. We performed a retrospective cohort study among Thai laborers with eosinophilic meningitis who ate raw snails (Ampullarium canaliculatus), as well as an environmental surveillance of larvae in snails. We enrolled 17 Thai laborers in whom severe headache and eosinophilia developed within 4 to 23 days after eating raw snails. Twelve (71%) developed eosinophilic meningitis. Third-stage larvae were found in the cerebrospinal fluids of 2 patients and in all 12 tested snails. Specific antibodies to A. cantonensis were detected in serum from 16 of the patients and in cerebrospinal fluid from 5 of the patients. Central nervous system manifestations included headache (n = 17 [100%]), fever (n = 11 [65%]), Brudzinski's sign/stiff neck (n = 11 [65%]), hyperesthesia (n = 3 [18%]), cranial nerve palsy (n = 2 [12%]), diplopia (n = 2 [12%]), and ataxia (n = 1 [6%]). Laboratory findings included peripheral eosinophilia (n = 15 [88%]) and cerebrospinal fluid eosinophilia (n = 12 [71%]); elevated immunoglobulin (Ig) E levels (n = 13 [100%]); and transient increases in white blood cell count (n = 7 [41%]) and in serum levels of creatine kinase (n = 7 [41%]), transaminase (n = 3 [18%]), and lactate dehydrogenase (n = 2 [12%]). The severity of illness and eosinophilia were correlated with the number of ingested snails. Meningeal and basal ganglion enhancement was noted on magnetic resonance imaging in several patients. Treatment with mebendazole combined with glucocorticosteroids appeared to shorten the course of the infection, but not the number of relapses. The eosinophil count fell to normal within 3 months, but IgE levels remained elevated for as long as 6 months. All patients recovered with minimal neurologic sequelae. Eosinophilic meningitis caused by A. cantonensis should be considered in patients who have headache or central nervous system manifestations after eating raw snails.


Effect of Royal Jelly on spatial learning and memory in rat model of streptozotocin-induced sporadic Alzheimer's disease.
It has been recently demonstrated that Royal jelly (RJ) has a beneficial role on neural functions. Alzheimer's disease (AD) is associated with impairments of learning and memory. Therefore, the present study was designed to examine the effect of RJ on spatial learning and memory in rats after intracerebroventricular injection of streptozotocin (icv-STZ). Rats were infused bilaterally with an icv injection of STZ, while sham rats received vehicle only. The rats were feed with RJ-contained food (3% w/w) (lyophilized RJ mixed with powdered regular food) or regular food for 10 days. Then spatial learning and memory was tested in the rats by Morris water maze test. Results showed that in icv-STZ group latency and path length were increased as compared to sham group, also icv-STZ rats less remembered the target quadrant that previously the platform was located; however, these were protected significantly in STZ group that received RJ-containing food. Our findings support the potential neuroprotective role of RJ and its helpful effects in AD.


Safety and Efficacy of Ferric Carboxymaltose in Anemic Pregnant Women: A Retrospective Case Control Study.
Background. Anemia during pregnancy is commonly caused by iron deficiency and can have severe consequences for both the mother and the developing fetus. The aim of this retrospective study was to assess the safety and efficacy of intravenous ferric carboxymaltose (FCM) in pregnant women. Methods. All women treated with FCM for anemia during pregnancy between 2010 and 2012 at our institution were included. A matched control group was selected, including women who either were nonanemic or had anemia but were not considered for intravenous iron. Main outcome measures were maternal safety and pregnancy outcomes. Results. The study included 128 patients (FCM: 64; control: 64). Median FCM dose was 1000 mg and median gestational age at the time of first treatment was 34 weeks and 6 days. Median Hb increased from 8.4 g/dL (interquartile range 7.7; 8.9 g/dL) at the first FCM administration to 10.7 g/dL (9.8; 11.5 g/dL; n = 46 with available Hb at delivery) at the time of delivery, achieving levels similar to those in the control group (10.8 g/dL [9.8; 11.8 g/dL; n = 48]). No treatment-related adverse events were reported and no statistically significant differences in pregnancy outcomes were observed between groups. Conclusions. Within the limitations of this case control study, FCM was a safe and efficient treatment of anemia during pregnancy.


Development, standardization, and testing of a lexicon for reporting contrast-enhanced breast magnetic resonance imaging studies.
The purpose of this study was to develop, standardize, and test reproducibility of a lexicon for reporting contrast-enhanced breast magnetic resonance imaging (MRI) examinations. To standardize breast MRI lesion description and reporting, seven radiologists with extensive breast MRI experience developed consensus on technical detail, clinical history, and terminology reporting to describe kinetic and architectural features of lesions detected on contrast-enhanced breast MR images. This lexicon adapted American College of Radiology Breast Imaging and Data Reporting System terminology for breast MRI reporting, including recommendations for reporting clinical history, technical parameters for breast MRI, descriptions for general breast composition, morphologic and kinetic characteristics of mass lesions or regions of abnormal enhancement, and overall impression and management recommendations. To test morphology reproducibility, seven radiologists assessed morphology characteristics of 85 contrast-enhanced breast MRI studies. Data from each independent reader were used to compute weighted and unweighted kappa (kappa) statistics for interobserver agreement among readers. The MR lexicon differentiates two lesion types, mass and non-mass-like enhancement based on morphology and geographical distribution, with descriptors of shape, margin, and internal enhancement. Lexicon testing showed substantial agreement for breast density (kappa = 0.63) and moderate agreement for lesion type (kappa = 0.57), mass margins (kappa = 0.55), and mass shape (kappa = 0.42). Agreement was fair for internal enhancement characteristics. Unweighted kappa statistics showed highest agreement for the terms dense in the breast composition category, mass in lesion type, spiculated and smooth in mass margins, irregular in mass shape, and both dark septations and rim enhancement for internal enhancement characteristics within a mass. The newly developed breast MR lexicon demonstrated moderate interobserver agreement. While breast density and lesion type appear reproducible, other terms require further refinement and testing to lead to a uniform standard language and reporting system for breast MRI. J. Magn. Reson. Imaging 2001;13:889-895.


Growth/differentiation factor-5: a candidate therapeutic agent for periodontal regeneration? A review of pre-clinical data.
Therapeutic concepts involving the application of matrix, growth and differentiation factors have been advocated in support of periodontal wound healing/regeneration. Growth/differentiation factor-5 (GDF-5), a member of the bone morphogenetic protein family, represents one such factor. The purpose of this review is to provide a background of the therapeutic effects of GDF-5 expressed in various musculoskeletal settings using small and large animal platforms. A comprehensive literature search was conducted to identify all reports in the English language evaluating GDF-5 using the PubMed and Google search engines, and a manual search of the reference lists from the electronically retrieved reports. Two reviewers independently screened the titles and abstracts from a total of 69 reports, 22 of which were identified as pre-clinical (in vivo) evaluations of GDF-5. The full-length article of the 22 pre-clinical reports was then reviewed. Various applications including cranial and craniofacial bone formation, spine fusion, long bone fracture healing, cartilage, and tendon/ligament repair using a variety of small and large animal platforms evaluating GDF-5 as a therapeutic agent were identified. A majority of studies, using biomechanical, radiographic, and histological analysis, demonstrated significant dose-dependent effects of GDF-5. These include increased/enhanced local bone formation, fracture healing/repair, and cartilage and tendon/ligament formation. GDF-5 frequently was shown to accelerate wound maturation. Several studies demonstrated GDF-5 to be a realistic alternative to autograft bone. Studies using pre-clinical models and human histology suggest GDF-5 may also increase/enhance periodontal wound healing/regeneration. GDF-5 appears a promising therapeutic agent for periodontal wound healing/regeneration as GDF-5 supports/accelerates bone and tendon/ligament formation in several musculoskeletal settings including periodontal tissues.


Gone with the wind: the origin of S0 galaxies in clusters
We present three-dimensional, high-resolution hydrodynamical simulations of the interaction between the hot ionized intracluster medium and the cold interstellar medium of spiral galaxies. Ram pressure and turbulent/viscous stripping remove 100% of the atomic hydrogen content of luminous galaxies like the Milky Way within 100 million years. These mechanisms naturally account for the morphology of S0 galaxies and the rapid truncation of star formation implied by spectroscopic observations, as well as a host of observational data on the neutral hydrogen (HI) morphology of galaxies in clusters.


Identification of a cyclin-cdk2 recognition motif present in substrates and p21-like cyclin-dependent kinase inhibitors.
Understanding how cyclin-cdk complexes recognize their substrates is a central problem in cell cycle biology. We identified an E2F1-derived eight-residue peptide which blocked the binding of cyclin A and E-cdk2 complexes to E2F1 and p21. Short peptides spanning similar sequences in p107, p130, and p21-like cdk inhibitors likewise bound to cyclin A-cdk2 and cyclin E-cdk2. In addition, these peptides promoted formation of stable cyclin A-cdk2 complexes in vitro but inhibited the phosphorylation of the retinoblastoma protein by cyclin A- but not cyclin B-associated kinases. Mutation of the cyclin-cdk2 binding motifs in p107 and E2F1 likewise prevented their phosphorylation by cyclin A-associated kinases in vitro. The cdk inhibitor p21 was found to contain two functional copies of this recognition motif, as determined by in vitro kinase binding/inhibition assays and in vivo growth suppression assays. Thus, these studies have identified a cyclin A- and E-cdk2 substrate recognition motif. Furthermore, these data suggest that p21-like cdk inhibitors function, at least in part, by blocking the interaction of substrates with cyclin-cdk2 complexes.


Effects of different head-neck positions on the larynges of ridden horses.
Hyperflexion, that is the strong deflection of the horse's head, poll and neck, is a prevalent training technique in equitation. Hyperflexion has come under criticism in recent years for being suspected of affecting the horses' well-being contrary to animal welfare. The goal of the present study is a comparison between the impacts of different poll-neck positions on findings in the upper respiratory tract of ridden horses. For this purpose, video recordings of the larynges of 14 horses were taken using an overground endoscope. The videos were recorded at rest and during three different riding phases: firstly, in a stretching posture, secondly, in a working position and, thirdly, in hyperflexion. A comparison between the analyses of the working position and hyperflexion phases revealed a significant reduction in the laryngeal opening area (p = 0.001) with a value of 8.2 ± 5.0%. Furthermore, other parameters of the larynx evaluated also showed a significant diminishment. These changes did not correlate with the age of the horses or their level of education, and they were independent of the individual anatomical conditions of the poll-neck region. In summary, it can be stated that hyperflexion causes a considerable compression of the larynx.


A new type of conjugative transposon encodes resistance to sulfamethoxazole, trimethoprim, and streptomycin in Vibrio cholerae O139.
Vibrio cholerae O139 is the first non-O1 serogroup of V. cholerae to give rise to epidemic cholera. Apparently, this new serogroup arose from an El Tor O1 strain of V cholerae, but V. cholerae O139 is distinguishable from V. cholerae El Tor O1 by virtue of its novel antigenic structure and also its characteristic pattern of resistances to the antibiotics sulfamethoxazole, trimethoprim, streptomycin, and furazolidone. We found that the first three of these antibiotic resistances are carried on an approximately 62-kb self-transmissible, chromosomally integrating genetic element which we have termed the SXT element. This novel conjugative transposon-like element could be conjugally transferred from V. cholerae O139 to V cholerae O1 and Escherichia coli strains, where it integrated into the recipient chromosomes in a site-specific manner independent of recA. To study the potential virulence properties of the SXT element as well as to improve upon the live attenuated O139 vaccine strain Bengal-2, a large internal deletion in the SXT element was crossed on to the Bengal-2 chromosome. The resulting strain, Bengal-2.SXT(s), is sensitive to sulfamethoxazole and trimethoprim and colonizes the intestines of suckling mice as well as wild-type strains do, suggesting that the SXT element does not encode a colonization factor. Derivatives of Bengal-2.SXT(s) are predicted to be safe, antibiotic-sensitive, live attenuated vaccines for cholera due to the O139 serogroup.


Analysis and design of modified Wollaston prisms.
An exact analysis of the working parameters of a modified Wollaston prism is presented. Parameters include the output splitting angle, the retardation, and the location of the plane of the interference fringes (plane of apparent splitting). Results are presented for the entire range of optical axis inclinations and wedge angles. Approximate expressions from the literature are evaluated. An angle of incidence that causes the plane of the interference fringes to be perpendicular to the axis of the optical system is found for each configuration analyzed. This is then applied to the design of modified Wollaston prisms for Nomarski differential interference contrast microscopy.


Purulent pericarditis and pneumonia caused by Streptococcus equi subsp. zooepidemicus.
Purulent pericarditis is a life-threatening disease that usually manifests following bacteraemia or through spreading from an intrathoracic focus. Only a few cases of this disease have been reported with Lancefield group C streptococci as aetiological agents, and the primary focus in these infections remains unknown. We report a case of purulent pericarditis with septic and cardiogenic shock, caused by Streptococcus equi subsp. zooepidemicus (group C) in a 51-year-old patient. The pathogen was possibly contracted through contact with horses. Most probably, it caused initially pneumonia before spreading to the pericardium, either directly or via the bloodstream. A combined therapeutic approach, consisting of antibiotic therapy and repeated pericardial drainage, was necessary to ensure a clinical cure. After discharge, long-term follow-up for development of constrictive pericarditis is considered mandatory.


Microfibrous carriers for cell culture: a comparative study.
Small patches of polyethylene terephthalate (PET) nonwoven microfibrous matrices have excellent properties and can be used as carriers for culturing cells in agitated bioreactors. The microfibrous carriers are highly porous and can provide large surface areas and three-dimensional space for high-density cell growth. In this work, the microfibrous carriers and several commercial microcarriers were used to study cell attachment kinetics, growth, and monoclonal antibody production with Chinese hamster ovary cells. Compared with commercial solid and macroporous microcarriers, the microfibrous carriers showed better or similar performances. In addition, the microfibrous carriers provided a wider operable range for agitation rate than commercial microcarriers, effectively protecting cells from shear stress and carrier collisions. In addition, the microfibrous carriers are available at a much lower cost than commercial microcarriers, providing an attractive alternative to microcarrier-based large-scale cell cultures.


Renal failure after high-dose methotrexate in a child homozygous for MTHFR C677T polymorphism.
We report the case of an 11-year-old female treated for mediastinal T-cell lymphoma who presented renal failure following the second cycle of high-dose methotrexate (HDMTX). Because of life threatening plasma methotrexate (MTX) levels, carboxypeptidase G2 (CPDG2) was administered resulting in a dramatic decrease within 1 hr. The patient recovered from renal failure and no other side effects were observed. Homozygosity for the methylentetrahydrofolate reductase (MTHFR) C677T polymorphism diagnosed by molecular genetic analysis was the only explanation for this toxicity.


Relationship between body image disturbance and incidence of depression: the SUN prospective cohort.
Body image disturbance is an increasing problem in Western societies and is associated with a number of mental health outcomes including anorexia, bulimia, body dysmorphia, and depression. The aim of this study was to assess the association between body image disturbance and the incidence of depression. This study included 10,286 participants from a dynamic prospective cohort of Spanish university graduates, who were followed-up for a median period of 4.2 years (Seguimiento Universidad de Navarra - the SUN study). The key characteristic of the study is the permanently open recruitment that started in 1999. The baseline questionnaire included information about body mass index (BMI) and the nine figure schemes that were used to assess body size perception. These variables were grouped according to recommended classifications and the difference between BMI and body size perception was considered as a proxy of body image disturbance. A subject was classified as an incident case of depression if he/she was initially free of depression and reported a physician-made diagnosis of depression and/or the use of antidepressant medication in at least one of the follow-up questionnaires. The association between body image disturbance and the incidence of depression was estimated by calculating the multivariable adjusted Odds Ratio (OR) and its 95% Confidence Interval (95% CI), using logistic regression models. The cumulative incidence of depression during follow-up in the cohort was 4.8%. Men who underestimated their body size had a high percentage of overweight and obesity (50.1% and 12.6%, respectively), whereas women who overestimated their body size had a high percentage of underweight (87.6%). The underestimation exhibited a negative association with the incidence of depression among women (OR: 0.72, 95% CI: 0.54 - 0.95), but this effect disappeared after adjusting for possible confounding variables. The proportion of participants who correctly perceived their body size was high (53.3%) and gross misperception was seldom found, with most cases selecting only one silhouette below (42.7%) or above (2.6%) their actual BMI. We found no association between body image disturbance and subsequent depression in a cohort of university graduates in Spain.


Antibacterial activity of honey against community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA).
Community-associated methicillin-resistant Staphylococcus aureus (CA-MRSA) has now been described globally, as a clinically significant pathogen, particularly associated with skin and soft tissue infections, including abscesses, cellulitis and furunculosis. The recent emergence of CA-MRSA combined with its predominant presentation associated with skin and soft tissue infection, the previous literature indicating honey as an effective treatment of healthcare-associated HA-MRSA-related wound infection, as well as honey's ease of topical application, make the current study timely and of interest to healthcare practitioners involved with wound management. Although previous studies have examined the antimicrobial activity of honey against HA-MRSA, such data are limited regarding the activity of honey against this emerging type of MRSA. CA-MRSA (n=6 isolates), was examined for its susceptibility to natural honey (n=3 honey produced from bees in Northern Ireland and one commercial French honey). Results demonstrated that all honey was able to reduce the cultural count of all CA-MRSA from approximately 10(6) colony-forming units (cfus) (mean = 6.46 log10 cfu/g) to none detectable within 24h of co-culture of separate CA-MRSA organisms individually with all four-honey types examined. Subsequent non-selective enrichment of honey demonstrated that inoculated honey remained positive for CA-MRSA until 72h postinoculation, after which point no culturable organisms could be detected. This study demonstrated that, in vitro, these natural products had an antimicrobial activity against the CA-MRSA organisms tested. Further studies are now required to demonstrate if this antimicrobial activity has any clinical application.


Pick's disease: clinicopathologic characterization of 21 cases.
Pick's disease (PiD) is a unique subtype of frontotemporal lobar degeneration characterized pathologically by aggregates of 3-Repeat tau. Few studies have examined the clinical variability and disease progression in PiD. We describe the clinical features, neuropsychological profiles and coexistent pathologies in 21 cases of autopsy-confirmed PiD. This study was a retrospective analysis of patients with Pick's disease evaluated at Mayo Clinic, Rochester or Jacksonville (1995-2018), and identified through an existing database. Twenty-one cases with sufficient clinical data were identified. Behavioral variant FTD (bvFTD; 12/21) was the most common phenotype, followed by primary progressive aphasia (PPA; 7/21), corticobasal syndrome (CBS; 1/21) and amnestic dementia (1/21). Median age at disease onset was 54 years, with PPA cases (median = 52 years) presenting earlier than bvFTD (median = 59). Median disease duration (onset-death) overall was 10 years and did not differ significantly between bvFTD (median = 9.5 years) and PPA (median = 13). Age at death was not significantly different in PPA (median = 66) compared to bvFTD (median = 68.5). A third of the cases (n = 7/21) demonstrated pure PiD pathology, while the remainder showed co-existent other pathologies including Alzheimer's type (n = 6), cerebral amyloid angiopathy (n = 3), combined Alzheimer's and amyloid angiopathy (n = 4), and Lewy body disease (n = 1). Our study shows that bvFTD and PPA are the most common clinical phenotypes associated with PiD, although rare presentations such as CBS were also seen. Coexisting non-Pick's pathology was also present in many cases. Our study highlights the clinical and pathologic heterogeneity in PiD.


Low chitinase activity in Acacia myrmecophytes: a potential trade-off between biotic and chemical defences?
We determined chitinase activity in leaves of four myrmecophytic and four non-myrmecophytic leguminous species at the plants' natural growing sites in Mexico. Myrmecophytic plants (or 'ant plants') have obligate mutualisms with ants protecting them against herbivores and pathogenic fungi. Plant chitinases can be considered a reliable measure of plant resistance to pathogenic fungi. The myrmecophytic Acacia species, which were colonised by mutualistic ants, exhibited at least six-fold lower levels of chitinase activity compared with the non-myrmecophytic Acacia farnesiana and three other non-myrmecophytes. Though belonging to different phylogenetic groups, the myrmecophytic Acacia species formed one distinct group in the data set, which was clearly separated from the non-myrmecophytic species. These findings allowed for comparison between two recent hypotheses that attempt to explain low chitinase activity in ant plants. Most probably, chitinases are reduced in myrmecophytic plant species because these are effectively defended indirectly due to their symbiosis with mutualistic ants.


Bone formation within a breast abscess.
We present a rare case of osseous metaplasia in a poorly healing breast abscess. An 87-year-old woman was referred to the breast surgery clinic with a painful lump in her right breast. Initial imaging and core biopsy suggested a breast abscess. Despite several courses of antibiotics and repeated attempts at aspiration the painful lesion persisted. It was eventually surgically excised in its entirety and final histopathology showed the presence of bone formation within the abscess. The patient's symptoms subsequently resolved. To the best of our knowledge, this is the first case in the literature, of osseous metaplasia within a breast abscess in the absence of malignancy.


Rituximab versus cyclophosphamide in ANCA-associated renal vasculitis.
Cyclophosphamide induction regimens for antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis are effective in 70 to 90% of patients, but they are associated with high rates of death and adverse events. Treatment with rituximab has led to remission rates of 80 to 90% among patients with refractory ANCA-associated vasculitis and may be safer than cyclophosphamide regimens. We compared rituximab with cyclophosphamide as induction therapy in ANCA-associated vasculitis. We randomly assigned, in a 3:1 ratio, 44 patients with newly diagnosed ANCA-associated vasculitis and renal involvement to a standard glucocorticoid regimen plus either rituximab at a dose of 375 mg per square meter of body-surface area per week for 4 weeks, with two intravenous cyclophosphamide pulses (33 patients, the rituximab group), or intravenous cyclophosphamide for 3 to 6 months followed by azathioprine (11 patients, the control group). Primary end points were sustained remission rates at 12 months and severe adverse events. The median age was 68 years, and the glomerular filtration rate (GFR) was 18 ml per minute per 1.73 m(2) of body-surface area. A total of 25 patients in the rituximab group (76%) and 9 patients in the control group (82%) had a sustained remission (P=0.68). Severe adverse events occurred in 14 patients in the rituximab group (42%) and 4 patients in the control group (36%) (P=0.77). Six of the 33 patients in the rituximab group (18%) and 2 of the 11 patients in the control group (18%) died (P=1.00). The median increase in the GFR between 0 and 12 months was 19 ml per minute in the rituximab group and 15 ml per minute in the control group (P=0.14). A rituximab-based regimen was not superior to standard intravenous cyclophosphamide for severe ANCA-associated vasculitis. Sustained-remission rates were high in both groups, and the rituximab-based regimen was not associated with reductions in early severe adverse events. (Funded by Cambridge University Hospitals National Health Service Foundation Trust and F. Hoffmann-La Roche; Current Controlled Trials number, ISRCTN28528813.)


Does litigation influence medical practice? The influence of community radiologists' medical malpractice perceptions and experience on screening mammography.
To assess the relationship between radiologists' perception of and experience with medical malpractice and their patient-recall rates in actual community-based clinical settings. All study activities were approved by the institutional review boards of the involved institutions, and patient and radiologist informed consent was obtained where necessary. This study was performed in three regions of the United States (Washington, Colorado, and New Hampshire). Radiologists who routinely interpret mammograms completed a mailed survey that included questions on demographic data, practice environment, and medical malpractice. Survey responses were linked to interpretive performance for all screening mammography examinations performed between January 1, 1996, and December 31, 2001. The odds of recall were modeled by using logistic regression analysis based on generalized estimating equations that adjust for study region. Of 181 eligible radiologists, 139 (76.8%) returned the survey with full consent. The analysis included 124 radiologists who had interpreted a total of 557 143 screening mammograms. Approximately half (64 of 122 [52.4%]) of the radiologists reported a prior malpractice claim, with 18 (14.8%) reporting mammography-related claims. The majority (n = 51 [81.0%]) of the 63 radiologists who responded to a question regarding the degree of stress caused by a medical malpractice claim described the experience as very or extremely stressful. More than three of every four radiologists (ie, 94 [76.4%] of 123) expressed concern about the impact medical malpractice has on mammography practice, with over half (72 [58.5%] of 123) indicating that their concern moderately to greatly increased the number of their recommendations for breast biopsies. Radiologists' estimates of their future malpractice risk were substantially higher than the actual historical risk. Almost one of every three radiologists (43 of 122 [35.3%]) had considered withdrawing from mammogram interpretation because of malpractice concerns. No significant association was found between recall rates and radiologists' experiences or perceptions of medical malpractice. U.S. radiologists are extremely concerned about medical malpractice and report that this concern affects their recall rates and biopsy recommendations. However, medical malpractice experience and concerns were not associated with recall or false-positive rates. Heightened concern of almost all radiologists may be a key reason that recall rates are higher in the United States than in other countries, but this hypothesis requires further study.


A flux-sensing mechanism could regulate the switch between respiration and fermentation.
The yeast Saccharomyces cerevisiae can show different metabolic phenotypes (e.g. fermentation and respiration). Based on data from the literature, we argue that the substrate uptake rate is the core variable in the system that controls the global metabolic phenotype. Consequently the metabolic phenotype that the cell expresses is not dependent on the type of the sugar or its concentration, but only on the rate at which the sugar enters the cell. As this requires the cells to 'measure' metabolic flux, we discuss the existing clues toward a flux-sensing mechanism in this organism and also outline several aspects of the involved flux-dependent regulation system. It becomes clear that the sensing and regulation system that divides the taken up carbon flux into the respiratory or fermentative pathways is complex with many molecular components interacting on multiple levels. To obtain a true understanding about how the global metabolic phenotype of S. cerevisiae is controlled by the glucose uptake rate, different tools and approaches from systems biology will be required.


Dopamine receptor expression and function in corticotroph ectopic tumors.
Dopamine receptor (DR) expression and dopamine agonist (DA) effectiveness have never been demonstrated in neuroendocrine tumors associated with ectopic ACTH syndrome (EAS). The aim of the current study was to evaluate DR and particularly D2 subtype expression in neuroendocrine tumors associated with EAS and to evaluate the in vivo effectiveness of the DA cabergoline in the treatment of EAS. Six ACTH-secreting neuroendocrine tumors, including four lung, one pancreatic, and one thymic carcinoid, were used for the evaluation of D2 expression by immunohistochemistry. DR subtypes and D2 isoforms and number were evaluated by RT-PCR in three cases of persistent EAS after surgery. These patients were treated with cabergoline at the dose of 3.5 mg/wk for 6 months. Clinical parameters, hormonal levels, and tumor size were monitored during the treatment period. At immunohistochemistry, D2 was expressed in five (83.3%) tumors. At RT-PCR, D2 was confirmed in all three cases but at variable numbers, whereas D4 was expressed in two cases. D(2long) was expressed in all three cases, together with D(2short) in one case. A normalization of urinary cortisol levels was found in two patients (66.7%) after 3 months of treatment. However, treatment escape was demonstrated in one of these patients afterward. The results of this study demonstrated that DR are expressed in neuroendocrine tumors associated with EAS and that cabergoline treatment could be effective in controlling cortisol excess in a subgroup of patients with EAS. Further studies on a larger number of patients are mandatory to confirm the usefulness of DA in EAS.


International NGOs and primary health care in Mozambique: the need for a new model of collaboration.
In keeping with the neo-liberal emphasis on privatization, international aid has been increasingly channeled through non-governmental organizations (NGOs) and their expatriate technical experts to support primary health care (PHC) in the developing world. Relationships between international aid workers and their local counterparts have thus become critical aspects of PHC and its effectiveness. However, these important social dynamics of PHC remain understudied by social scientists. Based on three years of participant-observation in Mozambique, this paper presents an ethnographic case study of these relationships in one central province. The Mozambique experience reveals that the deluge of NGOs and their expatriate workers over the last decade has fragmented the local health system, undermined local control of health programs, and contributed to growing local social inequality. Since national health system salaries plummeted over the same period as a result of structural adjustment, health workers became vulnerable to financial favors offered by NGOs seeking to promote their projects in turf struggles with other agencies. It is argued that new aid management strategies, while necessary, will not be sufficient to remedy the fragmentation of the health sector. A new model for collaboration between expatriate aid workers and their local counterparts in the developing world is urgently needed that centers on the building of long-term equitable professional relationships in a sustainable adequately funded public sector. The case study presented here illustrates how the NGO model undermines the establishment of these relationships that are so vital to successful development assistance.


An intersubunit trigger of channel gating in the muscle nicotinic receptor.
Binding of neurotransmitter triggers gating of synaptic receptor channels, but our understanding of the structures that link the binding site to the channel is just beginning to develop. Here, we identify an intersubunit triggering element required for rapid and efficient gating of muscle nicotinic receptors using a structural model of the Torpedo receptor at 4 A resolution, recordings of currents through single receptor channels, measurements of inter-residue energetic coupling, and functional consequences of disulfide trapping. Mutation of the conserved residues, alphaTyr 127, epsilonAsn 39, and deltaAsn 41, located at the two subunit interfaces that form the agonist binding sites, markedly attenuates acetylcholine-elicited channel gating; mutant cycle analyses based on changes in the channel gating equilibrium constant reveal strong energetic coupling among these residues. After each residue is substituted with Cys, oxidizing conditions that promote disulfide bond formation attenuate gating of mutant, but not wild-type receptors. Gating is similarly attenuated when the Cys substitutions are confined to either of the binding-site interfaces, but can be restored by reducing conditions that promote disulfide bond breakage. Thus, the Tyr-Asn pair is an intersubunit trigger of rapid and efficient gating of muscle nicotinic receptors.


The use of prostheses in pelvic reconstructive surgery: joy or toy?
The high recurrence rate of pelvic organ prolapse (POP) of up to 30% after pelvic reconstructive surgery makes a more refined surgery imperative, as well as the need for either biological or synthetic prostheses as adjuvant treatment. Patients with recurrence risks may benefit from the adjuvant treatment: (1) to substitute for the lack of supportive tissue; (2) to reinforce inadequate tissue; (3) to induce new supportive tissue; and (4) to consolidate and complement the insufficient surgical techniques. However, some debatable issues in use of the prosthetics remain. The use of prosthetics enables the simultaneous repair of all vaginal defects of POP and concomitant anti-incontinence surgery to be faster, easier and more precise. Nevertheless, great care should be devoted to the actual and theoretical short- and long-term risks, many of which have not been fully elucidated. Despite the lack of various ideal characteristics, the type I monofilament, macroporous polypropylene, has been suggested to have the lowest incidence of infection and erosion among the nonabsorbable prostheses. There is good evidence to support the use of nonabsorbable synthetic mesh for abdominal sacrocolpopexy, while the use of prostheses for repairing isolated anterior and posterior compartment defects remains controversial. There have been no long-term studies with sufficient patient numbers to prove whether synthetic or biological prostheses are superior during vaginal surgery. Tension-free vaginal mesh techniques with procedural kits are being adopted increasingly, despite the paucity of data. Although short-term follow-up studies have shown tension-free vaginal mesh to be a safe and effective technique to correct POP, anatomic and functional results of long-term follow-up studies, however, have not yet confirmed the effectiveness and safety. Mesh erosion remains a concern, with variable rates according to different materials and approaches. Newly developed prostheses offer an alternative option to pelvic reconstructive surgery. However, some questions remain: (1) Should prostheses be considered for primary repairs, secondary repairs, or solely in patients with risk factors for recurrence? (2) Which prosthetic material is better: synthetic or biological ones; absorbable or nonabsorbable ones? (3) Do the benefits of prosthetics in pelvic reconstructive surgery outweigh the risks of complications? These questions are explored and reports in the literature reviewed.


Protocol for in vitro somatic embryogenesis and regeneration of rice (Oryza sativa L.).
Development of highly efficient and reproducible plant regeneration system has tremendous potential to provide improved technology to assist in genetic transformation of indica rice cultivars for their further exploitation in selection. For the development of a highly reproducible regeneration system through somatic embryogenesis, mature embryos of highly popular rice cultivars i.e., Govind (for rainfed areas), Pusa Basmati-1 (aromatic basmati) and Jaya (for irrigated areas) were used. Optimum callus formation (%) to MS medium supplemented with 2, 4-D was obtained at 12.0 microM in Govind, 14.0 microM in Jaya and 15.0 microM in Pusa Basmati-1. All the cultivars showed good proliferation on MS medium without hormone. In Govind, highest embryogenic response was observed in MS medium supplemented with 2, 4-D (0.4 microM) + kinetin (0.4 microM), while in Pusa Basmati-1 with 2, 4-D (0.4 microM) + kinetin (2.0 microM) and in Jaya on hormone-free MS medium. Excellent embryo regeneration in Govind was observed on MS medium supplemented with low concentrations (1.1 microM) of BAP or hormone-free MS medium, while in Pusa Basmati-1 and Jaya embryogenesis was observed on MS medium supplemented with higher concentration of BAP (2.2 microM). Similarly, maximum plantlets with proliferated roots were observed in Govind on hormone-free MS medium, while in Pusa Basmati-1 and Jaya on MS medium supplemented with high concentration of NAA (4.0 microM). Developed plantlets were further successfully acclimatized and grown under pot culture up to maturity. Further the yield potential of in vitro developed plants was accessed at par to the direct seeded one under pot culture. Present, protocol standardizes somatic embryogenesis and efficient regeneration of agronomically important, high yielding and diverse indica rice cultivars which can be utilized as an efficient tool for molecular studies and genetic transformation in future.


Cyclosporin A markedly enhances superantigen-induced peripheral T cell deletion and inhibits anergy induction.
Cyclosporin A (CsA) is a well-known immunosuppressive agent that modulates immune tolerance in many ways. CsA can give rise to a state of long-term nonimmunosuppressed transplantation tolerance, but it can also aggravate autoimmune diseases, and provoke specific forms of autoimmunity. These effects, which are often paradoxical, remain largely unexplained. In this study, we investigated the effects of CsA on superantigen (superAg)-reactive peripheral T cells. The intravenous injection of either staphylococcal enterotoxin B (SEB), or Mls-1a cells into Mls-1b recipients, causes long-term in vitro nonresponsiveness (anergy) and partial elimination of the peripheral T cell receptor (TCR) V beta 8+/CD4+ and -V beta 6+/CD4+ T cell subsets, respectively. We report that CsA markedly enhances the peripheral elimination of SEB- and Mls-1a-reactive T cells such that up to 90% of the targeted CD4+/V beta subpopulations are deleted. The degree of deletion depends on the dose and the schedule of CsA administration, and the number of superAg injections. In situations where the extent of deletion is only moderate, we find that the remaining superAg-reactive T cells fail to develop anergy, unlike the T cells of control superAg-immunized mice. Higher doses of CsA are required to enhance T cell deletion (greater than or equal to 25 mg/kg/d, i.p.) than to impair anergy induction (greater than or equal to 6.25 mg/kg/d, i.p.). In view of these results, it appears that the degree of tolerance in CsA/superAg-treated mice depends on the balance between these opposing effects, i.e., enhancement of peripheral elimination versus the abrogation of anergy. The possibility of enhancing or preventing immune tolerance with a drug may have important clinical implications.


Proteomic response of marine invertebrate larvae to ocean acidification and hypoxia during metamorphosis and calcification.
Calcifying marine invertebrates with complex life cycles are particularly at risk to climate changes as they undergo an abrupt ontogenetic shift during larval metamorphosis. Although our understanding of the larval response to climate changes is rapidly advancing, the proteome plasticity involved in a compensatory response to climate change is still unknown. In this study, we investigated the proteomic response of metamorphosing larvae of the tubeworm Hydroides elegans, challenged with two climate change stressors, ocean acidification (OA; pH 7.6) and hypoxia (HYP; 2.8 mg O2 l(-1)), and with both combined. Using a two-dimensional gel electrophoresis (2-DE)-based approach coupled with mass spectrometry, we found that climate change stressors did not affect metamorphosis except under OA, but altered the larval proteome and phosphorylation status. Metabolism and various stress and calcification-related proteins were downregulated in response to OA. In OA and HYP combined, HYP restored the expression of the calcification-related proteins to the control levels. We speculate that mild HYP stress could compensate for the negative effects of OA. This study also discusses the potential functions of selected proteins that might play important roles in larval acclimation and adaption to climate change.


Impact of Diabetes Control on Subclinical Atherosclerosis: Analysis from Coronary Computed Tomographic Angiography Registry.
There are limited data on the impact of diabetes control on the risk of subclinical coronary atherosclerosis. We analyzed 6,434 consecutive asymptomatic individuals without previous history of coronary artery disease who underwent coronary computed tomographic angiography (CCTA) (mean age, 53.7±7.6 years and 4,694 men [73.0%]). The degree and extent of subclinical coronary atherosclerosis were assessed by CCTA, and ≥50% diameter stenosis was defined as significant. A cardiac event was defined as a composite of all-cause death, myocardial infarction, unstable angina, or coronary revascularization. Study participants were categorized as normal (n=5,319), controlled diabetes (glycosylated hemoglobin [HbA1c] <7%, n=747), or uncontrolled diabetes (HbA1c ≥7%, n=368), respectively. Compared with normal individuals, there were no statistically significant differences in the risk of for any atherosclerotic plaque (odds ratio [OR], 1.16; 95% confidence interval [CI], 0.98 to 1.38; P=0.086) and significant coronary artery stenosis (OR, 1.08; 95% CI, 0.82 to 1.42; P=0.583) in controlled diabetic individuals. In contrast, uncontrolled diabetic individuals had consistently higher risks of any atherosclerotic plaque (OR, 2.16; 95% CI, 1.70 to 2.75; P<0.001) and significant coronary artery stenosis (OR, 3.34; 95% CI, 2.52 to 4.43; P<0.001) than normal individuals. During a follow-up of median 5.4 years, there was no significant difference in cardiac events between normal and controlled diabetic individuals (P=0.365). However, uncontrolled diabetes was associated with an increased risk of cardiac events compared with normal individuals (P<0.001) and controlled diabetic individuals (P=0.023). Asymptomatic uncontrolled diabetes was associated with significant subclinical coronary atherosclerosis with subsequent high risk for cardiac events.


Reduction in behavior problems with omega-3 supplementation in children aged 8-16 years: a randomized, double-blind, placebo-controlled, stratified, parallel-group trial.
While limited evidence suggests that omega-3 supplementation may reduce antisocial behavior in children, studies have not reported on posttreatment follow-up and most treatment periods have been of short duration. This study tests the hypothesis that omega-3 supplementation over 6 months will reduce behavior problems in children both at the end of treatment and at 6 months post treatment. In this randomized, double-blind, placebo-controlled, stratified, parallel-group trial, a community sample of 8-16 year old children were randomized into a treatment group (N = 100) and a placebo-control group (N = 100). The supplementation consisted of a fruit drink containing 1 g/day of omega-3 or a placebo consisting of the same fruit drink without omega-3. Participants, caregivers, and research assistants were blinded to group assignment. The primary outcome measures of externalizing and internalizing behavior problems were reported by both caregivers and their children in a laboratory setting at 0 months (baseline), 6 months (end of treatment) and 12 months (6 months post treatment), together with the secondary outcome measures of parental antisocial behavior. Data were analyzed on an intention-to-treat basis including all participants. ClinicalTrials.gov: http://clinicaltrials.gov/ct2/show/NCT02016079?term=mauritius&rank=2 RESULTS: Significant group × time interactions were observed with the treatment group showing long-term improvements in child behavior problems. The average posttreatment effect size was d = -.59. Effects were documented for parent reports, but with the exception of proactive and reactive aggression, child-report data were nonsignificant. Parents whose children took omega-3 showed significant posttreatment reductions in their own antisocial and aggressive behavior. This improvement in caregiver behavior partly mediated the improvements observed in child behavior. Findings provide initial evidence that omega-3 supplementation can produce sustained reductions in externalizing and internalizing behavior problems. Results are the first to report improvements in caregiver behavior, and to establish this improvement as a part-mechanism for the efficacy of omega-3.


Tibial articular cartilage and meniscus geometries combine to influence female risk of anterior cruciate ligament injury.
Tibial plateau subchondral bone geometry has been associated with the risk of sustaining a non-contact ACL injury; however, little is known regarding the influence of the meniscus and articular cartilage interface geometry on risk. We hypothesized that geometries of the tibial plateau articular cartilage surface and meniscus were individually associated with the risk of non-contact ACL injury. In addition, we hypothesized that the associations were independent of the underlying subchondral bone geometry. MRI scans were acquired on 88 subjects that suffered non-contact ACL injuries (27 males, 61 females) and 88 matched control subjects that were selected from the injured subject's teammates and were thus matched on sex, sport, level of play, and exposure to risk of injury. Multivariate analysis of the female data revealed that increased posterior-inferior directed slope of the middle articular cartilage region and decreased height of the posterior horn of the meniscus in the lateral compartment were associated with increased risk of sustaining a first time, non-contact ACL injury, independent of each other and of the slope of the tibial plateau subchondral bone. No measures were independently related to risk of non-contact ACL injury among males.


Early clinical experience with networked system for promoting patient self-management.
To gain early experience with a networked system designed to assess a patient's adherence to oral medication and physiologic metrics in an ambulatory, at-home setting. Prospective, observational studies. This networked system for patient self-management consists of ingestible markers and a wearable, personal monitor. When a marker is ingested, it communicates to a monitor that time-stamps the ingestion and identifies the marker as unique. The monitor also records heart rate and activity. Data from third-party monitoring equipment (eg, sphygmomanometer, weight scale) can be integrated into the system. Collected data are summarized for patient and physician review. Directly observed ingestion (DOI) of placebo tablet markers was used to assess the system's technical performance. Markers were also coencapsulated with drugs to capture at-home adherence. A performance criterion of <95% was set as the objective for system performance. A total of 111 subjects ingested 7144 ingestible markers; 3298 were DOIs. The system's positive detection accuracy and negative detection accuracy in detecting ingested markers were 97.1% and 97.7%, respectively. It differentiated 100% of multiple drugs and doses taken simultaneously by type and by dose. Medication adherence was >85%. The most common adverse effect was mild skin rash from the monitor's electrodes. No definitive marker-related adverse effects were reported. The system appears to be safe and effective in capturing and integrating adherence and physiologic data. Efforts are under way to enhance system functionalities and refine user interfaces. By providing context-rich information, this system may enhance patient-provider collaboration.


Propranolol blocks cardiac and neuronal voltage-gated sodium channels.
Propranolol is a widely used, non-selective β-adrenergic receptor antagonist with proven efficacy in treating cardiovascular disorders and in the prevention of migraine headaches. At plasma concentrations exceeding those required for β-adrenergic receptor inhibition, propranolol also exhibits anti-arrhythmic ("membrane stabilizing") effects that are not fully explained by β-blockade. Previous in vitro studies suggested that propranolol may have local anesthetic effects. We directly tested the effects of propranolol on heterologously expressed recombinant human cardiac (NaV1.5) and brain (NaV1.1, NaV1.2, NaV1.3) sodium channels using whole-cell patch-clamp recording. We found that block was not stereospecific as we observed approximately equal IC50 values for tonic and use-dependent block by R-(+) and S-(-) propranolol (tonic block: R: 21.4 μM vs S: 23.6 μM; use-dependent block: R: 2.7 μM vs S: 2.6 μM). Metoprolol and nadolol did not block NaV1.5 indicating that sodium channel block is not a class effect of β-blockers. The biophysical effects of R-(+)-propranolol on NaV1.5 and NaV1.1 resembled that of the prototypical local anesthetic lidocaine including the requirement for a critical phenylalanine residue (F1760 in NaV1.5) in the domain 4 S6 segment. Finally, we observed that brain sodium channels exhibited less sensitivity to R-(+)-propranolol than NaV1.5 channels. Our findings establish sodium channels as targets for propranolol and may help explain some beneficial effects of the drug in treating cardiac arrhythmias, and may explain certain adverse central nervous system effects.