Individualized glycaemic targets and pharmacotherapy in type 2 diabetes.
The Global Partnership for Effective Diabetes Management, established to provide practical guidance to improve patient outcomes in diabetes, has developed and modified recommendations to improve glycaemic control in type 2 diabetes. The Global Partnership advocates an individualized therapeutic approach and, as part of the process to customize therapy, has previously identified specific type 2 diabetes patient subgroups that require special consideration. This article builds on earlier publications, expanding the scope of practical guidance to include newly diagnosed individuals with complications and women with diabetes in pregnancy. Good glycaemic control remains the cornerstone of managing type 2 diabetes, and plays a vital role in preventing or delaying the onset and progression of diabetic complications. Individualizing therapeutic goals and treatments to meet glycaemic targets safely and without delay remains paramount, in addition to a wider programme of care to reduce cardiovascular risk factors and improve patient outcomes.


Multiplex genomic test of mutation and fusion genes in small biopsy specimen of lung cancer.
We evaluated multiple oncogenic mutations and fusion genes in small specimen obtained by bronchoscopy. Eight patients with lung cancer were recruited, 3 small cell lung cancer, 3 non-small cell lung cancer, 1 adenocarcinoma and 1 squamous cell carcinoma. A median value of extracted RNA and DNA amounts from specimen was 1573 ng (range 367.5 to 8900) and 6700 ng (range 550 to 68000 ng), respectively. We applied amplicon sequencing panels that cover exon regions of 41 genes related to lung tumorigenesis as well as total 61 major variants of ALK, ROS, RET or NTRK1 fusion transcripts. Nineteen of 41 gene mutations were detected in our isolated DNAs of 8 patients. We could detect four to eleven mutations in each specimen; however the mutation combination in each 8 patients were different. The most common genetic alterations were TP53, KMT2D, MET, NOTCH2 and SETD2, which were detected in 4 to 6 patients. We did not detect fusion transcripts of ALK, ROS, RET and NTRK1 in every specimen. In conclusion, multiplex genomic test was performed on small amounts specimen of bronchoscopy biopsy with a 100% success rate. Such testing is considered to be able to assist physicians in matching patients with approved or experimental targeted treatments.


Traffic flow in the operating room: an explorative and descriptive study on air quality during orthopedic trauma implant surgery.
Understanding the protective potential of operating room (OR) ventilation under different conditions is crucial to optimizing the surgical environment. This study investigated the air quality, expressed as colony-forming units (CFU)/m(3), during orthopedic trauma surgery in a displacement-ventilated OR; explored how traffic flow and the number of persons present in the OR affects the air contamination rate in the vicinity of surgical wounds; and identified reasons for door openings in the OR. Data collection, consisting of active air sampling and observations, was performed during 30 orthopedic procedures. In 52 of the 91 air samples collected (57%), the CFU/m(3) values exceeded the recommended level of <10 CFU/m(3). In addition, the data showed a strongly positive correlation between the total CFU/m(3) per operation and total traffic flow per operation (r = 0.74; P = .001; n = 24), after controlling for duration of surgery. A weaker, yet still positive correlation between CFU/m(3) and the number of persons present in the OR (r = 0.22; P = .04; n = 82) was also found. Traffic flow, number of persons present, and duration of surgery explained 68% of the variance in total CFU/m(3) (P = .001). Traffic flow has a strong negative impact on the OR environment. The results of this study support interventions aimed at preventing surgical site infections by reducing traffic flow in the OR.


Protein-DNA interactions at the H4-site III upstream transcriptional element of a cell cycle regulated histone H4 gene: differences in normal versus tumor cells.
Upstream sequences of the H4 histone gene FO108 located between nt -418 to -213 are stimulatory for in vivo transcription. This domain contains one protein/DNA interaction site (H4-Site III) that binds factor H4UA-1. Based on methylation interference, copper-phenanthroline protection, and competition assays, we show that H4UA-1 interacts with sequences between nt -345 to -332 containing an element displaying sequence-similarity with the thyroid hormone response element (TRE). Using gel retardation assays, we also demonstrate that H4UA-1 binding activity is abolished at low concentrations of Zn2+ (0.75 mM), a characteristic shared with the thyroid hormone (TH) receptor DNA binding protein. Interestingly, phosphatase-treatment of nuclear proteins inhibits formation of the H4UA-1 protein/DNA complex, although a complex with higher mobility (H4UA-1b) can be detected; both complexes share identical protein-DNA contacts and competition behaviors. These findings suggest that phosphorylation may be involved in the regulation of H4-Site III protein/DNA interactions by directly altering protein/protein associations. H4-Site III interactions were examined in several cell culture systems during cell growth and differentiation. We find that H4UA-1 binding activity is present during the cell cycle of both normal diploid and transformed cells. However, during differentiation of normal diploid rat calvarial osteoblasts, we observe a selective loss of the H4UA-1/H4-Site III interaction, concomitant with an increase of the H4UA-1b/H4-Site III complex, indicating modifications in the heteromeric nature of protein/DNA interactions during downregulation of transcription at the cessation of proliferation. Transformed cells have elevated levels of H4UA-1, whereas H4UA-1b is predominantly present in normal diploid cells; this alteration in the ratio of H4UA-1 and H4UA-1b binding activities may reflect deregulation of H4-Site III interactions in transformed cells. We propose that H4-Site III interactions may contribute, together with protein/DNA interactions at proximal regulatory sequences, in determining the level of H4-FO108 histone gene transcription.


Epidemic forecasts as a tool for public health: interpretation and (re)calibration.
Recent studies have used Bayesian methods to predict timing of influenza epidemics many weeks in advance, but there is no documented evaluation of how such forecasts might support the day-to-day operations of public health staff. During the 2015 influenza season in Melbourne, Australia, weekly forecasts were presented at Health Department surveillance unit meetings, where they were evaluated and updated in light of expert opinion to improve their accuracy and usefulness. Predictive capacity of the model was substantially limited by delays in reporting and processing arising from an unprecedented number of notifications, disproportionate to seasonal intensity. Adjustment of the predictive algorithm to account for these delays and increased reporting propensity improved both current situational awareness and forecasting accuracy. Collaborative engagement with public health practitioners in model development improved understanding of the context and limitations of emerging surveillance data. Incorporation of these insights in a quantitative model resulted in more robust estimates of disease activity for public health use. Implications for public health: In addition to predicting future disease trends, forecasting methods can quantify the impact of delays in data availability and variable reporting practice on the accuracy of current epidemic assessment. Such evidence supports investment in systems capacity.


Childhood cancer in El Salvador: A preliminary exploration of parental concerns in the abandonment of treatment.
In El Salvador, children under 12 diagnosed with cancer have access to free treatment at a specialized national facility. Until recently, 13 percent of patients annually abandoned therapy--a serious loss of lives and scarce resources. This qualitative study explores how some parents perceived their child's cancer and treatment, and what led them to stop bringing their child for chemotherapy. In in-depth interviews, parents of six children who abandoned their child's cancer treatment discussed sickness and life circumstances during the course of treatment. Poverty, effects of treatment, mistrust, emotions and religious convictions all figured in the parents' explanation of their actions. However, each family weighed these concerns differently. It was the interaction of the concerns, and not the concern per se, that represented the explanatory frameworks the families used to explain stopping their child's treatment. This finding illustrates the parents' navigation among a collection of variable concerns, rather than exposing one fixed cause for their behavior. For example, poverty affects a parent's worldview as well as concrete living conditions, and therefore has a complex relationship with abandonment of treatment. Thus, it follows that strategies to reduce treatment abandonment (and increase a child's chance for survival) must be multidimensional. Qualitative studies of how families perceive childhood cancer and treatment can illuminate the processes and relationships involved in abandonment of treatment. This approach can also show how families' living circumstances frame their perceptions and inform strategies to improve how medical services are provided, thus reducing abandonment of treatment.


Reactivity of the organometallic fac-[(CO)3ReI(H2O)3]+ aquaion. Kinetic and thermodynamic properties of H2O substitution.
The water exchange process on [(CO)(3)Re(H(2)O)(3)](+) (1) was kinetically investigated by (17)O NMR. The acidity dependence of the observed rate constant k(obs) was analyzed with a two pathways model in which k(ex) (k(ex)(298) = (6.3 +/- 0.1) x 10(-3) s(-1)) and k(OH) (k(OH)(298)= 27 +/- 1 s(-1)) denote the water exchange rate constants on 1 and on the monohydroxo species [(CO)(3)Re(I)(H(2)O)(2)(OH)], respectively. The kinetic contribution of the basic form was proved to be significant only at [H(+)] < 3 x 10(-3) M. Above this limiting [H(+)] concentration, kinetic investigations can be unambiguously conducted on the triaqua cation (1). The variable temperature study has led to the determination of the activation parameters Delta H(++)(ex) = 90 +/- 3 kJ mol(-1), Delta S(++)(ex) = +14 +/- 10 J K(-1) mol(-1), the latter being indicative of a dissociative activation mode for the water exchange process. To support this assumption, water substitution reaction on 1 has been followed by (17)O/(1)H/(13)C/(19)F NMR with ligands of various nucleophilicities (TFA, Br(-), CH(3)CN, Hbipy(+), Hphen(+), DMS, TU). With unidentate ligands, except Br(-), the mono-, bi-, and tricomplexes were formed by water substitution. With bidentate ligands, bipy and phen, the chelate complexes [(CO)(3)Re(H(2)O)(bipy)]CF(3)SO(3) (2) and [(CO)(3)Re(H(2)O)(phen)](NO(3))(0.5)(CF(3)SO(3))(0.5).H(2)O (3) were isolated and X-ray characterized. For each ligand, the calculated interchange rate constants k'(i) (2.9 x 10(-3) (TFA) < k'(I) < 41.5 x 10(-3) (TU) s(-1)) were found in the same order as the water exchange rate constant k(ex), the S-donor ligands being slightly more reactive. This result is indicative of I(d) mechanism for water exchange and complex formation, since larger variations of k'(i) are expected for an associatively activated mechanism.


Electrophysiology and plasticity in isolated postsynaptic densities.
The organization and regulation of excitatory synapses in the mammalian CNS entails complex molecular and cellular processes. In the postsynaptic membrane, scaffolding proteins bring together glutamate receptors with multiple regulatory proteins involved in signal transduction. This gives rise to an elaborate postsynaptic structure known as the postsynaptic density (PSD). This protein network plays a critical role in the regulation of glutamate receptor function and thus in synaptic plasticity. To study this regulation, we have developed a system in which ionotropic glutamate receptors (iGluRs) can be recorded, in the steady state, by the patch clamp technique in isolated PSDs incorporated into giant liposomes. In this preparation, ionotropic glutamate receptors maintain their characteristic physiological and pharmacological properties. The recordings reflect the presence of channel clusters, as multiple conductance and subconductance states are observed. Each of the receptor subtypes is activated by a specific set of kinases that are activated differentially by Ca(2+): the "kainate receptor kinases" are active even in the presence of EGTA, i.e. they are not calcium-dependent; the "N-methyl-D-aspartate receptor (NMDAR) channel kinases" are active in the presence of submicromolar calcium concentrations, whereas the "alpha-amino-3- hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptor kinases" need microM calcium for activation. The NMDA receptor showed its characteristic voltage-dependent Mg(2+) blockade, and activation by phosphorylation was in part a consequence of a relief of Mg(2+) blockade. These results allow us to propose a model in which phosphorylation of NMDA receptors can contribute to a long-lasting and self-maintained change in synaptic function. The experimental approach we present will allow us to test the functional consequence of activation of the multiple signal transduction pathways thought to regulate excitatory neurotransmission in the adult CNS.


Climate and human water use diminish wetland networks supporting continental waterbird migration.
Migrating waterbirds moving between upper and lower latitudinal breeding and wintering grounds rely on a limited network of endorheic lakes and wetlands when crossing arid continental interiors. Recent drying of global endorheic water stores raises concerns over deteriorating migratory pathways, yet few studies have considered these effects at the scale of continental flyways. Here, we investigate the resiliency of waterbird migration networks across western North America by reconstructing long-term patterns (1984-2018) of terminal lake and wetland surface water area in 26 endorheic watersheds. Findings were partitioned regionally by snowmelt- and monsoon-driven hydrologies and combined with climate and human water-use data to determine their importance in predicting surface water trends. Nonlinear patterns of lake and wetland drying were apparent along latitudinal flyway gradients. Pervasive surface water declines were prevalent in northern snowmelt watersheds (lakes -27%, wetlands -47%) while largely stable in monsoonal watersheds to the south (lakes -13%, wetlands +8%). Monsoonal watersheds represented a smaller proportion of total lake and wetland area, but their distribution and frequency of change within highly arid regions of the continental flyway increased their value to migratory waterbirds. Irrigated agriculture and increasing evaporative demands were the most important drivers of surface water declines. Underlying agricultural and wetland relationships however were more complex. Approximately 7% of irrigated lands linked to flood irrigation and water storage practices supported 61% of all wetland inundation in snowmelt watersheds. In monsoonal watersheds, small earthen dams, meant to capture surface runoff for livestock watering, were a major component of wetland resources (67%) that supported networks of isolated wetlands surrounding endorheic lakes. Ecological trends and human impacts identified herein underscore the importance of assessing flyway-scale change as our model depictions likely reflect new and emerging bottlenecks to continental migration.


Genetic variants in hormone-related genes and risk of breast cancer.
Sex hormones play a key role in the development of breast cancer. Certain polymorphic variants (SNPs and repeat polymorphisms) in hormone-related genes are associated with sex hormone levels. However, the relationship observed between these genetic variants and breast cancer risk has been inconsistent. We conducted a case-control study nested within two prospective cohorts to assess the relationship between specific genetic variants in hormone-related genes and breast cancer risk. In total, 1164 cases and 2111 individually-matched controls were included in the study. We did not observe an association between potential functional genetic polymorphisms in the estrogen pathway, SHBG rs6259, ESR1 rs2234693, CYP19 rs10046 and rs4775936, and UGT1A1 rs8175347, or the progesterone pathway, PGR rs1042838, with the risk of breast cancer. Our results suggest that these genetic variants do not have a strong effect on breast cancer risk.


Common and distinct neural substrates of attentional control in an integrated Simon and spatial Stroop task as assessed by event-related fMRI.
The purpose of this experiment was to directly examine the neural mechanisms of attentional control involved in the Simon task as compared to a spatial Stroop task using event-related fMRI. The Simon effect typically refers to the interference people experience when there is a stimulus-response conflict. The Stroop effect refers to the interference people experience when two attributes of the same stimulus conflict with each other. Although previous imaging studies have compared the brain activation for each of these tasks performed separately, none had done so in an integrated task that incorporates both types of interference, as was done in the current experiment. Both tasks activated brain regions that serve as a source of attentional control (dorsolateral prefrontal cortex) and posterior regions that are sites of attentional control (the visual processing stream-middle occipital and inferior temporal cortices). In addition, there were also specific brain regions activated to a significantly greater degree by one task and/or only by a single task. The brain regions significantly more activated by the Simon task were those sensitive to detection of response conflict, response selection, and planning (anterior cingulate cortex, supplementary motor areas, and precuneus), and visuospatial-motor association areas. In contrast, the regions significantly more activated by the Stroop task were those involved in biasing the processing toward the task-relevant attribute (inferior parietal cortex). These findings suggest that the interference effects of these two tasks are caused by different types of conflict (stimulus-response conflict for the Simon effect and stimulus-stimulus conflict for the Stroop effect) but both invoke similar sources of top-down modulation.


Binding of NIR-conPK and NIR-6T to astrocytomas and microglial cells: evidence for a protein related to TSPO.
PK 11195 and DAA1106 bind with high-affinity to the translocator protein (TSPO, formerly known as the peripheral benzodiazepine receptor). TSPO expression in glial cells increases in response to cytokines and pathological stimuli. Accordingly, [(11)C]-PK 11195 and [(11)C]-DAA1106 are recognized molecular imaging (MI) agents capable of monitoring changes in TSPO expression occurring in vivo and in response to various neuropathologies.Here we tested the pharmacological characteristics and TSPO-monitoring potential of two novel MI agents: NIR-conPK and NIR-6T. NIR-conPK is an analogue of PK 11195 conjugated to the near-infrared (NIR) emitting fluorophore: IRDye 800CW. NIR-6T is a DAA1106 analogue also conjugated to IRDye 800CW.We found that NIR-6T competed for [(3)H]-PK 11195 binding in astrocytoma cell homogenates with nanomolar affinity, but did not exhibit specific binding in intact astrocytoma cells in culture, indicating that NIR-6T is unlikely to constitute a useful MI agent for monitoring TSPO expression in intact cells. Conversely, we found that NIR-conPK did not compete for [(3)H]-PK 11195 binding in astrocytoma cell homogenate, but exhibited specific binding in intact astrocytoma cells in culture with nanomolar affinity, suggesting that NIR-conPK binds to a protein distinct, but related to, TSPO. Accordingly, treating intact astrocytoma cells and microglia in culture with cytokines led to significant changes in the amount of NIR-conPK specific binding without corresponding change in TSPO expression. Remarkably, the cytokine-induced changes in the protein targeted by NIR-conPK in intact microglia were selective, since IFN-gamma (but not TNFalpha and TGFbeta) increased the amount of NIR-conPK specific binding in these cells.Together these results suggest that NIR-conPK binds to a protein that is related to TSPO, and expressed by astrocytomas and microglia. Our results also suggest that the expression of this protein is increased by specific cytokines, and thus allows for the monitoring of a particular subtype of microglia activation.


Dynamics of acquisition and loss of carriage of Staphylococcus aureus strains in the community: the effect of clonal complex.
Staphylococcus aureus nasal carriage increases infection risk. However, few studies have investigated S. aureus acquisition/loss over >1 year, and fewer still used molecular typing. 1123 adults attending five Oxfordshire general practices had nasal swabs taken. 571 were re-swabbed after one month then every two months for median two years. All S. aureus isolates were spa-typed. Risk factors were collected from interviews and medical records. 32% carried S. aureus at recruitment (<1% MRSA). Rates of spa-type acquisition were similar in participants S. aureus positive (1.4%/month) and negative (1.8%/month, P = 0.13) at recruitment. Rates were faster in those carrying clonal complex (CC)15 (adjusted (a)P = 0.03) or CC8 (including USA300) (aP = 0.001) at recruitment versus other CCs. 157/274 (57%) participants S. aureus positive at recruitment returning ≥ 12 swabs carried S. aureus consistently, of whom 135 carried the same spa-type. CC22 (including EMRSA-15) was more prevalent in long-term than intermittent spa-type carriers (aP = 0.03). Antibiotics transiently reduced carriage, but no other modifiable risk factors were found. Both transient and longer-term carriage exist; however, the approximately constant rates of S. aureus gain and loss suggest that 'never' or truly 'persistent' carriage are rare. Long-term carriage varies by strain, offering new explanations for the success of certain S. aureus clones.


Reversible phosphorylation differentially affects nuclear and cytoplasmic functions of splicing factor 2/alternative splicing factor.
The Ser/Arg-rich (SR) proteins constitute a family of highly conserved nuclear phosphoproteins that are involved in many steps of mRNA metabolism. Previously, we demonstrated that shuttling SR proteins can associate with translating ribosomes and enhance translation of reporter mRNAs both in vivo and in vitro. Here, we show that endogenous, cytoplasmic splicing factor 2/alternative splicing factor (SF2/ASF) associated with the translation machinery is hypophosphorylated, suggesting that the phosphorylation state of the Arg-Ser-rich (RS) domain may influence the role of SF2/ASF in cytoplasmic RNA processing. In agreement, we show that mutations mimicking a hypophosphorylated RS domain strongly increased SF2/ASF binding to cytoplasmic mRNA and its activity in translation. We also demonstrate that, whereas the RS domain is not required for the function of SF2/ASF in mRNA translation in vivo or in vitro, its second RNA recognition motif (RRM)2 plays a critical role in this process. Taken together, these data suggest that RS-domain phosphorylation may influence the association of SF2/ASF with mRNA, whereas RRM2 may play an important role in mediating protein-protein interactions during translation. These data are consistent with a model whereby reversible protein phosphorylation differentially regulates the subcellular localization and activity of shuttling SR proteins.


A phase 2 study of MK-5442, a calcium-sensing receptor antagonist, in postmenopausal women with osteoporosis after long-term use of oral bisphosphonates.
In women with osteoporosis treated with alendronate for >12 months and oral bisphosphonates for >3 of the last 4 years, switching to MK-5442, a calcium receptor antagonist, stimulated endogenous parathyroid hormone (PTH) secretion and increased bone turnover marker levels, but produced a decline in bone mineral density (BMD) at all sites. This study assessed the effects of switching from long-term oral bisphosphonate therapy to the calcium-sensing receptor antagonist MK-5442 on BMD and bone turnover markers (BTMs) in post-menopausal women with osteoporosis. This randomized, active and placebo-controlled, dose-ranging study enrolled 526 postmenopausal women, who had taken alendronate (ALN) for ≥12 months preceding the trial and any oral bisphosphonate for ≥3 of the preceding 4 years and had spine or hip BMD T-scores ≤-2.5 or ≤-1.5 with ≥1 prior fragility fracture. Women were randomized to continue ALN 70 mg weekly or switch to MK-5442 (5, 7.5, 10, or 15 mg daily) or placebo. Switching from ALN to MK-5442 produced a dose-dependent parathyroid hormone (PTH) pulse of threefold to sixfold above baseline at 1 h, with PTH levels that remained twofold to threefold above baseline at 4 h and returned to baseline by 24 h. Switching to MK-5442 or placebo increased BTM levels compared to baseline within 3 months and MK-5442 10 mg increased BTM levels compared to placebo by 6 months. With all MK-5442 doses and placebo, spine and hip BMD declined from baseline, and at 12 months, BMD levels were below those who continued ALN (all groups P < 0.05 vs ALN). There was also a dose-dependent increase in the incidence of hypercalcemia with MK-5442. Switching from ALN to MK-5442 resulted in a pulsatile increase in PTH and increases in BTMs, but a decline in BMD compared with continued ALN. MK-5442 is not a viable option for the treatment of osteoporosis.


Hyperoxic treatment induces mesenchymal-to-epithelial transition in a rat adenocarcinoma model.
Tumor hypoxia is relevant for tumor growth, metabolism and epithelial-to-mesenchymal transition (EMT). We report that hyperbaric oxygen (HBO) treatment induced mesenchymal-to-epithelial transition (MET) in a dimethyl-alpha-benzantracene induced mammary rat adenocarcinoma model, and the MET was associated with extensive coordinated gene expression changes and less aggressive tumors. One group of tumor bearing rats was exposed to HBO (2 bar, pO(2) = 2 bar, 4 exposures à 90 minutes), whereas the control group was housed under normal atmosphere (1 bar, pO(2) = 0.2 bar). Treatment effects were determined by assessment of tumor growth, tumor vascularisation, tumor cell proliferation, cell death, collagen fibrils and gene expression profile. Tumor growth was significantly reduced (approximately 16%) after HBO treatment compared to day 1 levels, whereas control tumors increased almost 100% in volume. Significant decreases in tumor cell proliferation, tumor blood vessels and collagen fibrils, together with an increase in cell death, are consistent with tumor growth reduction and tumor stroma influence after hyperoxic treatment. Gene expression profiling showed that HBO induced MET. In conclusion, hyperoxia induced MET with coordinated expression of gene modules involved in cell junctions and attachments together with a shift towards non-tumorigenic metabolism. This leads to more differentiated and less aggressive tumors, and indicates that oxygen per se might be an important factor in the "switches" of EMT and MET in vivo. HBO treatment also attenuated tumor growth and changed tumor stroma, by targeting the vascular system, having anti-proliferative and pro-apoptotic effects.


Induced expression of nucleolin phosphorylation-deficient mutant confers dominant-negative effect on cell proliferation.
Nucleolin (NCL) is a major nucleolar phosphoprotein that has pleiotropic effects on cell proliferation and is elevated in a variety of tumors. NCL is highly phosphorylated at the N-terminus by two major kinases: interphase casein kinase 2 (CK2) and mitotic cyclin-dependent kinase 1 (CDK1). Earlier we demonstrated that a NCL-mutant that is partly defective in undergoing phosphorylation by CK2 inhibits chromosomal replication through its interactions with Replication Protein A, mimicking the cellular response to DNA damage. We further delineated that the N-terminus of NCL associates with Hdm2, the most common E3 ubiquitin ligase of p53. We reported that NCL antagonizes Hdm2 to stabilize p53 and stimulates p53 transcriptional activity. Although NCL-phosphorylation by CK2 and ribosomal DNA transcription are closely coordinated during interphase, the role of NCL phosphorylation in regulating cell proliferation remains unexplored. We have therefore engineered unique human cells that specifically induce expression of NCL-wild type (WT) or a phosphorylation-deficient NCL-mutant, 6/S*A where all the six CK2 consensus serine sites residing in the N-terminus NCL were mutated to alanine. Here we show that this NCL-mutant is defective in undergoing phosphorylation by CK2. We also demonstrate that NCL-phosphorylation by CK2 is required through the S-phase progression in cell cycle and hence proliferation. Induced expression of NCL with mutated CK2 phosphorylation sites stabilizes p53, results in higher expression of Bcl2 (B-cell lymphoma 2) homology 3 (BH3)-only apoptotic markers and causes a dominant-negative effect on cell viability. Our unique cellular system thus provides the first evidential support to delineate phospho-specific functions of NCL on cell proliferation.


Cloning and characterization of a Lambert-Eaton myasthenic syndrome antigen.
Lambert-Eaton myasthenic syndrome is a paraneoplastic neuromuscular disorder in which an immune response directed against a small-cell lung tumor crossreacts with antigens in the neuromuscular junction. To isolate and characterize the antigens, we screened a human fetal brain expression library with a high-titer serum from a patient with Lambert-Eaton myasthenic syndrome. This screening resulted in the isolation of a complementary DNA clone encoding an antigen we call myasthenic syndrome antigen B (MysB). Approximately 43% (3 of 7) of Lambert-Eaton myasthenic syndrome sera specifically recognized MysB fusion protein, whereas none of 34 control sera did. The predicted amino acid sequence of this clone shows a high degree of homology to the beta subunit of calcium channel complexes. The MysB pre-messenger RNA is alternatively spliced to yield 3 forms of the protein differing in the domain between two highly conserved alpha-helical segments.


Contributions of the Ventral Striatum to Conscious Perception: An Intracranial EEG Study of the Attentional Blink.
The brain is limited in its capacity to consciously process information, necessitating gating of information. While conscious perception is robustly associated with sustained, recurrent interactions between widespread cortical regions, subcortical regions, including the striatum, influence cortical activity. Here, we examined whether the ventral striatum, given its ability to modulate cortical information flow, contributes to conscious perception. Using intracranial EEG, we recorded ventral striatum activity while 7 patients performed an attentional blink task in which they had to detect two targets (T1 and T2) in a stream of distractors. Typically, when T2 follows T1 within 100-500 ms, it is often not perceived (i.e., the attentional blink). We found that conscious T2 perception was influenced and signaled by ventral striatal activity. Specifically, the failure to perceive T2 was foreshadowed by a T1-induced increase in α and low β oscillatory activity as early as 80 ms after T1, indicating that the attentional blink to T2 may be due to very early T1-driven attentional capture. Moreover, only consciously perceived targets were associated with an increase in θ activity between 200 and 400 ms. These unique findings shed new light on the mechanisms that give rise to the attentional blink by revealing that conscious target perception may be determined by T1 processing at a much earlier processing stage than traditionally believed. More generally, they indicate that ventral striatum activity may contribute to conscious perception, presumably by gating cortical information flow. What determines whether we become aware of a piece of information or not? Conscious access has been robustly associated with activity within a distributed network of cortical regions. Using intracranial electrophysiological recordings during an attentional blink task, we tested the idea that the ventral striatum, because of its ability to modulate cortical information flow, may contribute to conscious perception. We find that conscious perception is influenced and signaled by ventral striatal activity. Short-latency (80-140 ms) striatal responses to a first target determined conscious perception of a second target. Moreover, conscious perception of the second target was signaled by longer-latency (200-400 ms) striatal activity. These results suggest that the ventral striatum may be part of a subcortical network that influences conscious experience.


Clinical Application of Prognostic Gene Expression Signature in Fusion Gene-Negative Rhabdomyosarcoma: A Report from the Children's Oncology Group.
Pediatric rhabdomyosarcoma (RMS) has two common histologic subtypes: embryonal (ERMS) and alveolar (ARMS). PAX-FOXO1 fusion gene status is a more reliable prognostic marker than alveolar histology, whereas fusion gene-negative (FN) ARMS patients are clinically similar to ERMS patients. A five-gene expression signature (MG5) previously identified two diverse risk groups within the fusion gene-negative RMS (FN-RMS) patients, but this has not been independently validated. The goal of this study was to test whether expression of the MG5 metagene, measured using a technical platform that can be applied to routine pathology material, would correlate with outcome in a new cohort of patients with FN-RMS. Cases were taken from the Children's Oncology Group (COG) D9803 study of children with intermediate-risk RMS, and gene expression profiling for the MG5 genes was performed using the nCounter assay. The MG5 score was correlated with clinical and pathologic characteristics as well as overall and event-free survival. MG5 standardized score showed no significant association with any of the available clinicopathologic variables. The MG5 signature score showed a significant correlation with overall (N = 57; HR, 7.3; 95% CI, 1.9-27.0; P = 0.003) and failure-free survival (N = 57; HR, 6.1; 95% CI, 1.9-19.7; P = 0.002). This represents the first, validated molecular prognostic signature for children with FN-RMS who otherwise have intermediate-risk disease. The capacity to measure the expression of a small number of genes in routine pathology material and apply a simple mathematical formula to calculate the MG5 metagene score provides a clear path toward better risk stratification in future prospective clinical trials.


Snoring as a risk factor for disease: an epidemiological survey.
In a study conducted in four family practice units in Toronto, Canada, 2001 subjects reported on snoring and medical conditions in members of their households. For spouses the prevalence of snoring increased with age up to the seventh decade, with a higher prevalence of nearly 85% in husbands. For 11 medical problems an association existed between snoring, its frequency, and the presence of the condition. This association continued when the data were corrected for sex, age, and marital state. For hypertension both men and women who snored between the fifth and 10th decades had a twofold increase over non-snorers. The prevalence of heart disease and other conditions, except for diabetes and asthma, also increased in snorers in this age group. When corrected for smoking and obesity the association between snoring, hypertension, and heart disease persisted. These findings extend those of Lugaresi et al, and if they could be confirmed snoring as a risk factor for conditions other than sleep apnoea and sleep disorders might be considered. Methods of alleviating the acoustic annoyance of snoring may also provide direct medical benefits.


Synthesis, characterization and evaluation of novel triblock copolymer based nanoparticles for vaccine delivery against hepatitis B.
Poly lactic acid (PLA) is one of widely used biodegradable polymer in vaccine delivery. However, the use is restricted due to hydrophobic nature and generation of acidic microenvironment upon its degradation, rendering it unfavorable to the encapsulated antigen. In the present study we have synthesized PEG derivatized block copolymers of PLA for development of nanoparticles encapsulating HBsAg for mucosal vaccination against hepatitis B. The copolymers of compositions AB, ABA and BAB (PLA as A-block and PEG as B-block) were synthesized and characterized by 1H NMR spectroscopy and gel permeation chromatography. Nanoparticles were characterized to determine the effect of copolymer. Among all, BAB produced nanoparticles of smallest size and lowest zeta potential, suggesting highest PEG density on their surface. The in vitro release experiments were performed in PBS (pH7.4). SDS-PAGE analysis confirmed the structural stability and integrity of the released antigen. Results were compared for immunogenicity with plain PLA nanoparticles and conventional alum-HBsAg based vaccine. BAB nanoparticles produced better humoral response as compared to other polymeric nanoparticles. The extent of humoral response obtained in single dose of BAB nanoparticles was comparable to the response produced by alum based vaccine (which received a booster dose). Block copolymeric nanoparticles also produced better sIgA level at all local and distal mucosal sites as compare of PLA nanoparticles, where alum based formulation failed to give any considerable response. Additionally, IgG1 and IgG2a isotype were determined to confirm the T(H)1/T(H)2 mixed immune response. These data demonstrate the potential of BAB nanoparticles as mucosal vaccine delivery system capable of eliciting high and prolonged immune response.


Adsorption behavior and adhesive properties of biopolyelectrolyte multilayers formed from cationic and anionic starch.
Cationic starch (D.S. 0.065) and anionic starch (D.S. 0.037) were used to form biopolyelectrolyte multilayers. The influence of the solution concentration of NaCl on the adsorption of starch onto silicon oxide substrates and on the formation of multilayers was investigated using stagnation point adsorption reflectometry (SPAR) and quartz crystal microbalance with dissipation (QCM-D). The wet adhesive properties of the starch multilayers were examined by measuring pull-off forces with the AFM colloidal probe technique. It was shown that polyelectrolyte multilayers (PEM) can be successfully constructed from cationic starch and anionic starch at electrolyte concentrations of 1 mM NaCl and 10 mM NaCl. The water content of the PEMs was approximately 80% at both electrolyte concentrations. However, the thickness of the PEMs formed at 10 mM NaCl was approximately twice the thickness formed at 1 mM NaCl. The viscoelastic properties of the starch PEMs, modeled as Voigt elements, were dependent on the polyelectrolyte that was adsorbed in the outermost layer. The PEMs appeared to be more rigid when capped by anionic starch than when capped by cationic starch. The wet adhesive pull-off forces increased with layer number and were also dependent on the polyelectrolyte adsorbed in the outermost layer. Thus, starch PEM treatment has a large potential for increasing the adhesive interaction between solid substrates to levels higher than can be reached by a single layer of cationic starch.


The mammalian gene function resource: the International Knockout Mouse Consortium.
In 2007, the International Knockout Mouse Consortium (IKMC) made the ambitious promise to generate mutations in virtually every protein-coding gene of the mouse genome in a concerted worldwide action. Now, 5 years later, the IKMC members have developed high-throughput gene trapping and, in particular, gene-targeting pipelines and generated more than 17,400 mutant murine embryonic stem (ES) cell clones and more than 1,700 mutant mouse strains, most of them conditional. A common IKMC web portal (www.knockoutmouse.org) has been established, allowing easy access to this unparalleled biological resource. The IKMC materials considerably enhance functional gene annotation of the mammalian genome and will have a major impact on future biomedical research.


Measures assessing spirituality as more than religiosity: a methodological review of nursing and health-related literature.
This paper is a report of a methodological review conducted to analyse, evaluate and synthesize the rigour of measures found in nursing and health-related literature used to assess and evaluate patient spirituality as more than religiosity. Holistic healthcare practitioners recognize important distinctions exist about what constitutes spiritual care needs and preferences and what constitutes religious care needs and preferences in patient care practice. Databases searched, limited to the years 1982 and 2009, included AMED, Alt Health Watch, CINAHL Plus with Full Text, EBSCO Host, EBSCO Host Religion and Philosophy, ERIC, Google Scholar, HAPI, HUBNET, IngentaConnect, Mental Measurements Yearbook Online, Ovid MEDLINE, Social Work Abstracts and Hill and Hood's Measures of Religiosity text. A methodological review was carried out. Measures assessing spirituality as more than religiosity were critically reviewed including quality appraisal, relevant data extraction and a narrative synthesis of findings. Ten measures fitting inclusion criteria were included in the review. Despite agreement among nursing and health-related disciplines that spirituality and religiosity are distinct and diverse concepts, the concept of spirituality was often used interchangeably with the concept religion to assess and evaluate patient spirituality. The term spiritual or spirituality was used in a preponderance of items to assess or evaluate spirituality. Measures differentiating spirituality from religiosity are grossly lacking in nursing and health-related literature.


Expression of hepatocyte growth factor and its receptor c-met proto-oncogene in hepatocellular carcinoma.
The c-met proto-oncogene encodes the tyrosine kinase receptor for hepatocyte growth factor (HGF), a potent mitogen and motogen for epithelial cells. Because of its profound effects on cell growth and motility, HGF may be important in the development of cancer metastases in hepatocellular carcinoma (HCC). In this study, we examined HGF concentration and expression of the c-met-proto-oncogene product (c-met) in 62 patients with HCC to determine the relationship between the level of expression and clinicopathological features, and patient outcome following hepatectomy. Western blotting was used to examine the c-met expression, and HGF concentration in tumors was measured using an enzyme-linked immunosorbent assay. c-met was found to be overexpressed in HCC compared with nontumorous liver tissue (P < .01), and correlated with an increased incidence of intrahepatic metastases (P = .039). Patients were divided into two groups, low c-met HCC and high c-met HCC. Patients with high c-met HCC had a significantly shorter 5-year survival than patients with low c-met HCC (33.5% vs. 80.3%, respectively; P < .05). However, there was no correlation between HGF concentration in the tumor tissue and clinicopathological factors and patient survival. These results indicate that expression of c-met played an important role in tumor growth and metastases in patients who underwent hepatectomy for HCC.


Vigilance, visual search and attention in an agricultural task.
In a fragile agricultural environment, such as Western Australia (WA), introduced exotic plant species present a serious environmental and economic threat. Skeleton weed, centaurea juncea, a Mediterranean daisy, was accidentally introduced into WA in 1963. It competes with cash crops such as wheat. When observed in the fields, farms are quarantined and mechanised teams search for the infestations in order to destroy them. Since the search process requires attention, visual search and vigilance, the present investigators conducted a number of controlled field trials to identify the importance of these factors in detection of the weed. The paper describes the basic hit rate, vigilance decrement, effect of search party size, effect of target size, and some data on the effect of solar illumination of the target. Several recommendations have been made and incorporated in the search programme and some laboratory studies undertaken to answer questions arising.


Comparative efficacy of recombinant modified vaccinia virus Ankara expressing simian immunodeficiency virus (SIV) Gag-Pol and/or Env in macaques challenged with pathogenic SIV.
Prior studies demonstrated that immunization of macaques with simian immunodeficiency virus (SIV) Gag-Pol and Env recombinants of the attenuated poxvirus modified vaccinia virus Ankara (MVA) provided protection from high levels of viremia and AIDS following challenge with a pathogenic strain of SIV (V. M. Hirsch et al., J. Virol. 70:3741-3752, 1996). This MVA-SIV recombinant expressed relatively low levels of the Gag-Pol portion of the vaccine. To optimize protection, second-generation recombinant MVAs that expressed high levels of either Gag-Pol (MVA-gag-pol) or Env (MVA-env), alone or in combination (MVA-gag-pol-env), were generated. A cohort of 24 macaques was immunized with recombinant or nonrecombinant MVA (four groups of six animals) and was challenged with 50 times the dose at which 50% of macaques are infected with uncloned pathogenic SIVsmE660. Although all animals became infected postchallenge, plasma viremia was significantly reduced in animals that received the MVA-SIV recombinant vaccines as compared with animals that received nonrecombinant MVA (P = 0.0011 by repeated-measures analysis of variance). The differences in the degree of virus suppression achieved by the three MVA-SIV vaccines were not significant. Most importantly, the reduction in levels of viremia resulted in a significant increase in median (P < 0.05 by Student's t test) and cumulative (P = 0.010 by log rank test) survival. These results suggest that recombinant MVA has considerable potential as a vaccine vector for human AIDS.


The human RAD54 recombinational DNA repair protein is a double-stranded DNA-dependent ATPase.
DNA double-strand break repair through the RAD52 homologous recombination pathway in the yeast Saccharomyces cerevisiae requires, among others, the RAD51, RAD52, and RAD54 genes. The biological importance of homologous recombination is underscored by the conservation of the RAD52 pathway from fungi to humans. The critical roles of the RAD52 group proteins in the early steps of recombination, the search for DNA homology and strand exchange, are now becoming apparent. Here, we report the purification of the human Rad54 protein. We showed that human Rad54 has ATPase activity that is absolutely dependent on double-stranded DNA. Unexpectedly, the ATPase activity appeared not absolutely required for the DNA repair function of human Rad54 in vivo. Despite the presence of amino acid sequence motifs that are conserved in a large family of DNA helicases, no helicase activity of human Rad54 was observed on a variety of different DNA substrates. Possible functions of human Rad54 in homologous recombination that couple the energy gained from ATP hydrolysis to translocation along DNA, rather than disruption of base pairing, are discussed.


Efficient production of transgenic citrus plants using isopentenyl transferase positive selection and removal of the marker gene by site-specific recombination.
The presence of marker genes conferring antibiotic resistance in transgenic plants represents a serious obstacle for their public acceptance and future commercialization. In citrus, selection using the selectable marker gene nptII, that confers resistance to the antibiotic kanamycin, is in general very effective. An attractive alternative is offered by the MAT system (Multi-Auto-Transformation), which combines the ipt gene for positive selection with the recombinase system R/RS for removal of marker genes from transgenic cells after transformation. Transformation with a MAT vector has been attempted in two citrus genotypes, Pineapple sweet orange (Citrus sinensis L. Osb.) and Carrizo citrange (C. sinensis L. Osb. x Poncirus trifoliata L. Raf.). Results indicated that the IPT phenotype was clearly distinguishable in sweet orange but not in citrange, and that excision was not always efficient and precise. Nevertheless, the easy visual detection of the IPT phenotype combined with the higher transformation efficiency achieved in sweet orange using this system open interesting perspectives for the generation of marker-free transgenic citrus plants.


Endodontic treatment-related antibiotic prescribing patterns of South African oral health professionals.
To assess the antibiotic prescribing patterns of South African dentists for patients undergoing endodontic treatment. This study used data from 2013 health insurance claims submitted by South African oral health professionals to determine the antibiotic prescribing patterns related to endodontic treatment. A logistic regression model was used to test the fully adjusted statistical significance of the association between the exploratory variables (gender, age group, event type, abscess treatment, chronic health) and the dependent variable (antibiotic prescription). Odds ratios with 95% confidence intervals (CI) are reported, and a 95% CI excluding 1 was considered statistically significant. Almost 10% of endodontic treatments were prescribed an antibiotic. There were no significant differences in prescribing patterns by gender, age and chronic health status. Prescriptions were more common at the preparatory stage (9.4%) of root canal treatment compared to the therapy (4.7%) and canal filling (2%) stages. Patients who received apical surgery (OR = 2.28; 95% CI 1.38-3.76) and treatment of an abscess (OR = 2.57; 95% 1.82-3.63) had a significantly increased odds of being prescribed an antibiotic. Almost three-quarters of prescriptions were for narrow spectrum antibiotics. The frequency of antibiotic prescribing by South African dental practitioners for patients undergoing endodontic treatment is relatively low and predominantly involved narrow spectrum antibiotics. It, however, remains important that antibiotics are only prescribed when clinically essential, such as when there are obvious systemic effects. These include fever above 37 degrees, malaise, lymphadenopathy, trismus, increase swelling, cellulitis, osteomyelitis and persistent infection. The wider dissemination and adherence to clear evidence-based prescribing guidelines for antibiotics in this clinical area are important.


Word learning from a tablet app: Toddlers perform better in a passive context.
In recent years, the popularity of tablets has skyrocketed and there has been an explosive growth in apps designed for children. Howhever, many of these apps are released without tests for their effectiveness. This is worrying given that the factors influencing children's learning from touchscreen devices need to be examined in detail. In particular, it has been suggested that children learn less from passive video viewing relative to equivalent live interaction, which would have implications for learning from such digital tools. However, this so-called video deficit may be reduced by allowing children greater influence over their learning environment. Across two touchscreen-based experiments, we examined whether 2- to 4-year-olds benefit from actively choosing what to learn more about in a digital word learning task. We designed a tablet study in which "active" participants were allowed to choose which objects they were taught the label of, while yoked "passive" participants were presented with the objects chosen by their active peers. We then examined recognition of the learned associations across different tasks. In Experiment 1, children in the passive condition outperformed those in the active condition (n = 130). While Experiment 2 replicated these findings in a new group of Malay-speaking children (n = 32), there were no differences in children's learning or recognition of the novel word-object associations using a more implicit looking time measure. These results suggest that there may be performance costs associated with active tasks designed as in the current study, and at the very least, there may not always be systematic benefits associated with active learning in touchscreen-based word learning tasks. The current studies add to the evidence that educational apps need to be evaluated before release: While children might benefit from interactive apps under certain conditions, task design and requirements need to consider factors that may detract from successful performance.


Phosphomonoester is associated with proliferation in human breast cancer: a 31P MRS study.
Phospholipid metabolism of human breast cancer was studied by 31P magnetic resonance spectroscopy (MRS). In vivo localised 31P MR spectra were obtained from the tumour alone using phase modulated rotating frame imaging. For 31 tumours, median (range) phosphomonoester (PME) to ATP ratio was 1.48 (0.57-3.78) and phosphodiester (PDE) to ATP ratio was 1.65 (0.44-3.89). DNA index and S phase fraction (SPF) were measured by flow cytometry of paraffin embedded tissue. Twelve (39%) tumours were diploid and 19 aneuploid. Median (range) SPF for 29 assessable tumours was 5.3% (0.6-28%), with significantly greater median SPF for aneuploid tumours (9.3%) than diploid (3.8%, P = 0.007). There was a significant association between PME/ATP and SPF (P = 0.03) due to a significant correlation for aneuploid tumours (P = 0.01). High resolution 31P MRS of extracts from 18 tumours (including seven studied in vivo) demonstrated that the PME peak consists predominantly of phosphoethanolamine (PE) with a smaller contribution from phosphocholine (PC) (median (range) PE/PC: 3.02 (1.13-5.09)). Changes in PME/ATP were observed for two tumours where tamoxifen stablized disease and may be consistent with the cytostatic effects of this drug.


Targeting Ubiquitin-Proteasome Pathway by Natural Products: Novel Therapeutic Strategy for Treatment of Neurodegenerative Diseases.
Misfolded proteins are the main common feature of neurodegenerative diseases, thereby, normal proteostasis is an important mechanism to regulate the neural survival and the central nervous system functionality. The ubiquitin-proteasome system (UPS) is a non-lysosomal proteolytic pathway involved in numerous normal functions of the nervous system, modulation of neurotransmitter release, synaptic plasticity, and recycling of membrane receptors or degradation of damaged and regulatory intracellular proteins. Aberrant accumulation of intracellular ubiquitin-positive inclusions has been implicated to a variety of neurodegenerative disorders such as Alzheimer's disease (AD), Parkinson's disease (PD), Huntington disease (HD), Amyotrophic Lateral Sclerosis (ALS), and Multiple Myeloma (MM). Genetic mutation in deubiquitinating enzyme could disrupt UPS and results in destructive effects on neuron survival. To date, various agents were characterized with proteasome-inhibitory potential. Proteins of the ubiquitin-proteasome system, and in particular, E3 ubiquitin ligases, may be promising molecular targets for neurodegenerative drug discovery. Phytochemicals, specifically polyphenols (PPs), were reported to act as proteasome-inhibitors or may modulate the proteasome activity. PPs modify the UPS by means of accumulation of ubiquitinated proteins, suppression of neuronal apoptosis, reduction of neurotoxicity, and improvement of synaptic plasticity and transmission. This is the first comprehensive review on the effect of PPs on UPS. Here, we review the recent findings describing various aspects of UPS dysregulation in neurodegenerative disorders. This review attempts to summarize the latest reports on the neuroprotective properties involved in the proper functioning of natural polyphenolic compounds with implication for targeting ubiquitin-proteasome pathway in the neurodegenerative diseases. We highlight the evidence suggesting that polyphenolic compounds have a dose and disorder dependent effects in improving neurological dysfunctions, and so their mechanism of action could stimulate the UPS, induce the protein degradation or inhibit UPS and reduce protein degradation. Future studies should focus on molecular mechanisms by which PPs can interfere this complex regulatory system at specific stages of the disease development and progression.


Sequence variants at CHRNB3-CHRNA6 and CYP2A6 affect smoking behavior.
Smoking is a common risk factor for many diseases. We conducted genome-wide association meta-analyses for the number of cigarettes smoked per day (CPD) in smokers (n = 31,266) and smoking initiation (n = 46,481) using samples from the ENGAGE Consortium. In a second stage, we tested selected SNPs with in silico replication in the Tobacco and Genetics (TAG) and Glaxo Smith Kline (Ox-GSK) consortia cohorts (n = 45,691 smokers) and assessed some of those in a third sample of European ancestry (n = 9,040). Variants in three genomic regions associated with CPD (P < 5 x 10(-8)), including previously identified SNPs at 15q25 represented by rs1051730[A] (effect size = 0.80 CPD, P = 2.4 x 10(-69)), and SNPs at 19q13 and 8p11, represented by rs4105144[C] (effect size = 0.39 CPD, P = 2.2 x 10(-12)) and rs6474412-T (effect size = 0.29 CPD, P = 1.4 x 10(-8)), respectively. Among the genes at the two newly associated loci are genes encoding nicotine-metabolizing enzymes (CYP2A6 and CYP2B6) and nicotinic acetylcholine receptor subunits (CHRNB3 and CHRNA6), all of which have been highlighted in previous studies of smoking and nicotine dependence. Nominal associations with lung cancer were observed at both 8p11 (rs6474412[T], odds ratio (OR) = 1.09, P = 0.04) and 19q13 (rs4105144[C], OR = 1.12, P = 0.0006).


A level-set approach to joint image segmentation and registration with application to CT lung imaging.
Automated analysis of structural imaging such as lung Computed Tomography (CT) plays an increasingly important role in medical imaging applications. Despite significant progress in the development of image registration and segmentation methods, lung registration and segmentation remain a challenging task. In this paper, we present a novel image registration and segmentation approach, for which we develop a new mathematical formulation to jointly segment and register three-dimensional lung CT volumes. The new algorithm is based on a level-set formulation, which merges a classic Chan-Vese segmentation with the active dense displacement field estimation. Combining registration with segmentation has two key advantages: it allows to eliminate the problem of initializing surface based segmentation methods, and to incorporate prior knowledge into the registration in a mathematically justified manner, while remaining computationally attractive. We evaluate our framework on a publicly available lung CT data set to demonstrate the properties of the new formulation. The presented results show the improved accuracy for our joint segmentation and registration algorithm when compared to registration and segmentation performed separately.


Cerebellar cleft: a form of prenatal cerebellar disruption.
In contrast to malformations, cerebellar disruptions have attracted little interest in the literature. We draw attention for the first time to the hypothesis that cerebellar clefts are residual changes following a prenatal cerebellar insult, and represent disruptions. We reviewed the clinical records and MR findings of six patients with a cerebellar cleft, two of whom also had prenatal MRI at 24 weeks of gestation. The clefts were located in the left cerebellar hemisphere in five cases, in the right in one patient. Other typical findings included disorderly alignment of the cerebellar folia and fissures, irregular gray/white matter junction, and abnormal arborization of the white matter in all patients. The cerebellar cleft extended into the fourth ventricle in three cases, and in two children cystic cortical lesions were seen. Supratentorial schizencephaly was found in two patients. In two patients there was a documented fetal cerebellar hemorrhage at 24 weeks of gestation. We conclude that cerebellar clefts are residual changes resulting from a prenatal cerebellar insult and consequently represent disruptions rather than primary malformations. The supratentorial findings are also in agreement with an acquired lesion. The outcome in these children was variable, mainly depending of the presence of supratentorial lesions.


Corpus callosum volume and neurocognition in autism.
The corpus callosum has recently been considered as an index of interhemispheric connectivity. This study applied a novel volumetric method to examine the size of the corpus callosum in 32 individuals with autism and 34 age-, gender- and IQ-matched controls and to investigate the relationship between this structure and cognitive measures linked to interhemispheric functioning. Participants with autism displayed reductions in total corpus callosum volume and in several of its subdivisions. Relationships were also observed between volumetric alterations and performance on several cognitive tests including the Tower of Hanoi test. These findings provide further evidence for anatomical alterations in the corpus callosum in autism, but warrant additional studies examining the relationship of this structure and specific measures of interhemispheric connectivity.


A blue spectral shift of the hemoglobin soret band correlates with the age (time since deposition) of dried bloodstains.
The ability to determine the time since deposition of a bloodstain found at a crime scene could prove invaluable to law enforcement investigators, defining the time frame in which the individual depositing the evidence was present. Although various methods of accomplishing this have been proposed, none has gained widespread use due to poor time resolution and weak age correlation. We have developed a method for the estimation of the time since deposition (TSD) of dried bloodstains using UV-VIS spectrophotometric analysis of hemoglobin (Hb) that is based upon its characteristic oxidation chemistry. A detailed study of the Hb Soret band (λmax=412 nm) in aged bloodstains revealed a blue shift (shift to shorter wavelength) as the age of the stain increases. The extent of this shift permits, for the first time, a distinction to be made between bloodstains that were deposited minutes, hours, days and weeks prior to recovery and analysis. The extent of the blue shift was found to be a function of ambient relative humidity and temperature. The method is extremely sensitive, requiring as little as a 1 µl dried bloodstain for analysis. We demonstrate that it might be possible to perform TSD measurements at the crime scene using a portable low-sample-volume spectrophotometer.


Long-term beneficial effects of vanadate, tungstate, and molybdate on insulin secretion and function of cultured beta cells.
The ultratrace elements vanadate, tungstate, and molybdate exhibit significant antihyperglycemic effects in both type 1 and 2 diabetic animals, but possible effects on the function of pancreatic beta cells are understudied. In the present study, clonal BRIN BD11 cells were cultured for 3 days with each ultratrace element to establish doses lacking detrimental effects on viable beta cell mass. Vanadate treatment (4 micromol/L) had no effect on cellular insulin content but improved glucose-induced insulin secretory responsiveness. However, insulin secretion mediated by PKA and PKC activation was desensitized in vanadate-treated cells. Culture with tungstate (300 micromol/L) and molybdate (1 mmol/L) increased cellular insulin content and enhanced basal insulin release and the responsiveness to glucose and a wide range of other secretagogues. These observations suggest significant effects of ultratrace elements on pancreatic beta cells that may contribute to their antihyperglycemic action.


Interaction between antibody-diversification enzyme AID and spliceosome-associated factor CTNNBL1.
Activation-induced deaminase (AID) deaminates deoxycytidine residues in immunoglobulin genes, triggering antibody diversification. Here, by use of two-hybrid and coimmunoprecipitation assays, we identify CTNNBL1 (also known as NAP) as an AID-specific interactor. Mutants of AID that interfere with CTNNBL1 interaction yield severely diminished hypermutation and class switching. Targeted inactivation of CTNNBL1 in DT40 B cells also considerably diminishes IgV diversification. CTNNBL1 is a widely expressed nuclear protein that associates with the Prp19 complex of the spliceosome, interacting with its CDC5L component. The results, therefore, identify residues in AID involved in its in vivo targeting and suggest they might act through interaction with CTNNBL1, giving possible insight into the linkage between AID recruitment and target-gene transcription.


Characterization and pharmacokinetics of coenzyme Q10 nanoparticles prepared by a rapid expansion of supercritical solution process.
Coenzyme Q10 (CoQ10) has been found to be effective in cardiovascular diseases and neurodegenerative diseases. However, the extremely poor solubility of CoQ10 in water is hampering its bioavailability as a therapeutic agent. To overcome solubility problem, we micronized the CoQ10 powder to the nanometer level by the supercritical solution (RESS) process, which does not employ any toxic organic solvent. The obtained CoQ10 nanoparticles were 147.9 +/- 27.3nm in diameter and their physicochemical properties were characterized by scanning electron microscopy (SEM), dynamic light scattering (DLS), liquid chromatography-mass spectrometry (LC-MS), X-ray diffractometry (XRD) and differential scanning calorimetry (DSC) analyzes. Moreover, the pharmacokinetics of the CoQ10 nanoparticles, in comparison with the unprocessed CoQ10 powder, were investigated in rats. From the results of physicochemical and pharmacokinetic studies, the CoQ10 nanoparticles had high solubility in water and possessed less crystalline structure, which can enhance the bioavailability of CoQ10, and provide a water-soluble solid dosage form of CoQ10.


Influence of viral transport media and freeze-thaw cycling on the sensitivity of qRT-PCR detection of SARS-CoV-2 nucleic acids.
Objective: The events of the last year have highlighted the complexity of implementing large-scale molecular diagnostic testing for novel pathogens. The purpose of this study was to determine the chemical influences of sample collection media and storage on the stability and detection of viral nucleic acids by qRT-PCR. We studied the mechanism(s) through which viral transport media (VTM) and number of freeze-thaw cycles influenced the analytical sensitivity of qRT-PCR detection of SARS-CoV-2. Our goal is to reinforce testing capabilities and identify weaknesses that could arise in resource-limited environments that do not have well-controlled cold chains. Method: The sensitivity of qRT-PCR analysis was studied in four VTM for synthetic single-stranded RNA (ssRNA) and double-stranded DNA (dsDNA) simulants of the SARS-CoV-2 genome. Results: The sensitivity and reproducibility of qRT-PCR for the synthetic ssRNA and dsDNA were found to be highly sensitive to VTM with the best results observed for ssRNA in HBSS and PBS-G. Surprisingly, the presence of epithelial cellular material with the ssRNA increased the sensitivity of the qRT-PCR assay. Repeated freeze-thaw cycling decreased the sensitivity of the qRT-PCR with two noted exceptions. Conclusions: The choice of VTM is critically important to defining the sensitivity of COVID-19 molecular diagnostics assays and this study suggests they can impact upon the stability of the SARS-CoV-2 viral genome. This becomes increasingly important if the virus structure is destabilised before analysis, which can occur due to poor storage conditions. This study suggests that COVID-19 testing performed with glycerol-containing PBS will produce a high level of stability and sensitivity. These results are in agreement with clinical studies reported for patient-derived samples.


Epigenetic Control of Phenotypic Plasticity in the Filamentous Fungus Neurospora crassa.
Phenotypic plasticity is the ability of a genotype to produce different phenotypes under different environmental or developmental conditions. Phenotypic plasticity is a ubiquitous feature of living organisms, and is typically based on variable patterns of gene expression. However, the mechanisms by which gene expression is influenced and regulated during plastic responses are poorly understood in most organisms. While modifications to DNA and histone proteins have been implicated as likely candidates for generating and regulating phenotypic plasticity, specific details of each modification and its mode of operation have remained largely unknown. In this study, we investigated how epigenetic mechanisms affect phenotypic plasticity in the filamentous fungus Neurospora crassa By measuring reaction norms of strains that are deficient in one of several key physiological processes, we show that epigenetic mechanisms play a role in homeostasis and phenotypic plasticity of the fungus across a range of controlled environments. In general, effects on plasticity are specific to an environment and mechanism, indicating that epigenetic regulation is context dependent and is not governed by general plasticity genes. Specifically, we found that, in Neurospora, histone methylation at H3K36 affected plastic response to high temperatures, H3K4 methylation affected plastic response to pH, but H3K27 methylation had no effect. Similarly, DNA methylation had only a small effect in response to sucrose. Histone deacetylation mainly decreased reaction norm elevation, as did genes involved in histone demethylation and acetylation. In contrast, the RNA interference pathway was involved in plastic responses to multiple environments.


Limited validity of diagnosis codes in Medicare claims for identifying cancer metastases and inferring stage.
Researchers are using diagnosis codes from health claims to identify metastatic disease in cancer patients. The validity of this approach has not been established. We used the linked 2005-2007 Surveillance, Epidemiology and End Results (SEER)-Medicare data to assess the validity of metastasis codes at diagnosis from claims compared with stage reported by SEER cancer registries. The cohort included 80,052 incident breast, lung, and colorectal cancer patients aged 65 years and older. Using gold-standard SEER data, we evaluated sensitivity, specificity, positive predictive value, and negative predictive value of claims-based stage, survival by stage classification, and patient factors associated with stage misclassification using multivariable regression. For patients with a registry report of distant metastatic cancer, the sensitivity, specificity, and positive predictive value of claims never simultaneously exceeded 80% for any cancer: lung (42.7%, 94.8%, and 88.1%), breast (51.0%, 98.3%, and 65.8%), and colorectal (72.8%, 93.8%, and 68.5%). Misclassification of stage from Medicare claims was significantly associated with inaccurate estimates of stage-specific survival (P < .001). In adjusted analysis, patients who were older, black, or living in low-income areas were more likely to have their stage misclassified in claims. Diagnosis codes in Medicare claims have limited validity for inferring cancer stage and metastatic disease.


Clinical effectiveness and cost-effectiveness of issuing longer versus shorter duration (3-month vs. 28-day) prescriptions in patients with chronic conditions: systematic review and economic modelling.
To reduce expenditure on, and wastage of, drugs, some commissioners have encouraged general practitioners to issue shorter prescriptions, typically 28 days in length; however, the evidence base for this recommendation is uncertain. To evaluate the evidence of the clinical effectiveness and cost-effectiveness of shorter versus longer prescriptions for people with stable chronic conditions treated in primary care. The design of the study comprised three elements. First, a systematic review comparing 28-day prescriptions with longer prescriptions in patients with chronic conditions treated in primary care, evaluating any relevant clinical outcomes, adherence to treatment, costs and cost-effectiveness. Databases searched included MEDLINE (PubMed), EMBASE, Cumulative Index to Nursing and Allied Health Literature, Web of Science and Cochrane Central Register of Controlled Trials. Searches were from database inception to October 2015 (updated search to June 2016 in PubMed). Second, a cost analysis of medication wastage associated with < 60-day and ≥ 60-day prescriptions for five patient cohorts over an 11-year period from the Clinical Practice Research Datalink. Third, a decision model adapting three existing models to predict costs and effects of differing adherence levels associated with 28-day versus 3-month prescriptions in three clinical scenarios. In the systematic review, from 15,257 unique citations, 54 full-text papers were reviewed and 16 studies were included, five of which were abstracts and one of which was an extended conference abstract. None was a randomised controlled trial: 11 were retrospective cohort studies, three were cross-sectional surveys and two were cost studies. No information on health outcomes was available. An exploratory meta-analysis based on six retrospective cohort studies suggested that lower adherence was associated with 28-day prescriptions (standardised mean difference -0.45, 95% confidence interval -0.65 to -0.26). The cost analysis showed that a statistically significant increase in medication waste was associated with longer prescription lengths. However, when accounting for dispensing fees and prescriber time, longer prescriptions were found to be cost saving compared with shorter prescriptions. Prescriber time was the largest component of the calculated cost savings to the NHS. The decision modelling suggested that, in all three clinical scenarios, longer prescription lengths were associated with lower costs and higher quality-adjusted life-years. The available evidence was found to be at a moderate to serious risk of bias. All of the studies were conducted in the USA, which was a cause for concern in terms of generalisability to the UK. No evidence of the direct impact of prescription length on health outcomes was found. The cost study could investigate prescriptions issued only; it could not assess patient adherence to those prescriptions. Additionally, the cost study was based on products issued only and did not account for underlying patient diagnoses. A lack of good-quality evidence affected our decision modelling strategy. Although the quality of the evidence was poor, this study found that longer prescriptions may be less costly overall, and may be associated with better adherence than 28-day prescriptions in patients with chronic conditions being treated in primary care. There is a need to more reliably evaluate the impact of differing prescription lengths on adherence, on patient health outcomes and on total costs to the NHS. The priority should be to identify patients with particular conditions or characteristics who should receive shorter or longer prescriptions. To determine the need for any further research, an expected value of perfect information analysis should be performed. This study is registered as PROSPERO CRD42015027042. The National Institute for Health Research Health Technology Assessment programme.


Aspermatogenesis in the guinea pig induced by testicular tissue and adjuvants.
The injection into the dorsal skin of a suspension of guinea pig testis or spermia incorporated in a water-in-oil emulsion containing killed mycobacteria induces aspermatogenesis in guinea pigs. The injury begins with the inhibition of the maturation of spermia and proceeds through the degeneration and exfoliation of spermatids, spermatocytes, and finally spermatogonia. These germinal cells pass from the seminiferous tubules into the epididymis. The process is not associated with inflammation. No significant changes occur in the intertubular spaces and the Leydig cells do not seem to be affected. The seminal vesicles and the prostate remain normal. The aspermatogenesis may begin in 10 days and it lasts for more than 5 months. The process may lead to atrophy of the seminiferous tubules and fibrosis. Guinea pigs which receive a suspension of their own testis or spermia and adjuvants develop a similar injury. The "mitochondrial" fraction of the testis of guinea pig is effective while repeated injections of alcoholic extract of testis emulsified with paraffin oil containing mycobacteria do not cause aspermatogenesis. The presence of acid-fast bacilli in the water-in-oil emulsion containing testis or spermia seems to be essential for the production of testicular lesions; the injection of antigen and mycobacteria into different sites is ineffective. When guinea pig testis is replaced by guinea pig liver or kidney or rabbit testis no testicular damage occurs. The injection of rabbit spinal cord combined with adjuvants results in allergic encephalomyelitis in a large proportion of guinea pigs, accompanied by a great loss of weight. The testes of a few of these animals show a varying degree of aspermatogenesis. When guinea pig brain is combined with adjuvants and administered subcutaneously the incidence of testicular injury is high, although the damage is, in general, mild. From the standpoint of mechanism, the inhibition of spermatogenesis which occurs in these animals may be unrelated to the injury which follows the injection of germinal cells. Aspermatogenesis follows the injection of killed mycobacteria in paraffin oil into the testis as well as into certain sites related to the gonad: the abdominal cavity, the subcutaneous tissue over the abdomen, and the skin of the inguinal region. Antibodies fixing complement in the presence of spermia are demonstrable in the sera of guinea pigs injected with testis or spermia and adjuvants. When the mycobacteria are omitted the titers are low and no testicular injury occurs. Although there seems to be a correlation between testicular damage and complement-fixing titer, this may not be a causal relationship. Antibodies which neutralize guinea pig hyaluronidase and those which immobilize spermia have also been demonstrated in the sera of these guinea pigs.


Hydrogen Sulfide-a potent multichannel anti-arrhythmic drug.
-Arrhythmia is a big headache for cardiologist for a long time. A new antiarrythmic drug with good effects and few side effects should be explored. Hydrogen sulfide, a newly found gaseous transmitter, was found to have multi-effects on a variety of ion channels. It may be a promising antiarrythmic drug. This article reviews the different effects of hydrogen sulfide on different ion channels.


Postprandial lipoprotein lipase, insulin and gastric inhibitory polypeptide responses to test meals of different fatty acid composition: comparison of saturated, n-6 and n-3 polyunsaturated fatty acids.
The present study was carried out to investigate effects of meals, rich in either saturated fatty acids (SFA), or n-6 or n-3 fatty acids, on postprandial plasma lipid and hormone concentrations as well as post-heparin plasma lipoprotein lipase (LPL) activity. The study was a randomized single-blind study comparing responses to three test meals. The volunteers attended the Clinical Investigation Unit of the Royal Surrey County Hospital on three separate occasions in order to consume the meals. Twelve male volunteers with an average age of 22.5 +/- 1.4 years (mean +/- SD), were selected from the University of Surrey student population; one subject dropped out of the study because he found the test meal unpalatable. Three meals were given in the early evening and postprandial responses were followed overnight for 11h. The oils used to prepare each of the three test meals were: a mixed oil rich in saturated fatty acids (SFA) which mimicked the fatty acid composition of the current UK diet, corn oil, rich in n-6 fatty acids and a fish oil concentrate (MaxEPA) rich in n-3 fatty acids. The oil under investigation (40 g) was incorporated into the test meals which were otherwise identical [208 g carbohydrates, 35 g protein, 5.65 MJ (1350 kcal) energy]. Postprandial plasma triacylglycerol (TAG), gastric inhibitory polypeptide (GIP), and insulin responses, as well as post-heparin LPL activity (measured at 12 h postprandially only) were investigated. Fatty acids of the n-3 series significantly reduced plasma TAG responses compared to the mixed oil meal (P < 0.05) and increased post-heparin LPL activity 15 min after the injection of heparin (P < 0.01). A biphasic response was observed in TAG, with peak responses occurring at 1 h and between 3-7 h postprandially. GIP and insulin showed similar responses to the three test meals and no significant differences were observed. We conclude that fish oils can decrease postprandial plasma TAG levels partly through an increase in post-heparin LPL activity, which however, is not due to increased GIP or insulin concentrations.


"Double crap!" abuse and harmed identity in Fifty Shades of Grey.
While intimate partner violence (IPV) affects 25% of women and impairs health, current societal conditions-including the normalization of abuse in popular culture such as novels, film, and music-create the context to support such violence. Fifty Shades of Grey, a best-selling novel, depicts a "romantic" and "erotic" relationship involving 28-year-old megamillionaire, Christian Grey, and a 22-year-old college student, Anastasia Steele. We argue that the relationship is characterized by IPV, which is harmful to Anastasia. All authors engaged in iterative readings of the text, and wrote narrative summaries to elucidate themes. Validity checks included double review of the first eight chapters of the novel to establish consistency in our analysis approach, iterative discussions in-person and electronically to arbitrate discrepancies, and review of our analysis with other abuse and sexual practice experts. To characterize IPV, we used the U.S. Centers for Disease Control and Prevention's definitions of emotional abuse (intimidation/threats; isolation; stalking; and humiliation) and sexual violence (forced sex acts/contact against a person's will, including using alcohol/drugs or intimidation/pressure). To characterize harm, we used Smith's conceptualizations of perceived threat, managing, altered identity, yearning, entrapment, and disempowerment experienced by abused women. Emotional abuse is present in nearly every interaction, including: stalking (Christian deliberately follows Anastasia and appears in unusual places, uses a phone and computer to track Anastasia's whereabouts, and delivers expensive gifts); intimidation (Christian uses intimidating verbal and nonverbal behaviors, such as routinely commanding Anastasia to eat and threatening to punish her); and isolation (Christian limits Anastasia's social contact). Sexual violence is pervasive-including using alcohol to compromise Anastasia's consent, as well as intimidation (Christian initiates sexual encounters when genuinely angry, dismisses Anastasia's requests for boundaries, and threatens her). Anastasia experiences reactions typical of abused women, including: constant perceived threat ("my stomach churns from his threats"); altered identity (describes herself as a "pale, haunted ghost"); and stressful managing (engages in behaviors to "keep the peace," such as withholding information about her social whereabouts to avoid Christian's anger). Anastasia becomes disempowered and entrapped in the relationship as her behaviors become mechanized in response to Christian's abuse. Our analysis identified patterns in Fifty Shades that reflect pervasive intimate partner violence-one of the biggest problems of our time. Further, our analysis adds to a growing body of literature noting dangerous violence standards being perpetuated in popular culture.


Repeat deliberate self-harm: a link with childhood sexual abuse?
The purpose of this study was to identify whether a past history of childhood sexual abuse is a risk factor for repeated Deliberate Self-Harm (DSH). The study was a 6-month prospective study of 178 patients responsible for 190 consecutive cases of DSH seen during a 3-month censoring period. Patients were identified by review of the in-patient and accident and emergency (A&E) records of all cases of DSH at the A&E department of a major teaching hospital. Patients with a history of childhood sexual abuse showed a marked clustering of four major risk factors for repeat DSH (unemployment, past deliberate self-poisoning, self-injury and psychiatric illness) and were significantly more likely to repeat DSH within the 6-month follow-up period.


Role of primary stability for successful osseointegration of dental implants: Factors of influence and evaluation.
A secure implant primary (mechanical) stability is positively associated with a successful implant integration and long-term successful clinical outcome. Therefore, it is essential to assess the initial stability at different time-points to ensure a successful osseointegration. The present study critically reviews the factors that may play a role in achieving a successful initial stability in dental implants. Databases were searched from 1983 up to and including October 2013 using different combinations of various keywords. Bone quality and quantity, implant geometry, and surgical technique adopted may significantly influence primary stability and overall success rate of dental implants.


Polygenic Risk Score, Parental Socioeconomic Status, Family History of Psychiatric Disorders, and the Risk for Schizophrenia: A Danish Population-Based Study and Meta-analysis.
Schizophrenia has a complex etiology influenced both by genetic and nongenetic factors but disentangling these factors is difficult. To estimate (1) how strongly the risk for schizophrenia relates to the mutual effect of the polygenic risk score, parental socioeconomic status, and family history of psychiatric disorders; (2) the fraction of cases that could be prevented if no one was exposed to these factors; (3) whether family background interacts with an individual's genetic liability so that specific subgroups are particularly risk prone; and (4) to what extent a proband's genetic makeup mediates the risk associated with familial background. We conducted a nested case-control study based on Danish population-based registers. The study consisted of 866 patients diagnosed as having schizophrenia between January 1, 1994, and December 31, 2006, and 871 matched control individuals. Genome-wide data and family psychiatric and socioeconomic background information were obtained from neonatal biobanks and national registers. Results from a separate meta-analysis (34,600 cases and 45,968 control individuals) were applied to calculate polygenic risk scores. Polygenic risk scores, parental socioeconomic status, and family psychiatric history. Odds ratios (ORs), attributable risks, liability R2 values, and proportions mediated. Schizophrenia was associated with the polygenic risk score (OR, 8.01; 95% CI, 4.53-14.16 for highest vs lowest decile), socioeconomic status (OR, 8.10; 95% CI, 3.24-20.3 for 6 vs no exposures), and a history of schizophrenia/psychoses (OR, 4.18; 95% CI, 2.57-6.79). The R2 values were 3.4% (95% CI, 2.1-4.6) for the polygenic risk score, 3.1% (95% CI, 1.9-4.3) for parental socioeconomic status, and 3.4% (95% CI, 2.1-4.6) for family history. Socioeconomic status and psychiatric history accounted for 45.8% (95% CI, 36.1-55.5) and 25.8% (95% CI, 21.2-30.5) of cases, respectively. There was an interaction between the polygenic risk score and family history (P = .03). A total of 17.4% (95% CI, 9.1-26.6) of the effect associated with family history of schizophrenia/psychoses was mediated through the polygenic risk score. Schizophrenia was associated with the polygenic risk score, family psychiatric history, and socioeconomic status. Our study demonstrated that family history of schizophrenia/psychoses is partly mediated through the individual's genetic liability.


Analysis of the [RNQ+] prion reveals stability of amyloid fibers as the key determinant of yeast prion variant propagation.
Variation in pathology of human prion disease is believed to be caused, in part, by distinct conformations of aggregated protein resulting in different prion strains. Several prions also exist in yeast and maintain different self-propagating structures, referred to as prion variants. Investigation of the yeast prion [PSI(+)] has been instrumental in deciphering properties of prion variants and modeling the physical basis of their formation. Here, we describe the generation of specific variants of the [RNQ(+)] prion in yeast transformed with fibers formed in vitro in different conditions. The fibers of the Rnq1p prion-forming domain (PFD) that induce different variants in vivo have distinct biochemical properties. The physical basis of propagation of prion variants has been previously correlated to rates of aggregation and disaggregation. With [RNQ(+)] prion variants, we found that the prion variant does not correlate with the rate of aggregation as anticipated but does correlate with stability. Interestingly, we found that there are differences in the ability of the [RNQ(+)] prion variants to faithfully propagate themselves and to template the aggregation of other proteins. Incorporating the mechanism of variant formation elucidated in this study with that previously proposed for [PSI(+)] variants has provided a framework to separate general characteristics of prion variant properties from those specific to individual prion proteins.


Subclinical persisting pulmonary hypertension in chronic neonatal lung disease.
The development of pulmonary hypertension is one of the adverse factors in the outcome of infants with chronic neonatal lung disese (CNLD). The purpose of this cross sectional study was to evaluate the prevalence and degree of pulmonary hypertension in a cohort of survivors of CNLD stable in air. Pulmonary artery pressure was assessed using its inverse correlation with the ratio of time to peak velocity and right ventricular ejection time (TPV:RVET) as measured from Doppler velocity time signals in the main pulmonary artery. A normal ratio is > or = 0.35, a possibly low ratio lies between 0.31 and 0.35, and a definitely low ratio is < 0.31. The subjects were divided into three groups. Group A comprised 58 infants with oxygen dependence and an abnormal chest radiograph at 28 days of age; group B comprised 18 infants with oxygen dependence and a normal chest radiograph at 28 days of age; and group C (controls) comprised 21 siblings without oxygen dependence by 10 days and a normal chest radiograph. There were significant differences in mean (SD) TPV:RVET ratio between group A 0.346 (0.045), group B 0.335 (0.057), and groups A + B 0.344 (0.048) when compared with group C controls 0.385 (0.034). The prevalence of a definitely low TPV:RVET ratio suggesting a raised pulmonary artery pressure was 19% in group A, 39% in group B, 24% in groups A + B, and none in group C. There were no clinical signs of pulmonary hypertension in any patient studied. Stepwise multiple linear regression failed to find significant associations with antenatal or neonatal putative risk factors. Additionally, there were no associations with childhood respiratory morbidity. These data suggest a high prevalence of subclinical pulmonary hypertension in CNLD patients. It is speculated that occult hypoxaemia may be occurring in this group of infants.


Loss of the Coffin-Lowry syndrome-associated gene RSK2 alters ERK activity, synaptic function and axonal transport in Drosophila motoneurons.
Plastic changes in synaptic properties are considered as fundamental for adaptive behaviors. Extracellular-signal-regulated kinase (ERK)-mediated signaling has been implicated in regulation of synaptic plasticity. Ribosomal S6 kinase 2 (RSK2) acts as a regulator and downstream effector of ERK. In the brain, RSK2 is predominantly expressed in regions required for learning and memory. Loss-of-function mutations in human RSK2 cause Coffin-Lowry syndrome, which is characterized by severe mental retardation and low IQ scores in affected males. Knockout of RSK2 in mice or the RSK ortholog in Drosophila results in a variety of learning and memory defects. However, overall brain structure in these animals is not affected, leaving open the question of the pathophysiological consequences. Using the fly neuromuscular system as a model for excitatory glutamatergic synapses, we show that removal of RSK function causes distinct defects in motoneurons and at the neuromuscular junction. Based on histochemical and electrophysiological analyses, we conclude that RSK is required for normal synaptic morphology and function. Furthermore, loss of RSK function interferes with ERK signaling at different levels. Elevated ERK activity was evident in the somata of motoneurons, whereas decreased ERK activity was observed in axons and the presynapse. In addition, we uncovered a novel function of RSK in anterograde axonal transport. Our results emphasize the importance of fine-tuning ERK activity in neuronal processes underlying higher brain functions. In this context, RSK acts as a modulator of ERK signaling.


NMR analysis of the fibronectin cell-adhesive sequence, Arg-Gly-Asp, in a recombinant silk-like protein and a model peptide.
It is well established that by introducing the cell-adhesive sequence Arg-Gly-Asp (RGD) from fibronectin into Bombyx mori silk fibroin by covalent coupling or bioengineering techniques, excellent biomaterials have been developed with the modified silk fibroin. However, there is no report about the structure and dynamics of the RGD moiety in the silk fibroin. To clarify the origin of such a high cell adhesion character and to design new recombinant silk protein with higher cell adhesion ability, it is necessary to characterize the structure and dynamics of the RGD moiety introduced into silk fibroin. In this study, the structure and dynamics of the RGD moiety in a recombinant silk-like protein, SLPF(10), consisting of the repeated silk fibroin sequence (AGSGAG)(3) and the sequence ASTGRGDSPA including the RGD moiety, were studied using solution NMR. The (1)H, (15)N, and (13)C chemical shifts indicate that the RGD moiety, as well as the silk fibroin sequence, takes a random coil form with high mobility in aqueous solution. Next, a (13)C solid-state NMR study was performed on a (13)C selectively labeled model peptide, AGSGAG[3-(13)C]A(7)GSGAGAGSGGT[2-(13)C]G(19)R[1-(13)C]G(21)DSPAGGGAGAGSGAG. After formic acid treatment, an increase in the β-sheet fraction for the AGSGAG sequence and peak narrowing of the residues around the RGD moiety were observed in the dry state. The latter indicates a decrease in the chemical shift distribution although the RGD moiety is still in random coil. A decrease in the peak intensities of the RGD moiety in the swollen state after immersing it in distilled water was observed, indicating high mobility of the RGD sequence in the peptide in the swollen state. Thus, the random coil state of the RGD moiety in the recombinant silk-like protein is maintained in aqueous solution and also in both dry and swollen state. This is similar to the case of the RGD moiety in fibronectin. The presence of the linker ASTG at the N-terminus and SPAGG at the C-terminus seems important to maintain the random coil form and the flexible state of the RGD sequence in order to permit access for binding to various integrins.


Study protocol for a multicentre, controlled non-randomised trial: benefits of exercise physiology services for type 2 diabetes (BEST).
Controlled trials support the efficacy of exercise as a treatment modality for chronic conditions, yet effectiveness of real-world Exercise Physiology services is yet to be determined. This study will investigate the efficacy and cost-effectiveness of services provided by Accredited Exercise Physiologists (AEPs) for clients with type 2 diabetes (T2D) in clinical practice. A non-randomised, opportunistic control, longitudinal design trial will be conducted at ten Exercise Physiology Clinics. Participants will be individuals with T2D attending one of the Exercise Physiology Clinics for routine AEP services (exercise prescription and counselling) (intervention) or individuals with T2D not receiving AEP services (usual care) (control). The experimental period will be 6 months with measurements performed at baseline and at 6 months. Primary outcome measures will be glycosylated haemoglobin (HbA1c), resting brachial blood pressure (BP), body mass index, waist circumference, 6 min walk test, grip strength, 30 s sit to stand, Medical Outcomes Short-Form 36-Item Health Survey and Active Australia Questionnaire. Secondary outcomes will be medication usage, out-of-pocket expenses, incidental, billable and non-billable health professional encounters and work missed through ill health. Healthcare utilisation will be measured for 12 months prior to, during and 12 months after trial participation using linked data from Medicare Benefits Schedule and Pharmaceutical Benefits Scheme data. The study is a multicentre trial comprising: University of Tasmania, University of New South Wales Lifestyle Clinic, University of Canberra, Baker Heart and Diabetes Institute (covered under the ethics approval of University of Tasmania Health and Medical Ethics Committee H0015266), Deakin University (Approval number: 2016-187), Australian Catholic University (2016-304R), Queensland University of Technology (1600000049), University of South Australia (0000035306), University of Western Australia (RA/4/1/8282) and Canberra Hospital (ETH.8.17.170). The findings of this clinical trial will be communicated via peer-reviewed journal articles, conference presentations, social media and broadcast media. ACTRN12616000264482.