MicroRNA-335-5p suppresses voltage-gated sodium channel expression and may be a target for seizure control.

There remains an urgent need for new therapies for treatment-resistant epilepsy. Sodium channel blockers are effective for seizure control in common forms of epilepsy, but loss of sodium channel function underlies some genetic forms of epilepsy. Approaches that provide bidirectional control of sodium channel expression are needed. MicroRNAs (miRNA) are small noncoding RNAs which negatively regulate gene expression. Here we show that genome-wide miRNA screening of hippocampal tissue from a rat epilepsy model, mice treated with the antiseizure medicine cannabidiol, and plasma from patients with treatment-resistant epilepsy, converge on a single target-miR-335-5p. Pathway analysis on predicted and validated miR-335-5p targets identified multiple voltage-gated sodium channels (VGSCs). Intracerebroventricular injection of antisense oligonucleotides against miR-335-5p resulted in upregulation of Scn1a, Scn2a, and Scn3a in the mouse brain and an increased action potential rising phase and greater excitability of hippocampal pyramidal neurons in brain slice recordings, consistent with VGSCs as functional targets of miR-335-5p. Blocking miR-335-5p also increased voltage-gated sodium currents and SCN1A, SCN2A, and SCN3A expression in human induced pluripotent stem cell-derived neurons. Inhibition of miR-335-5p increased susceptibility to tonic-clonic seizures in the pentylenetetrazol seizure model, whereas adeno-associated virus 9-mediated overexpression of miR-335-5p reduced seizure severity and improved survival. These studies suggest modulation of miR-335-5p may be a means to regulate VGSCs and affect neuronal excitability and seizures. Changes to miR-335-5p may reflect compensatory mechanisms to control excitability and could provide biomarker or therapeutic strategies for different types of treatment-resistant epilepsy.

Genome-wide microRNA profiling of plasma from three different animal models identifies biomarkers of temporal lobe epilepsy.

Epilepsy diagnosis is complex, requires a team of specialists and relies on in-depth patient and family history, MRI-imaging and EEG monitoring. There is therefore an unmet clinical need for a non-invasive, molecular-based, biomarker to either predict the development of epilepsy or diagnose a patient with epilepsy who may not have had a witnessed seizure. Recent studies have demonstrated a role for microRNAs in the pathogenesis of epilepsy. MicroRNAs are short non-coding RNA molecules which negatively regulate gene expression, exerting profound influence on target pathways and cellular processes. The presence of microRNAs in biofluids, ease of detection, resistance to degradation and functional role in epilepsy render them excellent candidate biomarkers. Here we performed the first multi-model, genome-wide profiling of plasma microRNAs during epileptogenesis and in chronic temporal lobe epilepsy animals. From video-EEG monitored rats and mice we serially sampled blood samples and identified a set of dysregulated microRNAs comprising increased miR-93-5p, miR-142-5p, miR-182-5p, miR-199a-3p and decreased miR-574-3p during one or both phases. Validation studies found miR-93-5p, miR-199a-3p and miR-574-3p were also dysregulated in plasma from patients with intractable temporal lobe epilepsy. Treatment of mice with common anti-epileptic drugs did not alter the expression levels of any of the five miRNAs identified, however administration of an anti-epileptogenic microRNA treatment prevented dysregulation of several of these miRNAs. The miRNAs were detected within the Argonuate2-RISC complex from both neurons and microglia indicating these miRNA biomarker candidates can likely be traced back to specific brain cell types. The current studies identify additional circulating microRNA biomarkers of experimental and human epilepsy which may support diagnosis of temporal lobe epilepsy via a quick, cost-effective rapid molecular-based test.

Diagnosis and management of tropomyosin receptor kinase (TRK) fusion sarcomas: expert recommendations from the World Sarcoma Network.

Sarcomas are a heterogeneous group of malignancies with mesenchymal lineage differentiation. The discovery of neurotrophic tyrosine receptor kinase (NTRK) gene fusions as tissue-agnostic oncogenic drivers has led to new personalized therapies for a subset of patients with sarcoma in the form of tropomyosin receptor kinase (TRK) inhibitors. NTRK gene rearrangements and fusion transcripts can be detected with different molecular pathology techniques, while TRK protein expression can be demonstrated with immunohistochemistry. The rarity and diagnostic complexity of NTRK gene fusions raise a number of questions and challenges for clinicians. To address these challenges, the World Sarcoma Network convened two meetings of expert adult oncologists and pathologists and subsequently developed this article to provide practical guidance on the management of patients with sarcoma harboring NTRK gene fusions. We propose a diagnostic strategy that considers disease stage and histologic and molecular subtypes to facilitate routine testing for TRK expression and subsequent testing for NTRK gene fusions.

First Evidence of Local E×B Drift in the Divertor Influencing the Structure and Stability of Confined Plasma near the Edge of Fusion Devices.

The structure of the edge plasma in a magnetic confinement system has a strong impact on the overall plasma performance. We uncover for the first time a magnetic-field-direction dependent density shelf, i.e., local flattening of the density radial profile near the magnetic separatrix, in high confinement plasmas with low edge collisionality in the DIII-D tokamak. The density shelf is correlated with a doubly peaked density profile near the divertor target plate, which tends to occur for operation with the ion B×∇B drift direction away from the X-point, as currently employed for DIII-D advanced tokamak scenarios. This double-peaked divertor plasma profile is connected via the E×B drifts, arising from a strong radial electric field induced by the radial electron temperature gradient near the divertor target. The drifts lead to the reversal of the poloidal flow above the divertor target, resulting in the formation of the density shelf. The edge density shelf can be further enhanced at higher heating power, preventing large, periodic bursts of the plasma, i.e., edge-localized modes, in the edge region, consistent with ideal magnetohydrodynamics calculations.

MicroRNAs as biomarkers and treatment targets in status epilepticus.

Microribonucleic acids (miRNAs) are short noncoding ribonucleic acids (RNAs) that have been proposed as potential biomarkers for epilepsy, acute seizures, and status epilepticus. Various properties support their potential in this regard, including relative stability and amenability to rapid quantitation in biofluids. Several miRNAs are enriched in the brain and within specific cell types. Dysregulation of miRNAs has been reported in brain regions damaged by status epilepticus and in resected brain tissue from patients with drug-resistant epilepsy. Silencing miRNAs using antisense-like oligonucleotides termed antagomirs has been reported to suppress evoked and spontaneous seizures in animal models, indicating therapeutic applications. The prospect of miRNAs as mechanistic biomarkers is supported by recent studies showing blood levels of brain-enriched miRNAs increase after status epilepticus in rodents, and clinical studies have identified miRNAs upregulated in human cerebrospinal fluid after status epilepticus. It remains unproven, however, whether there are miRNAs that uniquely identify acute seizures, chronic epilepsy, or the process of epileptogenesis. Finally, efforts have turned to the challenge of proving that some of the circulating miRNAs actually originate from the brain. New models that feature a biochemically-labeled protein involved in miRNA function and restricted to specific brain cell types offer opportunities to resolve this issue. This review summarizes recent progress on miRNAs as diagnostic biomarkers of status epilepticus and considers some of the unanswered questions and future directions. This article is part of the Special Issue "Proceedings of the 7th London-Innsbruck Colloquium on Status Epilepticus and Acute Seizures.

Sociodemographic, psychosocial and clinical factors associated with uptake of genetic counselling for hereditary cancer: a systematic review.

Evidence suggests that a significant proportion of individuals referred to cancer genetic counselling (GC) do not attend, and thus may not be engaged in adequate cancer risk management. We aimed to review the literature to better understand barriers to accessing GC and how they may be overcome. We conducted a systematic literature search for articles examining factors influencing cancer GC uptake as well as motivators and barriers to GC attendance. Factors were categorised as sociodemographic, psychosocial or clinical. The literature search identified 1413 citations, 35 of which met the inclusion criteria. GC uptake ranged from 19% to 88%. With the exceptions of education level, socioeconomic status, cancer-specific distress, personal cancer diagnosis and actual and perceived risk of cancer, support was lacking for most sociodemographic, clinical and psychosocial factors as predictors of GC uptake. Cost and logistical barriers, emotional concerns, family concerns and low perceived personal relevance were reported as important considerations for those declining GC. We conclude that there is poor understanding of GC and a lack of decision support among those referred to GC. Research into ways of providing education and support to referred individuals will be important as the scope and availability of GC and genetic testing broaden.

Assessing the clinical value of targeted massively parallel sequencing in a longitudinal, prospective population-based study of cancer patients.

INTRODUCTION: Recent discoveries in cancer research have revealed a plethora of clinically actionable mutations that provide therapeutic, prognostic and predictive benefit to patients. The feasibility of screening mutations as part of the routine clinical care of patients remains relatively unexplored as the demonstration of massively parallel sequencing (MPS) of tumours in the general population is required to assess its value towards the health-care system. METHODS: Cancer 2015 study is a large-scale, prospective, multisite cohort of newly diagnosed cancer patients from Victoria, Australia with 1094 patients recruited. MPS was performed using the Illumina TruSeq Amplicon Cancer Panel. RESULTS: Overall, 854 patients were successfully sequenced for 48 common cancer genes. Accurate determination of clinically relevant mutations was possible including in less characterised cancer types; however, technical limitations including formalin-induced sequencing artefacts were uncovered. Applying strict filtering criteria, clinically relevant mutations were identified in 63% of patients, with 26% of patients displaying a mutation with therapeutic implications. A subset of patients was validated for canonical mutations using the Agena Bioscience MassARRAY system with 100% concordance. Whereas the prevalence of mutations was consistent with other institutionally based series for some tumour streams (breast carcinoma and colorectal adenocarcinoma), others were different (lung adenocarcinoma and head and neck squamous cell carcinoma), which has significant implications for health economic modelling of particular targeted agents. Actionable mutations in tumours not usually thought to harbour such genetic changes were also identified. CONCLUSIONS: Reliable delivery of a diagnostic assay able to screen for a range of actionable mutations in this cohort was achieved, opening unexpected avenues for investigation and treatment of cancer patients.

A phase Ib/II translational study of sunitinib with neoadjuvant radiotherapy in soft-tissue sarcoma.

BACKGROUND: Preoperative radiotherapy (RT) is commonly used to treat localised soft-tissue sarcomas (STS). Hypoxia is an important determinant of radioresistance. Whether antiangiogenic therapy can 'normalise' tumour vasculature, thereby improving oxygenation, remains unknown. METHODS: Two cohorts were prospectively enrolled. Cohort A evaluated the implications of hypoxia in STS, using the hypoxic tracer (18)F-azomycin arabinoside (FAZA-PET). In cohort B, sunitinib was added to preoperative RT in a dose-finding phase 1b/2 design. RESULTS: In cohort A, 13 out of 23 tumours were hypoxic (FAZA-PET), correlating with metabolic activity (r(2)=0.85; P<0.001). Two-year progression-free (PFS) and overall (OS) survival were 61% (95% CI: 0.44-0.84) and 87% (95% CI: 0.74-1.00), respectively. Hypoxia was associated with radioresistance (P=0.012), higher local recurrence (Hazard ratio (HR): 10.2; P=0.02), PFS (HR: 8.4; P=0.02), and OS (HR: 41.4; P<0.04). In Cohort B, seven patients received sunitinib at dose level (DL): 0 (50 mg per day for 2 weeks before RT; 25 mg per day during RT) and two patients received DL: -1 (37.5 mg per day for entire period). Dose-limiting toxicities were observed in 4 out of 7 patients at DL 0 and 2 out of 2 patients at DL -1, resulting in premature study closure. Although there was no difference in PFS or OS, patients receiving sunitinib had higher local failure (HR: 8.1; P=0.004). CONCLUSION: In STS, hypoxia is associated with adverse outcomes. The combination of sunitinib with preoperative RT resulted in unacceptable toxicities, and higher local relapse rates.

Can a weight loss of one pound a week be achieved with a 3500-kcal deficit? Commentary on a commonly accepted rule.

Despite theoretical evidence that the model commonly referred to as the 3500-kcal rule grossly overestimates actual weight loss, widespread application of the 3500-kcal formula continues to appear in textbooks, on respected government- and health-related websites, and scientific research publications. Here we demonstrate the risk of applying the 3500-kcal rule even as a convenient estimate by comparing predicted against actual weight loss in seven weight loss experiments conducted in confinement under total supervision or objectively measured energy intake. We offer three newly developed, downloadable applications housed in Microsoft Excel and Java, which simulates a rigorously validated, dynamic model of weight change. The first two tools available at http://www.pbrc.edu/sswcp, provide a convenient alternative method for providing patients with projected weight loss/gain estimates in response to changes in dietary intake. The second tool, which can be downloaded from the URL http://www.pbrc.edu/mswcp, projects estimated weight loss simultaneously for multiple subjects. This tool was developed to inform weight change experimental design and analysis. While complex dynamic models may not be directly tractable, the newly developed tools offer the opportunity to deliver dynamic model predictions as a convenient and significantly more accurate alternative to the 3500-kcal rule.

Brain cell-specific origin of circulating microRNA biomarkers in experimental temporal lobe epilepsy.

The diagnosis of epilepsy is complex and challenging and would benefit from the availability of molecular biomarkers, ideally measurable in a biofluid such as blood. Experimental and human epilepsy are associated with altered brain and blood levels of various microRNAs (miRNAs). Evidence is lacking, however, as to whether any of the circulating pool of miRNAs originates from the brain. To explore the link between circulating miRNAs and the pathophysiology of epilepsy, we first sequenced argonaute 2 (Ago2)-bound miRNAs in plasma samples collected from mice subject to status epilepticus induced by intraamygdala microinjection of kainic acid. This identified time-dependent changes in plasma levels of miRNAs with known neuronal and microglial-cell origins. To explore whether the circulating miRNAs had originated from the brain, we generated mice expressing FLAG-Ago2 in neurons or microglia using tamoxifen-inducible Thy1 or Cx3cr1 promoters, respectively. FLAG immunoprecipitates from the plasma of these mice after seizures contained miRNAs, including let-7i-5p and miR-19b-3p. Taken together, these studies confirm that a portion of the circulating pool of miRNAs in experimental epilepsy originates from the brain, increasing support for miRNAs as mechanistic biomarkers of epilepsy.

Anti-seizure effects of JNJ-54175446 in the intra-amygdala kainic acid model of drug-resistant temporal lobe epilepsy in mice.

There remains a need for new drug targets for treatment-resistant temporal lobe epilepsy. The ATP-gated P2X7 receptor coordinates neuroinflammatory responses to tissue injury. Previous studies in mice reported that the P2X7 receptor antagonist JNJ-47965567 suppressed spontaneous seizures in the intraamygdala kainic acid model of epilepsy and reduced attendant gliosis in the hippocampus. The drug-resistance profile of this model is not fully characterised, however, and newer P2X7 receptor antagonists with superior pharmacokinetic profiles have recently entered clinical trials. Using telemetry-based continuous EEG recordings in mice, we demonstrate that spontaneous recurrent seizures in the intraamygdala kainic acid model are refractory to the common anti-seizure medicine levetiracetam. In contrast, once-daily dosing of JNJ-54175446 (30 mg/kg, intraperitoneal) resulted in a significant reduction in spontaneous recurrent seizures which lasted several days after the end of drug administration. Using a combination of immunohistochemistry and ex vivo radiotracer assay, we find that JNJ-54175446-treated mice at the end of recordings display a reduction in astrogliosis and altered microglia process morphology within the ipsilateral CA3 subfield of the hippocampus, but no difference in P2X7 receptor surface expression. The present study extends the characterisation of the drug-resistance profile of the intraamygdala kainic acid model in mice and provides further evidence that targeting the P2X7 receptor may have therapeutic applications in the treatment of temporal lobe epilepsy.

BICS01 Mediates Reversible Anti-seizure Effects in Brain Slice Models of Epilepsy.

Drug-resistant epilepsy remains a significant clinical and societal burden, with one third of people with epilepsy continuing to experience seizures despite the availability of around 30 anti-seizure drugs (ASDs). Further, ASDs often have substantial adverse effects, including impacts on learning and memory. Therefore, it is important to develop new ASDs, which may be more potent or better tolerated. Here, we report the preliminary preclinical evaluation of BICS01, a synthetic product based on a natural compound, as a potential ASD. To model seizure-like activity in vitro, we prepared hippocampal slices from adult male Sprague Dawley rats, and elicited epileptiform bursting using high extracellular potassium. BICS01 (200 µM) rapidly and reversibly reduced the frequency of epileptiform bursting but did not change broad measures of network excitability or affect short-term synaptic facilitation. BICS01 was well tolerated following systemic injection at up to 1,000 mg/kg. However, we did not observe any protective effect of systemic BICS01 injection against acute seizures evoked by pentylenetetrazol. These results indicate that BICS01 is able to acutely reduce epileptiform activity in hippocampal networks. Further preclinical development studies to enhance pharmacokinetics and accumulation in the brain, as well as studies to understand the mechanism of action, are now required.

Integration of full divertor detachment with improved core confinement for tokamak fusion plasmas.

Divertor detachment offers a promising solution to the challenge of plasma-wall interactions for steady-state operation of fusion reactors. Here, we demonstrate the excellent compatibility of actively controlled full divertor detachment with a high-performance (ßN ~ 3, H98 ~ 1.5) core plasma, using high-ßp (poloidal beta, ßp > 2) scenario characterized by a sustained core internal transport barrier (ITB) and a modest edge transport barrier (ETB) in DIII-D tokamak. The high-ßp high-confinement scenario facilitates divertor detachment which, in turn, promotes the development of an even stronger ITB at large radius with a weaker ETB. This self-organized synergy between ITB and ETB, leads to a net gain in energy confinement, in contrast to the net confinement loss caused by divertor detachment in standard H-modes. These results show the potential of integrating excellent core plasma performance with an efficient divertor solution, an essential step towards steady-state operation of reactor-grade plasmas.

Dual actions of sphingosine-1-phosphate: extracellular through the Gi-coupled receptor Edg-1 and intracellular to regulate proliferation and survival.

Sphingosine-1-phosphate (SPP), a bioactive lipid, acts both intracellularly and extracellularly to cause pleiotropic biological responses. Recently, we identified SPP as a ligand for the G protein-coupled receptor Edg-1 (Lee, M.-J., J.R. Van Brocklyn, S. Thangada, C.H. Liu, A.R. Hand, R. Menzeleev, S. Spiegel, and T. Hla. 1998. Science. 279:1552-1555). Edg-1 binds SPP with remarkable specificity as only sphinganine-1-phosphate displaced radiolabeled SPP, while other sphingolipids did not. Binding of SPP to Edg-1 resulted in inhibition of forskolin-stimulated cAMP accumulation, in a pertussis toxin-sensitive manner. In contrast, two well-characterized biological responses of SPP, mitogenesis and prevention of apoptosis, were clearly unrelated to binding to Edg-1 and correlated with intracellular uptake. SPP also stimulated signal transduction pathways, including calcium mobilization, activation of phospholipase D, and tyrosine phosphorylation of p125(FAK), independently of edg-1 expression. Moreover, DNA synthesis in Swiss 3T3 fibroblasts was significantly and specifically increased by microinjection of SPP. Finally, SPP suppresses apoptosis of HL-60 and pheochromocytoma PC12 cells, which do not have specific SPP binding or expression of Edg-1 mRNA. Conversely, sphinganine-1-phosphate, which binds to and signals via Edg-1, does not have any significant cytoprotective effect. Thus, SPP is a prototype for a novel class of lipid mediators that act both extracellularly as ligands for cell surface receptors and intracellularly as second messengers.

Helium/Carbon dioxide ratios as premonitors of volcanic activity.

The composition of the gaseous emissions of two fumaroles at the summit of Kilauea Volcano was monitored for môre than 2 years. Magma was released from the summit reservoir on three occasions during this period; prior to or during each event the ratios of helium to carbon dioxide in the fumarole gases decreased substantially from that observed during periods of quiescence.

Altered Biogenesis and MicroRNA Content of Hippocampal Exosomes Following Experimental Status Epilepticus.

Repetitive or prolonged seizures (status epilepticus) can damage neurons within the hippocampus, trigger gliosis, and generate an enduring state of hyperexcitability. Recent studies have suggested that microvesicles including exosomes are released from brain cells following stimulation and tissue injury, conveying contents between cells including microRNAs (miRNAs). Here, we characterized the effects of experimental status epilepticus on the expression of exosome biosynthesis components and analyzed miRNA content in exosome-enriched fractions. Status epilepticus induced by unilateral intra-amygdala kainic acid in mice resulted in acute subfield-specific, bi-directional changes in hippocampal transcripts associated with exosome biosynthesis including up-regulation of endosomal sorting complexes required for transport (ESCRT)-dependent and -independent pathways. Increased expression of exosome components including Alix were detectable in samples obtained 2 weeks after status epilepticus and changes occurred in both the ipsilateral and contralateral hippocampus. RNA sequencing of exosome-enriched fractions prepared using two different techniques detected a rich diversity of conserved miRNAs and showed that status epilepticus selectively alters miRNA contents. We also characterized editing sites of the exosome-enriched miRNAs and found six exosome-enriched miRNAs that were adenosine-to-inosine (ADAR) edited with the majority of the editing events predicted to occur within miRNA seed regions. However, the prevalence of these editing events was not altered by status epilepticus. These studies demonstrate that status epilepticus alters the exosome pathway and its miRNA content, but not editing patterns. Further functional studies will be needed to determine if these changes have pathophysiological significance for epileptogenesis.

Estimating the performance of lithium beam measurements of current density and electron density in an H-mode pedestal.

The lithium beam is an effective diagnostic tool for investigation of stability and particle transport in the pedestal. It was used successfully to measure edge current density on DIII-D, achieving qualitative agreement with neoclassical models. Electron density profiles were also measured. Proposed upgrades will continue these measurements with higher reliability as well as explore edge current measurements using spectroscopy. The optics will be redesigned to optimize throughput and aperture broadening and to replace the photomultiplier tubes with avalanche photodiodes. The new system will yield detailed measurements of the pedestal, complementing existing diagnostics for investigating pedestal stability, edge localized mode cycle, and particle transport through the pedestal.

A review of machine learning in obesity.

Rich sources of obesity-related data arising from sensors, smartphone apps, electronic medical health records and insurance data can bring new insights for understanding, preventing and treating obesity. For such large datasets, machine learning provides sophisticated and elegant tools to describe, classify and predict obesity-related risks and outcomes. Here, we review machine learning methods that predict and/or classify such as linear and logistic regression, artificial neural networks, deep learning and decision tree analysis. We also review methods that describe and characterize data such as cluster analysis, principal component analysis, network science and topological data analysis. We introduce each method with a high-level overview followed by examples of successful applications. The algorithms were then applied to National Health and Nutrition Examination Survey to demonstrate methodology, utility and outcomes. The strengths and limitations of each method were also evaluated. This summary of machine learning algorithms provides a unique overview of the state of data analysis applied specifically to obesity.