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Endogenous levels of glucocorticoids rise during pregnancy to warrant development and maturation of the fetal organs close to birth. However, during most of the gestation, the fetus is protected from excessive biologically active endogenous glucocorticoids by placental and fetal expression of 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2). Maternal stress, which may overwhelm placental 11β-HSD2 activity with high glucocorticoid levels, or administration of synthetic glucocorticoids to improve the survival chances of the premature newborn, are associated to postnatal increased risk for immune diseases. Fetal exposure to excessive glucocorticoids may underlie this altered postnatal immunity. Here, we revise the role that placental and fetal 11β-HSD2, fetal glucocorticoid exposure, and programming of the offspring's the hypothalamic-pituitary-adrenal (HPA) axis play on concerted steps in immune fetal development. We could identify gaps in knowledge about glucocorticoid-induced programming of immune diseases. Finally, based on current evidence about glucocorticoid and HPA axis-mediated immune regulation, we hypothesize on mechanisms that could drive the enhanced risk for atopies, infections, and type I diabetes in offspring that were prenatally exposed to glucocorticoids.

Insufficient alveolar gas exchange capacity is a major contributor to lung disease. During lung development, a population of distal epithelial progenitors first produce bronchiolar-fated and subsequently alveolar-fated progeny. The mechanisms controlling this bronchiolar-to-alveolar developmental transition remain largely unknown. We developed a novel grafting assay to test if lung epithelial progenitors are intrinsically programmed or if alveolar cell identity is determined by environmental factors. These experiments revealed that embryonic lung epithelial identity is extrinsically determined. We show that both glucocorticoid and STAT3 signalling can control the timing of alveolar initiation, but that neither pathway is absolutely required for alveolar fate specification; rather, glucocorticoid receptor and STAT3 work in parallel to promote alveolar differentiation. Thus, developmental acquisition of lung alveolar fate is a robust process controlled by at least two independent extrinsic signalling inputs. Further elucidation of these pathways might provide therapeutic opportunities for restoring alveolar capacity., Competing Interests: The authors declare no competing or financial interests., (© 2016. Published by The Company of Biologists Ltd.)

Objective: To describe pediatric patients with convulsive refractory status epilepticus in whom there is intention to use an IV anesthetic for seizure control., Design: Two-year prospective observational study evaluating patients (age range, 1 mo to 21 yr) with refractory status epilepticus not responding to two antiepileptic drug classes and treated with continuous infusion of anesthetic agent., Setting: Nine pediatric hospitals in the United States., Patients: In a cohort of 111 patients with refractory status epilepticus (median age, 3.7 yr; 50% male), 54 (49%) underwent continuous infusion of anesthetic treatment., Main Results: The median (interquartile range) ICU length of stay was 10 (3-20) days. Up to four "cycles" of serial anesthetic therapy were used, and seizure termination was achieved in 94% by the second cycle. Seizure duration in controlled patients was 5.9 (1.9-34) hours for the first cycle and longer when a second cycle was required (30 [4-120] hr; p = 0.048). Midazolam was the most frequent first-line anesthetic agent (78%); pentobarbital was the most frequently used second-line agent after midazolam failure (82%). An electroencephalographic endpoint was used in over half of the patients; higher midazolam dosing was used with a burst suppression endpoint. In midazolam nonresponders, transition to a second agent occurred after a median of 1 day. Most patients (94%) experienced seizure termination with these two therapies., Conclusions: Midazolam and pentobarbital remain the mainstay of continuous infusion therapy for refractory status epilepticus in the pediatric patient. The majority of patients experience seizure termination within a median of 30 hours. These data have implications for the design and feasibility of future intervention trials. That is, testing a new anesthetic anticonvulsant after failure of both midazolam and pentobarbital is unlikely to be feasible in a pediatric study, whereas a decision to test an alternative to pentobarbital, after midazolam failure, may be possible in a multicenter multinational study.

Background: The rapid production and incorporation of engineered nanomaterials into consumer products alongside research suggesting nanomaterials can cause cell death and DNA damage (genotoxicity) makes in vitro assays desirable for nanosafety screening. However, conflicting outcomes are often observed when in vitro and in vivo study results are compared, suggesting more physiologically representative in vitro models are required to minimise reliance on animal testing., Method: BASF Levasil® silica nanoparticles (16 and 85 nm) were used to adapt the 3D reconstructed skin micronucleus (RSMN) assay for nanomaterials administered topically or into the growth medium. 3D dose-responses were compared to a 2D micronucleus assay using monocultured human B cells (TK6) after standardising dose between 2D / 3D assays by total nanoparticle mass to cell number. Cryogenic vitrification, scanning electron microscopy and dynamic light scattering techniques were applied to characterise in-medium and air-liquid interface exposures. Advanced transmission electron microscopy imaging modes (high angle annular dark field) and X-ray spectrometry were used to define nanoparticle penetration / cellular uptake in the intact 3D models and 2D monocultured cells., Results: For all 2D exposures, significant (p < 0.002) increases in genotoxicity were observed (≥100 μg/mL) alongside cell viability decreases (p < 0.015) at doses ≥200 μg/mL (16 nm-SiO2) and ≥100 μg/mL (85 nm-SiO2). In contrast, 2D-equivalent exposures to the 3D models (≤300 μg/mL) caused no significant DNA damage or impact on cell viability. Further increasing dose to the 3D models led to probable air-liquid interface suffocation. Nanoparticle penetration / cell uptake analysis revealed no exposure to the live cells of the 3D model occurred due to the protective nature of the skin model's 3D cellular microarchitecture (topical exposures) and confounding barrier effects of the collagen cell attachment layer (in-medium exposures). 2D monocultured cells meanwhile showed extensive internalisation of both silica particles causing (geno)toxicity., Conclusions: The results establish the importance of tissue microarchitecture in defining nanomaterial exposure, and suggest 3D in vitro models could play a role in bridging the gap between in vitro and in vivo outcomes in nanotoxicology. Robust exposure characterisation and uptake assessment methods (as demonstrated) are essential to interpret nano(geno)toxicity studies successfully.

Childhood nephrotic syndrome, in which steroid-dependence occurs concurrently with steroid-resistance, requires aggressive therapy to prevent relapse. Predictive biomarkers that can be used to stratify treatment are urgently needed. Here we report that reciprocal regulation of the glucocorticoid metabolizing enzymes, 11β-hydroxysteroid dehydrogenase types 1 and 2, is associated with steroid-responsiveness and disease remission in childhood nephrotic syndrome, potentially providing a marker to identify patients in which aggressive therapy is required., (Copyright © 2016 by the American Academy of Pediatrics.)

Invasive pneumococcal disease (IPD), caused by infection with Streptococcus pneumoniae, has a substantial global burden. There are over 90 known serotypes of S. pneumoniae with a considerable body of evidence supporting serotype-specific mortality rates immediately following IPD. This is the first study to consider the association between serotype and longer-term mortality following IPD. Using enhanced surveillance data from the North East of England we assessed both the short-term (30-day) and longer-term (⩽7 years) independent adjusted associations between individual serotypes and mortality following IPD diagnosis using logistic regression and extended Cox proportional hazards models. Of the 1316 cases included in the analysis, 243 [18·5%, 95% confidence interval (CI) 16·4-20·7] died within 30 days of diagnosis. Four serotypes (3, 6A, 9N, 19 F) were significantly associated with overall increased 30-day mortality. Effects were observable only for older adults (⩾60 years). After extension of the window to 12 months and 36 months, one serotype was associated with significantly increased mortality at 12 months (19 F), but no individual serotypes were associated with increased mortality at 36 months. Two serotypes had statistically significant hazard ratios (HR) for longer-term mortality: serotype 1 for reduced mortality (HR 0·51, 95% CI 0·30-0·86) and serotype 9N for increased mortality (HR 2·30, 95% CI 1·29-4·37). The association with serotype 9N was no longer observed after limiting survival analysis to an observation period starting 30 days after diagnosis. This study supports the evidence for associations between serotype and short-term (30-day) mortality following IPD and provides the first evidence for the existence of statistically significant associations between individual serotypes and longer-term variation in mortality following IPD.

Endogenous glucocorticoid action within cells is enhanced by prereceptor metabolism by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which converts intrinsically inert cortisone and 11-dehydrocorticosterone into active cortisol and corticosterone, respectively. 11β-HSD1 is highly expressed in immune cells elicited to the mouse peritoneum during thioglycollate-induced peritonitis and is down-regulated as the inflammation resolves. During inflammation, 11β-HSD1-deficient mice show enhanced recruitment of inflammatory cells and delayed acquisition of macrophage phagocytic capacity. However, the key cells in which 11β-HSD1 exerts these effects remain unknown. Here we have identified neutrophils (CD11b(+),Ly6G(+),7/4(+) cells) as the thioglycollate-recruited cells that most highly express 11β-HSD1 and show dynamic regulation of 11β-HSD1 in these cells during an inflammatory response. Flow cytometry showed high expression of 11β-HSD1 in peritoneal neutrophils early during inflammation, declining at later states. In contrast, expression in blood neutrophils continued to increase during inflammation. Ablation of monocytes/macrophages by treatment of CD11b-diphtheria-toxin receptor transgenic mice with diphtheria toxin prior to thioglycollate injection had no significant effect on 11β-HSD1 activity in peritoneal cells, consistent with neutrophils being the predominant 11β-HSD1 expressing cell type at this time. Similar to genetic deficiency in 11β-HSD1, acute inhibition of 11β-HSD1 activity during thioglycollate-induced peritonitis augmented inflammatory cell recruitment to the peritoneum. These data suggest that neutrophil 11β-HSD1 increases during inflammation to contribute to the restraining effect of glucocorticoids upon neutrophil-mediated inflammation. In human neutrophils, lipopolysaccharide activation increased 11β-HSD1 expression, suggesting the antiinflammatory effects of 11β-HSD1 in neutrophils may be conserved in humans.

Fetoplacental glucocorticoid overexposure is a significant mechanism underlying fetal growth restriction and the programming of adverse health outcomes in the adult. Placental glucocorticoid inactivation by 11β-hydroxysteroid dehydrogenase type 2 (11β-HSD2) plays a key role. We previously discovered that Hsd11b2(-/-) mice, lacking 11β-HSD2, show marked underdevelopment of the placental vasculature. We now explore the consequences for fetal cardiovascular development and whether this is reversible. We studied Hsd11b2(+/+), Hsd11b2(+/-), and Hsd11b2(-/-) littermates from heterozygous (Hsd11b(+/-)) matings at embryonic day (E)14.5 and E17.5, where all three genotypes were present to control for maternal effects. Using high-resolution ultrasound, we found that umbilical vein blood velocity in Hsd11b2(-/-) fetuses did not undergo the normal gestational increase seen in Hsd11b2(+/+) littermates. Similarly, the resistance index in the umbilical artery did not show the normal gestational decline. Surprisingly, given that 11β-HSD2 absence is predicted to initiate early maturation, the E/A wave ratio was reduced at E17.5 in Hsd11b2(-/-) fetuses, suggesting impaired cardiac function. Pravastatin administration from E6.5, which increases placental vascular endothelial growth factor A and, thus, vascularization, increased placental fetal capillary volume, ameliorated the aberrant umbilical cord velocity, normalized fetal weight, and improved the cardiac function of Hsd11b2(-/-) fetuses. This improved cardiac function occurred despite persisting indications of increased glucocorticoid exposure in the Hsd11b2(-/-) fetal heart. Thus, the pravastatin-induced enhancement of fetal capillaries within the placenta and the resultant hemodynamic changes correspond with restored fetal cardiac function. Statins may represent a useful therapeutic approach to intrauterine growth retardation due to placental vascular hypofunction.

The efficacy of mineralocorticoid receptor (MR) antagonism in the treatment of certain patients with heart failure has highlighted the pivotal role of aldosterone and MR in heart disease. The glucocorticoid (GC) receptor (GR) is also expressed in heart, but the role of cardiac GR had received much less attention until recently. GR and MR are highly homologous in both structure and function, although not in cellular readout. Recent evidence in animal models has uncovered a tonic role for GC action via GR in cardiomyocytes in prevention of heart disease. Here, we review this evidence and the implications for a balance between GR and MR activation in the early life maturation of the heart and its subsequent health and disease., (Copyright © 2015 Elsevier Ltd. All rights reserved.)

Global deficiency of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), an enzyme that regenerates glucocorticoids within cells, promotes angiogenesis, and reduces acute infarct expansion after myocardial infarction (MI), suggesting that 11β-HSD1 activity has an adverse influence on wound healing in the heart after MI. The present study investigated whether 11β-HSD1 deficiency could prevent the development of heart failure after MI and examined whether 11β-HSD1 deficiency in cardiomyocytes and vascular smooth muscle cells confers this protection. Male mice with global deficiency in 11β-HSD1, or with Hsd11b1 disruption in cardiac and vascular smooth muscle (via SM22α-Cre recombinase), underwent coronary artery ligation for induction of MI. Acute injury was equivalent in all groups. However, by 8 weeks after induction of MI, relative to C57Bl/6 wild type, globally 11β-HSD1-deficient mice had reduced infarct size (34.7 ± 2.1% left ventricle [LV] vs 44.0 ± 3.3% LV, P = .02), improved function (ejection fraction, 33.5 ± 2.5% vs 24.7 ± 2.5%, P = .03) and reduced ventricular dilation (LV end-diastolic volume, 0.17 ± 0.01 vs 0.21 ± 0.01 mL, P = .01). This was accompanied by a reduction in hypertrophy, pulmonary edema, and in the expression of genes encoding atrial natriuretic peptide and β-myosin heavy chain. None of these outcomes, nor promotion of periinfarct angiogenesis during infarct repair, were recapitulated when 11β-HSD1 deficiency was restricted to cardiac and vascular smooth muscle. 11β-HSD1 expressed in cells other than cardiomyocytes or vascular smooth muscle limits angiogenesis and promotes infarct expansion with adverse ventricular remodeling after MI. Early pharmacological inhibition of 11β-HSD1 may offer a new therapeutic approach to prevent heart failure associated with ischemic heart disease.

Mice deficient in the glucocorticoid-regenerating enzyme 11β-HSD1 resist age-related spatial memory impairment. To investigate the mechanisms and pathways involved, we used microarrays to identify differentially expressed hippocampal genes that associate with cognitive ageing and 11β-HSD1. Aged wild-type mice were separated into memory-impaired and unimpaired relative to young controls according to their performance in the Y-maze. All individual aged 11β-HSD1-deficient mice showed intact spatial memory. The majority of differentially expressed hippocampal genes were increased with ageing (e.g. immune/inflammatory response genes) with no genotype differences. However, the neuronal-specific transcription factor, Npas4, and immediate early gene, Arc, were reduced (relative to young) in the hippocampus of memory-impaired but not unimpaired aged wild-type or aged 11β-HSD1-deficient mice. A quantitative reverse transcriptase-polymerase chain reaction and in situ hybridisation confirmed reduced Npas4 and Arc mRNA expression in memory-impaired aged wild-type mice. These findings suggest that 11β-HSD1 may contribute to the decline in Npas4 and Arc mRNA levels associated with memory impairment during ageing, and that decreased activity of synaptic plasticity pathways involving Npas4 and Arc may, in part, underlie the memory deficits seen in cognitively-impaired aged wild-type mice., (© 2015 The Authors. Journal of Neuroendocrinology published by John Wiley & Sons Ltd on behalf of British Society for Neuroendocrinology.)

Background: Transient early-life perturbations in glucocorticoids (GC) are linked with cardiovascular disease risk in later life. Here the impact of early life manipulations of GC on adult heart structure, function and gene expression were assessed., Methods and Results: Zebrafish embryos were incubated in dexamethasone (Dex) or injected with targeted glucocorticoid receptor (GR) morpholino knockdown (GR Mo) over the first 120 h post fertilisation (hpf); surviving embryos (>90%) were maintained until adulthood under normal conditions. Cardiac function, heart histology and cardiac genes were assessed in embryonic (120 hpf) and adult (120 days post fertilisation (dpf)) hearts. GR Mo embryos (120 hpf) had smaller hearts with fewer cardiomyocytes, less mature striation pattern, reduced cardiac function and reduced levels of vmhc and igf mRNA compared with controls. GR Mo adult hearts were smaller with diminished trabecular network pattern, reduced expression of vmhc and altered echocardiographic Doppler flow compared to controls. Dex embryos had larger hearts at 120 hpf (Dex 107.2 ± 3.1 vs. controls 90.2 ± 1.1 μm, p < 0.001) with a more mature trabecular network and larger cardiomyocytes (1.62 ± 0.13 cells/μm vs control 2.18 ± 0.13 cells/μm, p < 0.05) and enhanced cardiac performance compared to controls. Adult hearts were larger (1.02 ± 0.07 μg/mg vs controls 0.63 ± 0.06 μg/mg, p = 0.0007), had increased vmhc and gr mRNA levels., Conclusion: Perturbations in GR activity during embryonic development results in short and long-term alterations in the heart., (Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.)

In epileptic encephalopathy, the seizures and interictal epileptiform activity create additional neurocognitive dysfunction beyond that due to the underlying etiology. Treatment leading to a reduction in seizures or interictal abnormalities may help improve neurocognitive function in these situations. The focus of our discussion is reviewing data that support the concept that treatment can impact outcome independent of the etiology in some cases., (Wiley Periodicals, Inc. © 2015 International League Against Epilepsy.)

OBJECT Seizures may cause diagnostic confusion and be a source of metabolic stress after traumatic brain injury (TBI) in children. The incidence of electroencephalography (EEG)-confirmed seizures and of subclinical seizures in the pediatric population with TBI is not well known. METHODS A routine protocol for continuous EEG (cEEG) monitoring was initiated for all patients with moderate or severe TBI at a Level 1 pediatric trauma center. Over a 3.5-year period, all patients with TBI who underwent cEEG monitoring, both according to protocol and those with mild head injuries who underwent cEEG monitoring at the discretion of the treating team, were identified prospectively. Clinical data were collected and analyzed. RESULTS Over the study period, 594 children were admitted with TBI, and 144 of these children underwent cEEG monitoring. One hundred two (71%) of these 144 children had moderate or severe TBI. Abusive head trauma (AHT) was the most common mechanism of injury (65 patients, 45%) in children with cEEG monitoring. Seizures were identified on cEEG in 43 patients (30%). Forty (93%) of these 43 patients had subclinical seizures, including 17 (40%) with only subclinical seizures and 23 (53%) with both clinical and subclinical seizures. Fifty-three percent of patients with seizures experienced status epilepticus. Age less than 2.4 years and AHT mechanism were strongly correlated with presence of seizures (odds ratios 8.7 and 6.0, respectively). Those patients with only subclinical seizures had the same risk factors as the other groups. The presence of seizures did not correlate with discharge disposition but was correlated with longer hospital stay and intensive care unit stay. CONCLUSIONS Continuous EEG monitoring identifies a significant number of subclinical seizures acutely after TBI. Children younger than 2.4 years of age and victims of AHT are particularly vulnerable to subclinical seizures, and seizures in general. Continuous EEG monitoring allows for accurate diagnosis and timely treatment of posttraumatic seizures, and may mitigate secondary injury to the traumatized brain.

Glucocorticoid levels rise dramatically in late gestation to mature foetal organs in readiness for postnatal life. Immature heart function may compromise survival. Cardiomyocyte glucocorticoid receptor (GR) is required for the structural and functional maturation of the foetal heart in vivo, yet the molecular mechanisms are largely unknown. Here we asked if GR activation in foetal cardiomyocytes in vitro elicits similar maturational changes. We show that physiologically relevant glucocorticoid levels improve contractility of primary-mouse-foetal cardiomyocytes, promote Z-disc assembly and the appearance of mature myofibrils, and increase mitochondrial activity. Genes induced in vitro mimic those induced in vivo and include PGC-1α, a critical regulator of cardiac mitochondrial capacity. SiRNA-mediated abrogation of the glucocorticoid induction of PGC-1α in vitro abolished the effect of glucocorticoid on myofibril structure and mitochondrial oxygen consumption. Using RNA sequencing we identified a number of transcriptional regulators, including PGC-1α, induced as primary targets of GR in foetal cardiomyocytes. These data demonstrate that PGC-1α is a key mediator of glucocorticoid-induced maturation of foetal cardiomyocyte structure and identify other candidate transcriptional regulators that may play critical roles in the transition of the foetal to neonatal heart.

Objective: To describe the time elapsed from onset of pediatric convulsive status epilepticus (SE) to administration of antiepileptic drug (AED)., Methods: This was a prospective observational cohort study performed from June 2011 to June 2013. Pediatric patients (1 month-21 years) with convulsive SE were enrolled. In order to study timing of AED administration during all stages of SE, we restricted our study population to patients who failed 2 or more AED classes or needed continuous infusions to terminate convulsive SE., Results: We enrolled 81 patients (44 male) with a median age of 3.6 years. The first, second, and third AED doses were administered at a median (p25-p75) time of 28 (6-67) minutes, 40 (20-85) minutes, and 59 (30-120) minutes after SE onset. Considering AED classes, the initial AED was a benzodiazepine in 78 (96.3%) patients and 2 (2-3) doses of benzodiazepines were administered before switching to nonbenzodiazepine AEDs. The first and second doses of nonbenzodiazepine AEDs were administered at 69 (40-120) minutes and 120 (75-296) minutes. In the 64 patients with out-of-hospital SE onset, 40 (62.5%) patients did not receive any AED before hospital arrival. In the hospital setting, the first and second in-hospital AED doses were given at 8 (5-15) minutes and 16 (10-40) minutes after SE onset (for patients with in-hospital SE onset) or after hospital arrival (for patients with out-of-hospital SE onset)., Conclusions: The time elapsed from SE onset to AED administration and escalation from one class of AED to another is delayed, both in the prehospital and in-hospital settings., (© 2015 American Academy of Neurology.)

The glucocorticoid receptor (GR) is a member of the nuclear receptor superfamily, which controls programs regulating cell proliferation, differentiation, and apoptosis. We have identified an unexpected role for GR in mitosis. We discovered that specifically modified GR species accumulate at the mitotic spindle during mitosis in a distribution that overlaps with Aurora kinases. We found that Aurora A was required to mediate mitosis-driven GR phosphorylation, but not recruitment of GR to the spindle. GR was necessary for mitotic progression, with increased time to complete mitosis, frequency of mitotic aberrations, and death in mitosis observed following GR knockdown. Complementation studies revealed an essential role for the GR ligand-binding domain, but no clear requirement for ligand binding in regulating chromosome segregation. The GR N-terminal domain, and specifically phosphosites S203 and S211, were not required. Reduced GR expression results in a cell cycle phenotype, with isolated cells from mouse and human subjects showing changes in chromosome content over prolonged passage. Furthermore, GR haploinsufficient mice have an increased incidence of tumor formation, and, strikingly, these tumors are further depleted for GR, implying additional GR loss as a consequence of cell transformation. We identified reduced GR expression in a panel of human liver, lung, prostate, colon, and breast cancers. We therefore reveal an unexpected role for the GR in promoting accurate chromosome segregation during mitosis, which is causally linked to tumorigenesis, making GR an authentic tumor suppressor gene.

Risk assessment of human exposure to chemicals is crucial for understanding whether such agents can cause cancer. The current emphasis on avoidance of animal testing has placed greater importance on in vitro tests for the identification of genotoxicants. Selection of an appropriate in vitro dosing regime is imperative in determining the genotoxic effects of test chemicals. Here, the issue of dosing approaches was addressed by comparing acute and chronic dosing, uniquely using low-dose experiments. Acute 24 h exposures were compared with equivalent dosing every 24 h over 5-day, fractionated treatment periods. The in vitro micronucleus assay was used to measure clastogenicity induced by methyl methanesulfonate (MMS) and N-methyl-N-nitrosourea (MNU) in human lymphoblastoid cell line, TK6. Quantitative real-time (qRT) PCR was used to measure mRNA level induction of DNA repair enzymes. Lowest observed genotoxic effect levels (LOGELs) for MMS were obtained at 0.7 µg/ml for the acute study and 1.0 µg/ml for the chronic study. For acute MNU dosing, a LOGEL was observed at 0.46 µg/ml, yet genotoxicity was completely removed following the chronic study. Interestingly, acute MNU dosing demonstrated a statistically significant decrease at 0.009 µg/ml. Levels of selected DNA repair enzymes did not change significantly following doses tested. However, p53 deficiency (using the TK6-isogenic cell line, NH32) increased sensitivity to MMS during chronic dosing, causing this LOGEL to equate to the acute treatment LOGEL. In the context of the present data for 2 alkylating agents, chronic dosing could be a valuable in vitro supplement to acute dosing and could contribute to reduction of unnecessary in vivo follow-up tests., (© The Author 2015. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Objective: Oral cannabis extracts (OCEs) have been used in the treatment of epilepsy; however, no studies demonstrate clear efficacy. We report on a cohort of pediatric patients with epilepsy who were given OCE and followed in a single tertiary epilepsy center., Methods: A retrospective chart review of children and adolescents who were given OCE for treatment of their epilepsy was performed., Results: Seventy-five patients were identified of which 57% reported any improvement in seizure control and 33% reported a >50% reduction in seizures (responders). If the family had moved to CO for OCE treatment, the responder rate was 47% vs. 22% for children who already were in CO. The responder rate varied based on epilepsy syndrome: Dravet 23%, Doose 0%, and Lennox-Gastaut syndrome (LGS) 88.9%. The background EEG of the 8 responders where EEG data were available was not improved. Additional benefits reported included: improved behavior/alertness (33%), improved language (10%), and improved motor skills (10%). Adverse events (AEs) occurred in 44% of patients including increased seizures (13%) and somnolence/fatigue (12%). Rare adverse events included developmental regression, abnormal movements, status epilepticus requiring intubation, and death., Significance: Our retrospective study of OCE use in pediatric patients with epilepsy demonstrates that some families reported patient improvement with treatment; however, we also found a variety of challenges and possible confounding factors in studying OCE retrospectively in an open-labeled fashion. We strongly support the need for controlled, blinded studies to evaluate the efficacy and safety of OCE for treatment of pediatric epilepsies using accurate seizure counts, formal neurocognitive assessments, as well as EEG as a biomarker. This study provides Class III evidence that OCE is well tolerated by children and adolescents with epilepsy., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Purpose: Electrographic seizures are common in encephalopathic critically ill children, but identification requires continuous EEG monitoring (CEEG). Development of a seizure prediction model would enable more efficient use of limited CEEG resources. We aimed to develop and validate a seizure prediction model for use among encephalopathic critically ill children., Method: We developed a seizure prediction model using a retrospectively acquired multi-center database of children with acute encephalopathy without an epilepsy diagnosis, who underwent clinically indicated CEEG. We performed model validation using a separate prospectively acquired single center database. Predictor variables were chosen to be readily available to clinicians prior to the onset of CEEG and included: age, etiology category, clinical seizures prior to CEEG, initial EEG background category, and inter-ictal discharge category., Results: The model has fair to good discrimination ability and overall performance. At the optimal cut-off point in the validation dataset, the model has a sensitivity of 59% and a specificity of 81%. Varied cut-off points could be chosen to optimize sensitivity or specificity depending on available CEEG resources., Conclusion: Despite inherent variability between centers, a model developed using multi-center CEEG data and few readily available variables could guide the use of limited CEEG resources when applied at a single center. Depending on CEEG resources, centers could choose lower cut-off points to maximize identification of all patients with seizures (but with more patients monitored) or higher cut-off points to reduce resource utilization by reducing monitoring of lower risk patients (but with failure to identify some patients with seizures)., (Copyright © 2014 British Epilepsy Association. Published by Elsevier Ltd. All rights reserved.)

The 5-hydroxytryptamine2C (5-HT)2C receptor is widely implicated in the aetiology of affective and eating disorders as well as regulation of the hypothalamo-pituitary-adrenal axis. Signalling through this receptor is regulated by A-to-I RNA editing, affecting three amino acids in the protein sequence, with unedited transcripts encoding a receptor (INI) that, in vitro, is hyperactive compared with edited isoforms. Targeted alteration (knock-in) of the Htr2c gene to generate 'INI' mice with no alternate splicing, solely expressing the full-length unedited isoform, did not produce an overt metabolic phenotype or altered anxiety behaviour, but did display reduced depressive-like and fear-associated behaviours. INI mice exhibited a hyperactive hypothalamo-pituitary-adrenal axis, with increased nadir plasma corticosterone and corticotrophin-releasing hormone expression in the hypothalamus but responded normally to chronic stress and showed normal circadian activity and activity in a novel environment. The circadian patterns of 5-HT2C receptor mRNA and mbii52, a snoRNA known to regulate RNA editing and RNA splicing of 5-HT2C receptor pre-mRNA, were altered in INI mice compared with wild-type control mice. Moreover, levels of 5-HT1A receptor mRNA were increased in the hippocampus of INI mice. These gene expression changes may underpin the neuroendocrine and behavioural changes observed in INI mice. However, the phenotype of INI mice was not consistent with a globally hyperactive INI receptor encoded by the unedited transcript in the absence of alternate splicing. Hence, the in vivo outcome of RNA editing may be neuronal cell type specific., (© 2014 The Authors. European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Rasmussen encephalitis (RE) is an inflammatory unilateral progressive medically refractory epilepsy associated with hemiparesis, cognitive dysfunction, and hemispheric atrophy. Here, we present 2 cases from our institution that demonstrate the dual nature of RE in 2 similarly aged children. Overall, 2 types of RE have been described: type 1 has a short prodromal phase and more explosive onset and type 2 has a longer prodromal of partial seizures followed by hemiparesis and atrophy. Younger patients are more likely to fit into the type 1 presentation and have been described as more likely to have dual pathology. Perhaps the patients with a more acute onset are more likely to have a dual pathology as is found in our 2 cases. We review the typical findings in RE and discuss current treatment options, highlighting new experimental treatments., (Copyright © 2014 Elsevier Inc. All rights reserved.)

Genome-wide association studies have consistently implicated the interleukin-15 (IL-15) gene in acute lymphoblastic leukemia (ALL) biology, including associations with disease susceptibility, and increased risk of central nervous system (CNS) involvement. However, whether pre-B ALL blasts directly respond to IL-15 is unknown. Here, we show that most pre-B ALL primary samples and cell lines express IL-15 and components of its receptor and that primary pre-B ALL cells show increased growth in culture in response to IL-15. Investigation of mechanisms of action using IL-15-responsive SD-1 cells shows this growth advantage is maximal under low-serum conditions, mimicking those found in cerebrospinal fluid. IL-15 also upregulates PSGL-1 and CXCR3, molecules associated with CNS trafficking. Investigation of downstream signaling pathways indicates that IL-15 induces signal transducer and activator of transcription 5 (STAT5), extracellular signal-regulated kinase (ERK) 1/2, and to a lesser extent phosphatidylinositol 3-kinase (PI3K) and nuclear factor κB (NF-κB) phosphorylation. The IL-15-mediated growth advantage is abolished by mitogen-activated protein kinase kinase/extracellular signal-regulated kinase (MEK/ERK), PI3K, and NF-κB inhibitors but preserved in the presence of STAT5 inhibition. Together, these observations provide a mechanistic link between increased levels of IL-15 expression and leukemogenesis, high-risk disease, and CNS relapse and suggest potential therapeutic targets., (© 2014 by The American Society of Hematology.)

Micronucleus (MN) induction is an established cytogenetic end point for evaluating structural and numerical chromosomal alterations in genotoxicity testing. A semi-automated scoring protocol for the assessment of MN preparations from human cell lines and a 3D skin cell model has been developed and validated. Following exposure to a range of test agents, slides were stained with 4'-6-diamidino-2-phenylindole (DAPI) and scanned by use of the MicroNuc module of metafer 4, after the development of a modified classifier for selecting MN in binucleate cells. A common difficulty observed with automated systems is an artefactual output of high false positives, in the case of the metafer system this is mainly due to the loss of cytoplasmic boundaries during slide preparation. Slide quality is paramount to obtain accurate results. We show here that to avoid elevated artefactual-positive MN outputs, diffuse cell density and low-intensity nuclear staining are critical. Comparisons between visual (Giemsa stained) and automated (DAPI stained) MN frequencies and dose-response curves were highly correlated (R (2) = 0.70 for hydrogen peroxide, R (2) = 0.98 for menadione, R (2) = 0.99 for mitomycin C, R (2) = 0.89 for potassium bromate and R (2) = 0.68 for quantum dots), indicating the system is adequate to produce biologically relevant and reliable results. Metafer offers many advantages over conventional scoring including increased output and statistical power, and reduced scoring subjectivity, labour and costs. Further, the metafer system is easily adaptable for use with a range of different cells, both suspension and adherent human cell lines. Awareness of the points raised here reduces the automatic positive errors flagged and drastically reduces slide scoring time, making metafer an ideal candidate for genotoxic biomonitoring and population studies and regulatory genotoxic testing.

Purpose: Status epilepticus (SE) is a life-threatening condition that can be refractory to initial treatment. Randomized controlled studies to guide treatment choices, especially beyond first-line drugs, are not available. This report summarizes the evidence that guides the management of refractory convulsive SE (RCSE) in children, defines gaps in our clinical knowledge and describes the development and works of the 'pediatric Status Epilepticus Research Group' (pSERG)., Methods: A literature review was performed to evaluate current gaps in the pediatric SE and RCSE literature. In person and online meetings helped to develop and expand the pSERG network., Results: The care of pediatric RCSE is largely based on extrapolations of limited evidence derived from adult literature and supplemented with case reports and case series in children. No comparative effectiveness trials have been performed in the pediatric population. Gaps in knowledge include risk factors for SE, biomarkers of SE and RCSE, second- and third-line treatment options, and long-term outcome., Conclusion: The care of children with RCSE is based on limited evidence. In order to address these knowledge gaps, the multicenter pSERG was established to facilitate prospective collection, analysis, and sharing of de-identified data and biological specimens from children with RCSE. These data will allow identification of treatment strategies associated with better outcomes and delineate evidence-based interventions to improve the care of children with SE., (Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Objective: To describe the prevalence, characteristics, and predictors of electrographic seizures after convulsive status epilepticus (CSE)., Study Design: This was a multicenter retrospective study in which we describe clinical and electroencephalographic (EEG) features of children (1 month to 21 years) with CSE who underwent continuous EEG monitoring., Results: Ninety-eight children (53 males) with CSE (median age of 5 years) underwent subsequent continuous EEG monitoring after CSE. Electrographic seizures (with or without clinical correlate) were identified in 32 subjects (33%). Eleven subjects (34.4%) had electrographic-only seizures, 17 subjects (53.1%) had electroclinical seizures, and 4 subjects (12.5%) had an unknown clinical correlate. Of the 32 subjects with electrographic seizures, 15 subjects (46.9%) had electrographic status epilepticus. Factors associated with the occurrence of electrographic seizures after CSE were a previous diagnosis of epilepsy (P = .029) and the presence of interictal epileptiform discharges (P < .0005). The median (p25-p75) duration of stay in the pediatric intensive care unit was longer for children with electrographic seizures than for children without electrographic seizures (9.5 [3-22.5] vs 2 [2-5] days, Wilcoxon test, Z = 3.916, P = .0001). Four children (4.1%) died before leaving the hospital, and we could not identify a relationship between death and the presence or absence of electrographic seizures., Conclusions: After CSE, one-third of children who underwent EEG monitoring experienced electrographic seizures, and among these, one-third experienced entirely electrographic-only seizures. A previous diagnosis of epilepsy and the presence of interictal epileptiform discharges were risk factors for electrographic seizures., (Copyright © 2014 Mosby, Inc. All rights reserved.)

Glucocorticoids are steroid hormones, essential in mammals to prepare for life after birth. Blood levels of glucocorticoids (cortisol in most mammals including humans; corticosterone in rats and mice) rise dramatically shortly before birth. This is mimicked clinically in the routine administration of synthetic glucocorticoids to pregnant women threatened by a preterm birth or to preterm infants to improve neonatal survival. Whilst effects on lung are well documented and essential for postnatal survival, those on heart are less well known. In this study, we review recent evidence for a crucial role of glucocorticoids in late gestational heart maturation. Either insufficient or excessive glucocorticoid exposure before birth may alter the normal glucocorticoid-regulated trajectory of heart maturation with potential life-long consequences.

Context: Levels of 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1), which regenerates active glucocorticoids, are selectively elevated in adipose tissue in human obesity and metabolic syndrome, both conditions associated with chronic low-grade inflammation. 11β-HSD1 expression is induced by proinflammatory cytokines in a variety of cell types, including in human adipocytes differentiated in vitro., Objective: Our objective was to determine the mechanisms by which proinflammatory cytokines induce 11β-HSD1 in human adipocytes., Results: The proinflammatory cytokines IL-1α (10 ng/mL) and TNFα (20 ng/mL) increased 11β-HSD1 mRNA levels in human primary adipocyte fractions and Simpson-Golabi-Behmel syndrome (SGBS) adipocytes (P<.001). Inhibition of the MAPK/ERK kinase (MEK) attenuated CCAAT/enhancer binding protein (C/EBP) β phosphorylation at Thr235 and IL-1α/TNFα induction of 11β-HSD1 (P≤.007). The small interfering RNA-mediated knockdown of C/EBPβ and nuclear factor (NF)-κB/RelA or inhibition of NF-κB/RelA also attenuated cytokine induction of 11β-HSD1 (P≤.001). Moreover, induction of 11β-HSD1 by IL-1α in SGBS cells was associated with nuclear localization of C/EBPβ and NF-κB/RelA. Chromatin immunoprecipitation experiments showed C/EBPβ and NF-κB/RelA located to the 11β-HSD1 promoter in human adipose tissue. Treatment of adipocyte fractions or SGBS adipocytes with metformin or acetylsalicylic acid, which target C/EBPβ and NF-κB/RelA signaling, attenuated the IL-1α induction of 11β-HSD1 (P≤.002)., Conclusions: Increased proinflammatory signaling in inflamed adipose tissue may mediate elevated 11β-HSD1 expression at this site via MEK, C/EBPβ, and NF-κB/RelA. These molecules/signaling pathways are, therefore, potential targets for drugs, including metformin and acetylsalicylic acid, to prevent/decreased up-regulation of 11β-HSD1 in human obese/metabolic syndrome adipose tissue.

Background: Sarcopenia, the loss of muscle mass and function with age, is associated with increased morbidity and mortality. Current understanding of the underlying mechanisms is limited. Glucocorticoids (GC) in excess cause muscle weakness and atrophy. We hypothesized that GC may contribute to sarcopenia through elevated circulating levels or increased glucocorticoid receptor (GR) signaling by increased expression of either GR or the GC-amplifying enzyme 11 beta-hydroxysteroid dehydrogenase type 1 (11βHSD1) in muscle., Methods: There were 82 participants; group 1 comprised 33 older men (mean age 70.2 years, SD 4.4) and 19 younger men (22.2 years, 1.7) and group 2 comprised 16 older men (79.1 years, 3.4) and 14 older women (80.1 years, 3.7). We measured muscle strength, mid-thigh cross-sectional area, fasting morning plasma cortisol, quadriceps muscle GR and 11βHSD1 mRNA, and urinary glucocorticoid metabolites. Data were analysed using multiple linear regression adjusting for age, gender and body size., Results: Muscle strength and size were not associated with plasma cortisol, total urinary glucocorticoids or the ratio of urinary 5β-tetrahydrocortisol +5α-tetrahydrocortisol to tetrahydrocortisone (an index of systemic 11βHSD activity). Muscle strength was associated with 11βHSD1 mRNA levels (β -0.35, p = 0.04), but GR mRNA levels were not significantly associated with muscle strength or size., Conclusion: Although circulating levels of GC are not associated with muscle strength or size in either gender, increased cortisol generation within muscle by 11βHSD1 may contribute to loss of muscle strength with age, a key component of sarcopenia. Inhibition of 11βHSD1 may have therapeutic potential in sarcopenia.

Background: Some communicable diseases disproportionately affect poor and vulnerable groups. Invasive pneumococcal disease (IPD) is an important cause of morbidity and mortality; however, the relationship between IPD and deprivation has not been well described., Methods: Population based study assessing the relationship between incidence of IPD and deprivation in the North East of England using data from an enhanced IPD surveillance system and the 2010 Indices of Multiple Deprivation and the Rural and Urban Area Classification., Results: The incidence of IPD increased linearly with increasing deprivation from 7.0 per 100 000 population to 13.6 per 100 000 population. This association was demonstrated for the 16-64 and ≥65 year age groups, but not the <16 year age group. IPD incidence was strongly associated with all individual domains of deprivation except for the 'barriers to housing and services' domain. IPD incidence was higher in urban than rural areas., Conclusions: The risk of IPD is strongly associated with deprivation in adults, but not children. The mechanisms producing the associations observed remain unclear and require further investigation. Findings from this study reinforce the need to address social inequalities to reduce the burden of disease. Targeting vaccination at adults living in deprived areas could reduce the burden of IPD.

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) converts inert glucocorticoids into active forms, thereby increasing intracellular glucocorticoid levels, important to restrain acute inflammation. 11β-HSD1 is induced by pro-inflammatory cytokines in a variety of cells. Here, we show 11β-HSD1 expression in human A549 epithelial cells is increased by pro-inflammatory cytokines (IL-1α/TNFα) via the P2 promoter of the HSD11B1 gene. Inhibition of p38 MAPK attenuated the pro-inflammatory cytokine induction of mRNA encoding 11β-HSD1 as well as that encoding C/EBPβ. IL-1α/TNFα-induced phosphorylation of C/EBPβ at Thr235 was also attenuated by p38 MAPK inhibition suggesting involvement of a p38 MAPK-C/EBPβ pathway. siRNA-mediated knock-down of C/EBPβ and NF-κB/RelA implicated both transcription factors in the IL-1α/TNFα induction of HSD11B1 mRNA. Transient transfections of HSD11B1 promoter-reporter constructs identified the proximal region of the P2 promoter of HSD11B1 as essential for this induction. IL-1α increased binding of C/EBPβ to the HSD11B1 P2 promoter, but this was not observed for NF-κB/RelA, suggesting indirect regulation by NF-κB/RelA. Ectopic expression of mutant chicken C/EBPβ constructs unable to undergo phosphorylation at the threonine equivalent to Thr235 attenuated the IL-1α-induction of HSD11B1, whereas mimicking constitutive phosphorylation of Thr235 (by mutation to aspartate) increased basal expression of HSD11B1 mRNA without affecting IL-1α-induced levels. These data clearly demonstrate a role for both C/EBPβ and NF-κB/RelA in the pro-inflammatory cytokine induction of HSD11B1 in human epithelial cells and show that p38 MAPK-induced phosphorylation of C/EBPβ at Thr235 is critical in this.

Since the discovery of cortisone in the 1940s and its early success in treatment of rheumatoid arthritis, glucocorticoids have remained the mainstay of anti-inflammatory therapies. However, cortisone itself is intrinsically inert. To be effective, it requires conversion to cortisol, the active glucocorticoid, by the enzyme 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1). Despite the identification of 11β-HSD in liver in 1953 (which we now know to be 11β-HSD1), its physiological role has been little explored until recently. Over the past decade, however, it has become apparent that 11β-HSD1 plays an important role in shaping endogenous glucocorticoid action. Acute inflammation is more severe with 11β-HSD1-deficiency or inhibition, yet in some inflammatory settings such as obesity or diabetes, 11β-HSD1-deficiency/inhibition is beneficial, reducing inflammation. Current evidence suggests both beneficial and detrimental effects may result from 11β-HSD1 inhibition in chronic inflammatory disease. Here we review recent evidence pertaining to the role of 11β-HSD1 in inflammation. This article is part of a Special Issue entitled 'CSR 2013'., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

Glucocorticoids are vital for the structural and functional maturation of foetal organs, yet excessive foetal exposure is detrimental to adult cardiovascular health. To elucidate the role of glucocorticoid signalling in late-gestation cardiovascular maturation, we have generated mice with conditional disruption of glucocorticoid receptor (GR) in cardiomyocytes and vascular smooth muscle cells using smooth muscle protein 22-driven Cre recombinase (SMGRKO mice) and compared them with mice with global deficiency in GR (GR(-/-)). Echocardiography shows impaired heart function in both SMGRKO and GR(-/-) mice at embryonic day (E)17.5, associated with generalized oedema. Cardiac ultrastructure is markedly disrupted in both SMGRKO and GR(-/-) mice at E17.5, with short, disorganized myofibrils and cardiomyocytes that fail to align in the compact myocardium. Failure to induce critical genes involved in contractile function, calcium handling and energy metabolism underpins this common phenotype. However, although hearts of GR(-/-) mice are smaller, with 22% reduced ventricular volume at E17.5, SMGRKO hearts are normally sized. Moreover, while levels of mRNA encoding atrial natriuretic peptide are reduced in E17.5 GR(-/-) hearts, they are normal in foetal SMGRKO hearts. These data demonstrate that structural, functional and biochemical maturation of the foetal heart is dependent on glucocorticoid signalling within cardiomyocytes and vascular smooth muscle, though some aspects of heart maturation (size, ANP expression) are independent of GR at these key sites.

Purpose: Survey data indicate that continuous electroencephalography (EEG) (CEEG) monitoring is used with increasing frequency to identify electrographic seizures in critically ill children, but studies of current CEEG practice have not been conducted. We aimed to describe the clinical utilization of CEEG in critically ill children at tertiary care hospitals with a particular focus on variables essential for designing feasible prospective multicenter studies evaluating the impact of electrographic seizures on outcome., Methods: Eleven North American centers retrospectively enrolled 550 consecutive critically ill children who underwent CEEG. We collected data regarding subject characteristics, CEEG indications, and CEEG findings., Key Findings: CEEG indications were encephalopathy with possible seizures in 67% of subjects, event characterization in 38% of subjects, and management of refractory status epilepticus in 11% of subjects. CEEG was initiated outside routine work hours in 47% of subjects. CEEG duration was <12 h in 16%, 12-24 h in 34%, and >24 h in 48%. Substantial variability existed among sites in CEEG indications and neurologic diagnoses, yet within each acute neurologic diagnosis category a similar proportion of subjects at each site had electrographic seizures. Electrographic seizure characteristics including distribution and duration varied across sites and neurologic diagnoses., Significance: These data provide a systematic assessment of recent CEEG use in critically ill children and indicate variability in practice. The results suggest that multicenter studies are feasible if CEEG monitoring pathways can be standardized. However, the data also indicate that electrographic seizure variability must be considered when designing studies that address the impact of electrographic seizures on outcome., (Wiley Periodicals, Inc. © 2013 International League Against Epilepsy.)

Objectives: We aimed to determine the incidence of electrographic seizures in children in the pediatric intensive care unit who underwent EEG monitoring, risk factors for electrographic seizures, and whether electrographic seizures were associated with increased odds of mortality., Methods: Eleven sites in North America retrospectively reviewed a total of 550 consecutive children in pediatric intensive care units who underwent EEG monitoring. We collected data on demographics, diagnoses, clinical seizures, mental status at EEG onset, EEG background, interictal epileptiform discharges, electrographic seizures, intensive care unit length of stay, and in-hospital mortality., Results: Electrographic seizures occurred in 162 of 550 subjects (30%), of which 61 subjects (38%) had electrographic status epilepticus. Electrographic seizures were exclusively subclinical in 59 of 162 subjects (36%). A multivariable logistic regression model showed that independent risk factors for electrographic seizures included younger age, clinical seizures prior to EEG monitoring, an abnormal initial EEG background, interictal epileptiform discharges, and a diagnosis of epilepsy. Subjects with electrographic status epilepticus had greater odds of in-hospital death, even after adjusting for EEG background and neurologic diagnosis category., Conclusions: Electrographic seizures are common among children in the pediatric intensive care unit, particularly those with specific risk factors. Electrographic status epilepticus occurs in more than one-third of children with electrographic seizures and is associated with higher in-hospital mortality.

Murine 3T3-L1 adipocytes are widely used as a cellular model of obesity. However, whereas transfection of 3T3-L1 preadipocytes is straightforward, ectopic gene expression in mature 3T3-L1 adipocytes has proved challenging. Here, we used the pSLIK vector system to generate stable doxycycline-inducible expression of the liver-enriched inhibitor protein isoform of CCAAT/enhancer binding protein β (C/ebpβ (Cebpb)) (C/EBPβ-LIP) in fully differentiated 3T3-L1 adipocytes. Because overexpression of C/ebpβ-LIP impairs adipocyte differentiation, the C/ebpβ-LIP construct was first integrated in 3T3-L1 preadipocytes but expression was induced only when adipocytes were fully differentiated. Increased C/EBPβ-LIP in mature adipocytes down-regulated C/ebpβ target genes including 11β-hydroxysteroid dehydrogenase type 1, phosphoenolpyruvate carboxykinase and fatty acid binding protein 4 but had no effect on asparagine synthetase, demonstrating that transcriptional down-regulation by C/ebpβ-LIP in 3T3-L1 adipocytes is not a general effect. Importantly, these genes were modulated in a similar manner in adipose tissue of mice with genetically increased C/ebpβ-LIP levels. The use of the pSLIK system to conditionally express transgenes in 3T3-L1 cells could be a valuable tool to dissect adipocyte physiology.

Introduction: Nonconvulsive seizures and nonconvulsive status epilepticus commonly occur in patients with aneurysmal subarachnoid hemorrhages. When continuous EEG is used in patients in the neuro-intensive care unit, rhythmical and periodic patterns of uncertain significance are frequently encountered. It is unknown how these findings impact patient outcome., Methods: Patients were enrolled from a single tertiary care center with subarachnoid hemorrhages secondary to ruptured intracranial aneurysm, and either a witnessed seizure or significantly impaired mental status. Prospective clinical, laboratory, imaging, and short-term outcome data were collected. Continuous EEG monitoring was performed and scored according to American Clinical Neurophysiology Society (ACNS) Subcommittee on Research Terminology for Continuous EEG Monitoring., Results: Sixty-eight patients were enrolled. Fifty-four had a poor-grade subarachnoid hemorrhage upon admission. Fifty-one patients had rhythmical or periodic patterns: 33 with periodic discharges and 38 with rhythmic delta activity. Four patients had unequivocal electrographic seizures. Patients did poorly in the short term: 14 died and 42 were severely disabled at discharge. In hospital, mortality was 19.6% in patients with rhythmical or periodic patterns and 23.5% in patients without. Age, female gender, and endovascular treatment had a positive correlation with the occurrence of periodic discharges. However, there was no correlation between rhythmical and periodic patterns and outcome., Discussion: Using the ACNS Research Terminology, it is shown that rhythmical and periodic patterns are very common in critically ill patients with subarachnoid hemorrhage. However, the presence and the abundance of these patterns did not predict short-term outcome in this prospective, single-center observational study. We were unable to show that rhythmical and periodic EEG patterns are an independent predictor for outcome relative to other clinical features. Large multicenter studies will be required to determine if these patterns independently predict outcome and to demonstrate the impact of treatment interventions that are directed at rhythmical and periodic continuous EEG patterns.

Atherosclerosis is a chronic inflammatory disease in which initial vascular damage leads to extensive macrophage and lymphocyte infiltration. Although acutely glucocorticoids suppress inflammation, chronic glucocorticoid excess worsens atherosclerosis, possibly by exacerbating systemic cardiovascular risk factors. However, glucocorticoid action within the lesion may reduce neointimal proliferation and inflammation. Glucocorticoid levels within cells do not necessarily reflect circulating levels due to pre-receptor metabolism by 11β-hydroxysteroid dehydrogenases (11β-HSDs). 11β-HSD2 converts active glucocorticoids into inert 11-keto forms. 11β-HSD1 catalyses the reverse reaction, regenerating active glucocorticoids. 11β-HSD2-deficiency/inhibition causes hypertension, whereas deficiency/inhibition of 11β-HSD1 generates a cardioprotective lipid profile and improves glycemic control. Importantly, 11β-HSD1-deficiency/inhibition is atheroprotective, whereas 11β-HSD2-deficiency accelerates atherosclerosis. These effects are largely independent of systemic risk factors, reflecting modulation of glucocorticoid action and inflammation within the vasculature. Here, we consider whether evidence linking the 11β-HSDs to vascular inflammation suggests these isozymes are potential therapeutic targets in vascular injury and atherosclerosis.

Prolonged video-EEG (vEEG) monitoring helps characterize paroxysmal events and epilepsy. There is limited literature in pediatrics describing the safety and utility of vEEG. We retrospectively reviewed 454 pediatric epilepsy monitoring unit admissions over two years. Final event diagnoses, duration of seizures, and medical complications were analyzed. Two hundred twenty admissions (48.4%) captured epileptic seizures, 150 (33.0%) captured nonepileptic events, and 84 (18.5%) failed to capture any events. Medical complications were seen in 4 patients (1.8%) with no long-term complications. Seventeen episodes of status epilepticus occurred in 13 patients. This constituted 2.9% of all admissions and 5.9% of admissions with epileptic seizures. The median duration of status was 26 min, and three patients required transfer to the pediatric intensive care unit. Video-EEG monitoring had a high yield in capturing events and differentiating epileptic from nonepileptic events. Our pediatric patients experienced greater risk of status epilepticus but lesser risk of injury., (Copyright © 2013 Elsevier Inc. All rights reserved.)

An imbalance between central glucocorticoid (GR) and mineralocorticoid (MR) receptors is proposed to underlie the HPA axis dysregulation that associates with susceptibility to psychopathology (anxiety, PTSD). To test this 'balance hypothesis' we examined whether the impact of MR levels upon HPA-axis control and behaviour depended on the relative levels of GR and vice versa. Avoiding antenatal maternal 'programming' effects by using littermates, we generated mice with forebrain MR over-expression (MR(hi)) and/or simultaneous global GR under-expression (GR(lo)). We found a significant interaction between MR and GR in control of the HPA-axis under stressed but not basal conditions. With reduced GR levels, HPA-axis activity in response to restraint stress was enhanced, likely due to impaired negative feedback. However, high MR in concert with reduced GR minimised this HPA-axis overshoot in response to stress. MR:GR balance also played a role in determining strategies of spatial memory during a watermaze probe trial: when coupled with GR under-expression, MR(hi) show enhanced perseveration, suggesting enhanced spatial recall or reduced exploratory flexibility. Other alterations in cognitive functions were specific to a single receptor without interaction, with both MR(hi) and GR(lo) manipulations independently impairing reversal learning in spatial and fear memory tasks. Thus, MR and GR interact in specific domains of neuroendocrine and cognitive control, but for other limbic-associated behaviours each receptor mediates its own repertoire of responses. Since modulation of HPA-axis and behavioural dysfunction associated with high levels of MR, selective ligands or transcriptional regulators may afford novel therapeutic approaches to affective psychopathologies., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Purpose: To describe current continuous EEG monitoring (cEEG) utilization in critically ill children., Methods: An online survey of pediatric neurologists from 50 US and 11 Canadian institutions was conducted in August 2011., Results: Responses were received from 58 of 61 (95%) surveyed institutions. Common cEEG indications are altered mental status after a seizure or status epilepticus (97%), altered mental status of unknown etiology (88%), or altered mental status with an acute primary neurologic condition (88%). The median number of patients undergoing cEEG per month per center increased from August 2010 to August 2011 (6 to 10 per month in the United States; 2 to 3 per month in Canada). Few institutions have clinical pathways addressing cEEG use (31%). Physicians most commonly review cEEG twice per day (37%). There is variability regarding which services can order cEEG, the degree of neurology involvement, technologist availability, and whether technologists perform cEEG screening., Conclusions: Among the surveyed institutions, which included primarily large academic centers, cEEG use in pediatric intensive care units is increasing and is often considered indicated for children with altered mental status at risk for nonconvulsive seizures. However, there remains substantial variability in cEEG access and utilization among institutions.

11β-Hydroxysteroid dehydrogenase type-1 (11β-HSD1) converts inert cortisone into active cortisol, amplifying intracellular glucocorticoid action. 11β-HSD1 deficiency improves cardiovascular risk factors in obesity but exacerbates acute inflammation. To determine the effects of 11β-HSD1 deficiency on atherosclerosis and its inflammation, atherosclerosis-prone apolipoprotein E-knockout (ApoE-KO) mice were treated with a selective 11β-HSD1 inhibitor or crossed with 11β-HSD1-KO mice to generate double knockouts (DKOs) and challenged with an atherogenic Western diet. 11β-HSD1 inhibition or deficiency attenuated atherosclerosis (74-76%) without deleterious effects on plaque structure. This occurred without affecting plasma lipids or glucose, suggesting independence from classical metabolic risk factors. KO plaques were not more inflamed and indeed had 36% less T-cell infiltration, associated with 38% reduced circulating monocyte chemoattractant protein-1 (MCP-1) and 36% lower lesional vascular cell adhesion molecule-1 (VCAM-1). Bone marrow (BM) cells are key to the atheroprotection, since transplantation of DKO BM to irradiated ApoE-KO mice reduced atherosclerosis by 51%. 11β-HSD1-null macrophages show 76% enhanced cholesterol ester export. Thus, 11β-HSD1 deficiency reduces atherosclerosis without exaggerated lesional inflammation independent of metabolic risk factors. Selective 11β-HSD1 inhibitors promise novel antiatherosclerosis effects over and above their benefits for metabolic risk factors via effects on BM cells, plausibly macrophages.

Purpose: The terms "electrical status epilepticus during sleep (ESES)" and "continuous spikes and waves during sleep (CSWS)" have been used interchangeably when referring to related but different concepts. In addition, the quantification of epileptiform activity has not been standardized, and different approaches to quantification have been used. The aim of this study was to evaluate the extent to which pediatric neurologists and epileptologists use a homogeneous terminology and conceptualization in CSWS and ESES and to characterize the current understanding of these conditions., Methods: A survey addressing the use of terminology in "ESES" and "CSWS" and the understanding of related concepts was distributed online to all members of the Child Neurology Society and the American Epilepsy Society mailing lists. Surveys were self-administered and collected using an online survey website (http://www.surveymonkey.com)., Key Findings:   Two hundred nineteen surveys were completed, 137 from the Child Neurology Society mailing list and 82 from the American Epilepsy Society mailing list. ESES and CSWS were considered synonymous by 117 respondents, not synonymous by 61, 21 respondents did not know, and 20 did not respond. Most respondents (63.1%) considered CSWS as a devastating epileptic encephalopathy with severe sequelae even if treated correctly, but 25.1% of respondents indicated that it does not leave sequelae if epilepsy was treated early and another 11.8% noted that cognitive difficulties resolved with age. Cognitive and/or language regression were considered mandatory for the diagnosis of CSWS by only 27% of the respondents. The diagnosis of CSWS was based on electroencephalography (EEG) assessment alone by 31% of respondents. Respondents used different methods for calculation of the epileptiform activity, different EEG samples for calculation, and considered differently the lateralized epileptiform activity. The cut-off values for percentage of the sleep record occupied by spike-waves were variable depending on the respondent. There was no agreement on whether these cutoff values were mandatory for the diagnosis of ESES and CSWS., Significance:   Our data show that the professionals caring for children with ESES and CSWS in North America use the terms, concepts, and defining features heterogeneously. The lack of a common language may complicate communication among clinicians and jeopardize research in this field. We anticipate that our data will fuel the development of much needed common terminology and conceptualization of ESES and CSWS., Competing Interests: OF CONFLICT OF INTERESTS None of the authors has any conflict of interest to disclose. We confirm that we have read the Journal’s position on issues involved in ethical publication and affirm that this report is consistent with those guidelines., (Wiley Periodicals, Inc. © 2012 International League Against Epilepsy.)

Continuous electroencephalographic (CEEG) monitoring is used with increasing frequency in critically ill children to provide insight into brain function and to identify electrographic seizures. CEEG monitoring use often impacts clinical management, most often by identifying electrographic seizures and status epilepticus. Most electrographic seizures have no clinical correlate, and thus would not be identified without CEEG monitoring. There are increasing data showing that electrographic seizures and electrographic status epilepticus are associated with worse outcome. Seizure identification efficiency may be improved by further development of quantitative electroencephalography trends. This review describes the clinical impact of CEEG data, the epidemiology of electrographic seizures and status epilepticus, the impact of electrographic seizures on outcome, the utility of quantitative electroencephalographic trends for seizure identification, and practical considerations regarding CEEG monitoring.

The 7-valent pneumococcal conjugate vaccine (PCV7) has been included in the routine childhood immunization programme in the UK since September 2006. A population-based study of serotypes causing invasive pneumococcal disease (IPD) post-PCV7 in North East England was conducted using data from a regional enhanced IPD surveillance system. Overall, there was a 20% reduction [95% confidence interval (CI) 5-32] from 12·1 cases/100 000 population in 2006/2007 to 9·7 in 2009/2010. There was a fall in IPD caused by PCV7 serotypes in all age groups, with reductions of 90% (95% CI 61-99) in children aged <5 years, 50% (95% CI 4-75) in persons aged 5-64 years and 66% (95% CI 40-82) in adults aged ⩾65 years. There was a non-significant increase in IPD caused by non-PCV7 serotypes in children aged <5 years of 88% (95% CI -10 to 312) and adults aged ⩾65 years of 12% (95% CI -19 to 50), which was largely caused by serotypes 7F, 19A and 22F. Replacement disease appears to have reduced the benefits of PCV7 in North East England.

For a number of years, there has been a widespread view that the adverse side-effects of prolonged glucocorticoid (GC) treatment are a result of glucocorticoid receptor (GR)-mediated gene activation, whilst the beneficial anti-inflammatory effects result from GR-mediated 'transrepression'. Since the introduction of the dimerisation-deficient GR mutant, GR(dim), was apparently unable to activate gene transcription, yet still able to repress pro-inflammatory gene transcription, the search for novel GR modulators has centred on the separation of gene activation from repression by prevention of GR dimerisation. However, recent work has questioned the conclusions drawn from these early GR(dim) studies, with evidence that GR(dim) mutants not only activate gene transcription, but that, in direct contradiction to the initial GR(dim) work, are also capable of forming dimers. This review of the current literature highlights the versatility of the GR in forming homodimer interactions, as well as the ability to bind to alternate nuclear receptors, and investigates the potential implications such varying GR dimer conformations may have for the design of GR ligands with a safer side effect profile., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Continuous spikes and waves during sleep (CSWS) is an epileptic encephalopathy characterized in most patients by (1) difficult to control seizures, (2) interictal epileptiform activity that becomes prominent during sleep leading to an electroencephalogram (EEG) pattern of electrical status epilepticus in sleep (ESES), and (3) neurocognitive regression. In this paper, we will summarize current epidemiological, clinical, and EEG knowledge on CSWS and will provide suggestions for treatment. CSWS typically presents with seizures around 2-4 years of age. Neurocognitive regression occurs around 5-6 years of age, and it is accompanied by subacute worsening of EEG abnormalities and seizures. At approximately 6-9 years of age, there is a gradual resolution of seizures and EEG abnormalities, but the neurocognitive deficits persist in most patients. The cause of CSWS is unknown, but early developmental lesions play a major role in approximately half of the patients, and genetic associations have recently been described. High-dose benzodiazepines and corticosteroids have been successfully used to treat clinical and electroencephalographic features. Corticosteroids are often reserved for refractory disease because of adverse events. Valproate, ethosuximide, levetiracetam, sulthiame, and lamotrigine have been also used with some success. Epilepsy surgery may be considered in a few selected patients.