Exploring the relevance of NUP93 variants in steroid-resistant nephrotic syndrome using next generation sequencing and a fly kidney model.

BACKGROUND: Variants in genes encoding nuclear pore complex (NPC) proteins are a newly identified cause of paediatric steroid-resistant nephrotic syndrome (SRNS). Recent reports describing NUP93 variants suggest these could be a significant cause of paediatric onset SRNS. We report NUP93 cases in the UK and demonstrate in vivo functional effects of Nup93 depletion in a fly (Drosophila melanogaster) nephrocyte model. METHODS: Three hundred thirty-seven paediatric SRNS patients from the National cohort of patients with Nephrotic Syndrome (NephroS) were whole exome and/or whole genome sequenced. Patients were screened for over 70 genes known to be associated with Nephrotic Syndrome (NS). D. melanogaster Nup93 knockdown was achieved by RNA interference using nephrocyte-restricted drivers. RESULTS: Six novel homozygous and compound heterozygous NUP93 variants were detected in 3 sporadic and 2 familial paediatric onset SRNS characterised histologically by focal segmental glomerulosclerosis (FSGS) and progressing to kidney failure by 12 months from clinical diagnosis. Silencing of the two orthologs of human NUP93 expressed in D. melanogaster, Nup93-1, and Nup93-2 resulted in significant signal reduction of up to 82% in adult pericardial nephrocytes with concomitant disruption of NPC protein expression. Additionally, nephrocyte morphology was highly abnormal in Nup93-1 and Nup93-2 silenced flies surviving to adulthood. CONCLUSION: We expand the spectrum of NUP93 variants detected in paediatric onset SRNS and demonstrate its incidence within a national cohort. Silencing of either D. melanogaster Nup93 ortholog caused a severe nephrocyte phenotype, signaling an important role for the nucleoporin complex in podocyte biology. A higher resolution version of the Graphical abstract is available as Supplementary information.

In search for the most optimal EEG method: A practical evaluation of a water-based electrode EEG system.

The study assessed a mobile electroencephalography system with water-based electrodes for its applicability in cognitive and behavioural neuroscience. It was compared to a standard gel-based wired system. Electroencephalography was recorded on two occasions (first with gel-based, then water-based system) as participants completed the flanker task. Technical and practical considerations for the application of the water-based system are reported based on participant and experimenter experiences. Empirical comparisons focused on electroencephalography data noise levels, frequency power across four bands (theta, alpha, low beta and high beta) and event-related components (P300 and ERN). The water-based system registered more noise compared to the gel-based system which resulted in increased loss of data during artefact rejection. Signal-to-noise ratio was significantly lower for the water-based system in the parietal channels which affected the observed parietal beta power. It also led to a shift in topography of the maximal P300 activity from parietal to frontal regions. The water-based system may be prone to slow drift noise which may affect the reliability and consistency of low-frequency band analyses. Practical considerations for the use of water-based electrode electroencephalography systems are provided.

Affective responses to coherence in high and low risk scenarios.

Presenting information in a coherent fashion has been shown to increase processing fluency, which in turn influences affective responses. The pattern of responses have been explained by two apparently competing accounts: hedonic marking (response to fluency is positive) and fluency amplification (response to fluency can be positive or negative, depending on stimuli valence). This paper proposes that these accounts are not competing explanations, but separate mechanisms, serving different purposes. Therefore, their individual contributions to overall affective responses should be observable. In three experiments, participants were presented with businesses scenarios, with riskiness (valence) and coherence (fluency) manipulated, and affective responses recorded. Results suggested that increasing the fluency of stimuli increases positive affect. If the stimulus is negative, then increasing fluency simultaneously increases negative affect. These affective responses appeared to cancel each other out (Experiment 1) when measured using self-report bipolar scales. However, separate measurement of positive and negative affect, either using unipolar scales (Experiment 2) or using facial electromyography (Experiment 3), provided evidence for co-occurring positive and negative affective responses, and therefore the co-existence of hedonic marking and fluency amplification mechanisms.

Long-read nanopore sequencing resolves a TMEM231 gene conversion event causing Meckel-Gruber syndrome.

The diagnostic deployment of massively parallel short-read next-generation sequencing (NGS) has greatly improved genetic test availability, speed, and diagnostic yield, particularly for rare inherited disorders. Nonetheless, diagnostic approaches based on short-read sequencing have a poor ability to accurately detect gene conversion events. We report on the genetic analysis of a family in which 3 fetuses had clinical features consistent with the autosomal recessive disorder Meckel-Gruber syndrome (MKS). Targeted NGS of 29 known MKS-associated genes revealed a heterozygous TMEM231 splice donor variant c.929+1A>G. Comparative read-depth analysis, performed to identify a second pathogenic allele, revealed an apparent heterozygous deletion of TMEM231 exon 4. To verify this result we performed single-molecule long-read sequencing of a long-range polymerase chain reaction product spanning this locus. We identified four missense variants that were absent from the short-read dataset due to the preferential mapping of variant-containing reads to a downstream TMEM231 pseudogene. Consistent with the parental segregation analysis, we demonstrate that the single-molecule long reads could be used to show that the variants are arranged in trans. Our experience shows that robust validation of apparent dosage variants remains essential to avoid the pitfalls of short-read sequencing and that new third-generation long-read sequencing technologies can already aid routine clinical care.

Live Intravital Imaging of Cellular Trafficking in the Cardiac Microvasculature-Beating the Odds.

Although mortality rates from cardiovascular disease in the developed world are falling, the prevalence of cardiovascular disease (CVD) is not. Each year, the number of people either being diagnosed as suffering with CVD or undergoing a surgical procedure related to it, such as percutaneous coronary intervention, continues to increase. In order to ensure that we can effectively manage these diseases in the future, it is critical that we fully understand their basic physiology and their underlying causative factors. Over recent years, the important role of the cardiac microcirculation in both acute and chronic disorders of the heart has become clear. The recruitment of inflammatory cells into the cardiac microcirculation and their subsequent activation may contribute significantly to tissue damage, adverse remodeling, and poor outcomes during recovery. However, our basic understanding of the cardiac microcirculation is hampered by an historic inability to image the microvessels of the beating heart-something we have been able to achieve in other organs for over 100 years. This stems from a couple of clear and obvious difficulties related to imaging the heart-firstly, it has significant inherent contractile motion and is affected considerably by the movement of lungs. Secondly, it is located in an anatomically challenging position for microscopy. However, recent microscopic and technological developments have allowed us to overcome some of these challenges and to begin to answer some of the basic outstanding questions in cardiac microvascular physiology, particularly in relation to inflammatory cell recruitment. In this review, we will discuss some of the historic work that took place in the latter part of last century toward cardiac intravital, before moving onto the advanced work that has been performed since. This work, which has utilized technology such as spinning-disk confocal and multiphoton microscopy, has-along with some significant advancements in algorithms and software-unlocked our ability to image the "business end" of the cardiac vascular tree. This review will provide an overview of these techniques, as well as some practical pointers toward software and other tools that may be useful for other researchers who are considering utilizing this technique themselves.

Tify: A quality-based frame selection tool for improving the output of unstable biomedical imaging.

The ability to image biological tissues is critical to our understanding of a range of systems and processes. In the case of in situ living tissue, such imaging is hampered by the innate mechanical properties of the tissue. In many cases, this provides challenges in how to process large amounts of image data which may contain aberrations from movement. Generally, current tools require the provision of reference images and are unable to maintain temporal correlations within an image set. Here, we describe a tool-Tify-which can accurately predict a numerical quality score versus human scoring and can analyse image sets in a manner that allows the maintenance of temporal relationships. The tool uses regression-based techniques to link image statistics to image quality based on user provided scores from a sample of images. Scores calculated by the software correlate strongly with the scores provided by human users. We identified that, in most cases, the software requires users to score between 20-30 frames in order to be able to accurately calculate the remaining images. Importantly, our results suggest that the software can use coefficients generated from consolidated image sets to process images without the need for additional manual scoring. Finally, the tool is able to use a frame windowing technique to identify the highest quality frame from a moving window, thus retaining macro-chronological connections between frames. In summary, Tify is able to successfully predict the quality of images in an image set based on a small number of sample scores provided by end-users. This software has the potential to improve the effectiveness of biological imaging techniques where motion artefacts, even in the presence of stabilisation, pose a significant problem.

Therapy influences goal attainment following botulinum neurotoxin injection for focal spasticity in adults with neurological conditions.

OBJECTIVE: To determine whether therapy influenced goal attainment following botulinum toxin (BoNT-A) injection for focal spasticity in adults with neurological conditions. METHODS: A prospective observational cohort study conducted in a large metropolitan spasticity clinic on adults with focal spasticity of any origin. Participants were provided with a therapy programme, designed to maximise therapeutic outcome. The primary outcome measure was Goal Attainment Scaling. To measure adherence, participants completed a therapy-recording tool each day. Goal attainment, and the rate of adherence to the therapy programme, was evaluated after 10 weeks. RESULTS: Active indications for BoNT-A treatment made up the majority of the goals (80.30%). Goals were achieved in 43/76 cases (56.60%; 95% CI = 42.40 to 69.80%). Therapy adherence was associated with significantly greater goal attainment (OR = 1.02, p = 0.03, 95% CI = 1.00 to 1.04). Greater adherence to therapy increased the odds of goal achievement for active indications but not for passive indications, suggesting a possible statistical interaction between the indication for injection and adherence to therapy (p < 0.01). CONCLUSION: Therapy adherence was associated with greater goal attainment. Active indications for BoNT-A were more reliant on adherence to prescribed therapy programmes than passive indications, although further investigation is required.

Clinical genetic testing using a custom-designed steroid-resistant nephrotic syndrome gene panel: analysis and recommendations.

BACKGROUND: There are many single-gene causes of steroid-resistant nephrotic syndrome (SRNS) and the list continues to grow rapidly. Prompt comprehensive diagnostic testing is key to realising the clinical benefits of a genetic diagnosis. This report describes a bespoke-designed, targeted next-generation sequencing (NGS) diagnostic gene panel assay to detect variants in 37 genes including the ability to identify copy number variants (CNVs). METHODS: This study reports results of 302 patients referred for SRNS diagnostic gene panel analysis. Phenotype and clinical impact data were collected using a standard proforma. Candidate variants detected by NGS were confirmed by Sanger sequencing/Multiplex Ligation-dependent Probe Amplification with subsequent family segregation analysis where possible. RESULTS: Clinical presentation was nephrotic syndrome in 267 patients and suspected Alport syndrome (AS) in 35. NGS panel testing determined a likely genetic cause of disease in 44/220 (20.0%) paediatric and 10/47 (21.3%) adult nephrotic cases, and 17/35 (48.6%) of haematuria/AS patients. Of 71 patients with genetic disease, 32 had novel pathogenic variants without a previous disease association including two with deletions of one or more exons of NPHS1 or NPHS2. CONCLUSION: Gene panel testing provides a genetic diagnosis in a significant number of patients presenting with SRNS or suspected AS. It should be undertaken at an early stage of the care pathway and include the ability to detect CNVs as an emerging mechanism for genes associated with this condition. Use of clinical genetic testing after diagnosis of SRNS has the potential to stratify patients and assist decision-making regarding management.

MAGI2 Mutations Cause Congenital Nephrotic Syndrome.

Steroid-resistant nephrotic syndrome (SRNS), a heterogeneous disorder of the renal glomerular filtration barrier, results in impairment of glomerular permselectivity. Inheritance of genetic SRNS may be autosomal dominant or recessive, with a subset of autosomal recessive SRNS presenting as congenital nephrotic syndrome (CNS). Mutations in 53 genes are associated with human SRNS, but these mutations explain ≤30% of patients with hereditary cases and only 20% of patients with sporadic cases. The proteins encoded by these genes are expressed in podocytes, and malfunction of these proteins leads to a universal end point of podocyte injury, glomerular filtration barrier disruption, and SRNS. Here, we identified novel disease-causing mutations in membrane-associated guanylate kinase, WW, and PDZ domain-containing 2 (MAGI2) through whole-exome sequencing of a deeply phenotyped cohort of patients with congenital, childhood-onset SRNS. Although MAGI2 has been shown to interact with nephrin and regulate podocyte cytoskeleton and slit diaphragm dynamics, MAGI2 mutations have not been described in human SRNS. We detected two unique frameshift mutations and one duplication in three patients (two families); two siblings shared the same homozygous frameshift mutation, whereas one individual with sporadic SRNS exhibited compound heterozygosity. Two mutations were predicted to introduce premature stop codons, and one was predicted to result in read through of the normal translational termination codon. Immunohistochemistry in kidney sections from these patients revealed that mutations resulted in lack of or diminished podocyte MAGI2 expression. Our data support the finding that mutations in the MAGI2 gene are causal for congenital SRNS.

Potential for use of creatine supplementation following mild traumatic brain injury.

There is significant overlap between the neuropathology of mild traumatic brain injury (mTBI) and the cellular role of creatine, as well as evidence of neural creatine alterations after mTBI. Creatine supplementation has not been researched in mTBI, but shows some potential as a neuroprotective when administered prior to or after TBI. Consistent with creatine's cellular role, supplementation reduced neuronal damage, protected against the effects of cellular energy crisis and improved cognitive and somatic symptoms. A variety of factors influencing the efficacy of creatine supplementation are highlighted, as well as avenues for future research into the potential of supplementation as an intervention for mTBI. In particular, the slow neural uptake of creatine may mean that greater effects are achieved by pre-emptive supplementation in at-risk groups.

A Metal-Insulator Transition of the Buried MnO2 Monolayer in Complex Oxide Heterostructure.

A novel artificially created MnO2 monolayer system is demonstrated in atomically controlled epitaxial perovskite heterostructures. With careful design of different electrostatic boundary conditions, a magnetic transition as well as a metal-insulator transition of the MnO2 monolayer is unveiled, providing a fundamental understanding of dimensionality-confined strongly correlated electron systems and a direction to design new electronic devices.

Long-term structural changes after mTBI and their relation to post-concussion symptoms.

PRIMARY OBJECTIVE: To investigate sustained structural changes in the long-term (>1 year) after mild traumatic brain injury (mTBI) and their relationship to ongoing post-concussion syndrome (PCS). RESEARCH DESIGN: Morphological and structural connectivity magnetic resonance imaging (MRI) data were acquired from 16 participants with mTBI and nine participants without previous head injury. MAIN OUTCOMES AND RESULTS: Participants with mTBI had less prefrontal grey matter and lower fractional anisotropy (FA) in the anterior corona radiata and internal capsule. Furthermore, PCS severity was associated with less parietal lobe grey matter and lower FA in the corpus callosum. CONCLUSIONS: There is evidence for both white and grey matter damage in participants with mTBI over 1 year after injury. Furthermore, these structural changes are greater in those that report more PCS symptoms, suggesting a neurophysiological basis for these persistent symptoms.

Multimodal imaging of mild traumatic brain injury and persistent postconcussion syndrome.

BACKGROUND: Persistent postconcussion syndrome (PCS) occurs in around 5-10% of individuals after mild traumatic brain injury (mTBI), but research into the underlying biology of these ongoing symptoms is limited and inconsistent. One reason for this could be the heterogeneity inherent to mTBI, with individualized injury mechanisms and psychological factors. A multimodal imaging study may be able to characterize the injury better. AIM: To look at the relationship between functional (fMRI), structural (diffusion tensor imaging), and metabolic (magnetic resonance spectroscopy) data in the same participants in the long term (>1 year) after injury. It was hypothesized that only those mTBI participants with persistent PCS would show functional changes, and that these changes would be related to reduced structural integrity and altered metabolite concentrations. METHODS: Functional changes associated with persistent PCS after mTBI (>1 year postinjury) were investigated in participants with and without PCS (both n = 8) and non-head injured participants (n = 9) during performance of working memory and attention/processing speed tasks. Correlation analyses were performed to look at the relationship between the functional data and structural and metabolic alterations in the same participants. RESULTS: There were no behavioral differences between the groups, but participants with greater PCS symptoms exhibited greater activation in attention-related areas (anterior cingulate), along with reduced activation in temporal, default mode network, and working memory areas (left prefrontal) as cognitive load was increased from the easiest to the most difficult task. Functional changes in these areas correlated with reduced structural integrity in corpus callosum and anterior white matter, and reduced creatine concentration in right dorsolateral prefrontal cortex. CONCLUSION: These data suggest that the top-down attentional regulation and deactivation of task-irrelevant areas may be compensating for the reduction in working memory capacity and variation in white matter transmission caused by the structural and metabolic changes after injury. This may in turn be contributing to secondary PCS symptoms such as fatigue and headache. Further research is required using multimodal data to investigate the mechanisms of injury after mTBI, but also to aid individualized diagnosis and prognosis.

Comparing methods for determining motor-hand lateralization based on fTCD signals.

The lateralization index (LI) as determined from functional transcranial Doppler sonography (fTCD) can be used to determine the hemispheric organization of neural activation during a behavioral task. Previous studies have proposed different methods to determine this index, but to our knowledge no studies have compared the performance of these methods. In this study, we compare two established methods with a simpler method proposed here. The aim was to see whether similar results could be achieved with a simpler method and to give an indication of the analysis steps required to determine the LI. A simple unimanual motor task was performed while fTCD was acquired, and the LI determined by each of these methods was compared. In addition, LI determined by each method was related to behavioural output in the form of degree of handedness. The results suggest that although the methods differed in complexity, they yielded similar results when determining the lateralization of motor functions, and its correlation with behavior. Further investigation is needed to expand the conclusions of this preliminary study, however the new method proposed in the paper has great potential as it is much simpler than the more established methods yet yields similar results.

Alcohol use and substance use disorders in Gulf War, Afghanistan, and Iraq War veterans compared with nondeployed military personnel.

Although recent veterans have been found to be at increased risk of psychiatric disorders, limited research has focused on alcohol or substance use disorders. This systematic review and meta-analysis examined whether alcohol or substance use disorders were more common in Gulf War, Afghanistan, and Iraq War veterans compared with military comparison groups nondeployed to the corresponding conflict, including never deployed personnel. Literature was searched (1990-2014) in multiple electronic databases. Studies were assessed for eligibility and quality, including risk of bias. Eighteen studies (1997-2014) met inclusion criteria. Pooled analysis based on a random-effects model yielded a summary odds ratio of 1.33 (95% confidence interval (CI): 1.22, 1.46) for alcohol (7 studies) and 2.13 (95% CI: 0.96, 4.72) for substance use (3 studies) disorders among Gulf War veterans, as well as 1.36 (95% CI: 1.11, 1.66) for alcohol (7 studies) and 1.14 (95% CI: 1.04, 1.25) for substance use (4 studies) disorders among Iraq/Afghanistan veterans; meta-regressions found no statistically significant association between theater of war and alcohol use or substance use disorders. Our findings indicate that Gulf and Iraq/Afghanistan war veterans are at higher alcohol use disorder risk than nondeployed veterans, but further studies with increased power are needed to assess substance use disorder risk in Gulf War veteran populations.

CI Therapy is Beneficial to Patients with Chronic Low-Functioning Hemiparesis after Stroke.

CI therapy is effective in patients with relatively good levels of residual arm function but its applicability to patients with low-functioning hemiparesis is not entirely clear. In the present study, we examined the feasibility and efficacy of the CI therapy concept in patients with very limited upper arm function prior to treatment, and further tested how the length of daily shaping training and constraining the good arm affects treatment outcome. In a baseline-controlled design, 65 chronic patients were treated with 2 weeks of modified CI therapy. Patients were randomly allocated to four treatment groups receiving 90 or 180 min of daily shaping training applied with or without constraint, respectively. Outcome was measured through the Reliable Change Index, which was calculated for parameters of motor function, health, and psychological wellbeing. Follow-up data were collected at 6 and 12 months. Two analyses were conducted, a whole-group analysis across all 65 participants and a sub-group analysis contrasting the four treatment variants. The whole-group analysis showed a significant treatment effect, which was largely sustained after 1 year. The sub-group analysis revealed a mixed picture; while improvements against the baseline period were observed in all four subgroups, 180 min of daily shaping training coupled with the constraint yielded better outcome on the MAL but not the WMFT, while for 90 min of training the level of improvement was similar for those who wore the constraint and those who did not. Together these results suggest that, at least in those patients available for follow-up measures, modified CI therapy induces sustained improvements in motor function in patients with chronic low-functioning hemiparesis. The absence of clear differences between the four treatment variants points to a complex relationship between the length of daily shaping training and the constraint in this patient group, which is likely to be mediated by fatigue and/or compliance with the constraint.

Quantitative electroencephalography and behavioural correlates of daytime sleepiness in chronic stroke.

Sleepiness is common after stroke, but in contrast to its importance for rehabilitation, existing studies focus primarily on the acute state and often use subjective sleepiness measures only. We used quantitative electroencephalography (qEEG) to extract physiological sleepiness, as well as subjective reports, in response to motor-cognitive demand in stroke patients and controls. We hypothesised that (a) slowing of the EEG is chronically sustained after stroke; (b) increased power in lower frequencies and increased sleepiness are associated; and (c) sleepiness is modulated by motor-cognitive demand. QEEGs were recorded in 32 chronic stroke patients and 20 controls using a Karolinska Drowsiness Test protocol administered before and after a motor priming task. Subjective sleepiness was measured using the Karolinska Sleepiness Scale. The findings showed that power density was significantly increased in delta and theta frequency bands over both hemispheres in patients which were not associated with subjective sleepiness ratings. This effect was not observed in controls. The motor priming task induced differential hemispheric effects with greater increase in low-frequency bands and presumably compensatory increases in higher frequency bands. The results indicate sustained slowing in the qEEG in chronic stroke, but in contrast to healthy controls, these changes are not related to perceived sleepiness.

Pain-related musculoskeletal disorders, psychological comorbidity, and the relationship with physical and mental well-being in Gulf War veterans.

Occupational activities such as lifting loads, working in constrained spaces, and training increase the risk of pain-related musculoskeletal disorders (MSDs) in military veterans. Few studies have investigated MSD and psychological disorder in veterans, and previous studies had limitations. This cross-sectional study compared pain-related MSD and psychological comorbidity and well-being between 1381 male Australian 1990-1991 Gulf War veterans (veterans) and a military comparison group (n=1377, of whom 39.6% were serving and 32.7% had previously deployed). At a medical assessment, 2000-2002, reported doctor-diagnosed arthritis or rheumatism, back or neck problems, joint problems, and soft tissue disorders were rated by medical practitioners as nonmedical, unlikely, possible, or probable diagnoses. Only probable MSDs were analysed. Psychological disorders in the past 12 months were measured using the Composite International Diagnostic Interview. The Short-Form Health Survey (SF-12) assessed 4-week physical and mental well-being. Almost one-quarter of veterans (24.5%) and the comparison group (22.4%) reported an MSD. Having any or specific MSD was associated with depression and posttraumatic stress disorder (PTSD), but not alcohol disorders. Physical and mental well-being was poorer in those with an MSD compared to those without, in both study groups (eg, veterans with any MSD, difference in SF-12 physical component summary scale medians = -10.49: 95% confidence interval -12.40, -8.57), and in those with MSD and psychological comorbidity compared with MSD alone. Comorbidity of any MSD and psychological disorder was more common in veterans, but MSDs were associated with depression, PTSD, and poorer well-being in both groups. Psychological comorbidity needs consideration in MSD management. Longitudinal studies are needed to assess directionality and causality.