Seasonal variations of microplastic pollution in the German River Weser.

Rivers play a major role in the distribution of microplastics (MPs) in the environment, however, research on temporal variations in these highly dynamic systems is still in its infancy. To date, most studies dealing with the seasonality of MP contamination in rivers focus on bi-yearly analysis, while temporal fluctuations over the course of the year are rarely studied. To shed more light on seasonal variability of MP abundance and potential driving factors, we have thus sampled the water surface of one location in the Weser River in Germany monthly over one entire year. In our study, we targeted MP in the size range 10-5000 µm, using two different state-of-the-art sampling methods (manta net for large MP (l-MP; 500-5000 µm) and a filtration system for small MP (s-MP; 10-500 µm)) and analysis techniques (ATR-FTIR and FPA-µFTIR) for chemical identification. Our findings show a strong size-dependent positive correlation of the MP concentration with discharge rates (specifically direct runoff) and suspended particulate matter (SPM) for s-MPs, specifically in the size range 10-149 µm. L-MPs, however, show a different environmental behaviour and do not follow these patterns. With our study, we were able to deliver a much higher temporal resolution, covering a broader size range of MPs compared to most studies. Our findings point towards an interplay of two possible mechanisms: a) the riverbeds play an important role in large-scale MP and SPM release via resuspension during high discharge events, and b) precipitation-driven soil erosion and runoff from urban surfaces (e.g. rain sewers) introduce MP and SPM. Hence, our study serves as a basis for more detailed investigations of MP transport in and between ecosystems.

Understanding the role of the hematopoietic niche in Huntington's disease's phenotypic expression: in vivo evidence using a parabiosis model.

In a previous study, we have shown that parabiotic coupling of a knock-in mouse model (zQ175) of Huntington's disease (HD) to wild-type (WT) littermates resulted in a worsening of the normal phenotype as seen by detection of mutant huntingtin protein (mHTT) aggregates within peripheral organs and the cerebral cortex as well as vascular abnormalities in WT mice. In contrast, parabiosis improved disease features in the zQ175 mice such as reduction of mHTT aggregate number in the liver and cortex, decrease in blood-brain barrier (BBB) permeability and attenuation of mitochondrial impairments. While the shared circulation mediated these effects, no specific factor was identified. To better understand which blood elements were involved in the aforementioned changes, WT and zQ175 mice underwent parabiotic surgery prior to exposing one of the paired animals to irradiation. The irradiation procedure successfully eliminated the hematopoietic niche followed by repopulation with cells originating from the non-irradiated parabiont, as measured by the quantification of mHTT levels in peripheral blood mononuclear cells. Although irradiation of the WT parabiont, causing the loss of healthy hematopoietic cells, did lead to a few alterations in mitochondrial function in the muscle (TOM40 levels), and increased neuroinflammation in the striatum (GFAP levels), most of the changes observed were likely attributable to the irradiation procedure itself (e.g. mHTT aggregates in cortex and liver; cellular stress in peripheral organs). However, factors such as mHTT aggregation in the brain and periphery, and BBB leakage, which were improved in zQ175 mice when paired to WT littermates in the previous parabiosis experiment, were unaffected by perturbation of the hematopoietic niche. It would therefore appear that cells of the hematopoietic stem cell niche are largely uninvolved in the beneficial effects of parabiosis.

Design and Evaluation of [18F]CHDI-650 as a Positron Emission Tomography Ligand to Image Mutant Huntingtin Aggregates.

Therapeutic interventions are being developed for Huntington's disease (HD), a hallmark of which is mutant huntingtin protein (mHTT) aggregates. Following the advancement to human testing of two [11C]-PET ligands for aggregated mHTT, attributes for further optimization were identified. We replaced the pyridazinone ring of CHDI-180 with a pyrimidine ring and minimized off-target binding using brain homogenate derived from Alzheimer's disease patients. The major in vivo metabolic pathway via aldehyde oxidase was blocked with a 2-methyl group on the pyrimidine ring. A strategically placed ring-nitrogen on the benzoxazole core ensured high free fraction in the brain without introducing efflux. Replacing a methoxy pendant with a fluoro-ethoxy group and introducing deuterium atoms suppressed oxidative defluorination and accumulation of [18F]-signal in bones. The resulting PET ligand, CHDI-650, shows a rapid brain uptake and washout profile in non-human primates and is now being advanced to human testing.

Development of a ligand for in vivo imaging of mutant huntingtin in Huntington's disease.

Huntington's disease (HD) is a dominantly inherited neurodegenerative disorder caused by a CAG trinucleotide expansion in the huntingtin (HTT) gene that encodes the pathologic mutant HTT (mHTT) protein with an expanded polyglutamine (polyQ) tract. Whereas several therapeutic programs targeting mHTT expression have advanced to clinical evaluation, methods to visualize mHTT protein species in the living brain are lacking. Here, we demonstrate the development and characterization of a positron emission tomography (PET) imaging radioligand with high affinity and selectivity for mHTT aggregates. This small molecule radiolabeled with 11C ([11C]CHDI-180R) allowed noninvasive monitoring of mHTT pathology in the brain and could track region- and time-dependent suppression of mHTT in response to therapeutic interventions targeting mHTT expression in a rodent model. We further showed that in these animals, therapeutic agents that lowered mHTT in the striatum had a functional restorative effect that could be measured by preservation of striatal imaging markers, enabling a translational path to assess the functional effect of mHTT lowering.

Assessment of patient-reported outcomes after polytrauma - instruments and methods: a systematic review.

OBJECTIVES: We (1) collected instruments that assess health-related quality of life (HRQoL), activities of daily living (ADL) and social participation during follow-up after polytrauma, (2) described their use and (3) investigated other relevant patient-reported outcomes (PROs) assessed in the studies. DESIGN: Systematic Review using the Preferred Reporting Items for Systematic Review and Meta-Analysis guideline. DATA SOURCES: MEDLINE, Embase, CINAHL, PsycINFO, CENTRAL, as well as the trials registers ClinicalTrials.gov and WHO ICTRP were searched from January 2005 to April 2018. ELIGIBILITY CRITERIA: All original empirical research published in English or German including PROs of patients aged 18-75 years with an Injury Severity Score≥16 and/or an Abbreviated Injury Scale≥3. Studies with defined injuries or diseases (e.g. low-energy injuries) and some text types (e.g. grey literature and books) were excluded. Systematic reviews and meta-analyses were excluded, but references screened for appropriate studies. DATA EXTRACTION AND SYNTHESIS: Data extraction, narrative content analysis and a critical appraisal (e.g. UK National Institute for Health and Care Excellence) were performed by two reviewers independently. RESULTS: The search yielded 3496 hits; 54 publications were included. Predominantly, HRQoL was assessed, with Short Form-36 Health Survey applied most frequently. ADL and (social) participation were rarely assessed. The methods most used were postal surveys and single assessments of PROs, with a follow-up period of one to one and a half years. Other relevant PRO areas reported were function, mental disorders and pain. CONCLUSIONS: There is a large variation in the assessment of PROs after polytrauma, impairing comparability of outcomes. First efforts to standardise the collection of PROs have been initiated, but require further harmonisation between central players. Additional knowledge on rarely reported PRO areas (e.g. (social) participation, social networks) may lead to their consideration in health services provision. PROSPERO REGISTRATION NUMBER: CRD42017060825.

Pharmacological characterization of mutant huntingtin aggregate-directed PET imaging tracer candidates.

Huntington's disease (HD) is caused by a CAG trinucleotide repeat expansion in the first exon of the huntingtin (HTT) gene coding for the huntingtin (HTT) protein. The misfolding and consequential aggregation of CAG-expanded mutant HTT (mHTT) underpin HD pathology. Our interest in the life cycle of HTT led us to consider the development of high-affinity small-molecule binders of HTT oligomerized/amyloid-containing species that could serve as either cellular and in vivo imaging tools or potential therapeutic agents. We recently reported the development of PET tracers CHDI-180 and CHDI-626 as suitable for imaging mHTT aggregates, and here we present an in-depth pharmacological investigation of their binding characteristics. We have implemented an array of in vitro and ex vivo radiometric binding assays using recombinant HTT, brain homogenate-derived HTT aggregates, and brain sections from mouse HD models and humans post-mortem to investigate binding affinities and selectivity against other pathological proteins from indications such as Alzheimer's disease and spinocerebellar ataxia 1. Radioligand binding assays and autoradiography studies using brain homogenates and tissue sections from HD mouse models showed that CHDI-180 and CHDI-626 specifically bind mHTT aggregates that accumulate with age and disease progression. Finally, we characterized CHDI-180 and CHDI-626 regarding their off-target selectivity and binding affinity to beta amyloid plaques in brain sections and homogenates from Alzheimer's disease patients.

[11C]CHDI-626, a PET Tracer Candidate for Imaging Mutant Huntingtin Aggregates with Reduced Binding to AD Pathological Proteins.

The expanded polyglutamine-containing mutant huntingtin (mHTT) protein is implicated in neuronal degeneration of medium spiny neurons in Huntington's disease (HD) for which multiple therapeutic approaches are currently being evaluated to eliminate or reduce mHTT. Development of effective and orthogonal biomarkers will ensure accurate assessment of the safety and efficacy of pharmacologic interventions. We have identified and optimized a class of ligands that bind to oligomerized/aggregated mHTT, which is a hallmark in the HD postmortem brain. These ligands are potentially useful imaging biomarkers for HD therapeutic development in both preclinical and clinical settings. We describe here the optimization of the benzo[4,5]imidazo[1,2-a]pyrimidine series that show selective binding to mHTT aggregates over Aß- and/or tau-aggregates associated with Alzheimer's disease pathology. Compound [11C]-2 was selected as a clinical candidate based on its high free fraction in the brain, specific binding in the HD mouse model, and rapid brain uptake/washout in nonhuman primate positron emission tomography imaging studies.

Shedding a new light on Huntington's disease: how blood can both propagate and ameliorate disease pathology.

Huntington's disease (HD) is a monogenic neurodegenerative disorder resulting from a mutation in the huntingtin gene. This leads to the expression of the mutant huntingtin protein (mHTT) which provokes pathological changes in both the central nervous system (CNS) and periphery. Accumulating evidence suggests that mHTT can spread between cells of the CNS but here, we explored the possibility that mHTT could also propagate and cause pathology via the bloodstream. For this, we used a parabiosis approach to join the circulatory systems of wild-type (WT) and zQ175 mice. After surgery, we observed mHTT in the plasma and circulating blood cells of WT mice and post-mortem analyses revealed the presence of mHTT aggregates in several organs including the liver, kidney, muscle and brain. The presence of mHTT in the brain was accompanied by vascular abnormalities, such as a reduction of Collagen IV signal intensity and altered vessel diameter in the striatum, and changes in expression of Glutamic acid decarboxylase 65/67 (GAD65-67) in the cortex. Conversely, we measured reduced pathology in zQ175 mice by decreased mitochondrial impairments in peripheral organs, restored vessel diameter in the cortex and improved expression of Dopamine- and cAMP-regulated phosphoprotein 32 (DARPP32) in striatal neurons. Collectively, these results demonstrate that circulating mHTT can disseminate disease, but importantly, that healthy blood can dilute pathology. These findings have significant implications for the development of therapies in HD.

Imaging Mutant Huntingtin Aggregates: Development of a Potential PET Ligand.

Mutant huntingtin (mHTT) protein carrying the elongated N-terminal polyglutamine (polyQ) tract misfolds and forms protein aggregates characteristic of Huntington's disease (HD) pathology. A high-affinity ligand specific for mHTT aggregates could serve as a positron emission tomography (PET) imaging biomarker for HD therapeutic development and disease progression. To identify such compounds with binding affinity for polyQ aggregates, we embarked on systematic structural activity studies; lead optimization of aggregate-binding affinity, unbound fractions in brain, permeability, and low efflux culminated in the discovery of compound 1, which exhibited target engagement in autoradiography (ARG) studies in brain slices from HD mouse models and postmortem human HD samples. PET imaging studies with 11C-labeled 1 in both HD mice and WT nonhuman primates (NHPs) demonstrated that the right-hand-side labeled ligand [11C]-1R (CHDI-180R) is a suitable PET tracer for imaging of mHTT aggregates. [11C]-1R is now being advanced to human trials as a first-in-class HD PET radiotracer.

Expression of mutant exon 1 huntingtin fragments in human neural stem cells and neurons causes inclusion formation and mitochondrial dysfunction.

Robust cellular models are key in determining pathological mechanisms that lead to neurotoxicity in Huntington's disease (HD) and for high throughput pre-clinical screening of potential therapeutic compounds. Such models exist but mostly comprise non-human or non-neuronal cells that may not recapitulate the correct biochemical milieu involved in pathology. We have developed a new human neuronal cell model of HD, using neural stem cells (ReNcell VM NSCs) stably transduced to express exon 1 huntingtin (HTT) fragments with variable length polyglutamine (polyQ) tracts. Using a system with matched expression levels of exon 1 HTT fragments, we investigated the effect of increasing polyQ repeat length on HTT inclusion formation, location, neuronal survival, and mitochondrial function with a view to creating an in vitro screening platform for therapeutic screening. We found that expression of exon 1 HTT fragments with longer polyQ tracts led to the formation of intra-nuclear inclusions in a polyQ length-dependent manner during neurogenesis. There was no overt effect on neuronal viability, but defects of mitochondrial function were found in the pathogenic lines. Thus, we have a human neuronal cell model of HD that may recapitulate some of the earliest stages of HD pathogenesis, namely inclusion formation and mitochondrial dysfunction.

Huntingtin Aggregates and Mitochondrial Pathology in Skeletal Muscle but not Heart of Late-Stage R6/2 Mice.

BACKGROUND: Cell or tissue specific background may influence the consequences of expressing the Huntington's disease (HD) mutation. Aggregate formation is known to occur in skeletal muscle, but not heart of the R6/2 fragment HD model. OBJECTIVE: We asked whether aggregate formation and the expression and subcellular localization of huntingtin species was associated with mitochondrial dysfunction. METHODS: We analyzed levels of soluble HTT and HTT aggregates, as well as important fission and fusion proteins and mitochondrial respiratory chain activities, in quadriceps and heart of the R6/2 N-terminal fragment mouse model (12 weeks, 160±10 CAG repeats). RESULTS: Soluble mutant HTT was present in both tissues with expression higher in cytoplasmic/mitochondrial than nuclear fractions. HTT aggregates were only detectable in R6/2 quadriceps, in association with increased levels of the pro-fission factor DRP1 and its phosphorylated active form, and decreased levels of the pro-fusion factor MFN2. In addition, respiratory chain complex activities were decreased. In heart that was without detectable HTT aggregates, we found no evidence for mitochondrial dysfunction. CONCLUSION: Tissue specific factors may exist that protect the R6/2 heart from HTT aggregate formation and mitochondrial pathology.

Meso scale discovery-based assays for the detection of aggregated huntingtin.

Huntington's disease (HD) is a monogenic neurodegenerative disorder caused by an expansion of the CAG trinucleotide repeat domain in the huntingtin (HTT) gene, leading to an expanded poly-glutamine (polyQ) stretch in the HTT protein. This mutant HTT (mHTT) protein is highly prone to intracellular aggregation, causing significant damage and cellular loss in the striatal, cortical, and other regions of the brain. Therefore, modulation of mHTT levels in these brain regions in order to reduce intracellular mHTT and aggregate levels represents a direct approach in the development of HD therapeutics. To this end, assays that can be used to detect changes in HTT levels in biological samples are invaluable tools to assess target engagement and guide dose selection in clinical trials. The Meso Scale Discovery (MSD) ELISA-based assay platform is a robust and sensitive method previously employed for the quantification of HTT. However, the currently available MSD assays for HTT are primarily detecting the monomeric soluble form of the protein, but not aggregated species. In this study, we describe the development of novel MSD assays preferentially detecting mHTT in an aggregated form. Recombinant monomeric HTT(1-97)-Q46, which forms aggregates in a time-dependent manner, was used to characterize the ability of each established assay to distinguish between HTT monomers and HTT in a higher assembly state. Further validation of these assays was performed using brain lysates from R6/2, zQ175 knock-in, and BACHD mouse models, to replicate a previously well-characterized age-dependent increase in brain aggregate signals, as well as a significant reduction of aggregate levels in the striatum following mHTT knockdown with a CAG-directed allele-specific zinc-finger repressor protein (ZFP). Lastly, size exclusion chromatography was used to separate and characterize HTT species from brain tissue lysates to demonstrate specificity of the assays for the fractions containing aggregated HTT. In summary, we demonstrate that the newly developed assays preferentially detect aggregated HTT with improved performance in comparison to previous assay technologies. These assays complement the existing MSD platform assays specific for soluble HTT monomers, allowing for a more comprehensive analysis of disease-relevant HTT species in preclinical models of HD.

Assessment of patient-reported outcomes after polytrauma: protocol for a systematic review.

INTRODUCTION: Survivors of polytrauma experience long-term and short-term burden that influences their lives. The patients' view of relevant short-term and long-term outcomes should be captured in instruments that measure quality of life and other patient-reported outcomes (PROs) after a polytrauma. The aim of this systematic review is to (1) collect instruments that assess PROs (quality of life, social participation and activities of daily living) during follow-up after polytrauma, (2) describe the instruments' application (eg, duration of period of follow-up) and (3) investigate other relevant PROs that are also assessed in the included studies (pain, depression, anxiety and cognitive function). METHODS AND ANALYSIS: The systematic review protocol is developed in line with the Preferred Reporting Items for Systematic Review and Meta-Analysis Protocols statement. MEDLINE, EMBASE, Cumulative Index to Nursing and Allied Health Literature, PsycINFO, Cochrane Central Register of Controlled Trials and the trials registers ClinicalTrials.gov and WHO International Clinical Trials Registry Platform will be searched. Keywords, for example, 'polytrauma', 'multiple trauma', 'quality of life', 'activities of daily living' or 'pain' will be used. Publications published between January 2005 and the most recent date (currently: August 2016) will be included. In order to present the latest possible results, an update of the search is conducted before publication. The data extraction and a content analysis will be carried out systematically. A critical appraisal will be performed. ETHICS AND DISSEMINATION: Formal ethical approval is not required as primary data will not be collected. The results will be published in a peer-reviewed publication. PROSPERO REGISTRATION NUMBER: CRD42017060825.

A standardised frankincense extract reduces disease activity in relapsing-remitting multiple sclerosis (the SABA phase IIa trial).

OBJECTIVE: To investigate whether oral administration of a standardised frankincense extract (SFE) is safe and reduces disease activity in patients with relapsing-remitting multiple sclerosis (RRMS). METHODS: We performed an investigator-initiated, bicentric phase IIa, open-label, baseline-to-treatment pilot study with an oral SFE in patients with RRMS (NCT01450124). After a 4-month baseline observation phase, patients were treated for 8 months with an option to extend treatment for up to 36 months. The primary outcome measures were the number and volume of contrast-enhancing lesions (CEL) measured in MRI during the 4-month treatment period compared with the 4-month baseline period. Eighty patients were screened at two centres, 38 patients were included in the trial, 28 completed the 8-month treatment period and 18 of these participated in the extension period. RESULTS: The SFE significantly reduced the median number of monthly CELs from 1.00 (IQR 0.75-3.38) to 0.50 (IQR 0.00-1.13; difference -0.625, 95% CI -1.25 to -0.50; P<0.0001) at months 5-8. We observed significantly less brain atrophy as assessed by parenchymal brain volume change (P=0.0081). Adverse events were generally mild (57.7%) or moderate (38.6%) and comprised mainly gastrointestinal symptoms and minor infections. Mechanistic studies showed a significant increase in regulatory CD4+ T cell markers and a significant decrease in interleukin-17A-producing CD8+ T cells indicating a distinct mechanism of action of the study drug. INTERPRETATION: The oral SFE was safe, tolerated well and exhibited beneficial effects on RRMS disease activity warranting further investigation in a controlled phase IIb or III trial. CLINICAL TRIAL REGISTRATION: NCT01450124; Results.

Phonological awareness in German-speaking preschool children with cochlear implants - 3 case examples.

OBJECTIVE: The aim was to explore PA skills German-speaking preschool children with cochlea implants (CIs) and how these skills may be related to their speech and language skills. METHODS: Three monolingual German-speaking pre-school children aged 5;04-6;01 with bilateral CIs were tested. Their cognitive, speech and language skills were assessed. Six subtests of a standardized PA test battery were administered (i.e. rhyme identification, rhyme production; phoneme identification- input and -output; phoneme blending-input and -output). RESULTS: All three children showed distinctive PA profiles. One boy, who had no spoken language deficits, struggled to complete the rhyme tasks but performed well on three phoneme tasks. However, he showed a discrepancy between expressive and receptive phoneme blending skills, scoring poorly on the expressive subtest. The second boy, who displayed grammar comprehension and expressive vocabulary difficulties, showed a mixed profile, with a below average performance on rhyme production. The girl who had significant speech and language deficits scored below average on all six PA subtests. CONCLUSIONS: PA profiles in children with CI vary considerably and PA testing should include a range of different PA tasks. The assumed link between spoken language deficits and PA difficulties shown in children with normal hearing could be confirmed.

Simulation of future groundwater recharge using a climate model ensemble and SAR-image based soil parameter distributions - A case study in an intensively-used Mediterranean catchment.

We used observed climate data, an ensemble of four GCM-RCM combinations (global and regional climate models) and the water balance model mGROWA to estimate present and future groundwater recharge for the intensively-used Thau lagoon catchment in southern France. In addition to a highly resolved soil map, soil moisture distributions obtained from SAR-images (Synthetic Aperture Radar) were used to derive the spatial distribution of soil parameters covering the full simulation domain. Doing so helped us to assess the impact of different soil parameter sources on the modelled groundwater recharge levels. Groundwater recharge was simulated in monthly time steps using the ensemble approach and analysed in its spatial and temporal variability. The soil parameters originating from both sources led to very similar groundwater recharge rates, proving that soil parameters derived from SAR images may replace traditionally used soil maps in regions where soil maps are sparse or missing. Additionally, we showed that the variance in different GCM-RCMs influences the projected magnitude of future groundwater recharge change significantly more than the variance in the soil parameter distributions derived from the two different sources. For the period between 1950 and 2100, climate change impacts based on the climate model ensemble indicated that overall groundwater recharge will possibly show a low to moderate decrease in the Thau catchment. However, as no clear trend resulted from the ensemble simulations, reliable recommendations for adapting the regional groundwater management to changed available groundwater volumes could not be derived.

Onset and phoneme awareness and its relationship to letter knowledge in German-speaking preschool children.

OBJECTIVES: The aim was to explore whether word-initial onset awareness is acquired before phoneme awareness and whether onset complexity influences performance on identification tasks. In addition, the relationship between onset and phoneme awareness and letter knowledge was investigated. METHOD: In this study, 22 monolingual German-speaking preschool children aged 5;00-5;11 were tested. Onset, phoneme identification, and letter knowledge tasks were administered. The children were presented with pictures of word pairs. Both words in each pair shared a single-consonant onset, a two-consonant onset cluster or the first consonant of a consonant cluster. The children were asked to pronounce the shared sound(s). Additionally, they were asked to name all 26 upper-case letters. RESULTS: Onset awareness tasks were significantly easier to complete than phoneme awareness tasks. However, no influence of onset complexity on onset awareness performance was found. Moreover, letter knowledge correlated with all phonological awareness tasks. CONCLUSIONS: The results corroborate that phoneme awareness develops already at preschool age irrespective of explicit literacy tuition. Nevertheless, letter knowledge is closely related and should be linked to onset/phoneme awareness tasks.

Quantification assays for total and polyglutamine-expanded huntingtin proteins.

The expansion of a CAG trinucleotide repeat in the huntingtin gene, which produces huntingtin protein with an expanded polyglutamine tract, is the cause of Huntington's disease (HD). Recent studies have reported that RNAi suppression of polyglutamine-expanded huntingtin (mutant HTT) in HD animal models can ameliorate disease phenotypes. A key requirement for such preclinical studies, as well as eventual clinical trials, aimed to reduce mutant HTT exposure is a robust method to measure HTT protein levels in select tissues. We have developed several sensitive and selective assays that measure either total human HTT or polyglutamine-expanded human HTT proteins on the electrochemiluminescence Meso Scale Discovery detection platform with an increased dynamic range over other methods. In addition, we have developed an assay to detect endogenous mouse and rat HTT proteins in pre-clinical models of HD to monitor effects on the wild type protein of both allele selective and non-selective interventions. We demonstrate the application of these assays to measure HTT protein in several HD in vitro cellular and in vivo animal model systems as well as in HD patient biosamples. Furthermore, we used purified recombinant HTT proteins as standards to quantitate the absolute amount of HTT protein in such biosamples.