Search

The characterisation of resting-state networks (RSNs) using neuroimaging techniques has significantly contributed to our understanding of the organisation of brain activity. Prior work has demonstrated the electrophysiological basis of RSNs and their dynamic nature, revealing transient activations of brain networks with millisecond timescales. While previous research has confirmed the comparability of RSNs identified by electroencephalography (EEG) to those identified by magnetoencephalography (MEG) and functional magnetic resonance imaging (fMRI), most studies have utilised static analysis techniques, ignoring the dynamic nature of brain activity. Often, these studies use high-density EEG systems, which limit their applicability in clinical settings. Addressing these gaps, our research studies RSNs using medium-density EEG systems (61 sensors), comparing both static and dynamic brain network features to those obtained from a high-density MEG system (306 sensors). We assess the qualitative and quantitative comparability of EEG-derived RSNs to those from MEG, including their ability to capture age-related effects, and explore the reproducibility of dynamic RSNs within and across the modalities. Our findings suggest that both MEG and EEG offer comparable static and dynamic network descriptions, albeit with MEG offering some increased sensitivity and reproducibility. Such RSNs and their comparability across the two modalities remained consistent qualitatively but not quantitatively when the data were reconstructed without subject-specific structural MRI images., (© 2024 The Author(s). Human Brain Mapping published by Wiley Periodicals LLC.)

Purpose: Speech-language pathology (SLP) is considered an essential intervention due to the high prevalence of dysphagia and dysarthria in paediatric neuromuscular disorders (pNMD). Evidence-based guidelines for SLP in pNMD are missing and children could be deprived the best of care. This study aimed to achieve consensus and present best practice recommendations on SLP intervention in pNMD., Method: A modified Delphi technique was used with a panel of experienced Dutch speech-language pathologists. In two online survey rounds and a face-to-face consensus meeting, the SLP experts proposed intervention items for cases of four types of pNMD (congenital myopathy, Duchenne muscular dystrophy, myotonic dystrophy type 1, and spinal muscular atrophy type 2), covering symptoms of dysphagia, dysarthria, drooling, and oral hygiene problems. They rated the level of agreement., Result: Intervention items that achieved consensus were incorporated into best practice recommendations. These recommendations cover six core intervention components (wait and see, explanation and advice, training and treatment, aids and adjustments, referral to other disciplines, and monitoring) suitable for the described symptoms., Conclusion: Insight into treatment options is essential to facilitate speech-language pathologists in clinical decision-making. The current study led to best practice recommendations for speech-language pathologists working within the field of pNMD.

Background & Aim: Irradiation of the salivary glands during head and neck cancer treatment induces cellular senescence in response to DNA damage and contributes to radiation-induced hyposalivation by affecting the salivary gland stem/progenitor cell (SGSC) niche. Cellular senescence, such as that induced by radiation, is a state of cell-cycle arrest, accompanied by an altered pro-inflammatory secretome known as the senescence-associated secretory phenotype (SASP) with potential detrimental effects on the surrounding microenvironment. We hypothesized that the pro-regenerative properties of mesenchymal stem cells (MSCs) may attenuate cellular senescence post-irradiation. Therefore, here we evaluated the effects of adipose-derived MSCs (ADSCs) on the radiation-induced response of salivary gland organoids (SGOs)., Methods: Proteomic analyses to identify soluble mediators released by ADSCs co-cultured with SGOS revealed secretion of hepatocyte growth factor (HGF) in ADSCs, suggesting a possible role in the stem cell crosstalk. Next, the effect of recombinant HGF in the culture media of ex vivo grown salivary gland cells was tested in 2D monolayers and 3D organoid models., Results: Treatment with HGF robustly increased salivary gland cell proliferation. Importantly, HGF supplementation post-irradiation enhanced proliferation at lower doses of radiation (0, 3, 7 Gy), but not at higher doses (10, 14 Gy) where most cells stained positive for senescence-associated beta-galactosidase. Furthermore, HGF had no effect on the senescence-associated secretory phenotype (SASP) of irradiated SGOs, suggesting there may be compensatory proliferation by cell-division competent cells instead of a reversal of cellular senescence after irradiation., Conclusion: ADSCs may positively influence radiation recovery through HGF secretion and can promote the ex vivo expansion of salivary gland stem/progenitor cells to enhance the effects of co-transplanted SGSC., Competing Interests: Declaration of competing interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Coppes R.P. reports financial support was provided by Dutch Cancer Society. Coppes R.P. serves as an editor for Radiother Oncol., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

The aim of this study was to investigate the surgical and long-term neurological outcomes of patients with acetylcholine-receptor-antibody-associated myasthenia gravis (AChR-MG) who underwent robotic thymectomy (RATS). We retrospectively analyzed the clinical-pathological data of all patients with AChR-MG who underwent RATS using the DaVinci® Robotic System at the MUMC+ between April 2004 and December 2018. Follow-up data were collected from 60 referring Dutch hospitals. In total, 230 myasthenic patients including 76 patients with a thymoma (33.0%) were enrolled in this study. Mean follow-up time, procedure time and hospitalization were, respectively 65.7 ± 43.1 months, 111±52.5 min and 3.3 ± 2.2 days. Thymomatous patients had significantly more frequently and more severe complications than nonthymomatous patients (18.4% vs. 3.9%, p<0.001). Follow up data was available in 71.7% of the included patients. The Myasthenia Gravis Foundation of America postintervention score showed any kind of improvement of MG-symptoms after RATS in 82.4% of the patients. Complete stable remission (CSR) or pharmacological remission (PR) of MG was observed in 8.4% and 39.4% of the patients, respectively. Mean time till CSR/PR remission after thymectomy was 26.2 ± 29.2 months. No statistical difference was found in remission or improvement in MGFA scale between thymomatous and nonthymomatous patients. RATS is safe and feasible in patients with MG. The majority of the patients (82.4%) improved after thymectomy. CSR and PR were observed in 8.4% and 39.4% of the patients, respectively, with a mean of 26.2 months after thymectomy. Thymomatous patients had more frequently and more severe complications compared to nonthymomatous patients., Competing Interests: Declaration of Competing Interest None., (Copyright © 2023 The Author(s). Published by Elsevier B.V. All rights reserved.)

Since the original outbreak of the SARS-CoV-2 virus, several rapidly spreading SARS-CoV-2 variants of concern (VOC) have emerged. Here, we show that a single dose of Ad26.COV2.S (based on the Wuhan-Hu-1 spike variant) protects against the Gamma and Delta variants in naive hamsters, supporting the observed maintained vaccine efficacy in humans against these VOC. Adapted spike-based booster vaccines targeting Omicron variants have now been authorized in the absence of human efficacy data. We evaluated the immunogenicity and efficacy of Ad26.COV2.S.529 (encoding a stabilized Omicron BA.1 spike) in naive mice and in hamsters with pre-existing immunity to the Wuhan-Hu-1 spike. In naive mice, Ad26.COV2.S.529 elicited higher neutralizing antibody titers against SARS-CoV-2 Omicron BA.1 and BA.2, compared with Ad26.COV2.S. However, neutralizing titers against the SARS-CoV-2 B.1 (D614G) and Delta variants were lower after primary vaccination with Ad26.COV2.S.529 compared with Ad26.COV2.S. In contrast, we found comparable Omicron BA.1 and BA.2 neutralizing titers in hamsters with pre-existing Wuhan-Hu-1 spike immunity after vaccination with Ad26.COV2.S, Ad26.COV2.S.529 or a combination of the two vaccines. Moreover, all three vaccine modalities induced equivalent protection against Omicron BA.2 challenge in these animals. Overall, our data suggest that an Omicron BA.1-based booster in rodents does not improve immunogenicity and efficacy against Omicron BA.2 over an Ad26.COV2.S booster in a setting of pre-existing immunity to SARS-CoV-2., (© 2023. The Author(s).)

Purpose: The primary aim was to describe sports participation of Dutch children and adolescents with lower limb deficiencies (LLD). The secondary aim was to explore perceived limitations concerning sports participation., Methods: A total of 103 children and adolescents with LLD, aged 8-18 years (mean 11.7 years), were asked about their sports participation using a study-specific self-report questionnaire., Results: Children and adolescents with LLD frequently (78%) participated in sports activities, and most of them (68%) participated in the sport of their preference. Just over half of all children (52%) perceived an inability to participate in specific sports. Physical performance (running) and endurance were mentioned as the most limiting factors in participating in certain sports., Conclusion: Children and adolescents with LLD in the Netherlands participate in a variety of sports. Despite dependency on lower limb prostheses in most cases, children and adolescents with LLD have a high potentiality of participating in sports.

Background: Mortality caused by Traumatic Brain Injury (TBI) remains high, despite improvements in trauma and critical care. Polytrauma is naturally associated with high mortality. This study compared mortality rates between isolated TBI ( I TBI) patients and polytrauma patients with TBI ( P TBI) admitted to ICU to investigate if concomitant injuries lead to higher mortality amongst TBI patients., Methods: A 3-year cohort study compared polytrauma patients with TBI ( P TBI) with AIS head ≥3 (and AIS of other body regions ≥3) from a prospective collected database to isolated TBI ( I TBI) patients from a retrospective collected database with AIS head ≥3 (AIS of other body regions ≤2), both admitted to a single level-I trauma center ICU. Patients <16 years of age, injury caused by asphyxiation, drowning, burns and ICU transfers from and to other hospitals were excluded. Patient demographics, shock and resuscitation parameters, multiple organ dysfunction syndrome (MODS), acute respiratory distress syndrome (ARDS), and mortality data were collected and analyzed for group differences., Results: 259 patients were included; 111 P TBI and 148 I TBI patients. The median age was 54 [33-67] years, 177 (68%) patients were male, median ISS was 26 [20-33]. Seventy-nine (31%) patients died. Patients with P TBI developed more ARDS (7% vs. 1%, p = 0.041) but had similar MODS rates (18% vs. 10%, p = 0.066). They also stayed longer on the ventilator (7 vs. 3 days, p=<0.001), longer in ICU (9 vs. 4 days, p=<0.001) and longer in hospital (24 vs. 11 days, p=<0.001). TBI was the most prevalent cause of death in polytrauma patients. Patients with P TBI showed no higher in-hospital mortality rate. Moreover, mortality rates were skewed towards I TBI patients (24% vs. 35%, p = 0.06)., Discussion: There was no difference in mortality rates between P TBI and I TBI patients, suggesting TBI-severity as the predominant factor for ICU mortality in an era of ever improving acute trauma care., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interest, financially or otherwise., (Copyright © 2022. Published by Elsevier Ltd.)

Objective: The role of the survival of motor neuron (SMN) gene in amyotrophic lateral sclerosis (ALS) is unclear, with several conflicting reports. A decisive result on this topic is needed, given that treatment options are available now for SMN deficiency., Methods: In this largest multicenter case control study to evaluate the effect of SMN1 and SMN2 copy numbers in ALS, we used whole genome sequencing data from Project MinE data freeze 2. SMN copy numbers of 6,375 patients with ALS and 2,412 controls were called from whole genome sequencing data, and the reliability of the calls was tested with multiplex ligation-dependent probe amplification data., Results: The copy number distribution of SMN1 and SMN2 between cases and controls did not show any statistical differences (binomial multivariate logistic regression SMN1 p = 0.54 and SMN2 p = 0.49). In addition, the copy number of SMN did not associate with patient survival (Royston-Parmar; SMN1 p = 0.78 and SMN2 p = 0.23) or age at onset (Royston-Parmar; SMN1 p = 0.75 and SMN2 p = 0.63)., Interpretation: In our well-powered study, there was no association of SMN1 or SMN2 copy numbers with the risk of ALS or ALS disease severity. This suggests that changing SMN protein levels in the physiological range may not modify ALS disease course. This is an important finding in the light of emerging therapies targeted at SMN deficiencies. ANN NEUROL 2021;89:686-697., (© 2021 The Authors. Annals of Neurology published by Wiley Periodicals LLC on behalf of American Neurological Association.)

Introduction: An increasing number of women becomes pregnant while suffering from chronic kidney disease (CKD). As a result of decreased renal function, uremic solutes circulate at high levels in the maternal circulation. This study aimed to acquire more knowledge about the placental transfer of uremic solutes across the human placenta., Methods: Placental transfer was studied in healthy term placentas, via the ex vivo dual-side human cotyledon perfusion technique (closed-closed set-up for both maternal and fetal circulations). Uremic solute concentrations in maternal and fetal perfusates were measured via LC-MS/MS over 180 min of perfusion., Results: We found that the studied compounds demonstrated different degrees of placental transfer. Fetal-to-maternal perfusate ratios at t = 180 min were for anthranilic acid 1.00 ± 0.02, indole-3-acetic acid 0.47 ± 0.08, hippuric acid 0.36 ± 0.18, l-arabinitol 0.33 ± 0.04, indoxyl sulfate 0.33 ± 0.11, neopterin 0.28 ± 0.14 and kynurenic acid 0.13 ± 0.03. All uremic solutes studied also emerged in the perfusates when cotyledons were perfused in the absence of uremic solute concentrations added to the maternal reservoir. For kynurenin these concentrations were so high, it complicated the calculation of a transfer ratio for the exogenously administered compound., Discussion: After 180 min of exposure the extent of placental transfer differs substantially for the solutes studied, reflecting different transfer rates. Future studies should investigate to what extent specific uremic solutes reach the fetal circulation in vivo and how they may interfere with organ function and development of the unborn child., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: Dysphagia and dysarthria are frequently described in pediatric neuromuscular diseases (pNMD). The consequences can be substantial: failure to thrive, malnutrition, aspiration pneumonia, or communication problems. Early detection and identification of risk factors and etiology support preventing complications and morbidity, including impact on quality of life. Information about the prevalence of dysphagia and dysarthria in pNMD is scarce., Objective: To describe the pooled prevalence of dysphagia and dysarthria in pNMD in the Netherlands. In addition, we describe the prevalence of dysphagia and dysarthria each, and the prevalence of chewing (oral) and swallowing problems per diagnostic group, based on their anatomic origin., Methods: Data were collected from 295 children (mean age 11;0 years, range 2;6-18;0) with pNMD in 12 hospitals and rehabilitation centers in the Netherlands. A speech language therapist established whether dysphagia and dysarthria were present or not., Results: In almost all the 14 diagnostic groups of pNMD, dysphagia and dysarthria were present. Pooled overall prevalence of dysphagia and dysarthria was 47.2% and 31.5%, respectively. Of 114 children with dysphagia, 90.0% had chewing problems, 43.0% showed swallowing problems and 33.3% showed both chewing and swallowing problems., Conclusions: The overall pooled prevalence of dysphagia and dysarthria was high in the population of pNMD. It can be argued that periodic monitoring of dysphagia and dysarthria and early referral to a speech language therapist should be a necessity from the start of the diagnosis in the whole pNMD population.

The characterization of buried nanoscale structures nondestructively is an important challenge in a number of applications, such as defect detection and metrology in the semiconductor industry. A promising technique is Subsurface Scanning Probe Microscopy (SSPM), which combines ultrasound with Atomic Force Microscopy (AFM). Initially, SSPM was used to measure the viscoelastic contrast between a subsurface feature and its surrounding medium. However, by increasing the ultrasonic frequency to >1 GHz, it has been shown that SSPM can also measure acoustic impedance based contrasts. At these frequencies, it becomes difficult to reliably couple the sound into the sample such that the AFM is able to pick up the scattered sound field. The cause is the existence of strong acoustic resonances in the sample, the transducer, and the coupling layer-the liquid layer used to couple the sound energy from the transducer into the sample-in combination with the nonlinearity of the tip-sample interaction. Thus, it is essential to control and measure the thickness of the coupling layer with nanometer accuracy. Here, we present the design of a mechanical clamp to ensure a stable acoustic coupling. Moreover, an acoustic method is presented to measure the coupling layer thickness in real-time. Stable coupling layers with thicknesses of 700 ± 2 nm were achieved over periods of 2-4 h. Measurements of the downmixed AFM signals showed stable signal intensities for >1 h. The clamp and monitoring method introduced here makes scattering based SSPM practical, robust, and reliable and enables measurement periods of hours.

Background and Purpose: Previous studies have reported that many patients with a severe head injury are not transported to a higher-level trauma centre where the necessary round-the-clock neurosurgical care is available. The aim of this study was to analyse the diagnostic value of emergency medical services (EMS) provider judgement in the identification of a head injury., Methods: In this multicentre cohort study, all trauma patients aged 16 years and over who were transported with highest priority to a trauma centre were evaluated. The diagnostic value of EMS provider judgement was determined using an Abbreviated Injury Scale score of ≥1 in the head region as reference standard., Results: A total of 980 (35.4%) of the 2766 patients who were included had a head injury. EMS provider judgement (Abbreviated Injury Scale score ≥1) had a sensitivity of 67.9% and a specificity of 87.7%. In the cohort, 208 (7.5%) patients had a severe head injury. Of these, 68% were transported to a level I trauma centre., Conclusions: Identification of a head injury on-scene is challenging. EMS providers could not identify 32% of the patients with a head injury and 21% of the patients with a severe head injury. Additional education, training and a supplementary protocol with predictors of a severe head injury could help EMS providers in the identification of these patients., (© 2018 EAN.)

Introduction: Traumatic brain injury (TBI) remains a major cause of death. Withdrawal of life-sustaining treatment (WLST) can be initiated if there is little anticipated chance of recovery to an acceptable quality of life. The aim of this study was firstly to investigate WLST rates in patients with moderate to severe isolated TBI and secondly to assess outcome data in the survivor group., Material and Methods: A retrospective cohort study was performed. Patients aged ≥ 18 years with moderate or severe isolated TBI admitted to the ICU of a single academic hospital between 2011 and 2015 were included. Exclusion criteria were isolated spinal cord injury and referrals to and from other hospitals. Gathered data included demographics, mortality, cause of death, WLST, and Glasgow Outcome Scale (GOS) score after three months. Good functional outcome was defined as GOS > 3., Results: Of 367 patients, 179 patients were included after applying inclusion and exclusion criteria. 55 died during admission (33%), of whom 45 (82%) after WLST. Patients undergoing WLST were older, had worse neurological performance at presentation, and had more radiological abnormalities than patients without WLST. The decision to withdraw life-sustaining treatment was made on the day of admission in 40% of patients. In 33% of these patients, this decision was made while the patient was in the Emergency Department. 71% of survivors had a good functional outcome after three months. No patient left hospital with an unresponsive wakefulness syndrome (UWS) or suffered from UWS after three months. One patient died within three months of discharge., Conclusion: In-hospital mortality in isolated brain injured patients was 33%. The vast majority died after a decision to withdraw life-sustaining treatment. None of the patients were discharged with an unresponsive wakefulness syndrome.

Frontotemporal dementia (FTD) is a neurodegenerative disorder predominantly affecting the frontal and temporal lobes. Genome-wide association studies (GWAS) on FTD identified only a few risk loci. One of the possible explanations is that FTD is clinically, pathologically, and genetically heterogeneous. An important open question is to what extent epigenetic factors contribute to FTD and whether these factors vary between FTD clinical subgroup. We compared the DNA-methylation levels of FTD cases (n = 128), and of FTD cases with Amyotrophic Lateral Sclerosis (FTD-ALS; n = 7) to those of unaffected controls (n = 193), which resulted in 14 and 224 candidate genes, respectively. Cluster analysis revealed significant class separation of FTD-ALS from controls. We could further specify genes with increased susceptibility for abnormal gene-transcript behavior by jointly analyzing DNA-methylation levels with the presence of mutations in a GWAS FTD-cohort. For FTD-ALS, this resulted in 9 potential candidate genes, whereas for FTD we detected 1 candidate gene (ELP2). Independent validation-sets confirmed the genes DLG1, METTL7A, KIAA1147, IGHMBP2, PCNX, UBTD2, WDR35, and ELP2/SLC39A6 among others. We could furthermore demonstrate that genes harboring mutations and/or displaying differential DNA-methylation, are involved in common pathways, and may therefore be critical for neurodegeneration in both FTD and FTD-ALS.

Sodium channel Nav1.7, encoded by SCN9A, is expressed in DRG neurons and regulates their excitability. Genetic and functional studies have established a critical contribution of Nav1.7 to human pain disorders. We have now characterized a novel Nav1.7 mutation (R1279P) from a female human subject with distal limb pain, in which depolarized fast inactivation overrides impaired activation to produce hyperexcitability and spontaneous firing in DRG neurons. Whole-cell voltage-clamp recordings in human embryonic kidney (HEK) 293 cells demonstrated that R1279P significantly depolarizes steady-state fast-, slow-, and closed-state inactivation. It accelerates deactivation, decelerates inactivation, and facilitates repriming. The mutation increases ramp currents in response to slow depolarizations. Our voltage-clamp analysis showed that R1279P depolarizes channel activation, a change that was supported by our multistate structural modeling. Because this mutation confers both gain-of-function and loss-of-function attributes on the Nav1.7 channel, we tested the impact of R1279P expression on DRG neuron excitability. Current-clamp studies reveal that R1279P depolarizes resting membrane potential, decreases current threshold, and increases firing frequency of evoked action potentials within small DRG neurons. The populations of spontaneously firing and repetitively firing neurons were increased by expressing R1279P. These observations indicate that the dominant proexcitatory gating changes associated with this mutation, including depolarized steady-state fast-, slow-, and closed-state inactivation, faster repriming, and larger ramp currents, override the depolarizing shift of activation, to produce hyperexcitability and spontaneous firing of nociceptive neurons that underlie pain., (Copyright © 2014 the authors 0270-6474/14/3412328-13$15.00/0.)

We present a new method to analyse simultaneous Topography and RECognition Atomic Force Microscopy data such that it becomes possible to measure single molecule binding rates of surface bound proteins. We have validated this method on a model system comprising a S-layer surface modified with Strep-tagII for binding sites and strep-tactin bound to an Atomic Force Microscope tip through a flexible Poly-Ethylene-Glycol linker. At larger distances, the binding rate is limited by the linker, which limits the diffusion of the strep-tactin molecule, but at lateral distances below 3 nm, the binding rate is solely determined by the intrinsic molecular characteristics and the surface geometry and chemistry of the system. In this regime, Kon as determined from single molecule TREC data is in agreement with Kon determined using traditional biochemical methods., (Copyright © 2014 The Authors. Published by Elsevier B.V. All rights reserved.)

Hepatitis E virus (HEV) is classified within the family Hepeviridae, genus Hepevirus. HEV genotype 3 (Gt3) infections are endemic in pigs in Western Europe and in North and South America and cause zoonotic infections in humans. Several serological assays to detect HEV antibodies in pigs have been developed, at first mainly based on HEV genotype 1 (Gt1) antigens. To develop a sensitive HEV Gt3 ELISA, a recombinant baculovirus expression product of HEV Gt3 open reading frame-2 was produced and coated onto polystyrene ELISA plates. After incubation of porcine sera, bound HEV antibodies were detected with anti-porcine anti-IgG and anti-IgM conjugates. For primary estimation of sensitivity and specificity of the assay, sets of sera were used from pigs experimentally infected with HEV Gt3. For further validation of the assay and to set the cutoff value, a batch of 1100 pig sera was used. All pig sera were tested using the developed HEV Gt3 assay and two other serologic assays based on HEV Gt1 antigens. Since there is no gold standard available for HEV antibody testing, further validation and a definite setting of the cutoff of the developed HEV Gt3 assay were performed using a statistical approach based on Bayes' theorem. The developed and validated HEV antibody assay showed effective detection of HEV-specific antibodies. This assay can contribute to an improved detection of HEV antibodies and enable more reliable estimates of the prevalence of HEV Gt3 in swine in different regions.

Genome-wide association studies have been successful in identifying common variants that influence the susceptibility to complex diseases. From these studies, it has emerged that there is substantial overlap in susceptibility loci between diseases. In line with those findings, we hypothesized that shared genetic pathways may exist between multiple sclerosis (MS) and amyotrophic lateral sclerosis (ALS). While both diseases may have inflammatory and neurodegenerative features, epidemiological studies have indicated an increased co-occurrence within individuals and families. To this purpose, we combined genome-wide data from 4088 MS patients, 3762 ALS patients and 12 030 healthy control individuals in whom 5 440 446 single-nucleotide polymorphisms (SNPs) were successfully genotyped or imputed. We tested these SNPs for the excess association shared between MS and ALS and also explored whether polygenic models of SNPs below genome-wide significance could explain some of the observed trait variance between diseases. Genome-wide association meta-analysis of SNPs as well as polygenic analyses fails to provide evidence in favor of an overlap in genetic susceptibility between MS and ALS. Hence, our findings do not support a shared genetic background of common risk variants in MS and ALS.

Atomic force microscopy (AFM) is widely used in the study of both morphology and mechanical properties of living cells under physiologically relevant conditions. However, quantitative experiments on timescales of minutes to hours are generally limited by thermal drift in the instrument, particularly in the vertical (z) direction. In addition, we demonstrate the necessity to remove all air-liquid interfaces within the system for measurements in liquid environments, which may otherwise result in perturbations in the measured deflection. These effects severely limit the use of AFM as a practical tool for the study of long-term cell behavior, where precise knowledge of the tip-sample distance is a crucial requirement. Here we present a readily implementable, cost effective method of minimizing z-drift and liquid instabilities by utilizing active temperature control combined with a customized fluid cell system. Long-term whole cell mechanical measurements were performed using this stabilized AFM by attaching a large sphere to a cantilever in order to approximate a parallel plate system. An extensive examination of the effects of sphere attachment on AFM data is presented. Profiling of cantilever bending during substrate indentation revealed that the optical lever assumption of free ended cantilevering is inappropriate when sphere constraining occurs, which applies an additional torque to the cantilevers "free" end. Here we present the steps required to accurately determine force-indentation measurements for such a scenario. Combining these readily implementable modifications, we demonstrate the ability to investigate long-term whole cell mechanics by performing strain controlled cyclic deformation of single osteoblasts.

The objective of this study was to investigate the nature and biomechanical properties of collagen fibers within the human myocardium. Targeting cardiac interstitial abnormalities will likely become a major focus of future preventative strategies with regard to the management of cardiac dysfunction. Current knowledge regarding the component structures of myocardial collagen networks is limited, further delineation of which will require application of more innovative technologies. We applied a novel methodology involving combined confocal laser scanning and atomic force microscopy to investigate myocardial collagen within ex-vivo right atrial tissue from 10 patients undergoing elective coronary bypass surgery. Immuno-fluorescent co-staining revealed discrete collagen I and III fibers. During single fiber deformation, overall median values of stiffness recorded in collagen III were 37±16% lower than in collagen I [p<0.001]. On fiber retraction, collagen I exhibited greater degrees of elastic recoil [p<0.001; relative percentage increase in elastic recoil 7±3%] and less energy dissipation than collagen III [p<0.001; relative percentage increase in work recovered 7±2%]. In atrial biopsies taken from patients in permanent atrial fibrillation (n=5) versus sinus rhythm (n=5), stiffness of both collagen fiber subtypes was augmented (p<0.008). Myocardial fibrillar collagen fibers organize in a discrete manner and possess distinct biomechanical differences; specifically, collagen I fibers exhibit relatively higher stiffness, contrasting with higher susceptibility to plastic deformation and less energy efficiency on deformation with collagen III fibers. Augmented stiffness of both collagen fiber subtypes in tissue samples from patients with atrial fibrillation compared to those in sinus rhythm are consistent with recent published findings of increased collagen cross-linking in this setting., (Copyright © 2011 Elsevier Ltd. All rights reserved.)

Background: The microtubule-associated protein tau is thought to play a pivotal role in neurodegeneration. Mutations in the tau coding gene MAPT are a cause of frontotemporal dementia, and the H1/H1 genotype of MAPT, giving rise to higher tau expression levels, is associated with progressive supranuclear palsy, corticobasal degeneration, and Parkinson disease (PD). Furthermore, tau hyperphosphorylation and aggregation is a hallmark of Alzheimer disease (AD), and reducing endogenous tau has been reported to ameliorate cognitive impairment in a mouse model for AD. Tau hyperphosphorylation and aggregation have also been described in amyotrophic lateral sclerosis (ALS), both in human patients and in the mutant SOD1 mouse model for this disease. However, the precise role of tau in motor neuron degeneration remains uncertain., Methods: The possible association between ALS and the MAPT H1/H2 polymorphism was studied in 3,540 patients with ALS and 8,753 controls. Furthermore, the role of tau in the SOD1(G93A) mouse model for ALS was studied by deleting Mapt in this model., Results: The MAPT genotype of the H1/H2 polymorphism did not influence ALS susceptibility (odds ratio = 1.08 [95% confidence interval 0.99-1.18], p = 0.08) and did not affect the clinical phenotype. Lowering tau levels in the SOD1(G93A) mouse failed to delay disease onset (p = 0.302) or to increase survival (p = 0.557)., Conclusion: These findings suggest that the H1/H2 polymorphism in MAPT is not associated with human amyotrophic lateral sclerosis, and that lowering tau levels in the mutant SOD1 mouse does not affect the motor neuron degeneration in these animals.

A compact, two-stage nanomanipulator was designed and built for use inside a scanning electron microscope. It consists of a fine stage employing piezostacks that provide a 15 microm range in three dimensions and a coarse stage based on commercially available stick-slip motors. Besides the fabrication of enhanced probes for scanning probe microscopy and the enhancement of electron field emitters, other novel manipulation processes were developed, such as locating, picking up, and positioning small nanostructures with an accuracy of approximately 10 nm. In combination with in situ I-V experiments, welding, and etching, this results in a multipurpose nanofactory, enabling a new range of experiments.

An aneurysm of the aorta is a common pathology characterized by segmental weakening of the artery. Although it is generally accepted that the vessel-wall weakening is caused by an impaired collagen metabolism, a clear association has been demonstrated only for rare syndromes such as the vascular type Ehlers-Danlos syndrome. Here we show that vessel-wall failure in growing aneurysms of patients who have aortic abdominal aneurysm (AAA) or Marfan syndrome is not related to a collagen defect at the molecular level. On the contrary our findings indicate similar (Marfan) or even higher collagen concentrations (AAA) and increased collagen cross-linking in the aneurysms. Using 3D confocal imaging we show that the two conditions are associated with profound defects in collagen microarchitecture. Reconstructions of normal vessel wall show that adventitial collagen fibers are organized in a loose braiding of collagen ribbons. These ribbons encage the vessel, allowing the vessel to dilate easily but preventing overstretching. AAA and aneurysms in Marfan syndrome show dramatically altered collagen architectures with loss of the collagen knitting. Evaluations of the functional characteristics by atomic force microscopy showed that the wall has lost its ability to stretch easily and revealed a second defect: although vascular collagen in normal aortic wall behaves as a coherent network, in AAA and Marfan tissues it does not. As result, mechanical forces loaded on individual fibers are not distributed over the tissue. These studies demonstrate that the mechanical properties of tissue are strongly influenced by collagen microarchitecture and that perturbations in the collagen networks may lead to mechanical failure.

Background: Six candidate gene studies report a genetic association of DNA variants within the paraoxonase locus with sporadic amyotrophic lateral sclerosis (ALS). However, several other large studies, including five genome-wide association studies, have not duplicated this finding., Methods: We conducted a meta-analysis of 10 published studies and one unpublished study of the paraoxonase locus, encompassing 4,037 ALS cases and 4,609 controls, including genome-wide association data from 2,018 ALS cases and 2,425 controls., Results: The combined fixed effects odds ratio (OR) for rs662 (PON1 Q192R) was 1.09 (95% confidence interval [CI], 1.02-1.16, p = 0.01); the genotypic OR for RR homozygotes at Q192R was 1.25 (95% CI, 1.07-1.45, p = 0.0004); the combined OR for rs854560 (PON1 L55M) was 0.97 (95% CI, 0.86-1.10, p = 0.62); the OR for rs10487132 (PON2) was 1.08 (95% CI, 0.92-1.27, p = 0.35). Although the rs662 polymorphism reached a nominal level of significance, no polymorphism was significant after multiple testing correction. In the subanalysis of samples with genome-wide data from which population outliers were removed, rs662 had an OR of 1.06 (95% CI, 0.97-1.16, p = 0.22)., Conclusions: In contrast to previous positive smaller studies, our genetic meta-analysis showed no significant association of amyotrophic lateral sclerosis (ALS) with the PON locus. This is the largest meta-analysis of a candidate gene in ALS to date and the first ALS meta-analysis to include data from whole genome association studies. The findings reinforce the need for much larger and more collaborative investigations of the genetic determinants of ALS.

Objectives: Null mutations in progranulin (PGRN) cause ubiquitin-positive frontotemporal dementia (FTD) linked to chromosome 17q21 (FTDU-17). Here we examined PGRN genetic variability in amyotrophic lateral sclerosis (ALS), a neurodegenerative motor neuron disease that overlaps with FTD at a clinical, pathologic, and epidemiologic level., Methods: We sequenced all exons, exon-intron boundaries, and 5' and 3' regulatory regions of PGRN in a Belgian sample of 230 patients with ALS. The frequency of observed genetic variants was determined in 436 healthy control individuals. The contribution of eight frequent polymorphisms to ALS risk, onset age, and survival was assessed in an association study in the Belgian sample and a replication series of 308 Dutch patients with ALS and 345 Dutch controls., Results: In patients with ALS we identified 11 mutations, 5 of which were predicted to affect PGRN protein sequence or levels (four missense mutations and one 5' regulatory variant). Moreover, common variants (rs9897526, rs34424835, and rs850713) and haplotypes were significantly associated with a reduction in age at onset and a shorter survival after onset of ALS in both the Belgian and the Dutch studies., Conclusion: PGRN acts as a modifier of the course of disease in patients with amyotrophic lateral sclerosis, through earlier onset and shorter survival.

The aim of the experiment was to study whether bovine herpesvirus 1 (BHV1) marker vaccine batches known to be contaminated with bovine virus diarrhoea virus (BVDV) type 1 could cause BVD in cattle. For this purpose, four groups of cattle were used. The first group (n = 4 calves, the positive control group), was vaccinated with vaccine from a batch contaminated with BVDV type 2. The second group (n = 4 calves, the negative control group), was vaccinated with vaccine from a batch that was not contaminated with BVDV. The third group (n = 39 calves), was vaccinated with a vaccine from one of four batches contaminated with BVDV type 1 (seronegative experimental group). The fourth group (n = 6 seropositive heifers), was vaccinated with a vaccine from one of three batches known to be contaminated with BVDV type 1. All cattle were vaccinated with an overdose of the BHV1 marker vaccine. At the start of the experiment, all calves except those from group 4 were seronegative for BVDV and BHV1. The calves from group 4 had antibodies against BVDV, were BVDV-free and seronegative to BHV1. After vaccination, the positive control calves became severely ill, had fever for several days, and BVDV was isolated from nasal swabs and white blood cells. In addition, these calves produced antibodies to BVDV and BHV1. No difference in clinical scores of the other groups was seen, nor were BVDV or BVDV-specific antibody responses detected in these calves; however, they did produce antibodies against BHV1. The remainder of each vaccine vial used was examined for the presence of infectious BVDV in cell culture. From none of the vials was BVDV isolated after three subsequent passages. This indicates that BVDV was either absent from the vials or was present in too low an amount to be isolated. Thus vaccination of calves with vaccines from BHV1 marker vaccine batches contaminated with BVDV type 1 did not result in BVDV infections.

We measured energy expenditure with the doubly labeled water technique during heavy sustained exercise in the Tour de France, a bicycle race lasting more than 3 wk. Four subjects were observed for consecutive intervals of 7, 8, and 7 days. Each interval started with an oral isotope dose to reach an excess isotope level of 200 ppm 18O and 130 ppm 2H. The biological half-lives of the isotopes were between 2.25 and 3.80 days. Energy expenditure was compared with simultaneous measurements of energy intake, and body mass and body composition did not change significantly. The doubly labeled water technique gave higher values for energy expenditure than the food record technique. The discrepancy showed a systematic increment from the first to the third interval, being 12.9 +/- 7.9, 21.4 +/- 9.8, and 35.3 +/- 4.4% of the energy expenditure calculated from dietary intake, respectively. Possible explanations for the discrepancy are discussed. The subjects reached an average daily metabolic rate of 3.4-3.9 or 4.3-5.3 times basal metabolic rate based on the food record technique and the doubly labeled water technique, respectively. Thus, when measured with the same technique, the energetic ceiling for performance in humans is comparable with that of animals like birds.