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White matter (WM) atrophy is a significant feature of Huntington disease (HD), although its aetiology and early pathological manifestations remain poorly defined. In this study, we aimed to characterize WM-related features in the transgenic YAC128 and BACHD models of HD. Using diffusion tensor magnetic resonance imaging (DT-MRI), we demonstrate that microstructural WM abnormalities occur from an early age in YAC128 mice. Similarly, electron microscopy analysis of myelinated fibres of the corpus callosum indicated that myelin sheaths are thinner in YAC128 mice as early as 1.5 months of age, well before any neuronal loss can be detected. Transcript levels of myelin-related genes in striatal and cortical tissues were significantly lower in YAC128 mice from 2 weeks of age, and these findings were replicated in differentiated primary oligodendrocytes from YAC128 mice, suggesting a possible mechanistic explanation for the observed structural deficits. Concordant with these observations, we demonstrate reduced expression of myelin-related genes at 3 months of age and WM microstructural abnormalities using DT-MRI at 12 months of age in the BACHD rats. These findings indicate that WM deficits in HD are an early phenotype associated with cell-intrinsic effects of mutant huntingtin on myelin-related transcripts in oligodendrocytes, and raise the possibility that WM abnormalities may be an early contributing factor to the pathogenesis of HD., (© The Author 2016. Published by Oxford University Press.)

Olesoxime, a small molecule drug candidate, has recently attracted attention due to its significant beneficial effects in models of several neurodegenerative disorders including Huntington's disease. Olesoxime's neuroprotective effects have been assumed to be conveyed through a direct, positive influence on mitochondrial function. In a long-term treatment study in BACHD rats, the latest rat model of Huntington's disease, olesoxime revealed a positive influence on mitochondrial function and improved specific behavioral and neuropathological phenotypes. Moreover, a novel target of the compound was discovered, as olesoxime was found to suppress the activation of the calpain proteolytic system, a major contributor to the cleavage of the disease-causing mutant huntingtin protein into toxic fragments, and key player in degenerative processes in general. Results from a second model of Huntington's disease, the Hdh (Q111) knock-in mouse, confirm olesoxime's calpain-suppressing effects and support the therapeutic value of olesoxime for Huntington's disease and other disorders involving calpain overactivation.

Impulsivity trait was characterized in 3-5 months old BACHD rats, a transgenic model of Huntington disease, using (1) the delay discounting task to assess cognitive/choice impulsivity, and (2) the Differential Reinforcement of Low Rate of Responding task to evaluate motor/action impulsivity. Transgenic animals showed a high level of choice impulsivity and, to a lesser extent, action impulsivity. Our results provide the first evidence that the transgenic BACHD rat (TG5 line) displays impulsivity disorder as early as 3 months old, as described in early symptomatic HD patients, thus adding to the face validity of the rat model., (Copyright © 2015 Elsevier B.V. All rights reserved.)

Parkinson's disease (PD) is a multisystem disorder, involving several monoaminergic neurotransmitter systems resulting in a broad range of motor and non-motor symptoms. Pathological hallmarks of PD are the loss of dopaminergic neurons and the accumulation of alpha-synuclein, however also being present in the serotonergic raphe nuclei early in the disease course. The dysfunction of the serotonergic system projecting to the hippocampus may contribute to early non-motor symptoms such as anxiety and depression. The adult hippocampal dentate gyrus (DG), a unique niche of the forebrain continuously generating new neurons, may particularly present enhanced susceptibility towards accumulating alpha-synuclein levels. The underlying molecular mechanisms in the context of neuronal maturation and survival of new-born neurons are yet not well understood. To characterize the effects of overexpression of human full-length alpha-synuclein on hippocampal cellular and synaptic plasticity, we used a recently generated BAC alpha-synuclein transgenic rat model showing important features of PD such as widespread and progressive alpha-synuclein aggregation pathology, dopamine loss and age-dependent motor decline. At the age of four months, thus prior to the occurrence of the motor phenotype, we observed a profoundly impaired dendritogenesis of neuroblasts in the hippocampal DG resulting in severely reduced survival of adult new-born neurons. Diminished neurogenesis concurred with a serotonergic deficit in the hippocampus as defined by reduced levels of serotonin (5-HT) 1B receptor, decreased 5-HT neurotransmitter levels, and a loss of serotonergic nerve terminals innervating the DG/CA3 subfield, while the number of serotonergic neurons in the raphe nuclei remained unchanged. Moreover, alpha-synuclein overexpression reduced proteins involved in vesicle release, in particular synapsin-1 and Rab3 interacting molecule (RIM3), in conjunction with an altered ultrastructural architecture of hippocampal synapses. Importantly, BAC alpha-synuclein rats showed an early anxiety-like phenotype consisting of reduced exploratory behavior and feeding. Taken together, these findings imply that accumulating alpha-synuclein severely affects hippocampal neurogenesis paralleled by impaired 5-HT neurotransmission prior to the onset of aggregation pathology and overt motor deficits in this transgenic rat model of PD., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Huntington's disease is a fatal human neurodegenerative disorder caused by a CAG repeat expansion in the HTT gene, which translates into a mutant huntingtin protein. A key event in the molecular pathogenesis of Huntington's disease is the proteolytic cleavage of mutant huntingtin, leading to the accumulation of toxic protein fragments. Mutant huntingtin cleavage has been linked to the overactivation of proteases due to mitochondrial dysfunction and calcium derangements. Here, we investigated the therapeutic potential of olesoxime, a mitochondria-targeting, neuroprotective compound, in the BACHD rat model of Huntington's disease. BACHD rats were treated with olesoxime via the food for 12 months. In vivo analysis covered motor impairments, cognitive deficits, mood disturbances and brain atrophy. Ex vivo analyses addressed olesoxime's effect on mutant huntingtin aggregation and cleavage, as well as brain mitochondria function. Olesoxime improved cognitive and psychiatric phenotypes, and ameliorated cortical thinning in the BACHD rat. The treatment reduced cerebral mutant huntingtin aggregates and nuclear accumulation. Further analysis revealed a cortex-specific overactivation of calpain in untreated BACHD rats. Treated BACHD rats instead showed significantly reduced levels of mutant huntingtin fragments due to the suppression of calpain-mediated cleavage. In addition, olesoxime reduced the amount of mutant huntingtin fragments associated with mitochondria, restored a respiration deficit, and enhanced the expression of fusion and outer-membrane transport proteins. In conclusion, we discovered the calpain proteolytic system, a key player in Huntington's disease and other neurodegenerative disorders, as a target of olesoxime. Our findings suggest that olesoxime exerts its beneficial effects by improving mitochondrial function, which results in reduced calpain activation. The observed alleviation of behavioural and neuropathological phenotypes encourages further investigations on the use of olesoxime as a therapeutic for Huntington's disease., (© The Author (2015). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

X-linked intellectual disability (XLID) is a genetically heterogeneous disorder with more than 100 genes known to date. Most genes are responsible for a small proportion of patients only, which has hitherto hampered the systematic screening of large patient cohorts. We performed targeted enrichment and next-generation sequencing of 107 XLID genes in a cohort of 150 male patients. Hundred patients had sporadic intellectual disability, and 50 patients had a family history suggestive of XLID. We also analysed a sporadic female patient with severe ID and epilepsy because she had strongly skewed X-inactivation. Target enrichment and high parallel sequencing allowed a diagnostic coverage of >10 reads for ~96% of all coding bases of the XLID genes at a mean coverage of 124 reads. We found 18 pathogenic variants in 13 XLID genes (AP1S2, ATRX, CUL4B, DLG3, IQSEC2, KDM5C, MED12, OPHN1, SLC9A6, SMC1A, UBE2A, UPF3B and ZDHHC9) among the 150 male patients. Thirteen pathogenic variants were present in the group of 50 familial patients (26%), and 5 pathogenic variants among the 100 sporadic patients (5%). Systematic gene dosage analysis for low coverage exons detected one pathogenic hemizygous deletion. An IQSEC2 nonsense variant was detected in the female ID patient, providing further evidence for a role of this gene in encephalopathy in females. Skewed X-inactivation was more frequently observed in mothers with pathogenic variants compared with those without known X-linked defects. The mutation rate in the cohort of sporadic patients corroborates previous estimates of 5-10% for X-chromosomal defects in male ID patients.

Executive dysfunction and psychiatric symptoms are hallmarks of Huntington disease (HD), a neurodegenerative disorder genetically characterized by expanded CAG repeats in the HTT gene. Using the BACHD rat model of HD (97 CAG-CAA repeats), the present research seeks to characterize the progressive emergence of decision-making impairments in a rat version of the Iowa Gambling Task (RGT) and the impact of emotional modulation, whether positive or negative, on choice behavior. The choice efficiency shown both by WT rats (independent of their age) and the youngest BACHD rats (2 and 8months old) evidenced that they are able to integrate outcomes of past decisions to determine expected reward values for each option. However, 18months old BACHD rats made fewer choices during the RGT session and were less efficient in choosing advantageous options than younger animals. Presenting either chocolate pellets or electrical footshocks half-way through a second RGT session reduced exploratory activity (inefficient nose-poking) and choices with a weaker effect on BACHD animals than on WT. Choice efficiency was left intact in transgenic rats. Our results bring new knowledge on executive impairments and impact of emotional state on decision-making at different stages of the disease, increasing the face-validity of the BACHD rat model., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Together with point mutations, homozygous deletions or duplications in PARK2 are responsible for the majority of autosomal recessive juvenile Parkinsonism. It is debated, however, whether heterozygous carriers of these mutations are at increased risk of Parkinson's disease (PD). Our goal was to determine whether heterozygous carriers of copy number variants (CNVs) affecting exons of the PARK2 gene are at risk of PD that is greater than that of non-carriers. We searched for CNVs affecting exons of PARK2 in a sample of 105 749 genotyped Icelanders. In total, 989 carriers, including 24 diagnosed with PD, were identified. The heterozygous carriers were tested for association in a sample of 1415 PD patients and 40 474 controls ≥65 years of age. PD patients were more often heterozygous carriers of PARK2 CNVs than controls [odds ratio (OR) = 1.69, P = 0.03] and compound heterozygous PD patients for a CNV and a missense mutation were not found. Furthermore, we conducted a meta-analysis of studies reporting on case-control samples screened for heterozygous PARK2 CNVs. Ten studies were included in the final analysis, with 4538 cases and 4213 controls. The pooled OR and P-value for the published and Icelandic results showed significant association between PARK2 CNVs and risk of PD (OR = 2.11, P = 2.54 × 10(-6)). Our analysis shows that heterozygous carriers of CNVs affecting exons of PARK2 have greater risk of PD than non-carriers., (© The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: Dental implants have been widely used to replace missing teeth, accomplishing aesthetics and function. Due to its large use worldwide, the small percentage of implant loss becomes significant in number of cases. Lactotransferrin (LTF) is a pleiotropic protein, expressed in various body tissues and fluids, which modulates the host immune-inflammatory response and bone metabolism, and might be involved in dental implant osseointegration. Recently, a few studies have been investigating genetic aspects underlying dental implant failure., Purpose: This case-control study aimed to investigate the association of genetic markers (tag SNPs) in LTF gene and clinical parameters with dental implant loss., Material and Methods: 278 patients, both sexes, mean age 51 years old, divided into 184 without and 94 with implant loss, were genotyped for sixteen tag SNPs, representative of the whole LTF gene. Also, clinical oral and systemic parameters were analyzed. Univariate and Multivariate Logistic Regression model were used to analyze the results (p < .05)., Results: No association was found between the tag SNPs and implant loss in the study population. Clinical association was found with medical treatment, hormonal reposition, edentulism, number of placed implants, plaque, calculus, and mobility., Conclusion: Clinical variables, but not LTF gene polymorphisms, were associated with implant loss., (© 2014 Wiley Periodicals, Inc.)

As the second most frequent neurodegenerative disorder of the aging population, Parkinson's disease (PD) is characterized by progressive deficits in spontaneous movement, atrophy of dopaminergic midbrain neurons and aggregation of the protein alpha-synuclein (SNCA). To elucidate molecular events before irreversible cell death, we studied synucleinopathy-induced expression changes in mouse brain and identified 49 midbrain/brainstem-specific transcriptional dysregulations. In particular complexin-1 (Cplx1), Rabl2a and 14-3-3epsilon (Ywhae) downregulation, as well as upregulation of the midbrain-specific factor forkhead box P1 (Foxp1) and of Rabgef1, were interesting as early mRNA level effects of alpha-synuclein triggered pathology. The protein levels of complexin-1 were elevated in midbrain/brainstem tissue of mice with A53T-SNCA overexpression and of mice with SNCA-knockout. The response of CPLX1 and Foxp1 levels to SNCA deficiency supports the notion that these factors are regulated by altered physiological function of alpha-synuclein. Thus, their analysis might be useful in PD stages before the advent of Lewy pathology. Because both alpha-synuclein and complexin-1 modulate vesicle release, our findings support presynaptic dysfunction as an early event in PD pathology.

Multi-gene panels are used to identify genetic causes of hereditary breast and ovarian cancer (HBOC) in large patient cohorts. This study compares the diagnostic workflow in two centers and gives valuable insights into different next-generation sequencing (NGS) strategies. Moreover, we present data from 620 patients sequenced at both centers. Both sequencing centers are part of the German consortium for hereditary breast and ovarian cancer (GC-HBOC). All 620 patients included in this study were selected following standard BRCA1/2 testing guidelines. A set of 10 sequenced genes was analyzed per patient. Twelve samples were exchanged and sequenced at both centers. NGS results were highly concordant in 12 exchanged samples (205/206 variants = 99.51 %). One non-pathogenic variant was missed at center B due to a sequencing gap (no technical coverage). The custom enrichment at center B was optimized during this study; for example, the average number of missing bases was reduced by a factor of four (vers. 1: 1939.41, vers. 4: 506.01 bp). There were no sequencing gaps at center A, but four CCDS exons were not included in the enrichment. Pathogenic mutations were found in 12.10 % (75/620) of all patients: 4.84 % (30/620) in BRCA1, 4.35 % in BRCA2 (27/620), 0.97 % in CHEK2 (6/620), 0.65 % in ATM (4/620), 0.48 % in CDH1 (3/620), 0.32 % in PALB2 (2/620), 0.32 % in NBN (2/620), and 0.16 % in TP53 (1/620). NGS diagnostics for HBOC-related genes is robust, cost effective, and the method of choice for genetic testing in large cohorts. Adding 8 genes to standard BRCA1- and BRCA2-testing increased the mutation detection rate by one-third.

There is a growing international agreement on the need to provide greater access to research data and bio-specimen collections to optimize their long-term value and exploit their potential for health discovery and validation. This is especially evident for rare disease research. Currently, the rising value of data and bio-specimen collections does not correspond with an equal increase in data/sample-sharing and data/sample access. Contradictory legal and ethical frameworks across national borders are obstacles to effective sharing: more specifically, the absence of an integrated model proves to be a major logistical obstruction. The Charter intends to amend the obstacle by providing both the ethical foundations on which data sharing should be based, as well as a general Material and Data Transfer Agreement (MTA/DTA). This Charter is the result of a careful negotiation of different stakeholders' interest and is built on earlier consensus documents and position statements, which provided the general international legal framework. Further to this, the Charter provides tools that may help accelerate sharing. The Charter has been formulated to serve as an enabling tool for effective and transparent data and bio-specimen sharing and the general MTA/DTA constitutes a mechanism to ensure uniformity of access across projects and countries, and may be regarded as a consistent basic agreement for addressing data and material sharing globally. The Charter is forward looking in terms of emerging issues from the perspective of a multi-stakeholder group, and where possible, provides strategies that may address these issues.

Huntington's disease (HD) is an inherited progressive neurodegenerative disorder caused by an expanded CAG repeat in exon 1 of the huntingtin gene (HTT). The primary neuropathology of HD has been attributed to the preferential degeneration of medium spiny neurons (MSN) in the striatum. Reports on striatal neurogenesis have been a subject of debate; nevertheless, it should be considered as an endogenous attempt to repair the brain. The subventricular zone (SVZ) might offer a close-by region to supply the degenerated striatum with new cells. Previously, we have demonstrated that R6/2 mice, a widely used preclinical model representing an early onset HD, showed reduced olfactory bulb (OB) neurogenesis but induced striatal migration of neuroblasts without affecting the proliferation of neural progenitor cell (NPCs) in the SVZ. The present study revisits these findings, using a clinically more relevant transgenic rat model of late onset HD (tgHD rats) carrying the human HTT gene with 51 CAG repeats and mimicking many of the neuropathological features of HD seen in patients. We demonstrate that cell proliferation is reduced in the SVZ and OB of tgHD rats compared to WT rats. In the OB of tgHD rats, although cell survival was reduced, the frequency of neuronal differentiation was not altered in the granule cell layer (GCL) compared to the WT rats. However, an increased frequency of dopamenergic neuronal differentiation was noticed in the glomerular layer (GLOM) of tgHD rats. Besides this, we observed a selective proliferation of neuroblasts in the adjacent striatum of tgHD rats. There was no evidence for neuronal maturation and survival of these striatal neuroblasts. Therefore, the functional role of these invading neuroblasts still needs to be determined, but they might offer an endogenous alternative for stem or neuronal cell transplantation strategies.

Objective: To find a potential underlying cause for Mayer-Rokitansky-Küster-Hauser syndrome (MRKHS) discordance in monozygotic twins., Design: Prospective comparative study., Setting: University hospital., Patient(s): Our study genetically analyzed 5 MRKHS-discordant monozygotic twin pairs with the unique opportunity to include saliva and rudimentary uterine tissue., Intervention(s): Blood, saliva, or rudimentary uterine tissue from five MRKHS-discordant twins was analyzed and compared between twin pairs as well as within the same individual where applicable. We used copy number variations (CNVs) to identify differences., Main Outcome Measure(s): CNVs in blood, rudimentary uterine tissue, and saliva, network analysis, and review of the literature., Result(s): One duplication found in the affected twin included two genes, matrix metalloproteinase 14 (MMP14) and low-density lipoprotein receptor-related protein 10 (LRP10), which have known functions in the embryonic development of the uterus and endometrium. The duplicated region was detected in rudimentary uterine tissue from the same individual but not in saliva, making a tissue-specific mosaicism a possible explanation for twin discordance. Additional network analysis revealed important connections to differentially expressed genes from previous studies. These genes encode several molecules involved in extracellular matrix (ECM) remodeling and neoangiogenesis., Conclusion(s): MMP-14, LRP-10, ECM, and neoangiogenesis genes are identified as candidate genes in a tissue-specific mosaicism. The detected clusters provide evidence of deficient vascularization during uterine development and/or disturbed reorganization of ECM components, potentially during müllerian duct elongation signaled by the embryologically relevant phosphatidylinositol 3-kinase/protein kinase B pathway. Therefore, we consider these genes to be new candidates in the manifestation of MRKHS., (Copyright © 2015 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.)

Background: FDG-PET detects hypometabolism in premanifest and symptomatic Huntington's disease (HD). A cross-sectional study suggested that whole-brain FDG-PET is capable to detect a phenotype in transgenic (tg) HD rats. Recently, a longitudinal follow-up study showed no FDG-PET changes in tgHD rats. Both studies applied small sample sizes and analysis was limited to whole-brain or striatum., Objective: We therefore performed a follow-up study in a larger cohort of tgHD and wild-type (wt) rats encompassing several pre-defined regions of interest (ROIs) and hypothesis free voxel-by-voxel SPM analysis to clarify whether FDG-PET can detect a phenotype in tgHD rats and to determine onset …and effect sizes of changes over time., Methods: N = 19 tgHD- and n = 20 wt-rats, mixed gender, were included. Repeated small animal FDG-μPET and MRI were performed at 5,10,15, and 20 months of age. ROIs encompassing entire brain, cortex, striatum, thalamus, subventricular-zone, and cerebellum were placed manually on the MRI and transferred to the co-registered μPET. Mean and maximal FDG-PET activities within ROIs were calculated and normalized to cerebellar FDG uptake. Activity and spatially normalized FDG-μPET were compared between groups on a hypothesis-free voxel-by-voxel basis using SPM., Results: FDG uptake showed changes over time in both tgHD- and wt-rats, however, there was no consistent difference between tgHD- and wt-rats in both the manual ROI and SPM analysis., Conclusions: In this transgenic rat model of HD FDG-μPET imaging does not detect significant alterations at the ages investigated. Further investigations are warranted employing other age groups and alternative imaging biomarkers for neuronal degeneration, respectively.

Background: Missense mutations in the eukaryotic translation initiation factor 4-γ 1 (EIF4G1) gene have previously been implicated in familial Parkinson's disease (PD). A large PD family with autosomal-dominant segregation showed a heterozygous missense mutation and additional patients were found to have unique sequence variants that have not been observed in controls. Subsequent studies have reported contradictory findings., Methods: We assessed the relevance of EIF4G1 mutations in a European cohort of 2146 PD patients. Of these, 2051 sporadic PD patients were screened for the reported p.Ala502Val and p.Arg1205His mutations. In addition, the complete coding region of EIF4G1 was directly sequenced in 95 familial PD patients with autosomal-dominant inheritance. Moreover, we imputed the p.Arg1205His substitution and tested for association with PD in the Icelandic population (93 698 samples)., Results: We did not observe the presence of the p.Ala502Val substitution in our cohort; however, the p.Arg1205His mutation was identified in one sporadic PD patient. The same mutation was also found in 76 Icelandic subjects older than 65 years using haplotype imputing. Only five of these subjects reported PD symptoms (OR 1.3, p=0.50). Thus, if causal, the p.Arg1205His EIF4G1 mutation has a low penetrance or a late onset manifestation. A novel variant p.Arg566Cys found in a patient with familial PD did not cosegregate with PD in all three affected siblings. All further recently published EIF4G1 mutations found in our cohort are likely to be benign polymorphisms., Conclusions: This is the largest genetic study of EIF4G1 mutations in PD. Our data do not support the EIF4G1 gene as a high-risk PD locus, neither for the familial nor the sporadic condition. Furthermore, the p.Arg1205His mutation is not significantly associated with increased risk of PD in the Icelandic population. Therefore, caution should be exercised when interpreting EIF4G1 genotyping results in isolated patients and PD families. In summary, diagnostic testing of EIF4G1 should not be recommended in clinical settings., (Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://group.bmj.com/group/rights-licensing/permissions.)

Dental caries is a common multifactorial disease, resulting from the interaction of biofilm, cariogenic diet and host response over time. Lactotransferrin (LTF) is a main salivary glycoprotein, which modulates the host immune-inflammatory and antibacterial response. Although a genetic component for caries outcome has been identified, little is known over the genetic aspects underlying its susceptibility. Thus, the aim of this study was to investigate the association between LTF polymorphisms and caries susceptibility. Six hundred seventy seven 12-year-old students were selected: 346 with (DMFT ≥ 1) and 331 without caries experience (DMFT = 0). Also, individuals concentrating higher levels of disease (polarization group, DMFT ≥ 2, n = 253) were tested against those with DMFT ≤ 1 (n = 424). Along with clinical parameters, three representative LTF tag SNPs (rs6441989, rs2073495, rs11716497) were genotyped and the results were evaluated using univariate and multivariate analyses. Allele A for tag SNP rs6441989 was found to be significantly less frequent in the polarization group, conferring a protective effect against caries experience [AA + AG × GG (OR: 0.710, 95% CI: 0.514-0.980, p = 0.045)], and remained significantly associated with caries protection in the presence of gingivitis (p = 0.020) and plaque (p = 0.035). These results might contribute to the understanding of the genetic control of caries susceptibility in humans., (© 2015 S. Karger AG, Basel.)

Uniparental disomy (UPD) describes the inheritance of a pair of chromosomes from only one parent. It may occur as isodisomy, heterodisomy or a combination of both and may involve only chromosome segments. UPD can affect each chromosome. The incidence is estimated to be around 1:3500 in live births. Some parts of chromosomes are subject to 'parent-of-origin imprinting' and the phenotypic effect in UPD syndromes is mainly due to functional imbalance of imprinted genes. Isodisomy can result in mutation homozygosity in autosomal-recessive inherited diseases. UPD causes several well-defined imprinting syndromes associated with intellectual disability (ID). Although knowledge on frequency and size of UPDs in patients with unexplained ID remains largely unknown as no efficient genome-wide screening technique was available for detection of both isodisomic and heterodisomic UPDs. SNP microarrays have been proven to be capable to detect UPDs through Mendelian errors. The correct subclassification of UPD requires child-parent trio experiments. To further elucidate the role of UPD in patients with unexplained ID, we analyzed a total of 322 child-parent trios. We were not able to detect UPDs (isodisomies and heterodisomies) within our cohort spanning whole chromosomes or chromosomal segments. We conclude that UPD is rare in patients with unexplained ID.

Polyglutamine-coding (CAG)n repeat expansions in seven different genes cause spinocerebellar ataxias. Although the size of the expansion is negatively correlated with age at onset, it accounts for only 50-70% of its variability. To find other factors involved in this variability, we performed a regression analysis in 1255 affected individuals with identified expansions (spinocerebellar ataxia types 1, 2, 3, 6 and 7), recruited through the European Consortium on Spinocerebellar Ataxias, to determine whether age at onset is influenced by the size of the normal allele in eight causal (CAG)n-containing genes (ATXN1-3, 6-7, 17, ATN1 and HTT). We confirmed the negative effect of the expanded allele and detected threshold effects reflected by a quadratic association between age at onset and CAG size in spinocerebellar ataxia types 1, 3 and 6. We also evidenced an interaction between the expanded and normal alleles in trans in individuals with spinocerebellar ataxia types 1, 6 and 7. Except for individuals with spinocerebellar ataxia type 1, age at onset was also influenced by other (CAG)n-containing genes: ATXN7 in spinocerebellar ataxia type 2; ATXN2, ATN1 and HTT in spinocerebellar ataxia type 3; ATXN1 and ATXN3 in spinocerebellar ataxia type 6; and ATXN3 and TBP in spinocerebellar ataxia type 7. This suggests that there are biological relationships among these genes. The results were partially replicated in four independent populations representing 460 Caucasians and 216 Asian samples; the differences are possibly explained by ethnic or geographical differences. As the variability in age at onset is not completely explained by the effects of the causative and modifier sister genes, other genetic or environmental factors must also play a role in these diseases., (© The Author (2014). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

SHORT syndrome (OMIM 269880) is a rare autosomal-dominant disorder characterized by short stature, hyperextensibility of joints, hernias, ocular depression, ophthalmic anomalies (Rieger anomaly, posterior embryotoxon, glaucoma), teething delay, partial lipodystrophy, insulin resistance and facial dysmorphic signs. Heterozygous mutations in PIK3R1 were recently identified in 14 families with SHORT syndrome. Eight of these families had a recurrent missense mutation (c.1945C>T; p.Arg649Trp). We report on two unrelated patients with typical clinical features of SHORT syndrome and additional problems such as pulmonary stenosis and ectopic kidney. Analysis of PIK3R1 revealed the mutation c.1945C>T; p.Arg649Trp de novo in both patients. These two patients not only provide additional evidence that PIK3R1 mutations cause SHORT syndrome, but also broaden the clinical spectrum of this syndrome and further confirm that the amino acid exchange c.1945C>T; p.Arg649Trp is a hotspot mutation in this gene., (© 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Hereditary spastic paraplegias (HSP) constitute a rare and highly heterogeneous group of neurodegenerative disorders, defined clinically by progressive lower limb spasticity and pyramidal weakness. Autosomal recessive HSP as well as sporadic cases present a significant diagnostic challenge. Mutations in AP5Z1, a gene playing a role in intracellular membrane trafficking, have been recently reported to be associated with spastic paraplegia type 48 (SPG48). Our objective was to determine the relative frequency and clinical relevance of AP5Z1 mutations in a large cohort of 127 HSP patients. We applied a targeted next-generation sequencing approach to analyze all coding exons of the AP5Z1 gene. With the output of high-quality reads and a mean coverage of 51-fold, we demonstrated a robust detection of variants. One 43-year-old female with sporadic complicated paraplegia showed two heterozygous nonsynonymous variants of unknown significance (VUS3; p.[R292W];[(T756I)]). Thus, AP5Z1 gene mutations are rare, at least in Europeans. Due to its low frequency, systematic genetic testing for AP5Z1 mutations is not recommended until larger studies are performed to add further evidence. Our findings demonstrate that amplicon-based deep sequencing is technically feasible and allows a compact molecular characterization of multiple HSP patients with high accuracy.

Background: The need for improving throughput, validity, and reliability in the behavioral characterization of rodents may benefit from integrating automated intra-home-cage-screening systems allowing the simultaneous detection of multiple behavioral and physiological parameters in parallel., New Method: To test this hypothesis, transgenic Huntington's disease (tgHD) rats were repeatedly screened within phenotyping home-cages (PhenoMaster and IntelliCage for rats), where spontaneous activity, feeding, drinking, temperature, and metabolic performance were continuously measured. Cognition and emotionality were evaluated within the same environment by means of operant learning procedures and refined analysis of the behavioral display under conditions of novelty. This investigator-independent approach was further correlated with behavioral display of the animals in classical behavioral assays. Multivariate analysis (MVA) including Principle Component Analysis (PCA) and Partial Least Squares Discriminant Analysis (PLS-DA) was used to explore correlation patterns of variables within and across the two genotypes., Results: The automated systems traced previously undetected aspects in the phenotype of tgHD rats (circadian activity, energy metabolism, rearing), and out of those spontaneous free rearing correlated with individual performance in the accelerod test. PCA revealed a segregation by genotype in juvenile tgHD rats that differed from adult animals, being further resolved by PLS-DA detecting "temperature" (juvenile) and "rearing" (adult) as phenotypic key variables in the tgHD model., Conclusions: Intra-home-cage phenotyping in combination with MVA, is capable of characterizing a complex phenotype by detecting novel physiological and behavioral markers with high sensitivity and standardization using fewer human resources. A broader application of automated systems for large-scale screening is encouraged., (Copyright © 2014. Published by Elsevier B.V.)

Huntington disease (HD) is an inherited neurodegenerative disease characterized by motor, cognitive, psychiatric and metabolic symptoms. Animal models of HD show phenotypes that can be divided into similar categories, with the metabolic phenotype of certain models being characterized by obesity. Although interesting in terms of modeling metabolic symptoms of HD, the obesity phenotype can be problematic as it might confound the results of certain behavioral tests. This concerns the assessment of cognitive function in particular, as tests for such phenotypes are often based on food depriving the animals and having them perform tasks for food rewards. The BACHD rat is a recently established animal model of HD, and in order to ensure that behavioral characterization of these rats is done in a reliable way, a basic understanding of their physiology is needed. Here, we show that BACHD rats are obese and suffer from discrete developmental deficits. When assessing the motivation to lever push for a food reward, BACHD rats were found to be less motivated than wild type rats, although this phenotype was dependent on the food deprivation strategy. Specifically, the phenotype was present when rats of both genotypes were deprived to 85% of their respective free-feeding body weight, but not when deprivation levels were adjusted in order to match the rats' apparent hunger levels. The study emphasizes the importance of considering metabolic abnormalities as a confounding factor when performing behavioral characterization of HD animal models.

Lewy bodies, a pathological hallmark of Parkinson's disease (PD), contain aggregated alpha-synuclein (αSyn), which is found in several modified forms and can be discovered phosphorylated, ubiquitinated and truncated. Aggregation-prone truncated species of αSyn caused by aberrant cleavage of this fibrillogenic protein are hypothesized to participate in its sequestration into inclusions subsequently leading to synaptic dysfunction and neuronal death. Here, we investigated the role of calpain cleavage of αSyn in vivo by generating two opposing mouse models. We crossed into human [A30P]αSyn transgenic (i) mice deficient for calpastatin, a calpain-specific inhibitor, thus enhancing calpain activity (SynCAST(-)) and (ii) mice overexpressing human calpastatin leading to reduced calpain activity (SynCAST(+)). As anticipated, a reduced calpain activity led to a decreased number of αSyn-positive aggregates, whereas loss of calpastatin led to increased truncation of αSyn in SynCAST(-). Furthermore, overexpression of calpastatin decreased astrogliosis and the calpain-dependent degradation of synaptic proteins, potentially ameliorating the observed neuropathology in [A30P]αSyn and SynCAST(+) mice. Overall, our data further support a crucial role of calpains, particularly of calpain 1, in the pathogenesis of PD and in disease-associated aggregation of αSyn, indicating a therapeutic potential of calpain inhibition in PD., (© The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Background: GNAL mutations have been shown to cause adult-onset isolated dystonia, a disabling movement disorder characterized by involuntary muscle contractions causing twisting and repetitive movements or abnormal postures., Methods: To test the frequency of GNAL mutations in a series of 137 German patients with sporadic dystonia patients we used next-generation sequencing of amplicon-derived barcoded NexteraXT libraries for the coding exons and adjacent intronic sequences of GNAL., Results: In our cohort we identified 1 pathogenic nonsense mutation (c.733C>T, p.R245*) in a patient with cervical dystonia. In a second patient a synonymous coding nonsynonymous variant (c.G252A, p.E84E) was detected, which is predicted to alter a splice site., Conclusions: Our findings further support GNAL as causative gene in adult-onset isolated dystonia., (© 2014 International Parkinson and Movement Disorder Society.)

The mitochondrial chaperone mortalin was implicated in Parkinson's disease (PD) because of its reduced levels in the brains of PD patients and disease-associated rare genetic variants that failed to rescue impaired mitochondrial integrity in cellular knockdown models. To uncover the molecular mechanisms underlying mortalin-related neurodegeneration, we dissected the cellular surveillance mechanisms related to mitochondrial quality control, defined the effects of reduced mortalin function at the molecular and cellular levels and investigated the functional interaction of mortalin with Parkin and PINK1, two PD-related proteins involved in mitochondrial homeostasis. We found that reduced mortalin function leads to: (1) activation of the mitochondrial unfolded protein response (UPR(mt)), (2) increased susceptibility towards intramitochondrial proteolytic stress, (3) increased autophagic degradation of fragmented mitochondria and (4) reduced mitochondrial mass in human cells in vitro and ex vivo. These alterations caused increased vulnerability toward apoptotic cell death. Proteotoxic perturbations induced by either partial loss of mortalin or chemical induction were rescued by complementation with native mortalin, but not disease-associated mortalin variants, and were independent of the integrity of autophagic pathways. However, Parkin and PINK1 rescued loss of mortalin phenotypes via increased lysosomal-mediated mitochondrial clearance and required intact autophagic machinery. Our results on loss of mortalin function reveal a direct link between impaired mitochondrial proteostasis, UPR(mt) and PD and show that effective removal of dysfunctional mitochondria via either genetic (PINK1 and Parkin overexpression) or pharmacological intervention (rapamycin) may compensate mitochondrial phenotypes.

Intensive scientific research devoted in the recent years to understand the molecular mechanisms or neurodegeneration in spinocerebellar ataxias (SCAs) are identifying new pathways and targets providing new insights and a better understanding of the molecular pathogenesis in these diseases. In this consensus manuscript, the authors discuss their current views on the identified molecular processes causing or modulating the neurodegenerative phenotype in spinocerebellar ataxias with the common opinion of translating the new knowledge acquired into candidate targets for therapy. The following topics are discussed: transcription dysregulation, protein aggregation, autophagy, ion channels, the role of mitochondria, RNA toxicity, modulators of neurodegeneration and current therapeutic approaches. Overall point of consensus includes the common vision of neurodegeneration in SCAs as a multifactorial, progressive and reversible process, at least in early stages. Specific points of consensus include the role of the dysregulation of protein folding, transcription, bioenergetics, calcium handling and eventual cell death with apoptotic features of neurons during SCA disease progression. Unresolved questions include how the dysregulation of these pathways triggers the onset of symptoms and mediates disease progression since this understanding may allow effective treatments of SCAs within the window of reversibility to prevent early neuronal damage. Common opinions also include the need for clinical detection of early neuronal dysfunction, for more basic research to decipher the early neurodegenerative process in SCAs in order to give rise to new concepts for treatment strategies and for the translation of the results to preclinical studies and, thereafter, in clinical practice.

Background and Objectives: The ataxias are a challenging group of neurological diseases due the aetiological heterogeneity and the complexity of the genetic subtypes. This guideline focuses on the heredodegenerative ataxias. The aim is to provide a peer-reviewed evidence-based guideline for clinical neurologists and other specialist physicians responsible for the care of patients with ataxia., Methods: This guideline is based on systematic evaluations of the relevant literature and on three consensus meetings of the task force., Diagnosis: If acquired causes are ruled out, and if the disease course is rather slowly progressive, a (heredo)degenerative disease is likely. A positive family history gives much guidance. In the case of a dominant family history, first line genetic screening is recommended for spinocerebellar ataxia (SCA) 1, 2, 3, 6, 7 and 17 (level B), and in Asian patients also for dentatorubral-pallidoluysian atrophy (DRPLA). In the case of recessive disease, a stepwise diagnostic work-up is recommended, including both biochemical markers and targeted genetic testing, particularly aimed at Friedreich's ataxia, ataxia telangiectasia, ataxia due to vitamin E deficiency, polymerase gamma gene (POLG gene, various mutations), autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) and ataxia with oculomotor apraxia (AOA) types 1 and 2. If family history is negative, we still advise to screen for the more common dominant and recessive ataxias. In addition, if onset is below 45 years we recommend the full work-up for recessive ataxias; if onset is above 45 years we recommend to screen for fragile X mental retardation 1 FMR1 premutations (good practice points). In sporadic cases with an onset after 30 years, a diagnosis of multiple system atrophy should be considered (good practice point). In particular the genetic work-up will change over the upcoming years due to the diagnostic utility of new techniques such as gene panel diagnostics based on next generation sequencing for routine work-up, or even whole exome and genome sequencing for selected cases., Treatment: Some of the rare recessive ataxias are treatable, but for most of the heredodegenerative ataxias treatment is purely symptomatic. Idebenone is not effective in Friedreich's ataxia (level A). Riluzole (level B) and amantadine (level C) might provide symptomatic relief, irrespective of exact etiology. Also, varenicline for SCA3 patients (level B) can be considered. There is level Class II evidence to recommend physiotherapy, and Class III data to support occupational therapy., (© 2014 The Author(s) European Journal of Neurology © 2014 EFNS.)

Behavioral characterization is an important part of establishing novel animal models, but classical behavioral tests struggle to reveal conclusive results due to problems with both reproducibility and validity. On the contrary, automated homecage observations are believed to produce robust outcomes that relate more to natural animal behavior. However, information on the behavior of background strains from such observations, which could provide important reference material, is rare. For this reason, we compared the behavior of the commonly used Lister Hooded, Lewis, Fischer 344 and Wistar rats during 70 h of exposure to an automated homecage system at 2, 4 and 6 months of age. We found considerable strain differences in metabolic parameters, novelty-induced and baseline activity-related behavior as well as differences in the development of these parameters with age. The results are discussed in terms of advantages and disadvantages of the system compared to classical behavioral tests, as well as the system's ability to recreate common findings in literature., (© 2013 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.)

Lewy bodies and neurites are the pathological hallmark of Parkinson's disease. These structures are composed of fibrillized and ubiquitinated alpha-synuclein suggesting that impaired protein clearance is an important event in aggregate formation. The A30P mutation is known for its fast oligomerization, but slow fibrillization rate. Despite its toxicity to neurons, mechanisms involved in either clearance or conversion of A30P alpha-synuclein from its soluble state into insoluble fibrils and their effects in vivo are poorly understood. Synphilin-1 is present in Lewy bodies, interacting with alpha-synuclein in vivo and in vitro and promotes its sequestration into aggresomes, which are thought to act as cytoprotective agents facilitating protein degradation. We therefore crossed animals overexpressing A30P alpha-synuclein with synphilin-1 transgenic mice to analyze its impact on aggregation, protein clearance and phenotype progression. We observed that co-expression of synphilin-1 mildly delayed the motor phenotype caused by A30P alpha-synuclein. Additionally, the presence of N- and C-terminal truncated alpha-synuclein species and fibrils were strongly reduced in double-transgenic mice when compared with single-transgenic A30P mice. Insolubility of mutant A30P and formation of aggresomes was still detectable in aged double-transgenic mice, paralleled by an increase of ubiquitinated proteins and high autophagic activity. Hence, this study supports the notion that co-expression of synphilin-1 promotes formation of autophagic-susceptible aggresomes and consecutively the degradation of human A30P alpha-synuclein. Notably, although synphilin-1 overexpression significantly reduced formation of fibrils and astrogliosis in aged animals, a similar phenotype is present in single- and double-transgenic mice suggesting additional neurotoxic processes in disease progression.

Background: A major risk-factor for developing Parkinson's disease (PD) is genetic variability in leucine-rich repeat kinase 2 (LRRK2), most notably the p.G2019S mutation. Examination of the effects of this mutation is necessary to determine the etiology of PD and to guide therapeutic development., Objective: Assess the behavioral consequences of LRRK2 p.G2019S overexpression in transgenic rats as they age and test the functional integrity of the nigro-striatal dopamine system. Conduct positron emission tomography (PET) neuroimaging to compare transgenic rats with previous data from human LRRK2 mutation carriers., Methods: Rats overexpressing human LRRK2 p.G2019S were generated by BAC transgenesis and compared to non-transgenic (NT) littermates. Motor skill tests were performed at 3, 6 and 12 months-of-age. PET, performed at 12 months, assessed the density of dopamine and vesicular monoamine transporters (DAT and VMAT2, respectively) and measured dopamine synthesis, storage and availability. Brain tissue was assayed for D2, DAT, dopamine and cAMP-regulated phosphoprotein (DARPP32) and tyrosine hydroxylase (TH) expression by Western blot, and TH by immunohistochemistry., Results: Transgenic rats had no abnormalities in measures of striatal dopamine function at 12 months. A behavioral phenotype was present, with LRRK2 p.G2019S rats performing significantly worse on the rotarod than non-transgenic littermates (26% reduction in average running duration at 6 months), but with normal performance in other motor tests., Conclusions: Neuroimaging using dopaminergic PET did not recapitulate prior studies in human LRRK2 mutation carriers. Consistently, LRRK2 p.G2019S rats do not develop overt neurodegeneration; however, they do exhibit behavioral abnormalities.

Mutations in the leucine-rich repeat kinase 2 (LRRK2) gene represent the most common genetic cause of Parkinson's disease (PD). However, LRRK2 function and molecular mechanisms causing the parkinsonian phenotype remain widely unknown. Most of LRRK2 knockdown and overexpression models strengthen the relevance of LRRK2 in regulating neurite outgrowth. We have recently identified ARHGEF7 as the first guanine nucleotide exchange factor (GEF) of LRRK2. This GEF is influencing neurite outgrowth through regulation of actin polymerization. Here, we examined the expression profile of neuroblastoma cells with reduced LRRK2 and ARHGEF7 levels to identify additional partners of LRRK2 in this process. Tropomyosins (TPMs), and in particular TPM4, were the most interesting candidates next to other actin cytoskeleton regulating transcripts in this dataset. Subsequently, enhanced neurite branching was shown using primary hippocampal neurons of LRRK2 knockdown animals. Furthermore, we observed an enhanced number of growth cones per neuron and a mislocalization and dysregulation of ARHGEF7 and TPM4 in these neuronal compartments. Our results reveal a fascinating connection between the neurite outgrowth phenotype of LRRK2 models and the regulation of actin polymerization directing further investigations of LRRK2-related pathogenesis., (© 2013.)

In Parkinson's disease (PD) patients, alpha-synuclein (α-syn) pathology advances in form of Lewy bodies and Lewy neurites throughout the brain. Clinically, PD is defined by motor symptoms that are predominantly attributed to the dopaminergic cell loss in the substantia nigra. However, motor deficits are frequently preceded by smell deficiency or neuropsychological symptoms, including increased anxiety and cognitive dysfunction. Accumulating evidence indicates that aggregation of α-syn impairs synaptic function and neurogenic capacity that may be associated with deficits in memory, learning and mood. Whether and how α-syn accumulation contributes to neuropathological events defining these earliest signs of PD is presently poorly understood. We used a tetracycline-suppressive (tet-off) transgenic mouse model that restricts overexpression of human A30P α-syn to neurons owing to usage of the neuron-specific CaMKIIα promoter. Abnormal accumulation of A30P correlated with a decreased survival of newly generated neurons in the hippocampus and olfactory bulb. Furthermore, when A30P α-syn expression was suppressed, we observed reduction of the human protein in neuronal soma. However, residual dox resistant A30P α-syn was detected in glial cells within the hippocampal neurogenic niche, concomitant with the failure to fully restore hippocampal neurogenesis. This finding is indicative to a potential spread of pathology from neuron to glia. In addition, mice expressing A30P α-syn show increased anxiety-related behavior that was reversed after dox treatment. This implies that glial A30P α-synucleinopathy within the dentate gyrus is part of a process leading to impaired hippocampal neuroplasticity, which is, however, not a sole critical event for circuits implicated in anxiety-related behavior., (© 2013.)

Huntington's disease (HD) is characterized by triad of motor, cognitive, and emotional symptoms along with neuropathology in fronto-striatal circuit and limbic system including amygdala. Emotional alterations, which have a negative impact on patient well-being, represent some of the earliest symptoms of HD and might be related to the onset of the neurodegenerative process. In the transgenic rat model (tgHD rats), evidence suggest emotional alterations at the symptomatic stage along with neuropathology of the central nucleus of amygdala (CE). Studies in humans and animals demonstrate that emotion can modulate time perception. The impact of emotion on time perception has never been tested in HD, nor is it known if that impact could be part of the presymptomatic emotional phenotype of the pathology. The aim of this paper was to characterize the effect of emotion on temporal discrimination in presymptomatic tgHD animals. In the first experiment, we characterized the acute effect of an emotion (fear) conditioned stimulus on temporal discrimination using a bisection procedure, and tested its dependency upon an intact central amygdala. The second experiment was aimed at comparing presymptomatic homozygous transgenic animals at 7-months of age and their wild-type littermates (WT) in their performance on the modulation of temporal discrimination by emotion. Our principal findings show that (1) a fear cue produces a short-lived decrease of temporal precision after its termination, and (2) animals with medial CE lesion and presymptomatic tgHD animals demonstrate an alteration of this emotion-evoked temporal distortion. The results contribute to our knowledge about the presymptomatic phenotype of this HD rat model, showing susceptibility to emotion that may be related to dysfunction of the central nucleus of amygdala.

Spinocerebellar Ataxia type 17 (SCA17) is an autosomal dominantly inherited, neurodegenerative disease characterized by ataxia, involuntary movements, and dementia. A novel SCA17 mouse model having a 71 polyglutamine repeat expansion in the TATA-binding protein (TBP) has shown age related motor deficit using a classic motor test, yet concomitant weight increase might be a confounding factor for this measurement. In this study we used an automated home cage system to test several motor readouts for this same model to confirm pathological behavior results and evaluate benefits of automated home cage in behavior phenotyping. Our results confirm motor deficits in the Tbp/Q71 mice and present previously unrecognized behavioral characteristics obtained from the automated home cage, indicating its use for high-throughput screening and testing, e.g. of therapeutic compounds., (Copyright © 2013 Elsevier B.V. All rights reserved.)

The cause of Huntington disease (HD) is a polyglutamine repeat expansion of more than 36 units in the huntingtin protein, which is inversely correlated with the age at onset of the disease. However, additional genetic factors are believed to modify the course and the age at onset of HD. Recently, we identified the V471A polymorphism in the autophagy-related gene ATG7, a key component of the autophagy pathway that plays an important role in HD pathogenesis, to be associated with the age at onset in a large group of European Huntington disease patients. To confirm this association in a second independent patient cohort, we analysed the ATG7 V471A polymorphism in additional 1,464 European HD patients of the "REGISTRY" cohort from the European Huntington Disease Network (EHDN). In the entire REGISTRY cohort we could not confirm a modifying effect of the ATG7 V471A polymorphism. However, analysing a modifying effect of ATG7 in these REGISTRY patients and in patients of our previous HD cohort according to their ethnic origin, we identified a significant effect of the ATG7 V471A polymorphism on the HD age at onset only in the Italian population (327 patients). In these Italian patients, the polymorphism is associated with a 6-years earlier disease onset and thus seems to have an aggravating effect. We could specify the role of ATG7 as a genetic modifier for HD particularly in the Italian population. This result affirms the modifying influence of the autophagic pathway on the course of HD, but also suggests population-specific modifying mechanisms in HD pathogenesis.

Unlabelled: UPDtool is a computational tool for detection and classification of uniparental disomy (UPD) in trio SNP-microarray experiments. UPDs are rare events of chromosomal malsegregation and describe the condition of two homologous chromosomes or homologous chromosomal segments that were inherited from one parent. The occurrence of UPD can be of major clinical relevance. Though high-throughput molecular screening techniques are widely used, detection of UPDs and especially the subclassification remains complex. We developed UPDtool to detect and classify UPDs from SNP microarray data of parent-child trios. The algorithm was tested using five positive controls including both iso- and heterodisomic segmental UPDs and 30 trios from the HapMap project as negative controls. With UPDtool, we were able to correctly identify all occurrences of non-mosaic UPD within our positive controls, whereas no occurrence of UPD was found within our negative controls. In addition, the chromosomal breakage points could be determined more precisely than by microsatellite analysis. Our results were compared with both the gold standard, microsatellite analysis and SNPtrio, another program available for UPD detection. UPDtool is platform independent, light weight and flexible. Because of its simple input format, UPDtool may also be used with other high-throughput technologies (e.g., next-generation sequencing)., Availability and Implementation: UPDtool executables, documentation and examples can be downloaded from http://www.uni-tuebingen.de/uni/thk/de/f-genomik-software.html.

Deletions of chromosome band 12q24.33 are rare. We report on a 17-year-old male patient with intellectual disability but no major malformations or dysmorphic features in whom a de novo interstitial 660 kb deletion in 12q24.33 was detected by SNP array analysis. This deletion was secondary to a translocation t(12;14)(q24.3;q13)dn that also led to a small deletion in 14q21.1 and a small duplication in 2p23.1. The deletion overlaps with two previously published larger deletions in patients who suffered from intellectual disability, obesity, and polycystic kidney disease, indicating that haploinsufficiency of one or several of the genes in the deleted interval of the patient reported here causes intellectual deficits, but not obesity or renal problems. The 14 RefSeq genes that are harbored by this deletion include P2RX2, which had previously been proposed as a candidate gene for intellectual disability. Thus, the patient reported here broadens our knowledge of the phenotypic consequences of deletions in 12q24.33 and facilitates genotype-phenotype correlations for chromosome aberrations of this region., (Copyright © 2013 Wiley Periodicals, Inc.)

Spinocerebellar ataxia 17 (SCA17) is an autosomal-dominant, late-onset neurodegenerative disorder caused by an expanded polyglutamine (polyQ) repeat in the TATA-box-binding protein (TBP). To further investigate this devastating disease, we sought to create a first transgenic rat model for SCA17 that carries a full human cDNA fragment of the TBP gene with 64 CAA/CAG repeats (TBPQ64). In line with previous observations in mouse models for SCA17, TBPQ64 rats show a severe neurological phenotype including ataxia, impairment of postural reflexes, and hyperactivity in early stages followed by reduced activity, loss of body weight, and early death. Neuropathologically, the severe phenotype of SCA17 rats was associated with neuronal loss, particularly in the cerebellum. Degeneration of Purkinje, basket, and stellate cells, changes in the morphology of the dendrites, nuclear TBP-positive immunoreactivity, and axonal torpedos were readily found by light and electron microscopy. While some of these changes are well recapitulated in existing mouse models for SCA17, we provide evidence that some crucial characteristics of SCA17 are better mirrored in TBPQ64 rats. Thus, this SCA17 model represents a valuable tool to pursue experimentation and therapeutic approaches that may be difficult or impossible to perform with SCA17 transgenic mice. We show for the first time positron emission tomography (PET) and diffusion tensor imaging (DTI) data of a SCA animal model that replicate recent PET studies in human SCA17 patients. Our results also confirm that DTI are potentially useful correlates of neuropathological changes in TBPQ64 rats and raise hope that DTI imaging could provide a biomarker for SCA17 patients.

Spinocerebellar Ataxia Type 3 (SCA3), also known as Machado-Joseph disease, is an autosomal dominantly inherited neurodegenerative disease caused by an expanded polyglutamine stretch in the ataxin-3 protein. A pathological hallmark of the disease is cerebellar and brainstem atrophy, which correlates with the formation of intranuclear aggregates in a specific subset of neurons. Several studies have demonstrated that the formation of aggregates depends on the generation of aggregation-prone and toxic intracellular ataxin-3 fragments after proteolytic cleavage of the full-length protein. Despite this observed increase in aggregated mutant ataxin-3, information on soluble mutant ataxin-3 levels in brain tissue is lacking. A quantitative method to analyze soluble levels will be a useful tool to characterize disease progression or to screen and identify therapeutic compounds modulating the level of toxic soluble ataxin-3. In the present study we describe the development and application of a quantitative and easily applicable immunoassay for quantification of soluble mutant ataxin-3 in human cell lines and brain samples of transgenic SCA3 mice. Consistent with observations in Huntington disease, transgenic SCA3 mice reveal a tendency for decrease of soluble mutant ataxin-3 during disease progression in fractions of the cerebellum, which is inversely correlated with aggregate formation and phenotypic aggravation. Our analyses demonstrate that the time-resolved Förster resonance energy transfer immunoassay is a highly sensitive and easy method to measure the level of soluble mutant ataxin-3 in biological samples. Of interest, we observed a tendency for decrease of soluble mutant ataxin-3 only in the cerebellum of transgenic SCA3 mice, one of the most affected brain regions in Spinocerebellar Ataxia Type 3 but not in whole brain tissue, indicative of a brain region selective change in mutant ataxin-3 protein homeostasis.

Mutations or deletions of ACSL4 (FACL4, OMIM 300157) are a rare cause of non-syndromic X-linked intellectual disability. We report on a 10-year-old male patient with moderate intellectual disability, sensorineural hearing loss, facial dysmorphism, pyloric stenosis, and intestinal obstruction in whom a de novo Xq22.3-q23 deletion was detected by SNP array analysis. The deleted 1.56 Mb interval harbored ACSL4 and eight neighboring genes (GUCY2F, NXT2, KCNE1L, TMEM164, MIR3978, AMMECR1, SNORD96B, and RGAG1). In contrast to previously reported patients with chromosome aberrations in the region of the AMME complex (Alport syndrome, intellectual disability, midface hypoplasia, and elliptocytosis, OMIM 300194), this deletion did not contain the Alport syndrome gene COL4A5, suggesting that loss of one or several of the other genes in this interval is responsible for the clinical problems. In summary, the patient reported here broadens our knowledge of the phenotypic consequences of deletions of chromosome region Xq22.3-q23 and provides further proof for ACSL4 as an X-linked intellectual disability gene., (Copyright © 2013 Wiley Periodicals, Inc.)

Spinocerebellar ataxia type 3 (SCA3) is pathologically characterized by the formation of intranuclear aggregates which contain ataxin-3, the mutated protein in SCA3, in a specific subtype of neurons. It has been proposed that ataxin-3 is cleaved by proteolytic enzymes, in particular by calpains and caspases, eventually leading to the formation of aggregates. In our study, we examined the ability of calpains to cleave ataxin-3 in vitro and in vivo. We demonstrated in cell culture and mouse brain homogenates that cleavage of overexpressed ataxin-3 by calpains and in particular by calpain-2 occur and that polyglutamine expanded ataxin-3 is more sensitive to calpain degradation. Based on these results, we investigated the influence of calpains on the pathogenesis of SCA3 in vivo. For this purpose, we enhanced calpain activity in a SCA3 transgenic mouse model by knocking out the endogenous calpain inhibitor calpastatin. Double-mutant mice demonstrated an aggravated neurological phenotype with an increased number of nuclear aggregates and accelerated neurodegeneration in the cerebellum. This study confirms the critical importance of calcium-dependent calpain-type proteases in the pathogenesis of SCA3 and suggests that the manipulation of the ataxin-3 cleavage pathway and the regulation of intracellular calcium homeostasis may represent novel targets for therapeutic intervention in SCA3.

Background and Purpose: Strokes have especially devastating implications if they occur early in life; however, only limited information exists on the characteristics of acute cerebrovascular disease in young adults. Although risk factors and manifestation of atherosclerosis are commonly associated with stroke in the elderly, recent data suggests different causes for stroke in the young. We initiated the prospective, multinational European study Stroke in Young Fabry Patients (sifap) to characterize a cohort of young stroke patients., Methods: Overall, 5023 patients aged 18 to 55 years with the diagnosis of ischemic stroke (3396), hemorrhagic stroke (271), transient ischemic attack (1071) were enrolled in 15 European countries and 47 centers between April 2007 and January 2010 undergoing a detailed, standardized, clinical, laboratory, and radiological protocol., Results: Median age in the overall cohort was 46 years. Definite Fabry disease was diagnosed in 0.5% (95% confidence interval, 0.4%-0.8%; n=27) of all patients; and probable Fabry disease in additional 18 patients. Males dominated the study population (2962/59%) whereas females outnumbered men (65.3%) among the youngest patients (18-24 years). About 80.5% of the patients had a first stroke. Silent infarcts on magnetic resonance imaging were seen in 20% of patients with a first-ever stroke, and in 11.4% of patients with transient ischemic attack and no history of a previous cerebrovascular event. The most common causes of ischemic stroke were large artery atherosclerosis (18.6%) and dissection (9.9%)., Conclusions: Definite Fabry disease occurs in 0.5% and probable Fabry disease in further 0.4% of young stroke patients. Silent infarcts, white matter intensities, and classical risk factors were highly prevalent, emphasizing the need for new early preventive strategies. Clinical Trial Registration Information- URL: http://www.clinicaltrials.gov.Unique identifier: NCT00414583.

Conversion of soluble α-synuclein into insoluble and fibrillar inclusions is a hallmark of Parkinson's disease and other synucleinopathies. Accumulating evidence points towards a relationship between its generation at nerve terminals and structural synaptic pathology. Little is known about the pathogenic impact of α-synuclein conversion and deposition at nigrostriatal dopaminergic synapses in transgenic mice, mainly owing to expression limitations of the α-synuclein construct. Here, we explore whether both the rat as a model and expression of the bacterial artificial chromosome construct consisting of human full-length wild-type α-synuclein could exert dopaminergic neuropathological effects. We found that the human promoter induced a pan-neuronal expression, matching the rodent α-synuclein expression pattern, however, with prominent C-terminally truncated fragments. Ageing promoted conversion of both full-length and C-terminally truncated α-synuclein species into insolube and proteinase K-resistant fibres, with strongest accumulation in the striatum, resembling biochemical changes seen in human Parkinson's disease. Transgenic rats develop early changes in novelty-seeking, avoidance and smell before the progressive motor deficit. Importantly, the observed pathological changes were associated with severe loss of the dopaminergic integrity, thus resembling more closely the human pathology.

The Roma people, living throughout Europe and West Asia, are a diverse population linked by the Romani language and culture. Previous linguistic and genetic studies have suggested that the Roma migrated into Europe from South Asia about 1,000-1,500 years ago. Genetic inferences about Roma history have mostly focused on the Y chromosome and mitochondrial DNA. To explore what additional information can be learned from genome-wide data, we analyzed data from six Roma groups that we genotyped at hundreds of thousands of single nucleotide polymorphisms (SNPs). We estimate that the Roma harbor about 80% West Eurasian ancestry-derived from a combination of European and South Asian sources-and that the date of admixture of South Asian and European ancestry was about 850 years before present. We provide evidence for Eastern Europe being a major source of European ancestry, and North-west India being a major source of the South Asian ancestry in the Roma. By computing allele sharing as a measure of linkage disequilibrium, we estimate that the migration of Roma out of the Indian subcontinent was accompanied by a severe founder event, which appears to have been followed by a major demographic expansion after the arrival in Europe.

Background: The genetic cause of intellectual disability in most patients is unclear because of the absence of morphological clues, information about the position of such genes, and suitable screening methods. Our aim was to identify de-novo variants in individuals with sporadic non-syndromic intellectual disability., Methods: In this study, we enrolled children with intellectual disability and their parents from ten centres in Germany and Switzerland. We compared exome sequences between patients and their parents to identify de-novo variants. 20 children and their parents from the KORA Augsburg Diabetes Family Study were investigated as controls., Findings: We enrolled 51 participants from the German Mental Retardation Network. 45 (88%) participants in the case group and 14 (70%) in the control group had de-novo variants. We identified 87 de-novo variants in the case group, with an exomic mutation rate of 1·71 per individual per generation. In the control group we identified 24 de-novo variants, which is 1·2 events per individual per generation. More participants in the case group had loss-of-function variants than in the control group (20/51 vs 2/20; p=0·022), suggesting their contribution to disease development. 16 patients carried de-novo variants in known intellectual disability genes with three recurrently mutated genes (STXBP1, SYNGAP1, and SCN2A). We deemed at least six loss-of-function mutations in six novel genes to be disease causing. We also identified several missense alterations with potential pathogenicity., Interpretation: After exclusion of copy-number variants, de-novo point mutations and small indels are associated with severe, sporadic non-syndromic intellectual disability, accounting for 45-55% of patients with high locus heterogeneity. Autosomal recessive inheritance seems to contribute little in the outbred population investigated. The large number of de-novo variants in known intellectual disability genes is only partially attributable to known non-specific phenotypes. Several patients did not meet the expected syndromic manifestation, suggesting a strong bias in present clinical syndrome descriptions., Funding: German Ministry of Education and Research, European Commission 7th Framework Program, and Swiss National Science Foundation., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Huntington disease (HD) is an inherited progressive neurodegenerative disorder, characterized by motor, cognitive, and psychiatric deficits as well as neurodegeneration and brain atrophy beginning in the striatum and the cortex and extending to other subcortical brain regions. The genetic cause is an expansion of the CAG repeat stretch in the HTT gene encoding huntingtin protein (htt). Here, we generated an HD transgenic rat model using a human bacterial artificial chromosome (BAC), which contains the full-length HTT genomic sequence with 97 CAG/CAA repeats and all regulatory elements. BACHD transgenic rats display a robust, early onset and progressive HD-like phenotype including motor deficits and anxiety-related symptoms. In contrast to BAC and yeast artificial chromosome HD mouse models that express full-length mutant huntingtin, BACHD rats do not exhibit an increased body weight. Neuropathologically, the distribution of neuropil aggregates and nuclear accumulation of N-terminal mutant huntingtin in BACHD rats is similar to the observations in human HD brains. Aggregates occur more frequently in the cortex than in the striatum and neuropil aggregates appear earlier than mutant htt accumulation in the nucleus. Furthermore, we found an imbalance in the striatal striosome and matrix compartments in early stages of the disease. In addition, reduced dopamine receptor binding was detectable by in vivo imaging. Our data demonstrate that this transgenic BACHD rat line may be a valuable model for further understanding the disease mechanisms and for preclinical pharmacological studies.

Synucleinopathies including Parkinson's disease (PD) are characterized by the accumulation of alpha-synuclein (α-syn) within neural cell bodies and their processes. Transgenic mice overexpressing human wild-type or mutant forms of α-syn under the control of different promoters were developed to analyse the underlying neuropathology of PD. One of the earliest clinical symptoms associated with PD is olfactory impairment. The generation of new neurons persists up to adulthood in mammals, in particular the olfactory bulb (OB). In order to assess this process in relation to α-syn accumulation, we used mice overexpressing human wild-type α-syn under the regulatable control (tet-off) of the calcium/calmodulin-dependent protein kinase IIα-promoter (CaMKII). We observed a decrease in OB neurogenesis in transgenic animals compared to controls using 5-bromo-2'-deoxyuridine (BrdU) to label newly generated cells (neuron-specific nuclear protein; NeuN). After cessation of transgene expression we detected an increase in newly generated cells both in granular (GCL) and glomerular (GLOM) layers of the OB. This led to a rescue of newly generated neurons (BrdU(+)/NeuN(+)) within the GLOM with a distinct specificity for the dopaminergic subpopulation. In contrast, we did not detect a cell-specific rescue of neuronal cells in the GCL suggesting diverse effects of alpha-synucleinopathy in both interneuronal layers of the OB. Colabelling of BrdU with glial markers showed that a differentiation into neither astroglia nor microglia attributed to the observed phenotype in the GCL. In particular, BrdU(+) particles located within microglial cells were predominantly associated close to the membrane therefore the resembling phagocytosed nuclear fragments of BrdU(+) cells. Thus, our study further contributes insights into α-syn accumulation as a causative player in the impairment of adult neurogenesis and emphasizes its diverse role in cell renewal of distinct OB cell layers., (Copyright © 2012 IBRO. All rights reserved.)