Birgitt Schüle, MD, is an Associate Professor in the Department of Pathology at Stanford University School of Medicine. Her research focuses on medical genetics and stem cell modeling to uncover disease mechanisms and pathways involved in neurodegeneration in Parkinson's disease and related disorders. She is dedicated to developing novel therapeutic strategies that contribute to the advancement of precision medicine.
Dr. Schüle obtained her medical training from the Georg-August University Göttingen and Medical University Lübeck, Germany, between 1993 and 2001. She earned her doctoral degree in medicine (Dr. med.) in neurophysiology from the Georg-August University Göttingen in 2001. During her neurology internship from 2001 to 2002 at the Medical University of Lübeck under the guidance of Prof. Christine Klein. Subsequently, she pursued a postdoctoral fellowship in human genetics with Prof. Uta Francke at Stanford University School of Medicine from 2003 to 2005.
From 2005 to 2019, Dr. Schüle demonstrated leadership in spearheading critical clinical research programs and establishing essential biospecimen repositories for neurogenetics, translational stem cell research, and brain donation at the Parkinson's Institute and Clinical Center.
Currently, Dr. Schüle serves as the Associate Core Leader, Neuropathology, within the Stanford Alzheimer Research Center (ADRC). Her contributions to ADRC include genetic characterization, biobanking, and the establishment of a human induced pluripotent stem cell and post-mortem leptomeninges tissue bank. These resources are shared with the data and tissue repositories at the National Institutes of Health (NIH), facilitating collaborative research and advancing our understanding of neurodegenerative diseases.
Dr. Schüle's expertise and dedication in the field of neurodegeneration contribute significantly to the advancement of medical knowledge. She is recognized as a respected member of the scientific community, playing an important role in the pursuit of effective treatments and precision medicine approaches.
Associate Professor (Research), Pathology
Member, SPARK at Stanford
Member, Wu Tsai Neurosciences Institute
Member, Justice, Equity, Diversity & Inclusion (JEDI) Committee, Stanford ADRC (2021 - Present)
Mentor, Pathways to Neurosciences Program (2021 - Present)
Member, Clin Gen, Clinical Domain Working Groups, Parkinson's Disease Gene Curation Expert Panel (2020 - Present)
Member, Diversity and Inclusion Committee, Department Pathology (2020 - Present)
Dr. med., Georg-August University Goettingen, Germany, Neurophysiology (2001)
MD, Medical University Luebeck, Germany, Human Medicine (2001)
Community and International Work
Clinico-genetic studies in spinocerebellar ataxia 10 (SCA-10) in Mexico, Guadalajara & Mexico City, Mexico
Genomic analysis and clinical phenotyping of SCA-10
Hospital San Javier, Guadalajara & UNAM, Mexico
Patients with movement disorders
Opportunities for Student Involvement
Independent Studies (2)
- Out-of-Department Advanced Research Laboratory in Bioengineering
BIOE 191X (Aut)
- Out-of-Department Undergraduate Research
BIO 199X (Aut, Win, Spr)
- Out-of-Department Advanced Research Laboratory in Bioengineering
Embryoid Body Cells from Human Embryonic Stem Cells Overexpressing Dopaminergic Transcription Factors Survive and Initiate Neurogenesis via Neural Rosettes in the Substantia Nigra.
2023; 13 (2)
Transplantation of immature dopaminergic neurons or neural precursors derived from embryonic stem cells (ESCs) into the substantia nigra pars compacta (SNpc) is a potential therapeutic approach for functional restitution of the nigrostriatal pathway in Parkinson's disease (PD). However, further studies are needed to understand the effects of the local microenvironment on the transplanted cells to improve survival and specific differentiation in situ. We have previously reported that the adult SNpc sustains a neurogenic microenvironment. Non-neuralized embryoid body cells (EBCs) from mouse ESCs (mESCs) overexpressing the dopaminergic transcription factor Lmx1a gave rise to many tyrosine hydroxylase (Th+) cells in the intact and damaged adult SNpc, although only for a short-term period. Here, we extended our study by transplanting EBCs from genetically engineered naive human ESC (hESC), overexpressing the dopaminergic transcription factors LMX1A, FOXA2, and OTX2 (hESC-LFO), in the SNpc. Unexpectedly, no graft survival was observed in wild-type hESC EBCs transplants, whereas hESC-LFO EBCs showed viability in the SNpc. Interestingly, neural rosettes, a developmental hallmark of neuroepithelial tissue, emerged at 7- and 15-days post-transplantation (dpt) from the hESC-LFO EBCs. Neural rosettes expressed specification dopaminergic markers (Lmx1a, Otx2), which gave rise to several Th+ cells at 30 dpt. Our results suggest that the SNpc enables the robust initiation of neural differentiation of transplanted human EBCs prompted to differentiate toward the midbrain dopaminergic phenotype.
View details for DOI 10.3390/brainsci13020329
View details for PubMedID 36831872
Nuclease-dead S. aureus Cas9 downregulates alpha-synuclein and reduces mtDNA damage and oxidative stress levels in patient-derived stem cell model of Parkinson's disease.
bioRxiv : the preprint server for biology
Parkinson's disease (PD) is one of the most common neurodegenerative diseases, but no disease modifying therapies have been successful in clinical translation presenting a major unmet medical need. A promising target is alpha-synuclein or its aggregated form, which accumulates in the brain of PD patients as Lewy bodies. While it is not entirely clear which alpha-synuclein protein species is disease relevant, mere overexpression of alpha-synuclein in hereditary forms leads to neurodegeneration. To specifically address gene regulation of alpha-synuclein, we developed a CRISPR interference (CRISPRi) system based on the nuclease dead S. aureus Cas9 (SadCas9) fused with the transcriptional repressor domain Krueppel-associated box to controllably repress alpha-synuclein expression at the transcriptional level. We screened single guide (sg)RNAs across the SNCA promoter and identified several sgRNAs that mediate downregulation of alpha-synuclein at varying levels. CRISPRi downregulation of alpha-synuclein in iPSC-derived neuronal cultures from a patient with an SNCA genomic triplication showed functional recovery by reduction of oxidative stress and mitochondrial DNA damage. Our results are proof-of-concept in vitro for precision medicine by targeting the SNCA gene promoter. The SNCA CRISPRi approach presents a new model to understand safe levels of alpha-synuclein downregulation and a novel therapeutic strategy for PD and related alpha-synucleinopathies.
View details for DOI 10.1101/2023.01.24.525105
View details for PubMedID 36747875
A reference human induced pluripotent stem cell line for large-scale collaborative studies.
Cell stem cell
2022; 29 (12): 1685-1702.e22
Human induced pluripotent stem cell (iPSC) lines are a powerful tool for studying development and disease, but the considerable phenotypic variation between lines makes it challenging to replicate key findings and integrate data across research groups. To address this issue, we sub-cloned candidate human iPSC lines and deeply characterized their genetic properties using whole genome sequencing, their genomic stability upon CRISPR-Cas9-based gene editing, and their phenotypic properties including differentiation to commonly used cell types. These studies identified KOLF2.1J as an all-around well-performing iPSC line. We then shared KOLF2.1J with groups around the world who tested its performance in head-to-head comparisons with their own preferred iPSC lines across a diverse range of differentiation protocols and functional assays. On the strength of these findings, we have made KOLF2.1J and its gene-edited derivative clones readily accessible to promote the standardization required for large-scale collaborative science in the stem cell field.
View details for DOI 10.1016/j.stem.2022.11.004
View details for PubMedID 36459969
Early embryonic lethality in complex I associated p.L104P Nubpl mutant mice.
Orphanet journal of rare diseases
2022; 17 (1): 386
BACKGROUND: Variants in the mitochondrial complex I assembly factor, NUBPL are associated with a rare cause of complex I deficiency mitochondrial disease. Patients affected by complex I deficiency harboring homozygous NUBPL variants typically have neurological problems including seizures, intellectual disability, and ataxia associated with cerebellar hypoplasia. Thus far only 19 cases have been reported worldwide, and no treatment is available for this rare disease.METHODS: To investigate the pathogenesis of NUBPL-associated complex I deficiency, and for translational studies, we generated a knock-in mouse harboring a patient-specific variant Nubpl c.311T>C; p. L104P reported in three families.RESULTS: Similar to Nubpl global knockout mice, the Nubpl p. L104P homozygous mice are lethal at embryonic day E10.5, suggesting that the Nubpl p. L104P variant is likely a hypomorph allele. Given the recent link between Parkinson's disease and loss-of-function NUBPL variants, we also explored aging-related behaviors and immunocytochemical changes in Nubpl hemizygous mice and did not find significant behavioral and pathological changes for alpha-synuclein and oxidative stress markers .CONCLUSION: Our data suggest that homozygotes with Nubpl variants, similar to the null mice, are lethal, and heterozygotes are phenotypically and neuropathologically normal. We propose that a tissue-specific knockout strategy is required to establish a mouse model of Nubpl-associated complex I deficiency disorder for future mechanistic and translational studies.
View details for DOI 10.1186/s13023-022-02446-y
View details for PubMedID 36280881
ATTCT and ATTCC repeat expansions in the ATXN10 gene affect disease penetrance of spinocerebellar ataxia type 10.
2022; 3 (4): 100137
Spinocerebellar ataxia type 10 (SCA10) is an autosomal-dominant disorder caused by an expanded pentanucleotide repeat in the ATXN10 gene. This repeat expansion, when fully penetrant, has a size of 850-4,500 repeats. It has been shown that the repeat composition can be a modifier of disease, e.g., seizures. Here, we describe a Mexican kindred in which we identified both pure (ATTCT)n and mixed (ATTCT)n-(ATTCC)n expansions in the same family. We used amplification-free targeted sequencing and optical genome mapping to decipher the composition of these repeat expansions. We found a considerable degree of mosaicism of the repeat expansion. This mosaicism was confirmed in skin fibroblasts from individuals with ATXN10 expansions with RNAScope in situ hybridization. All affected family members with the mixed ATXN10 repeat expansion showed typical clinical signs of spinocerebellar ataxia and epilepsy. In contrast, individuals with the pure ATXN10 expansion present with Parkinson's disease or are unaffected, even in individuals more than 20 years older than the average age at onset for SCA10. Our findings suggest that the pure (ATTCT)n expansion is non-pathogenic, while repeat interruptions, e.g., (ATTCC)n, are necessary to cause SCA10. This mechanism has been recently described for several other repeat expansions including SCA31 (BEAN1), SCA37 (DAB1), and three loci for benign adult familial myoclonic epilepsy BAFME (SAMD12, TNRC6A, RAPGEF2). Therefore, long-read sequencing and optical genome mapping of the entire genomic structure of repeat expansions are critical for clinical practice and genetic counseling, as variations in the repeat can affect disease penetrance, symptoms, and disease trajectory.
View details for DOI 10.1016/j.xhgg.2022.100137
View details for PubMedID 36092952
Gene Expression Analysis in Stem Cell-derived Cortical Neuronal Cultures Using Multi-well SYBR Green Quantitative PCR Arrays
2022; 12 (14)
View details for DOI 10.21769/BioProtoc.4476
View details for Web of Science ID 000834751400004
Multiplex imaging of human induced pluripotent stem cell-derived neurons with CO-Detection by indEXing (CODEX) technology.
Journal of neuroscience methods
BACKGROUND: Human induced pluripotent stem cell (iPSC) models have been hailed as a breakthrough for understanding disease and developing new therapeutics. The major advantage of iPSC-derived neurons is that they carry the genetic background of the donor, and as such could be more predictive for clinical translation. However, the development of these cell models is time-consuming and expensive and it is thus critical to maximize readout of markers for immunocytochemistry. One option is to use a highly multiplexed fluorescence imaging assay, like CO-Detection by indEXing (CODEX), which allows detection of 50+ targets in situ.NEW METHOD: This paper describes the development of CODEX in neuronal cell cultures derived from human iPSCs.RESULTS: We differentiated human iPSCs into mixed neuronal and glial cultures on glass coverslips. We then developed and optimized a panel of 21 antibodies to phenotype iPSC-derived neuronal subtypes of cortical, dopaminergic, and striatal neurons, as well as astrocytes, and pre-and postsynaptic proteins.COMPARISON WITH EXISTING METHODS: Compared to standard immunocytochemistry, CODEX oligo-conjugated fluorophores circumvent antibody host interactions and allow for highly customized multiplexing.CONCLUSION: We show that CODEX can be applied to iPSC neuronal cultures and developed fixation and staining protocols for the neurons to sustain the multiple wash-stain cycles of the technology. Furthermore, we demonstrate both cellular and subcellular resolution imaging of multiplexed markers in the same sample.
View details for DOI 10.1016/j.jneumeth.2022.109653
View details for PubMedID 35724898
Effect of LRRK2 protein and activity on stimulated cytokines in human monocytes and macrophages.
NPJ Parkinson's disease
2022; 8 (1): 34
Leucine-rich-repeat kinase 2 (LRRK2), a potential therapeutic target for the treatment of Parkinson's disease (PD), is highly expressed in monocytes and macrophages and may play a role in the regulation of inflammatory pathways. To determine how LRRK2 protein levels and/or its activity modulate inflammatory cytokine/chemokine levels in human immune cells, isogenic human induced pluripotent stem cells (iPSC) with the LRRK2-activating G2019S mutation, wild-type LRRK2, and iPSC deficient in LRRK2 were differentiated to monocytes and macrophages and stimulated with inflammatory toll-like receptor (TLR) agonists in the presence and absence of LRRK2 kinase inhibitors. The effect of LRRK2 inhibitors and the effect of increasing LRRK2 levels with interferon gamma on TLR-stimulated cytokines were also assessed in primary peripheral blood-derived monocytes. Monocytes and macrophages with the LRRK2 G2019S mutation had significantly higher levels of cytokines and chemokines in tissue culture media following stimulation with TLR agonists compared to isogenic controls. Knockout of LRRK2 impaired phagocytosis but did not significantly affect TLR-mediated cytokine levels. Interferon gamma significantly increased the levels of LRRK2 and phosphorylation of its downstream Rab10 substrate, and potentiated TLR-mediated cytokine levels. LRRK2 kinase inhibitors did not have a major effect on TLR-stimulated cytokine levels. Results suggest that the LRRK2 G2019S mutation may potentiate inflammation following activation of TLRs. However, this was not dependent on LRRK2 kinase activity. Indeed, LRRK2 kinase inhibitors had little effect on TLR-mediated inflammation under the conditions employed in this study.
View details for DOI 10.1038/s41531-022-00297-9
View details for PubMedID 35347144
Isogenic human SNCA gene dosage induced pluripotent stem cells to model Parkinson's disease.
Stem cell research
2022; 60: 102733
Alpha-synuclein overexpression and aggregation are critical factors in the pathogenesis of Parkinson's disease (PD). Clinical cases with alpha-synuclein (SNCA) multiplications or deletions indicate that gene expression levels are essential for neurodegeneration and neurodevelopment. Here, we developed an isogenic SNCA gene dosage model using CRISPR/Cas9 gene editing to introduce frameshift mutations into exon 2 of the SNCA coding region in human induced pluripotent stem cells (iPSCs) from a patient with an SNCA triplication. We derived and characterized clones with different frameshift mutations. This isogenic SNCA gene dosage panel will address the physiological and detrimental effects of varying alpha-synuclein expression levels.
View details for DOI 10.1016/j.scr.2022.102733
View details for PubMedID 35263701
Phenotypic Heterogeneity among GBA p.R202X Carriers in Lewy Body Spectrum Disorders.
1800; 10 (1)
We describe the clinical and neuropathologic features of patients with Lewy body spectrum disorder (LBSD) carrying a nonsense variant, c.604C>T; p.R202X, in the glucocerebrosidase 1 (GBA) gene. While this GBA variant is causative for Gaucher's disease, the pathogenic role of this mutation in LBSD is unclear. Detailed neuropathologic evaluation was performed for one index case and a structured literature review of other GBA p.R202X carriers was conducted. Through the systematic literature search, we identified three additional reported subjects carrying the same GBA mutation, including one Parkinson's disease (PD) patient with early disease onset, one case with neuropathologically-verified LBSD, and one unaffected relative of a Gaucher's disease patient. Among the affected subjects carrying the GBA p.R202X, all males were diagnosed with Lewy body dementia, while the two females presented as PD. The clinical penetrance of GBA p.R202X in LBSD patients and families argues strongly for a pathogenic role for this variant, although presenting with a striking phenotypic heterogeneity of clinical and pathological features.
View details for DOI 10.3390/biomedicines10010160
View details for PubMedID 35052839
The commercial genetic testing landscape for Parkinson's disease.
Parkinsonism & related disorders
INTRODUCTION: There have been no specific guidelines regarding which genes should be tested in the clinical setting for Parkinson's disease (PD) or parkinsonism. We evaluated the types of clinical genetic testing offered for PD as the first step of our gene curation.METHODS: The National Institutes of Health (NIH) Genetic Testing Registry (GTR) was queried on 12/7/2020 to identify current commercial PD genetic test offerings by clinical laboratories, internationally.RESULTS: We identified 502 unique clinical genetic tests for PD, from 28 Clinical Laboratory Improvement Amendments (CLIA)-approved clinical laboratories. These included 11 diagnostic PD panels. The panels were notable for their differences in size, ranging from 5 to 62 genes. Five genes for variant query were included in all panels (SNCA, PRKN, PINK-1, PARK7 (DJ1), and LRRK2). Notably, the addition of the VPS35 and GBA genes was variable. Panel size differences stemmed from inclusion of genes linked to atypical parkinsonism and dystonia disorders, and genes in which the link to PD causation is controversial.CONCLUSION: There is an urgent need for expert opinion regarding which genes should be included in a commercial laboratory multi-gene panel for PD.
View details for DOI 10.1016/j.parkreldis.2021.10.001
View details for PubMedID 34696975
Embryoid Body Formation from Mouse and Human Pluripotent Stem Cells for Transplantation to Study Brain Microenvironment and Cellular Differentiation.
Methods in molecular biology (Clifton, N.J.)
Human embryonic stem cell (hESC) and human-induced pluripotent stem cell (hiPSC) technologies have a critical role in regenerative strategies for personalized medicine. Both share the ability to differentiate into almost any cell type of the human body. The study of their properties and clinical applications requires the development of robust and reproducible cell culture paradigms that direct cell differentiation toward a specific phenotype in vitro and in vivo. Our group evaluated the potential of mouse ESCs (mESCs), hESCs, and hiPSCs (collectively named pluripotent stem cells, PSCs) to analyze brain microenvironments through the use of embryoid body (EB)-derived cells from these cell sources. EB are cell aggregates in 3D culture conditions that recapitulate embryonic development. Our approach focuses on studying the midbrain dopaminergic phenotype and transplanting EB into the substantia nigra pars compacta (SNpc) in a Parkinson's disease rodent model. Here, we describe cell culture protocols for EB generation from PSCs that show significant in vivo differentiation toward dopaminergic neurons.
View details for DOI 10.1007/7651_2021_433
View details for PubMedID 34611820
Short-term deceleration capacity of heart rate: a sensitive marker of cardiac autonomic dysfunction in idiopathic Parkinson's disease.
Clinical autonomic research : official journal of the Clinical Autonomic Research Society
PURPOSE: Cardiac autonomic dysfunction in idiopathic Parkinson's disease (PD) manifests as reduced heart rate variability (HRV). In the present study, we explored the deceleration capacity of heart rate (DC) in patients with idiopathic PD, an advanced HRV marker that has proven clinical utility.METHODS: Standard and advanced HRV measures derived from 7-min electrocardiograms in 20 idiopathic PD patients and 27 healthy controls were analyzed. HRV measures were compared using regression analysis, controlling for age, sex, and mean heart rate.RESULTS: Significantly reduced HRV was found only in the subcohort of PD patients older than 60years. Low- frequency power and global HRV measures were lower in patients than in controls, but standard beat-to-beat HRV markers (i.e., rMSSD and high-frequency power) were not significantly different between groups. DC was significantly reduced in the subcohort of PD patients older than 60years compared to controls.CONCLUSIONS: Deceleration-related oscillations of HRV were significantly reduced in the older PD patients compared to healthy controls, suggesting that short-term DC may be a sensitive marker of cardiac autonomic dysfunction in PD. DC may be complementary to traditional markers of short-term HRV for the evaluation of autonomic modulation in PD. Further study to examine the association between DC and cardiac adverse events in PD is needed to clarify the clinical relevance of DC in this population.
View details for DOI 10.1007/s10286-021-00815-4
View details for PubMedID 34251546
Genome-wide association studies of LRRK2 modifiers of Parkinson's disease.
Annals of neurology
OBJECTIVE: The aim of this study was to search for genes/variants that modify the effect of LRRK2 mutations in terms of penetrance and age-at-onset of Parkinson's disease.METHODS: We performed the first genome-wide association study of penetrance and age-at-onset of Parkinson's disease in LRRK2 mutation carriers (776 cases and 1,103 non-cases at their last evaluation). Cox proportional hazard models and linear mixed models were used to identify modifiers of penetrance and age-at-onset of LRRK2 mutations, respectively. We also investigated whether a polygenic risk score derived from a published genome-wide association study of Parkinson's disease was able to explain variability in penetrance and age-at-onset in LRRK2 mutation carriers.RESULTS: A variant located in the intronic region of CORO1C on chromosome 12 (rs77395454; P-value=2.5E-08, beta=1.27, SE=0.23, risk allele: C) met genome-wide significance for the penetrance model. Co-immunoprecipitation analyses of LRRK2 and CORO1C supported an interaction between these two proteins. A region on chromosome 3, within a previously reported linkage peak for Parkinson's disease susceptibility, showed suggestive associations in both models (penetrance top variant: P-value=1.1E-07; age-at-onset top variant: P-value=9.3E-07). A polygenic risk score derived from publicly available Parkinson's disease summary statistics was a significant predictor of penetrance, but not of age-at-onset.INTERPRETATION: This study suggests that variants within or near CORO1C may modify the penetrance of LRRK2 mutations. In addition, common Parkinson's disease associated variants collectively increase the penetrance of LRRK2 mutations. This article is protected by copyright. All rights reserved.
View details for DOI 10.1002/ana.26094
View details for PubMedID 33938021
Genetic and Environmental Factors Influence the Pleomorphy of LRRK2 Parkinsonism.
International journal of molecular sciences
2021; 22 (3)
Missense mutations in the LRRK2 gene were first identified as a pathogenic cause of Parkinson's disease (PD) in 2004. Soon thereafter, a founder mutation in LRRK2, p.G2019S (rs34637584), was described, and it is now estimated that there are approximately 100,000 people worldwide carrying this risk variant. While the clinical presentation of LRRK2 parkinsonism has been largely indistinguishable from sporadic PD, disease penetrance and age at onset can be quite variable. In addition, its neuropathological features span a wide range from nigrostriatal loss with Lewy body pathology, lack thereof, or atypical neuropathology, including a large proportion of cases with concomitant Alzheimer's pathology, hailing LRRK2 parkinsonism as the "Rosetta stone" of parkinsonian disorders, which provides clues to an understanding of the different neuropathological trajectories. These differences may result from interactions between the LRRK2 mutant protein and other proteins or environmental factors that modify LRRK2 function and, thereby, influence pathobiology. This review explores how potential genetic and biochemical modifiers of LRRK2 function may contribute to the onset and clinical presentation of LRRK2 parkinsonism. We review which genetic modifiers of LRRK2 influence clinical symptoms, age at onset, and penetrance, what LRRK2 mutations are associated with pleomorphic LRRK2 neuropathology, and which environmental modifiers can augment LRRK2 mutant pathophysiology. Understanding how LRRK2 function is influenced and modulated by other interactors and environmental factors-either increasing toxicity or providing resilience-will inform targeted therapeutic development in the years to come. This will allow the development of disease-modifying therapies for PD- and LRRK2-related neurodegeneration.
View details for DOI 10.3390/ijms22031045
View details for PubMedID 33494262
The Role of Alpha-Synuclein and Other Parkinson's Genes in Neurodevelopmental and Neurodegenerative Disorders.
International journal of molecular sciences
2020; 21 (16)
Neurodevelopmental and late-onset neurodegenerative disorders present as separate entities that are clinically and neuropathologically quite distinct. However, recent evidence has highlighted surprising commonalities and converging features at the clinical, genomic, and molecular level between these two disease spectra. This is particularly striking in the context of autism spectrum disorder (ASD) and Parkinson's disease (PD). Genetic causes and risk factors play a central role in disease pathophysiology and enable the identification of overlapping mechanisms and pathways. Here, we focus on clinico-genetic studies of causal variants and overlapping clinical and cellular features of ASD and PD. Several genes and genomic regions were selected for our review, including SNCA (alpha-synuclein), PARK2 (parkin RBR E3 ubiquitin protein ligase), chromosome 22q11 deletion/DiGeorge region, and FMR1 (fragile X mental retardation 1) repeat expansion, which influence the development of both ASD and PD, with converging features related to synaptic function and neurogenesis. Both PD and ASD display alterations and impairments at the synaptic level, representing early and key disease phenotypes, which support the hypothesis of converging mechanisms between the two types of diseases. Therefore, understanding the underlying molecular mechanisms might inform on common targets and therapeutic approaches. We propose to re-conceptualize how we understand these disorders and provide a new angle into disease targets and mechanisms linking neurodevelopmental disorders and neurodegeneration.
View details for DOI 10.3390/ijms21165724
View details for PubMedID 32785033
Establishing polygenic risk score reporting standards and a polygenic score catalog to improve validation, interpretation and reproducibility
WILEY. 2020: 496–97
View details for Web of Science ID 000540090000068
Report of the Phenotype of a Patient with Roberts Syndrome and a Rare ESCO2 Variant.
Journal of pediatric genetics
2020; 9 (1): 58–62
Roberts syndrome is a rare autosomal recessive genetic disease. In this report, we report a Brazilian patient with a rare ESCO2 variant. The patient manifested a broad range of clinical findings including the significant, bilateral shortening of the extremities. He deteriorated and passed away at 20 days of age. High-resolution GTG-banded karyotype showed lack of centromeric constriction in some chromosomes, premature centromere separation in others, and repulsion of the heterochromatin regions. Molecular analysis of the ESCO2 gene revealed a deletion of 4 bp involving exon 4 in homozygosity (NM_00107420.2:c.875_878delACAG), which causes loss of ESCO2 function. We describe the clinical presentation caused by a rare ESCO2 variant.
View details for DOI 10.1055/s-0039-1696636
View details for PubMedID 31976146
View details for PubMedCentralID PMC6976314
Pulse-Field capillary electrophoresis of repeat-primed PCR amplicons for analysis of large repeats in Spinocerebellar Ataxia Type 10.
2020; 15 (3): e0228789
Large expansions of microsatellite DNA cause several neurological diseases. In Spinocerebellar ataxia type 10 (SCA10), the repeat interruptions change disease phenotype; an (ATTCC)n or a (ATCCT)n/(ATCCC)n interruption within the (ATTCT)n repeat is associated with the robust phenotype of ataxia and epilepsy while mostly pure (ATTCT)n may have reduced penetrance. Large repeat expansions of SCA10, and many other microsatellite expansions, can exceed 10,000 base pairs (bp) in size. Conventional next generation sequencing (NGS) technologies are ineffective in determining internal sequence contents or size of these expanded repeats. Using repeat primed PCR (RP-PCR) in conjunction with a high-sensitivity pulsed-field capillary electrophoresis fragment analyzer (FEMTO-Pulse, Agilent, Santa Clara, CA) (RP-FEMTO hereafter), we successfully determined sequence content of large expansion repeats in genomic DNA of SCA10 patients and transformed yeast artificial chromosomes containing SCA10 repeats. This RP-FEMTO is a simple and economical methodology which could complement emerging NGS for very long sequence reads such as Single Molecule, Real-Time (SMRT) and nanopore sequencing technologies.
View details for DOI 10.1371/journal.pone.0228789
View details for PubMedID 32160188
Loss-of-Function NUBPL Mutation May Link Parkinson's Disease to Recessive Complex I Deficiency.
Frontiers in neurology
2020; 11: 555961
In an unbiased genome-wide screen for copy number variants (CNVs) on a cohort of Parkinson's disease (PD) patients, we identified in one patient a complex chromosomal rearrangement involving the nucleotide binding protein-like (NUBPL) gene on chromosome 14q12. We noted that mutations in the NUBPL gene had been reported as causing autosomal recessive (AR) mitochondrial Complex I (CI) deficiency in children. The precise breakpoints of the rearrangement in our PD case were found to be identical to those described in a patient with AR CI deficiency who also harbored a second pathogenic mutation in NUBPL. Mitochondrial dysfunction has long been considered a strong contributor to PD, and there is substantial evidence that decreased CI activity plays a central role in PD pathogenesis. We hypothesize that pathogenic NUBPL variants may increase the risk for PD analogous to variants in the glucosylceramidase beta (GBA) gene that increase the risk of developing PD in heterozygous carriers.
View details for DOI 10.3389/fneur.2020.555961
View details for PubMedID 33224084
Nonsteroidal Anti-Inflammatory Use and LRRK2 Parkinson's Disease Penetrance.
Movement disorders : official journal of the Movement Disorder Society
The penetrance of leucine rich repeat kinase 2 (LRRK2) mutations is incomplete and may be influenced by environmental and/or other genetic factors. Nonsteroidal anti-inflammatory drugs (NSAIDs) are known to reduce inflammation and may lower Parkinson's disease (PD) risk, but their role in LRRK2-associated PD is unknown.The objective of this study is to evaluate the association of regular NSAID use and LRRK2-associated PD.Symptomatic ("LRRK2-PD") and asymptomatic ("LRRK2-non-PD") participants with LRRK2 G2019S, R1441X, or I2020T variants (definitely pathogenic variant carriers) or G2385R or R1628P variants (risk variant carriers) from 2 international cohorts provided information on regular ibuprofen and/or aspirin use (≥2 pills/week for ≥6 months) prior to the index date (diagnosis date for PD, interview date for non-PD). Multivariate logistic regression was used to evaluate the relationship between regular NSAID use and PD for any NSAID, separately for ibuprofen and aspirin in all carriers and separately in pathogenic and risk variant groups.A total of 259 LRRK2-PD and 318 LRRK2-non-PD participants were enrolled. Regular NSAID use was associated with reduced odds of PD in the overall cohort (odds ratio [OR], 0.34; 95% confidence interval [CI], 0.21-0.57) and in both pathogenic and risk variant carriers (ORPathogenic , 0.38; 95% CI, 0.21-0.67 and ORRiskVariant , 0.19; 95% CI, 0.04-0.99). Similar associations were observed for ibuprofen and aspirin separately (ORIbuprofen , 0.19; 95% CI, 0.07-0.50 and ORAspirin , 0.51; 95% CI, 0.28-0.91).Regular NSAID use may be associated with reduced penetrance in LRRK2-associated PD. The LRRK2 protein is involved in inflammatory pathways and appears to be modulated by regular anti-inflammatory use. Longitudinal observational and interventional studies of NSAID exposure and LRRK2-PD are needed to confirm this association. © 2020 International Parkinson and Movement Disorder Society.
View details for DOI 10.1002/mds.28189
View details for PubMedID 32662532
Heart rate variability biomarkers of leucine-rich repeat kinase 2-associated Parkinson's disease
View details for Web of Science ID 000621172600099
Exosome/Microvesicle Content is Altered in LRRK2 Mutant iPSC-derived Neural Cells.
The Journal of comparative neurology
Extracellular vesicles, including exosomes and other microvesicles (EMVs), have been described as sensitive biomarkers that represent disease states and response to therapies. In light of recent reports of disease-mirroring EMV molecular signatures, the present study profiled 2 EMVs from different Parkinson's disease (PD) tissue sources: 1. neural progenitor cells derived from an endogenous adult stem/progenitor cell, called adult human neural progenitor cells (AHNP), that we found to be pathological when isolated from postmortem PD patients' substantia nigra; and 2. Leucine-Rich Kinase 2 (LRRK2) gene identified patient induced pluripotent stem cells (iPSCs), which were used to isolate EMVs and begin to characterize their cargoes. Initial characterization of EMVs derived from idiopathic patients (AHNPs), and mutant LRRK2 patients showed differences between both phenotypes and when compared to a sibling control in EMV size and release based on Nanosight analysis. Furthermore, molecular profiling disclosed that neurodegenerative-related gene pathways altered in PD can be reversed using gene-editing approaches. In fact, the EMV cargo genes exhibited normal expression patterns after gene editing. This study shows that EMVs have the potential to serve as sensitive biomarkers of disease state in both idiopathic and gene-identified PD patients and that following gene-editing EMVs reflect a corrected state. This is relevant for both prodromal and symptomatic patient populations where potential responses to therapies can be monitored via non-invasive liquid biopsies and EMV characterizations. This article is protected by copyright. All rights reserved.
View details for DOI 10.1002/cne.24819
View details for PubMedID 31743443
Large expert-curated database for benchmarking document similarity detection in biomedical literature search
DATABASE-THE JOURNAL OF BIOLOGICAL DATABASES AND CURATION
View details for DOI 10.1093/database/baz085
View details for Web of Science ID 000494411700001
Increased markers of cardiac vagal activity in leucine-rich repeat kinase 2-associated Parkinson's disease.
Clinical autonomic research : official journal of the Clinical Autonomic Research Society
PURPOSE: Cardiac autonomic dysfunction manifests as reduced heart rate variability (HRV) in idiopathic Parkinson's disease (PD), but no significant reduction has been found in PD patients who carry the LRRK2 mutation. Novel HRV features have not been investigated in these individuals. We aimed to assess cardiac autonomic modulation through standard and novel approaches to HRV analysis in individuals who carry the LRRK2 G2019S mutation.METHODS: Short-term electrocardiograms were recorded in 14 LRRK2-associated PD patients, 25 LRRK2-non-manifesting carriers, 32 related non-carriers, 20 idiopathic PD patients, and 27 healthy controls. HRV measures were compared using regression modeling, controlling for age, sex, mean heart rate, and disease duration. Discriminant analysis highlighted the feature combination that best distinguished LRRK2-associated PD from controls.RESULTS: Beat-to-beat and global HRV measures were significantly increased in LRRK2-associated PD patients compared with controls (e.g., deceleration capacity of heart rate: p=0.006) and idiopathic PD patients (e.g., 8th standardized moment of the interbeat interval distribution: p=0.0003), respectively. LRRK2-associated PD patients also showed significantly increased irregularity of heart rate dynamics, as quantified by Renyi entropy, when compared with controls (p=0.002) and idiopathic PD patients (p=0.0004). Ordinal pattern statistics permitted the identification of LRRK2-associated PD individuals with 93% sensitivity and 93% specificity. Consistent results were found in a subgroup of LRRK2-non-manifesting carriers when compared with controls.CONCLUSIONS: Increased beat-to-beat HRV in LRRK2 G2019S mutation carriers compared with controls and idiopathic PD patients may indicate augmented cardiac autonomic cholinergic activity, suggesting early impairment of central vagal feedback loops in LRRK2-associated PD.
View details for DOI 10.1007/s10286-019-00632-w
View details for PubMedID 31444591
LRRK2 modifies alpha-syn pathology and spread in mouse models and human neurons
2019; 137 (6): 961–80
View details for DOI 10.1007/s00401-019-01995-0
View details for Web of Science ID 000468746900006
Alpha-Synuclein Physiology and Pathology: A Perspective on Cellular Structures and Organelles.
Frontiers in neuroscience
2019; 13: 1399
Alpha-synuclein (alpha-syn) is localized in cellular organelles of most neurons, but many of its physiological functions are only partially understood. alpha-syn accumulation is associated with Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy as well as other synucleinopathies; however, the exact pathomechanisms that underlie these neurodegenerative diseases remain elusive. In this review, we describe what is known about alpha-syn function and pathophysiological changes in different cellular structures and organelles, including what is known about its behavior as a prion-like protein. We summarize current knowledge of alpha-syn and its pathological forms, covering its effect on each organelle, including aggregation and toxicity in different model systems, with special interest on the mitochondria due to its relevance during the apoptotic process of dopaminergic neurons. Moreover, we explore the effect that alpha-syn exerts by interacting with chromatin remodeling proteins that add or remove histone marks, up-regulate its own expression, and resume the impairment that alpha-syn induces in vesicular traffic by interacting with the endoplasmic reticulum. We then recapitulate the events that lead to Golgi apparatus fragmentation, caused by the presence of alpha-syn. Finally, we report the recent findings about the accumulation of alpha-syn, indirectly produced by the endolysosomal system. In conclusion, many important steps into the understanding of alpha-syn have been made using in vivo and in vitro models; however, the time is right to start integrating observational studies with mechanistic models of alpha-syn interactions, in order to look at a more complete picture of the pathophysiological processes underlying alpha-synucleinopathies.
View details for DOI 10.3389/fnins.2019.01399
View details for PubMedID 32038126
LRRK2-mediated Rab10 phosphorylation in immune cells from Parkinson's disease patients.
Movement disorders : official journal of the Movement Disorder Society
BACKGROUND: Leucine-rich repeat kinase 2 is a potential therapeutic target for the treatment of Parkinson's disease, and clinical trials of leucine-rich repeat kinase 2 inhibitors are in development. The objective of this study was to evaluate phosphorylation of a new leucine-rich repeat kinase 2 substrate, Rab10, for potential use as a target engagement biomarker and/or patient enrichment biomarker for leucine-rich repeat kinase 2 inhibitor clinical trials.METHODS: Peripheral blood mononuclear cells and neutrophils were isolated from Parkinson's disease patients and matched controls, and treated ex vivo with a leucine-rich repeat kinase 2 inhibitor. Immunoblotting was used to measure levels of leucine-rich repeat kinase 2 and Rab10 and their phosphorylation. Plasma inflammatory cytokines were measured by multiplex enzyme-linked immunosorbent assay.RESULTS: Mononuclear cells and neutrophils of both controls and Parkinson's disease patients responded the same to leucine-rich repeat kinase 2 inhibitor treatment. Leucine-rich repeat kinase 2 levels in mononuclear cells were the same in controls and Parkinson's disease patients, whereas leucine-rich repeat kinase 2 was significantly increased in Parkinson's disease neutrophils. Rab10 T73 phosphorylation levels were similar in controls and Parkinson's disease patients and did not correlate with leucine-rich repeat kinase 2 levels. Immune-cell levels of leucine-rich repeat kinase 2 and Rab10 T73 phosphorylation were associated with plasma inflammatory cytokine levels.CONCLUSIONS: Rab10 T73 phosphorylation appears to be a valid target engagement biomarker for potential use in leucine-rich repeat kinase 2 inhibitor clinical trials. However, a lack of association between leucine-rich repeat kinase 2 and Rab10 phosphorylation complicates the potential use of Rab10 phosphorylation as a patient enrichment biomarker. Although replication is required, increased leucine-rich repeat kinase 2 levels in neutrophils from Parkinson's disease patients may have the potential for patient stratification. leucine-rich repeat kinase 2 activity in peripheral immune cells may contribute to an inflammatory phenotype. © 2018 International Parkinson and Movement Disorder Society.
View details for DOI 10.1002/mds.27601
View details for PubMedID 30597610
A pathway for Parkinson's Disease LRRK2 kinase to block primary cilia and Sonic hedgehog signaling in the brain.
Parkinson's disease-associated LRRK2 kinase phosphorylates multiple Rab GTPases, including Rab8A and Rab10. We show here that LRRK2 kinase interferes with primary cilia formation in cultured cells, human LRRK2 G2019S iPS cells and in the cortex of LRRK2 R1441C mice. Rab10 phosphorylation strengthens its intrinsic ability to block ciliogenesis by enhancing binding to RILPL1. Importantly, the ability of LRRK2 to interfere with ciliogenesis requires both Rab10 and RILPL1 proteins. Pathogenic LRRK2 influences the ability of cells to respond to cilia-dependent, Hedgehog signaling as monitored by Gli1 transcriptional activation. Moreover, cholinergic neurons in the striatum of LRRK2 R1441C mice show decreased ciliation, which will decrease their ability to sense Sonic hedgehog in a neuro-protective circuit that supports dopaminergic neurons. These data reveal a molecular pathway for regulating cilia function that likely contributes to Parkinson's disease-specific pathology.Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter).
View details for PubMedID 30398148
Genetic fine-mapping of the Iowan SNCA gene triplication in a patient with Parkinson's disease
NPJ PARKINSONS DISEASE
2018; 4: 18
The "Iowa kindred," a large Iowan family with autosomal-dominant Parkinson's disease, has been followed clinically since the 1920s at the Mayo Clinic. In 2003, the genetic cause was determined to be a 1.7 Mb triplication of the alpha-synuclein genomic locus. Affected individuals present with an early-onset, severe parkinsonism-dementia syndrome. Here, we present a descendant of the Iowa kindred with novel, disease-associated non-motor findings of reduced heart rate variability, complete anosmia, and a rare skin condition called colloid milium. At autopsy, key neuropathological findings were compatible with diffuse Lewy body disease. Using high-resolution comparative genomic hybridization (CGH) array analysis to fine-map the genomic breakpoints, we observed two independent recombination events of the SNCA locus that resulted in a genomic triplication of twelve genes, including SNCA, and the disruption of two genes, HERC6 and CCSER1, at the genomic breakpoints. In conclusion, we provide further evidence that the mere two-fold overexpression of alpha-synuclein leads to a fulminant alpha-synucleinopathy with rapid progression and severe clinical and neuropathological features.
View details for PubMedID 29928688
Clustering of Motor and Nonmotor Traits in Leucine-Rich Repeat Kinase 2 G2019S Parkinson's Disease Nonparkinsonian Relatives: A Multicenter Family Study
2018; 33 (6): 960–65
The objective of this study was to determine phenotypic features that differentiate nonparkinsonian first-degree relatives of PD leucine-rich repeat kinase 2 (LRRK2) G2019S multiplex families, regardless of carrier status, from healthy controls because nonparkinsonian individuals in multiplex families seem to share a propensity to present neurological features.We included nonparkinsonian first-degree relatives of LRRK2 G2019S familial PD cases and unrelated healthy controls participating in established multiplex family LRRK2 cohorts. Study participants underwent neurologic assessment including cognitive screening, olfaction testing, and questionnaires for daytime sleepiness, depression, and anxiety. We used a multiple logistic regression model with backward variable selection, validated with bootstrap resampling, to establish the best combination of motor and nonmotor features that differentiates nonparkinsonian first-degree relatives of LRRK2 G2019S familial PD cases from unrelated healthy controls.We included 142 nonparkinsonian family members and 172 unrelated healthy controls. The combination of past or current symptoms of anxiety (adjusted odds ratio, 4.16; 95% confidence interval, 2.01-8.63), less daytime sleepiness (adjusted odds ratio [1 unit], 0.90; 95% confidence interval, 0.83-0.97], and worse motor UPDRS score (adjusted odds ratio [1 unit], 1.4; 95% confidence interval, 1.20-1.67) distinguished nonparkinsonian family members, regardless of LRRK2 G2019S mutation status, from unrelated healthy controls. The model accuracy was good (area under the curve = 79.3%).A set of motor and nonmotor features distinguishes first-degree relatives of LRRK2 G2019S probands, regardless of mutation status, from unrelated healthy controls. Environmental or non-LRRK2 genetic factors in LRRK2-associated PD may influence penetrance of the LRRK2 G2019S mutation. The relationship of these features to actual PD risk requires longitudinal observation of LRRK2 familial PD cohorts. © 2018 International Parkinson and Movement Disorder Society.
View details for DOI 10.1002/mds.27272
View details for Web of Science ID 000439829100012
View details for PubMedID 29665080
Advancing Stem Cell Models of Alpha-Synuclein Gene Regulation in Neurodegenerative Disease
FRONTIERS IN NEUROSCIENCE
2018; 12: 199
Alpha-synuclein (non A4 component of amyloid precursor, SNCA, NM_000345.3) plays a central role in the pathogenesis of Parkinson's disease (PD) and related Lewy body disorders such as Parkinson's disease dementia, Lewy body dementia, and multiple system atrophy. Since its discovery as a disease-causing gene in 1997, alpha-synuclein has been a central point of scientific interest both at the protein and gene level. Mutations, including copy number variants, missense mutations, short structural variants, and single nucleotide polymorphisms, can be causative for PD and affect conformational changes of the protein, can contribute to changes in expression of alpha-synuclein and its isoforms, and can influence regulation of temporal as well as spatial levels of alpha-synuclein in different tissues and cell types. A lot of progress has been made to understand both the physiological transcriptional and epigenetic regulation of the alpha-synuclein gene and whether changes in transcriptional regulation could lead to disease and neurodegeneration in PD and related alpha-synucleinopathies. Although the histopathological changes in these neurodegenerative disorders are similar, the temporal and spatial presentation and progression distinguishes them which could be in part due to changes or disruption of transcriptional regulation of alpha-synuclein. In this review, we describe different genetic alterations that contribute to PD and neurodegenerative conditions and review aspects of transcriptional regulation of the alpha-synuclein gene in the context of the development of PD. New technologies, advanced gene engineering and stem cell modeling, are on the horizon to shed further light on a better understanding of gene regulatory processes and exploit them for therapeutic developments.
View details for DOI 10.3389/fnins.2018.00199
View details for Web of Science ID 000429480400001
View details for PubMedID 29686602
View details for PubMedCentralID PMC5900030
LRRK2 p. Ile1371Val Mutation in a Case with Neuropathologically Confirmed Multi-System Atrophy
JOURNAL OF PARKINSONS DISEASE
2018; 8 (1): 93–100
Mutations in the leucine rich repeat kinase 2 (LRRK2) gene are among the most common genetic causes of Lewy body Parkinson's disease (PD). However, LRRK2 mutations can also lead to a variety of pathological phenotypes other than typical PD, including relatively pure nigrostriatal cell loss without alpha-synuclein-positive Lewy bodies or Lewy neurites, progressive supranuclear palsy (PSP), and multiple system atrophy (MSA). The mechanisms behind this remarkable pleomorphic pathology are currently unclear.To genetically and pathologically characterize a case with a LRRK2, p.Ile1371Val rare variant and pathologically proven MSA.From the brain donation program at the Parkinson's Institute and Clinical Center, we selected 26 brains with family history and a with clinicopathological diagnosis of PD (n = 20), MSA (n = 4), or PSP (n = 2). We performed neuropathological evaluation, including alpha-synuclein and tau immunohistochemistry and sequenced 188 genes that have been reported as causative for or associated with neurodegenerative diseases.We identified a known LRRK2, p.Ile1371Val genetic variant in a case with clinically diagnosed and pathologically proven MSA. Neuropathology revealed that the olivopontocerebellar system was more affected than the striatonigral system.Our data suggest that genetic variants in the LRRK2 gene can present clinically and neuropathologically as MSA. One other LRRK2 genetic variant (LRRK2, p.Ile2020Thr) has been reported with a neuropathological diagnosis of MSA. Interestingly, the LRRK2 variant (LRRK2, p.Ile1371Val) identified here has been reported previously in a postmortem case with Lewy body PD.Future studies are critical to discover the mechanisms leading to different neurodegenerative trajectories both in neuronal and glial cell populations.
View details for DOI 10.3233/JPD-171237
View details for Web of Science ID 000425657100009
View details for PubMedID 29480226
LRRK2 G2019S-induced mitochondrial DNA damage is LRRK2 kinase dependent and inhibition restores mtDNA integrity in Parkinson's disease
HUMAN MOLECULAR GENETICS
2017; 26 (22): 4340–51
Mutations in leucine-rich repeat kinase 2 (LRRK2) are associated with increased risk for developing Parkinson's disease (PD). Previously, we found that LRRK2 G2019S mutation carriers have increased mitochondrial DNA (mtDNA) damage and after zinc finger nuclease-mediated gene mutation correction, mtDNA damage was no longer detectable. While the mtDNA damage phenotype can be unambiguously attributed to the LRRK2 G2019S mutation, the underlying mechanism(s) is unknown. Here, we examine the role of LRRK2 kinase function in LRRK2 G2019S-mediated mtDNA damage, using both genetic and pharmacological approaches in cultured neurons and PD patient-derived cells. Expression of LRRK2 G2019S induced mtDNA damage in primary rat midbrain neurons, but not in cortical neuronal cultures. In contrast, the expression of LRRK2 wild type or LRRK2 D1994A mutant (kinase dead) had no effect on mtDNA damage in either midbrain or cortical neuronal cultures. In addition, human LRRK2 G2019S patient-derived lymphoblastoid cell lines (LCL) demonstrated increased mtDNA damage relative to age-matched controls. Importantly, treatment of LRRK2 G2019S expressing midbrain neurons or patient-derived LRRK2 G2019S LCLs with the LRRK2 kinase inhibitor GNE-7915, either prevented or restored mtDNA damage to control levels. These findings support the hypothesis that LRRK2 G2019S-induced mtDNA damage is LRRK2 kinase activity dependent, uncovering a novel pathological role for this kinase. Blocking or reversing mtDNA damage via LRRK2 kinase inhibition or other therapeutic approaches may be useful to slow PD-associated pathology.
View details for DOI 10.1093/hmg/ddx320
View details for Web of Science ID 000414403900003
View details for PubMedID 28973664
View details for PubMedCentralID PMC5886254
Penetrance Estimate of LRRK2 p.G2019S Mutation in Individuals of Non-Ashkenazi Jewish Ancestry
2017; 32 (10): 1432–38
Penetrance estimates of the leucine-rich repeat kinase 2 (LRRK2) p.G2019S mutation for PD vary widely (24%-100%). The p.G2019S penetrance in individuals of Ashkenazi Jewish ancestry has been estimated as 25%, adjusted for multiple covariates. It is unknown whether penetrance varies among different ethnic groups. The objective of this study was to estimate the penetrance of p.G2019S in individuals of non-Ashkenazi Jewish ancestry and compare penetrance between Ashkenazi Jews and non-Ashkenazi Jews to age 80.The kin-cohort method was used to estimate penetrance in 474 first-degree relatives of 69 non-Ashkenazi Jewish LRRK2 p.G2019S carrier probands at 8 sites from the Michael J. Fox LRRK2 Cohort Consortium. An identical validated family history interview was administered to assess age at onset of PD, current age, or age at death for relatives in different ethnic groups at each site. Neurological examination and LRRK2 genotype of relatives were included when available.Risk of PD in non-Ashkenazi Jewish relatives who carry a LRRK2 p.G2019S mutation was 42.5% (95% confidence interval [CI]: 26.3%-65.8%) to age 80, which is not significantly higher than the previously estimated 25% (95% CI: 16.7%-34.2%) in Ashkenazi Jewish carrier relatives. The penetrance of PD to age 80 in LRRK2 p.G2019S mutation carrier relatives was significantly higher than the noncarrier relatives, as seen in Ashkenazi Jewish relatives.The similar penetrance of LRRK2 p.G2019S estimated in Ashkenazi Jewish carriers and non-Ashkenazi Jewish carriers confirms that p.G2019S penetrance is 25% to 42.5% at age 80 in all populations analyzed. © 2017 International Parkinson and Movement Disorder Society.
View details for DOI 10.1002/mds.27059
View details for Web of Science ID 000412934600016
View details for PubMedID 28639421
View details for PubMedCentralID PMC5656509
Parkinson's disease associated with pure ATXN10 repeat expansion
NPJ PARKINSONS DISEASE
2017; 3: 27
Large, non-coding pentanucleotide repeat expansions of ATTCT in intron 9 of the ATXN10 gene typically cause progressive spinocerebellar ataxia with or without seizures and present neuropathologically with Purkinje cell loss resulting in symmetrical cerebellar atrophy. These ATXN10 repeat expansions can be interrupted by sequence motifs which have been attributed to seizures and are likely to act as genetic modifiers. We identified a Mexican kindred with multiple affected family members with ATXN10 expansions. Four affected family members showed clinical features of spinocerebellar ataxia type 10 (SCA10). However, one affected individual presented with early-onset levodopa-responsive parkinsonism, and one family member carried a large repeat ATXN10 expansion, but was clinically unaffected. To characterize the ATXN10 repeat, we used a novel technology of single-molecule real-time (SMRT) sequencing and CRISPR/Cas9-based capture. We sequenced the entire span of ~5.3-7.0 kb repeat expansions. The Parkinson's patient carried an ATXN10 expansion with no repeat interruption motifs as well as an unaffected sister. In the siblings with typical SCA10, we found a repeat pattern of ATTCC repeat motifs that have not been associated with seizures previously. Our data suggest that the absence of repeat interruptions is likely a genetic modifier for the clinical presentation of l-Dopa responsive parkinsonism, whereas repeat interruption motifs contribute clinically to epilepsy. Repeat interruptions are important genetic modifiers of the clinical phenotype in SCA10. Advanced sequencing techniques now allow to better characterize the underlying genetic architecture for determining accurate phenotype-genotype correlations.
View details for DOI 10.1038/s41531-017-0029-x
View details for Web of Science ID 000411056400001
View details for PubMedID 28890930
View details for PubMedCentralID PMC5585403
beta(2)-Adrenoreceptor is a regulator of the alpha-synuclein gene driving risk of Parkinson's disease
2017; 357 (6354): 891–97
Copy number mutations implicate excess production of α-synuclein as a possibly causative factor in Parkinson's disease (PD). Using an unbiased screen targeting endogenous gene expression, we discovered that the β2-adrenoreceptor (β2AR) is a regulator of the α-synuclein gene (SNCA). β2AR ligands modulate SNCA transcription through histone 3 lysine 27 acetylation of its promoter and enhancers. Over 11 years of follow-up in 4 million Norwegians, the β2AR agonist salbutamol, a brain-penetrant asthma medication, was associated with reduced risk of developing PD (rate ratio, 0.66; 95% confidence interval, 0.58 to 0.76). Conversely, a β2AR antagonist correlated with increased risk. β2AR activation protected model mice and patient-derived cells. Thus, β2AR is linked to transcription of α-synuclein and risk of PD in a ligand-specific fashion and constitutes a potential target for therapies.
View details for DOI 10.1126/science.aaf3934
View details for Web of Science ID 000408734900037
View details for PubMedID 28860381
View details for PubMedCentralID PMC5761666
Heart Rate Variability in leucine-rich repeat kinase 2-Associated Parkinson's Disease
2017; 32 (4): 610–14
Heart rate variability is reduced in idiopathic PD, indicating cardiac autonomic dysfunction likely resulting from peripheral autonomic synucleinopathy. Little is known about heart rate variability in leucine-rich repeat kinase 2-associated PD.This study investigated heart rate variability in LRRK2-associated PD.Resting electrocardiograms were obtained from 20 individuals with LRRK2-associated PD, 37 nonmanifesting carriers, 48 related noncarriers, 26 idiopathic PD patients, and 32 controls. Linear regression modelling compared time and frequency domain values, adjusting for age, sex, heart rate, and disease duration.Low-frequency power and the ratio of low-high frequency power were reduced in idiopathic PD versus controls (P < .008, P < .029 respectively). In contrast, individuals with LRRK2-associated PD were not statistically different from controls in any parameter measured. Furthermore, all parameters trended toward being higher in LRRK2-associated PD when compared with idiopathic PD.Heart rate variability may remain intact in LRRK2-associated PD, adding to a growing literature supporting clinical-pathologic differences between LRRK2-associated and idiopathic PD. © 2017 International Parkinson and Movement Disorder Society.
View details for DOI 10.1002/mds.26896
View details for Web of Science ID 000399758800020
View details for PubMedID 28071824
Inflammatory profile discriminates clinical subtypes in LRRK2-associated Parkinson's disease
EUROPEAN JOURNAL OF NEUROLOGY
2017; 24 (2): 427–E6
The presentation of Parkinson's disease patients with mutations in the LRRK2 gene (PDLRRK2 ) is highly variable, suggesting a strong influence of modifying factors. In this context, inflammation is a potential candidate inducing clinical subtypes.An extensive battery of peripheral inflammatory markers was measured in human serum in a multicentre cohort of 142 PDLRRK2 patients from the MJFF LRRK2 Consortium, stratified by three different subtypes as recently proposed for idiopathic Parkinson's disease: diffuse/malignant, intermediate and mainly pure motor.Patients classified as diffuse/malignant presented with the highest levels of the pro-inflammatory proteins interleukin 8 (IL-8), monocyte chemotactic protein 1 (MCP-1) and macrophage inflammatory protein 1-β (MIP-1-β) paralleled by high levels of the neurotrophic protein brain-derived neurotrophic factor (BDNF). It was also possible to distinguish the clinical subtypes based on their inflammatory profile by using discriminant and area under the receiver operating characteristic curve analysis.Inflammation seems to be associated with the presence of a specific clinical subtype in PDLRRK2 that is characterized by a broad and more severely affected spectrum of motor and non-motor symptoms. The pro-inflammatory metabolites IL-8, MCP-1 and MIP-1-β as well as BDNF are interesting candidates to be included in biomarker panels that aim to differentiate subtypes in PDLRRK2 and predict progression.
View details for DOI 10.1111/ene.13223
View details for Web of Science ID 000392804300032
View details for PubMedID 28102045
Derivation of Leptomeninges Explant Cultures from Postmortem Human Brain Donors
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
Even though great progress has been made in the clinical characterization of Parkinson's disease, several studies report that the diagnosis of Parkinson's disease is not pathologically confirmed in up to 25% of clinically diagnosed Parkinson's disease. Therefore, tissue collected from clinically diagnosed patients with idiopathic Parkinson's disease can have a high rate of misdiagnosis; hence in vitro studies from such tissues to study Parkinson's disease as a preclinical model can become futile. By collecting postmortem human leptomeninges with a confirmed neuropathological diagnosis of Parkinson's disease and characterized by nigrostriatal cell loss and intracellular protein inclusions called Lewy bodies, one can be certain that clinically observed parkinsonism is not caused by another underlying disease process (e.g. tumor, arteriosclerosis). This protocol presents the dissection and preparation of postmortem human leptomeninges for derivation of a meningeal fibroblast culture. This procedure is robust and has a high success rate. The challenge of the culture is sterility as the brain procurement is generally not performed under sterile conditions. Therefore, it is important to supplement the culture media with a cocktail of penicillin, streptomycin, and amphotericin B. The derivation of meningeal fibroblasts from autopsy-confirmed cases with Parkinson's disease provides the foundation for in vitro modeling of Parkinson's disease. Meningeal fibroblasts appear 3-9 days after sample preparation and about 20-30 million cells can be cryopreserved in 6-8 weeks. The meningeal fibroblast culture is homogenous and the cells express fibronectin, a commonly used marker to identify meninges.
View details for DOI 10.3791/55045
View details for Web of Science ID 000397847200042
View details for PubMedID 28190070
View details for PubMedCentralID PMC5352281
Functional Impairment in Miro Degradation and Mitophagy Is a Shared Feature in Familial and Sporadic Parkinson's Disease
CELL STEM CELL
2016; 19 (6): 709-724
Mitochondrial movements are tightly controlled to maintain energy homeostasis and prevent oxidative stress. Miro is an outer mitochondrial membrane protein that anchors mitochondria to microtubule motors and is removed to stop mitochondrial motility as an early step in the clearance of dysfunctional mitochondria. Here, using human induced pluripotent stem cell (iPSC)-derived neurons and other complementary models, we build on a previous connection of Parkinson's disease (PD)-linked PINK1 and Parkin to Miro by showing that a third PD-related protein, LRRK2, promotes Miro removal by forming a complex with Miro. Pathogenic LRRK2G2019S disrupts this function, delaying the arrest of damaged mitochondria and consequently slowing the initiation of mitophagy. Remarkably, partial reduction of Miro levels in LRRK2G2019S human neuron and Drosophila PD models rescues neurodegeneration. Miro degradation and mitochondrial motility are also impaired in sporadic PD patients. We reveal that prolonged retention of Miro, and the downstream consequences that ensue, may constitute a central component of PD pathogenesis.
View details for DOI 10.1016/j.stem.2016.08.002
View details for Web of Science ID 000389474900010
View details for PubMedCentralID PMC5135570
Inflammatory profile in LRRK2-associated prodromal and clinical PD
JOURNAL OF NEUROINFLAMMATION
2016; 13: 122
There is evidence for a relevant role of inflammation in the pathogenesis of Parkinson's disease (PD). Mutations in the LRRK2 gene represent the most frequent genetic cause for autosomal dominant PD. LRRK2 is highly expressed in macrophages and microglia suggesting an involvement in inflammatory pathways. The objectives are to test (1) whether idiopathic PD and LRRK2-associated PD share common inflammatory pathways or present distinct profiles and (2) whether non-manifesting LRRK2 mutation carriers present with similar aspects of inflammatory profiles as seen in PD-affected patients.We assessed serum profiles of 23 immune-associated markers and the brain-derived neurotrophic factor in 534 individuals from the MJFF LRRK2 consortium.A large proportion of inflammatory markers were gender-dependent. Both PD-affected cohorts showed increased levels of the pro-inflammatory marker fatty-acid-binding protein. Additionally, idiopathic PD but not LRRK2-associated PD patients showed increased levels of the pro-inflammatory marker interleukin-12-p40 as well as the anti-inflammatory species interleukin-10, brain-derived neurotrophic factor, and stem cell factor. Non-manifesting LRRK2 mutation carriers including those with prodromal characteristics of PD presented with control-like inflammatory profiles.Concomitant inflammation seems to be associated with idiopathic and LRRK2-associated PD. Identifying PD patients in whom inflammatory processes play a major role in their pathophysiology might offer a new therapeutic window at least for a subgroup of patients. Since non-manifesting LRRK2 mutation carriers with symptoms of the prodromal phase of PD did not show inflammatory profiles, activation of the immune system seems not an early event in the disease cascade.
View details for DOI 10.1186/s12974-016-0588-5
View details for Web of Science ID 000377286500007
View details for PubMedID 27220776
View details for PubMedCentralID PMC4879729
IS PARKIN PARKINSONISM A CANCER PREDISPOSITION SYNDROME?
2015; 1 (4): e31
Mutations in the PARKIN gene (chromosome 6q25-27) were first described in 1998 in families with "juvenile" autosomal recessive parkinsonism. More than 180 causative variants in the PARKIN gene have been identified; point mutations and copy number variants (i.e., exon deletions or duplications) occur at nearly equal frequencies.(1) PARKIN is one of the largest genes in the human genome (1.3 Mb) and contains a chromosomal fragile site (CFS) FRA6E (6q26) between exons 2 and 8. This is of interest regarding the etiology of cancer because CFSs are prone to spontaneous breaks leading to chromosome alterations. Therefore, it is not surprising that PARKIN mutations have also been found in various cancer cell lines and primary tumors.(2,3) Mutations in PARKIN show decreased PARKIN E3 ligase function with resultant accumulation of cyclin E, creating the potential for mitotic instability in dividing cells.
View details for DOI 10.1212/NXG.0000000000000031
View details for Web of Science ID 000445305700005
View details for PubMedID 27066568
View details for PubMedCentralID PMC4811379
Multisystem Lewy body disease and the other parkinsonian disorders
2015; 47 (12): 1378–84
Here we prioritize as multisystem Lewy body disease (MLBD) those genetic forms of Parkinson's disease that point the way toward a mechanistic understanding of the majority of sporadic disease. Pathological diagnosis of genetic subtypes offers the prospect of distinguishing different mechanistic trajectories with a common mutational etiology, differing outcomes from varying allelic bases, and those disease-associated variants that can be used in gene-environment analysis. Clearly delineating parkinsonian disorders into subclasses on the basis of molecular mechanisms with well-characterized outcome expectations is the basis for refining these forms of neurodegeneration as research substrate through the use of cell models derived from affected individuals while ensuring that clinically collected data can be used for therapeutic decisions and research without increasing the noise and confusion engendered by the collection of data against a range of historically defined criteria.
View details for DOI 10.1038/ng.3454
View details for Web of Science ID 000365813200006
View details for PubMedID 26620112
Elevated alpha-synuclein caused by SNCA gene triplication impairs neuronal differentiation and maturation in Parkinson's patient-derived induced pluripotent stem cells
CELL DEATH & DISEASE
2015; 6: e1994
We have assessed the impact of α-synuclein overexpression on the differentiation potential and phenotypic signatures of two neural-committed induced pluripotent stem cell lines derived from a Parkinson's disease patient with a triplication of the human SNCA genomic locus. In parallel, comparative studies were performed on two control lines derived from healthy individuals and lines generated from the patient iPS-derived neuroprogenitor lines infected with a lentivirus incorporating a small hairpin RNA to knock down the SNCA mRNA. The SNCA triplication lines exhibited a reduced capacity to differentiate into dopaminergic or GABAergic neurons and decreased neurite outgrowth and lower neuronal activity compared with control cultures. This delayed maturation phenotype was confirmed by gene expression profiling, which revealed a significant reduction in mRNA for genes implicated in neuronal differentiation such as delta-like homolog 1 (DLK1), gamma-aminobutyric acid type B receptor subunit 2 (GABABR2), nuclear receptor related 1 protein (NURR1), G-protein-regulated inward-rectifier potassium channel 2 (GIRK-2) and tyrosine hydroxylase (TH). The differentiated patient cells also demonstrated increased autophagic flux when stressed with chloroquine. We conclude that a two-fold overexpression of α-synuclein caused by a triplication of the SNCA gene is sufficient to impair the differentiation of neuronal progenitor cells, a finding with implications for adult neurogenesis and Parkinson's disease progression, particularly in the context of bioenergetic dysfunction.
View details for DOI 10.1038/cddis.2015.318
View details for Web of Science ID 000367155300027
View details for PubMedID 26610207
View details for PubMedCentralID PMC4670926
Comparative Genomic Hybridization Solves a 14-Year-Old PARKIN Mystery
ANNALS OF NEUROLOGY
2015; 78 (4): 663–64
View details for DOI 10.1002/ana.24478
View details for Web of Science ID 000362668100020
View details for PubMedID 26179350
Clinical Correlations With Lewy Body Pathology in LRRK2-Related Parkinson Disease
2015; 72 (1): 100–105
Mutations in leucine-rich repeat kinase 2 (LRRK2) are the most common cause of genetic Parkinson disease (PD) known to date. The clinical features of manifesting LRRK2 mutation carriers are generally indistinguishable from those of patients with sporadic PD. However, some PD cases associated with LRRK2 mutations lack Lewy bodies (LBs), a neuropathological hallmark of PD. We investigated whether the presence or absence of LBs correlates with different clinical features in LRRK2-related PD.We describe genetic, clinical, and neuropathological findings of 37 cases of LRRK2-related PD including 33 published and 4 unpublished cases through October 2013. Among the different mutations, the LRRK2 p.G2019S mutation was most frequently associated with LB pathology. Nonmotor features of cognitive impairment/dementia, anxiety, and orthostatic hypotension were correlated with the presence of LBs. In contrast, a primarily motor phenotype was associated with a lack of LBs.To our knowledge, this is the first report of clinicopathological correlations in a series of LRRK2-related PD cases. Findings from this selected group of patients with PD demonstrated that parkinsonian motor features can occur in the absence of LBs. However, LB pathology in LRRK2-related PD may be a marker for a broader parkinsonian symptom complex including cognitive impairment.
View details for DOI 10.1001/jamaneurol.2014.2704
View details for Web of Science ID 000347815300016
View details for PubMedID 25401511
View details for PubMedCentralID PMC4399368
Higher Vulnerability and Stress Sensitivity of Neuronal Precursor Cells Carrying an Alpha-Synuclein Gene Triplication
2014; 9 (11): e112413
Parkinson disease (PD) is a multi-factorial neurodegenerative disorder with loss of dopaminergic neurons in the substantia nigra and characteristic intracellular inclusions, called Lewy bodies. Genetic predisposition, such as point mutations and copy number variants of the SNCA gene locus can cause very similar PD-like neurodegeneration. The impact of altered α-synuclein protein expression on integrity and developmental potential of neuronal stem cells is largely unexplored, but may have wide ranging implications for PD manifestation and disease progression. Here, we investigated if induced pluripotent stem cell-derived neuronal precursor cells (NPCs) from a patient with Parkinson's disease carrying a genomic triplication of the SNCA gene (SNCA-Tri). Our goal was to determine if these cells these neuronal precursor cells already display pathological changes and impaired cellular function that would likely predispose them when differentiated to neurodegeneration. To achieve this aim, we assessed viability and cellular physiology in human SNCA-Tri NPCs both under normal and environmentally stressed conditions to model in vitro gene-environment interactions which may play a role in the initiation and progression of PD. Human SNCA-Tri NPCs displayed overall normal cellular and mitochondrial morphology, but showed substantial changes in growth, viability, cellular energy metabolism and stress resistance especially when challenged by starvation or toxicant challenge. Knockdown of α-synuclein in the SNCA-Tri NPCs by stably expressed short hairpin RNA (shRNA) resulted in reversal of the observed phenotypic changes. These data show for the first time that genetic alterations such as the SNCA gene triplication set the stage for decreased developmental fitness, accelerated aging, and increased neuronal cell loss. The observation of this "stem cell pathology" could have a great impact on both quality and quantity of neuronal networks and could provide a powerful new tool for development of neuroprotective strategies for PD.
View details for DOI 10.1371/journal.pone.0112413
View details for Web of Science ID 000349144400078
View details for PubMedID 25390032
View details for PubMedCentralID PMC4229205
Systems-Based Analyses of Brain Regions Functionally Impacted in Parkinson's Disease Reveals Underlying Causal Mechanisms
2014; 9 (8): e102909
Detailed analysis of disease-affected tissue provides insight into molecular mechanisms contributing to pathogenesis. Substantia nigra, striatum, and cortex are functionally connected with increasing degrees of alpha-synuclein pathology in Parkinson's disease. We undertook functional and causal pathway analysis of gene expression and proteomic alterations in these three regions, and the data revealed pathways that correlated with disease progression. In addition, microarray and RNAseq experiments revealed previously unidentified causal changes related to oligodendrocyte function and synaptic vesicle release, and these and other changes were reflected across all brain regions. Importantly, subsets of these changes were replicated in Parkinson's disease blood; suggesting peripheral tissue may provide important avenues for understanding and measuring disease status and progression. Proteomic assessment revealed alterations in mitochondria and vesicular transport proteins that preceded gene expression changes indicating defects in translation and/or protein turnover. Our combined approach of proteomics, RNAseq and microarray analyses provides a comprehensive view of the molecular changes that accompany functional loss and alpha-synuclein pathology in Parkinson's disease, and may be instrumental to understand, diagnose and follow Parkinson's disease progression.
View details for DOI 10.1371/journal.pone.0102909
View details for Web of Science ID 000341127500006
View details for PubMedID 25170892
View details for PubMedCentralID PMC4149353
Michael J. Fox Foundation LRRK2 Consortium: geographical differences in returning genetic research data to study participants
GENETICS IN MEDICINE
2014; 16 (8): 644–45
View details for DOI 10.1038/gim.2014.55
View details for Web of Science ID 000340024500013
View details for PubMedID 25093570
View details for PubMedCentralID PMC4682659
DICE, an efficient system for iterative genomic editing in human pluripotent stem cells.
Nucleic acids research
2014; 42 (5)
To reveal the full potential of human pluripotent stem cells, new methods for rapid, site-specific genomic engineering are needed. Here, we describe a system for precise genetic modification of human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). We identified a novel human locus, H11, located in a safe, intergenic, transcriptionally active region of chromosome 22, as the recipient site, to provide robust, ubiquitous expression of inserted genes. Recipient cell lines were established by site-specific placement of a 'landing pad' cassette carrying attP sites for phiC31 and Bxb1 integrases at the H11 locus by spontaneous or TALEN-assisted homologous recombination. Dual integrase cassette exchange (DICE) mediated by phiC31 and Bxb1 integrases was used to insert genes of interest flanked by phiC31 and Bxb1 attB sites at the H11 locus, replacing the landing pad. This system provided complete control over content, direction and copy number of inserted genes, with a specificity of 100%. A series of genes, including mCherry and various combinations of the neural transcription factors LMX1a, FOXA2 and OTX2, were inserted in recipient cell lines derived from H9 ESC, as well as iPSC lines derived from a Parkinson's disease patient and a normal sibling control. The DICE system offers rapid, efficient and precise gene insertion in ESC and iPSC and is particularly well suited for repeated modifications of the same locus.
View details for DOI 10.1093/nar/gkt1290
View details for PubMedID 24304893
LRRK2 mutations cause mitochondrial DNA damage in iPSC-derived neural cells from Parkinson's disease patients: Reversal by gene correction
NEUROBIOLOGY OF DISEASE
2014; 62: 381–86
Parkinson's disease associated mutations in leucine rich repeat kinase 2 (LRRK2) impair mitochondrial function and increase the vulnerability of induced pluripotent stem cell (iPSC)-derived neural cells from patients to oxidative stress. Since mitochondrial DNA (mtDNA) damage can compromise mitochondrial function, we examined whether LRRK2 mutations can induce damage to the mitochondrial genome. We found greater levels of mtDNA damage in iPSC-derived neural cells from patients carrying homozygous or heterozygous LRRK2 G2019S mutations, or at-risk individuals carrying the heterozygous LRRK2 R1441C mutation, than in cells from unrelated healthy subjects who do not carry LRRK2 mutations. After zinc finger nuclease-mediated repair of the LRRK2 G2019S mutation in iPSCs, mtDNA damage was no longer detected in differentiated neuroprogenitor and neural cells. Our results unambiguously link LRRK2 mutations to mtDNA damage and validate a new cellular phenotype that can be used for examining pathogenic mechanisms and screening therapeutic strategies.
View details for DOI 10.1016/j.nbd.2013.10.013
View details for Web of Science ID 000330553600035
View details for PubMedID 24148854
View details for PubMedCentralID PMC3877733
Discovery of functional non-coding conserved regions in the alpha-synuclein gene locus.
2014; 3: 259
Several single nucleotide polymorphisms (SNPs) and the Rep-1 microsatellite marker of the alpha-synuclein ( SNCA) gene have consistently been shown to be associated with Parkinson's disease, but the functional relevance is unclear. Based on these findings we hypothesized that conserved cis-regulatory elements in the SNCA genomic region regulate expression of SNCA, and that SNPs in these regions could be functionally modulating the expression of SNCA, thus contributing to neuronal demise and predisposing to Parkinson's disease. In a pair-wise comparison of a 206kb genomic region encompassing the SNCA gene, we revealed 34 evolutionary conserved DNA sequences between human and mouse. All elements were cloned into reporter vectors and assessed for expression modulation in dual luciferase reporter assays. We found that 12 out of 34 elements exhibited either an enhancement or reduction of the expression of the reporter gene. Three elements upstream of the SNCA gene displayed an approximately 1.5 fold (p<0.009) increase in expression. Of the intronic regions, three showed a 1.5 fold increase and two others indicated a 2 and 2.5 fold increase in expression (p<0.002). Three elements downstream of the SNCA gene showed 1.5 fold and 2.5 fold increase (p<0.0009). One element downstream of SNCA had a reduced expression of the reporter gene of 0.35 fold (p<0.0009) of normal activity. Our results demonstrate that the SNCA gene contains cis-regulatory regions that might regulate the transcription and expression of SNCA. Further studies in disease-relevant tissue types will be important to understand the functional impact of regulatory regions and specific Parkinson's disease-associated SNPs and its function in the disease process.
View details for DOI 10.12688/f1000research.3281.2
View details for PubMedID 25566351
Identification and Rescue of alpha-Synuclein Toxicity in Parkinson Patient-Derived Neurons
2013; 342 (6161): 983–87
The induced pluripotent stem (iPS) cell field holds promise for in vitro disease modeling. However, identifying innate cellular pathologies, particularly for age-related neurodegenerative diseases, has been challenging. Here, we exploited mutation correction of iPS cells and conserved proteotoxic mechanisms from yeast to humans to discover and reverse phenotypic responses to α-synuclein (αsyn), a key protein involved in Parkinson's disease (PD). We generated cortical neurons from iPS cells of patients harboring αsyn mutations, who are at high risk of developing PD dementia. Genetic modifiers from unbiased screens in a yeast model of αsyn toxicity led to identification of early pathogenic phenotypes in patient neurons. These included nitrosative stress, accumulation of endoplasmic reticulum (ER)-associated degradation substrates, and ER stress. A small molecule identified in a yeast screen (NAB2), and the ubiquitin ligase Nedd4 it affects, reversed pathologic phenotypes in these neurons.
View details for DOI 10.1126/science.1245296
View details for Web of Science ID 000327223500044
View details for PubMedID 24158904
View details for PubMedCentralID PMC4022187
Elevated Alpha-Synuclein Impairs Innate Immune Cell Function and Provides a Potential Peripheral Biomarker for Parkinson's Disease
2013; 8 (8): e71634
Alpha-synuclein protein is strongly implicated in the pathogenesis Parkinson's disease. Increased expression of α-synuclein due to genetic multiplication or point mutations leads to early onset disease. While α-synuclein is known to modulate membrane vesicle dynamics, it is not clear if this activity is involved in the pathogenic process or if measurable physiological effects of α-synuclein over-expression or mutation exist in vivo. Macrophages and microglia isolated from BAC α-synuclein transgenic mice, which overexpress α-synuclein under regulation of its own promoter, express α-synuclein and exhibit impaired cytokine release and phagocytosis. These processes were affected in vivo as well, both in peritoneal macrophages and microglia in the CNS. Extending these findings to humans, we found similar results with monocytes and fibroblasts isolated from idiopathic or familial Parkinson's disease patients compared to age-matched controls. In summary, this paper provides 1) a new animal model to measure α-synuclein dysfunction; 2) a cellular system to measure synchronized mobilization of α-synuclein and its functional interactions; 3) observations regarding a potential role for innate immune cell function in the development and progression of Parkinson's disease and other human synucleinopathies; 4) putative peripheral biomarkers to study and track these processes in human subjects. While altered neuronal function is a primary issue in PD, the widespread consequence of abnormal α-synuclein expression in other cell types, including immune cells, could play an important role in the neurodegenerative progression of PD and other synucleinopathies. Moreover, increased α-synuclein and altered phagocytosis may provide a useful biomarker for human PD.
View details for DOI 10.1371/journal.pone.0071634
View details for Web of Science ID 000324403200014
View details for PubMedID 24058406
View details for PubMedCentralID PMC3751933
Skin Punch Biopsy Explant Culture for Derivation of Primary Human Fibroblasts
JOVE-JOURNAL OF VISUALIZED EXPERIMENTS
Tissues and cell lines derived from an individual with disease are ideal sources to study disease-related cellular phenotypes. Patient-derived fibroblasts in this protocol have been successfully used in the derivation of induced pluripotent stem cells to model disease(1). Early passages of these fibroblasts can also be used for cell-based functional assays to study specific disease pathways, mechanisms(2) and subsequent drug screening approaches. The advantage of the presented protocol over enzymatic procedures are 1) the reproducibility of the technique from small amounts of tissue derived from older patients, e.g. patients affected with Parkinson's disease, 2) the technically simple approach over more challenging methodologies using enzymatic treatments, and 3) the time consideration: this protocol takes 15-20 min and can be performed immediately after biopsy arrival. Enzymatic treatments can take up to 4 hr and have the problems of overdigestion, reduction of cell viability and subsequent attachment of cells when not handled properly. This protocol describes the dissection and preparation of a 4-mm human skin biopsy for derivation of a fibroblast culture and has a very high success rate which is important when dealing with patient-derived tissue samples. In this culture, keratinocytes migrate out of the biopsy tissue within the first week after preparation. Fibroblasts appear 7-10 days after the first outgrowth of keratinocytes. DMEM high glucose media supplemented with 20% FBS favors the growth of fibroblasts over keratinocytes and fibroblasts will overgrow the keratinocytes. After 2 passages keratinocytes have been diluted out resulting in relatively homogenous fibroblast cultures which expresses the fibroblast marker SERPINH1 (HSP-47). Using this approach, 15-20 million fibroblasts can be derived in 4-8 weeks for cell banking. The skin dissection takes about 15-20 min, cells are then monitored once a day under the microscope, and media is changed every 2-3 days after attachment and outgrowth of cells.
View details for DOI 10.3791/3779
View details for Web of Science ID 000209227900001
View details for PubMedID 23852182
View details for PubMedCentralID PMC3731437
Clinical correlations with lewy body pathology in LRRK2-related Parkinson's disease
WILEY-BLACKWELL. 2013: S405
View details for Web of Science ID 000320940505046
Small molecules greatly improve conversion of human-induced pluripotent stem cells to the neuronal lineage.
Stem cells international
2012; 2012: 140427-?
Efficient in vitro differentiation into specific cell types is more important than ever after the breakthrough in nuclear reprogramming of somatic cells and its potential for disease modeling and drug screening. Key success factors for neuronal differentiation are the yield of desired neuronal marker expression, reproducibility, length, and cost. Three main neuronal differentiation approaches are stromal-induced neuronal differentiation, embryoid body (EB) differentiation, and direct neuronal differentiation. Here, we describe our neurodifferentiation protocol using small molecules that very efficiently promote neural induction in a 5-stage EB protocol from six induced pluripotent stem cells (iPSC) lines from patients with Parkinson's disease and controls. This protocol generates neural precursors using Dorsomorphin and SB431542 and further maturation into dopaminergic neurons by replacing sonic hedgehog with purmorphamine or smoothened agonist. The advantage of this approach is that all patient-specific iPSC lines tested in this study were successfully and consistently coaxed into the neural lineage.
View details for DOI 10.1155/2012/140427
View details for PubMedID 22567022
View details for PubMedCentralID PMC3339118
SNCA Triplication Parkinson's Patient's iPSC-derived DA Neurons Accumulate alpha-Synuclein and Are Susceptible to Oxidative Stress
2011; 6 (11)
Parkinson's disease (PD) is an incurable age-related neurodegenerative disorder affecting both the central and peripheral nervous systems. Although common, the etiology of PD remains poorly understood. Genetic studies infer that the disease results from a complex interaction between genetics and environment and there is growing evidence that PD may represent a constellation of diseases with overlapping yet distinct underlying mechanisms. Novel clinical approaches will require a better understanding of the mechanisms at work within an individual as well as methods to identify the specific array of mechanisms that have contributed to the disease. Induced pluripotent stem cell (iPSC) strategies provide an opportunity to directly study the affected neuronal subtypes in a given patient. Here we report the generation of iPSC-derived midbrain dopaminergic neurons from a patient with a triplication in the α-synuclein gene (SNCA). We observed that the iPSCs readily differentiated into functional neurons. Importantly, the PD-affected line exhibited disease-related phenotypes in culture: accumulation of α-synuclein, inherent overexpression of markers of oxidative stress, and sensitivity to peroxide induced oxidative stress. These findings show that the dominantly-acting PD mutation is intrinsically capable of perturbing normal cell function in culture and confirm that these features reflect, at least in part, a cell autonomous disease process that is independent of exposure to the entire complexity of the diseased brain.
View details for DOI 10.1371/journal.pone.0026159
View details for Web of Science ID 000297555400007
View details for PubMedID 22110584
View details for PubMedCentralID PMC3217921
Call for participation in the neurogenetics consortium within the Human Variome Project
2011; 12 (3): 169–73
The rate of DNA variation discovery has accelerated the need to collate, store and interpret the data in a standardised coherent way and is becoming a critical step in maximising the impact of discovery on the understanding and treatment of human disease. This particularly applies to the field of neurology as neurological function is impaired in many human disorders. Furthermore, the field of neurogenetics has been proven to show remarkably complex genotype-to-phenotype relationships. To facilitate the collection of DNA sequence variation pertaining to neurogenetic disorders, we have initiated the "Neurogenetics Consortium" under the umbrella of the Human Variome Project. The Consortium's founding group consisted of basic researchers, clinicians, informaticians and database creators. This report outlines the strategic aims established at the preliminary meetings of the Neurogenetics Consortium and calls for the involvement of the wider neurogenetic community in enabling the development of this important resource.
View details for DOI 10.1007/s10048-011-0287-4
View details for Web of Science ID 000294059600001
View details for PubMedID 21630033
Phenotype in parkinsonian and nonparkinsonian LRRK2 G2019S mutation carriers
2011; 77 (4): 325-333
Using a family study design, we describe the motor and nonmotor phenotype in probands with LRRK2 G2019S mutations and family members and compare these individuals to patients with idiopathic Parkinson disease (iPD) and unrelated controls.Probands with G2019S mutations and their first-degree relatives, subjects with iPD, and unrelated control subjects were identified from 4 movement disorders centers. All underwent neurologic examinations and tests of olfaction, color vision, anxiety, and depression inventories.Tremor was more often a presenting feature among 25 individuals with LRRK2-associated PD than among 84 individuals with iPD. Subjects with LRRK2-PD had better olfactory identification compared with subjects with iPD, higher Beck Depression Inventory scores, and higher error scores on Farnsworth-Munsell 100-Hue test of color discrimination. Postural or action tremor was more common among 29 nonmanifesting mutation carriers compared with 53 noncarriers within the families. Nonparkinsonian family members had higher Unified Parkinson's Disease Rating Scale motor scores, more constipation, and worse color discrimination than controls, regardless of mutation status.Although tremor is a more common presenting feature of LRRK2-PD than iPD and some nonmotor features differed in degree, the phenotype is largely overlapping. Postural or action tremor may represent an early sign. Longitudinal evaluation of a large sample of nonmanifesting carriers will be required to describe any premotor phenotype that may allow early diagnosis.
View details for Web of Science ID 000293123300010
View details for PubMedID 21753163
View details for PubMedCentralID PMC3140802
LRRK2 Mutant iPSC-Derived DA Neurons Demonstrate Increased Susceptibility to Oxidative Stress
CELL STEM CELL
2011; 8 (3): 267-280
Studies of Parkinson's disease (PD) have been hindered by lack of access to affected human dopaminergic (DA) neurons. Here, we report generation of induced pluripotent stem cells that carry the p.G2019S mutation (G2019S-iPSCs) in the Leucine-Rich Repeat Kinase-2 (LRRK2) gene, the most common PD-related mutation, and their differentiation into DA neurons. The high penetrance of the LRRK2 mutation and its clinical resemblance to sporadic PD suggest that these cells could provide a valuable platform for disease analysis and drug development. We found that DA neurons derived from G2019S-iPSCs showed increased expression of key oxidative stress-response genes and α-synuclein protein. The mutant neurons were also more sensitive to caspase-3 activation and cell death caused by exposure to stress agents, such as hydrogen peroxide, MG-132, and 6-hydroxydopamine, than control DA neurons. This enhanced stress sensitivity is consistent with existing understanding of early PD phenotypes and represents a potential therapeutic target.
View details for DOI 10.1016/j.stem.2011.01.013
View details for Web of Science ID 000288404400009
View details for PubMedCentralID PMC3578553
Mitochondrial Dysfunction in Skin Fibroblasts from a Parkinson's Disease Patient with an alpha-Synuclein Triplication
JOURNAL OF PARKINSONS DISEASE
2011; 1 (2): 175–83
Mitochondrial dysfunction has been frequently implicated in the neurodegenerative process that underlies Parkinson's disease (PD), but the basis for this impairment is not fully understood. The goal of this study was to investigate the effects of α-synuclein (α-syn) gene multiplication on mitochondrial function in human tissue. To investigate this question, human fibroblasts were taken from a patient with parkinsonism carrying a triplication in the α-syn gene. Unexpectedly, the cells showed a significant decrease in cell growth compared to matched healthy controls. With regard to mitochondrial function, α-syn triplication fibroblasts exhibited a 39% decrease in ATP production, a 40% reduction in mitochondrial membrane potential, and a 49% reduction in complex I activity. Furthermore, they proved to be more sensitive to the effects of the nigrostrial toxicant paraquat compared to controls. Finally, siRNA knockdown of α-syn resulted in a partial rescue of mitochondrial impairment and reduction of paraquat-induced cell toxicity, suggesting that α-syn plays a causative role for mitochondrial dysfunction in these patient-derived peripheral skin fibroblasts.
View details for DOI 10.3233/JPD-2011-11025
View details for Web of Science ID 000308482400005
View details for PubMedID 23934919
Alpha-synuclein-glucocerebrosidase interactions in pharmacological Gaucher models: A biological link between Gaucher disease and parkinsonism
2009; 30 (6): 1127–32
A growing body of experimental and clinical literature indicates an association between Gaucher disease and parkinsonism, raising the possibility that convergent mechanisms may contribute to neurodegeneration in these disorders. The aim of this study was to determine whether there is a relationship between alpha-synuclein (alpha-syn), a key protein in Parkinson's disease pathogenesis, and abnormalities in glucocerebroside (GC) catabolism that lead to the development of Gaucher disease. We inhibited glucocerebrosidase (GCase) with conduritol B epoxide (CBE) in neuroblastoma cells and mice to test whether a biological link exists between GCase activity and alpha-syn. After CBE exposure, enhanced alpha-syn protein was detected in differentiated cells challenged with CBE as compared to vehicle, with no change in alpha-syn mRNA. In the mouse model, after one injection of CBE, elevated nigral alpha-syn levels were also detected. Analyses by Western blot and confocal microscopy revealed that normal alpha-syn distribution was perturbed after CBE exposure with its accumulation apparent within nigral cell bodies as well as astroglia. These findings raise the possibility that alpha-syn may contribute to the cascade of events that promote neuronal dysfunction in Gaucher disease and are the first to implicate this protein as a plausible biological intersection between Gaucher disease and parkinsonism using a pharmacological model.
View details for DOI 10.1016/j.neuro.2009.06.009
View details for Web of Science ID 000272873700033
View details for PubMedID 19576930
Can cellular models revolutionize drug discovery in Parkinson's disease?
BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR BASIS OF DISEASE
2009; 1792 (11): 1043–51
The study of mechanisms that underlie Parkinson's disease (PD), as well as translational drug development, has been hindered by the lack of appropriate models. Both cell culture systems and animal models have limitations, and to date none faithfully recapitulate all of the clinical and pathological phenotypes of the disease. In this review we examine the various cell culture model systems of PD, with a focus on different stem cell models that can be used for investigating disease mechanisms as well as drug discovery for PD. We conclude with a discussion of recent discoveries in the field of stem cell biology that have led to the ability to reprogram somatic cells to a pluripotent state via the use of a combination of genetic factors; these reprogrammed cells are termed "induced pluripotent stem cells" (iPSCs). This groundbreaking technique allows for the derivation of patient-specific cell lines from individuals with sporadic forms of PD and also those with known disease-causing mutations. Such cell lines have the potential to serve as a human cellular model of neurodegeneration and PD when differentiated into dopaminergic neurons. The hope is that these iPSC-derived dopaminergic neurons can be used to replicate the key molecular aspects of neural degeneration associated with PD. If so, this approach could lead to transformative new tools for the study of disease mechanisms. In addition, such cell lines can be potentially used for high-throughput drug screening. While not the focus of this review, ultimately it is envisioned that techniques for reprogramming of somatic cells may be optimized to a point sufficient to provide potential new avenues for stem cell-based restorative therapies.
View details for DOI 10.1016/j.bbadis.2009.08.014
View details for Web of Science ID 000271807300001
View details for PubMedID 19733239
Severe congenital encephalopathy caused by MECP2 null mutations in males: central hypoxia and reduced neuronal dendritic structure
2008; 74 (2): 116-126
Non-mosaic males with a 46,XY karyotype and a MECP2 null mutation display a phenotype of severe neonatal-onset encephalopathy that is distinctly different from Rett syndrome (RTT). To increase awareness of this rare disorder, we are reporting novel findings in a sporadic case, compare them to 16 previously reported cases and establish salient criteria for clinical diagnosis. The proband suffered from general hypotonia and hypoxia caused by hypoventilation and irregular breathing. He developed abnormal movements, seizures and electroencephalogram abnormalities. He failed to thrive and to reach any motor milestones and died at 15 months from central respiratory failure without a diagnosis. In a muscle biopsy, type II fibers were reduced in diameter, indicating central hypoxia. At autopsy, the brain was small with disproportionate reduction of the frontal and temporal lobes. Synaptophysin staining of synaptic vesicles was greatly reduced in cerebellar and spinal cord sections. Analysis of Golgi-stained pyramidal neurons from cortical layers III and V of the frontal and temporal lobes revealed drastically diminished dendritic trees. Post-mortem MECP2 mutation analysis on DNA and RNA from fibroblasts revealed a novel de novo 9-nucleotide deletion including the intron 3/exon 4 splice junction. The two nucleotides flanking the deletion form a new splice site, and the aberrantly spliced transcript lacks seven nucleotides (r.378_384delTCCCCAG), causing a frameshift and premature termination codon (p.I126fsX11). Males with congenital encephalopathy, not females with RTT, represent the true human counterpart for the commonly studied Mecp2-/y mouse model and provide unique insight into the mechanisms of MeCP2 deficiency.
View details for DOI 10.1111/j.1399-0004.2008.01005.x
View details for Web of Science ID 000257476200003
View details for PubMedID 18477000
Pure akinesia as initial presentation of PSP: A clinicopathological study
PARKINSONISM & RELATED DISORDERS
2008; 14 (6): 517–19
Pure akinesia (PA) is a rare neurodegenerative condition that may represent a limited expression of progressive supranuclear palsy (PSP). Only a few pathological studies have been reported and its classification remains unclear. We report the case of a 57-year-old Caucasian man who was initially clinically diagnosed with classical PA. After four years the patient developed additional symptoms and signs compatible with the diagnosis of clinically probable PSP. The diagnosis of PSP was confirmed by post-mortem examination. Genetic analysis of the MAPT gene revealed an A0/A0 genotype, which has been repeatedly associated with the PSP phenotype, and might discriminate between PA and other gait disorders. Our case strengthens the hypothesis that PA should be considered as initial manifestation of PSP.
View details for DOI 10.1016/j.parkreldis.2007.11.004
View details for Web of Science ID 000259653100012
View details for PubMedID 18325816
Phenotypic spectrum and sex effects in eleven myoclonus-dystonia families with epsilon-sarcoglycan mutations
2008; 23 (4): 588–92
Myoclonus-dystonia (M-D) due to SGCE mutations is characterized by early onset myoclonic jerks, often associated with dystonia. Penetrance is influenced by parental sex, but other sex effects have not been established. In 42 affected individuals from 11 families with identified mutations, we found that sex was highly associated with age at onset regardless of mutation type; the median age onset for girls was 5 years versus 8 years for boys (P < 0.0097). We found no association between mutation type and phenotype.
View details for DOI 10.1002/mds.21785
View details for Web of Science ID 000254808000017
View details for PubMedID 18175340
DLX5 and DLX6 expression is biallelic and not modulated by MeCP2 deficiency
AMERICAN JOURNAL OF HUMAN GENETICS
2007; 81 (3): 492-506
Mutations in MECP2 and Mecp2 (encoding methyl-CpG binding protein 2 [MeCP2]) cause distinct neurological phenotypes in humans and mice, respectively, but the molecular pathology is unclear. Recent literature claimed that the developmental homeobox gene DLX5 is imprinted and that its imprinting status is modulated by MeCP2, leading to biallelic expression in Rett syndrome and twofold overexpression of Dlx5 and Dlx6 in Mecp2-null mice. The conclusion that DLX5 is a direct target of MeCP2 has implications for research on the molecular bases of Rett syndrome, autism, and genomic imprinting. Attempting to replicate the reported data, we evaluated allele-specific expression of DLX5 and DLX6 in mouse x human somatic cell hybrids, lymphoblastoid cell lines, and frontal cortex from controls and individuals with MECP2 mutations. We identified novel single-nucleotide polymorphisms in DLX5 and DLX6, enabling the first imprinting studies of DLX6. We found that DLX5 and DLX6 are biallelically expressed in somatic cell hybrids and in human cell lines and brain, with no differences between affected and control samples. We also determined expression levels of Dlx5 and Dlx6 in forebrain from seven male Mecp2-mutant mice and eight wild-type littermates by real-time quantitative reverse-transcriptase polymerase chain reaction assays. Expression of Dlx5 and Dlx6, as well as of the imprinted gene Peg3, in mouse forebrain was highly variable, with no consistent differences between Mecp2-null mutants and controls. We conclude that DLX5 and DLX6 are not imprinted in humans and are not likely to be direct targets of MeCP2 modulation. In contrast, the imprinting status of PEG3 and PEG10 is maintained in MeCP2-deficient tissues. Our results confirm that MeCP2 plays no role in the maintenance of genomic imprinting and add PEG3 and PEG10 to the list of studied imprinted genes.
View details for DOI 10.1086/520063
View details for Web of Science ID 000249128200006
View details for PubMedID 17701895
View details for PubMedCentralID PMC1950824
Phenotypic variation in a large Swedish pedigree due to SNCA duplication and triplication
2007; 68 (12): 916–22
The "Lister family complex," an extensive Swedish family with autosomal dominant Parkinson disease, was first described by Henry Mjönes in 1949. On the basis of clinical, molecular, and genealogic findings on a Swedish and an American family branch, we provide genetic evidence that explains the parkinsonism in this extended pedigree.Clinical methods included a detailed neurologic exam of the proband of the Swedish family branch, MRI, and (I)-beta-CIT SPECT imaging. Genomic analysis included alpha-synuclein sequencing, SNCA real-time PCR dosage, chromosome 4q21 microsatellite analysis, and high-resolution microarray genotyping. The geographic origin and ancestral genealogy of each pedigree were researched in the medical literature and Swedish Parish records.The proband of the Swedish family branch presented with early dysautonomia followed by progressive parkinsonism suggestive of multiple system atrophy. Molecular analysis identified a genomic duplication of <0.9 Mb encompassing alpha-synuclein and multimerin 1 (SNCA-MMRN1), flanked by long interspersed repeat sequences (LINE L1). Microsatellite variability within the genomic interval was identical to that previously described for a Swedish American family with an alpha-synuclein triplication. Subsequent genealogic investigation suggested that both kindreds are ancestrally related to the Lister family complex.Our findings extend clinical, genetic, and genealogical research on the Lister family complex. The genetic basis for familial parkinsonism is an SNCA-MMRN11 multiplication, but whereas SNCA-MMRN1 duplication in the Swedish proband (Branch J) leads to late-onset autonomic dysfunction and parkinsonism, SNCA-MMRN1 triplication in the Swedish American family (Branch I) leads to early-onset Parkinson disease and dementia.
View details for DOI 10.1212/01.wnl.0000254458.17630.c5
View details for Web of Science ID 000245034400009
View details for PubMedID 17251522
Parkin gene variations and parkinsonism: Association does not imply causation
ANNALS OF NEUROLOGY
2007; 61 (1): 4–6
View details for DOI 10.1002/ana.21069
View details for Web of Science ID 000244028000004
View details for PubMedID 17262853
Novel features in a patient homozygous for the L347P mutation in the PINK1 gene
PARKINSONISM & RELATED DISORDERS
2007; 13 (6): 359–61
The purpose of this study was to assess the genotype-phenotype of PINK1 mutations. We genotyped eight known mutations in three clinic-based cohorts with Parkinsonism and found one homozygous p.L347P mutation in PINK1. Clinically, hypo-osmia and profound diurnal variation of symptoms were identified as novel features; fluorodopa positron emission tomography revealed striking decline in striatal fluorodopa uptake. We suggest that it may be possible to clinically separate this form of Parkinsonism from dopa-responsive dystonia and Parkin-related Parkinsonism. Furthermore, as this mutation has only been reported in Filipinos (two originated from Luzon island), our results support the hypothesis of a common founder.
View details for DOI 10.1016/j.parkreldis.2006.08.009
View details for Web of Science ID 000249633400007
View details for PubMedID 17055324
Intrafamilial phenotypic and genetic heterogeneity of dystonia
JOURNAL OF THE NEUROLOGICAL SCIENCES
2006; 250 (1-2): 92–96
Most cases of early-onset primary torsion dystonia are caused by the same 3-bp (GAG) deletion in the DYT1 gene. We describe a large Serbian family with significant intrafamilial variability of the DYT1 phenotype, from asymptomatic carrier status to late-onset focal, and generalized jerky dystonia. Seven mutation carriers (six proven by direct analysis and one by inferred haplotype) were identified, but only two of them were affected by dystonia (penetrance reduced to 29%). In addition, three GAG-deletion-negative family members also developed dystonia (two multifocal dystonia and one torticollis), suggesting that their involuntary movements are due to some other etiological factor(s) (i.e., another dystonia gene), or may be psychogenic.
View details for DOI 10.1016/j.jns.2006.07.010
View details for Web of Science ID 000242471500015
View details for PubMedID 17027035
Inactivating mutations in ESCO2 cause SC phocomelia and Roberts syndrome: No phenotype-genotype correlation
AMERICAN JOURNAL OF HUMAN GENETICS
2005; 77 (6): 1117-1128
The rare, autosomal recessive Roberts syndrome (RBS) is characterized by tetraphocomelia, profound growth deficiency of prenatal onset, craniofacial anomalies, microcephaly, and mental deficiency. SC phocomelia (SC) has a milder phenotype, with a lesser degree of limb reduction and with survival to adulthood. Since heterochromatin repulsion (HR) is characteristic for both disorders and is not complemented in somatic-cell hybrids, it has been hypothesized that the disorders are allelic. Recently, mutations in ESCO2 (establishment of cohesion 1 homolog 2) on 8p21.1 have been reported in RBS. To determine whether ESCO2 mutations are also responsible for SC, we studied three families with SC and two families in which variable degrees of limb and craniofacial abnormalities, detected by fetal ultrasound, led to pregnancy terminations. All cases were positive for HR. We identified seven novel mutations in exons 3-8 of ESCO2. In two families, affected individuals were homozygous--for a 5-nucleotide deletion in one family and a splice-site mutation in the other. In three nonconsanguineous families, probands were compound heterozygous for a single-nucleotide insertion or deletion, a nonsense mutation, or a splice-site mutation. Abnormal splice products were characterized at the RNA level. Since only protein-truncating mutations were identified, regardless of clinical severity, we conclude that genotype does not predict phenotype. Having established that RBS and SC are caused by mutations in the same gene, we delineated the clinical phenotype of the tetraphocomelia spectrum that is associated with HR and ESCO2 mutations and differentiated it from other types of phocomelia that are negative for HR.
View details for Web of Science ID 000233241200020
View details for PubMedID 16380922
PINK1, Parkin, and DJ-1 mutations in Italian patients with early-onset parkinsonism
EUROPEAN JOURNAL OF HUMAN GENETICS
2005; 13 (9): 1086–93
Recessively inherited early-onset parkinsonism (EOP) has been associated with mutations in the Parkin, DJ-1, and PINK1 genes. We studied the prevalence of mutations in all three genes in 65 Italian patients (mean age of onset: 43.2+/-5.4 years, 62 sporadic, three familial), selected by age at onset equal or younger than 51 years. Clinical features were compatible with idiopathic Parkinson's disease in all cases. To detect small sequence alterations in Parkin, DJ-1, and PINK1, we performed a conventional mutational analysis (SSCP/dHPLC/sequencing) of all coding exons of these genes. To test for the presence of exon rearrangements in PINK1, we established a new quantitative duplex PCR assay. Gene dosage alterations in Parkin and DJ-1 were excluded using previously reported protocols. Five patients (8%; one woman/four men; mean age at onset: 38.2+/-9.7 (range 25-49) years) carried mutations in one of the genes studied: three cases had novel PINK1 mutations, one of which occurred twice (homozygous c.1602_1603insCAA; heterozygous c.1602_1603insCAA; heterozygous c.836G>A), and two patients had known Parkin mutations (heterozygous c.734A>T and c.924C>T; heterozygous c.924C>T). Family history was negative for all mutation carriers, but one with a history of tremor. Additionally, we detected one novel polymorphism (c.344A>T) and four novel PINK1 changes of unknown pathogenic significance (-21G/A; IVS1+97A/G; IVS3+38_40delTTT; c.852C>T), but no exon rearrangements. No mutations were found in the DJ-1 gene. The number of mutation carriers in both the Parkin and the PINK1 gene in our cohort is low but comparable, suggesting that PINK1 has to be considered in EOP.
View details for DOI 10.1038/sj.ejhg.5201455
View details for Web of Science ID 000231395500016
View details for PubMedID 15970950
Molecular breakpoint cloning and gene expression studies of a novel translocation t(4;15)(q27;q11.2) associated with Prader-Willi syndrome
BMC MEDICAL GENETICS
Prader-Willi syndrome (MIM #176270; PWS) is caused by lack of the paternally-derived copies, or their expression, of multiple genes in a 4 Mb region on chromosome 15q11.2. Known mechanisms include large deletions, maternal uniparental disomy or mutations involving the imprinting center. De novo balanced reciprocal translocations in 5 reported individuals had breakpoints clustering in SNRPN intron 2 or exon 20/intron 20. To further dissect the PWS phenotype and define the minimal critical region for PWS features, we have studied a 22 year old male with a milder PWS phenotype and a de novo translocation t(4;15)(q27;q11.2).We used metaphase FISH to narrow the breakpoint region and molecular analyses to map the breakpoints on both chromosomes at the nucleotide level. The expression of genes on chromosome 15 on both sides of the breakpoint was determined by RT-PCR analyses.Pertinent clinical features include neonatal hypotonia with feeding difficulties, hypogonadism, short stature, late-onset obesity, learning difficulties, abnormal social behavior and marked tolerance to pain, as well as sticky saliva and narcolepsy. Relative macrocephaly and facial features are not typical for PWS. The translocation breakpoints were identified within SNRPN intron 17 and intron 10 of a spliced non-coding transcript in band 4q27. LINE and SINE sequences at the exchange points may have contributed to the translocation event. By RT-PCR of lymphoblasts and fibroblasts, we find that upstream SNURF/SNRPN exons and snoRNAs HBII-437 and HBII-13 are expressed, but the downstream snoRNAs PWCR1/HBII-85 and HBII-438A/B snoRNAs are not.As part of the PWCR1/HBII-85 snoRNA cluster is highly conserved between human and mice, while no copy of HBII-438 has been found in mouse, we conclude that PWCR1/HBII-85 snoRNAs is likely to play a major role in the PWS- phenotype.
View details for DOI 10.1186/1471-2350-6-18
View details for Web of Science ID 000230403600001
View details for PubMedID 15877813
View details for PubMedCentralID PMC1142316
Genetic heterogeneity in ten families with myoclonus-dystonia
JOURNAL OF NEUROLOGY NEUROSURGERY AND PSYCHIATRY
2004; 75 (8): 1181-1185
Myoclonus-dystonia (M-D) is a movement disorder with autosomal dominant inheritance and reduced penetrance but may also occur sporadically. Recently, mutations in the epsilon-sarcoglycan gene (SGCE) were shown to cause M-D. Furthermore, single variants in the dopamine D2 receptor (DRD2) and DYT1 genes were found in combination with SGCE mutations in two M-D families, and another M-D locus was recently mapped to chromosome 18p11 in one family.The authors clinically and genetically characterised ten consecutive cases with myoclonus-dystonia; seven familial and three sporadic. Twenty nine M-D patients and 40 unaffected family members underwent a standardised clinical examination by a movement disorder specialist. Index cases were screened for mutations in the SGCE, DYT1, and DRD2 genes and for deletions of the SGCE gene. Suitable mutation negative families were tested for linkage to the SGCE region and to chromosome 18p11.Two SGCE mutations were detected among the seven familial but no mutation in the sporadic cases. Haplotype analysis at the new M-D locus was compatible with linkage in two families and excluded in another family, suggesting at least one additional M-D gene. There were no obvious clinical differences between M-D families with and without detected mutations.M-D is genetically heterogeneous with SGCE mutations accounting for the disease in only part of the clinically typical cases.
View details for DOI 10.1136/jnnp.2003.027177
View details for Web of Science ID 000222727800024
View details for PubMedID 15258227
View details for PubMedCentralID PMC1739169
Parkin gene alterations in hepatocellular carcinoma
GENES CHROMOSOMES & CANCER
2004; 40 (2): 85–96
The Parkin gene is an extremely large gene (1.5 Mb) within the highly unstable FRA6E common fragile site (CFS) region, which is frequently altered in ovarian, breast, and hepatocellular carcinomas. Because Parkin/FRA6E has genomic similarities to FHIT/FRA3B and WWOX/FRA16D, two other large tumor-suppressor genes that are within CFS regions, we were interested in characterizing Parkin gene alterations and their possible association with cancer. After analyzing 50 cancer-derived cell lines including 11 hepatocellular carcinoma (HCC) cell lines, we found that one HCC cell line, PLC/PRF/5, had a detectable homozygous deletion encompassing exon 3. Using quantitative duplex PCR and fluorescence in situ hybridization analysis to characterize the copy number changes of Parkin exons in HCC cell lines, we found that 4 of 11 HCC cell lines had heterozygous deletions of Parkin exons and one, Hep3B, had an exon duplication. Parkin protein expression was significantly decreased or absent in all 11 HCC cell lines. Furthermore, more than 50% of HCC primary tumors had decreased Parkin expression compared to that in normal liver tissue. Parkin gene-transfected PLC5 and Hep3B cells grew more slowly than vector-only transfectants and also showed increased sensitivity to apoptosis induced by cell-cycle inhibitors. Therefore, we suggest that Parkin may be involved in tumor suppression and that the loss of Parkin contributes to the development of hepatocarcinoma.
View details for DOI 10.1002/gcc.20020
View details for Web of Science ID 000221154800001
View details for PubMedID 15101042
Myoclonus-dystonia: Detection of novel, recurrent, and de novo SGCE mutations
2004; 62 (7): 1229–31
View details for DOI 10.1212/01.WNL.0000118286.75059.35
View details for Web of Science ID 000220769300042
View details for PubMedID 15079037
Clinical and genetic features of myoclonus-dystonia in 3 cases: A video presentation
2004; 19 (2): 231–34
Many cases of myoclonus-dystonia (M-D) are caused by mutations in the epsilon-sarcoglycan (SGCE) gene. We describe 3 children with a similar clinical picture of autosomal dominant M-D and an SGCE mutation in only one of them, suggesting that M-D is genetically heterogeneous.
View details for DOI 10.1002/mds.10635
View details for Web of Science ID 000189160700019
View details for PubMedID 14978685
Alterations in the common fragile site gene Parkin in ovarian and other cancers
2003; 22 (51): 8370–78
The cloning and characterization of the common fragile site (CFS) FRA6E (6q26) identified Parkin, the gene involved in the pathogenesis of many cases of juvenile, early-onset and, rarely, late-onset Parkinson's disease, as the third large gene to be localized within a large CFS. Initial analyses of Parkin indicated that in addition to playing a role in Parkinson's disease, it might also be involved in the development and/or progression of ovarian cancer. These analyses also indicated striking similarities among the large CFS-locus genes: fragile histidine triad gene (FHIT; 3p14.2), WW domain-containing oxidoreductase gene (WWOX; 16q23), and Parkin (6q26). Analyses of FHIT and WWOX in a variety of different cancer types have identified the presence of alternative transcripts with whole exon deletions. Interestingly, various whole exon duplications and deletions have been identified for Parkin in juvenile and early-onset Parkinson's patients. Therefore, we performed mutational/exon rearrangement analysis of Parkin in ovarian cancer cell lines and primary tumors. Four (66.7%) cell lines and four (18.2%) primary tumors were identified as being heterozygous for the duplication or deletion of a Parkin exon. Additionally, three of 23 (13.0%) nonovarian tumor-derived cell lines were also identified as having a duplication or deletion of one or more Parkin exons. Analysis of Parkin protein expression with antibodies revealed that most of the ovarian cancer cell lines and primary tumors had diminished or absent Parkin expression. While functional analyses have not yet been performed for Parkin, these data suggest that like FHIT and WWOX, Parkin may represent a tumor suppressor gene.
View details for DOI 10.1038/sj.onc.1207072
View details for Web of Science ID 000186541500015
View details for PubMedID 14614460
Hereditary myoclonus-dystonia associated with epilepsy
2003; 60 (12): 1988–90
A five-generation Dutch family with inherited myoclonus-dystonia (M-D) is described. Genetic analysis revealed a novel truncating mutation within the epsilon-sarcoglycan gene (SGCE). In three of five gene carriers, epilepsy and/or EEG abnormalities were associated with the symptoms of myoclonus and dystonia. The genetic and clinical heterogeneity of M-D is extended. EEG changes and epilepsy should not be considered exclusion criteria for the clinical diagnosis of M-D.
View details for DOI 10.1212/01.WNL.0000066020.99191.76
View details for Web of Science ID 000183696500023
View details for PubMedID 12821748
Evidence that paternal expression of the epsilon-Sarcoglycan gene accounts for reduced penetrance in myoclonus-dystonia
AMERICAN JOURNAL OF HUMAN GENETICS
2002; 71 (6): 1303–11
Myoclonus-dystonia (M-D) is a movement disorder characterized by rapid muscle contractions and sustained twisting and repetitive movements and has recently been associated with mutations in the epsilon-sarcoglycan gene (SGCE). The mode of inheritance is autosomal dominant with reduced penetrance upon maternal transmission, suggesting a putative maternal imprinting mechanism. We present an apparently sporadic M-D case and two patients from an M-D family with seemingly autosomal recessive inheritance. In both families, we detected an SGCE mutation that was inherited from the patients' clinically unaffected fathers in an autosomal dominant fashion. Whereas, in the first family, RNA expression studies revealed expression of only the mutated allele in affected individuals and expression of the normal allele exclusively in unaffected mutation carriers, the affected individual of the second family expressed both alleles. In addition, we identified differentially methylated regions in the promoter region of the SGCE gene as a characteristic feature of imprinted genes. Using a rare polymorphism in the promoter region in a family unaffected with M-D as a marker, we demonstrated methylation of the maternal allele, in keeping with maternal imprinting of the SGCE gene. Loss of imprinting in the patient with M-D who had biallelic expression of the SGCE gene was associated with partial loss of methylation at several CpG dinucleotides.
View details for DOI 10.1086/344531
View details for Web of Science ID 000179586800005
View details for PubMedID 12444570
View details for PubMedCentralID PMC378568
epsilon-sarcoglycan mutations found in combination with other dystonia gene mutations
ANNALS OF NEUROLOGY
2002; 52 (5): 675–79
Myoclonus-dystonia is a movement disorder associated with mutations in the epsilon-sarcoglycan gene (SGCE) in most families and in the DRD2 and DYT1 genes in two single families. In both of the latter families, we also found a mutation of SGCE. The molecular mechanisms through which the detected mutations may contribute to myoclonus-dystonia remain to be determined.
View details for DOI 10.1002/ana.10358
View details for Web of Science ID 000178914000026
View details for PubMedID 12402271
Phenotypic features of myoclonus-dystonia in three kindreds
2002; 59 (8): 1187–96
Myoclonus-dystonia (M-D) is a movement disorder with involuntary jerks and dystonic contractions. Autosomal dominant alcohol-responsive M-D is associated with mutations in the epsilon-sarcoglycan gene (SGCE) (six families) and with a missense change in the D2 dopamine receptor (DRD2)gene (one family).To investigate the clinical phenotype associated with M-D including motor symptoms, psychiatric disorders, and neuropsychological deficits.Fifty individuals in three M-D families were evaluated and a standardized neurologic examination and DNA analysis were performed. Psychiatric profiles were established with the Diagnostic Interviews for Genetic Studies (DIGS) and the Yale-Brown Obsessive-Compulsive Scale (YBOCS). Cognition was evaluated with standardized neuropsychological tests.Distinct truncating mutations in the SGCE gene were identified in each family. Additionally, a missense alteration in the DRD2 gene was previously found in one family. Motor expression was variable, with onset of myoclonus or dystonia or both affecting the upper body and progression to myoclonus and dystonia in most cases. Psychiatric profiles revealed depression, obsessive-compulsive disorder, substance abuse, anxiety/panic/phobic disorders, and psychosis in two families, and depression only in the third family. Averaged scores from cognitive testing showed impaired verbal learning and memory in one family, impaired memory in the second family, and no cognitive deficits in the third family.Cognitive deficits may be associated with M-D. Psychiatric abnormalities correlate with the motor symptoms in affected individuals. Assessment of additional M-D families with known mutations is needed to determine whether these are characteristic phenotypic manifestations of M-D.
View details for DOI 10.1212/WNL.59.8.1187
View details for Web of Science ID 000178726700013
View details for PubMedID 12391346
Dopamine transmission in DYT1 dystonia: A biochemical and autoradiographical study
2002; 59 (3): 445-448
Indices of dopamine transmission were measured in the postmortem striatum of DYT1 dystonia brains. A significant increase in the striatal 3,4-dihydroxyphenylacetic acid/dopamine ratio was found. Quantitative autoradiography revealed no differences in the density of dopamine transporter or vesicular monoamine transporter-2 binding; however, there was a trend toward a reduction in D(1) receptor and D(2) receptor binding. One brain with DYT1 parkinsonism was similarly evaluated and marked reductions in striatal dopamine, 3,4-dihydroxyphenylacetic acid, and homovanillic acid content as well as the density of binding of all four dopaminergic ligands were measured.
View details for Web of Science ID 000177335800027
View details for PubMedID 12177384
Role of SC42 mutations in early- and late-onset dopa-responsive parkinsonism
ANNALS OF NEUROLOGY
2002; 52 (2): 257–58
View details for DOI 10.1002/ana.10270
View details for Web of Science ID 000177140000024
View details for PubMedID 12210804
The parkin gene is not involved in late-onset Parkinson's disease
2002; 58 (5): 835
View details for DOI 10.1212/WNL.58.5.835
View details for Web of Science ID 000174301300033
View details for PubMedID 11889262
Slice culture of the olfactory bulb of Xenopus laevis tadpoles
2001; 26 (4): 399–407
We report on the development of a slice culture of amphibian brain tissue. In particular, we cultured slices from Xenopus laevis tadpoles that contain the olfactory mucosae, the olfactory nerves, the olfactory bulb and the telencephalon. During 6 days in roller tubes the slices flattened, starting from 250 microm and decreasing to approximately 40 microm, corresponding to about three cell layers. Dendritic processes could be followed over distances as long as 200 microm. Neurons in the cultured slice could be recorded using the patch clamp technique and simultaneously imaged using an inverted laser scanning microscope. We characterized the main neuron types of the olfactory bulb, i.e. mitral cells and granule cells, by correlating their typical morphological features in the acute slice with the electrophysiological properties in both the acute slice and slice culture. This correlation allowed unambiguous identification of mitral cells and granule cells in the slice culture.
View details for DOI 10.1093/chemse/26.4.399
View details for Web of Science ID 000168752800005
View details for PubMedID 11369674
Noradrenergic modulation of calcium currents and synaptic transmission in the olfactory bulb of Xenopus laevis tadpoles
EUROPEAN JOURNAL OF NEUROSCIENCE
2001; 13 (6): 1093–1100
Norepinephrine (NE) has various modulatory roles in both the peripheral and the central nervous systems. Here we investigate the function of the locus coeruleus efferent fibres in the olfactory bulb of Xenopus laevis tadpoles. In order to distinguish unambiguously between mitral cells and granule cells of the main olfactory bulb and the accessory olfactory bulb, we used a slice preparation. The two neuron types were distinguished on the basis of their location in the slice, their typical branching pattern and by electrophysiological criteria. At NE concentrations lower than 5 microM there was only one effect of NE upon voltage-gated conductances; NE blocked a high-voltage-activated Ca(2+)-current in mitral cells of both the main and the accessory olfactory bulbs. No such effect was observed in granule cells. The effect of NE upon mitral cell Ca(2+)-currents was mimicked by the alpha(2)-receptor agonists clonidine and alpha-methyl-NE. As a second effect, NE or clonidine blocked spontaneous synaptic activity in granule cells of both the main and the accessory olfactory bulbs. NE or clonidine also blocked the spontaneous synaptic activity in mitral cells of either olfactory bulb. The amplitude of glutamate-induced currents in granule cells was modulated neither by clonidine nor by alpha-methyl-NE. Taken together, the main effect of the noradrenergic, presynaptic, alpha(2)-receptor-mediated block of Ca(2)+-currents in mitral cells appeared to be a wide-spread disinhibition of mitral cells in the accessory olfactory bulb as well as in the main olfactory bulb.
View details for DOI 10.1046/j.0953-816x.2001.01479.x
View details for Web of Science ID 000167615400004
View details for PubMedID 11285006