Project description:The pathophysiology of neurodegenerative diseases is poorly understood, and therapeutic options are few. Neurodegenerative diseases are hallmarked by progressive neuronal dysfunction and loss, associated with chronic glial and immune activation1. Microglia are the resident macrophages of the nervous system thought to be important for the clearance of debris after neuronal damage and aid to repair and regeneration. Their activation is viewed in general as a reactive process in neurodegeneration1-3. Whether microglia itself may be causative of neurodegenerative diseases is unclear. Late-onset neurodegenerative disease is frequently observed in patients with histiocytoses4-10, which are clonal myeloid diseases associated with somatic mutations in the RAS/MEK/ERK pathway such as BRAFV600E 5,11,12, suggesting a possible role of somatic mutations in neurodegeneration. Yet, expression of BRAFV600E in the hematopoietic stem cell (HSC) lineage does not cause neurodegeneration13,14. However, recent works from our laboratories and other have revealed that microglia belong to a lineage of adult tissue-resident myeloid cells that develop during organogenesis from yolk sac erythro-myeloid progenitors (EMP) distinct from HSC15-18. We thus hypothesized that somatic BRAFV600E mutations in the resident macrophage lineage may cause neurodegeneration. We found that while BRAFV600E expression in the HSC lineage causes leukemic and tumoral diseases13,14, its expression in the EMP lineage instead results in a severe late-onset neurodegenerative disorder. Neurological symptoms, neuronal death, astrogliosis, immune activation, and amyloid precursor protein deposition were driven by ERK-activated microglia clones and preventable by RAF inhibition. These results identify the fetal precursors of tissue-resident macrophages as a potential cell-of-origin for histiocytoses, and demonstrate that a somatic mutation in the EMP lineage can drive late-onset neurodegeneration. Moreover, these data identify activation of the MAP kinase pathway in microglia as a cause of neurodegenerative disease, and provide opportunities for therapeutic intervention aimed at preventing neuronal death in neurodegenerative diseases

Project description:Neurodegenerative diseases of the central nervous system are characterised by pathogenetic cellular and molecular changes in specific areas of the brain that lead to the dysfunction and/or loss of explicit neuronal populations. Despite exhibiting different clinical profiles and selective neuronal loss, common features such as abnormal protein deposition, dysfunctional cellular transport, mitochondrial deficits, glutamate excitotoxicity and inflammation are observed in most, if not all, neurodegenerative disorders, suggesting converging pathways of neurodegeneration. We have generated comparative genome-wide gene expression data for Alzheimer’s disease, amyotrophic lateral sclerosis, Huntington’s disease, multiple sclerosis, Parkinson’s disease and schizophrenia using an extensive cohort of well characterised post-mortem CNS tissues. The analysis of whole genome expression patterns across these major disorders offers an outstanding opportunity not only to look into exclusive disease specific changes, but more importantly to uncover potential common molecular pathogenic mechanisms that could be targeted for therapeutic gain. Surprisingly, no dysregulated gene that passed our selection criteria was found in common across all 6 diseases using our primary method of analysis. However, 61 dysregulated genes were shared when comparing five and four diseases. Our analysis indicates firstly the involvement of common neuronal homeostatic, survival and synaptic plasticity pathways. Secondly, we report changes to immunoregulatory and immunomodulatory pathways in all diseases. Our secondary method of analysis confirmed significant up-regulation of a number of genes in diseases presenting degeneration and showed that somatostatin was downregulated in all 6 diseases. The latter is supportive of a general role for neuroinflammation in the pathogenesis and/or response to neurodegeneration. Unravelling the detailed nature of the molecular changes regulating inflammation in the CNS is key to the development of novel therapeutic approaches for these chronic conditions. A total of 113 cases were selected retrospectively on the basis of a confirmed clinical and neuropathological diagnosis and snap-frozen brain blocks were provided by various tissue banks within the BrainNet Europe network. Total RNA was extracted from dissected snap-frozen tissue (< 100 mg) by the individual laboratories according to a BNE optimised common protocol using the RNeasy(r) tissue lipid mini kit (Qiagen Ltd, Crawley, UK) according to the manufacturer's instructions, and was stored at -80C until further use. Gene expression analysis was performed on the RNA samples using the Illumina whole genome HumanRef8 v2 BeadChip (Illumina, London, UK). All the labelling and hybridisation of the samples was carried out in a single experiment by the Imperial College group to reduce the technical variability. RNA samples were prepared for array analysis using the Illumina TotalPrep(tm)-96 RNA Amplification Kit (Ambion/Applied Biosystems, Warrington, UK). Finally, the BeadChips we re scanned using the Illumina BeadArray Reader. The data was extracted using BeadStudio 3.2 (Illumina). Data normalisation and gene differential expression analyses were conducted using the Rosetta error models available in the Rosetta Resolver(r) system (Rosetta Biosoftware, Seattle, Wa, USA). Two samples presented very low signal expression most likely due to hybridization problems and did not pass the quality control test. They are not represented here. One of the 2 samples was a replicate, therefore there was loss of only 1 case bringing the grand total of cases used to 112 (total of samples of 118 including 6 replicates).

Project description:Neurodegenerative diseases of the central nervous system are characterised by pathogenetic cellular and molecular changes in specific areas of the brain that lead to the dysfunction and/or loss of explicit neuronal populations. Despite exhibiting different clinical profiles and selective neuronal loss, common features such as abnormal protein deposition, dysfunctional cellular transport, mitochondrial deficits, glutamate excitotoxicity and inflammation are observed in most, if not all, neurodegenerative disorders, suggesting converging pathways of neurodegeneration. We have generated comparative genome-wide gene expression data for Alzheimer’s disease, amyotrophic lateral sclerosis, Huntington’s disease, multiple sclerosis, Parkinson’s disease and schizophrenia using an extensive cohort of well characterised post-mortem CNS tissues. The analysis of whole genome expression patterns across these major disorders offers an outstanding opportunity not only to look into exclusive disease specific changes, but more importantly to uncover potential common molecular pathogenic mechanisms that could be targeted for therapeutic gain. Surprisingly, no dysregulated gene that passed our selection criteria was found in common across all 6 diseases using our primary method of analysis. However, 61 dysregulated genes were shared when comparing five and four diseases. Our analysis indicates firstly the involvement of common neuronal homeostatic, survival and synaptic plasticity pathways. Secondly, we report changes to immunoregulatory and immunomodulatory pathways in all diseases. Our secondary method of analysis confirmed significant up-regulation of a number of genes in diseases presenting degeneration and showed that somatostatin was downregulated in all 6 diseases. The latter is supportive of a general role for neuroinflammation in the pathogenesis and/or response to neurodegeneration. Unravelling the detailed nature of the molecular changes regulating inflammation in the CNS is key to the development of novel therapeutic approaches for these chronic conditions.

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Somatic mutations in Single Human Neurons from Patients with Congenital Neurodegenerative Diseases

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