|
1
|
Arber C, Casey JM, Crawford S, Rambarack N, Yaman U, Wiethoff S, Augustin E, Piers TM, Price M, Rostagno A, Ghiso J, Lewis PA, Revesz T, Hardy J, Pocock JM, Houlden H, Schott JM, Salih DA, Lashley T, Wray S. Microglia contribute to the production of the amyloidogenic ABri peptide in familial British dementia. Acta Neuropathol 2024; 148:65. [PMID: 39546024 PMCID: PMC11568029 DOI: 10.1007/s00401-024-02820-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/10/2024] [Revised: 10/21/2024] [Accepted: 10/30/2024] [Indexed: 11/17/2024]
Abstract
Mutations in ITM2B cause familial British, Danish, Chinese, and Korean dementias. In familial British dementia (FBD), a mutation in the stop codon of the ITM2B gene (also known as BRI2) causes a C-terminal cleavage fragment of the ITM2B/BRI2 protein to be extended by 11 amino acids. This fragment, termed amyloid-Bri (ABri), is highly insoluble and forms extracellular plaques in the brain. ABri plaques are accompanied by tau pathology, neuronal cell death and progressive dementia, with striking parallels to the aetiology and pathogenesis of Alzheimer's disease. The molecular mechanisms underpinning FBD are ill-defined. Using patient-derived induced pluripotent stem cells, we show that expression of ITM2B/BRI2 is 34-fold higher in microglia than neurons and 15-fold higher in microglia compared with astrocytes. This cell-specific enrichment is supported by expression data from both mouse and human brain tissue. ITM2B/BRI2 protein levels are higher in iPSC-microglia compared with neurons and astrocytes. The ABri peptide was detected in patient iPSC-derived microglial lysates and conditioned media but was undetectable in patient-derived neurons and control microglia. The pathological examination of post-mortem tissue supports the presence of ABri in microglia that are in proximity to pre-amyloid deposits. Finally, gene co-expression analysis supports a role for ITM2B/BRI2 in disease-associated microglial responses. These data demonstrate that microglia are major contributors to the production of amyloid forming peptides in FBD, potentially acting as instigators of neurodegeneration. Additionally, these data also suggest ITM2B/BRI2 may be part of a microglial response to disease, motivating further investigations of its role in microglial activation. These data have implications for our understanding of the role of microglia and the innate immune response in the pathogenesis of FBD and other neurodegenerative dementias including Alzheimer's disease.
Collapse
Affiliation(s)
- Charles Arber
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
| | - Jackie M Casey
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
| | - Samuel Crawford
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
- Dementia Research Institute at UCL, London, UK
| | | | - Umran Yaman
- Dementia Research Institute at UCL, London, UK
| | - Sarah Wiethoff
- Klinik für Neurologie mit Institut für Translationale Neurologie Albert Schweitzer Campus, Gebäude A1, 48149, Münster, Germany
| | - Emma Augustin
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
| | - Thomas M Piers
- Department of Neuroinflammation, UCL Queen Square Institute of Neurology, London, UK
| | - Matthew Price
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
| | - Agueda Rostagno
- Department of Pathology, New York University Grossman School of Medicine, New York, USA
| | - Jorge Ghiso
- Department of Pathology, New York University Grossman School of Medicine, New York, USA
| | - Patrick A Lewis
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
- Royal Veterinary College, Royal College Street, London, UK
| | - Tamas Revesz
- The Queen Square Brain Bank for Neurological Disorders, Department of Clinical and Movement Neuroscience, UCL Queen Square Institute of Neurology, London, UK
| | - John Hardy
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK
- Dementia Research Institute at UCL, London, UK
| | - Jennifer M Pocock
- Department of Neuroinflammation, UCL Queen Square Institute of Neurology, London, UK
| | - Henry Houlden
- Department of Neuromuscular Disorders, UCL Queen Square Institute of Neurology, London, UK
| | - Jonathan M Schott
- Dementia Research Centre, UCL Queen Square Institute of Neurology, London, UK
| | | | - Tammaryn Lashley
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK.
| | - Selina Wray
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, London, UK.
| |
Collapse
|
|
2
|
Bhagat R, Minaya MA, Renganathan A, Mehra M, Marsh J, Martinez R, Eteleeb AM, Nana AL, Spina S, Seeley WW, Grinberg LT, Karch CM. Long non-coding RNA SNHG8 drives stress granule formation in tauopathies. Mol Psychiatry 2023; 28:4889-4901. [PMID: 37730840 PMCID: PMC10914599 DOI: 10.1038/s41380-023-02237-2] [Citation(s) in RCA: 9] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/28/2023] [Revised: 08/17/2023] [Accepted: 08/24/2023] [Indexed: 09/22/2023]
Abstract
Tauopathies are a heterogenous group of neurodegenerative disorders characterized by tau aggregation in the brain. In a subset of tauopathies, rare mutations in the MAPT gene, which encodes the tau protein, are sufficient to cause disease; however, the events downstream of MAPT mutations are poorly understood. Here, we investigate the role of long non-coding RNAs (lncRNAs), transcripts >200 nucleotides with low/no coding potential that regulate transcription and translation, and their role in tauopathy. Using stem cell derived neurons from patients carrying a MAPT p.P301L, IVS10 + 16, or p.R406W mutation and CRISPR-corrected isogenic controls, we identified transcriptomic changes that occur as a function of the MAPT mutant allele. We identified 15 lncRNAs that were commonly differentially expressed across the three MAPT mutations. The commonly differentially expressed lncRNAs interact with RNA-binding proteins that regulate stress granule formation. Among these lncRNAs, SNHG8 was significantly reduced in a mouse model of tauopathy and in FTLD-tau, progressive supranuclear palsy, and Alzheimer's disease brains. We show that SNHG8 interacts with tau and stress granule-associated RNA-binding protein TIA1. Overexpression of mutant tau in vitro is sufficient to reduce SNHG8 expression and induce stress granule formation. Rescuing SNHG8 expression leads to reduced stress granule formation and reduced TIA1 levels in immortalized cells and in MAPT mutant neurons, suggesting that dysregulation of this non-coding RNA is a causal factor driving stress granule formation via TIA1 in tauopathies.
Collapse
Affiliation(s)
- Reshma Bhagat
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Miguel A Minaya
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Arun Renganathan
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Muneshwar Mehra
- Department of Neuroscience, Washington University in St Louis, St Louis, MO, USA
| | - Jacob Marsh
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Rita Martinez
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Abdallah M Eteleeb
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA
| | - Alissa L Nana
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA
| | - Salvatore Spina
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA
| | - William W Seeley
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA
- Department of Pathology, University of California, San Francisco, San Francisco, CA, USA
| | - Lea T Grinberg
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, USA
- Department of Pathology, University of California, San Francisco, San Francisco, CA, USA
- Department of Pathology, University of Sao Paulo, São Paulo, Brazil
| | - Celeste M Karch
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, USA.
- Knight Alzheimer Disease Research Center, Washington University in St Louis, St Louis, MO, USA.
| |
Collapse
|
|
3
|
Patani R, Hardingham GE, Liddelow SA. Functional roles of reactive astrocytes in neuroinflammation and neurodegeneration. Nat Rev Neurol 2023; 19:395-409. [PMID: 37308616 DOI: 10.1038/s41582-023-00822-1] [Citation(s) in RCA: 192] [Impact Index Per Article: 96.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 05/11/2023] [Indexed: 06/14/2023]
Abstract
Despite advances in uncovering the mechanisms that underlie neuroinflammation and neurodegenerative disease, therapies that prevent neuronal loss remain elusive. Targeting of disease-defining markers in conditions such as Alzheimer disease (amyloid-β and tau) or Parkinson disease (α-synuclein) has been met with limited success, suggesting that these proteins do not act in isolation but form part of a pathological network. This network could involve phenotypic alteration of multiple cell types in the CNS, including astrocytes, which have a major neurosupportive, homeostatic role in the healthy CNS but adopt reactive states under acute or chronic adverse conditions. Transcriptomic studies in human patients and disease models have revealed the co-existence of many putative reactive sub-states of astrocytes. Inter-disease and even intra-disease heterogeneity of reactive astrocytic sub-states are well established, but the extent to which specific sub-states are shared across different diseases is unclear. In this Review, we highlight how single-cell and single-nuclei RNA sequencing and other 'omics' technologies can enable the functional characterization of defined reactive astrocyte states in various pathological scenarios. We provide an integrated perspective, advocating cross-modal validation of key findings to define functionally important sub-states of astrocytes and their triggers as tractable therapeutic targets with cross-disease relevance.
Collapse
Affiliation(s)
- Rickie Patani
- Department of Neuromuscular Disease, UCL Queen Square Institute of Neurology, London, UK
- The Francis Crick Institute, Human Stem Cells and Neurodegeneration Laboratory, London, UK
| | - Giles E Hardingham
- Euan MacDonald Centre for MND, University of Edinburgh, Edinburgh, UK
- UK Dementia Research Institute at the University of Edinburgh, University of Edinburgh, Edinburgh, UK
- Centre for Discovery Brain Sciences, University of Edinburgh, Edinburgh, UK
| | - Shane A Liddelow
- Neuroscience Institute, NYU Grossman School of Medicine, New York, NY, USA.
- Department of Neuroscience & Physiology, NYU Grossman School of Medicine, New York, NY, USA.
- Department of Ophthalmology, NYU Grossman School of Medicine, New York, NY, USA.
- Parekh Center for Interdisciplinary Neurology, NYU Grossman School of Medicine, New York, NY, USA.
| |
Collapse
|
|
4
|
Bhagat R, Minaya MA, Renganathan A, Mehra M, Marsh J, Martinez R, Nana AL, Spina S, Seeley WW, Grinberg LT, Karch CM. Long non-coding RNA SNHG8 drives stress granule formation in tauopathies. MEDRXIV : THE PREPRINT SERVER FOR HEALTH SCIENCES 2023:2023.02.27.23286548. [PMID: 36909621 PMCID: PMC10002771 DOI: 10.1101/2023.02.27.23286548] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 03/06/2023]
Abstract
Tauopathies are a heterogenous group of neurodegenerative disorders characterized by tau aggregation in the brain. In a subset of tauopathies, rare mutations in the MAPT gene, which encodes the tau protein, are sufficient to cause disease; however, the events downstream of MAPT mutations are poorly understood. Here, we investigate the role of long non-coding RNAs (lncRNAs), transcripts >200 nucleotides with low/no coding potential that regulate transcription and translation, and their role in tauopathy. Using stem cell derived neurons from patients carrying a MAPT p.P301L, IVS10+16, or p.R406W mutation, and CRISPR-corrected isogenic controls, we identified transcriptomic changes that occur as a function of the MAPT mutant allele. We identified 15 lncRNAs that were commonly differentially expressed across the three MAPT mutations. The commonly differentially expressed lncRNAs interact with RNA-binding proteins that regulate stress granule formation. Among these lncRNAs, SNHG8 was significantly reduced in a mouse model of tauopathy and in FTLD-tau, progressive supranuclear palsy, and Alzheimer’s disease brains. We show that SNHG8 interacts with tau and stress granule-associated RNA-binding protein TIA1. Overexpression of mutant tau in vitro is sufficient to reduce SNHG8 expression and induce stress granule formation. Rescuing SNHG8 expression leads to reduced stress granule formation and reduced TIA1 levels, suggesting that dysregulation of this non-coding RNA is a causal factor driving stress granule formation via TIA1 in tauopathies.
Collapse
|
|
5
|
Minaya MA, Mahali S, Iyer AK, Eteleeb AM, Martinez R, Huang G, Budde J, Temple S, Nana AL, Seeley WW, Spina S, Grinberg LT, Harari O, Karch CM. Conserved gene signatures shared among MAPT mutations reveal defects in calcium signaling. Front Mol Biosci 2023; 10:1051494. [PMID: 36845551 PMCID: PMC9948093 DOI: 10.3389/fmolb.2023.1051494] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/22/2022] [Accepted: 01/13/2023] [Indexed: 02/11/2023] Open
Abstract
Introduction: More than 50 mutations in the MAPT gene result in heterogeneous forms of frontotemporal lobar dementia with tau inclusions (FTLD-Tau). However, early pathogenic events that lead to disease and the degree to which they are common across MAPT mutations remain poorly understood. The goal of this study is to determine whether there is a common molecular signature of FTLD-Tau. Methods: We analyzed genes differentially expressed in induced pluripotent stem cell-derived neurons (iPSC-neurons) that represent the three major categories of MAPT mutations: splicing (IVS10 + 16), exon 10 (p.P301L), and C-terminal (p.R406W) compared with isogenic controls. The genes that were commonly differentially expressed in MAPT IVS10 + 16, p.P301L, and p.R406W neurons were enriched in trans-synaptic signaling, neuronal processes, and lysosomal function. Many of these pathways are sensitive to disruptions in calcium homeostasis. One gene, CALB1, was significantly reduced across the three MAPT mutant iPSC-neurons and in a mouse model of tau accumulation. We observed a significant reduction in calcium levels in MAPT mutant neurons compared with isogenic controls, pointing to a functional consequence of this disrupted gene expression. Finally, a subset of genes commonly differentially expressed across MAPT mutations were also dysregulated in brains from MAPT mutation carriers and to a lesser extent in brains from sporadic Alzheimer disease and progressive supranuclear palsy, suggesting that molecular signatures relevant to genetic and sporadic forms of tauopathy are captured in a dish. The results from this study demonstrate that iPSC-neurons capture molecular processes that occur in human brains and can be used to pinpoint common molecular pathways involving synaptic and lysosomal function and neuronal development, which may be regulated by disruptions in calcium homeostasis.
Collapse
Affiliation(s)
- Miguel A. Minaya
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Sidhartha Mahali
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Abhirami K. Iyer
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Abdallah M. Eteleeb
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Rita Martinez
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Guangming Huang
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - John Budde
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
| | - Sally Temple
- Neural Stem Cell Institute, Rensselaer, NY, United States
| | - Alissa L. Nana
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, United States
| | - William W. Seeley
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, United States
| | - Salvatore Spina
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, United States
| | - Lea T. Grinberg
- Department of Neurology, UCSF Weill Institute for Neurosciences, University of California, San Francisco, San Francisco, CA, United States
- Department of Pathology, University of Sao Paulo, Sao Paulo, Brazil
| | - Oscar Harari
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
- Hope Center for Neurological Disorders, Washington University in St Louis, St Louis, MO, United States
- NeuroGenomics and Informatics Center, Washington University in St Louis, St Louis, MO, United States
| | - Celeste M. Karch
- Department of Psychiatry, Washington University in St Louis, St Louis, MO, United States
- Hope Center for Neurological Disorders, Washington University in St Louis, St Louis, MO, United States
- NeuroGenomics and Informatics Center, Washington University in St Louis, St Louis, MO, United States
| |
Collapse
|
|
6
|
Thiry L, Clément JP, Haag R, Kennedy TE, Stifani S. Optimization of Long-Term Human iPSC-Derived Spinal Motor Neuron Culture Using a Dendritic Polyglycerol Amine-Based Substrate. ASN Neuro 2022; 14:17590914211073381. [PMID: 35023784 PMCID: PMC8784909 DOI: 10.1177/17590914211073381] [Citation(s) in RCA: 6] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/17/2021] [Revised: 12/14/2021] [Accepted: 12/22/2021] [Indexed: 11/30/2022] Open
Abstract
Human induced pluripotent stem cells (hiPSCs) derived from healthy and diseased individuals can give rise to many cell types, facilitating the study of mechanisms of development, human disease modeling, and early drug target validation. In this context, experimental model systems based on hiPSC-derived motor neurons (MNs) have been used to study MN diseases such as spinal muscular atrophy and amyotrophic lateral sclerosis. Modeling MN disease using hiPSC-based approaches requires culture conditions that can recapitulate in a dish the events underlying differentiation, maturation, aging, and death of MNs. Current hiPSC-derived MN-based applications are often hampered by limitations in our ability to monitor MN morphology, survival, and other functional properties over a prolonged timeframe, underscoring the need for improved long-term culture conditions. Here we describe a cytocompatible dendritic polyglycerol amine (dPGA) substrate-based method for prolonged culture of hiPSC-derived MNs. We provide evidence that MNs cultured on dPGA-coated dishes are more amenable to long-term study of cell viability, molecular identity, and spontaneous network electrophysiological activity. The present study has the potential to improve hiPSC-based studies of human MN biology and disease.We describe the use of a new coating substrate providing improved conditions for long-term cultures of human iPSC-derived motor neurons, thus allowing evaluation of cell viability, molecular identity, spontaneous network electrophysiological activity, and single-cell RNA sequencing of mature motor neurons.
Collapse
Affiliation(s)
- Louise Thiry
- Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, Canada
| | - Jean-Pierre Clément
- Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, Canada
| | - Rainer Haag
- Institute of Chemistry and Biochemistry, Freie Universität Berlin, Berlin, Germany
| | - Timothy E Kennedy
- Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, Canada
| | - Stefano Stifani
- Department of Neurology and Neurosurgery, Montreal Neurological Institute-Hospital, McGill University, Montreal, Canada
| |
Collapse
|
|
7
|
Potjewyd G, Kellett K, Hooper N. 3D hydrogel models of the neurovascular unit to investigate blood-brain barrier dysfunction. Neuronal Signal 2021; 5:NS20210027. [PMID: 34804595 PMCID: PMC8579151 DOI: 10.1042/ns20210027] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2021] [Revised: 10/20/2021] [Accepted: 10/22/2021] [Indexed: 12/16/2022] Open
Abstract
The neurovascular unit (NVU), consisting of neurons, glial cells, vascular cells (endothelial cells, pericytes and vascular smooth muscle cells (VSMCs)) together with the surrounding extracellular matrix (ECM), is an important interface between the peripheral blood and the brain parenchyma. Disruption of the NVU impacts on blood-brain barrier (BBB) regulation and underlies the development and pathology of multiple neurological disorders, including stroke and Alzheimer's disease (AD). The ability to differentiate induced pluripotent stem cells (iPSCs) into the different cell types of the NVU and incorporate them into physical models provides a reverse engineering approach to generate human NVU models to study BBB function. To recapitulate the in vivo situation such NVU models must also incorporate the ECM to provide a 3D environment with appropriate mechanical and biochemical cues for the cells of the NVU. In this review, we provide an overview of the cells of the NVU and the surrounding ECM, before discussing the characteristics (stiffness, functionality and porosity) required of hydrogels to mimic the ECM when incorporated into in vitro NVU models. We summarise the approaches available to measure BBB functionality and present the techniques in use to develop robust and translatable models of the NVU, including transwell models, hydrogel models, 3D-bioprinting, microfluidic models and organoids. The incorporation of iPSCs either without or with disease-specific genetic mutations into these NVU models provides a platform in which to study normal and disease mechanisms, test BBB permeability to drugs, screen for new therapeutic targets and drugs or to design cell-based therapies.
Collapse
Affiliation(s)
- Geoffrey Potjewyd
- Division of Neuroscience and Experimental Psychology, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, U.K
| | - Katherine A.B. Kellett
- Division of Neuroscience and Experimental Psychology, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, U.K
| | - Nigel M. Hooper
- Division of Neuroscience and Experimental Psychology, School of Biological Sciences, Faculty of Biology, Medicine and Health, University of Manchester, Manchester M13 9PT, U.K
- Geoffrey Jefferson Brain Research Centre, Manchester Academic Health Science Centre, Northern Care Alliance and University of Manchester, Manchester, U.K
| |
Collapse
|
|
8
|
Bergström P, Trybala E, Eriksson CE, Johansson M, Satir TM, Widéhn S, Fruhwürth S, Michno W, Nazir FH, Hanrieder J, Paludan SR, Agholme L, Zetterberg H, Bergström T. Herpes Simplex Virus 1 and 2 Infections during Differentiation of Human Cortical Neurons. Viruses 2021; 13:v13102072. [PMID: 34696502 PMCID: PMC8540961 DOI: 10.3390/v13102072] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2021] [Revised: 10/06/2021] [Accepted: 10/08/2021] [Indexed: 01/02/2023] Open
Abstract
Herpes simplex virus 1 (HSV-1) and 2 (HSV-2) can infect the central nervous system (CNS) with dire consequences; in children and adults, HSV-1 may cause focal encephalitis, while HSV-2 causes meningitis. In neonates, both viruses can cause severe, disseminated CNS infections with high mortality rates. Here, we differentiated human induced pluripotent stem cells (iPSCs) towards cortical neurons for infection with clinical CNS strains of HSV-1 or HSV-2. Progenies from both viruses were produced at equal quantities in iPSCs, neuroprogenitors and cortical neurons. HSV-1 and HSV-2 decreased viability of neuroprogenitors by 36.0% and 57.6% (p < 0.0001), respectively, 48 h post-infection, while cortical neurons were resilient to infection by both viruses. However, in these functional neurons, both HSV-1 and HSV-2 decreased gene expression of two markers of synaptic activity, CAMK2B and ARC, and affected synaptic activity negatively in multielectrode array experiments. However, unaltered secretion levels of the neurodegeneration markers tau and NfL suggested intact axonal integrity. Viral replication of both viruses was found after six days, coinciding with 6-fold and 22-fold increase in gene expression of cellular RNA polymerase II by HSV-1 and HSV-2, respectively. Our results suggest a resilience of human cortical neurons relative to the replication of HSV-1 and HSV-2.
Collapse
Affiliation(s)
- Petra Bergström
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-405 30 Gothenburg, Sweden; (P.B.); (T.M.S.); (S.F.); (F.H.N.); (L.A.)
| | - Edward Trybala
- Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden; (E.T.); (C.E.E.); (M.J.); (S.W.)
| | - Charlotta E. Eriksson
- Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden; (E.T.); (C.E.E.); (M.J.); (S.W.)
| | - Maria Johansson
- Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden; (E.T.); (C.E.E.); (M.J.); (S.W.)
| | - Tugce Munise Satir
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-405 30 Gothenburg, Sweden; (P.B.); (T.M.S.); (S.F.); (F.H.N.); (L.A.)
| | - Sibylle Widéhn
- Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden; (E.T.); (C.E.E.); (M.J.); (S.W.)
| | - Stefanie Fruhwürth
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-405 30 Gothenburg, Sweden; (P.B.); (T.M.S.); (S.F.); (F.H.N.); (L.A.)
- Department of Rheumatology and Inflammation Research, Institute of Medicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden;
| | - Wojciech Michno
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-431 80 Mölndal, Sweden; (W.M.); (J.H.); (H.Z.)
- Department of Neuroscience, Physiology and Pharmacology, University College London, London WC1E 6BT, UK
| | - Faisal Hayat Nazir
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-405 30 Gothenburg, Sweden; (P.B.); (T.M.S.); (S.F.); (F.H.N.); (L.A.)
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-431 80 Mölndal, Sweden; (W.M.); (J.H.); (H.Z.)
| | - Jörg Hanrieder
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-431 80 Mölndal, Sweden; (W.M.); (J.H.); (H.Z.)
- Department of Neurodegenerative Disease, Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK
| | - Soren Riis Paludan
- Department of Rheumatology and Inflammation Research, Institute of Medicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden;
- Department of Biomedicine, Aarhus University, 8000 Aarhus, Denmark
| | - Lotta Agholme
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-405 30 Gothenburg, Sweden; (P.B.); (T.M.S.); (S.F.); (F.H.N.); (L.A.)
| | - Henrik Zetterberg
- Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, SE-431 80 Mölndal, Sweden; (W.M.); (J.H.); (H.Z.)
- Department of Neurodegenerative Disease, Institute of Neurology, University College London, Queen Square, London WC1N 3BG, UK
- UK Dementia Research Institute at University College London, London WC1E 6BT, UK
- Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, SE-431 80 Mölndal, Sweden
| | - Tomas Bergström
- Department of Infectious Diseases, Institute of Biomedicine, Sahlgrenska Academy, University of Gothenburg, SE-413 46 Gothenburg, Sweden; (E.T.); (C.E.E.); (M.J.); (S.W.)
- Correspondence:
| |
Collapse
|
|
9
|
De Los Angeles A, Fernando MB, Hall NAL, Brennand KJ, Harrison PJ, Maher BJ, Weinberger DR, Tunbridge EM. Induced Pluripotent Stem Cells in Psychiatry: An Overview and Critical Perspective. Biol Psychiatry 2021; 90:362-372. [PMID: 34176589 PMCID: PMC8375580 DOI: 10.1016/j.biopsych.2021.04.008] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/17/2021] [Revised: 03/16/2021] [Accepted: 04/07/2021] [Indexed: 02/08/2023]
Abstract
A key challenge in psychiatry research is the development of high-fidelity model systems that can be experimentally manipulated to explore and test pathophysiological mechanisms of illness. In this respect, the emerging capacity to derive neural cells and circuits from human induced pluripotent stem cells (iPSCs) has generated significant excitement. This review aims to provide a critical appraisal of the potential for iPSCs in illuminating pathophysiological mechanisms in the context of other available technical approaches. We discuss the selection of iPSC phenotypes relevant to psychiatry, the information that researchers can draw on to help guide these decisions, and how researchers choose between the use of 2-dimensional cultures and the use of more complex 3-dimensional model systems. We discuss the strengths and limitations of current models and the challenges and opportunities that they present. Finally, we discuss the potential of iPSC-based model systems for clarifying the mechanisms underlying genetic risk for psychiatry and the steps that will be needed to ensure that robust and reliable conclusions can be drawn. We argue that while iPSC-based models are ideally placed to study fundamental processes occurring within and between neural cells, they are often less well suited for case-control studies, given issues relating to statistical power and the challenges in identifying which cellular phenotypes are meaningful at the level of the whole individual. Our aim is to highlight the importance of considering the hypotheses of a given study to guide decisions about which, if any, iPSC-based system is most appropriate to address it.
Collapse
Affiliation(s)
- Alejandro De Los Angeles
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom; Oxford Health NHS Foundation Trust, Oxford, United Kingdom
| | - Michael B Fernando
- Graduate School of Biomedical Science, Icahn School of Medicine at Mount Sinai, New York, New York; Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, New York
| | - Nicola A L Hall
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom; Oxford Health NHS Foundation Trust, Oxford, United Kingdom
| | - Kristen J Brennand
- Nash Family Department of Neuroscience, Icahn School of Medicine at Mount Sinai, New York, New York; Department of Genetics and Genomic Sciences, Icahn School of Medicine at Mount Sinai, New York, New York; Department of Psychiatry, Icahn School of Medicine at Mount Sinai, New York, New York
| | - Paul J Harrison
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom; Oxford Health NHS Foundation Trust, Oxford, United Kingdom
| | - Brady J Maher
- Lieber Institute for Brain Development, Baltimore, Maryland; Department of Psychiatry, Johns Hopkins University School of Medicine, Baltimore, Maryland
| | - Daniel R Weinberger
- Lieber Institute for Brain Development, Baltimore, Maryland; Department of Psychiatry, Johns Hopkins University School of Medicine, Baltimore, Maryland; Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland; Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, Maryland; Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland
| | - Elizabeth M Tunbridge
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom; Oxford Health NHS Foundation Trust, Oxford, United Kingdom.
| |
Collapse
|
|
10
|
Habekost M, Qvist P, Denham M, Holm IE, Jørgensen AL. Directly Reprogrammed Neurons Express MAPT and APP Splice Variants Pertinent to Ageing and Neurodegeneration. Mol Neurobiol 2021; 58:2075-2087. [PMID: 33415685 PMCID: PMC8018937 DOI: 10.1007/s12035-020-02258-w] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/29/2020] [Accepted: 12/09/2020] [Indexed: 12/12/2022]
Abstract
Neurons produced by reprogramming of other cell types are used to study cellular mechanisms of age-related neurodegenerative diseases. To model Alzheimer's disease and other tauopathies, it is essential that alternative splicing of the MAPT transcript in these neurons produces the relevant tau isoforms. Human neurons derived from induced pluripotent stem cells, however, express tau isoform compositions characteristic of foetal neurons rather than of adult neurons unless cultured in vitro for extended time periods. In this study, we characterised the dynamics of the MAPT and APP alternative splicing during a developmental time-course of porcine and murine cerebral cortices. We found age-dependent and species-specific isoform composition of MAPT, including 3R and 4R isoforms in the porcine adult brain similar to that of the adult human brain. We converted adult and embryonic fibroblasts directly into induced neurons and found similar developmental patterns of isoform composition, notably, the 3R and 4R isoforms relevant to the pathogenesis of Alzheimer's disease. Also, we observed cell-type-specific isoform expression of APP transcripts during the conversion. The approach was further used to generate induced neurons from transgenic pigs carrying Alzheimer's disease-causing mutations. We show that such neurons authentically model the first crucial steps in AD pathogenesis.
Collapse
Affiliation(s)
- Mette Habekost
- Department of Biomedicine, Aarhus University, 8000C, Aarhus, Denmark.
- Danish Research Institute of Translational Neuroscience, Nordic EMBL Partnership for Molecular Medicine, Aarhus University, 8000C, Aarhus, Denmark.
| | - Per Qvist
- Department of Biomedicine, Aarhus University, 8000C, Aarhus, Denmark
- iPSYCH, Lundbeck Foundation Initiative for Integrative Psychiatric Research, 8000C, Aarhus, Denmark
- Center for Genomics and Personalized Medicine, 8000C, Aarhus, Denmark
| | - Mark Denham
- Department of Biomedicine, Aarhus University, 8000C, Aarhus, Denmark
- Danish Research Institute of Translational Neuroscience, Nordic EMBL Partnership for Molecular Medicine, Aarhus University, 8000C, Aarhus, Denmark
| | - Ida E Holm
- Department of Pathology, Randers Hospital, 8930, Randers, Denmark
- Department of Clinical Medicine, Aarhus University, 8000C, Aarhus, Denmark
| | | |
Collapse
|
|
11
|
Arber C, Alatza A, Leckey CA, Paterson RW, Zetterberg H, Wray S. Mass spectrometry analysis of tau and amyloid-beta in iPSC-derived models of Alzheimer's disease and dementia. J Neurochem 2021; 159:305-317. [PMID: 33539581 PMCID: PMC8613538 DOI: 10.1111/jnc.15315] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2020] [Revised: 01/18/2021] [Accepted: 01/25/2021] [Indexed: 12/11/2022]
Abstract
Induced pluripotent stem cell (iPSC) technology enables the generation of human neurons in vitro, which contain the precise genome of the cell donor, therefore permitting the generation of disease models from individuals with a disease-associated genotype of interest. This approach has been extensively used to model inherited forms of Alzheimer's disease and frontotemporal dementia. The combination of iPSC-derived neuronal models with targeted mass spectrometry analysis has provided unprecedented insights into the regulation of specific proteins in human neuronal physiology and pathology. For example enabling investigations into tau and APP/Aβ, specifically: protein isoform expression, relative levels of cleavage fragments, aggregated species and functionally critical post-translational modifications. The use of mass spectrometry has enabled a determination of how closely iPSC-derived models recapitulate disease profiles observed in the human brain. This review will highlight the progress to date in studies using iPSCs and mass spectrometry to model Alzheimer's disease and dementia. We go on to convey our optimism, as studies in the near future will make use of this precedent, together with novel techniques such as genome editing and stable isotope labelling, to provide real progress towards an in depth understanding of early neurodegenerative processes and development of novel therapeutic agents.
Collapse
Affiliation(s)
- Charles Arber
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK
| | - Argyro Alatza
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK
| | - Claire A Leckey
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK.,Translational Mass Spectrometry Research Group, UCL Great Ormond Street Institute of Child Health, University College London, London, UK.,UK Dementia Research Institute at UCL, London, UK
| | - Ross W Paterson
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK
| | - Henrik Zetterberg
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK.,UK Dementia Research Institute at UCL, London, UK.,Department of Psychiatry and Neurochemistry, Institute of Neuroscience and Physiology, Sahlgrenska Academy at the University of Gothenburg, Mölndal, Sweden.,Clinical Neurochemistry Laboratory, Sahlgrenska University Hospital, Mölndal, Sweden
| | - Selina Wray
- Department of Neurodegenerative Disease, UCL Queen Square Institute of Neurology, University College London, London, UK
| |
Collapse
|
|
12
|
Franklin H, Clarke BE, Patani R. Astrocytes and microglia in neurodegenerative diseases: Lessons from human in vitro models. Prog Neurobiol 2020; 200:101973. [PMID: 33309801 PMCID: PMC8052192 DOI: 10.1016/j.pneurobio.2020.101973] [Citation(s) in RCA: 27] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2020] [Revised: 11/06/2020] [Accepted: 12/06/2020] [Indexed: 12/16/2022]
Abstract
Astrocytes and microglia key fulfil homeostatic and immune functions in the CNS. Dysfunction of these cell types is implicated in neurodegenerative diseases. Understanding cellular autonomy and early pathogenic changes is a key goal. New human iPSC models will inform on disease mechanisms and therapy development. Both astrocytes and microglia fulfil homeostatic and immune functions in the healthy CNS. Dysfunction of these cell types have been implicated in the pathomechanisms of several neurodegenerative diseases. Understanding the cellular autonomy and early pathological changes in these cell types may inform drug screening and therapy development. While animal models and post-mortem tissue have been invaluable in understanding disease processes, the advent of human in vitro models provides a unique insight into disease biology as a manipulable model system obtained directly from patients. Here, we discuss the different human in vitro models of astrocytes and microglia and outline the phenotypes that have been recapitulated in these systems.
Collapse
Affiliation(s)
- Hannah Franklin
- The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK; Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, Queen Square, London, UK
| | - Benjamin E Clarke
- The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK; Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, Queen Square, London, UK
| | - Rickie Patani
- The Francis Crick Institute, 1 Midland Road, London, NW1 1AT, UK; Department of Neuromuscular Diseases, UCL Queen Square Institute of Neurology, Queen Square, London, UK.
| |
Collapse
|
|
13
|
Modelling frontotemporal dementia using patient-derived induced pluripotent stem cells. Mol Cell Neurosci 2020; 109:103553. [PMID: 32956830 DOI: 10.1016/j.mcn.2020.103553] [Citation(s) in RCA: 20] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/08/2020] [Revised: 08/27/2020] [Accepted: 09/12/2020] [Indexed: 12/12/2022] Open
Abstract
Frontotemporal dementia (FTD) describes a group of clinically heterogeneous conditions that frequently affect people under the age of 65 (Le Ber et al., 2013). There are multiple genetic causes of FTD, including coding or splice-site mutations in MAPT, GRN mutations that lead to haploinsufficiency of progranulin protein, and a hexanucleotide GGGGCC repeat expansion in C9ORF72. Pathologically, FTD is characterised by abnormal protein accumulations in neurons and glia. These aggregates can be composed of the microtubule-associated protein tau (observed in FTD with MAPT mutations), the DNA/RNA-binding protein TDP-43 (seen in FTD with mutations in GRN or C9ORF72 repeat expansions) or dipeptide proteins generated by repeat associated non-ATG translation of the C9ORF72 repeat expansion. There are currently no disease-modifying therapies for FTD and the availability of in vitro models that recapitulate pathologies in a disease-relevant cell type would accelerate the development of novel therapeutics. It is now possible to generate patient-specific stem cells through the reprogramming of somatic cells from a patient with a genotype/phenotype of interest into induced pluripotent stem cells (iPSCs). iPSCs can subsequently be differentiated into a plethora of cell types including neurons, astrocytes and microglia. Using this approach has allowed researchers to generate in vitro models of genetic FTD in human cell types that are largely inaccessible during life. In this review we explore the recent progress in the use of iPSCs to model FTD, and consider the merits, limitations and future prospects of this approach.
Collapse
|
|
14
|
Wang Y, Patani R. Novel therapeutic targets for amyotrophic lateral sclerosis: ribonucleoproteins and cellular autonomy. Expert Opin Ther Targets 2020; 24:971-984. [PMID: 32746659 DOI: 10.1080/14728222.2020.1805734] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
INTRODUCTION Amyotrophic lateral sclerosis (ALS) is a devastating disease with a lifetime risk of approximately 1:400. It is incurable and invariably fatal. Average survival is between 3 and 5 years and patients become increasingly paralyzed, losing the ability to speak, eat, and breathe. Therapies in development either (i) target specific familial forms of ALS (comprising a minority of around 10% of cases) or ii) emanate from (over)reliance on animal models or non-human/non-neuronal cell models. There is a desperate and unmet clinical need for effective treatments. Deciphering the primacy and relative contributions of defective protein homeostasis and RNA metabolism in ALS across different model systems will facilitate the identification of putative therapeutic targets. AREAS COVERED This review examines the putative common primary molecular events that lead to ALS pathogenesis. We focus on deregulated RNA metabolism, protein mislocalization/pathological aggregation and the role of glia in ALS-related motor neuron degeneration. Finally, we describe promising targets for therapeutic evaluation. EXPERT OPINION Moving forward, an effective strategy could be achieved by a poly-therapeutic approach which targets both deregulated RNA metabolism and protein dyshomeostasis in the relevant cell types, at the appropriate phase of disease.
Collapse
Affiliation(s)
- Yiran Wang
- Department of Neuromuscular Diseases, Queen Square Institute of Neurology, University College London , London, UK.,Human Stem Cells and Neurodegeneration Laboratory, The Francis Crick Institute , London, UK
| | - Rickie Patani
- Department of Neuromuscular Diseases, Queen Square Institute of Neurology, University College London , London, UK.,Human Stem Cells and Neurodegeneration Laboratory, The Francis Crick Institute , London, UK
| |
Collapse
|
|
15
|
Karagiannis P, Inoue H. ALS, a cellular whodunit on motor neuron degeneration. Mol Cell Neurosci 2020; 107:103524. [PMID: 32629110 DOI: 10.1016/j.mcn.2020.103524] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2020] [Revised: 06/09/2020] [Accepted: 06/25/2020] [Indexed: 12/24/2022] Open
Abstract
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease that primarily targets motor neurons. Motor neurons from ALS patients show cytoplasmic inclusions that are reflective of an altered RNA metabolism and protein degradation. Causal gene mutations are found in all cell types even though patient motor neurons are by far the most susceptible to the degeneration. Using induced pluripotent stem cell (iPSC) technology, researchers have generated motor neurons with the same genotype as the patient including sporadic ones. They have also generated other cell types associated with the disease such as astrocytes, microglia and oligodendrocytes. These cells provide not only new insights on the mechanisms of the disease from the early stage, but also a platform for drug screening that has led to several clinical trials. This review examines the knowledge gained from iPSC studies using patient cells on the gene mutations and cellular networks in ALS and relevant experimental therapies.
Collapse
Affiliation(s)
- Peter Karagiannis
- Center for iPS Cell Research and Application, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan.
| | - Haruhisa Inoue
- Center for iPS Cell Research and Application, Kyoto University, 53 Kawahara-cho, Shogoin, Sakyo-ku, Kyoto 606-8507, Japan; iPSC-based Drug Discovery and Development Team, RIKEN BioResource Research Center (BRC), Kyoto, Japan; Medical-risk Avoidance based on iPS Cells Team, RIKEN Center for Advanced Intelligence Project (AIP), Kyoto, Japan.
| |
Collapse
|
|
16
|
Cairns DM, Rouleau N, Parker RN, Walsh KG, Gehrke L, Kaplan DL. A 3D human brain-like tissue model of herpes-induced Alzheimer's disease. SCIENCE ADVANCES 2020; 6:eaay8828. [PMID: 32494701 PMCID: PMC7202879 DOI: 10.1126/sciadv.aay8828] [Citation(s) in RCA: 162] [Impact Index Per Article: 32.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 07/25/2019] [Accepted: 02/18/2020] [Indexed: 05/23/2023]
Abstract
Alzheimer's disease (AD) is a neurodegenerative disorder that causes cognitive decline, memory loss, and inability to perform everyday functions. Hallmark features of AD-including generation of amyloid plaques, neurofibrillary tangles, gliosis, and inflammation in the brain-are well defined; however, the cause of the disease remains elusive. Growing evidence implicates pathogens in AD development, with herpes simplex virus type I (HSV-1) gaining increasing attention as a potential causative agent. Here, we describe a multidisciplinary approach to produce physiologically relevant human tissues to study AD using human-induced neural stem cells (hiNSCs) and HSV-1 infection in a 3D bioengineered brain model. We report a herpes-induced tissue model of AD that mimics human disease with multicellular amyloid plaque-like formations, gliosis, neuroinflammation, and decreased functionality, completely in the absence of any exogenous mediators of AD. This model will allow for future studies to identify potential downstream drug targets for treating this devastating disease.
Collapse
Affiliation(s)
- Dana M. Cairns
- Department of Biomedical Engineering, Tufts University, Medford, MA 02155, USA
- Allen Discovery Center, Tufts University, Medford, MA 02155, USA
| | - Nicolas Rouleau
- Department of Biomedical Engineering, Tufts University, Medford, MA 02155, USA
- Allen Discovery Center, Tufts University, Medford, MA 02155, USA
| | - Rachael N. Parker
- Department of Biomedical Engineering, Tufts University, Medford, MA 02155, USA
| | | | - Lee Gehrke
- Institute for Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA 02142, USA
| | - David L. Kaplan
- Department of Biomedical Engineering, Tufts University, Medford, MA 02155, USA
- Allen Discovery Center, Tufts University, Medford, MA 02155, USA
| |
Collapse
|
|
17
|
Neural In Vitro Models for Studying Substances Acting on the Central Nervous System. Handb Exp Pharmacol 2020; 265:111-141. [PMID: 32594299 DOI: 10.1007/164_2020_367] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Animal models have been greatly contributing to our understanding of physiology, mechanisms of diseases, and toxicity. Yet, their limitations due to, e.g., interspecies variation are reflected in the high number of drug attrition rates, especially in central nervous system (CNS) diseases. Therefore, human-based neural in vitro models for studying safety and efficacy of substances acting on the CNS are needed. Human iPSC-derived cells offer such a platform with the unique advantage of reproducing the "human context" in vitro by preserving the genetic and molecular phenotype of their donors. Guiding the differentiation of hiPSC into cells of the nervous system and combining them in a 2D or 3D format allows to obtain complex models suitable for investigating neurotoxicity or brain-related diseases with patient-derived cells. This chapter will give an overview over stem cell-based human 2D neuronal and mixed neuronal/astrocyte models, in vitro cultures of microglia, as well as CNS disease models and considers new developments in the field, more specifically the use of brain organoids and 3D bioprinted in vitro models for safety and efficacy evaluation.
Collapse
|
|
18
|
Vanneste J, Vercruysse T, Boeynaems S, Sicart A, Van Damme P, Daelemans D, Van Den Bosch L. C9orf72-generated poly-GR and poly-PR do not directly interfere with nucleocytoplasmic transport. Sci Rep 2019; 9:15728. [PMID: 31673013 PMCID: PMC6823349 DOI: 10.1038/s41598-019-52035-6] [Citation(s) in RCA: 43] [Impact Index Per Article: 7.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2019] [Accepted: 10/11/2019] [Indexed: 12/29/2022] Open
Abstract
Repeat expansions in the C9orf72 gene cause amyotrophic lateral sclerosis and frontotemporal dementia characterized by dipeptide-repeat protein (DPR) inclusions. The toxicity associated with two of these DPRs, poly-GR and poly-PR, has been associated with nucleocytoplasmic transport. To investigate the causal role of poly-GR or poly-PR on active nucleocytoplasmic transport, we measured nuclear import and export in poly-GR or poly-PR expressing Hela cells, neuronal-like SH-SY5Y cells and iPSC-derived motor neurons. Our data strongly indicate that poly-GR and poly-PR do not directly impede active nucleocytoplasmic transport.
Collapse
Affiliation(s)
- Joni Vanneste
- KU Leuven - University of Leuven, Department of Neurosciences, Experimental Neurology and Leuven Brain Institute (LBI), Leuven, Belgium
- VIB, Center for Brain & Disease Research, Laboratory of Neurobiology, Leuven, Belgium
| | - Thomas Vercruysse
- KU Leuven Department of Microbiology, Immunology and Transplantation, Laboratory of Virology and Chemotherapy, Rega Institute for Medical Research, Leuven, Belgium
| | - Steven Boeynaems
- Department of Genetics, Stanford University School of Medicine, Stanford, USA
| | - Adria Sicart
- KU Leuven - University of Leuven, Department of Neurosciences, Experimental Neurology and Leuven Brain Institute (LBI), Leuven, Belgium
- VIB, Center for Brain & Disease Research, Laboratory of Neurobiology, Leuven, Belgium
| | - Philip Van Damme
- KU Leuven - University of Leuven, Department of Neurosciences, Experimental Neurology and Leuven Brain Institute (LBI), Leuven, Belgium
- VIB, Center for Brain & Disease Research, Laboratory of Neurobiology, Leuven, Belgium
- University Hospitals Leuven, Department of Neurology, Leuven, Belgium
| | - Dirk Daelemans
- KU Leuven Department of Microbiology, Immunology and Transplantation, Laboratory of Virology and Chemotherapy, Rega Institute for Medical Research, Leuven, Belgium
| | - Ludo Van Den Bosch
- KU Leuven - University of Leuven, Department of Neurosciences, Experimental Neurology and Leuven Brain Institute (LBI), Leuven, Belgium.
- VIB, Center for Brain & Disease Research, Laboratory of Neurobiology, Leuven, Belgium.
| |
Collapse
|
|
19
|
McComish SF, Caldwell MA. Generation of defined neural populations from pluripotent stem cells. Philos Trans R Soc Lond B Biol Sci 2019; 373:rstb.2017.0214. [PMID: 29786550 DOI: 10.1098/rstb.2017.0214] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 03/06/2018] [Indexed: 12/25/2022] Open
Abstract
Effective and efficient generation of human neural stem cells and subsequently functional neural populations from pluripotent stem cells has facilitated advancements in the study of human development and disease modelling. This review will discuss the established protocols for the generation of defined neural populations including regionalized neurons and astrocytes, oligodendrocytes and microglia. Early protocols were established in embryonic stem cells (ESC) but the discovery of induced pluripotent stem cells (iPSC) in 2006 provided a new platform for modelling human disorders of the central nervous system (CNS). The ability to produce patient- and disease-specific iPSC lines has created a new age of disease modelling. Human iPSC may be derived from adult somatic cells and subsequently patterned into numerous distinct cell types. The ability to derive defined and regionalized neural populations from iPSC provides a powerful in vitro model of CNS disorders.This article is part of the theme issue 'Designer human tissue: coming to a lab near you'.
Collapse
Affiliation(s)
- Sarah F McComish
- Department of Physiology, Trinity College Institute for Neuroscience, Trinity College Dublin, Dublin 2, Ireland
| | - Maeve A Caldwell
- Department of Physiology, Trinity College Institute for Neuroscience, Trinity College Dublin, Dublin 2, Ireland
| |
Collapse
|
|
20
|
TCW J. Human iPSC application in Alzheimer’s disease and Tau-related neurodegenerative diseases. Neurosci Lett 2019; 699:31-40. [DOI: 10.1016/j.neulet.2019.01.043] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2018] [Revised: 11/23/2018] [Accepted: 01/23/2019] [Indexed: 12/11/2022]
|
|
21
|
Ziff OJ, Patani R. Harnessing cellular aging in human stem cell models of amyotrophic lateral sclerosis. Aging Cell 2019; 18:e12862. [PMID: 30565851 PMCID: PMC6351881 DOI: 10.1111/acel.12862] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/15/2018] [Accepted: 09/17/2018] [Indexed: 02/06/2023] Open
Abstract
Amyotrophic lateral sclerosis (ALS) is a relentlessly progressive neurodegenerative condition that is invariably fatal, usually within 3 to 5 years of diagnosis. The etiology of ALS remains unresolved and no effective treatments exist. There is therefore a desperate and unmet need for discovery of disease mechanisms to guide novel therapeutic strategies. The single major risk factor for ALS is aging, yet the molecular consequences of cell type‐specific aging remain understudied in this context. Induced pluripotent stem cells (iPSCs) have transformed the standard approach of examining human disease, generating unlimited numbers of disease‐relevant cells from patients, enabling analysis of disease mechanisms and drug screening. However, reprogramming patient cells to iPSCs reverses key hallmarks of cellular age. Therefore, although iPSC models recapitulate some disease hallmarks, a crucial challenge is to address the disparity between the advanced age of onset of neurodegenerative diseases and the fetal‐equivalent maturational state of iPSC‐derivatives. Increasing recognition of cell type‐specific aging paradigms underscores the importance of heterogeneity in ultimately tipping the balance from a state of compensated dysfunction (clinically pre‐symptomatic) to decompensation and progression (irreversible loss of neurological functions). In order to realize the true promise of iPSC technology in ALS, efforts need to prioritize faithfully recapitulating the clinical pathophysiological state, with proportionate emphasis on capturing the molecular sequelae of both cellular age and non‐cell‐autonomous disease mechanisms within this context.
Collapse
Affiliation(s)
- Oliver J. Ziff
- The Institute of Neurology; University College London; London UK
- The Francis Crick Institute; London UK
| | - Rickie Patani
- The Institute of Neurology; University College London; London UK
- The Francis Crick Institute; London UK
| |
Collapse
|
|
22
|
Lee SY, George JH, Nagel DA, Ye H, Kueberuwa G, Seymour LW. Optogenetic control of iPS cell-derived neurons in 2D and 3D culture systems using channelrhodopsin-2 expression driven by the synapsin-1 and calcium-calmodulin kinase II promoters. J Tissue Eng Regen Med 2019; 13:369-384. [PMID: 30550638 PMCID: PMC6492196 DOI: 10.1002/term.2786] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/01/2017] [Revised: 09/04/2018] [Accepted: 11/30/2018] [Indexed: 01/01/2023]
Abstract
Development of an optogenetically controllable human neural network model in three-dimensional (3D) cultures can provide an investigative system that is more physiologically relevant and better able to mimic aspects of human brain function. Light-sensitive neurons were generated by transducing channelrhodopsin-2 (ChR2) into human induced pluripotent stem cell (hiPSC) derived neural progenitor cells (Axol) using lentiviruses and cell-type specific promoters. A mixed population of human iPSC-derived cortical neurons, astrocytes and progenitor cells were obtained (Axol-ChR2) upon neural differentiation. Pan-neuronal promoter synapsin-1 (SYN1) and excitatory neuron-specific promoter calcium-calmodulin kinase II (CaMKII) were used to drive reporter gene expression in order to assess the differentiation status of the targeted cells. Expression of ChR2 and characterisation of subpopulations in differentiated Axol-ChR2 cells were evaluated using flow cytometry and immunofluorescent staining. These cells were transferred from 2D culture to 3D alginate hydrogel functionalised with arginine-glycine-aspartate (RGD) and small molecules (Y-27632). Improved RGD-alginate hydrogel was physically characterised and assessed for cell viability to serve as a generic 3D culture system for human pluripotent stem cells (hPSCs) and neuronal cells. Prior to cell encapsulation, neural network activities of Axol-ChR2 cells and primary neurons were investigated using calcium imaging. Results demonstrate that functional activities were successfully achieved through expression of ChR2- by both the CaMKII and SYN1 promoters. The RGD-alginate hydrogel system supports the growth of differentiated Axol-ChR2 cells whilst allowing detection of ChR2 expression upon light stimulation. This allows precise and non-invasive control of human neural networks in 3D.
Collapse
Affiliation(s)
- Si-Yuen Lee
- Department of Oncology, Old Road Campus Research Building, University of Oxford, Oxford, UK.,Institute of Biomedical Engineering, Old Road Campus Research Building, University of Oxford, Oxford, UK
| | - Julian H George
- Institute of Biomedical Engineering, Old Road Campus Research Building, University of Oxford, Oxford, UK
| | - David A Nagel
- School of Life and Health Sciences, University of Aston, Birmingham, UK
| | - Hua Ye
- Institute of Biomedical Engineering, Old Road Campus Research Building, University of Oxford, Oxford, UK
| | - Gray Kueberuwa
- Department of Cancer Sciences, Manchester Cancer Research Centre, University of Manchester, Manchester, UK
| | - Leonard W Seymour
- Department of Oncology, Old Road Campus Research Building, University of Oxford, Oxford, UK
| |
Collapse
|
|
23
|
Rowland HA, Hooper NM, Kellett KAB. Modelling Sporadic Alzheimer's Disease Using Induced Pluripotent Stem Cells. Neurochem Res 2018; 43:2179-2198. [PMID: 30387070 PMCID: PMC6267251 DOI: 10.1007/s11064-018-2663-z] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2018] [Revised: 09/11/2018] [Accepted: 10/15/2018] [Indexed: 12/24/2022]
Abstract
Developing cellular models of sporadic Alzheimer's disease (sAD) is challenging due to the unknown initiator of disease onset and the slow disease progression that takes many years to develop in vivo. The use of human induced pluripotent stem cells (iPSCs) has revolutionised the opportunities to model AD pathology, investigate disease mechanisms and screen potential drugs. The majority of this work has, however, used cells derived from patients with familial AD (fAD) where specific genetic mutations drive disease onset. While these provide excellent models to investigate the downstream pathways involved in neuronal toxicity and ultimately neuronal death that leads to AD, they provide little insight into the causes and mechanisms driving the development of sAD. In this review we compare the data obtained from fAD and sAD iPSC-derived cell lines, identify the inconsistencies that exist in sAD models and highlight the potential role of Aβ clearance mechanisms, a relatively under-investigated area in iPSC-derived models, in the study of AD. We discuss the development of more physiologically relevant models using co-culture and three-dimensional culture of iPSC-derived neurons with glial cells. Finally, we evaluate whether we can develop better, more consistent models for sAD research using genetic stratification of iPSCs and identification of genetic and environmental risk factors that could be used to initiate disease onset for modelling sAD. These considerations provide exciting opportunities to develop more relevant iPSC models of sAD which can help drive our understanding of disease mechanisms and identify new therapeutic targets.
Collapse
Affiliation(s)
- Helen A Rowland
- Division of Neuroscience & Experimental Psychology, School of Biological Sciences, Faculty of Biology Medicine and Health, University of Manchester, Manchester, UK
| | - Nigel M Hooper
- Division of Neuroscience & Experimental Psychology, School of Biological Sciences, Faculty of Biology Medicine and Health, University of Manchester, Manchester, UK
| | - Katherine A B Kellett
- Division of Neuroscience & Experimental Psychology, School of Biological Sciences, Faculty of Biology Medicine and Health, University of Manchester, Manchester, UK.
| |
Collapse
|
|
24
|
Grainger AI, King MC, Nagel DA, Parri HR, Coleman MD, Hill EJ. In vitro Models for Seizure-Liability Testing Using Induced Pluripotent Stem Cells. Front Neurosci 2018; 12:590. [PMID: 30233290 PMCID: PMC6127295 DOI: 10.3389/fnins.2018.00590] [Citation(s) in RCA: 43] [Impact Index Per Article: 6.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/14/2018] [Accepted: 08/06/2018] [Indexed: 12/14/2022] Open
Abstract
The brain is the most complex organ in the body, controlling our highest functions, as well as regulating myriad processes which incorporate the entire physiological system. The effects of prospective therapeutic entities on the brain and central nervous system (CNS) may potentially cause significant injury, hence, CNS toxicity testing forms part of the “core battery” of safety pharmacology studies. Drug-induced seizure is a major reason for compound attrition during drug development. Currently, the rat ex vivo hippocampal slice assay is the standard option for seizure-liability studies, followed by primary rodent cultures. These models can respond to diverse agents and predict seizure outcome, yet controversy over the relevance, efficacy, and cost of these animal-based methods has led to interest in the development of human-derived models. Existing platforms often utilize rodents, and so lack human receptors and other drug targets, which may produce misleading data, with difficulties in inter-species extrapolation. Current electrophysiological approaches are typically used in a low-throughput capacity and network function may be overlooked. Human-derived induced pluripotent stem cells (iPSCs) are a promising avenue for neurotoxicity testing, increasingly utilized in drug screening and disease modeling. Furthermore, the combination of iPSC-derived models with functional techniques such as multi-electrode array (MEA) analysis can provide information on neuronal network function, with increased sensitivity to neurotoxic effects which disrupt different pathways. The use of an in vitro human iPSC-derived neural model for neurotoxicity studies, combined with high-throughput techniques such as MEA recordings, could be a suitable addition to existing pre-clinical seizure-liability testing strategies.
Collapse
Affiliation(s)
| | - Marianne C King
- Life and Health Sciences, Aston University, Birmingham, United Kingdom
| | - David A Nagel
- Life and Health Sciences, Aston University, Birmingham, United Kingdom
| | - H Rheinallt Parri
- Life and Health Sciences, Aston University, Birmingham, United Kingdom
| | - Michael D Coleman
- Life and Health Sciences, Aston University, Birmingham, United Kingdom
| | - Eric J Hill
- Life and Health Sciences, Aston University, Birmingham, United Kingdom
| |
Collapse
|
|
25
|
Robbins JP, Perfect L, Ribe EM, Maresca M, Dangla-Valls A, Foster EM, Killick R, Nowosiad P, Reid MJ, Polit LD, Nevado AJ, Ebner D, Bohlooly-Y M, Buckley N, Pangalos MN, Price J, Lovestone S. Clusterin Is Required for β-Amyloid Toxicity in Human iPSC-Derived Neurons. Front Neurosci 2018; 12:504. [PMID: 30090055 PMCID: PMC6068261 DOI: 10.3389/fnins.2018.00504] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/05/2018] [Accepted: 07/04/2018] [Indexed: 02/01/2023] Open
Abstract
Our understanding of the molecular processes underlying Alzheimer's disease (AD) is still limited, hindering the development of effective treatments, and highlighting the need for human-specific models. Advances in identifying components of the amyloid cascade are progressing, including the role of the protein clusterin in mediating β-amyloid (Aβ) toxicity. Mutations in the clusterin gene (CLU), a major genetic AD risk factor, are known to have important roles in Aβ processing. Here we investigate how CLU mediates Aβ-driven neurodegeneration in human induced pluripotent stem cell (iPSC)-derived neurons. We generated a novel CLU-knockout iPSC line by CRISPR/Cas9-mediated gene editing to investigate Aβ-mediated neurodegeneration in cortical neurons differentiated from wild type and CLU knockout iPSCs. We measured response to Aβ using an imaging assay and measured changes in gene expression using qPCR and RNA sequencing. In wild type neurons imaging indicated that neuronal processes degenerate following treatment with Aβ25-35 peptides and Aβ1-42 oligomers, in a dose dependent manner, and that intracellular levels of clusterin are increased following Aβ treatment. However, in CLU knockout neurons Aβ exposure did not affect neurite length, suggesting that clusterin is an important component of the amyloid cascade. Transcriptomic data were analyzed to elucidate the pathways responsible for the altered response to Aβ in neurons with the CLU deletion. Four of the five genes previously identified as downstream to Aβ and Dickkopf-1 (DKK1) proteins in an Aβ-driven neurotoxic pathway in rodent cells were also dysregulated in human neurons with the CLU deletion. AD and lysosome pathways were the most significantly dysregulated pathways in the CLU knockout neurons, and pathways relating to cytoskeletal processes were most dysregulated in Aβ treated neurons. The absence of neurodegeneration in the CLU knockout neurons in response to Aβ compared to the wild type neurons supports the role of clusterin in Aβ-mediated AD pathogenesis.
Collapse
Affiliation(s)
| | - Leo Perfect
- Department of Basic and Clinical Neuroscience, King's College London, London, United Kingdom
| | - Elena M Ribe
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom
| | - Marcello Maresca
- Translational Genomics, Discovery Sciences, Innovative Medicines and Early Development Biotech Unit, AstraZeneca, Gothenburg, Sweden
| | | | | | - Richard Killick
- Department of Old Age Psychiatry, King's College London, London, United Kingdom
| | - Paulina Nowosiad
- Department of Basic and Clinical Neuroscience, King's College London, London, United Kingdom
| | - Matthew J Reid
- Department of Basic and Clinical Neuroscience, King's College London, London, United Kingdom
| | - Lucia Dutan Polit
- Department of Basic and Clinical Neuroscience, King's College London, London, United Kingdom
| | - Alejo J Nevado
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom
| | - Daniel Ebner
- Nuffield Department of Medicine, Target Discovery Institute, University of Oxford, Oxford, United Kingdom
| | - Mohammad Bohlooly-Y
- Translational Genomics, Discovery Sciences, Innovative Medicines and Early Development Biotech Unit, AstraZeneca, Gothenburg, Sweden
| | - Noel Buckley
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom
| | - Menelas N Pangalos
- Innovative Medicines and Early Development Biotech Unit, AstraZeneca, Cambridge, United Kingdom
| | - Jack Price
- Department of Basic and Clinical Neuroscience, King's College London, London, United Kingdom
| | - Simon Lovestone
- Department of Psychiatry, University of Oxford, Oxford, United Kingdom
| |
Collapse
|
|
26
|
Wray S. Modeling tau pathology in human stem cell derived neurons. Brain Pathol 2018; 27:525-529. [PMID: 28585382 DOI: 10.1111/bpa.12521] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/18/2017] [Accepted: 04/25/2017] [Indexed: 01/01/2023] Open
Abstract
Tau pathology is a defining characteristic of multiple neurodegenerative disorders including Alzheimer's disease (AD) and Frontotemporal Dementia (FTD) with tau pathology. There is strong evidence from genetics and experimental models to support a central role for tau dysfunction in neuronal death, suggesting tau is a promising therapeutic target for AD and FTD. However, the development of tau pathology can precede symptom onset by several years, so understanding the earliest molecular events in tauopathy is a priority area of research. Induced pluripotent stem cells (iPSC) derived from patients with genetic causes of tauopathy provide an opportunity to derive limitless numbers of human neurons with physiologically appropriate expression levels of mutated genes for in vitro studies into disease mechanisms. This review discusses the progress made to date using this approach and highlights some of the challenges and unanswered questions this technology has the potential to address.
Collapse
Affiliation(s)
- Selina Wray
- Department of Molecular Neuroscience, UCL Institute of Neurology, London, WC1N 1PJ, UK
| |
Collapse
|
|
27
|
Serio A, Patani R. Concise Review: The Cellular Conspiracy of Amyotrophic Lateral Sclerosis. Stem Cells 2018; 36:293-303. [PMID: 29235200 DOI: 10.1002/stem.2758] [Citation(s) in RCA: 35] [Impact Index Per Article: 5.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2017] [Revised: 11/18/2017] [Accepted: 12/04/2017] [Indexed: 12/12/2022]
Abstract
Amyotrophic lateral sclerosis (ALS) is incurable and devastating. A dearth of therapies has galvanized experimental focus onto the cellular and molecular mechanisms that both initiate and subsequently drive motor neuron degeneration. A traditional view of ALS pathogenesis posits that disease-specific injury to a subtype of neurons is mechanistically cell-autonomous. This "neuron-centric" view has biased past research efforts. However, a wealth of accumulating evidence now strongly implicates non-neuronal cells as being major determinants of ALS. Although animal models have proven invaluable in basic neuroscience research, a growing number of studies confirm fundamental interspecies differences between popular model organisms and the human condition. This may in part explain the failure of therapeutic translation from rodent preclinical models. It follows that integration of a human experimental model using patient-specific induced pluripotent stem cells may be necessary to capture the complexity of human neurodegeneration with fidelity. Integration of enriched human neuronal and glial experimental platforms into the existing repertoire of preclinical models might prove transformational for clinical trial outcomes in ALS. Such reductionist and integrated cross-modal approaches allow systematic elucidation of cell-autonomous and non-cell-autonomous mechanisms of disease, which may then provide novel cellular targets for therapeutic intervention. Stem Cells 2018;36:293-303.
Collapse
Affiliation(s)
- Andrea Serio
- Tissue Engineering and Biophotonics Division, Dental Institute, Kings College London, London, United Kingdom
| | - Rickie Patani
- Department of Molecular Neuroscience, Institute of Neurology, University College London, London, United Kingdom
- The Francis Crick Institute, London, United Kingdom
| |
Collapse
|
|
28
|
Use of Human Neurons Derived via Cellular Reprogramming Methods to Study Host-Parasite Interactions of Toxoplasma gondii in Neurons. Cells 2017; 6:cells6040032. [PMID: 28946615 PMCID: PMC5755492 DOI: 10.3390/cells6040032] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/13/2017] [Revised: 09/12/2017] [Accepted: 09/22/2017] [Indexed: 12/31/2022] Open
Abstract
Toxoplasma gondii is an intracellular protozoan parasite, with approximately one-third of the worlds' population chronically infected. In chronically infected individuals, the parasite resides in tissue cysts in neurons in the brain. The chronic infection in immunocompetant individuals has traditionally been considered to be asymptomatic, but increasing evidence indicates that chronic infection is associated with diverse neurological disorders such as schizophrenia, cryptogenic epilepsy, and Parkinson's Disease. The mechanisms by which the parasite exerts affects on behavior and other neuronal functions are not understood. Human neurons derived from cellular reprogramming methods offer the opportunity to develop better human neuronal models to study T. gondii in neurons. Results from two studies using human neurons derived via cellular reprogramming methods indicate these human neuronal models provide better in vitro models to study the effects of T. gondii on neurons and neurological functions. In this review, an overview of the current neural reprogramming methods will be given, followed by a summary of the studies using human induced pluripotent stem cell (hiPSC)-derived neurons and induced neurons (iNs) to study T. gondii in neurons. The potential of these neural reprogramming methods for further study of the host-parasite interactions of T. gondii in neurons will be discussed.
Collapse
|
|
29
|
Arber C, Lovejoy C, Wray S. Stem cell models of Alzheimer's disease: progress and challenges. ALZHEIMERS RESEARCH & THERAPY 2017; 9:42. [PMID: 28610595 PMCID: PMC5470327 DOI: 10.1186/s13195-017-0268-4] [Citation(s) in RCA: 86] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 03/29/2017] [Accepted: 05/22/2017] [Indexed: 02/08/2023]
Abstract
A major challenge to our understanding of the molecular mechanisms of Alzheimer’s disease (AD) has been the lack of physiologically relevant in vitro models which capture the precise patient genome, in the cell type of interest, with physiological expression levels of the gene(s) of interest. Induced pluripotent stem cell (iPSC) technology, together with advances in 2D and 3D neuronal differentiation, offers a unique opportunity to overcome this challenge and generate a limitless supply of human neurons for in vitro studies. iPSC-neuron models have been widely employed to model AD and we discuss in this review the progress that has been made to date using patient-derived neurons to recapitulate key aspects of AD pathology and how these models have contributed to a deeper understanding of AD molecular mechanisms, as well as addressing the key challenges posed by using this technology and what progress is being made to overcome these. Finally, we highlight future directions for the use of iPSC-neurons in AD research and highlight the potential value of this technology to neurodegenerative research in the coming years.
Collapse
Affiliation(s)
- Charles Arber
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London, WC1N 1PJ, UK
| | - Christopher Lovejoy
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London, WC1N 1PJ, UK
| | - Selina Wray
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London, WC1N 1PJ, UK.
| |
Collapse
|
|
30
|
Devine H, Patani R. The translational potential of human induced pluripotent stem cells for clinical neurology : The translational potential of hiPSCs in neurology. Cell Biol Toxicol 2016; 33:129-144. [PMID: 27915387 PMCID: PMC5325844 DOI: 10.1007/s10565-016-9372-7] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2016] [Accepted: 11/18/2016] [Indexed: 12/14/2022]
Abstract
The induced pluripotent state represents a decade-old Nobel prize-winning discovery. Human-induced pluripotent stem cells (hiPSCs) are generated by the nuclear reprogramming of any somatic cell using a variety of established but evolving methods. This approach offers medical science unparalleled experimental opportunity to model an individual patient’s disease “in a dish.” HiPSCs permit developmentally rationalized directed differentiation into any cell type, which express donor cell mutation(s) at pathophysiological levels and thus hold considerable potential for disease modeling, drug discovery, and potentially cell-based therapies. This review will focus on the translational potential of hiPSCs in clinical neurology and the importance of integrating this approach with complementary model systems to increase the translational yield of preclinical testing for the benefit of patients. This strategy is particularly important given the expected increase in prevalence of neurodegenerative disease, which poses a major burden to global health over the coming decades.
Collapse
Affiliation(s)
- Helen Devine
- Department of Molecular Neuroscience, UCL Institute of Neurology, Queen Square, London, WC1N3BG, UK.,Sobell Department of Motor Neuroscience and Movement Disorders, UCL Institute of Neurology, Queen Square, London, UK
| | - Rickie Patani
- Department of Molecular Neuroscience, UCL Institute of Neurology, Queen Square, London, WC1N3BG, UK. .,National Hospital for Neurology and Neurosurgery, UCL Institute of Neurology, Queen Square, London, WC1N 3BG, UK. .,Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK. .,Euan MacDonald Centre for MND, University of Edinburgh, Edinburgh, UK.
| |
Collapse
|
|
31
|
Csöbönyeiová M, Polák Š, Danišovič L. Recent approaches and challenges in iPSCs: modeling and cell-based therapy of Alzheimer's disease. Rev Neurosci 2016; 27:457-64. [PMID: 26812864 DOI: 10.1515/revneuro-2015-0054] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/03/2015] [Accepted: 12/13/2015] [Indexed: 12/21/2022]
Abstract
The lack of effective therapies for different neurodegenerative disorders has placed huge burdens on society. To overcome the restricted capacity of the central nervous system for regeneration, the promising alternative would be to use stem cells for more effective treatment of chronic degenerative and inflammatory neurological conditions and also of acute neuronal damage and from injuries or cerebrovascular diseases. The generation of induced pluripotent stem cells from somatic cells by the ectopic expression of specific transcription factors has provided the regenerative medicine field with a new tool for investigating and treating neurodegenerative diseases, including Alzheimer's disease (AD). This technology provides an alternative to traditional approaches, such as nuclear transfer and somatic cell fusion using embryonic stem cells. However, due to a problem in standardization of certain reprogramming techniques and systems research, the induced pluripotent stem cell-based technology is still in its infancy. The present paper is aimed at a brief review of the current status in modeling and cell-based therapies for AD.
Collapse
|
|
32
|
Abstract
Astrocytes abound in the human central nervous system (CNS) and play a multitude of indispensable roles in neuronal homeostasis and regulation of synaptic plasticity. While traditionally considered to be merely ancillary supportive cells, their complex yet fundamental relevance to brain physiology and pathology have only become apparent in recent times. Beyond their myriad canonical functions, previously unrecognised region-specific functional heterogeneity of astrocytes is emerging as an important attribute and challenges the traditional perspective of CNS-wide astrocyte homogeneity. Animal models have undeniably provided crucial insights into astrocyte biology, yet interspecies differences may limit the translational yield of such studies. Indeed, experimental systems aiming to understand the function of human astrocytes in health and disease have been hampered by accessibility to enriched cultures. Human induced pluripotent stem cells (hiPSCs) now offer an unparalleled model system to interrogate the role of astrocytes in neurodegenerative disorders. By virtue of their ability to convey mutations at pathophysiological levels in a human system, hiPSCs may serve as an ideal pre-clinical platform for both resolution of pathogenic mechanisms and drug discovery. Here, we review astrocyte specification from hiPSCs and discuss their role in modelling human neurological diseases.
Collapse
|
|
33
|
Balendra R, Patani R. Quo vadis motor neuron disease? World J Methodol 2016; 6:56-64. [PMID: 27019797 PMCID: PMC4804252 DOI: 10.5662/wjm.v6.i1.56] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/28/2015] [Revised: 11/17/2015] [Accepted: 01/11/2016] [Indexed: 02/06/2023] Open
Abstract
Motor neuron disease (MND), also known as amyotrophic lateral sclerosis, is a relentlessly progressive neurodegenerative condition that is invariably fatal, usually within 3 to 5 years of diagnosis. The aetio-pathogenesis of MND remains unresolved and no effective treatments exist. The only Food and Drug Administration approved disease modifying therapy is riluzole, a glutamate antagonist, which prolongs survival by up to 3 mo. Current management is largely symptomatic/supportive. There is therefore a desperate and unmet clinical need for discovery of disease mechanisms to guide novel therapeutic strategy. In this review, we start by introducing the organizational anatomy of the motor system, before providing a clinical overview of its dysfunction specifically in MND. We then summarize insights gained from pathological, genetic and animal models and conclude by speculating on optimal strategies to drive the step change in discovery, which is so desperately needed in this arena.
Collapse
|
|
34
|
Neural Conversion and Patterning of Human Pluripotent Stem Cells: A Developmental Perspective. Stem Cells Int 2016; 2016:8291260. [PMID: 27069483 PMCID: PMC4812494 DOI: 10.1155/2016/8291260] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2015] [Accepted: 01/24/2016] [Indexed: 01/19/2023] Open
Abstract
Since the reprogramming of adult human terminally differentiated somatic cells into induced pluripotent stem cells (hiPSCs) became a reality in 2007, only eight years have passed. Yet over this relatively short period, myriad experiments have revolutionized previous stem cell dogmata. The tremendous promise of hiPSC technology for regenerative medicine has fuelled rising expectations from both the public and scientific communities alike. In order to effectively harness hiPSCs to uncover fundamental mechanisms of disease, it is imperative to first understand the developmental neurobiology underpinning their lineage restriction choices in order to predictably manipulate cell fate to desired derivatives. Significant progress in developmental biology provides an invaluable resource for rationalising directed differentiation of hiPSCs to cellular derivatives of the nervous system. In this paper we begin by reviewing core developmental concepts underlying neural induction in order to provide context for how such insights have guided reductionist in vitro models of neural conversion from hiPSCs. We then discuss early factors relevant in neural patterning, again drawing upon crucial knowledge gained from developmental neurobiological studies. We conclude by discussing open questions relating to these concepts and how their resolution might serve to strengthen the promise of pluripotent stem cells in regenerative medicine.
Collapse
|
|
35
|
Generating Diverse Spinal Motor Neuron Subtypes from Human Pluripotent Stem Cells. Stem Cells Int 2015; 2016:1036974. [PMID: 26823667 PMCID: PMC4707335 DOI: 10.1155/2016/1036974] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/01/2015] [Accepted: 09/14/2015] [Indexed: 12/18/2022] Open
Abstract
Resolving the mechanisms underlying human neuronal diversification remains a major challenge in developmental and applied neurobiology. Motor neurons (MNs) represent a diverse pool of neuronal subtypes exhibiting differential vulnerability in different human neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS) and spinal muscular atrophy (SMA). The ability to predictably manipulate MN subtype lineage restriction from human pluripotent stem cells (PSCs) will form the essential basis to establishing accurate, clinically relevant in vitro disease models. I first overview motor neuron developmental biology to provide some context for reviewing recent studies interrogating pathways that influence the generation of MN diversity. I conclude that motor neurogenesis from PSCs provides a powerful reductionist model system to gain insight into the developmental logic of MN subtype diversification and serves more broadly as a leading exemplar of potential strategies to resolve the molecular basis of neuronal subclass differentiation within the nervous system. These studies will in turn permit greater mechanistic understanding of differential MN subtype vulnerability using in vitro human disease models.
Collapse
|
|
36
|
Wiethoff S, Arber C, Li A, Wray S, Houlden H, Patani R. Using human induced pluripotent stem cells to model cerebellar disease: hope and hype. J Neurogenet 2015; 29:95-102. [PMID: 25985846 PMCID: PMC4673530 DOI: 10.3109/01677063.2015.1053478] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/27/2015] [Accepted: 05/18/2015] [Indexed: 12/19/2022]
Abstract
The cerebellum forms a highly ordered and indispensible component of motor function within the adult neuraxis, consisting of several distinct cellular subtypes. Cerebellar disease, through a variety of genetic and acquired causes, results in the loss of function of defined subclasses of neurons, and remains a significant and untreatable health care burden. The scarcity of therapies in this arena can partially be explained by unresolved disease mechanisms due to inaccessibility of human cerebellar neurons in a relevant experimental context where initiating disease mechanisms could be functionally elucidated, or drug screens conducted. In this review we discuss the potential promise of human induced pluripotent stem cells (hiPSCs) for regenerative neurology, with a particular emphasis on in vitro modelling of cerebellar degeneration. We discuss progress made thus far using hiPSC-based models of neurodegeneration, noting the relatively slower pace of discovery made in modelling cerebellar dysfunction. We conclude by speculating how strategies attempting cerebellar differentiation from hiPSCs can be refined to allow the generation of accurate disease models. This in turn will permit a greater understanding of cerebellar pathophysiology to inform mechanistically rationalised therapies, which are desperately needed in this field.
Collapse
Affiliation(s)
- Sarah Wiethoff
- National Hospital for Neurology and Neurosurgery, UCL Institute of Neurology, London, UK
- Center for Neurology and Hertie Institute for Clinical Brain Research, Eberhard-Karls-University, Tübingen, Germany
| | - Charles Arber
- Department of Molecular Neuroscience and Queen Square Brain Bank, UCL Institute of Neurology, London, UK
| | - Abi Li
- Department of Molecular Neuroscience and Queen Square Brain Bank, UCL Institute of Neurology, London, UK
| | - Selina Wray
- Department of Molecular Neuroscience and Queen Square Brain Bank, UCL Institute of Neurology, London, UK
| | - Henry Houlden
- National Hospital for Neurology and Neurosurgery, UCL Institute of Neurology, London, UK
| | - Rickie Patani
- National Hospital for Neurology and Neurosurgery, UCL Institute of Neurology, London, UK
- Department of Molecular Neuroscience and Queen Square Brain Bank, UCL Institute of Neurology, London, UK
- Department of Clinical Neurosciences, University of Cambridge, Cambridge, UK
- Euan MacDonald Centre for MND, University of Edinburgh, Edinburgh, UK
| |
Collapse
|
|
37
|
Zhu L, Dong C, Sun C, Ma R, Yang D, Zhu H, Xu J. Rejuvenation of MPTP-induced human neural precursor cell senescence by activating autophagy. Biochem Biophys Res Commun 2015; 464:526-33. [PMID: 26159917 DOI: 10.1016/j.bbrc.2015.06.174] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/29/2015] [Accepted: 06/30/2015] [Indexed: 12/15/2022]
Abstract
Aging of neural stem cell, which can affect brain homeostasis, may be caused by many cellular mechanisms. Autophagy dysfunction was found in aged and neurodegenerative brains. However, little is known about the relationship between autophagy and human neural stem cell (hNSC) aging. The present study used 1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine (MPTP) to treat neural precursor cells (NPCs) derived from human embryonic stem cell (hESC) line H9 and investigate related molecular mechanisms involved in this process. MPTP-treated NPCs were found to undergo premature senescence [determined by increased senescence-associated-β-galactosidase (SA-β-gal) activity, elevated intracellular reactive oxygen species level, and decreased proliferation] and were associated with impaired autophagy. Additionally, the cellular senescence phenotypes were manifested at the molecular level by a significant increase in p21 and p53 expression, a decrease in SOD2 expression, and a decrease in expression of some key autophagy-related genes such as Atg5, Atg7, Atg12, and Beclin 1. Furthermore, we found that the senescence-like phenotype of MPTP-treated hNPCs was rejuvenated through treatment with a well-known autophagy enhancer rapamycin, which was blocked by suppression of essential autophagy gene Beclin 1. Taken together, these findings reveal the critical role of autophagy in the process of hNSC aging, and this process can be reversed by activating autophagy.
Collapse
Affiliation(s)
- Liang Zhu
- East Hospital, Tongji University School of Medicine, Shanghai, China
| | - Chuanming Dong
- East Hospital, Tongji University School of Medicine, Shanghai, China; Department of Anatomy and Neurobiology, The Jiangsu Key Laboratory of Neuroregeneration, Nantong University, Nantong, China
| | - Chenxi Sun
- East Hospital, Tongji University School of Medicine, Shanghai, China
| | - Rongjie Ma
- East Hospital, Tongji University School of Medicine, Shanghai, China
| | - Danjing Yang
- East Hospital, Tongji University School of Medicine, Shanghai, China
| | - Hongwen Zhu
- Tianjin Hospital, Tianjin Academy of Integrative Medicine, Tianjin, China.
| | - Jun Xu
- East Hospital, Tongji University School of Medicine, Shanghai, China.
| |
Collapse
|
|
38
|
Sposito T, Preza E, Mahoney CJ, Setó-Salvia N, Ryan NS, Morris HR, Arber C, Devine MJ, Houlden H, Warner TT, Bushell TJ, Zagnoni M, Kunath T, Livesey FJ, Fox NC, Rossor MN, Hardy J, Wray S. Developmental regulation of tau splicing is disrupted in stem cell-derived neurons from frontotemporal dementia patients with the 10 + 16 splice-site mutation in MAPT. Hum Mol Genet 2015; 24:5260-9. [PMID: 26136155 PMCID: PMC4550814 DOI: 10.1093/hmg/ddv246] [Citation(s) in RCA: 110] [Impact Index Per Article: 11.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/03/2015] [Accepted: 06/23/2015] [Indexed: 12/13/2022] Open
Abstract
The alternative splicing of the tau gene, MAPT, generates six protein isoforms in the adult human central nervous system (CNS). Tau splicing is developmentally regulated and dysregulated in disease. Mutations in MAPT that alter tau splicing cause frontotemporal dementia (FTD) with tau pathology, providing evidence for a causal link between altered tau splicing and disease. The use of induced pluripotent stem cell (iPSC)-derived neurons has revolutionized the way we model neurological disease in vitro. However, as most tau mutations are located within or around the alternatively spliced exon 10, it is important that iPSC–neurons splice tau appropriately in order to be used as disease models. To address this issue, we analyzed the expression and splicing of tau in iPSC-derived cortical neurons from control patients and FTD patients with the 10 + 16 intronic mutation in MAPT. We show that control neurons only express the fetal tau isoform (0N3R), even at extended time points of 100 days in vitro. Neurons from FTD patients with the 10 + 16 mutation in MAPT express both 0N3R and 0N4R tau isoforms, demonstrating that this mutation overrides the developmental regulation of exon 10 inclusion in our in vitro model. Further, at extended time points of 365 days in vitro, we observe a switch in tau splicing to include six tau isoforms as seen in the adult human CNS. Our results demonstrate the importance of neuronal maturity for use in in vitro modeling and provide a system that will be important for understanding the functional consequences of altered tau splicing.
Collapse
Affiliation(s)
- Teresa Sposito
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Elisavet Preza
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Colin J Mahoney
- Dementia Research Centre, Department of Neurodegenerative Disease, UCL Institute of Neurology, Queen Square, London WC1N 3BG, UK
| | - Núria Setó-Salvia
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Natalie S Ryan
- Dementia Research Centre, Department of Neurodegenerative Disease, UCL Institute of Neurology, Queen Square, London WC1N 3BG, UK
| | - Huw R Morris
- Department of Clinical Neuroscience, UCL Institute of Neurology, Queen Square, London WC1N 3BG, UK
| | - Charles Arber
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Michael J Devine
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK, Division of Brain Sciences, Imperial College London, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK
| | - Henry Houlden
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Thomas T Warner
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Trevor J Bushell
- Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, Glasgow G4 0RE, UK
| | - Michele Zagnoni
- Centre for Microsystems and Photonics, Electronic and Electrical Engineering, University of Strathclyde, Glasgow G1 1XW, UK
| | - Tilo Kunath
- MRC Centre for Regenerative Medicine, School of Biological Sciences, University of Edinburgh, 5 Little France Drive, Edinburgh EH16 4UU, UK and
| | - Frederick J Livesey
- Gurdon Institute, Cambridge Stem Cell Institute and Department of Biochemistry, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK
| | - Nick C Fox
- Dementia Research Centre, Department of Neurodegenerative Disease, UCL Institute of Neurology, Queen Square, London WC1N 3BG, UK
| | - Martin N Rossor
- Dementia Research Centre, Department of Neurodegenerative Disease, UCL Institute of Neurology, Queen Square, London WC1N 3BG, UK
| | - John Hardy
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK
| | - Selina Wray
- Department of Molecular Neuroscience, UCL Institute of Neurology, 1 Wakefield Street, London WC1N 1PJ, UK,
| |
Collapse
|
|
39
|
Smith I, Silveirinha V, Stein JL, de la Torre-Ubieta L, Farrimond JA, Williamson EM, Whalley BJ. Human neural stem cell-derived cultures in three-dimensional substrates form spontaneously functional neuronal networks. J Tissue Eng Regen Med 2015; 11:1022-1033. [PMID: 25712225 DOI: 10.1002/term.2001] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/29/2014] [Revised: 12/12/2014] [Accepted: 12/17/2014] [Indexed: 12/12/2022]
Abstract
Differentiated human neural stem cells were cultured in an inert three-dimensional (3D) scaffold and, unlike two-dimensional (2D) but otherwise comparable monolayer cultures, formed spontaneously active, functional neuronal networks that responded reproducibly and predictably to conventional pharmacological treatments to reveal functional, glutamatergic synapses. Immunocytochemical and electron microscopy analysis revealed a neuronal and glial population, where markers of neuronal maturity were observed in the former. Oligonucleotide microarray analysis revealed substantial differences in gene expression conferred by culturing in a 3D vs a 2D environment. Notable and numerous differences were seen in genes coding for neuronal function, the extracellular matrix and cytoskeleton. In addition to producing functional networks, differentiated human neural stem cells grown in inert scaffolds offer several significant advantages over conventional 2D monolayers. These advantages include cost savings and improved physiological relevance, which make them better suited for use in the pharmacological and toxicological assays required for development of stem cell-based treatments and the reduction of animal use in medical research. Copyright © 2015 John Wiley & Sons, Ltd.
Collapse
Affiliation(s)
- Imogen Smith
- Cellular and Molecular Neuroscience Group, Department of Pharmacy, University of Reading, UK
| | - Vasco Silveirinha
- Cellular and Molecular Neuroscience Group, Department of Pharmacy, University of Reading, UK
| | - Jason L Stein
- Neurogenetics Program, Department of Neurology, David Geffen School of Medicine, University of California, Los Angeles, CA, USA
| | - Luis de la Torre-Ubieta
- Neurogenetics Program, Department of Neurology, David Geffen School of Medicine, University of California, Los Angeles, CA, USA
| | | | - Elizabeth M Williamson
- Cellular and Molecular Neuroscience Group, Department of Pharmacy, University of Reading, UK
| | - Benjamin J Whalley
- Cellular and Molecular Neuroscience Group, Department of Pharmacy, University of Reading, UK
| |
Collapse
|
|
40
|
Li X, Teng S. RNA Sequencing in Schizophrenia. Bioinform Biol Insights 2015; 9:53-60. [PMID: 27053919 PMCID: PMC4818022 DOI: 10.4137/bbi.s28992] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/12/2015] [Revised: 02/01/2016] [Accepted: 02/06/2016] [Indexed: 12/11/2022] Open
Abstract
Schizophrenia (SCZ) is a serious psychiatric disorder that affects 1% of general population and places a heavy burden worldwide. The underlying genetic mechanism of SCZ remains unknown, but studies indicate that the disease is associated with a global gene expression disturbance across many genes. Next-generation sequencing, particularly of RNA sequencing (RNA-Seq), provides a powerful genome-scale technology to investigate the pathological processes of SCZ. RNA-Seq has been used to analyze the gene expressions and identify the novel splice isoforms and rare transcripts associated with SCZ. This paper provides an overview on the genetics of SCZ, the advantages of RNA-Seq for transcriptome analysis, the accomplishments of RNA-Seq in SCZ cohorts, and the applications of induced pluripotent stem cells and RNA-Seq in SCZ research.
Collapse
Affiliation(s)
- Xin Li
- Department of Biology, Howard University, Washington, DC, USA
| | - Shaolei Teng
- Department of Biology, Howard University, Washington, DC, USA
| |
Collapse
|
|
41
|
Physiological characterisation of human iPS-derived dopaminergic neurons. PLoS One 2014; 9:e87388. [PMID: 24586273 PMCID: PMC3931621 DOI: 10.1371/journal.pone.0087388] [Citation(s) in RCA: 122] [Impact Index Per Article: 11.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2013] [Accepted: 12/24/2013] [Indexed: 11/19/2022] Open
Abstract
Human induced pluripotent stem cells (hiPSCs) offer the potential to study otherwise inaccessible cell types. Critical to this is the directed differentiation of hiPSCs into functional cell lineages. This is of particular relevance to research into neurological disease, such as Parkinson's disease (PD), in which midbrain dopaminergic neurons degenerate during disease progression but are unobtainable until post-mortem. Here we report a detailed study into the physiological maturation over time of human dopaminergic neurons in vitro. We first generated and differentiated hiPSC lines into midbrain dopaminergic neurons and performed a comprehensive characterisation to confirm dopaminergic functionality by demonstrating dopamine synthesis, release, and re-uptake. The neuronal cultures include cells positive for both tyrosine hydroxylase (TH) and G protein-activated inward rectifier potassium channel 2 (Kir3.2, henceforth referred to as GIRK2), representative of the A9 population of substantia nigra pars compacta (SNc) neurons vulnerable in PD. We observed for the first time the maturation of the slow autonomous pace-making (<10 Hz) and spontaneous synaptic activity typical of mature SNc dopaminergic neurons using a combination of calcium imaging and electrophysiology. hiPSC-derived neurons exhibited inositol tri-phosphate (IP3) receptor-dependent release of intracellular calcium from the endoplasmic reticulum in neuronal processes as calcium waves propagating from apical and distal dendrites, and in the soma. Finally, neurons were susceptible to the dopamine neuron-specific toxin 1-methyl-4-phenylpyridinium (MPP+) which reduced mitochondrial membrane potential and altered mitochondrial morphology. Mature hiPSC-derived dopaminergic neurons provide a neurophysiologically-defined model of previously inaccessible vulnerable SNc dopaminergic neurons to bridge the gap between clinical PD and animal models.
Collapse
|
|
42
|
The more, the better: modeling dravet syndrome with induced pluripotent stem cell-derived neurons. Epilepsy Curr 2014; 14:33-4. [PMID: 24526875 DOI: 10.5698/1535-7597-14.1.33] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2023] Open
|
|
43
|
Imaizumi Y, Okano H. Modeling human neurological disorders with induced pluripotent stem cells. J Neurochem 2013; 129:388-99. [PMID: 24286589 DOI: 10.1111/jnc.12625] [Citation(s) in RCA: 82] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2013] [Revised: 11/18/2013] [Accepted: 11/22/2013] [Indexed: 02/06/2023]
Abstract
Human induced pluripotent stem (iPS) cells obtained by reprogramming technology are a source of great hope, not only in terms of applications in regenerative medicine, such as cell transplantation therapy, but also for modeling human diseases and new drug development. In particular, the production of iPS cells from the somatic cells of patients with intractable diseases and their subsequent differentiation into cells at affected sites (e.g., neurons, cardiomyocytes, hepatocytes, and myocytes) has permitted the in vitro construction of disease models that contain patient-specific genetic information. For example, disease-specific iPS cells have been established from patients with neuropsychiatric disorders, including schizophrenia and autism, as well as from those with neurodegenerative diseases, including Parkinson's disease and Alzheimer's disease. A multi-omics analysis of neural cells originating from patient-derived iPS cells may thus enable investigators to elucidate the pathogenic mechanisms of neurological diseases that have heretofore been unknown. In addition, large-scale screening of chemical libraries with disease-specific iPS cells is currently underway and is expected to lead to new drug discovery. Accordingly, this review outlines the progress made via the use of patient-derived iPS cells toward the modeling of neurological disorders, the testing of existing drugs, and the discovery of new drugs. The production of human induced pluripotent stem (iPS) cells from the patients' somatic cells and their subsequent differentiation into specific cells have permitted the in vitro construction of disease models that contain patient-specific genetic information. Furthermore, innovations of gene-editing technologies on iPS cells are enabling new approaches for illuminating the pathogenic mechanisms of human diseases. In this review article, we outlined the current status of neurological diseases-specific iPS cell research and described recently obtained knowledge in the form of actual examples.
Collapse
Affiliation(s)
- Yoichi Imaizumi
- Department of Physiology, Keio University School of Medicine, Tokyo, Japan; Next Generation Systems CFU, Eisai Co. Ltd., Ibaraki, Japan
| | | |
Collapse
|
|
44
|
Pluripotent stem cell for modeling neurological diseases. Exp Cell Res 2013; 319:177-84. [DOI: 10.1016/j.yexcr.2012.11.007] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2012] [Accepted: 11/01/2012] [Indexed: 12/31/2022]
|
|
45
|
Abstract
Multiple Sclerosis (MS) is an inflammatory demyelinating neurodegenerative disorder of the brain and spinal cord that causes significant disability in young adults. Although the precise aetiopathogenesis of MS remains unresolved, its pathological hallmarks include inflammation, demyelination, axonal injury (acute and chronic), astrogliosis and variable remyelination. Despite major recent advances in therapeutics for the early stage of the disease there are currently no disease modifying treatments for the progressive stage of disease, whose pathological substrate is axonal degeneration. This represents the great and unmet clinical need in MS. Against this background, human stem cells offer promise both to improve understanding of disease mechanism(s) through in-vitro modeling as well as potentially direct use to supplement and promote remyelination, an endogenous reparative process where entire myelin sheaths are restored to demyelinated axons. Conceptually, stem cells can act directly to myelinate axons or indirectly through different mechanisms to promote endogenous repair; importantly these two mechanisms of action are not mutually exclusive. We propose that discovery of novel methods to invoke or enhance remyelination in MS may be the most effective therapeutic strategy to limit axonal damage and instigate restoration of structure and function in this debilitating condition. Human stem cell derived neurons and glia, including patient specific cells derived through reprogramming, provide an unprecedented experimental system to model MS “in a dish” as well as enable high-throughput drug discovery. Finally, we speculate upon the potential role for stem cell based therapies in MS.
Collapse
|