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Cantó-Santos J, Valls-Roca L, Tobías E, García-García FJ, Guitart-Mampel M, Andújar-Sánchez F, Vilaseca-Capel A, Esteve-Codina A, Martín-Mur B, Padrosa J, Peruga E, Madrigal I, Segalés P, García-Ruiz C, Fernández-Checa JC, Moreno-Lozano PJ, O'Callaghan AS, Sevilla A, Milisenda JC, Garrabou G. Human induced pluripotent stem cell-derived myotubes to model inclusion body myositis. Acta Neuropathol Commun 2025; 13:38. [PMID: 39985015 PMCID: PMC11844183 DOI: 10.1186/s40478-025-01933-0] [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: 11/25/2024] [Accepted: 01/23/2025] [Indexed: 02/23/2025] Open
Abstract
Inclusion body myositis (IBM) is an inflammatory myopathy that displays proximal and distal muscle weakness. At the histopathological level, the muscles of IBM patients show inflammatory infiltrates, rimmed vacuoles and mitochondrial changes. The etiology of IBM remains unknown, and there is a lack of validated disease models, biomarkers and effective treatments. To contribute to unveil disease underpins we developed a cell model based on myotubes derived from induced pluripotent stem cells (iPSC-myotubes) from IBM patients and compared the molecular phenotype vs. age and sex-paired controls (n = 3 IBM and 4 CTL). We evaluated protein histological findings and the gene expression profile by mRNA-seq, alongside functional analysis of inflammation, degeneration and mitochondrial function. Briefly, IBM iPSC-myotubes replicated relevant muscle histopathology features of IBM, including aberrant expression of HLA, TDP-43 and COX markers. mRNA seq analysis identified 1007 differentially expressed genes (DEGs) (p-value adj < 0.01; 789 upregulated and 218 downregulated), associated with myopathy, muscle structure and developmental changes. Among these, 1 DEG was related to inflammation, 28 to autophagy and 28 to mitochondria. At the functional level, inflammation was similar between the IBM and CTL groups under basal conditions (mean cytokine expression in IBM 4.6 ± 1.4 vs. 6.7 ± 3.4 in CTL), but increased in IBM iPSC-myotubes after lipopolysaccharide treatment (72.5 ± 21.8 in IBM vs. 13.0 ± 6.7 in CTL). Additionally, autophagy was disturbed, with 40.14% reduction in autophagy mediators. Mitochondrial dysfunction was strongly manifested, showing a conserved respiratory profile and antioxidant capacity, but a 56.33% lower cytochrome c oxidase/citrate synthase ratio and a 66.59% increase in lactate secretion. Overall, these findings support patient-derived iPSC-myotubes as a relevant model for IBM, reflecting the main muscle hallmarks, including inflammation, autophagy dysfunction and mitochondrial alterations at transcriptomic, protein and functional levels.
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Affiliation(s)
- Judith Cantó-Santos
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Laura Valls-Roca
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Ester Tobías
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Francesc Josep García-García
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Mariona Guitart-Mampel
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Félix Andújar-Sánchez
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Adrià Vilaseca-Capel
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Anna Esteve-Codina
- Centro Nacional de Análisis Genómico, CNAG, Barcelona Institute of Science and Technology, Barcelona, Spain
- Universitat Pompeu Fabra (UPF), Barcelona, Spain
| | - Beatriz Martín-Mur
- Centro Nacional de Análisis Genómico, CNAG, Barcelona Institute of Science and Technology, Barcelona, Spain
| | - Joan Padrosa
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Emma Peruga
- Biochemistry and Molecular Genetics Department, Hospital Clinic of Barcelona and Institut d'Investigacions Biomèdiques August Pi I Sunyer (IDIBAPS), Barcelona, 08036, Spain
| | - Irene Madrigal
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
- Biochemistry and Molecular Genetics Department, Hospital Clinic of Barcelona and Institut d'Investigacions Biomèdiques August Pi I Sunyer (IDIBAPS), Barcelona, 08036, Spain
| | - Paula Segalés
- Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB-CSIC), Liver Unit- HCB-IDIBAPS, Barcelona, Spain
- CIBEREHD-Spanish Biomedical Research Centre in Hepatic and Digestive Diseases, Madrid, Spain
| | - Carmen García-Ruiz
- Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB-CSIC), Liver Unit- HCB-IDIBAPS, Barcelona, Spain
- CIBEREHD-Spanish Biomedical Research Centre in Hepatic and Digestive Diseases, Madrid, Spain
| | - José Carlos Fernández-Checa
- Department of Cell Death and Proliferation, Institute of Biomedical Research of Barcelona (IIBB-CSIC), Liver Unit- HCB-IDIBAPS, Barcelona, Spain
- CIBEREHD-Spanish Biomedical Research Centre in Hepatic and Digestive Diseases, Madrid, Spain
| | - Pedro J Moreno-Lozano
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain
| | - Albert Selva O'Callaghan
- Vall d'Hebrón Systemic Autoimmune Diseases Unit. Internal Medicine Service, Hospital Universitari Vall d'Hebrón (HVH), Universitat Autònoma de Barcelona (UAB), Barcelona, Spain
| | - Ana Sevilla
- Departament de Biologia Cel·lular, Fisiologia i Immunologia, Facultat de Biologia, Institute of Neuroscience, Universitat de Barcelona, Barcelona, Spain.
- Institute of Biomedicine (IBUB), University of Barcelona, Barcelona, Spain.
| | - José César Milisenda
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain.
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain.
| | - Glòria Garrabou
- Inherited Metabolic Diseases and Muscular Disorders Research Group, Department of Internal Medicine, Faculty of Medicine and Health Sciences, Institut d'Investigacions Biomèdiques August Pi i Sunyer (IDIBAPS), Hospital Clinic of Barcelona, University of Barcelona, Barcelona, Spain.
- CIBERER- Spanish Biomedical Research Centre in Rare Diseases - ISCIII, Madrid, Spain.
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Javed S, Fersini M, Bernardini G. Unleashing the Power of Induced Pluripotent stem Cells in in vitro Modelling of Lesch-Nyhan Disease. Stem Cell Rev Rep 2025; 21:304-318. [PMID: 39495466 DOI: 10.1007/s12015-024-10821-4] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 10/29/2024] [Indexed: 11/05/2024]
Abstract
Lesch-Nyhan disease (LND) is a monogenic rare neurodevelopmental disorder caused by a deficiency in hypoxanthine-guanine phosphoribosyltransferase (HPRT), the key enzyme of the purines salvage pathway. Beyond its well-documented metabolic consequences, HPRT deficiency leads to a distinctive neurobehavioral syndrome characterized by motor disabilities, cognitive deficits, and self-injurious behavior. Although various cell and animal models have been developed to investigate LND pathology, none have adequately elucidated the underlying mechanisms of its neurological alterations. Recent advances in human pluripotent stem cell research and in vitro differentiation techniques have ushered in a new era in rare neurodevelopmental disorders research. Pluripotent stem cells, with their ability to propagate indefinitely and to differentiate into virtually any cell type, offer a valuable alternative for modeling rare diseases, allowing for the detection of pathological events from the earliest stages of neuronal network development. Furthermore, the generation of patient-derived induced pluripotent stem cells using reprogramming technology provides an opportunity to develop a disease-relevant model within the context of a patient-specific genome. In this review, we examine current stem cell-based models of LND and assess their potential as optimal models for exploring key pathological molecular events during neurogenesis and for the discovering novel treatment options. We also address the limitations, challenges, and future prospects for improving the use of iPSCs in LND research.
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Affiliation(s)
- Sundas Javed
- Dipartimento di Biotecnologie, Chimica e Farmacia, Università degli Studi di Siena, via Aldo Moro 2, Siena, 53100, Italy
| | - Marco Fersini
- Dipartimento di Biotecnologie, Chimica e Farmacia, Università degli Studi di Siena, via Aldo Moro 2, Siena, 53100, Italy
| | - Giulia Bernardini
- Dipartimento di Biotecnologie, Chimica e Farmacia, Università degli Studi di Siena, via Aldo Moro 2, Siena, 53100, Italy.
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Yahyazadeh R, Baradaran Rahimi V, Askari VR. Stem cell and exosome therapies for regenerating damaged myocardium in heart failure. Life Sci 2024; 351:122858. [PMID: 38909681 DOI: 10.1016/j.lfs.2024.122858] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2024] [Revised: 06/13/2024] [Accepted: 06/18/2024] [Indexed: 06/25/2024]
Abstract
Finding novel treatments for cardiovascular diseases (CVDs) is a hot topic in medicine; cell-based therapies have reported promising news for controlling dangerous complications of heart disease such as myocardial infarction (MI) and heart failure (HF). Various progenitor/stem cells were tested in various in-vivo, in-vitro, and clinical studies for regeneration or repairing the injured tissue in the myocardial to accelerate the healing. Fetal, adult, embryonic, and induced pluripotent stem cells (iPSC) have revealed the proper potency for cardiac tissue repair. As an essential communicator among cells, exosomes with specific contacts (proteins, lncRNAs, and miRNAs) greatly promote cardiac rehabilitation. Interestingly, stem cell-derived exosomes have more efficiency than stem cell transplantation. Therefore, stem cells induced pluripotent stem cells (iPSCs), embryonic stem cells (ESCs), cardiac stem cells (CDC), and skeletal myoblasts) and their-derived exosomes will probably be considered an alternative therapy for CVDs remedy. In addition, stem cell-derived exosomes have been used in the diagnosis/prognosis of heart diseases. In this review, we explained the advances of stem cells/exosome-based treatment, their beneficial effects, and underlying mechanisms, which will present new insights in the clinical field in the future.
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Affiliation(s)
- Roghayeh Yahyazadeh
- Pharmacological Research Center of Medicinal Plants, Mashhad University of Medical Sciences, Mashhad, Iran
| | - Vafa Baradaran Rahimi
- Department of Cardiovascular Diseases, Faculty of Medicine, Mashhad University of Medical Sciences, Mashhad, Iran.
| | - Vahid Reza Askari
- Pharmacological Research Center of Medicinal Plants, Mashhad University of Medical Sciences, Mashhad, Iran.
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HPRT1 Deficiency Induces Alteration of Mitochondrial Energy Metabolism in the Brain. Mol Neurobiol 2023; 60:3147-3157. [PMID: 36802322 PMCID: PMC10122629 DOI: 10.1007/s12035-023-03266-2] [Citation(s) in RCA: 7] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2022] [Accepted: 02/12/2023] [Indexed: 02/23/2023]
Abstract
Alterations in function of hypoxanthine guanine phosphoribosyl transferase (HPRT), one of the major enzymes involved in purine nucleotide exchange, lead to overproduction of uric acid and produce various symptoms of Lesch-Nyhan syndrome (LNS). One of the hallmarks of LNS is maximal expression of HPRT in the central nervous system with the highest activity of this enzyme in the midbrain and basal ganglia. However, the nature of neurological symptoms has yet to be clarified in details. Here, we studied whether HPRT1 deficiency changes mitochondrial energy metabolism and redox balance in murine neurons from the cortex and midbrain. We found that HPRT1 deficiency inhibits complex I-dependent mitochondrial respiration resulting in increased levels of mitochondrial NADH, reduction of the mitochondrial membrane potential, and increased rate of reactive oxygen species (ROS) production in mitochondria and cytosol. However, increased ROS production did not induce oxidative stress and did not decrease the level of endogenous antioxidant glutathione (GSH). Thus, disruption of mitochondrial energy metabolism but not oxidative stress could play a role of potential trigger of brain pathology in LNS.
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Yin S, Wang J, Zhu Y, Song L, Wu T, Zhang Z, Zhang X, Li F, Chen G. A novel uric acid biosensor based on regular Prussian blue nanocrystal/ upright graphene oxide array nanocomposites. CURR ANAL CHEM 2022. [DOI: 10.2174/1573411018666220117155419] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Objective:
Regular Prussian blue nanocrystals (RPB) were grown upright on graphene oxide flakes (GO), which were on the surface of a custom screen-printed carbon electrode (SPCE), using a spray method assisted by a constant magnetic field (CMF).
Method:
After immobilizing uricase, the uric acid biosensor Uricase/RPB/CMF-GO/SPCE was obtained. The detection range of the sensor response to UA was 0.005~2.525 mM, and the detection limit was as low as 3.6 μM. The cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS) results showed that compared to amorphous electrodeposited Prussian blue (EDPB), RPB more favorably accelerated electron transport.
Result:
This novel uric acid biosensor exhibits high sensitivity over, a wide concentration range, strong anti- interference ability, and good stability and reproducibility.
Conclusion:
Thus, it has good application prospects for determining uric acid in physiological samples
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Affiliation(s)
- Shiyu Yin
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Jikui Wang
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Yongbao Zhu
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Lingyu Song
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Tingxia Wu
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Zhiyi Zhang
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Xianbo Zhang
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Fan Li
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
| | - Guosong Chen
- School of Chemistry and Molecular Engineering, Nanjing Tech University, Nanjing, China; bCollege of chemical engineering, Nanjing Tech University, Nanjing, China
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Azar J, Bahmad HF, Daher D, Moubarak MM, Hadadeh O, Monzer A, Al Bitar S, Jamal M, Al-Sayegh M, Abou-Kheir W. The Use of Stem Cell-Derived Organoids in Disease Modeling: An Update. Int J Mol Sci 2021; 22:7667. [PMID: 34299287 PMCID: PMC8303386 DOI: 10.3390/ijms22147667] [Citation(s) in RCA: 38] [Impact Index Per Article: 9.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/29/2021] [Revised: 06/17/2021] [Accepted: 06/18/2021] [Indexed: 02/06/2023] Open
Abstract
Organoids represent one of the most important advancements in the field of stem cells during the past decade. They are three-dimensional in vitro culturing models that originate from self-organizing stem cells and can mimic the in vivo structural and functional specificities of body organs. Organoids have been established from multiple adult tissues as well as pluripotent stem cells and have recently become a powerful tool for studying development and diseases in vitro, drug screening, and host-microbe interaction. The use of stem cells-that have self-renewal capacity to proliferate and differentiate into specialized cell types-for organoids culturing represents a major advancement in biomedical research. Indeed, this new technology has a great potential to be used in a multitude of fields, including cancer research, hereditary and infectious diseases. Nevertheless, organoid culturing is still rife with many challenges, not limited to being costly and time consuming, having variable rates of efficiency in generation and maintenance, genetic stability, and clinical applications. In this review, we aim to provide a synopsis of pluripotent stem cell-derived organoids and their use for disease modeling and other clinical applications.
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Affiliation(s)
- Joseph Azar
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Hisham F. Bahmad
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
- Arkadi M. Rywlin M.D. Department of Pathology and Laboratory Medicine, Mount Sinai Medical Center, Miami Beach, FL 33140, USA
| | - Darine Daher
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Maya M. Moubarak
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Ola Hadadeh
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Alissar Monzer
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Samar Al Bitar
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
| | - Mohamed Jamal
- Hamdan Bin Mohammed College of Dental Medicine, Mohammed Bin Rashid University of Medicine and Health Sciences, Dubai 66566, United Arab Emirates
| | - Mohamed Al-Sayegh
- Biology Division, New York University Abu Dhabi, Abu Dhabi 2460, United Arab Emirates
| | - Wassim Abou-Kheir
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of Beirut, Beirut 1107 2260, Lebanon; (J.A.); (H.F.B.); (D.D.); (M.M.M.); (O.H.); (A.M.); (S.A.B.)
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Duchesne de Lamotte J, Perrier A, Martinat C, Nicoleau C. Emerging Opportunities in Human Pluripotent Stem-Cells Based Assays to Explore the Diversity of Botulinum Neurotoxins as Future Therapeutics. Int J Mol Sci 2021; 22:7524. [PMID: 34299143 PMCID: PMC8308099 DOI: 10.3390/ijms22147524] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2021] [Revised: 07/02/2021] [Accepted: 07/07/2021] [Indexed: 02/07/2023] Open
Abstract
Botulinum neurotoxins (BoNTs) are produced by Clostridium botulinum and are responsible for botulism, a fatal disorder of the nervous system mostly induced by food poisoning. Despite being one of the most potent families of poisonous substances, BoNTs are used for both aesthetic and therapeutic indications from cosmetic reduction of wrinkles to treatment of movement disorders. The increasing understanding of the biology of BoNTs and the availability of distinct toxin serotypes and subtypes offer the prospect of expanding the range of indications for these toxins. Engineering of BoNTs is considered to provide a new avenue for improving safety and clinical benefit from these neurotoxins. Robust, high-throughput, and cost-effective assays for BoNTs activity, yet highly relevant to the human physiology, have become indispensable for a successful translation of engineered BoNTs to the clinic. This review presents an emerging family of cell-based assays that take advantage of newly developed human pluripotent stem cells and neuronal function analyses technologies.
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Affiliation(s)
- Juliette Duchesne de Lamotte
- IPSEN Innovation, 91940 Les Ulis, France;
- I-STEM, INSERM UMR861, Université Evry-Paris Saclay, 91100 Corbeil-Essonne, France
| | - Anselme Perrier
- I-STEM, INSERM UMR861, Université Evry-Paris Saclay, 91100 Corbeil-Essonne, France
- Laboratoire des Maladies Neurodégénératives: Mécanismes, Thérapies, Imagerie, CEA/CNRS UMR9199, Université Paris Saclay, 92265 Fontenay-aux-Roses, France
| | - Cécile Martinat
- I-STEM, INSERM UMR861, Université Evry-Paris Saclay, 91100 Corbeil-Essonne, France
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Wang Y, Lei W, Yang J, Ni X, Ye L, Shen Z, Hu S. The updated view on induced pluripotent stem cells for cardiovascular precision medicine. Pflugers Arch 2021; 473:1137-1149. [DOI: 10.1007/s00424-021-02530-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2020] [Revised: 01/06/2021] [Accepted: 01/29/2021] [Indexed: 12/14/2022]
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Wang L, Ye Z, Jang YY. Convergence of human pluripotent stem cell, organoid, and genome editing technologies. Exp Biol Med (Maywood) 2021; 246:861-875. [PMID: 33467883 DOI: 10.1177/1535370220985808] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022] Open
Abstract
The last decade has seen many exciting technological breakthroughs that greatly expanded the toolboxes for biological and biomedical research, yet few have had more impact than induced pluripotent stem cells and modern-day genome editing. These technologies are providing unprecedented opportunities to improve physiological relevance of experimental models, further our understanding of developmental processes, and develop novel therapies. One of the research areas that benefit greatly from these technological advances is the three-dimensional human organoid culture systems that resemble human tissues morphologically and physiologically. Here we summarize the development of human pluripotent stem cells and their differentiation through organoid formation. We further discuss how genetic modifications, genome editing in particular, were applied to answer basic biological and biomedical questions using organoid cultures of both somatic and pluripotent stem cell origins. Finally, we discuss the potential challenges of applying human pluripotent stem cell and organoid technologies for safety and efficiency evaluation of emerging genome editing tools.
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Affiliation(s)
- Lin Wang
- Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, US Food and Drug Administration, Silver Spring, MD 20993, USA
| | - Zhaohui Ye
- Division of Cellular and Gene Therapies, Center for Biologics Evaluation and Research, US Food and Drug Administration, Silver Spring, MD 20993, USA
| | - Yoon-Young Jang
- Department of Oncology, Sidney Kimmel Comprehensive Cancer Center, John Hopkins University, Baltimore, MD 21218, USA
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Induced Pluripotent Stem Cells to Understand Mucopolysaccharidosis. I: Demonstration of a Migration Defect in Neural Precursors. Cells 2020; 9:cells9122593. [PMID: 33287330 PMCID: PMC7761689 DOI: 10.3390/cells9122593] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/13/2020] [Revised: 11/27/2020] [Accepted: 11/30/2020] [Indexed: 02/06/2023] Open
Abstract
Background: Mucopolysaccharidosis type I-Hurler (MPS1-H) is a severe genetic lysosomal storage disorder due to loss-of-function mutations in the IDUA gene. The subsequent complete deficiency of alpha l-iduronidase enzyme is directly responsible of a progressive accumulation of glycosaminoglycans (GAG) in lysosomes which affects the functions of many tissues. Consequently, MPS1 is characterized by systemic symptoms (multiorgan dysfunction) including respiratory and cardiac dysfunctions, skeletal abnormalities and early fatal neurodegeneration. Methods: To understand mechanisms underlying MPS1 neuropathology, we generated induced pluripotent stem cells (iPSC) from a MPS1-H patient with loss-of-function mutations in both IDUA alleles. To avoid variability due to different genetic background of iPSC, we established an isogenic control iPSC line by rescuing IDUA expression by a lentivectoral approach. Results: Marked differences between MPS1-H and IDUA-corrected isogenic controls were observed upon neural differentiation. A scratch assay revealed a strong migration defect of MPS1-H cells. Also, there was a massive impact of IDUA deficiency on gene expression (340 genes with an FDR <0.05). Conclusions: Our results demonstrate a hitherto unknown connection between lysosomal degradation, gene expression and neural motility, which might account at least in part for the phenotype of MPS1-H patients.
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Salsi V, Magdinier F, Tupler R. Does DNA Methylation Matter in FSHD? Genes (Basel) 2020; 11:E258. [PMID: 32121044 PMCID: PMC7140823 DOI: 10.3390/genes11030258] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/23/2020] [Revised: 02/17/2020] [Accepted: 02/25/2020] [Indexed: 12/13/2022] Open
Abstract
Facioscapulohumeral muscular dystrophy (FSHD) has been associated with the genetic and epigenetic molecular features of the CpG-rich D4Z4 repeat tandem array at 4q35. Reduced DNA methylation of D4Z4 repeats is considered part of the FSHD mechanism and has been proposed as a reliable marker in the FSHD diagnostic procedure. We considered the assessment of D4Z4 DNA methylation status conducted on distinct cohorts using different methodologies. On the basis of the reported results we conclude that the percentage of DNA methylation detected at D4Z4 does not correlate with the disease status. Overall, data suggest that in the case of FSHD1, D4Z4 hypomethylation is a consequence of the chromatin structure present in the contracted allele, rather than a proxy of its function. Besides, CpG methylation at D4Z4 DNA is reduced in patients presenting diseases unrelated to muscle progressive wasting, like Bosma Arhinia and Microphthalmia syndrome, a developmental disorder, as well as ICF syndrome. Consistent with these observations, the analysis of epigenetic reprogramming at the D4Z4 locus in human embryonic and induced pluripotent stem cells indicate that other mechanisms, independent from the repeat number, are involved in the control of the epigenetic structure at D4Z4.
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Affiliation(s)
- Valentina Salsi
- Department of Life Sciences, University of Modena and Reggio Emilia, 4, 41121 Modena, Italy;
| | | | - Rossella Tupler
- Department of Biomedical, Metabolic and Neural Sciences, University of Modena and Reggio Emilia, 4, 41121 Modena, Italy
- Center for Neuroscience and Neurotechnology, University of Modena and Reggio Emilia, 4, 41121 Modena, Italy
- Department of Molecular Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA 01003, USA
- Li Weibo Institute for Rare Diseases Research at the University of Massachusetts Medical School, Worcester, MA 01003, USA
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12
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Seranova E, Palhegyi AM, Verma S, Dimova S, Lasry R, Naama M, Sun C, Barrett T, Rosenstock TR, Kumar D, Cohen MA, Buganim Y, Sarkar S. Human Induced Pluripotent Stem Cell Models of Neurodegenerative Disorders for Studying the Biomedical Implications of Autophagy. J Mol Biol 2020; 432:2754-2798. [PMID: 32044344 DOI: 10.1016/j.jmb.2020.01.024] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/20/2019] [Revised: 01/22/2020] [Accepted: 01/23/2020] [Indexed: 12/12/2022]
Abstract
Autophagy is an intracellular degradation process that is essential for cellular survival, tissue homeostasis, and human health. The housekeeping functions of autophagy in mediating the clearance of aggregation-prone proteins and damaged organelles are vital for post-mitotic neurons. Improper functioning of this process contributes to the pathology of myriad human diseases, including neurodegeneration. Impairment in autophagy has been reported in several neurodegenerative diseases where pharmacological induction of autophagy has therapeutic benefits in cellular and transgenic animal models. However, emerging studies suggest that the efficacy of autophagy inducers, as well as the nature of the autophagy defects, may be context-dependent, and therefore, studies in disease-relevant experimental systems may provide more insights for clinical translation to patients. With the advancements in human stem cell technology, it is now possible to establish disease-affected cellular platforms from patients for investigating disease mechanisms and identifying candidate drugs in the appropriate cell types, such as neurons that are otherwise not accessible. Towards this, patient-derived human induced pluripotent stem cells (hiPSCs) have demonstrated considerable promise in constituting a platform for effective disease modeling and drug discovery. Multiple studies have utilized hiPSC models of neurodegenerative diseases to study autophagy and evaluate the therapeutic efficacy of autophagy inducers in neuronal cells. This review provides an overview of the regulation of autophagy, generation of hiPSCs via cellular reprogramming, and neuronal differentiation. It outlines the findings in various neurodegenerative disorders where autophagy has been studied using hiPSC models.
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Affiliation(s)
- Elena Seranova
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom
| | - Adina Maria Palhegyi
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom
| | - Surbhi Verma
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom; Cellular Immunology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, 110067, India
| | - Simona Dimova
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom
| | - Rachel Lasry
- Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, The Hebrew University Hadassah Medical School, Jerusalem, 91120, Israel
| | - Moriyah Naama
- Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, The Hebrew University Hadassah Medical School, Jerusalem, 91120, Israel
| | - Congxin Sun
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom
| | - Timothy Barrett
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom
| | - Tatiana Rosado Rosenstock
- Department of Physiological Science, Santa Casa de São Paulo School of Medical Sciences, São Paulo, SP, 01221-020, Brazil
| | - Dhiraj Kumar
- Cellular Immunology Group, International Centre for Genetic Engineering and Biotechnology, Aruna Asaf Ali Marg, New Delhi, 110067, India
| | - Malkiel A Cohen
- Whitehead Institute for Biomedical Research, Cambridge, MA, 02142, USA
| | - Yosef Buganim
- Department of Developmental Biology and Cancer Research, Institute for Medical Research Israel-Canada, The Hebrew University Hadassah Medical School, Jerusalem, 91120, Israel
| | - Sovan Sarkar
- Institute of Cancer and Genomic Sciences, Institute of Biomedical Research, College of Medical and Dental Sciences, University of Birmingham, Edgbaston, Birmingham B15 2TT, United Kingdom.
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13
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Marotta N, Kim S, Krainc D. Organoid and pluripotent stem cells in Parkinson's disease modeling: an expert view on their value to drug discovery. Expert Opin Drug Discov 2020; 15:427-441. [PMID: 31899983 DOI: 10.1080/17460441.2020.1703671] [Citation(s) in RCA: 24] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
Introduction: Parkinson's disease is a devastating neurodegenerative disorder preferentially involving loss of dopaminergic neurons in the substantia nigra, leading to typical motor symptoms. While there are still no therapeutics to modify disease course, recent work using induced pluripotent stem cell (iPSC) and 3D brain organoid models have provided further insight into Parkinson's disease pathogenesis and potential therapeutic targets.Areas covered: This review highlights the generation of iPSC neurons and neural organoids as models for studying Parkinson's disease. It further discusses the recent work using patient-derived neurons from both familial and sporadic forms of Parkinson's to study disease pathogenic phenotypes and pathways. It additionally provides an evaluation of iPSC neurons and organoid models for therapeutic development in Parkinson's.Expert opinion: The use of Parkinson's disease patient-derived neurons and organoids provides us with the exciting opportunity to directly investigate pathogenic mechanisms and test drug compounds in human neurons. Future studies will involve generating more sophisticated models of brain organoids, studying neuronal pathways using larger patient cohorts, and routinely assessing therapeutics in these models.
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Affiliation(s)
- Nick Marotta
- Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA
| | - Soojin Kim
- Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA
| | - Dimitri Krainc
- Department of Neurology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA
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14
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Costamagna G, Andreoli L, Corti S, Faravelli I. iPSCs-Based Neural 3D Systems: A Multidimensional Approach for Disease Modeling and Drug Discovery. Cells 2019; 8:E1438. [PMID: 31739555 PMCID: PMC6912470 DOI: 10.3390/cells8111438] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/30/2019] [Revised: 10/26/2019] [Accepted: 11/11/2019] [Indexed: 02/06/2023] Open
Abstract
Induced pluripotent stem cells (iPSCs)-based two-dimensional (2D) protocols have offered invaluable insights into the pathophysiology of neurological diseases. However, these systems are unable to reproduce complex cytoarchitectural features, cell-cell and tissue-tissue interactions like their in vivo counterpart. Three-dimensional (3D)-based culture protocols, though in their infancy, have offered new insights into modeling human diseases. Human neural organoids try to recapitulate the cellular diversity of complex tissues and can be generated from iPSCs to model the pathophysiology of a wide spectrum of pathologies. The engraftment of iPSCs into mice models and the improvement of differentiation protocols towards 3D cultures has enabled the generation of more complex multicellular systems. Consequently, models of neuropsychiatric disorders, infectious diseases, brain cancer and cerebral hypoxic injury can now be investigated from new perspectives. In this review, we consider the advancements made in modeling neuropsychiatric and neurological diseases with iPSC-derived organoids and their potential use to develop new drugs.
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Affiliation(s)
| | | | | | - Irene Faravelli
- Dino Ferrari Centre, Neuroscience Section, Department of Pathophysiology and Transplantation (DEPT), University of Milan, Neurology Unit, IRCCS Foundation Ca’ Granda Ospedale Maggiore Policlinico, 20122 Milan, Italy; (G.C.); (L.A.); (S.C.)
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15
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Abstract
The derivation of induced pluripotent stem cells (iPSCs) over a decade ago sparked widespread enthusiasm for the development of new models of human disease, enhanced platforms for drug discovery and more widespread use of autologous cell-based therapy. Early studies using directed differentiation of iPSCs frequently uncovered cell-level phenotypes in monogenic diseases, but translation to tissue-level and organ-level diseases has required development of more complex, 3D, multicellular systems. Organoids and human-rodent chimaeras more accurately mirror the diverse cellular ecosystems of complex tissues and are being applied to iPSC disease models to recapitulate the pathobiology of a broad spectrum of human maladies, including infectious diseases, genetic disorders and cancer.
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16
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Mani C, Reddy PH, Palle K. DNA repair fidelity in stem cell maintenance, health, and disease. Biochim Biophys Acta Mol Basis Dis 2019; 1866:165444. [PMID: 30953688 DOI: 10.1016/j.bbadis.2019.03.017] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2018] [Revised: 12/20/2018] [Accepted: 01/06/2019] [Indexed: 12/13/2022]
Abstract
Stem cells are a sub population of cell types that form the foundation of our body, and have the potential to replicate, replenish and repair limitlessly to maintain the tissue and organ homeostasis. Increased lifetime and frequent replication set them vulnerable for both exogenous and endogenous agents-induced DNA damage compared to normal cells. To counter these damages and preserve genetic information, stem cells have evolved with various DNA damage response and repair mechanisms. Furthermore, upon experiencing irreparable DNA damage, stem cells mostly prefer early senescence or apoptosis to avoid the accumulation of damages. However, the failure of these mechanisms leads to various diseases, including cancer. Especially, given the importance of stem cells in early development, DNA repair deficiency in stem cells leads to various disabilities like developmental delay, premature aging, sensitivity to DNA damaging agents, degenerative diseases, etc. In this review, we have summarized the recent update about how DNA repair mechanisms are regulated in stem cells and their association with disease progression and pathogenesis.
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Affiliation(s)
- Chinnadurai Mani
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Centre, Lubbock, TX 79430, United States of America
| | - P Hemachandra Reddy
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Centre, Lubbock, TX 79430, United States of America
| | - Komaraiah Palle
- Department of Cell Biology and Biochemistry, Texas Tech University Health Sciences Centre, Lubbock, TX 79430, United States of America.
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17
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Eguizabal C, Aran B, Chuva de Sousa Lopes SM, Geens M, Heindryckx B, Panula S, Popovic M, Vassena R, Veiga A. Two decades of embryonic stem cells: a historical overview. Hum Reprod Open 2019; 2019:hoy024. [PMID: 30895264 PMCID: PMC6396646 DOI: 10.1093/hropen/hoy024] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2018] [Accepted: 12/10/2018] [Indexed: 12/12/2022] Open
Abstract
STUDY QUESTION How did the field of stem cell research develop in the years following the derivation of the first human embryonic stem cell (hESC) line? SUMMARY ANSWER Supported by the increasing number of clinical trials to date, significant technological advances in the past two decades have brought us ever closer to clinical therapies derived from pluripotent cells. WHAT IS KNOWN ALREADY Since their discovery 20 years ago, the use of human pluripotent stem cells has progressed tremendously from bench to bedside. Here, we provide a concise review of the main keystones of this journey and focus on ongoing clinical trials, while indicating the most relevant future research directions. STUDY DESIGN, SIZE, DURATION This is a historical narrative, including relevant publications in the field of pluripotent stem cells (PSC) derivation and differentiation, recounted both through scholarly research of published evidence and interviews of six pioneers who participated in some of the most relevant discoveries in the field. PARTICIPANTS/MATERIALS, SETTING, METHODS The authors all contributed by researching the literature and agreed upon body of works. Portions of the interviews of the field pioneers have been integrated into the review and have also been included in full for advanced reader interest. MAIN RESULTS AND THE ROLE OF CHANCE The stem cell field is ever expanding. We find that in the 20 years since the derivation of the first hESC lines, several relevant developments have shaped the pluripotent cell field, from the discovery of different states of pluripotency, the derivation of induced PSC, the refinement of differentiation protocols with several clinical trials underway, as well as the recent development of organoids. The challenge for the years to come will be to validate and refine PSCs for clinical use, from the production of highly defined cell populations in clinical grade conditions to the possibility of creating replacement organoids for functional, if not anatomical, function restoration. LIMITATIONS, REASONS FOR CAUTION This is a non-systematic review of current literature. Some references may have escaped the experts’ analysis due to the exceedingly diverse nature of the field. As the field of regenerative medicine is rapidly advancing, some of the most recent developments may have not been captured entirely. WIDER IMPLICATIONS OF THE FINDINGS The multi-disciplinary nature and tremendous potential of the stem cell field has important implications for basic as well as translational research. Recounting these activities will serve to provide an in-depth overview of the field, fostering a further understanding of human stem cell and developmental biology. The comprehensive overview of clinical trials and expert opinions included in this narrative may serve as a valuable scientific resource, supporting future efforts in translational approaches. STUDY FUNDING/COMPETING INTEREST(S) ESHRE provided funding for the authors’ on-site meeting and discussion during the preparation of this manuscript. S.M.C.S.L. is funded by the European Research Council Consolidator (ERC-CoG-725722-OVOGROWTH). M.P. is supported by the Special Research Fund, Bijzonder Onderzoeksfonds (BOF01D08114). M.G. is supported by the Methusalem grant of Vrije Universiteit Brussel, in the name of Prof. Karen Sermon and by Innovation by Science and Technology in Flanders (IWT, Project Number: 150042). A.V. and B.A. are supported by the Plataforma de Proteomica, Genotipado y Líneas Celulares (PT1770019/0015) (PRB3), Instituto de Salud Carlos III. Research grant to B.H. by the Research Foundation—Flanders (FWO) (FWO.KAN.2016.0005.01 and FWO.Project G051516N). There are no conflicts of interest to declare. TRIAL REGISTRATION NUMBER Not applicable. ESHRE Pages are not externally peer reviewed. This article has been approved by the Executive Committee of ESHRE.
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Affiliation(s)
- C Eguizabal
- Cell Therapy and Stem Cell Group, Basque Center for Blood Transfusion and Human Tissues, Barrio Labeaga S/N, Galdakao, Spain
| | - B Aran
- Barcelona Stem Cell Bank, Centre of Regenerative Medicine in Barcelona, Barcelona, Spain
| | - S M Chuva de Sousa Lopes
- Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, Leiden, The Netherlands.,Ghent Fertility and Stem cell Team (G-FaST), Department for Reproductive Medicine, Ghent University Hospital, Ghent, Belgium
| | - M Geens
- Research Group Reproduction and Genetics, Vrije Univeristeit Brussel, Laarbeeklaan 103, Jette (Brussels), Belgium
| | - B Heindryckx
- Ghent Fertility and Stem cell Team (G-FaST), Department for Reproductive Medicine, Ghent University Hospital, Ghent, Belgium
| | - S Panula
- Department of Clinical Science, Intervention and Technology, Karolinska Institutet, Stockholm, Sweden
| | - M Popovic
- Ghent Fertility and Stem cell Team (G-FaST), Department for Reproductive Medicine, Ghent University Hospital, Ghent, Belgium
| | | | - A Veiga
- Barcelona Stem Cell Bank, Centre of Regenerative Medicine in Barcelona, Barcelona, Spain.,Dexeus Mujer, Hospital Universitari Dexeus, Barcelona, Spain
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18
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Kaindl J, Winner B. Disease Modeling of Neuropsychiatric Brain Disorders Using Human Stem Cell-Based Neural Models. Curr Top Behav Neurosci 2019; 42:159-183. [PMID: 31407242 DOI: 10.1007/7854_2019_111] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/10/2023]
Abstract
Human pluripotent stem (PS) cells are a relevant platform to model human-specific neurological disorders. In this chapter, we focus on human stem cell models for neuropsychiatric disorders including induced pluripotent stem (iPS) cell-derived neural precursor cells (NPCs), neurons and cerebral organoids. We discuss crucial steps for planning human disease modeling experiments. We introduce the different strategies of human disease modeling including transdifferentiation, human embryonic stem (ES) cell-based models, iPS cell-based models and genome editing options. Analysis of disease-relevant phenotypes is discussed. In more detail, we provide exemplary insight into modeling of the neurodevelopmental defects in autism spectrum disorder (ASD) and the process of neurodegeneration in Alzheimer's disease (AD). Besides monogenic diseases, iPS cell-derived models also generated data from idiopathic and sporadic cases.
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Affiliation(s)
- Johanna Kaindl
- Department of Stem Cell Biology, Friedrich-Alexander-University (FAU) Erlangen-Nürnberg, Erlangen, Germany
| | - Beate Winner
- Department of Stem Cell Biology, Friedrich-Alexander-University (FAU) Erlangen-Nürnberg, Erlangen, Germany.
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19
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Omole AE, Fakoya AOJ. Ten years of progress and promise of induced pluripotent stem cells: historical origins, characteristics, mechanisms, limitations, and potential applications. PeerJ 2018; 6:e4370. [PMID: 29770269 PMCID: PMC5951134 DOI: 10.7717/peerj.4370] [Citation(s) in RCA: 101] [Impact Index Per Article: 14.4] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2017] [Accepted: 01/24/2018] [Indexed: 12/11/2022] Open
Abstract
The discovery of induced pluripotent stem cells (iPSCs) by Shinya Yamanaka in 2006 was heralded as a major breakthrough of the decade in stem cell research. The ability to reprogram human somatic cells to a pluripotent embryonic stem cell-like state through the ectopic expression of a combination of embryonic transcription factors was greeted with great excitement by scientists and bioethicists. The reprogramming technology offers the opportunity to generate patient-specific stem cells for modeling human diseases, drug development and screening, and individualized regenerative cell therapy. However, fundamental questions have been raised regarding the molecular mechanism of iPSCs generation, a process still poorly understood by scientists. The efficiency of reprogramming of iPSCs remains low due to the effect of various barriers to reprogramming. There is also the risk of chromosomal instability and oncogenic transformation associated with the use of viral vectors, such as retrovirus and lentivirus, which deliver the reprogramming transcription factors by integration in the host cell genome. These challenges can hinder the therapeutic prospects and promise of iPSCs and their clinical applications. Consequently, extensive studies have been done to elucidate the molecular mechanism of reprogramming and novel strategies have been identified which help to improve the efficiency of reprogramming methods and overcome the safety concerns linked with iPSC generation. Distinct barriers and enhancers of reprogramming have been elucidated, and non-integrating reprogramming methods have been reported. Here, we summarize the progress and the recent advances that have been made over the last 10 years in the iPSC field, with emphasis on the molecular mechanism of reprogramming, strategies to improve the efficiency of reprogramming, characteristics and limitations of iPSCs, and the progress made in the applications of iPSCs in the field of disease modelling, drug discovery and regenerative medicine. Additionally, this study appraises the role of genomic editing technology in the generation of healthy iPSCs.
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Affiliation(s)
- Adekunle Ebenezer Omole
- Department of Basic Sciences, American University of Antigua College of Medicine, St. John's, Antigua
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20
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Giacomelli E, Mummery CL, Bellin M. Human heart disease: lessons from human pluripotent stem cell-derived cardiomyocytes. Cell Mol Life Sci 2017; 74:3711-3739. [PMID: 28573431 PMCID: PMC5597692 DOI: 10.1007/s00018-017-2546-5] [Citation(s) in RCA: 46] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2016] [Revised: 05/09/2017] [Accepted: 05/23/2017] [Indexed: 02/07/2023]
Abstract
Technical advances in generating and phenotyping cardiomyocytes from human pluripotent stem cells (hPSC-CMs) are now driving their wider acceptance as in vitro models to understand human heart disease and discover therapeutic targets that may lead to new compounds for clinical use. Current literature clearly shows that hPSC-CMs recapitulate many molecular, cellular, and functional aspects of human heart pathophysiology and their responses to cardioactive drugs. Here, we provide a comprehensive overview of hPSC-CMs models that have been described to date and highlight their most recent and remarkable contributions to research on cardiovascular diseases and disorders with cardiac traits. We conclude discussing immediate challenges, limitations, and emerging solutions.
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Affiliation(s)
- E Giacomelli
- Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands
| | - C L Mummery
- Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands
- Department of Applied Stem Cell Technologies, University of Twente, Building Zuidhorst, 7500 AE, Enschede, The Netherlands
| | - M Bellin
- Department of Anatomy and Embryology, Leiden University Medical Center, Einthovenweg 20, 2333 ZC, Leiden, The Netherlands.
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21
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Chandrasekaran AP, Song M, Ramakrishna S. Genome editing: a robust technology for human stem cells. Cell Mol Life Sci 2017; 74:3335-3346. [PMID: 28405721 PMCID: PMC11107609 DOI: 10.1007/s00018-017-2522-0] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/21/2016] [Revised: 04/05/2017] [Accepted: 04/07/2017] [Indexed: 12/20/2022]
Abstract
Human pluripotent stem cells comprise induced pluripotent and embryonic stem cells, which have tremendous potential for biological and therapeutic applications. The development of efficient technologies for the targeted genome alteration of stem cells in disease models is a prerequisite for utilizing stem cells to their full potential. Genome editing of stem cells is possible with the help of synthetic nucleases that facilitate site-specific modification of a gene of interest. Recent advances in genome editing techniques have improved the efficiency and speed of the development of stem cells for human disease models. Zinc finger nucleases, transcription activator-like effector nucleases, and clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated system are powerful tools for editing DNA at specific loci. Here, we discuss recent technological advances in genome editing with site-specific nucleases in human stem cells.
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Affiliation(s)
| | - Minjung Song
- Division of Bioindustry, Department of Food Biotechnology, College of Medical and Life Science, Silla University, Seoul, Republic of Korea.
| | - Suresh Ramakrishna
- Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul, Republic of Korea.
- College of Medicine, Hanyang University, Seoul, Republic of Korea.
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22
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Abstract
It is extremely rare for a single experiment to be so impactful and timely that it shapes and forecasts the experiments of the next decade. Here, we review how two such experiments-the generation of human induced pluripotent stem cells (iPSCs) and the development of CRISPR/Cas9 technology-have fundamentally reshaped our approach to biomedical research, stem cell biology, and human genetics. We will also highlight the previous knowledge that iPSC and CRISPR/Cas9 technologies were built on as this groundwork demonstrated the need for solutions and the benefits that these technologies provided and set the stage for their success.
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Affiliation(s)
- Dirk Hockemeyer
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
| | - Rudolf Jaenisch
- The Whitehead Institute for Biomedical Research and Department of Biology, MIT, Cambridge, MA 02142, USA
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23
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May I Cut in? Gene Editing Approaches in Human Induced Pluripotent Stem Cells. Cells 2017; 6:cells6010005. [PMID: 28178187 PMCID: PMC5371870 DOI: 10.3390/cells6010005] [Citation(s) in RCA: 32] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2016] [Revised: 01/20/2017] [Accepted: 01/30/2017] [Indexed: 12/16/2022] Open
Abstract
In the decade since Yamanaka and colleagues described methods to reprogram somatic cells into a pluripotent state, human induced pluripotent stem cells (hiPSCs) have demonstrated tremendous promise in numerous disease modeling, drug discovery, and regenerative medicine applications. More recently, the development and refinement of advanced gene transduction and editing technologies have further accelerated the potential of hiPSCs. In this review, we discuss the various gene editing technologies that are being implemented with hiPSCs. Specifically, we describe the emergence of technologies including zinc-finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 that can be used to edit the genome at precise locations, and discuss the strengths and weaknesses of each of these technologies. In addition, we present the current applications of these technologies in elucidating the mechanisms of human development and disease, developing novel and effective therapeutic molecules, and engineering cell-based therapies. Finally, we discuss the emerging technological advances in targeted gene editing methods.
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24
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Human Embryonic Stem Cells: A Model for the Study of Neural Development and Neurological Diseases. Stem Cells Int 2016; 2016:2958210. [PMID: 27239201 PMCID: PMC4864561 DOI: 10.1155/2016/2958210] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2015] [Accepted: 03/14/2016] [Indexed: 01/05/2023] Open
Abstract
Although the mechanism of neurogenesis has been well documented in other organisms, there might be fundamental differences between human and those species referring to species-specific context. Based on principles learned from other systems, it is found that the signaling pathways required for neural induction and specification of human embryonic stem cells (hESCs) recapitulated those in the early embryo development in vivo at certain degree. This underscores the usefulness of hESCs in understanding early human neural development and reinforces the need to integrate the principles of developmental biology and hESC biology for an efficient neural differentiation.
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25
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Avior Y, Sagi I, Benvenisty N. Pluripotent stem cells in disease modelling and drug discovery. Nat Rev Mol Cell Biol 2016; 17:170-82. [DOI: 10.1038/nrm.2015.27] [Citation(s) in RCA: 413] [Impact Index Per Article: 45.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
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26
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Salazar-Noratto GE, Barry FP, Guldberg RE. Application of biomaterials to in vitro pluripotent stem cell disease modeling of the skeletal system. J Mater Chem B 2016; 4:3482-3489. [DOI: 10.1039/c5tb02645h] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Disease-specific pluripotent stem cells can be derived through genetic manipulation of embryonic stem cells or by reprogramming somatic cells (induced pluripotent stem cells).
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Affiliation(s)
- Giuliana E. Salazar-Noratto
- Wallace H. Coulter Department of Biomedical Engineering
- Georgia Institute of Technology and Emory University
- Atlanta
- USA
- Parker H. Petit Institute for Bioengineering and Bioscience
| | - Frank P. Barry
- Regenerative Medicine Institute
- National University of Ireland Galway
- Biosciences
- Dangan
- Ireland
| | - Robert E. Guldberg
- Parker H. Petit Institute for Bioengineering and Bioscience
- Georgia Institute of Technology
- Atlanta
- USA
- Woodruff School of Mechanical Engineering
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27
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Denning C, Borgdorff V, Crutchley J, Firth KSA, George V, Kalra S, Kondrashov A, Hoang MD, Mosqueira D, Patel A, Prodanov L, Rajamohan D, Skarnes WC, Smith JGW, Young LE. Cardiomyocytes from human pluripotent stem cells: From laboratory curiosity to industrial biomedical platform. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2015; 1863:1728-48. [PMID: 26524115 PMCID: PMC5221745 DOI: 10.1016/j.bbamcr.2015.10.014] [Citation(s) in RCA: 219] [Impact Index Per Article: 21.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/02/2015] [Revised: 10/12/2015] [Accepted: 10/20/2015] [Indexed: 12/14/2022]
Abstract
Cardiomyocytes from human pluripotent stem cells (hPSCs-CMs) could revolutionise biomedicine. Global burden of heart failure will soon reach USD $90bn, while unexpected cardiotoxicity underlies 28% of drug withdrawals. Advances in hPSC isolation, Cas9/CRISPR genome engineering and hPSC-CM differentiation have improved patient care, progressed drugs to clinic and opened a new era in safety pharmacology. Nevertheless, predictive cardiotoxicity using hPSC-CMs contrasts from failure to almost total success. Since this likely relates to cell immaturity, efforts are underway to use biochemical and biophysical cues to improve many of the ~30 structural and functional properties of hPSC-CMs towards those seen in adult CMs. Other developments needed for widespread hPSC-CM utility include subtype specification, cost reduction of large scale differentiation and elimination of the phenotyping bottleneck. This review will consider these factors in the evolution of hPSC-CM technologies, as well as their integration into high content industrial platforms that assess structure, mitochondrial function, electrophysiology, calcium transients and contractility. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Integration of Developmental and Environmental Cues in the Heart edited by Marcus Schaub and Hughes Abriel.
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Affiliation(s)
- Chris Denning
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom.
| | - Viola Borgdorff
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - James Crutchley
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Karl S A Firth
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Vinoj George
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Spandan Kalra
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Alexander Kondrashov
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Minh Duc Hoang
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Diogo Mosqueira
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Asha Patel
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Ljupcho Prodanov
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Divya Rajamohan
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - William C Skarnes
- Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, United Kingdom
| | - James G W Smith
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
| | - Lorraine E Young
- Department of Stem Cell Biology, Centre for Biomolecular Sciences, University of Nottingham, NG7 2RD, United Kingdom
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Hotta A, Yamanaka S. From Genomics to Gene Therapy: Induced Pluripotent Stem Cells Meet Genome Editing. Annu Rev Genet 2015; 49:47-70. [PMID: 26407033 DOI: 10.1146/annurev-genet-112414-054926] [Citation(s) in RCA: 91] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
The advent of induced pluripotent stem (iPS) cells has opened up numerous avenues of opportunity for cell therapy, including the initiation in September 2014 of the first human clinical trial to treat dry age-related macular degeneration. In parallel, advances in genome-editing technologies by site-specific nucleases have dramatically improved our ability to edit endogenous genomic sequences at targeted sites of interest. In fact, clinical trials have already begun to implement this technology to control HIV infection. Genome editing in iPS cells is a powerful tool and enables researchers to investigate the intricacies of the human genome in a dish. In the near future, the groundwork laid by such an approach may expand the possibilities of gene therapy for treating congenital disorders. In this review, we summarize the exciting progress being made in the utilization of genomic editing technologies in pluripotent stem cells and discuss remaining challenges toward gene therapy applications.
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Affiliation(s)
- Akitsu Hotta
- Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto 606-8501, Japan; .,Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Kyoto 606-8501, Japan
| | - Shinya Yamanaka
- Center for iPS Cell Research and Application (CiRA), Kyoto University, Kyoto 606-8501, Japan; .,Institute for Integrated Cell-Material Sciences (iCeMS), Kyoto University, Kyoto 606-8501, Japan.,Gladstone Institute of Cardiovascular Disease, San Francisco, California 94158
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29
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Telias M, Ben-Yosef D. Modeling neurodevelopmental disorders using human pluripotent stem cells. Stem Cell Rev Rep 2015; 10:494-511. [PMID: 24728983 DOI: 10.1007/s12015-014-9507-2] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Neurodevelopmental disorders (NDs) are impairments that affect the development and growth of the brain and the central nervous system during embryonic and early postnatal life. Genetically manipulated animals have contributed greatly to the advancement of ND research, but many of them differ considerably from the human phenotype. Cellular in vitro models are also valuable, but the availability of human neuronal cells is limited and their lifespan in culture is short. Human pluripotent stem cells (hPSCs), including embryonic stem cells and induced pluripotent stem cells, comprise a powerful tool for studying developmentally regulated diseases, including NDs. We reviewed all recent studies in which hPSCs were used as in vitro models for diseases and syndromes characterized by impairment of neurogenesis or synaptogenesis leading to intellectual disability and delayed neurodevelopment. We analyzed their methodology and results, focusing on the data obtained following in vitro neural differentiation and gene expression and profiling of the derived neurons. Electrophysiological recording of action potentials, synaptic currents and response to neurotransmitters is pivotal for validation of the neuronal fate as well as for assessing phenotypic dysfunctions linked to the disease in question. We therefore focused on the studies which included electrophysiological recordings on the in vitro-derived neurons. Finally, we addressed specific issues that are critical for the advancement of this area of research, specifically in providing a reliable human pre-clinical research model and drug screening platform.
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Affiliation(s)
- Michael Telias
- The Wolfe PGD-Stem Cell Lab, Racine IVF Unit, Lis Maternity Hospital, Tel-Aviv Sourasky Medical Center, Department of Cell and Developmental Biology, Sackler Faculty of Medicine, Tel-Aviv University, Tel-Aviv, Israel
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Cerbini T, Luo Y, Rao MS, Zou J. Transfection, selection, and colony-picking of human induced pluripotent stem cells TALEN-targeted with a GFP gene into the AAVS1 safe harbor. J Vis Exp 2015. [PMID: 25741760 DOI: 10.3791/52504] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
Targeted transgene addition can provide persistent gene expression while circumventing the gene silencing and insertional mutagenesis caused by viral vector mediated random integration. This protocol describes a universal and efficient transgene targeted addition platform in human iPSCs based on utilization of validated open-source TALENs and a gene-trap-like donor to deliver transgenes into a safe harbor locus. Importantly, effective gene editing is rate-limited by the delivery efficiency of gene editing vectors. Therefore, this protocol first focuses on preparation of iPSCs for transfection to achieve high nuclear delivery efficiency. When iPSCs are dissociated into single cells using a gentle-cell dissociation reagent and transfected using an optimized program, >50% cells can be induced to take up the large gene editing vectors. Because the AAVS1 locus is located in the intron of an active gene (PPP1R12C), a splicing acceptor (SA)-linked puromycin resistant gene (PAC) was used to select targeted iPSCs while excluding random integration-only and untransfected cells. This strategy greatly increases the chance of obtaining targeted clones, and can be used in other active gene targeting experiments as well. Two weeks after puromycin selection at the dose adjusted for the specific iPSC line, clones are ready to be picked by manual dissection of large, isolated colonies into smaller pieces that are transferred to fresh medium in a smaller well for further expansion and genetic and functional screening. One can follow this protocol to readily obtain multiple GFP reporter iPSC lines that are useful for in vivo and in vitro imaging and cell isolation.
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Affiliation(s)
- Trevor Cerbini
- National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health
| | - Yongquan Luo
- National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health
| | | | - Jizhong Zou
- National Institute of Arthritis, Musculoskeletal and Skin Diseases, National Institutes of Health;
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31
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Halevy T, Urbach A. Comparing ESC and iPSC-Based Models for Human Genetic Disorders. J Clin Med 2014; 3:1146-62. [PMID: 26237596 PMCID: PMC4470175 DOI: 10.3390/jcm3041146] [Citation(s) in RCA: 48] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/10/2014] [Revised: 09/29/2014] [Accepted: 09/30/2014] [Indexed: 12/19/2022] Open
Abstract
Traditionally, human disorders were studied using animal models or somatic cells taken from patients. Such studies enabled the analysis of the molecular mechanisms of numerous disorders, and led to the discovery of new treatments. Yet, these systems are limited or even irrelevant in modeling multiple genetic diseases. The isolation of human embryonic stem cells (ESCs) from diseased blastocysts, the derivation of induced pluripotent stem cells (iPSCs) from patients' somatic cells, and the new technologies for genome editing of pluripotent stem cells have opened a new window of opportunities in the field of disease modeling, and enabled studying diseases that couldn't be modeled in the past. Importantly, despite the high similarity between ESCs and iPSCs, there are several fundamental differences between these cells, which have important implications regarding disease modeling. In this review we compare ESC-based models to iPSC-based models, and highlight the advantages and disadvantages of each system. We further suggest a roadmap for how to choose the optimal strategy to model each specific disorder.
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Affiliation(s)
- Tomer Halevy
- Stem Cell Unit, Department of Genetics, Institute of Life Sciences, The Hebrew University of Jerusalem, Jerusalem 91904, Israel.
| | - Achia Urbach
- Mina and Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 5290002, Israel.
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32
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Diecke S, Jung SM, Lee J, Ju JH. Recent technological updates and clinical applications of induced pluripotent stem cells. Korean J Intern Med 2014; 29:547-57. [PMID: 25228828 PMCID: PMC4164716 DOI: 10.3904/kjim.2014.29.5.547] [Citation(s) in RCA: 27] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/21/2014] [Accepted: 07/22/2014] [Indexed: 12/23/2022] Open
Abstract
Induced pluripotent stem cells (iPSCs) were first described in 2006 and have since emerged as a promising cell source for clinical applications. The rapid progression in iPSC technology is still ongoing and directed toward increasing the efficacy of iPSC production and reducing the immunogenic and tumorigenic potential of these cells. Enormous efforts have been made to apply iPSC-based technology in the clinic, for drug screening approaches and cell replacement therapy. Moreover, disease modeling using patient-specific iPSCs continues to expand our knowledge regarding the pathophysiology and prospective treatment of rare disorders. Furthermore, autologous stem cell therapy with patient-specific iPSCs shows great propensity for the minimization of immune reactions and the provision of a limitless supply of cells for transplantation. In this review, we discuss the recent updates in iPSC technology and the use of iPSCs in disease modeling and regenerative medicine.
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Affiliation(s)
- Sebastian Diecke
- Division of Cardiology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
- Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA, USA
- Stanford Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Seung Min Jung
- Division of Rheumatology, Department of Internal Medicine, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
| | - Jaecheol Lee
- Division of Cardiology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA
- Institute for Stem Cell Biology and Regenerative Medicine, Stanford University School of Medicine, Stanford, CA, USA
- Stanford Cardiovascular Institute, Stanford University School of Medicine, Stanford, CA, USA
| | - Ji Hyeon Ju
- Division of Rheumatology, Department of Internal Medicine, Seoul St. Mary's Hospital, College of Medicine, The Catholic University of Korea, Seoul, Korea
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33
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Sherman SL, Curnow EC, Easley CA, Jin P, Hukema RK, Tejada MI, Willemsen R, Usdin K. Use of model systems to understand the etiology of fragile X-associated primary ovarian insufficiency (FXPOI). J Neurodev Disord 2014; 6:26. [PMID: 25147583 PMCID: PMC4139715 DOI: 10.1186/1866-1955-6-26] [Citation(s) in RCA: 47] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/16/2013] [Accepted: 08/13/2014] [Indexed: 01/04/2023] Open
Abstract
Fragile X-associated primary ovarian insufficiency (FXPOI) is among the family of disorders caused by the expansion of a CGG repeat sequence in the 5' untranslated region of the X-linked gene FMR1. About 20% of women who carry the premutation allele (55 to 200 unmethylated CGG repeats) develop hypergonadotropic hypogonadism and cease menstruating before age 40. Some proportion of those who are still cycling show hormonal profiles indicative of ovarian dysfunction. FXPOI leads to subfertility and an increased risk of medical conditions associated with early estrogen deficiency. Little progress has been made in understanding the etiology of this clinically significant disorder. Understanding the molecular mechanisms of FXPOI requires a detailed knowledge of ovarian FMR1 mRNA and FMRP’s function. In humans, non-invasive methods to discriminate the mechanisms of the premutation on ovarian function are not available, thus necessitating the development of model systems. Vertebrate (mouse and rat) and invertebrate (Drosophila melanogaster) animal studies for the FMR1 premutation and ovarian function exist and have been instrumental in advancing our understanding of the disease phenotype. For example, rodent models have shown that FMRP is highly expressed in oocytes where it is important for folliculogenesis. The two premutation mouse models studied to date show evidence of ovarian dysfunction and, together, suggest that the long repeat in the transcript itself may have some pathological effect quite apart from any effect of the toxic protein. Further, ovarian morphology in young animals appears normal and the primordial follicle pool size does not differ from that of wild-type animals. However, there is a progressive premature decline in the levels of most follicle classes. Observations also include granulosa cell abnormalities and altered gene expression patterns. Further comparisons of these models are now needed to gain insight into the etiology of the ovarian dysfunction. Premutation model systems in non-human primates and those based on induced pluripotent stem cells show particular promise and will complement current models. Here, we review the characterization of the current models and describe the development and potential of the new models. Finally, we will discuss some of the molecular mechanisms that might be responsible for FXPOI.
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Affiliation(s)
- Stephanie L Sherman
- Department of Human Genetics, Emory University, 615 Michael St, Emory University, Atlanta, GA 30322, USA
| | - Eliza C Curnow
- Washington National Primate Center, University of Washington, Seattle, WA, USA
| | - Charles A Easley
- Laboratory of Translational Cell Biology, Department of Cell Biology, Emory University, Atlanta, GA, USA
| | - Peng Jin
- Department of Human Genetics, Emory University, 615 Michael St, Emory University, Atlanta, GA 30322, USA
| | - Renate K Hukema
- Department of Clinical Genetics, Erasmus MC, Rotterdam, The Netherlands
| | - Maria Isabel Tejada
- Molecular Genetics Laboratory, Genetics Service, BioCruces Health Research Institute, Hospital Universitario Cruces, Barakaldo, Biscay, Spain
| | - Rob Willemsen
- Department of Clinical Genetics, Erasmus MC, Rotterdam, The Netherlands
| | - Karen Usdin
- Laboratory of Molecular and Cellular Biology, NIDDK, National Institutes of Health, Bethesda, MD, USA
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Induced pluripotent stem cell technology in regenerative medicine and biology. ADVANCES IN BIOCHEMICAL ENGINEERING/BIOTECHNOLOGY 2014; 123:127-41. [PMID: 20549468 DOI: 10.1007/10_2010_72] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Subscribe] [Scholar Register] [Indexed: 10/19/2022]
Abstract
The potential of human embryonic stem cells (ESCs) for regenerative medicine is unquestionable, but practical and ethical considerations have hampered clinical application and research. In an attempt to overcome these issues, the conversion of somatic cells into pluripotent stem cells similar to ESCs, commonly termed nuclear reprogramming, has been a top objective of contemporary biology. More than 40 years ago, King, Briggs, and Gurdon pioneered somatic cell nuclear reprogramming in frogs, and in 1981 Evans successfully isolated mouse ESCs. In 1997 Wilmut and collaborators produced the first cloned mammal using nuclear transfer, and then Thomson obtained human ESCs from in vitro fertilized blastocysts in 1998. Over the last 2 decades we have also seen remarkable findings regarding how ESC behavior is controlled, the importance of which should not be underestimated. This knowledge allowed the laboratory of Shinya Yamanaka to overcome brilliantly conceptual and technical barriers in 2006 and generate induced pluripotent stem cells (iPSCs) from mouse fibroblasts by overexpressing defined combinations of ESC-enriched transcription factors. Here, we discuss some important implications of human iPSCs for biology and medicine and also point to possible future directions.
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35
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Abstract
One of the great advantages of embryonic stem (ES) cells over other cell types is their accessibility to genetic manipulation. They can easily undergo genetic modifications while remaining pluripotent, and can be selectively propagated, allowing the clonal expansion of genetically altered cells in culture. Since the first isolation of ES cells in mice, many effective techniques have been developed for gene delivery and manipulation of ES cells. These include transfection, electroporation, and infection protocols, as well as different approaches for inserting, deleting, or changing the expression of genes. These methods proved to be extremely useful in mouse ES cells, for monitoring and directing differentiation, discovering unknown genes, and studying their function, and are now being extensively implemented in human ES cells (HESCs). This chapter describes the different approaches and methodologies that have been applied for the genetic manipulation of HESCs and their applications. Detailed protocols for generating clones of genetically modified HESCs by transfection, electroporation, and infection will be described, with special emphasis on the important technical details that are required for this purpose. All protocols are equally effective in human-induced pluripotent stem (iPS) cells.
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Affiliation(s)
- Rachel Eiges
- Stem Cell Research Laboratory, Medical Genetics Institute, Shaare Zedek Medical Center, 3235, Jerusalem, 91031, Israel,
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36
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Burbacher TM, Grant KS, Worlein J, Ha J, Curnow E, Juul S, Sackett GP. Four decades of leading-edge research in the reproductive and developmental sciences: the Infant Primate Research Laboratory at the University of Washington National Primate Research Center. Am J Primatol 2013; 75:1063-83. [PMID: 23873400 PMCID: PMC5452618 DOI: 10.1002/ajp.22175] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/21/2013] [Revised: 05/31/2013] [Accepted: 05/31/2013] [Indexed: 12/20/2022]
Abstract
The Infant Primate Research Laboratory (IPRL) was established in 1970 at the University of Washington as a visionary project of Dr. Gene (Jim) P. Sackett. Supported by a collaboration between the Washington National Primate Research Center and the Center on Human Development and Disability, the IPRL operates under the principle that learning more about the causes of abnormal development in macaque monkeys will provide important insights into the origins and treatment of childhood neurodevelopmental disabilities. Over the past 40 years, a broad range of research projects have been conducted at the IPRL. Some have described the expression of normative behaviors in nursery-reared macaques while others have focused on important biomedical themes in child health and development. This article details the unique scientific history of the IPRL and the contributions produced by research conducted in the laboratory. Past and present investigations have explored the topics of early rearing effects, low-birth-weight, prematurity, birth injury, epilepsy, prenatal neurotoxicant exposure, viral infection (pediatric HIV), diarrheal disease, vaccine safety, and assisted reproductive technologies. Data from these studies have helped advance our understanding of both risk and resiliency in primate development. New directions of research at the IPRL include the production of transgenic primate models using our embryonic stem cell-based technology to better understand and treat heritable forms of human intellectual disabilities such as fragile X.
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Affiliation(s)
- Thomas M. Burbacher
- Department of Environmental and Occupational Health Sciences, School of Public Health, University of Washington, Seattle, WA, 98195 USA
- Center on Human Development and Disability, University of Washington, Seattle, WA, 98195 USA
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
| | - Kimberly S. Grant
- Department of Environmental and Occupational Health Sciences, School of Public Health, University of Washington, Seattle, WA, 98195 USA
- Center on Human Development and Disability, University of Washington, Seattle, WA, 98195 USA
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
| | - Julie Worlein
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
| | - James Ha
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
- Department of Psychology, School of Arts and Sciences, University of Washington, Seattle, WA, 98195 USA
| | - Eliza Curnow
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
| | - Sandra Juul
- Center on Human Development and Disability, University of Washington, Seattle, WA, 98195 USA
- Department of Pediatrics, School of Medicine, University of Washington, Seattle, WA, 98195 USA
| | - Gene P. Sackett
- Center on Human Development and Disability, University of Washington, Seattle, WA, 98195 USA
- Washington National Primate Research Center, University of Washington, Seattle, WA 98195 USA
- Department of Psychology, School of Arts and Sciences, University of Washington, Seattle, WA, 98195 USA
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Moretti A, Laugwitz KL, Dorn T, Sinnecker D, Mummery C. Pluripotent stem cell models of human heart disease. Cold Spring Harb Perspect Med 2013; 3:3/11/a014027. [PMID: 24186488 DOI: 10.1101/cshperspect.a014027] [Citation(s) in RCA: 59] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
Abstract
Understanding the molecular basis of many cardiac diseases has been hampered by the lack of appropriate in vitro cell culture models that accurately reflect the human disease phenotypes. In the past few years, remarkable advances in stem cell biology have made possible this long-standing ambition-the generation of human and even patient-specific cellular models of diseases. Combined with other novel technologies in the fields of human genetics, tissue engineering, and gene-targeted manipulation, disease modeling with pluripotent stem cells has the promise to influence modern cardiovascular medicine on several fronts: molecular understanding of pathological mechanisms, early diagnosis, drug development, and effective treatment.
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Affiliation(s)
- Alessandra Moretti
- Klinikum rechts der Isar-Technische Universität München, I. Medical Department-Cardiology, 81675 Munich, Germany
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Ramamoorthi K, Curtis D, Asuri P. Advances in homology directed genetic engineering of human pluripotent and adult stem cells. World J Stem Cells 2013; 5:98-105. [PMID: 24179598 PMCID: PMC3812527 DOI: 10.4252/wjsc.v5.i4.98] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/26/2013] [Revised: 09/06/2013] [Accepted: 09/17/2013] [Indexed: 02/06/2023] Open
Abstract
The ability to introduce precise genomic modifications in human cells has profound implications for both basic and applied research in stem cells, ranging from identification of genes regulating stem cell self-renewal and multilineage differentiation to therapeutic gene correction and creation of in vitro models of human diseases. However, the overall efficiency of this process is challenged by several factors including inefficient gene delivery into stem cells and low rates of homology directed site-specific targeting. Recent studies report the development of novel techniques to improve gene targeting efficiencies in human stem cells; these methods include molecular engineering of viral vectors to efficiently deliver episomal genetic sequences that can participate in homology directed targeting, as well as the design of synthetic proteins that can introduce double-stranded breaks in DNA to initiate such recombination events. This review focuses on the potential of these new technologies to precisely alter the human stem cell genome and also highlights the possibilities offered by the combination of these complementary strategies.
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Viral and non-viral gene delivery and its role in pluripotent stem cell engineering. DRUG DISCOVERY TODAY. TECHNOLOGIES 2013; 5:e105-48. [PMID: 24125542 DOI: 10.1016/j.ddtec.2008.10.002] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/14/2023]
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Stem cell reprogramming: generation of patient-specific stem cells by somatic cell nuclear reprogramming. DRUG DISCOVERY TODAY. TECHNOLOGIES 2013; 5:e105-48. [PMID: 24125543 DOI: 10.1016/j.ddtec.2008.11.006] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
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Fontes A, Lakshmipathy U. Advances in genetic modification of pluripotent stem cells. Biotechnol Adv 2013; 31:994-1001. [PMID: 23856320 DOI: 10.1016/j.biotechadv.2013.07.003] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/03/2012] [Revised: 05/24/2013] [Accepted: 07/04/2013] [Indexed: 12/20/2022]
Abstract
Genetically engineered stem cells aid in dissecting basic cell function and are valuable tools for drug discovery, in vivo cell tracking, and gene therapy. Gene transfer into pluripotent stem cells has been a challenge due to their intrinsic feature of growing in clusters and hence not amenable to common gene delivery methods. Several advances have been made in the rapid assembly of DNA elements, optimization of culture conditions, and DNA delivery methods. This has lead to the development of viral and non-viral methods for transient or stable modification of cells, albeit with varying efficiencies. Most methods require selection and clonal expansion that demand prolonged culture and are not suited for cells with limited proliferative potential. Choosing the right platform based on preferred length, strength, and context of transgene expression is a critical step. Random integration of the transgene into the genome can be complicated due to silencing or altered regulation of expression due to genomic effects. An alternative to this are site-specific methods that target transgenes followed by screening to identify the genomic loci that support long-term expression with stem cell proliferation and differentiation. A highly precise and accurate editing of the genome driven by homology can be achieved using traditional methods as well as the newer technologies such as zinc finger nuclease, TAL effector nucleases and CRISPR. In this review, we summarize the different genetic engineering methods that have been successfully used to create modified embryonic and induced pluripotent stem cells.
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Affiliation(s)
- Andrew Fontes
- Primary and Stem Cell Systems, Life Technologies, 5781 Van Allen Way, Carlsbad, CA 92008, USA
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Sinnecker D, Goedel A, Laugwitz KL, Moretti A. Induced pluripotent stem cell-derived cardiomyocytes: a versatile tool for arrhythmia research. Circ Res 2013; 112:961-8. [PMID: 23569105 DOI: 10.1161/circresaha.112.268623] [Citation(s) in RCA: 41] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Induced pluripotent stem cells offer the possibility to generate patient-specific stem cell lines from individuals affected by inherited disorders. Cardiomyocytes differentiated from such patient-specific induced pluripotent stem cells lines have been used to study the pathophysiology of arrhythmogenic heart diseases, such as the long-QT syndrome or catecholaminergic polymorphic ventricular tachycardia. Testing for unwanted drug side effects or tailoring medical treatment to the specific needs of individual patients with arrhythmogenic disorders may become future applications of this emerging technology.
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Affiliation(s)
- Daniel Sinnecker
- Klinikum rechts der Isar, Technische Universität München, I. Medizinische Klinik, Kardiologie, Munich, Germany
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Zhu Z, Huangfu D. Human pluripotent stem cells: an emerging model in developmental biology. Development 2013; 140:705-17. [PMID: 23362344 DOI: 10.1242/dev.086165] [Citation(s) in RCA: 130] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
Abstract
Developmental biology has long benefited from studies of classic model organisms. Recently, human pluripotent stem cells (hPSCs), including human embryonic stem cells and human induced pluripotent stem cells, have emerged as a new model system that offers unique advantages for developmental studies. Here, we discuss how studies of hPSCs can complement classic approaches using model organisms, and how hPSCs can be used to recapitulate aspects of human embryonic development 'in a dish'. We also summarize some of the recently developed genetic tools that greatly facilitate the interrogation of gene function during hPSC differentiation. With the development of high-throughput screening technologies, hPSCs have the potential to revolutionize gene discovery in mammalian development.
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Affiliation(s)
- Zengrong Zhu
- Developmental Biology Program, Sloan-Kettering Institute, New York, NY 10065, USA.
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Modeling hereditary cardiac disease with patient-specific-induced pluripotent stem cells: opportunities and concerns. J Cardiovasc Pharmacol 2013; 60:406-7. [PMID: 22653418 DOI: 10.1097/fjc.0b013e31825f3f0f] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023]
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Umeda K, Suzuki K, Yamazoe T, Shiraki N, Higuchi Y, Tokieda K, Kume K, Mitani K, Kume S. Albumin gene targeting in human embryonic stem cells and induced pluripotent stem cells with helper-dependent adenoviral vector to monitor hepatic differentiation. Stem Cell Res 2013; 10:179-94. [PMID: 23276698 DOI: 10.1016/j.scr.2012.11.003] [Citation(s) in RCA: 20] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/31/2012] [Revised: 10/10/2012] [Accepted: 11/09/2012] [Indexed: 01/16/2023] Open
Abstract
Although progresses in developing differentiation procedures have been achieved, it remains challenging to generate hES/iPS cell-derived mature hepatocytes. We performed knock-in of a monomeric Kusabira orange (mKO1) cassette in the albumin (ALB) gene, in human embryonic stem (hES) cells and induced pluripotent stem (hiPS) cells, with the use of the helper-dependent adenovirus vector (HDAdV). Upon induction into the hepatic lineages, these knock-in hES/iPS cells differentiated into cells that displayed several known hepatic functions. The mKO1 knock-in (ALB/mKo1) hES/hiPS cells were used to visualize hepatic differentiation in vitro. mKO1 reporter expression recapitulated endogenous ALB transcriptional activity. ALB/mKo1 [Hi] population isolated by flow cytometry was confirmed to be enriched with ALB mRNA. Expression profile analyses revealed that characteristic hepatocyte genes and genes related to drug metabolism and many aspects of liver function were highly enriched in the ALB/mKo1 [Hi] population. Our data demonstrate that ALB/mKo1 knock-in hES/iPS cells are valuable resources for monitoring in vitro hepatic differentiation, isolation and analyses of hES and hiPS cells-derived hepatic cells that actively transcribing ALB. These knock-in hES/iPS cell lines could provide further insights into the mechanism of hepatic differentiation and molecular signatures of the hepatic cells derived from hES/iPS cells.
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Affiliation(s)
- Kahoko Umeda
- Department of Stem Cell Biology, Institute of Molecular Embryology and Genetics, Kumamoto University, Honjo 2-2-1, Kumamoto 860-0811, Japan
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Han D, Binas B. Monoallelic gene targeting in hypoblast stem cells reveals X chromosome inactivation. Biochem Biophys Res Commun 2012; 427:563-7. [PMID: 23022182 DOI: 10.1016/j.bbrc.2012.09.097] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/07/2012] [Accepted: 09/18/2012] [Indexed: 11/30/2022]
Abstract
We recently isolated hypoblast stem cells (HypoSC), which are related to embryonic stem (ES) cells. ES cells efficiently perform homologous recombination (HR) and lack X chromosome inactivation (Xi), but it is unknown whether the same applies to HypoSC. Using the X-linked hypoxanthine phosphoribosyl transferase (HPRT) gene, we find that HypoSC perform HR with similar frequency as ES cells. Monoallelic targeting in female HypoSC eliminated HPRT gene expression, implying epigenetic inactivation of the other allele. Although density-induced differentiation complicated selection, the targeted clones maintained their original properties. These results will facilitate targeted genetic manipulation of HypoSC and the study of Xi.
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Affiliation(s)
- Dongjun Han
- Division of Molecular & Life Science, College of Science & Technology, Hanyang University, Ansan 426-791, South Korea
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Abstract
The boundaries of embryonic stem cell (ESC) research have extended considerably in recent years in several important ways. Alongside a deeper understanding of the pluripotent state, ESCs have been successfully integrated into various fields, such as genomics, epigenetics, and disease modeling. Significant progress in cell fate control has pushed directed differentiation and tissue engineering further than ever before and promoted clinical trials. The geographical distribution of research activity has also expanded, especially for human ESCs. This review outlines these developments and future challenges that remain.
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Bar-Nur O. Harnessing induced pluripotent stem cells for the modeling and treatment of neurological disorders. FUTURE NEUROLOGY 2012. [DOI: 10.2217/fnl.12.57] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Affiliation(s)
- Ori Bar-Nur
- Stem Cell Unit, Department of Genetics, Institute of Life Sciences, The Hebrew University of Jerusalem, Givat Ram 91904, Israel
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Rajamohan D, Matsa E, Kalra S, Crutchley J, Patel A, George V, Denning C. Current status of drug screening and disease modelling in human pluripotent stem cells. Bioessays 2012; 35:281-98. [PMID: 22886688 PMCID: PMC3597971 DOI: 10.1002/bies.201200053] [Citation(s) in RCA: 83] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
The emphasis in human pluripotent stem cell (hPSC) technologies has shifted from cell therapy to in vitro disease modelling and drug screening. This review examines why this shift has occurred, and how current technological limitations might be overcome to fully realise the potential of hPSCs. Details are provided for all disease-specific human induced pluripotent stem cell lines spanning a dozen dysfunctional organ systems. Phenotype and pharmacology have been examined in only 17 of 63 lines, primarily those that model neurological and cardiac conditions. Drug screening is most advanced in hPSC-cardiomyocytes. Responses for almost 60 agents include examples of how careful tests in hPSC-cardiomyocytes have improved on existing in vitro assays, and how these cells have been integrated into high throughput imaging and electrophysiology industrial platforms. Such successes will provide an incentive to overcome bottlenecks in hPSC technology such as improving cell maturity and industrial scalability whilst reducing cost.
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Affiliation(s)
- Divya Rajamohan
- Department of Stem Cells, Tissue Engineering & Modelling, Centre for Biomolecular Sciences, University of Nottingham, Nottingham, UK
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