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1
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Marzoog BA. Transcription Factors in Brain Regeneration: A Potential Novel Therapeutic Target. Curr Drug Targets 2024; 25:46-61. [PMID: 38444255 DOI: 10.2174/0113894501279977231210170231] [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: 08/30/2023] [Revised: 11/21/2023] [Accepted: 11/23/2023] [Indexed: 03/07/2024]
Abstract
Transcription factors play a crucial role in providing identity to each cell population. To maintain cell identity, it is essential to balance the expression of activator and inhibitor transcription factors. Cell plasticity and reprogramming offer great potential for future therapeutic applications, as they can regenerate damaged tissue. Specific niche factors can modify gene expression and differentiate or transdifferentiate the target cell to the required fate. Ongoing research is being carried out on the possibilities of transcription factors in regenerating neurons, with neural stem cells (NSCs) being considered the preferred cells for generating new neurons due to their epigenomic and transcriptome memory. NEUROD1/ASCL1, BRN2, MYTL1, and other transcription factors can induce direct reprogramming of somatic cells, such as fibroblasts, into neurons. However, the molecular biology of transcription factors in reprogramming and differentiation still needs to be fully understood.
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Affiliation(s)
- Basheer Abdullah Marzoog
- World-Class Research Center, Digital Biodesign and Personalized Healthcare», I.M. Sechenov First Moscow State Medical University (Sechenov University), 119991 Moscow, Russia
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2
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Aversano S, Palladino R, Caiazzo M. Direct Cell Conversion of Somatic Cells into Dopamine Neurons: Achievements and Perspectives. Cell Reprogram 2022; 24:259-270. [PMID: 36137065 DOI: 10.1089/cell.2022.0065] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Abstract
In the last decade, direct reprogramming has emerged as a novel strategy to obtain mature and functional dopamine neurons from somatic cells. This approach could overcome issues linked to the use of human pluripotent stem cells such as ethical concerns and safety problems that can arise from the overgrowth of undifferentiated cells after transplantation. Several conversion methodologies have been developed to obtain induced DA neurons (iDANs) or induced DA neuron progenitors (iDPs). iDANs have also proved to successfully integrate in mice striatum, alleviating Parkinson's disease (PD) motor symptoms. In the next decade, human iDANs and/or iDPs could be translated to clinic to achieve a patient-tailored therapy, but current critical issues hinder this goal, such as the low conversion rate of adult human fibroblasts and the risks associated with lentiviral delivery of conversion factors. In this study, we summarize the strategies and recent improvements developed for the generation of mouse and human iDANs/iDPs. Furthermore, we discuss the more recent application of in vivo direct conversion, which may enable clinical therapies for PD by means of brain in situ delivery of dopaminergic reprogramming transcription factors.
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Affiliation(s)
- Simona Aversano
- Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II," Naples, Italy
| | - Renata Palladino
- Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II," Naples, Italy
| | - Massimiliano Caiazzo
- Department of Molecular Medicine and Medical Biotechnology, University of Naples "Federico II," Naples, Italy.,Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Utrecht, The Netherlands
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3
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Pascale E, Caiazza C, Paladino M, Parisi S, Passaro F, Caiazzo M. MicroRNA Roles in Cell Reprogramming Mechanisms. Cells 2022; 11:940. [PMID: 35326391 PMCID: PMC8946776 DOI: 10.3390/cells11060940] [Citation(s) in RCA: 11] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2022] [Revised: 02/28/2022] [Accepted: 03/08/2022] [Indexed: 02/01/2023] Open
Abstract
Cell reprogramming is a groundbreaking technology that, in few decades, generated a new paradigm in biomedical science. To date we can use cell reprogramming to potentially generate every cell type by converting somatic cells and suitably modulating the expression of key transcription factors. This approach can be used to convert skin fibroblasts into pluripotent stem cells as well as into a variety of differentiated and medically relevant cell types, including cardiomyocytes and neural cells. The molecular mechanisms underlying such striking cell phenotypes are still largely unknown, but in the last decade it has been proven that cell reprogramming approaches are significantly influenced by non-coding RNAs. Specifically, this review will focus on the role of microRNAs in the reprogramming processes that lead to the generation of pluripotent stem cells, neurons, and cardiomyocytes. As highlighted here, non-coding RNA-forced expression can be sufficient to support some cell reprogramming processes, and, therefore, we will also discuss how these molecular determinants could be used in the future for biomedical purposes.
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Affiliation(s)
- Emilia Pascale
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
| | - Carmen Caiazza
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
| | - Martina Paladino
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
| | - Silvia Parisi
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
| | - Fabiana Passaro
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
| | - Massimiliano Caiazzo
- Department of Molecular Medicine and Medical Biotechnology, University of Naples “Federico II”, Via Pansini 5, 80131 Naples, Italy; (E.P.); (C.C.); (M.P.); (S.P.)
- Department of Pharmaceutics, Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University, Universiteitsweg 99, 3584 CG Utrecht, The Netherlands
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4
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Notaras M, Lodhi A, Dündar F, Collier P, Sayles NM, Tilgner H, Greening D, Colak D. Schizophrenia is defined by cell-specific neuropathology and multiple neurodevelopmental mechanisms in patient-derived cerebral organoids. Mol Psychiatry 2022; 27:1416-1434. [PMID: 34789849 PMCID: PMC9095467 DOI: 10.1038/s41380-021-01316-6] [Citation(s) in RCA: 74] [Impact Index Per Article: 24.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/26/2021] [Revised: 08/03/2021] [Accepted: 09/22/2021] [Indexed: 01/02/2023]
Abstract
Due to an inability to ethically access developing human brain tissue as well as identify prospective cases, early-arising neurodevelopmental and cell-specific signatures of Schizophrenia (Scz) have remained unknown and thus undefined. To overcome these challenges, we utilized patient-derived induced pluripotent stem cells (iPSCs) to generate 3D cerebral organoids to model neuropathology of Scz during this critical period. We discovered that Scz organoids exhibited ventricular neuropathology resulting in altered progenitor survival and disrupted neurogenesis. This ultimately yielded fewer neurons within developing cortical fields of Scz organoids. Single-cell sequencing revealed that Scz progenitors were specifically depleted of neuronal programming factors leading to a remodeling of cell-lineages, altered differentiation trajectories, and distorted cortical cell-type diversity. While Scz organoids were similar in their macromolecular diversity to organoids generated from healthy controls (Ctrls), four GWAS factors (PTN, COMT, PLCL1, and PODXL) and peptide fragments belonging to the POU-domain transcription factor family (e.g., POU3F2/BRN2) were altered. This revealed that Scz organoids principally differed not in their proteomic diversity, but specifically in their total quantity of disease and neurodevelopmental factors at the molecular level. Single-cell sequencing subsequently identified cell-type specific alterations in neuronal programming factors as well as a developmental switch in neurotrophic growth factor expression, indicating that Scz neuropathology can be encoded on a cell-type-by-cell-type basis. Furthermore, single-cell sequencing also specifically replicated the depletion of BRN2 (POU3F2) and PTN in both Scz progenitors and neurons. Subsequently, in two mechanistic rescue experiments we identified that the transcription factor BRN2 and growth factor PTN operate as mechanistic substrates of neurogenesis and cellular survival, respectively, in Scz organoids. Collectively, our work suggests that multiple mechanisms of Scz exist in patient-derived organoids, and that these disparate mechanisms converge upon primordial brain developmental pathways such as neuronal differentiation, survival, and growth factor support, which may amalgamate to elevate intrinsic risk of Scz.
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Affiliation(s)
- Michael Notaras
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - Aiman Lodhi
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - Friederike Dündar
- Department of Physiology and Biophysics, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - Paul Collier
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - Nicole M Sayles
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - Hagen Tilgner
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA
| | - David Greening
- La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, Australia
- Central Clinical School, Monash University, Melbourne, VIC, Australia
- Baker Institute & Baker Department of Cardiometabolic Health, University of Melbourne, Melbourne, VIC, Australia
| | - Dilek Colak
- Center for Neurogenetics, Feil Family Brain and Mind Research Institute, Weill Cornell Medical College, Cornell University, New York, NY, USA.
- Gale and Ira Drukier Institute for Children's Health, Weill Cornell Medical College, Cornell University, New York, NY, USA.
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5
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Samoilova EM, Belopasov VV, Baklaushev VP. Transcription Factors of Direct Neuronal Reprogramming in Ontogenesis and Ex Vivo. Mol Biol 2021; 55:645-669. [DOI: 10.1134/s0026893321040087] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2020] [Revised: 12/14/2020] [Accepted: 12/15/2020] [Indexed: 03/07/2025]
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Abstract
Progressive aging is a physiological process that represents a central risk factor for the development of several human age-associated chronic diseases, including neurodegenerative diseases. A major focus in biomedical research is the pursuit for appropriate model systems to better model the biology of human aging and the interface between aging and disease mechanisms. Direct conversion of human fibroblasts into induced neurons (iNs) has emerged as a novel technology for the in vitro modeling of age-dependent neurological diseases. Similar to other cellular reprogramming techniques, e.g., iPSC-based cellular reprograming, direct conversion relies on the ectopic overexpression of transcription factors, typically including well-known pioneer factors. However, in contrast to alternative technologies to generate neurons, the entire process of direct conversion bypasses any proliferative or stem cell-like stage, which in fact renders it the unique aptitude of preserving age-associated hallmarks from the initial fibroblast source. In this chapter, we introduce direct conversion as a practical and easy-to-approach disease model for aging and neurodegenerative disease research. A focus here is to provide a stepwise protocol for the efficient and highly reproducible generation of iNs from adult dermal fibroblasts from human donors.
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7
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Vasan L, Park E, David LA, Fleming T, Schuurmans C. Direct Neuronal Reprogramming: Bridging the Gap Between Basic Science and Clinical Application. Front Cell Dev Biol 2021; 9:681087. [PMID: 34291049 PMCID: PMC8287587 DOI: 10.3389/fcell.2021.681087] [Citation(s) in RCA: 34] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/15/2021] [Accepted: 06/02/2021] [Indexed: 12/15/2022] Open
Abstract
Direct neuronal reprogramming is an innovative new technology that involves the conversion of somatic cells to induced neurons (iNs) without passing through a pluripotent state. The capacity to make new neurons in the brain, which previously was not achievable, has created great excitement in the field as it has opened the door for the potential treatment of incurable neurodegenerative diseases and brain injuries such as stroke. These neurological disorders are associated with frank neuronal loss, and as new neurons are not made in most of the adult brain, treatment options are limited. Developmental biologists have paved the way for the field of direct neuronal reprogramming by identifying both intrinsic cues, primarily transcription factors (TFs) and miRNAs, and extrinsic cues, including growth factors and other signaling molecules, that induce neurogenesis and specify neuronal subtype identities in the embryonic brain. The striking observation that postmitotic, terminally differentiated somatic cells can be converted to iNs by mis-expression of TFs or miRNAs involved in neural lineage development, and/or by exposure to growth factors or small molecule cocktails that recapitulate the signaling environment of the developing brain, has opened the door to the rapid expansion of new neuronal reprogramming methodologies. Furthermore, the more recent applications of neuronal lineage conversion strategies that target resident glial cells in situ has expanded the clinical potential of direct neuronal reprogramming techniques. Herein, we present an overview of the history, accomplishments, and therapeutic potential of direct neuronal reprogramming as revealed over the last two decades.
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Affiliation(s)
- Lakshmy Vasan
- Sunnybrook Research Institute, Biological Sciences Platform, Toronto, ON, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada
| | - Eunjee Park
- Sunnybrook Research Institute, Biological Sciences Platform, Toronto, ON, Canada
- Department of Biochemistry, University of Toronto, Toronto, ON, Canada
| | - Luke Ajay David
- Sunnybrook Research Institute, Biological Sciences Platform, Toronto, ON, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada
| | - Taylor Fleming
- Sunnybrook Research Institute, Biological Sciences Platform, Toronto, ON, Canada
| | - Carol Schuurmans
- Sunnybrook Research Institute, Biological Sciences Platform, Toronto, ON, Canada
- Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, ON, Canada
- Department of Biochemistry, University of Toronto, Toronto, ON, Canada
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8
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Sharif N, Calzolari F, Berninger B. Direct In Vitro Reprogramming of Astrocytes into Induced Neurons. Methods Mol Biol 2021; 2352:13-29. [PMID: 34324177 DOI: 10.1007/978-1-0716-1601-7_2] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/13/2023]
Abstract
Spontaneous neuronal replacement is almost absent in the postnatal mammalian nervous system. However, several studies have shown that both early postnatal and adult astroglia can be reprogrammed in vitro or in vivo by forced expression of proneural transcription factors, such as Neurogenin-2 or Achaete-scute homolog 1 (Ascl1), to acquire a neuronal fate. The reprogramming process stably induces properties such as distinctly neuronal morphology, expression of neuron-specific proteins, and the gain of mature neuronal functional features. Direct conversion of astroglia into neurons thus possesses potential as a basis for cell-based strategies against neurological diseases. In this chapter, we describe a well-established protocol used for direct reprogramming of postnatal cortical astrocytes into functional neurons in vitro and discuss available tools and approaches to dissect molecular and cell biological mechanisms underlying the reprogramming process.
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Affiliation(s)
- Nesrin Sharif
- Institute of Physiological Chemistry, University Medical Center Johannes Gutenberg University Mainz, Mainz, Germany
- International PhD Programme on Gene Regulation, Epigenetics and Genome Stability, Mainz, Germany
| | - Filippo Calzolari
- Institute of Physiological Chemistry, University Medical Center Johannes Gutenberg University Mainz, Mainz, Germany
| | - Benedikt Berninger
- Institute of Physiological Chemistry, University Medical Center Johannes Gutenberg University Mainz, Mainz, Germany.
- Institute of Psychiatry, Psychology, and Neuroscience, Centre for Developmental Neurobiology, King's College London, London, UK.
- MRC Centre for Neurodevelopmental Disorders, King's College London, London, UK.
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9
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Xu Z, Su S, Zhou S, Yang W, Deng X, Sun Y, Li L, Li Y. How to reprogram human fibroblasts to neurons. Cell Biosci 2020; 10:116. [PMID: 33062254 PMCID: PMC7549215 DOI: 10.1186/s13578-020-00476-2] [Citation(s) in RCA: 28] [Impact Index Per Article: 5.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/24/2020] [Accepted: 09/15/2020] [Indexed: 12/13/2022] Open
Abstract
Destruction and death of neurons can lead to neurodegenerative diseases. One possible way to treat neurodegenerative diseases and damage of the nervous system is replacing damaged and dead neurons by cell transplantation. If new neurons can replace the lost neurons, patients may be able to regain the lost functions of memory, motor, and so on. Therefore, acquiring neurons conveniently and efficiently is vital to treat neurological diseases. In recent years, studies on reprogramming human fibroblasts into neurons have emerged one after another, and this paper summarizes all these studies. Scientists find small molecules and transcription factors playing a crucial role in reprogramming and inducing neuron production. At the same time, both the physiological microenvironment in vivo and the physical and chemical factors in vitro play an essential role in the induction of neurons. Therefore, this paper summarized and analyzed these relevant factors. In addition, due to the unique advantages of physical factors in the process of reprogramming human fibroblasts into neurons, such as safe and minimally invasive, it has a more promising application prospect. Therefore, this paper also summarizes some successful physical mechanisms of utilizing fibroblasts to acquire neurons, which will provide new ideas for somatic cell reprogramming.
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Affiliation(s)
- Ziran Xu
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun, 130021 People's Republic of China
| | - Shengnan Su
- The Second Hospital of Jilin University, Jilin, Changchun, 130041 China
| | - Siyan Zhou
- Department of Stomatology, The First Hospital of Jilin University, Changchun, 130021 People's Republic of China
| | - Wentao Yang
- Norman Bethune College of Medicine, Jilin University, Changchun, 130021 People's Republic of China
| | - Xin Deng
- Norman Bethune College of Medicine, Jilin University, Changchun, 130021 People's Republic of China
| | - Yingying Sun
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun, 130021 People's Republic of China.,Department of Stomatology, The First Hospital of Jilin University, Changchun, 130021 People's Republic of China
| | - Lisha Li
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun, 130021 People's Republic of China
| | - Yulin Li
- The Key Laboratory of Pathobiology, Ministry of Education, College of Basic Medical Sciences, Jilin University, Changchun, 130021 People's Republic of China
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10
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Gordon A, Geschwind DH. Human in vitro models for understanding mechanisms of autism spectrum disorder. Mol Autism 2020; 11:26. [PMID: 32299488 PMCID: PMC7164291 DOI: 10.1186/s13229-020-00332-7] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/23/2019] [Accepted: 04/01/2020] [Indexed: 02/06/2023] Open
Abstract
Early brain development is a critical epoch for the development of autism spectrum disorder (ASD). In vivo animal models have, until recently, been the principal tool used to study early brain development and the changes occurring in neurodevelopmental disorders such as ASD. In vitro models of brain development represent a significant advance in the field. Here, we review the main methods available to study human brain development in vitro and the applications of these models for studying ASD and other psychiatric disorders. We discuss the main findings from stem cell models to date focusing on cell cycle and proliferation, cell death, cell differentiation and maturation, and neuronal signaling and synaptic stimuli. To be able to generalize the results from these studies, we propose a framework of experimental design and power considerations for using in vitro models to study ASD. These include both technical issues such as reproducibility and power analysis and conceptual issues such as the brain region and cell types being modeled.
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Affiliation(s)
- Aaron Gordon
- Department of Neurology, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA, USA
| | - Daniel H Geschwind
- Department of Neurology, David Geffen School of Medicine, University of California Los Angeles, Los Angeles, CA, USA.
- Program in Neurobehavioral Genetics, Semel Institute, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
- Center for Autism Research and Treatment, Semel Institute, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
- Department of Human Genetics, David Geffen School of Medicine, University of California, Los Angeles, CA, USA.
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11
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Traxler L, Edenhofer F, Mertens J. Next-generation disease modeling with direct conversion: a new path to old neurons. FEBS Lett 2019; 593:3316-3337. [PMID: 31715002 PMCID: PMC6907729 DOI: 10.1002/1873-3468.13678] [Citation(s) in RCA: 39] [Impact Index Per Article: 6.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/20/2019] [Revised: 10/20/2019] [Accepted: 11/07/2019] [Indexed: 12/13/2022]
Abstract
Within just over a decade, human reprogramming-based disease modeling has developed from a rather outlandish idea into an essential part of disease research. While iPSCs are a valuable tool for modeling developmental and monogenetic disorders, their rejuvenated identity poses limitations for modeling age-associated diseases. Direct cell-type conversion of fibroblasts into induced neurons (iNs) circumvents rejuvenation and preserves hallmarks of cellular aging. iNs are thus advantageous for modeling diseases that possess strong age-related and epigenetic contributions and can complement iPSC-based strategies for disease modeling. In this review, we provide an overview of the state of the art of direct iN conversion and describe the key epigenetic, transcriptomic, and metabolic changes that occur in converting fibroblasts. Furthermore, we summarize new insights into this fascinating process, particularly focusing on the rapidly changing criteria used to define and characterize in vitro-born human neurons. Finally, we discuss the unique features that distinguish iNs from other reprogramming-based neuronal cell models and how iNs are relevant to disease modeling.
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Affiliation(s)
- Larissa Traxler
- Department of GenomicsStem Cell Biology & Regenerative MedicineInstitute of Molecular Biology & CMBILeopold‐Franzens‐University InnsbruckInnsbruckAustria
- Laboratory of GeneticsThe Salk Institute for Biological StudiesLa JollaCAUSA
| | - Frank Edenhofer
- Department of GenomicsStem Cell Biology & Regenerative MedicineInstitute of Molecular Biology & CMBILeopold‐Franzens‐University InnsbruckInnsbruckAustria
| | - Jerome Mertens
- Department of GenomicsStem Cell Biology & Regenerative MedicineInstitute of Molecular Biology & CMBILeopold‐Franzens‐University InnsbruckInnsbruckAustria
- Laboratory of GeneticsThe Salk Institute for Biological StudiesLa JollaCAUSA
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12
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Flitsch LJ, Brüstle O. Evolving principles underlying neural lineage conversion and their relevance for biomedical translation. F1000Res 2019; 8. [PMID: 31559012 PMCID: PMC6743253 DOI: 10.12688/f1000research.18926.1] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Accepted: 08/23/2019] [Indexed: 12/19/2022] Open
Abstract
Scientific and technological advances of the past decade have shed light on the mechanisms underlying cell fate acquisition, including its transcriptional and epigenetic regulation during embryonic development. This knowledge has enabled us to purposefully engineer cell fates
in vitro by manipulating expression levels of lineage-instructing transcription factors. Here, we review the state of the art in the cell programming field with a focus on the derivation of neural cells. We reflect on what we know about the mechanisms underlying fate changes in general and on the degree of epigenetic remodeling conveyed by the distinct reprogramming and direct conversion strategies available. Moreover, we discuss the implications of residual epigenetic memory for biomedical applications such as disease modeling and neuroregeneration. Finally, we cover recent developments approaching cell fate conversion in the living brain and define questions which need to be addressed before cell programming can become an integral part of translational medicine.
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Affiliation(s)
- Lea Jessica Flitsch
- Institute of Reconstructive Neurobiology, University of Bonn School of Medicine & University Hospital Bonn, Bonn, North Rhine Wesphalia, 53127, Germany
| | - Oliver Brüstle
- Institute of Reconstructive Neurobiology, University of Bonn School of Medicine & University Hospital Bonn, Bonn, North Rhine Wesphalia, 53127, Germany
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13
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Rujanapun N, Heebkaew N, Promjantuek W, Sotthibundhu A, Kunhorm P, Chaicharoenaudomrung N, Noisa P. Small molecules re-establish neural cell fate of human fibroblasts via autophagy activation. In Vitro Cell Dev Biol Anim 2019; 55:622-632. [PMID: 31321620 DOI: 10.1007/s11626-019-00381-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/05/2019] [Accepted: 06/18/2019] [Indexed: 12/19/2022]
Abstract
The generation of neural cells is of great interest in medical research because of its promising in neurodegenerative diseases. Small chemical molecules have been used for inducing specific cell types across lineage boundaries. Therefore, to direct neural cell fate, small molecule is a feasible approach for generating clinically relevant cell types without genetic alterations. Human fibroblasts have been directly induced into neural cells with different combinations of small molecules; however, the mechanism underlying neural induction is still not fully understood. In this study, human fibroblasts were induced into neural cells by using only 4 small molecules in a short time period, 5 d. Small molecules used in this study included WNT activator, DNMT inhibitor, Notch inhibitor, and retinoic acid. Neural-specific genes, including NESTIN, TUJ1, and SOX2, were upregulated upon the induction for 5 d. Noteworthy, this neural induction process by small molecules coincided with the activation of autophagy. Autophagy-related genes, such as LC3, ATG12, and LAMP1, were enhanced upon neural induction, and the number of induced-neural cells decreased when autophagy was suppressed by chloroquine. The activation of autophagy was found to reduce ROS generation within the induced-neural cells, and the inhibition of autophagy by chloroquine suppressed the expression of antioxidant genes, CATALASE, SOD, and GPX. This implied that autophagy maintained the optimal level of ROS for neural induction of human fibroblasts. Altogether, this study presented the effective and convenient condition to induce neural cells from human fibroblasts and revealed the positive roles of autophagy in controlling neural cell induction.
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Affiliation(s)
- Narawadee Rujanapun
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
| | - Nudjanad Heebkaew
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
| | - Wilasinee Promjantuek
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
| | - Areechun Sotthibundhu
- Chulabhorn International College of Medicine, Thammasat University, Rungsit Campus, Rungsit, Patumthani, 12120, Thailand
| | - Phongsakorn Kunhorm
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
| | - Nipha Chaicharoenaudomrung
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand
| | - Parinya Noisa
- Laboratory of Cell-Based Assays and Innovations, School of Biotechnology, Institute of Agricultural Technology, Suranaree University of Technology, 111 University Avenue, Nakhon Ratchasima, 30000, Thailand.
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14
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Saul MC, Philip VM, Reinholdt LG, Chesler EJ. High-Diversity Mouse Populations for Complex Traits. Trends Genet 2019; 35:501-514. [PMID: 31133439 DOI: 10.1016/j.tig.2019.04.003] [Citation(s) in RCA: 121] [Impact Index Per Article: 20.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/19/2019] [Revised: 04/19/2019] [Accepted: 04/22/2019] [Indexed: 12/21/2022]
Abstract
Contemporary mouse genetic reference populations are a powerful platform to discover complex disease mechanisms. Advanced high-diversity mouse populations include the Collaborative Cross (CC) strains, Diversity Outbred (DO) stock, and their isogenic founder strains. When used in systems genetics and integrative genomics analyses, these populations efficiently harnesses known genetic variation for precise and contextualized identification of complex disease mechanisms. Extensive genetic, genomic, and phenotypic data are already available for these high-diversity mouse populations and a growing suite of data analysis tools have been developed to support research on diverse mice. This integrated resource can be used to discover and evaluate disease mechanisms relevant across species.
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Affiliation(s)
- Michael C Saul
- The Jackson Laboratory for Mammalian Genetics, Bar Harbor, ME, USA
| | - Vivek M Philip
- The Jackson Laboratory for Mammalian Genetics, Bar Harbor, ME, USA
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- The Jackson Laboratory for Mammalian Genetics, Bar Harbor, ME, USA; UNC Chapel Hill, Chapel Hill, NC, USA; SUNY Binghamton, Binghamton, NY, USA; Pittsburgh University, Pittsburgh, PA, USA
| | - Elissa J Chesler
- The Jackson Laboratory for Mammalian Genetics, Bar Harbor, ME, USA.
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15
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Pharmacological Transdifferentiation of Human Nasal Olfactory Stem Cells into Dopaminergic Neurons. Stem Cells Int 2019; 2019:2945435. [PMID: 31236114 PMCID: PMC6545791 DOI: 10.1155/2019/2945435] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/24/2018] [Accepted: 02/25/2019] [Indexed: 01/01/2023] Open
Abstract
The discovery of novel drugs for neurodegenerative diseases has been a real challenge over the last decades. The development of patient- and/or disease-specific in vitro models represents a powerful strategy for the development and validation of lead candidates in preclinical settings. The implementation of a reliable platform modeling dopaminergic neurons will be an asset in the study of dopamine-associated pathologies such as Parkinson's disease. Disease models based on cell reprogramming strategies, using either human-induced pluripotent stem cells or transcription factor-mediated transdifferentiation, are among the most investigated strategies. However, multipotent adult stem cells remain of high interest to devise direct conversion protocols and establish in vitro models that could bypass certain limitations associated with reprogramming strategies. Here, we report the development of a six-step chemically defined protocol that drives the transdifferentiation of human nasal olfactory stem cells into dopaminergic neurons. Morphological changes were progressively accompanied by modifications matching transcript and protein dopaminergic signatures such as LIM homeobox transcription factor 1 alpha (LMX1A), LMX1B, and tyrosine hydroxylase (TH) expression, within 42 days of differentiation. Phenotypic changes were confirmed by the production of dopamine from differentiated neurons. This new strategy paves the way to develop more disease-relevant models by establishing reprogramming-free patient-specific dopaminergic cell models for drug screening and/or target validation for neurodegenerative diseases.
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16
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Chang Y, Cho B, Kim S, Kim J. Direct conversion of fibroblasts to osteoblasts as a novel strategy for bone regeneration in elderly individuals. Exp Mol Med 2019; 51:1-8. [PMID: 31073120 PMCID: PMC6509166 DOI: 10.1038/s12276-019-0251-1] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/03/2018] [Revised: 12/24/2018] [Accepted: 01/28/2019] [Indexed: 12/31/2022] Open
Abstract
Mortality caused by age-related bone fractures or osteoporosis is steadily increasing worldwide as the population ages. The pace of the development of bone regeneration engineering to treat bone fractures has consequently increased in recent years. A range of techniques for bone regeneration, such as immunotherapy, allografts, and hydrogel therapy, have been devised. Cell-based therapies using bone marrow-derived mesenchymal stem cells and induced pluripotent stem cells derived from somatic cells are considered to be suitable approaches for bone repair. However, these cell-based therapies suffer from a number of limitations in terms of efficiency and safety. Somatic cells can also be directly differentiated into osteoblasts by several transcription factors. As osteoblasts play a central role in the process of bone formation, the direct reprogramming of fibroblasts into osteoblasts may hence be a new way to treat bone fractures in elderly individuals. Here, we review recent developments regarding the therapeutic potential of the direct reprogramming of cells for bone regeneration. Reprogramming cells that produce connective tissue to form bone instead could help prevent fractures in the elderly. Bones weaken with age, and fractures are a significant health risk in ageing populations. Most current bone regeneration treatments use stem cells, which can differentiate into any type of cell and have infinite capacity to divide; however, they are difficult to source and can lead to tumor formation. Jongpil Kim at Dongguk University in South Korea and coworkers have reviewed a new method that uses genetic signals to transform connective tissue-forming cells into bone-producing cells. The reprogrammed cells have been shown to generate new bone at the desired site, and because they have already lost their capacity for infinite division, tumor formation risk is greatly reduced. This method shows promise to expand treatment options for fractures and osteoporosis.
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Affiliation(s)
- Yujung Chang
- Department of Biomedical Engineering, Dongguk University, Pildong-ro 1-gil 30, Jung-gu, Seoul, 04620, Republic of Korea
| | - Byounggook Cho
- Department of Biomedical Engineering, Dongguk University, Pildong-ro 1-gil 30, Jung-gu, Seoul, 04620, Republic of Korea
| | - Siyoung Kim
- Department of Biomedical Engineering, Dongguk University, Pildong-ro 1-gil 30, Jung-gu, Seoul, 04620, Republic of Korea
| | - Jongpil Kim
- Department of Biomedical Engineering, Dongguk University, Pildong-ro 1-gil 30, Jung-gu, Seoul, 04620, Republic of Korea. .,Department of Chemistry, Dongguk University, 30, Pildong-ro 1-gil 30, Jung-gu, Seoul, 04620, Republic of Korea.
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17
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Mertens J, Reid D, Lau S, Kim Y, Gage FH. Aging in a Dish: iPSC-Derived and Directly Induced Neurons for Studying Brain Aging and Age-Related Neurodegenerative Diseases. Annu Rev Genet 2018; 52:271-293. [PMID: 30208291 DOI: 10.1146/annurev-genet-120417-031534] [Citation(s) in RCA: 189] [Impact Index Per Article: 27.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
Age-associated neurological diseases represent a profound challenge in biomedical research as we are still struggling to understand the interface between the aging process and the manifestation of disease. Various pathologies in the elderly do not directly result from genetic mutations, toxins, or infectious agents but are primarily driven by the many manifestations of biological aging. Therefore, the generation of appropriate model systems to study human aging in the nervous system demands new concepts that lie beyond transgenic and drug-induced models. Although access to viable human brain specimens is limited and induced pluripotent stem cell models face limitations due to reprogramming-associated cellular rejuvenation, the direct conversion of somatic cells into induced neurons allows for the generation of human neurons that capture many aspects of aging. Here, we review advances in exploring age-associated neurodegenerative diseases using human cell reprogramming models, and we discuss general concepts, promises, and limitations of the field.
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Affiliation(s)
- Jerome Mertens
- Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, California 92037, USA; .,Department of Genomics, Stem Cell Biology and Regenerative Medicine, Institute of Molecular Biology, and Center for Molecular Biosciences Innsbruck, University of Innsbruck, A-6020 Innsbruck, Austria
| | - Dylan Reid
- Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
| | - Shong Lau
- Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
| | - Yongsung Kim
- Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
| | - Fred H Gage
- Laboratory of Genetics, The Salk Institute for Biological Studies, La Jolla, California 92037, USA;
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18
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Loera-Valencia R, Piras A, Ismail MAM, Manchanda S, Eyjolfsdottir H, Saido TC, Johansson J, Eriksdotter M, Winblad B, Nilsson P. Targeting Alzheimer's disease with gene and cell therapies. J Intern Med 2018; 284:2-36. [PMID: 29582495 DOI: 10.1111/joim.12759] [Citation(s) in RCA: 41] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
Alzheimer's disease (AD) causes dementia in both young and old people affecting more than 40 million people worldwide. The two neuropathological hallmarks of the disease, amyloid beta (Aβ) plaques and neurofibrillary tangles consisting of protein tau are considered the major contributors to the disease. However, a more complete picture reveals significant neurodegeneration and decreased cell survival, neuroinflammation, changes in protein and energy homeostasis and alterations in lipid and cholesterol metabolism. In addition, gene and cell therapies for severe neurodegenerative disorders have recently improved technically in terms of safety and efficiency and have translated to the clinic showing encouraging results. Here, we review broadly current data within the field for potential targets that could modify AD through gene and cell therapy strategies. We envision that not only Aβ will be targeted in a disease-modifying treatment strategy but rather that a combination of treatments, possibly at different intervention times may prove beneficial in curing this devastating disease. These include decreased tau pathology, neuronal growth factors to support neurons and modulation of neuroinflammation for an appropriate immune response. Furthermore, cell based therapies may represent potential strategies in the future.
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Affiliation(s)
- R Loera-Valencia
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden
| | - A Piras
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden
| | - M A M Ismail
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden.,Theme Neuro, Diseases of the Nervous System Patient Flow, Karolinska University Hospital, Huddinge, Sweden
| | - S Manchanda
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden
| | - H Eyjolfsdottir
- Division of Clinical Geriatrics, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Huddinge, Sweden.,Theme Aging, Karolinska University Hospital, Huddinge, Sweden
| | - T C Saido
- RIKEN Brain Science Institute, Wako, Saitama, Japan
| | - J Johansson
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden
| | - M Eriksdotter
- Division of Clinical Geriatrics, Department of Neurobiology, Care Sciences and Society, Karolinska Institutet, Huddinge, Sweden.,Theme Aging, Karolinska University Hospital, Huddinge, Sweden
| | - B Winblad
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden.,Theme Aging, Karolinska University Hospital, Huddinge, Sweden
| | - P Nilsson
- Division of Neurogeriatrics, Department of Neurobiology, Care Sciences and Society, Center for Alzheimer Research, Karolinska Institutet, Solna, Sweden
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19
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Dhungel B, Ramlogan-Steel CA, Steel JC. MicroRNA-Regulated Gene Delivery Systems for Research and Therapeutic Purposes. Molecules 2018; 23:E1500. [PMID: 29933586 PMCID: PMC6099389 DOI: 10.3390/molecules23071500] [Citation(s) in RCA: 22] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/30/2018] [Revised: 06/18/2018] [Accepted: 06/20/2018] [Indexed: 12/18/2022] Open
Abstract
Targeted gene delivery relies on the ability to limit the expression of a transgene within a defined cell/tissue population. MicroRNAs represent a class of highly powerful and effective regulators of gene expression that act by binding to a specific sequence present in the corresponding messenger RNA. Involved in almost every aspect of cellular function, many miRNAs have been discovered with expression patterns specific to developmental stage, lineage, cell-type, or disease stage. Exploiting the binding sites of these miRNAs allows for construction of targeted gene delivery platforms with a diverse range of applications. Here, we summarize studies that have utilized miRNA-regulated systems to achieve targeted gene delivery for both research and therapeutic purposes. Additionally, we identify criteria that are important for the effectiveness of a particular miRNA for such applications and we also discuss factors that have to be taken into consideration when designing miRNA-regulated expression cassettes.
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Affiliation(s)
- Bijay Dhungel
- Gallipoli Medical Research Institute, Greenslopes Private Hospital, 102 Newdegate Street, Brisbane, QLD 4120, Australia.
- Faculty of Medicine, University of Queensland, 288 Herston Road, Herston, Brisbane, QLD 4006, Australia.
- University of Queensland Diamantina Institute, Translational Research Institute, 37 Kent Street, Woolloongabba, QLD 4102, Australia.
| | - Charmaine A Ramlogan-Steel
- Faculty of Medicine, University of Queensland, 288 Herston Road, Herston, Brisbane, QLD 4006, Australia.
- Layton Vision Foundation, Translational Research Institute, 37 Kent Street, Woolloongabba, QLD 4102, Australia.
| | - Jason C Steel
- Faculty of Medicine, University of Queensland, 288 Herston Road, Herston, Brisbane, QLD 4006, Australia.
- OcuGene, Translational Research Institute, 37 Kent Street, Woolloongabba, QLD 4102, Australia.
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20
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Ghaffari LT, Starr A, Nelson AT, Sattler R. Representing Diversity in the Dish: Using Patient-Derived in Vitro Models to Recreate the Heterogeneity of Neurological Disease. Front Neurosci 2018; 12:56. [PMID: 29479303 PMCID: PMC5812426 DOI: 10.3389/fnins.2018.00056] [Citation(s) in RCA: 28] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/13/2017] [Accepted: 01/23/2018] [Indexed: 12/14/2022] Open
Abstract
Neurological diseases, including dementias such as Alzheimer's disease (AD) and fronto-temporal dementia (FTD) and degenerative motor neuron diseases such as amyotrophic lateral sclerosis (ALS), are responsible for an increasing fraction of worldwide fatalities. Researching these heterogeneous diseases requires models that endogenously express the full array of genetic and epigenetic factors which may influence disease development in both familial and sporadic patients. Here, we discuss the two primary methods of developing patient-derived neurons and glia to model neurodegenerative disease: reprogramming somatic cells into induced pluripotent stem cells (iPSCs), which are differentiated into neurons or glial cells, or directly converting (DC) somatic cells into neurons (iNeurons) or glial cells. Distinct differentiation techniques for both models result in a variety of neuronal and glial cell types, which have been successful in displaying unique hallmarks of a variety of neurological diseases. Yield, length of differentiation, ease of genetic manipulation, expression of cell-specific markers, and recapitulation of disease pathogenesis are presented as determining factors in how these methods may be used separately or together to ascertain mechanisms of disease and identify therapeutics for distinct patient populations or for specific individuals in personalized medicine projects.
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Affiliation(s)
- Layla T Ghaffari
- Department of Neurobiology, Barrow Neurological Institute, Dignity Health-St. Joseph's Hospital and Medical Center, Phoenix, AZ, United States
| | - Alexander Starr
- Department of Neurobiology, Barrow Neurological Institute, Dignity Health-St. Joseph's Hospital and Medical Center, Phoenix, AZ, United States
| | - Andrew T Nelson
- Department of Neurobiology, Barrow Neurological Institute, Dignity Health-St. Joseph's Hospital and Medical Center, Phoenix, AZ, United States
| | - Rita Sattler
- Department of Neurobiology, Barrow Neurological Institute, Dignity Health-St. Joseph's Hospital and Medical Center, Phoenix, AZ, United States
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21
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Shrigley S, Pircs K, Barker RA, Parmar M, Drouin-Ouellet J. Simple Generation of a High Yield Culture of Induced Neurons from Human Adult Skin Fibroblasts. J Vis Exp 2018. [PMID: 29443113 PMCID: PMC5912349 DOI: 10.3791/56904] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/12/2023] Open
Abstract
Induced neurons (iNs), the product of somatic cells directly converted to neurons, are a way to obtain patient-derived neurons from tissue that is easily accessible. Through this route, mature neurons can be obtained in a matter of a few weeks. Here, we describe a straightforward and rapid one-step protocol to obtain iNs from dermal fibroblasts obtained through biopsy samples from adult human donors. We explain each step of the process, including the maintenance of the dermal fibroblasts, the freezing procedure to build a stock of the cell line, seeding of the cells for reprogramming, as well as the culture conditions during the conversion process. In addition, we describe the preparation of glass coverslips for electrophysiological recordings, long-term coating conditions, and fluorescence activated cell sorting (FACS). We also illustrate examples of the results to be expected. The protocol described here is easy to perform and can be applied to human fibroblasts derived from human skin biopsies from patients with various different diagnoses and ages. This protocol generates a sufficient amount of iNs which can be used for a wide array of biomedical applications, including disease modeling, drug screening, and target validation.
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Affiliation(s)
- Shelby Shrigley
- Department of Experimental Medical Science, Wallenberg Neuroscience Center, Division of Neurobiology and Lund Stem Cell Center, Lund University
| | - Karolina Pircs
- Department of Experimental Medical Science, Wallenberg Neuroscience Center, Division of Neurobiology and Lund Stem Cell Center, Lund University
| | - Roger A Barker
- Department of Experimental Medical Science, Wallenberg Neuroscience Center, Division of Neurobiology and Lund Stem Cell Center, Lund University; John van Geest Centre for Brain Repair & Department of Neurology, Department of Clinical Neurosciences and Cambridge Stem Cell Institute, University of Cambridge
| | - Malin Parmar
- Department of Experimental Medical Science, Wallenberg Neuroscience Center, Division of Neurobiology and Lund Stem Cell Center, Lund University
| | - Janelle Drouin-Ouellet
- Department of Experimental Medical Science, Wallenberg Neuroscience Center, Division of Neurobiology and Lund Stem Cell Center, Lund University;
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22
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Direct conversion from skin fibroblasts to functional dopaminergic neurons for biomedical application. BIOMEDICAL DERMATOLOGY 2017. [DOI: 10.1186/s41702-017-0004-5] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
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23
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MicroRNA-Directed Neuronal Reprogramming as a Therapeutic Strategy for Neurological Diseases. Mol Neurobiol 2017; 55:4428-4436. [PMID: 28664454 DOI: 10.1007/s12035-017-0671-7] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2017] [Accepted: 06/20/2017] [Indexed: 01/22/2023]
Abstract
The loss of neurons due to injury and disease results in a wide spectrum of highly disabling neurological and neurodegenerative conditions, given the apparent limited capacity of endogenous repair of the adult central nervous system (CNS). Therefore, it is important to develop technologies that can promote de novo neural stem cell and neuron generation. Current insights in CNS development and cellular reprogramming have provided the knowledge to finely modulate lineage-restricted transcription factors and microRNAs (miRNA) to elicit correct neurogenesis. Here, we discuss the current knowledge on the direct reprogramming of somatic non-neuronal cells into neural stem cells or subtype specific neurons in vitro and in vivo focusing on miRNA driven reprogramming. miRNA can allow rapid and efficient direct phenotype conversion by modulating gene networks active during development, which promote global shifts in the epigenetic landscape pivoting cell fate decisions. Furthermore, we critically present state-of-the-art and recent advances on miRNA therapeutics that can be applied to the diseased CNS. Together, the advances in our understanding of miRNA role in CNS development and disease, recent progress in miRNA-based therapeutic strategies, and innovative drug delivery methods create novel perspectives for meaningful therapies for neurodegenerative disorders.
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24
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Bahmad H, Hadadeh O, Chamaa F, Cheaito K, Darwish B, Makkawi AK, Abou-Kheir W. Modeling Human Neurological and Neurodegenerative Diseases: From Induced Pluripotent Stem Cells to Neuronal Differentiation and Its Applications in Neurotrauma. Front Mol Neurosci 2017; 10:50. [PMID: 28293168 PMCID: PMC5329035 DOI: 10.3389/fnmol.2017.00050] [Citation(s) in RCA: 44] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2016] [Accepted: 02/13/2017] [Indexed: 12/14/2022] Open
Abstract
With the help of several inducing factors, somatic cells can be reprogrammed to become induced pluripotent stem cell (iPSCs) lines. The success is in obtaining iPSCs almost identical to embryonic stem cells (ESCs), therefore various approaches have been tested and ultimately several ones have succeeded. The importance of these cells is in how they serve as models to unveil the molecular pathways and mechanisms underlying several human diseases, and also in its potential roles in the development of regenerative medicine. They further aid in the development of regenerative medicine, autologous cell therapy and drug or toxicity screening. Here, we provide a comprehensive overview of the recent development in the field of iPSCs research, specifically for modeling human neurological and neurodegenerative diseases, and its applications in neurotrauma. These are mainly characterized by progressive functional or structural neuronal loss rendering them extremely challenging to manage. Many of these diseases, including Parkinson's disease (PD), Huntington's disease (HD), Amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD) have been explored in vitro. The main purpose is to generate patient-specific iPS cell lines from the somatic cells that carry mutations or genetic instabilities for the aim of studying their differentiation potential and behavior. This new technology will pave the way for future development in the field of stem cell research anticipating its use in clinical settings and in regenerative medicine in order to treat various human diseases, including neurological and neurodegenerative diseases.
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Affiliation(s)
| | | | | | | | | | | | - Wassim Abou-Kheir
- Department of Anatomy, Cell Biology and Physiological Sciences, Faculty of Medicine, American University of BeirutBeirut, Lebanon
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25
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Abstract
Limited access to human neurons has posed a significant barrier to progress in biological and preclinical studies of the human nervous system. The advent of cell reprogramming technologies has widely disclosed unprecedented opportunities to generate renewable sources of human neural cells for disease modeling, drug discovery, and cell therapeutics. Both somatic reprogramming into induced pluripotent stem cells (iPSCs) and directly induced Neurons (iNeurons) rely on transcription factor-based cellular conversion processes. Nevertheless, they rely on very distinct mechanisms, biological barriers, technical limitations, different levels of efficiency, and generate neural cells with distinctive properties. Human iPSCs represent a long-term renewable source of neural cells, but over time genomic aberrations might erode the quality of the cultures and the in vitro differentiation process requires extensive time. Conversely, direct neuronal reprogramming ensures a fast and straightforward generation of iNeurons endowed with functional properties. However, in this last case, conversion efficiency is reduced when starting from adult human cells, and the molecular and functional fidelity of iNeurons with respect to their corresponding native neuronal subtype is yet to be fully ascertained in many cases. For any biomedical research application, it should be carefully pondered the reprogramming method to use for generating reprogrammed human neuronal subtypes that best fit with the following analysis considering the existing limitations and gap of knowledge still present in this young field of investigation.
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Affiliation(s)
- Vania Broccoli
- San Raffaele Scientific Institute, Milan, Italy; CNR-Institute of Neuroscience, Milan, Italy.
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26
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Xu Z, Chu X, Jiang H, Schilling H, Chen S, Feng J. Induced dopaminergic neurons: A new promise for Parkinson's disease. Redox Biol 2017; 11:606-612. [PMID: 28110217 PMCID: PMC5256671 DOI: 10.1016/j.redox.2017.01.009] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2017] [Revised: 01/06/2017] [Accepted: 01/11/2017] [Indexed: 12/28/2022] Open
Abstract
Motor symptoms that define Parkinson’s disease (PD) are caused by the selective loss of nigral dopaminergic (DA) neurons. Cell replacement therapy for PD has been focused on midbrain DA neurons derived from human fetal mesencephalic tissue, human embryonic stem cells (hESC) or human induced pluripotent stem cells (iPSC). Recent development in the direct conversion of human fibroblasts to induced dopaminergic (iDA) neurons offers new opportunities for transplantation study and disease modeling in PD. The iDA neurons are generated directly from human fibroblasts in a short period of time, bypassing lengthy differentiation process from human pluripotent stem cells and the concern for potentially tumorigenic mitotic cells. They exhibit functional dopaminergic neurotransmission and relieve locomotor symptoms in animal models of Parkinson’s disease. In this review, we will discuss this recent development and its implications to Parkinson’s disease research and therapy.
Fibroblasts can be directly converted to induced dopaminergic neurons by transcription factors. Many different types of cells can be converted to induced neurons in vitro and in vivo. Appropriate cell culture conditions enhance the direct conversion to induced neurons. The conversion to induced neurons is enhanced by G1 arrest and p53 attenuation. iDA neurons is a promising tool for PD research and therapy.
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Affiliation(s)
- Zhimin Xu
- Veterans Affairs Western New York Healthcare System, Buffalo, NY 14215, USA; Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, NY 14214, USA; Laboratory of Neurodegenerative Diseases, Institute of Health Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Science and Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Xingkun Chu
- Laboratory of Neurodegenerative Diseases, Institute of Health Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Science and Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Houbo Jiang
- Veterans Affairs Western New York Healthcare System, Buffalo, NY 14215, USA; Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, NY 14214, USA
| | - Haley Schilling
- Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, NY 14214, USA
| | - Shengdi Chen
- Laboratory of Neurodegenerative Diseases, Institute of Health Science, Shanghai Institutes for Biological Sciences, Chinese Academy of Science and Shanghai Jiao Tong University School of Medicine, Shanghai 200025, China
| | - Jian Feng
- Veterans Affairs Western New York Healthcare System, Buffalo, NY 14215, USA; Department of Physiology and Biophysics, State University of New York at Buffalo, Buffalo, NY 14214, USA.
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27
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Abstract
Over the past three decades, significant progress has been made in the development of potential regenerative cell-based therapies for neurodegenerative disease, with most success being seen in Parkinson's disease. Cell-based therapies face many challenges including ethical considerations, potential for immune-mediated rejection with allogeneic and xenogeneic tissue, pathological spread of protein-related disease into the grafted tissue as well as the risk of graft overgrowth and tumorigenesis in stem cell-derived transplants. Preclinical trials have looked at many tissue types of which the most successful to date have been those using fetal ventral mesencephalon grafts, which led to clinical trials, which have shown that in some cases they can work very well. With important proof-of-concept derived from these studies, there is now much interest in how dopaminergic neurons derived from stem cell sources could be used to develop cell-based therapies suitable for clinical use, with clinical trials poised to enter the clinic in the next couple of years.
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Affiliation(s)
- Thomas B Stoker
- John van Geest Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge, Forvie Site, Cambridge, CB2 0PY, UK.,Wellcome Trust - Medical Research Council Stem Cell Institute, University of Cambridge
| | - Roger A Barker
- John van Geest Centre for Brain Repair, Department of Clinical Neurosciences, University of Cambridge, Forvie Site, Cambridge, CB2 0PY, UK.,Wellcome Trust - Medical Research Council Stem Cell Institute, University of Cambridge
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28
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Geisler A, Fechner H. MicroRNA-regulated viral vectors for gene therapy. World J Exp Med 2016; 6:37-54. [PMID: 27226955 PMCID: PMC4873559 DOI: 10.5493/wjem.v6.i2.37] [Citation(s) in RCA: 60] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/27/2015] [Revised: 03/02/2016] [Accepted: 03/17/2016] [Indexed: 02/06/2023] Open
Abstract
Safe and effective gene therapy approaches require targeted tissue-specific transfer of a therapeutic transgene. Besides traditional approaches, such as transcriptional and transductional targeting, microRNA-dependent post-transcriptional suppression of transgene expression has been emerging as powerful new technology to increase the specificity of vector-mediated transgene expression. MicroRNAs are small non-coding RNAs and often expressed in a tissue-, lineage-, activation- or differentiation-specific pattern. They typically regulate gene expression by binding to imperfectly complementary sequences in the 3' untranslated region (UTR) of the mRNA. To control exogenous transgene expression, tandem repeats of artificial microRNA target sites are usually incorporated into the 3' UTR of the transgene expression cassette, leading to subsequent degradation of transgene mRNA in cells expressing the corresponding microRNA. This targeting strategy, first shown for lentiviral vectors in antigen presenting cells, has now been used for tissue-specific expression of vector-encoded therapeutic transgenes, to reduce immune response against the transgene, to control virus tropism for oncolytic virotherapy, to increase safety of live attenuated virus vaccines and to identify and select cell subsets for pluripotent stem cell therapies, respectively. This review provides an introduction into the technical mechanism underlying microRNA-regulation, highlights new developments in this field and gives an overview of applications of microRNA-regulated viral vectors for cardiac, suicide gene cancer and hematopoietic stem cell therapy, as well as for treatment of neurological and eye diseases.
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Evaluating cell reprogramming, differentiation and conversion technologies in neuroscience. Nat Rev Neurosci 2016; 17:424-37. [PMID: 27194476 DOI: 10.1038/nrn.2016.46] [Citation(s) in RCA: 214] [Impact Index Per Article: 23.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
The scarcity of live human brain cells for experimental access has for a long time limited our ability to study complex human neurological disorders and elucidate basic neuroscientific mechanisms. A decade ago, the development of methods to reprogramme somatic human cells into induced pluripotent stem cells enabled the in vitro generation of a wide range of neural cells from virtually any human individual. The growth of methods to generate more robust and defined neural cell types through reprogramming and direct conversion into induced neurons has led to the establishment of various human reprogramming-based neural disease models.
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Shi Z, Zhang J, Chen S, Li Y, Lei X, Qiao H, Zhu Q, Hu B, Zhou Q, Jiao J. Conversion of Fibroblasts to Parvalbumin Neurons by One Transcription Factor, Ascl1, and the Chemical Compound Forskolin. J Biol Chem 2016; 291:13560-70. [PMID: 27137935 DOI: 10.1074/jbc.m115.709808] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2015] [Indexed: 12/31/2022] Open
Abstract
Abnormalities in parvalbumin (PV)-expressing interneurons cause neurodevelopmental disorders such as epilepsy, autism, and schizophrenia. Unlike other types of neurons that can be efficiently differentiated from pluripotent stem cells, PV neurons were minimally generated using a conventional differentiation strategy. In this study we developed an adenovirus-based transdifferentiation strategy that incorporates an additional chemical compound for the efficient generation of induced PV (iPV) neurons. The chemical compound forskolin combined with Ascl1 induced ∼80% of mouse fibroblasts to iPV neurons. The iPV neurons generated by this procedure matured 5-7 days post infection and were characterized by electrophysiological properties and known neuronal markers, such as PV and GABA. Our studies, therefore, identified an efficient approach for generating PV neurons.
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Affiliation(s)
- Zixiao Shi
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Juan Zhang
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Shuangquan Chen
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Yanxin Li
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Xuepei Lei
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Huimin Qiao
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China, University of Chinese Academy of Sciences, Beijing 100049, China, and
| | - Qianwen Zhu
- Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China
| | - Baoyang Hu
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China
| | - Qi Zhou
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China
| | - Jianwei Jiao
- From the State Key Laboratory of Stem Cells and Reproductive Biology, Institute of Zoology, Chinese Academy of Sciences, Beijing 100101, China,
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31
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Hou S, Lu P. Direct reprogramming of somatic cells into neural stem cells or neurons for neurological disorders. Neural Regen Res 2016; 11:28-31. [PMID: 26981072 PMCID: PMC4774217 DOI: 10.4103/1673-5374.169602] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023] Open
Abstract
Direct reprogramming of somatic cells into neurons or neural stem cells is one of the most important frontier fields in current neuroscience research. Without undergoing the pluripotency stage, induced neurons or induced neural stem cells are a safer and timelier manner resource in comparison to those derived from induced pluripotent stem cells. In this prospective, we review the recent advances in generation of induced neurons and induced neural stem cells in vitro and in vivo and their potential treatments of neurological disorders.
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Affiliation(s)
- Shaoping Hou
- Spinal Cord Research Center, Department of Neurobiology & Anatomy, Drexel University College of Medicine, Philadelphia, PA, USA
| | - Paul Lu
- Veterans Administration Medical Center, San Diego, CA, USA; Department of Neurosciences, University of California at San Diego, La Jolla, CA, USA
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Gopalakrishnan S, Hor P, Ichida JK. New approaches for direct conversion of patient fibroblasts into neural cells. Brain Res 2015; 1656:2-13. [PMID: 26475975 DOI: 10.1016/j.brainres.2015.10.012] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2015] [Revised: 09/21/2015] [Accepted: 10/05/2015] [Indexed: 01/05/2023]
Abstract
Recent landmark studies have demonstrated the production of disease-relevant human cell types by two different methods; differentiation of stem cells using external morphogens or lineage conversion using genetic factors. Directed differentiation changes embryonic stem cells (ESCs) or induced pluripotent stem cells (iPSCs) into a desired cell type by providing developmental cues in an in vitro environment. Direct reprogramming is achieved by the introduction of exogenous lineage specific transcription factors to convert any somatic cell type into another, thereby bypassing an intermediate pluripotent stage. A variety of somatic cell types such as blood, keratinocytes and fibroblasts can be used to derive iPSC cells. However, the process is time consuming,laborious, expensive and gives rise to cells with reported epigenetic heterogeneity even amongst different iPSC lines from same patient which could propagate phenotypic variability. A major concern with the use of pluripotent cells as starting material for cell replacement therapy is their incomplete differentiation and their propensity to form tumors following transplantation. In comparison, transcription factor mediated reprogramming offers a direct route to target cell types. This could allow for rapid comparison of large cohorts of patient and control samples at a given time for disease modeling. Additionally, transcription factors that drive maturation may yield more functionally mature cells than directed differentiation. Several studies have demonstrated the feasibility of generating of cell types such as cardiomyocytes, hepatocytes, and neurons from fibroblasts. Here, we will discuss recent advances and key challenges regarding direct reprogramming of somatic cell types into diverse neural cells. This article is part of a Special Issue entitled SI: Exploiting human neurons.
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Affiliation(s)
- Suhasni Gopalakrishnan
- University of Southern California, Department of Stem Cells and Regenerative Medicine, Eli and Edythe Broad, CIRM Center for Regenerative Medicine and Stem Cell Research at USC, Los Angeles, CA 90033, USA
| | - Pooja Hor
- University of Southern California, Department of Stem Cells and Regenerative Medicine, Eli and Edythe Broad, CIRM Center for Regenerative Medicine and Stem Cell Research at USC, Los Angeles, CA 90033, USA
| | - Justin K Ichida
- University of Southern California, Department of Stem Cells and Regenerative Medicine, Eli and Edythe Broad, CIRM Center for Regenerative Medicine and Stem Cell Research at USC, Los Angeles, CA 90033, USA
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