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Xia Q, Chang KC, Sun Y, Nahmou M, Noro T, Cheng Y, Kong X, Mo X, Goldberg JL, Wu S. Retinal ganglion cells induce stem cell-derived neuroprotection via IL-12 to SCGF-β crosstalk. Stem Cell Res Ther 2025; 16:90. [PMID: 40001251 PMCID: PMC11863831 DOI: 10.1186/s13287-025-04198-5] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2024] [Accepted: 01/30/2025] [Indexed: 02/27/2025] Open
Abstract
BACKGROUND Stem cell-derived secreted factors could protect neurons in neurodegenerative disease or after injury. The exact neuroprotective components in the secretome remain challenging to discover. Here we developed a cell-to-cell interaction model to identify a retinal ganglion cell (RGC)-protective factor derived from induced pluripotent stem cells (iPSCs). METHODS Primary RGCs were co-cultured with iPSCs or treated with iPSC-conditioned media in vitro. Cell viability were assayed using live-cell staining, and culture supernatant were analyzed via multiplexed antibody-based assays and ELISA. In vivo tests were carried out under mouse optic nerve crush model and RGC transplantation study in rats. Paired t-tests were used for data analysis between two groups. RESULTS RGC viability was significantly enhanced when iPSCs were first stimulated with RGC-derived supernatant before iPSC-conditioned medium was collected and added into RGC culture. A significant increase of stem cell growth factor-beta (SCGF-β) concentration was detected in the latter conditioned medium. SCGF-β enhanced RGC survival in vitro and in vivo, and RGC-derived interleukin-12(p70) (IL-12[p70]) promotes secretion of iPSC-derived SCGF-β. Downstream of this IL-12(p70)-to-SCGF-β axis, ngn2 was significantly upregulated, and was found both necessary and sufficient for RGC survival. CONCLUSION This study addresses a longstanding question of how neurons and stem cells interact to promote neuroprotection, and define a novel molecular interaction pathway whereby RGC's secretion of IL-12(p70) enhances iPSCs' secretion of SCGF-β, and SCGF-β protects RGCs via upregulating ngn2, suggesting that neurons may call on stem cells for their own protection.
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Affiliation(s)
- Qing Xia
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China
| | - Kun-Che Chang
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA
- Department of Ophthalmology and Neurobiology, School of Medicine, University of Pittsburgh, Pittsburgh, PA, 15213, USA
| | - Yanan Sun
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China
| | - Michael Nahmou
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA
| | - Takahiko Noro
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA
- Department of Ophthalmology, Jikei University School of Medicine, Tokyo, 105-8461, Japan
| | - Yun Cheng
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China
| | - Xiangmei Kong
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China
| | - Xiaofen Mo
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China.
| | - Jeffrey L Goldberg
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA.
| | - Suqian Wu
- Shanghai Key Laboratory of Visual Impairment and Restoration, Department of Ophthalmology and Vision Science, Eye & ENT Hospital, Fudan University, Shanghai, 200031, China.
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA.
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Hammer J, Röppenack P, Yousuf S, Machate A, Fischer M, Hans S, Brand M. Blind But Alive - Congenital Loss of atoh7 Disrupts the Visual System of Adult Zebrafish. Invest Ophthalmol Vis Sci 2024; 65:42. [PMID: 39565303 PMCID: PMC11583992 DOI: 10.1167/iovs.65.13.42] [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: 04/29/2024] [Accepted: 09/30/2024] [Indexed: 11/21/2024] Open
Abstract
Purpose Vision is the predominant sense in most animal species. Loss of vision can be caused by a multitude of factors resulting in anatomic as well as behavioral changes. In mice and zebrafish, atoh7 mutants are completely blind as they fail to generate retinal ganglion cells (RGCs) during development. In contrast to mice, raising blind zebrafish to adulthood is challenging and this important model is currently missing in the field. Here, we report the phenotype of homozygous mutant adult zebrafish atoh7 mutants that have been raised using adjusted feeding and holding conditions. Methods The phenotype of adult mutants was characterized using classical histology and immunohistochemistry as well as optical coherence tomography. In addition, the optokinetic response was characterized. Results Adult atoh7 mutants display dark body pigmentation and significantly reduced body length. They fail to form RGCs, the resulting nerve fiber layer as well as the optic nerve, and consequently behave completely blindly. In contrast, increased amounts of other retinal neurons and Müller glia are formed. In addition, the optic tectum is anatomically reduced in size, presumably due to the missing retinal input. Conclusions Taken together, we provide a comprehensive characterization of a completely blind adult zebrafish mutant with focus on retinal and tectal morphology, as a useful model for glaucoma and optic nerve aplasia.
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Affiliation(s)
- Juliane Hammer
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Paul Röppenack
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Sarah Yousuf
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Anja Machate
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Marika Fischer
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Stefan Hans
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
| | - Michael Brand
- Center for Regenerative Therapies at TU Dresden(CRTD), Dresden, Germany
- Cluster of Excellence Physics of Life (PoL), TU Dresden, Dresden, Germany
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Jui J, Goldman D. Müller Glial Cell-Dependent Regeneration of the Retina in Zebrafish and Mice. Annu Rev Genet 2024; 58:67-90. [PMID: 38876121 DOI: 10.1146/annurev-genet-111523-102000] [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] [Indexed: 06/16/2024]
Abstract
Sight is one of our most precious senses. People fear losing their sight more than any other disability. Thus, restoring sight to the blind is an important goal of vision scientists. Proregenerative species, such as zebrafish, provide a system for studying endogenous mechanisms underlying retina regeneration. Nonregenerative species, such as mice, provide a system for testing strategies for stimulating retina regeneration. Key to retina regeneration in zebrafish and mice is the Müller glial cell, a malleable cell type that is amenable to a variety of regenerative strategies. Here, we review cellular and molecular mechanisms used by zebrafish to regenerate a retina, as well as the application of these mechanisms, and other strategies to stimulate retina regeneration in mice. Although our focus is on Müller glia (MG), niche components and their impact on MG reprogramming are also discussed.
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Affiliation(s)
- Jonathan Jui
- Molecular Neuroscience Institute and Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan, USA; ,
| | - Daniel Goldman
- Molecular Neuroscience Institute and Department of Biological Chemistry, University of Michigan, Ann Arbor, Michigan, USA; ,
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Liang S, Zhou J, Yu X, Lu S, Liu R. Neuronal conversion from glia to replenish the lost neurons. Neural Regen Res 2024; 19:1446-1453. [PMID: 38051886 PMCID: PMC10883502 DOI: 10.4103/1673-5374.386400] [Citation(s) in RCA: 1] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2023] [Accepted: 08/16/2023] [Indexed: 12/07/2023] Open
Abstract
ABSTRACT Neuronal injury, aging, and cerebrovascular and neurodegenerative diseases such as cerebral infarction, Alzheimer's disease, Parkinson's disease, frontotemporal dementia, amyotrophic lateral sclerosis, and Huntington's disease are characterized by significant neuronal loss. Unfortunately, the neurons of most mammals including humans do not possess the ability to self-regenerate. Replenishment of lost neurons becomes an appealing therapeutic strategy to reverse the disease phenotype. Transplantation of pluripotent neural stem cells can supplement the missing neurons in the brain, but it carries the risk of causing gene mutation, tumorigenesis, severe inflammation, and obstructive hydrocephalus induced by brain edema. Conversion of neural or non-neural lineage cells into functional neurons is a promising strategy for the diseases involving neuron loss, which may overcome the above-mentioned disadvantages of neural stem cell therapy. Thus far, many strategies to transform astrocytes, fibroblasts, microglia, Müller glia, NG2 cells, and other glial cells to mature and functional neurons, or for the conversion between neuronal subtypes have been developed through the regulation of transcription factors, polypyrimidine tract binding protein 1 (PTBP1), and small chemical molecules or are based on a combination of several factors and the location in the central nervous system. However, some recent papers did not obtain expected results, and discrepancies exist. Therefore, in this review, we discuss the history of neuronal transdifferentiation, summarize the strategies for neuronal replenishment and conversion from glia, especially astrocytes, and point out that biosafety, new strategies, and the accurate origin of the truly converted neurons in vivo should be focused upon in future studies. It also arises the attention of replenishing the lost neurons from glia by gene therapies such as up-regulation of some transcription factors or down-regulation of PTBP1 or drug interference therapies.
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Affiliation(s)
- Shiyu Liang
- National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, China
- School of Chemical Engineering, University of Chinese Academy of Sciences, Beijing, China
| | - Jing Zhou
- Department of Geriatric Rehabilitation, Beijing Rehabilitation Hospital, Capital Medical University, Beijing, China
| | - Xiaolin Yu
- National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, China
| | - Shuai Lu
- National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, China
| | - Ruitian Liu
- National Key Laboratory of Biochemical Engineering, Institute of Process Engineering, Chinese Academy of Sciences, Beijing, China
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Yuan F, Zhang R, Li J, Lei Q, Wang S, Jiang F, Guo Y, Xiang M. CCR5-overexpressing mesenchymal stem cells protect against experimental autoimmune uveitis: insights from single-cell transcriptome analysis. J Neuroinflammation 2024; 21:136. [PMID: 38802924 PMCID: PMC11131209 DOI: 10.1186/s12974-024-03134-3] [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: 01/09/2024] [Accepted: 05/17/2024] [Indexed: 05/29/2024] Open
Abstract
Autoimmune uveitis is a leading cause of severe vision loss, and animal models provide unique opportunities for studying its pathogenesis and therapeutic strategies. Here we employ scRNA-seq, RNA-seq and various molecular and cellular approaches to characterize mouse models of classical experimental autoimmune uveitis (EAU), revealing that EAU causes broad retinal neuron degeneration and marker downregulation, and that Müller glia may act as antigen-presenting cells. Moreover, EAU immune response is primarily driven by Th1 cells, and results in dramatic upregulation of CC chemokines, especially CCL5, in the EAU retina. Accordingly, overexpression of CCR5, a CCL5 receptor, in mesenchymal stem cells (MSCs) enhances their homing capacity and improves their immunomodulatory outcomes in preventing EAU, by reducing infiltrating T cells and activated microglia and suppressing Nlrp3 inflammasome activation. Taken together, our data not only provide valuable insights into the molecular characteristics of EAU but also open an avenue for innovative MSC-based therapy.
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Affiliation(s)
- Fa Yuan
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Rong Zhang
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Jiani Li
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Qiannan Lei
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Shuyi Wang
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Fanying Jiang
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Yanan Guo
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China
| | - Mengqing Xiang
- State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, 510060, China.
- Guangdong Provincial Key Laboratory of Brain Function and Disease, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, 510080, China.
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Sichani AS, Khoddam S, Shakeri S, Tavakkoli Z, Jafroodi AR, Dabbaghipour R, Sisakht M, Fallahi J. Partial Reprogramming as a Method for Regenerating Neural Tissues in Aged Organisms. Cell Reprogram 2024; 26:10-23. [PMID: 38381402 DOI: 10.1089/cell.2023.0123] [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] [Indexed: 02/22/2024] Open
Abstract
Aging causes numerous age-related diseases, leading the human species to death. Nevertheless, rejuvenating strategies based on cell epigenetic modifications are a possible approach to counteract disease progression while getting old. Cell reprogramming of adult somatic cells toward pluripotency ought to be a promising tool for age-related diseases. However, researchers do not have control over this process as cells lose their fate, and cause potential cancerous cells or unexpected cell phenotypes. Direct and partial reprogramming were introduced in recent years with distinctive applications. Although direct reprogramming makes cells lose their identity, it has various applications in regeneration medicine. Temporary and regulated in vivo overexpression of Yamanaka factors has been shown in several experimental contexts to be achievable and is used to rejuvenate mice models. This regeneration can be accomplished by altering the epigenetic adult cell signature to the signature of a younger cell. The greatest advantage of partial reprogramming is that this method does not allow cells to lose their identity when they are resetting their epigenetic clock. It is a regimen of short-term Oct3/4, Sox2, Klf4, and c-Myc expression in vivo that prevents full reprogramming to the pluripotent state and avoids both tumorigenesis and the presence of unwanted undifferentiated cells. We know that many neurological age-related diseases, such as Alzheimer's disease, stroke, dementia, and Parkinson's disease, are the main cause of death in the last decades of life. Therefore, scientists have a special tendency regarding neuroregeneration methods to increase human life expectancy.
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Affiliation(s)
- Ali Saber Sichani
- Department of Biology, Texas A&M University, College Station, Texas, USA
| | - Somayeh Khoddam
- Department of Medical Genetics, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Shayan Shakeri
- Department of Medical Genetics, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Zahra Tavakkoli
- Department of Medical Genetics, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Arad Ranji Jafroodi
- Department of Medical Genetics, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Reza Dabbaghipour
- Department of Medical Genetics, School of Medicine, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Mohsen Sisakht
- Department of Molecular Medicine, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran
| | - Jafar Fallahi
- Department of Molecular Medicine, School of Advanced Medical Sciences and Technologies, Shiraz University of Medical Sciences, Shiraz, Iran
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Nishino R, Nomura-Komoike K, Iida T, Fujieda H. Cell cycle-dependent activation of proneural transcription factor expression and reactive gliosis in rat Müller glia. Sci Rep 2023; 13:22712. [PMID: 38123648 PMCID: PMC10733309 DOI: 10.1038/s41598-023-50222-0] [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: 10/02/2023] [Accepted: 12/16/2023] [Indexed: 12/23/2023] Open
Abstract
Retinal Müller glia have a capacity to regenerate neurons in lower vertebrates like zebrafish, but such ability is extremely limited in mammals. In zebrafish, Müller glia proliferate after injury, which promotes their neurogenic reprogramming while inhibiting reactive gliosis. In mammals, however, how the cell cycle affects the fate of Müller glia after injury remains unclear. Here, we focused on the expression of proneural transcription factors, Ngn2 and Ascl1, and a gliosis marker glial fibrillary acidic protein (GFAP) in rat Müller glia after N-methyl-N-nitrosourea (MNU)-induced photoreceptor injury and analyzed the role of Müller glia proliferation in the regulation of their expression using retinal explant cultures. Thymidine-induced G1/S arrest of Müller glia proliferation significantly hampered the expression of Ascl1, Ngn2, and GFAP, and release from the arrest induced their upregulation. The migration of Müller glia nuclei into the outer nuclear layer was also shown to be cell cycle-dependent. These data suggest that, unlike the situation in zebrafish, cell cycle progression of Müller glia in mammals promotes both neurogenic reprogramming and reactive gliosis, which may be one of the mechanisms underlying the limited regenerative capacity of the mammalian retina.
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Affiliation(s)
- Reiko Nishino
- Department of Anatomy and Neurobiology, School of Medicine, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo, 162-8666, Japan
- Department of Ophthalmology, School of Medicine, Tokyo Women's Medical University, Tokyo, Japan
| | - Kaori Nomura-Komoike
- Department of Anatomy and Neurobiology, School of Medicine, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo, 162-8666, Japan
| | - Tomohiro Iida
- Department of Ophthalmology, School of Medicine, Tokyo Women's Medical University, Tokyo, Japan
| | - Hiroki Fujieda
- Department of Anatomy and Neurobiology, School of Medicine, Tokyo Women's Medical University, 8-1 Kawada-cho, Shinjuku-ku, Tokyo, 162-8666, Japan.
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Li J, Zeng Q. Trim9 regulates the directional differentiation of retinal Müller cells to retinal ganglion cells. ZHONG NAN DA XUE XUE BAO. YI XUE BAN = JOURNAL OF CENTRAL SOUTH UNIVERSITY. MEDICAL SCIENCES 2023; 48:1561-1571. [PMID: 38432885 PMCID: PMC10929896 DOI: 10.11817/j.issn.1672-7347.2023.230108] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Subscribe] [Scholar Register] [Received: 03/15/2023] [Indexed: 03/05/2024]
Abstract
OBJECTIVES Glaucoma is a leading cause of irreversible blindness, and effective therapies to reverse the visual system damage caused by glaucoma are still lacking. Recently, the stem cell therapy enable the repair and regeneration of the damaged retinal neurons, but challenges regarding the source of stem cells remain. This study aims to investigate a protocol that allows the dedifferentiation of Müller cells into retinal stem cells, following by directed differentiation into retinal ganglion cells with high efficiency, and to provide a new method of cellular acquisition for retinal stem cells. METHODS Epidermal cell growth factor and fibroblast growth factor 2 were used to induce the dedifferentiation of rat retinal Müller cells into retinal neural stem cells. Retinal stem cells derived from Müller cells were infected with a Trim9 overexpression lentiviral vector (PGC-FU-Trim9-GFP), and the efficiency of viral infection was assessed by fluorescence microscopy and flow cytometry. Retinoic acid and brain-derived neurotrophic factor treatments were used to induce the differentiation of the retinal stem cells into neurons and glial cells with or without the overexpression of Trim9. The expressions of each cellular marker (GLAST, GS, rhodopsin, PKC, HPC-1, Calbindin, Thy1.1, Brn-3b, Nestin, Pax6) were detected by immunofluorescence, PCR/real-time RT-PCR or Western blotting. RESULTS Rat retinal Müller cells expressed neural stem cells markers (Nestin and Pax6) with the treatment of epidermal cell growth factor and fibroblast growth factor 2. The Thy1.1 positive cell rate of retinal stem cells overexpressing Trim9 was significantly increased, indicating their directional differentiation into retinal ganglion cells after treatment with retinoic acid and brain-derived neurotrophic factor. CONCLUSIONS In this study, rat retinal Müller cells are dedifferentiated into retinal stem cells successfully, and Trim9 promotes the directional differentiation from retinal stem cells to retinal ganglion cells effectively.
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Affiliation(s)
- Jinxiang Li
- Department of Ophthalmology, First Hospital Affiliated with Hunan Normal University (Hunan Provincial People's Hospital), Changsha 410005, China.
| | - Qi Zeng
- Department of Ophthalmology, First Hospital Affiliated with Hunan Normal University (Hunan Provincial People's Hospital), Changsha 410005, China.
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Fafure AA, Edem EE, Obisesan AO, Enye LA, Adekeye AO, Adetunji AE, Nebo KE, Olusegun AA, Fafure OE. Fermented maize slurry (Ogi) and its supernatant (Omidun) mitigate elevated intraocular pressure by modulating BDNF expression and glial plasticity in the retina-gut axis of glaucomatous rats. JOURNAL OF COMPLEMENTARY & INTEGRATIVE MEDICINE 2022; 19:887-896. [PMID: 34380184 DOI: 10.1515/jcim-2021-0114] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 03/13/2021] [Accepted: 07/27/2021] [Indexed: 06/13/2023]
Abstract
OBJECTIVES Growing interest has been reported on the health benefits of fermented foods, which includes cognition enhancement and inflammation attenuation. BDNF is a known protectant against retinal degeneration, however, therapies that target this neurotrophic factor has been limited. Therefore, we assessed the reaction of BDNF and glial cells in glaucomatous rats and their response to treatment with fermented maize products. METHODS Thirty male adult rats were either injected via the episcleral vein with hypertonic saline to elevate intraocular pressure (IOP) or treated with fermented maize slurry (Ogi) or its supernatant (Omidun). Following sacrifice, the retina and duodenum were studied by immunohistochemical analysis using antibodies directed against GFAP, AIF-1 and BDNF. RESULTS Hypertonic saline injection produced hypertrophy of the Müller cells and increased GFAP and AIF-1 expression in the retina and gut when compared to the control. Treatment with Ogi and Omidun produced varying degrees of reduction of gliosis, protection against hypertonic saline-induced retinal ganglion cell loss, and reduced intraocular pressure. BDNF expression was downregulated following the hypertonic saline assault, while Omidun and Ogi treatment abrogated its reduction following the hypertonic saline assault. CONCLUSIONS Collectively, our findings suggest that acute elevation of IOP alters crosstalk between gut and retina with consequent aberrant activation of glial cells; and that probiotic bacteria like the lactic acid bacteria rich in fermented foods including Ogi and Omidun may offer neuroprotection to the ganglionic cells by attenuating the retinal glial reaction and improving BDNF activity.
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Affiliation(s)
- Adedamola Adediran Fafure
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Edem Ekpenyong Edem
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Abiola Oluwatosin Obisesan
- Department of Pharmaceutical Microbiology, Faculty of Pharmacy, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Linus Anderson Enye
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Adeshina Oloruntoba Adekeye
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Adedeji Enitan Adetunji
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Kate Eberechukwu Nebo
- Neuroscience Unit, Department of Anatomy, Faculty of Basic Medical Sciences, College of Medicine and Health Sciences, Afe Babalola University, Ado-Ekiti, Nigeria
| | - Adebayo Adeoluwa Olusegun
- Neuropharmacology Unit, Department of Pharmacology and Therapeutics, College of Medicine and Health Sciences, Afe-Babalola University, Ado-Ekiti, Nigeria
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Peña JS, Vazquez M. Harnessing the Neuroprotective Behaviors of Müller Glia for Retinal Repair. FRONT BIOSCI-LANDMRK 2022; 27:169. [PMID: 35748245 PMCID: PMC9639582 DOI: 10.31083/j.fbl2706169] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2022] [Revised: 04/18/2022] [Accepted: 05/05/2022] [Indexed: 11/29/2022]
Abstract
Progressive and irreversible vision loss in mature and aging adults creates a health and economic burden, worldwide. Despite the advancements of many contemporary therapies to restore vision, few approaches have considered the innate benefits of gliosis, the endogenous processes of retinal repair that precede vision loss. Retinal gliosis is fundamentally driven by Müller glia (MG) and is characterized by three primary cellular mechanisms: hypertrophy, proliferation, and migration. In early stages of gliosis, these processes have neuroprotective potential to halt the progression of disease and encourage synaptic activity among neurons. Later stages, however, can lead to glial scarring, which is a hallmark of disease progression and blindness. As a result, the neuroprotective abilities of MG have remained incompletely explored and poorly integrated into current treatment regimens. Bioengineering studies of the intrinsic behaviors of MG hold promise to exploit glial reparative ability, while repressing neuro-disruptive MG responses. In particular, recent in vitro systems have become primary models to analyze individual gliotic processes and provide a stepping stone for in vivo strategies. This review highlights recent studies of MG gliosis seeking to harness MG neuroprotective ability for regeneration using contemporary biotechnologies. We emphasize the importance of studying gliosis as a reparative mechanism, rather than disregarding it as an unfortunate clinical prognosis in diseased retina.
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Affiliation(s)
- Juan S. Peña
- Department of Biomedical Engineering, Rutgers, The State
University of New Jersey, Piscataway (08854), New Jersey, USA
| | - Maribel Vazquez
- Department of Biomedical Engineering, Rutgers, The State
University of New Jersey, Piscataway (08854), New Jersey, USA
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Gene-independent therapeutic interventions to maintain and restore light sensitivity in degenerating photoreceptors. Prog Retin Eye Res 2022; 90:101065. [PMID: 35562270 DOI: 10.1016/j.preteyeres.2022.101065] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/01/2022] [Revised: 04/08/2022] [Accepted: 04/18/2022] [Indexed: 12/14/2022]
Abstract
Neurodegenerative retinal diseases are a prime cause of blindness in industrialized countries. In many cases, there are no therapeutic treatments, although they are essential to improve patients' quality of life. A set of disease-causing genes, which primarily affect photoreceptors, has already been identified and is of major interest for developing gene therapies. Nevertheless, depending on the nature and the state of the disease, gene-independent strategies are needed. Various strategies to halt disease progression or maintain function of the retina are under research. These therapeutic interventions include neuroprotection, direct reprogramming of affected photoreceptors, the application of non-coding RNAs, the generation of artificial photoreceptors by optogenetics and cell replacement strategies. During recent years, major breakthroughs have been made such as the first optogenetic application to a blind patient whose visual function partially recovered by targeting retinal ganglion cells. Also, RPE cell transplantation therapies are under clinical investigation and show great promise to improve visual function in blind patients. These cells are generated from human stem cells. Similar therapies for replacing photoreceptors are extensively tested in pre-clinical models. This marks just the start of promising new cures taking advantage of developments in the areas of genetic engineering, optogenetics, and stem-cell research. In this review, we present the recent therapeutic advances of gene-independent approaches that are currently under clinical evaluation. Our main focus is on photoreceptors as these sensory cells are highly vulnerable to degenerative diseases, and are crucial for light detection.
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12
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Tworig JM, Coate C, Feller MB. Excitatory neurotransmission activates compartmentalized calcium transients in Müller glia without affecting lateral process motility. eLife 2021; 10:73202. [PMID: 34913435 PMCID: PMC8806189 DOI: 10.7554/elife.73202] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2021] [Accepted: 12/15/2021] [Indexed: 11/13/2022] Open
Abstract
Neural activity has been implicated in the motility and outgrowth of glial cell processes throughout the central nervous system. Here, we explore this phenomenon in Müller glia, which are specialized radial astroglia that are the predominant glial type of the vertebrate retina. Müller glia extend fine filopodia-like processes into retinal synaptic layers, in similar fashion to brain astrocytes and radial glia that exhibit perisynaptic processes. Using two-photon volumetric imaging, we found that during the second postnatal week, Müller glial processes were highly dynamic, with rapid extensions and retractions that were mediated by cytoskeletal rearrangements. During this same stage of development, retinal waves led to increases in cytosolic calcium within Müller glial lateral processes and stalks. These regions comprised distinct calcium compartments, distinguished by variable participation in waves, timing, and sensitivity to an M1 muscarinic acetylcholine receptor antagonist. However, we found that motility of lateral processes was unaffected by the presence of pharmacological agents that enhanced or blocked wave-associated calcium transients. Finally, we found that mice lacking normal cholinergic waves in the first postnatal week also exhibited normal Müller glial process morphology. Hence, outgrowth of Müller glial lateral processes into synaptic layers is determined by factors that are independent of neuronal activity. When it comes to studying the nervous system, neurons often steal the limelight; yet, they can only work properly thanks to an ensemble cast of cell types whose roles are only just emerging. For example, ‘glial cells’ – their name derives from the Greek word for glue – were once thought to play only a passive, supporting function in nervous tissues. Now, growing evidence shows that they are, in fact, integrated into neural circuits: their activity is influenced by neurons, and, in turn, they help neurons to function properly. The role of glial cells is becoming clear in the retina, the thin, light-sensitive layer that lines the back of the eye and relays visual information to the brain. There, beautifully intricate Müller glial cells display fine protrusions (or ‘processes') that intermingle with synapses, the busy space between neurons where chemical messengers are exchanged. These messengers can act on Müller cells, triggering cascades of molecular events that may influence the structure and function of glia. This is of particular interest during development: as Müller cells mature, they are exposed to chemicals released by more fully formed retinal neurons. Tworig et al. explored how neuronal messengers can influence the way Müller cells grow their processes. To do so, they tracked mouse retinal glial cells ‘live’ during development, showing that they were growing fine, highly dynamic processes in a region rich in synapses just as neurons and glia increased their communication. However, using drugs to disrupt this messaging for a short period did not seem to impact how the processes grew. Extending the blockade over a longer timeframe also did not change the way Müller cells developed, with the cells still acquiring their characteristic elaborate process networks. Taken together, these results suggest that the structural maturation of Müller glial cells is not impacted by neuronal signaling, giving a more refined understanding of how glia form in the retina and potentially in the brain.
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Affiliation(s)
- Joshua M Tworig
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
| | - Chandler Coate
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
| | - Marla B Feller
- Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, United States
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13
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Sherpa RD, Hui SP. An insight on established retinal injury mechanisms and prevalent retinal stem cell activation pathways in vertebrate models. Animal Model Exp Med 2021; 4:189-203. [PMID: 34557646 PMCID: PMC8446703 DOI: 10.1002/ame2.12177] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2020] [Accepted: 06/09/2021] [Indexed: 12/22/2022] Open
Abstract
Implementing different tools and injury mechanisms in multiple animal models of retina regeneration, researchers have discovered the existence of retinal stem/progenitor cells. Although they appear to be distributed uniformly across the vertebrate lineage, the reparative potential of the retina is mainly restricted to lower vertebrates. Regenerative repair post-injury requires the creation of a proliferative niche, vital for proper stem cell activation, propagation, and lineage differentiation. This seems to be lacking in mammals. Hence, in this review, we first discuss the many forms of retinal injuries that have been generated using animal models. Next, we discuss how they are utilized to stimulate regeneration and mimic eye disease pathologies. The key to driving stem cell activation in mammals relies on the information we can gather from these models. Lastly, we present a brief update about the genes, growth factors, and signaling pathways that have been brought to light using these models.
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Affiliation(s)
| | - Subhra Prakash Hui
- S. N. Pradhan Centre for NeurosciencesUniversity of CalcuttaKolkataIndia
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14
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Wang F, Cheng L, Zhang X. Reprogramming Glial Cells into Functional Neurons for Neuro-regeneration: Challenges and Promise. Neurosci Bull 2021; 37:1625-1636. [PMID: 34283396 DOI: 10.1007/s12264-021-00751-3] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2021] [Accepted: 04/24/2021] [Indexed: 01/02/2023] Open
Abstract
The capacity for neurogenesis in the adult mammalian brain is extremely limited and highly restricted to a few regions, which greatly hampers neuronal regeneration and functional restoration after neuronal loss caused by injury or disease. Meanwhile, transplantation of exogenous neuronal stem cells into the brain encounters several serious issues including immune rejection and the risk of tumorigenesis. Recent discoveries of direct reprogramming of endogenous glial cells into functional neurons have provided new opportunities for adult neuro-regeneration. Here, we extensively review the experimental findings of the direct conversion of glial cells to neurons in vitro and in vivo and discuss the remaining issues and challenges related to the glial subtypes and the specificity and efficiency of direct cell-reprograming, as well as the influence of the microenvironment. Although in situ glial cell reprogramming offers great potential for neuronal repair in the injured or diseased brain, it still needs a large amount of research to pave the way to therapeutic application.
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Affiliation(s)
- Fengchao Wang
- State Key Laboratory of Cognitive Neuroscience and Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing, 100875, China
| | - Leping Cheng
- Key Laboratory of Longevity and Aging-related Diseases of Chinese Ministry of Education, Guangxi-ASEAN Collaborative Innovation Center for Major Disease Prevention and Treatment, and Guangxi Key Laboratory of Regenerative Medicine, Center for Translational Medicine, Guangxi Medical University, Nanning, 530021, China. .,Department of Cell Biology and Genetics, School of Basic Medical Sciences, Guangxi Medical University, Nanning, 530021, China. .,Guangxi Health Commission Key Laboratory of Basic Research on Brain Function and Disease, Guangxi Medical University, Nanning, 530021, China.
| | - Xiaohui Zhang
- State Key Laboratory of Cognitive Neuroscience and Learning, IDG/McGovern Institute for Brain Research, Beijing Normal University, Beijing, 100875, China.
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15
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Zhu RL, Fang Y, Yu HH, Chen DF, Yang L, Cho KS. Absence of ephrin-A2/A3 increases retinal regenerative potential for Müller cells in Rhodopsin knockout mice. Neural Regen Res 2021; 16:1317-1322. [PMID: 33318411 PMCID: PMC8284269 DOI: 10.4103/1673-5374.301034] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/10/2020] [Revised: 05/11/2020] [Accepted: 09/28/2020] [Indexed: 11/25/2022] Open
Abstract
Müller cells (MC) are considered dormant retinal progenitor cells in mammals. Previous studies demonstrated ephrin-As act as negative regulators of neural progenitor cells in the retina and brain. It remains unclear whether the lack of ephrin-A2/A3 is sufficient to promote the neurogenic potential of MC. Here we investigated whether the MC is the primary retinal cell type expressing ephrin-A2/A3 and their role on the neurogenic potential of Müller cells. In this study, we showed that ephrin-A2/A3 and their receptor EphA4 were expressed in retina and especially enriched in MC. The level of ephrinAs/EphA4 expression increased as the retina matured that is correlated with the reduced proliferative and progenitor cell potential of MC. Next, we investigated the proliferation in primary MC cultures isolated from wild-type and A2-/- A3-/- mice by 5-ethynyl-2'-deoxyuridine (EdU) incorporation. We detected a significant increase of EdU+ cells in MC derived from A2-/- A3-/- mice. Next, we investigated the role of ephrin-A2/A3 in mice undergoing photoreceptor degeneration such as Rhodopsin knockout (Rho-/-) mice. To further evaluate the role of ephrin-A2/A3 in MC proliferation in vivo, EdU was injected intraperitoneally to adult wild-type, A2-/- A3-/- , Rho-/- and Rho-/- A2-/- A3-/- mice and the numbers of EdU+ cells distributed among different layers of the retina. EphrinAs/EphA4 expression was upregulated in the retina of Rho-/- mice compared to the wild-type mice. In addition, cultured MC derived from ephrin-A2-/- A3-/- mice also expressed higher levels of progenitor cell markers and exhibited higher proliferation potential than those from wild-type mice. Interestingly, we detected a significant increase of EdU+ cells in the retinas of adult ephrin-A2-/- A3-/- mice mainly in the inner nuclear layer; and these EdU+ cells were co-localized with MC marker, cellular retinaldehyde-binding protein, suggesting some proliferating cells are from MC. In Rhodopsin knockout mice (Rho-/- A2-/- A3-/- mice), a significantly greater amount of EdU+ cells were located in the ciliary body, retina and RPE than that of Rho-/- mice. Comparing between 6 and 12 weeks old Rho-/- A2-/- A3-/- mice, we recorded more EdU+ cells in the outer nuclear layer in the 12-week-old mice undergoing severe retinal degeneration. Taken together, Ephrin-A2/A3 are negative regulators of the proliferative and neurogenic potentials of MC. Absence of ephrin-A2/A3 promotes the migration of proliferating cells into the outer nuclear layer and may lead to retinal cell regeneration. All experimental procedures were approved by the Animal Care and Use Committee at Schepens Eye Research Institute, USA (approval No. S-353-0715) on October 24, 2012.
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Affiliation(s)
- Rui-Lin Zhu
- Department of Ophthalmology, Peking University First Hospital, Beijing, China
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School, Boston, MA, USA
| | - Yuan Fang
- Department of Ophthalmology and Visual Science, Eye & ENT Hospital, Shanghai Medical College, Fudan University, Shanghai, China
| | - Hong-Hua Yu
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School, Boston, MA, USA
- Guangdong Eye Institute, Department of Ophthalmology, Guangdong Provincial People's Hospital, Guangdong Academy of Medical Sciences, Guangzhou, Guangdong Province, China
| | - Dong F. Chen
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School, Boston, MA, USA
| | - Liu Yang
- Department of Ophthalmology, Peking University First Hospital, Beijing, China
| | - Kin-Sang Cho
- Schepens Eye Research Institute, Massachusetts Eye and Ear, Harvard Medical School, Boston, MA, USA
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16
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Advances in Regeneration of Retinal Ganglion Cells and Optic Nerves. Int J Mol Sci 2021; 22:ijms22094616. [PMID: 33924833 PMCID: PMC8125313 DOI: 10.3390/ijms22094616] [Citation(s) in RCA: 16] [Impact Index Per Article: 4.0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2021] [Revised: 04/20/2021] [Accepted: 04/21/2021] [Indexed: 02/07/2023] Open
Abstract
Glaucoma, the second leading cause of blindness worldwide, is an incurable neurodegenerative disorder due to the dysfunction of retinal ganglion cells (RGCs). RGCs function as the only output neurons conveying the detected light information from the retina to the brain, which is a bottleneck of vision formation. RGCs in mammals cannot regenerate if injured, and RGC subtypes differ dramatically in their ability to survive and regenerate after injury. Recently, novel RGC subtypes and markers have been uncovered in succession. Meanwhile, apart from great advances in RGC axon regeneration, some degree of experimental RGC regeneration has been achieved by the in vitro differentiation of embryonic stem cells and induced pluripotent stem cells or in vivo somatic cell reprogramming, which provides insights into the future therapy of myriad neurodegenerative disorders. Further approaches to the combination of different factors will be necessary to develop efficacious future therapeutic strategies to promote ultimate axon and RGC regeneration and functional vision recovery following injury.
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17
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Xia X, Teotia P, Patel H, Van Hook MJ, Ahmad I. Chemical induction of neurogenic properties in mammalian Müller glia. STEM CELLS (DAYTON, OHIO) 2021; 39:1081-1090. [PMID: 33764634 DOI: 10.1002/stem.3370] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Subscribe] [Scholar Register] [Received: 10/01/2020] [Accepted: 02/19/2021] [Indexed: 11/08/2022]
Abstract
Müller glia (MG), cells that maintain homeostasis in the retina, are dormant stem cells that can regenerate neurons upon injury. However, the regenerative property of MG, which is reproducibly displayed in the lower vertebrates, is not readily observed in the mammals even upon forced expression of regulatory genes or exposure to growth factors. Here, we demonstrate a reproducible unmasking of the neurogenic properties of enriched rodent MG by serial exposure to different combinations of small molecules. The enriched MG, in response to changing culture conditions, silenced glia-specific genes and acquired transcriptional signature of neurons, accompanied by upregulation of genes known to regulate neuronal potential of MG. The MG-derived neurons expressed immunoreactivities corresponding to neuronal proteins and displayed electrophysiological features of immature neurons. Our study presents a proof of principle of pharmacological activation of neurogenic properties of mammalian MG, which may be utilized for therapeutic regeneration.
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Affiliation(s)
- Xiaohuan Xia
- Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA.,Center for Translational Neurodegeneration and Regenerative Therapy, Shanghai Tenth People's Hospital affiliated to Tongji University School of Medicine, Shanghai, People's Republic of China
| | - Pooja Teotia
- Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA
| | - Hiren Patel
- Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA
| | - Matthew J Van Hook
- Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA
| | - Iqbal Ahmad
- Department of Ophthalmology and Visual Sciences, University of Nebraska Medical Center, Omaha, Nebraska, USA
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18
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You M, Rong R, Zeng Z, Li H, Xia X, Ji D. Single-cell RNA sequencing: A new opportunity for retinal research. WILEY INTERDISCIPLINARY REVIEWS-RNA 2021; 12:e1652. [PMID: 33754496 DOI: 10.1002/wrna.1652] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/25/2020] [Revised: 03/01/2021] [Accepted: 03/02/2021] [Indexed: 11/08/2022]
Abstract
Single-cell RNA sequencing (scRNA-seq) is a technology for single-cell transcriptome analysis that can be used to characterize complex dynamics of various retinal cell types. It provides deep scrutiny into the gene expression character of diverse cell types, lending insight into all the biological processes being carried out. The scRNA-seq is an alternative to regular RNA-seq, which does not achieve cellular heterogeneity. The retina, is a part of the central nervous system (CNS) and consists of six types of neurons and several types of glial cells. Studying retinal cell heterogeneity is important for understanding retinal diseases. Currently, scRNA-seq is employed to assess retina development and retinal disease pathogenesis and has improved our understanding of the relationship between the retina, its visual pathways, and the brain. Moreover, this technology provides new ideas on the sensitivity and molecular mechanisms of cell subtypes involved in retinal-related diseases. The application of scRNA-seq technology has given us a deeper understanding of the latest advancements and challenges in retinal development and diseases. We advocate scRNA-seq as one of the important tools for developing novel therapies for retinal diseases. This article is categorized under: RNA Methods > RNA Analyses in Cells RNA in Disease and Development > RNA in Development RNA in Disease and Development > RNA in Disease.
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Affiliation(s)
- Mengling You
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
| | - Rong Rong
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
| | - Zhou Zeng
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
| | - Haibo Li
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
| | - Xiaobo Xia
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
| | - Dan Ji
- Department of Ophthalmology, Xiangya Hospital, Central South University, Changsha, China.,Hunan Key Laboratory of Ophthalmology, Changsha, Hunan, China
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19
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Kim KM, Thaqi M, Peterson DA, Marr RA. Induced Neurons for Disease Modeling and Repair: A Focus on Non-fibroblastic Cell Sources in Direct Reprogramming. Front Bioeng Biotechnol 2021; 9:658498. [PMID: 33777923 PMCID: PMC7995206 DOI: 10.3389/fbioe.2021.658498] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/25/2021] [Accepted: 02/12/2021] [Indexed: 11/13/2022] Open
Abstract
Direct cellular reprogramming exhibits distinct advantages over reprogramming from an induced pluripotent stem cell intermediate. These include a reduced risk of tumorigenesis and the likely preservation of epigenetic data. In vitro direct reprogramming approaches primarily aim to model the pathophysiological development of neurological disease and identify therapeutic targets, while in vivo direct reprogramming aims to develop treatments for various neurological disorders, including cerebral injury and cancer. In both approaches, there is progress toward developing increased control of subtype-specific production of induced neurons. A majority of research primarily utilizes fibroblasts as the donor cells. However, there are a variety of other somatic cell types that have demonstrated the potential for reprogramming into induced neurons. This review highlights studies that utilize non-fibroblastic cell sources for reprogramming, such as astrocytes, olfactory ensheathing cells, peripheral blood cells, Müller glia, and more. We will examine benefits and obstructions for translation into therapeutics or disease modeling, as well as efficiency of the conversion. A summary of donor cells, induced neuron types, and methods of induction is also provided.
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Affiliation(s)
- Kathryn M. Kim
- Center for Neurodegenerative Disease and Therapeutics, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL, United States
| | - Mentor Thaqi
- Center for Neurodegenerative Disease and Therapeutics, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL, United States
- Scholl College of Podiatric Medicine, Rosalind Franklin University of Medicine and Science, North Chicago, IL, United States
| | - Daniel A. Peterson
- Center for Neurodegenerative Disease and Therapeutics, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL, United States
| | - Robert A. Marr
- Center for Neurodegenerative Disease and Therapeutics, Chicago Medical School, Rosalind Franklin University of Medicine and Science, North Chicago, IL, United States
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20
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Tolomeo AM, Laterza C, Grespan E, Michielin F, Canals I, Kokaia Z, Muraca M, Gagliano O, Elvassore N. NGN2 mmRNA-Based Transcriptional Programming in Microfluidic Guides hiPSCs Toward Neural Fate With Multiple Identities. Front Cell Neurosci 2021; 15:602888. [PMID: 33679325 PMCID: PMC7928329 DOI: 10.3389/fncel.2021.602888] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2020] [Accepted: 01/18/2021] [Indexed: 01/13/2023] Open
Abstract
Recent advancements in cell engineering have succeeded in manipulating cell identity with the targeted overexpression of specific cell fate determining transcription factors in a process named transcriptional programming. Neurogenin2 (NGN2) is sufficient to instruct pluripotent stem cells (PSCs) to acquire a neuronal identity when delivered with an integrating system, which arises some safety concerns for clinical applications. A non-integrating system based on modified messenger RNA (mmRNA) delivery method, represents a valuable alternative to lentiviral-based approaches. The ability of NGN2 mmRNA to instruct PSC fate change has not been thoroughly investigated yet. Here we aimed at understanding whether the use of an NGN2 mmRNA-based approach combined with a miniaturized system, which allows a higher transfection efficiency in a cost-effective system, is able to drive human induced PSCs (hiPSCs) toward the neuronal lineage. We show that NGN2 mRNA alone is able to induce cell fate conversion. Surprisingly, the outcome cell population accounts for multiple phenotypes along the neural development trajectory. We found that this mixed population is mainly constituted by neural stem cells (45% ± 18 PAX6 positive cells) and neurons (38% ± 8 βIIITUBULIN positive cells) only when NGN2 is delivered as mmRNA. On the other hand, when the delivery system is lentiviral-based, both providing a constant expression of NGN2 or only a transient pulse, the outcome differentiated population is formed by a clear majority of neurons (88% ± 1 βIIITUBULIN positive cells). Altogether, our data confirm the ability of NGN2 to induce neuralization in hiPSCs and opens a new point of view in respect to the delivery system method when it comes to transcriptional programming applications.
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Affiliation(s)
- Anna Maria Tolomeo
- Department of Industrial Engineering, University of Padua, Padua, Italy
- L.i.f.e.L.a.b. Program, Consorzio per la Ricerca Sanitaria (CORIS), Padua, Italy
| | - Cecilia Laterza
- Department of Industrial Engineering, University of Padua, Padua, Italy
- L.i.f.e.L.a.b. Program, Consorzio per la Ricerca Sanitaria (CORIS), Padua, Italy
- Veneto Institute of Molecular Medicine, Padua, Italy
| | - Eleonora Grespan
- Institute of Neuroscience, National Research Council, Padua, Italy
| | - Federica Michielin
- Department of Industrial Engineering, University of Padua, Padua, Italy
- Veneto Institute of Molecular Medicine, Padua, Italy
| | - Isaac Canals
- Stem Cells, Aging and Neurodegeneration Group, Lund Stem Cell Center, Lund University, Lund, Sweden
| | - Zaal Kokaia
- Laboratory of Stem Cells and Restorative Neurology, Lund Stem Cell Center, Lund University, Lund, Sweden
| | - Maurizio Muraca
- L.i.f.e.L.a.b. Program, Consorzio per la Ricerca Sanitaria (CORIS), Padua, Italy
- Department of Women’s and Children’s Health, Faculty of Medicine, University of Padua, Padua, Italy
| | - Onelia Gagliano
- Department of Industrial Engineering, University of Padua, Padua, Italy
- L.i.f.e.L.a.b. Program, Consorzio per la Ricerca Sanitaria (CORIS), Padua, Italy
- Veneto Institute of Molecular Medicine, Padua, Italy
| | - Nicola Elvassore
- Department of Industrial Engineering, University of Padua, Padua, Italy
- L.i.f.e.L.a.b. Program, Consorzio per la Ricerca Sanitaria (CORIS), Padua, Italy
- Veneto Institute of Molecular Medicine, Padua, Italy
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21
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Molecular Mechanisms Underlying Ascl1-Mediated Astrocyte-to-Neuron Conversion. Stem Cell Reports 2021; 16:534-547. [PMID: 33577795 PMCID: PMC7940254 DOI: 10.1016/j.stemcr.2021.01.006] [Citation(s) in RCA: 21] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2019] [Revised: 01/12/2021] [Accepted: 01/12/2021] [Indexed: 12/19/2022] Open
Abstract
Direct neuronal reprogramming potentially provides valuable sources for cell-based therapies. Proneural gene Ascl1 converts astrocytes into induced neuronal (iN) cells efficiently both in vitro and in vivo. However, the underlying mechanisms are largely unknown. By combining RNA sequencing and chromatin immunoprecipitation followed by high-throughput sequencing, we found that the expression of 1,501 genes was markedly changed during the early stages of Ascl1-induced astrocyte-to-neuron conversion and that the regulatory regions of 107 differentially expressed genes were directly bound by ASCL1. Among Ascl1's direct targets, Klf10 regulates the neuritogenesis of iN cells at the early stage, Myt1 and Myt1l are critical for the electrophysiological maturation of iN cells, and Neurod4 and Chd7 are required for the efficient conversion of astrocytes into neurons. Together, this study provides more insights into understanding the molecular mechanisms underlying Ascl1-mediated astrocyte-to-neuron conversion and will be of value for the application of direct neuronal reprogramming.
RNA-seq and ChIP-seq were used to study Ascl1-induced astrocyte-to-neuron conversion Early Klf10 regulates neuritogenesis and electrophysiological properties of iN cells Myt1 and Myt1l are critical for the electrophysiological maturation of iN cells Neurod4 and Chd7 are required for efficient conversion of astrocytes to neurons
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22
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DeOliveira-Mello L, Mack AF, Lara JM, Arévalo R. Cultures of glial cells from optic nerve of two adult teleost fish: Astatotilapia burtoni and Danio rerio. J Neurosci Methods 2021; 353:109096. [PMID: 33581217 DOI: 10.1016/j.jneumeth.2021.109096] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2020] [Revised: 12/20/2020] [Accepted: 02/06/2021] [Indexed: 10/22/2022]
Abstract
BACKGROUND In vitro studies are very useful to increase the knowledge of different cell types and could be the key to understand cell metabolism and function. Fish optic nerves (ON) can recover visual functions by reestablishing its structure and reconnecting the axons of ganglion cells. This is because fish show spontaneous regeneration of the central nervous system which does not occur in mammals. In addition, several studies have indicated that glial cells of ON have different properties in comparison to the glial cells from brain or retina. Consequently, providing an in vitro tool will be highly beneficial to increase the knowledge of these cells. NEW METHOD We developed a cell culture protocol to isolate glial cells from ON of two teleost fish species, Danio rerio and Astatotilapia burtoni. RESULTS The optimized protocol allowed us to obtain ON cells and brain-derived cells from adult teleost fish. These cells were characterized as glial cells and their proprieties in vitro were analyzed.Comparison with Existing Method(s): Although it is striking that ON glial cells show peculiarities, their study in vitro has been limited by the only published protocol going back to the 1990s. Our protocol makes glial cells of different fish species available for experiments and studies to increase the understanding of these glial cell types. CONCLUSIONS This validated and effective in vitro tool increases the possibilities on studies of glial cells from fish ON which implies a reduction in animal experimentation.
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Affiliation(s)
- Laura DeOliveira-Mello
- Dept. Cell Biology and Pathology, IBSAL-Institute of Neurosciences of Castilla and León University of Salamanca Salamanca, Spain.
| | - Andreas F Mack
- Institute of Clinical Anatomy and Cell Analysis University of Tübingen Tübingen, Germany
| | - Juan M Lara
- Dept. Cell Biology and Pathology, IBSAL-Institute of Neurosciences of Castilla and León University of Salamanca Salamanca, Spain
| | - Rosario Arévalo
- Dept. Cell Biology and Pathology, IBSAL-Institute of Neurosciences of Castilla and León University of Salamanca Salamanca, Spain
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23
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Fu Z, Kern TS, Hellström A, Smith LEH. Fatty acid oxidation and photoreceptor metabolic needs. J Lipid Res 2021; 62:100035. [PMID: 32094231 PMCID: PMC7905050 DOI: 10.1194/jlr.tr120000618] [Citation(s) in RCA: 57] [Impact Index Per Article: 14.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/13/2020] [Revised: 02/14/2020] [Indexed: 01/31/2023] Open
Abstract
Photoreceptors have high energy demands and a high density of mitochondria that produce ATP through oxidative phosphorylation (OXPHOS) of fuel substrates. Although glucose is the major fuel for CNS brain neurons, in photoreceptors (also CNS), most glucose is not metabolized through OXPHOS but is instead metabolized into lactate by aerobic glycolysis. The major fuel sources for photoreceptor mitochondria remained unclear for almost six decades. Similar to other tissues (like heart and skeletal muscle) with high metabolic rates, photoreceptors were recently found to metabolize fatty acids (palmitate) through OXPHOS. Disruption of lipid entry into photoreceptors leads to extracellular lipid accumulation, suppressed glucose transporter expression, and a duel lipid/glucose fuel shortage. Modulation of lipid metabolism helps restore photoreceptor function. However, further elucidation of the types of lipids used as retinal energy sources, the metabolic interaction with other fuel pathways, as well as the cross-talk among retinal cells to provide energy to photoreceptors is not fully understood. In this review, we will focus on the current understanding of photoreceptor energy demand and sources, and potential future investigations of photoreceptor metabolism.
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Affiliation(s)
- Zhongjie Fu
- Department of Ophthalmology, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA; Manton Center for Orphan Disease, Boston Children's Hospital, Boston, MA, USA.
| | - Timothy S Kern
- Center for Translational Vision Research, Gavin Herbert Eye Institute, Irvine, CA, USA
| | - Ann Hellström
- Section for Ophthalmology, Department of Clinical Neuroscience, Institute of Neuroscience and Physiology, Sahlgrenska Academy, University of Gothenburg, Göteborg, Sweden
| | - Lois E H Smith
- Department of Ophthalmology, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA.
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24
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Liu Y, Lee RK. Cell transplantation to replace retinal ganglion cells faces challenges - the Switchboard Dilemma. Neural Regen Res 2021; 16:1138-1143. [PMID: 33269762 PMCID: PMC8224141 DOI: 10.4103/1673-5374.300329] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/04/2022] Open
Abstract
The mammalian retina displays incomplete intrinsic regenerative capacities; therefore, retina degeneration is a major cause of irreversible blindness such as glaucoma, age-related macular degeneration and diabetic retinopathy. These diseases lead to the loss of retinal cells and serious vision loss in the late stage. Stem cell transplantation is a great promising novel treatment for these incurable retinal degenerative diseases and represents an exciting area of regenerative neurotherapy. Several suitable stem cell sources for transplantation including human embryonic stem cells, induced pluripotent stem cells and adult stem cells have been identified as promising target populations. However, the retina is an elegant neuronal complex composed of various types of cells with different functions. The replacement of these different types of cells by transplantation should be addressed separately. So far, retinal pigment epithelium transplantation has achieved the most advanced stage of clinical trials, while transplantation of retinal neurons such as retinal ganglion cells and photoreceptors has been mostly studied in pre-clinical animal models. In this review, we opine on the key problems that need to be addressed before stem cells transplantation, especially for replacing injured retinal ganglion cells, may be used practically for treatment. A key problem we have called the Switchboard Dilemma is a major block to have functional retinal ganglion cell replacement. We use the public switchboard telephone network as an example to illustrate different difficulties for replacing damaged components in the retina that allow for visual signaling. Retinal ganglion cell transplantation is confronted by significant hurdles, because retinal ganglion cells receive signals from different interneurons, integrate and send signals to the correct targets of the visual system, which functions similar to the switchboard in a telephone network – therefore the Switchboard Dilemma.
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Affiliation(s)
- Yuan Liu
- Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, FL, USA
| | - Richard K Lee
- Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, FL, USA
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25
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Abstract
Normal retina and its cell layers are essential for processing visual stimuli, and loss of its integrity has been documented in many disease processes. The numbers and the axonal processes of retinal ganglion cells are reduced substantially in glaucoma, leading to vision loss and blindness. Similarly, selective loss of photoreceptors in age-related macular degeneration and hereditary retinal dystrophies also results in the compromise of visual acuity. Development of genetically modified mice has led to increased understanding of the pathogenesis of many retinal diseases. Similarly, in this digital era, usage of modalities to quantify the retinal cell loss has grown exponentially leading to a better understanding of the suitability of animal models to study human retinal diseases. These quantification modalities provide valuable quantifiable data in studying pathogenesis and disease progression. This review will discuss the immunohistochemical markers for various retinal cells, available automated tools to quantify retinal cells, and present an example of retinal ganglion cell quantification using HALO image analysis platform. Additionally, we briefly review retinal cell types and subtypes, salient features of retina in various laboratory animal species, and a few of the main disease processes that affect retinal cell numbers in humans.
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Affiliation(s)
| | - Henry Chen
- 7845Regeneron Pharmaceuticals, Tarrytown, NY, USA
| | - Ying Hu
- 7845Regeneron Pharmaceuticals, Tarrytown, NY, USA
| | - Oliver C Turner
- Novartis, 98557Novartis Institutes for BioMedical Research, Preclinical Safety, East Hanover, NJ, USA
| | - Olulanu H Aina
- 426218Janssen Pharmaceutical Company of Johnson & Johnson, Spring House, PA, USA
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26
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Cao J, O'Day DR, Pliner HA, Kingsley PD, Deng M, Daza RM, Zager MA, Aldinger KA, Blecher-Gonen R, Zhang F, Spielmann M, Palis J, Doherty D, Steemers FJ, Glass IA, Trapnell C, Shendure J. A human cell atlas of fetal gene expression. Science 2020; 370:370/6518/eaba7721. [PMID: 33184181 DOI: 10.1126/science.aba7721] [Citation(s) in RCA: 419] [Impact Index Per Article: 83.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/03/2020] [Accepted: 09/10/2020] [Indexed: 12/14/2022]
Abstract
The gene expression program underlying the specification of human cell types is of fundamental interest. We generated human cell atlases of gene expression and chromatin accessibility in fetal tissues. For gene expression, we applied three-level combinatorial indexing to >110 samples representing 15 organs, ultimately profiling ~4 million single cells. We leveraged the literature and other atlases to identify and annotate hundreds of cell types and subtypes, both within and across tissues. Our analyses focused on organ-specific specializations of broadly distributed cell types (such as blood, endothelial, and epithelial), sites of fetal erythropoiesis (which notably included the adrenal gland), and integration with mouse developmental atlases (such as conserved specification of blood cells). These data represent a rich resource for the exploration of in vivo human gene expression in diverse tissues and cell types.
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Affiliation(s)
- Junyue Cao
- Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, USA
| | - Diana R O'Day
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA
| | - Hannah A Pliner
- Brotman Baty Institute for Precision Medicine, Seattle, WA, USA
| | - Paul D Kingsley
- Department of Pediatrics, University of Rochester Medical Center, Rochester, NY, USA
| | - Mei Deng
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA
| | - Riza M Daza
- Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, USA
| | - Michael A Zager
- Brotman Baty Institute for Precision Medicine, Seattle, WA, USA.,Center for Data Visualization, Fred Hutchinson Cancer Research Center, Seattle, WA, USA
| | - Kimberly A Aldinger
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA.,Center for Integrative Brain Research, Seattle Children's Research Institute, Seattle, WA, USA
| | - Ronnie Blecher-Gonen
- Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, USA
| | | | - Malte Spielmann
- Human Molecular Genomics Group, Max Planck Institute for Molecular Genetics, Berlin, Germany.,Institute of Human Genetics, University of Lübeck, Lübeck, Germany
| | - James Palis
- Department of Pediatrics, University of Rochester Medical Center, Rochester, NY, USA
| | - Dan Doherty
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA.,Brotman Baty Institute for Precision Medicine, Seattle, WA, USA.,Center for Integrative Brain Research, Seattle Children's Research Institute, Seattle, WA, USA
| | | | - Ian A Glass
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA.,Brotman Baty Institute for Precision Medicine, Seattle, WA, USA.,Center for Integrative Brain Research, Seattle Children's Research Institute, Seattle, WA, USA
| | - Cole Trapnell
- Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, USA. .,Brotman Baty Institute for Precision Medicine, Seattle, WA, USA.,Allen Discovery Center for Cell Lineage Tracing, Seattle, WA, USA
| | - Jay Shendure
- Department of Genome Sciences, University of Washington School of Medicine, Seattle, WA, USA. .,Brotman Baty Institute for Precision Medicine, Seattle, WA, USA.,Allen Discovery Center for Cell Lineage Tracing, Seattle, WA, USA.,Howard Hughes Medical Institute, Seattle, WA, USA
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27
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Oliveira-Valença VM, Bosco A, Vetter ML, Silveira MS. On the Generation and Regeneration of Retinal Ganglion Cells. Front Cell Dev Biol 2020; 8:581136. [PMID: 33043015 PMCID: PMC7527462 DOI: 10.3389/fcell.2020.581136] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/07/2020] [Accepted: 08/28/2020] [Indexed: 01/02/2023] Open
Abstract
Retinal development follows a conserved neurogenic program in vertebrates to orchestrate the generation of specific cell types from multipotent progenitors in sequential but overlapping waves. In this program, retinal ganglion cells (RGCs) are the first cell type generated. RGCs are the final output neurons of the retina and are essential for vision and circadian rhythm. Key molecular steps have been defined in multiple vertebrate species to regulate competence, specification, and terminal differentiation of this cell type. This involves neuronal-specific transcription factor networks, regulators of chromatin dynamics and miRNAs. In mammals, RGCs and their optic nerve axons undergo neurodegeneration and loss in glaucoma and other optic neuropathies, resulting in irreversible vision loss. The incapacity of RGCs and axons to regenerate reinforces the need for the design of efficient RGC replacement strategies. Here we describe the essential molecular pathways for the differentiation of RGCs in vertebrates, as well as experimental manipulations that extend the competence window for generation of this early cell type from late progenitors. We discuss recent advances in regeneration of retinal neurons in vivo in both mouse and zebrafish and discuss possible strategies and barriers to achieving RGC regeneration as a therapeutic approach for vision restoration in blinding diseases such as glaucoma.
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Affiliation(s)
- Viviane M Oliveira-Valença
- Laboratory of Neurogenesis, Neurobiology Program, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil
| | - Alejandra Bosco
- Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, UT, United States
| | - Monica L Vetter
- Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, UT, United States
| | - Mariana S Silveira
- Laboratory of Neurogenesis, Neurobiology Program, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro, Brazil.,Glial Cell Biology Group, Instituto de Investigação e Inovação em Saúde, Universidade do Porto, Porto, Portugal
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28
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de Melo Reis RA, Freitas HR, de Mello FG. Cell Calcium Imaging as a Reliable Method to Study Neuron-Glial Circuits. Front Neurosci 2020; 14:569361. [PMID: 33122991 PMCID: PMC7566175 DOI: 10.3389/fnins.2020.569361] [Citation(s) in RCA: 32] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/03/2020] [Accepted: 08/24/2020] [Indexed: 12/12/2022] Open
Abstract
Complex dynamic cellular networks have been studied in physiological and pathological processes under the light of single-cell calcium imaging (SCCI), a method that correlates functional data based on calcium shifts operated by different intracellular and extracellular mechanisms integrated with their cell phenotypes. From the classic synaptic structure to tripartite astrocytic model or the recent quadripartite microglia added ensemble, as well as other physiological tissues, it is possible to follow how cells signal spatiotemporally to cellular patterns. This methodology has been used broadly due to the universal properties of calcium as a second messenger. In general, at least two types of receptor operate through calcium permeation: a fast-acting ionotropic receptor channel and a slow-activating metabotropic receptor, added to exchangers/transporters/pumps and intracellular Ca2+ release activated by messengers. These prototypes have gained an enormous amount of information in dynamic signaling circuits. SCCI has also been used as a method to associate phenotypic markers during development and stage transitions in progenitors, stem, vascular cells, neuro- and glioblasts, neurons, astrocytes, oligodendrocytes, and microglia that operate through ion channels, transporters, and receptors. Also, cancer cells or inducible cell lines from human organoids characterized by transition stages are currently being used to model diseases or reconfigure healthy cells in terms of the expression of calcium-binding/permeable molecules and shed light on therapy.
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Affiliation(s)
- Ricardo Augusto de Melo Reis
- Laboratório de Neuroquímica, Instituto de Biofísica Carlos Chagas Filho, CCS, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
| | - Hércules Rezende Freitas
- Department of Pathology and Laboratory Medicine, MIND Institute, University of California, Davis, Sacramento, CA, United States
| | - Fernando Garcia de Mello
- Laboratório de Neuroquímica, Instituto de Biofísica Carlos Chagas Filho, CCS, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Brazil
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29
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Rzhanova LA, Kuznetsova AV, Aleksandrova MA. Reprogramming of Differentiated Mammalian and Human Retinal Pigment Epithelium: Current Achievements and Prospects. Russ J Dev Biol 2020. [DOI: 10.1134/s1062360420040062] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022]
Abstract
Abstract
Impairment of the homeostatic and functional integrity of the retina and retinal pigment epithelium (RPE) is the main cause of some degenerative diseases of the human eye, which are accompanied by loss of eyesight. Despite the significant progress made over the past decades in the development of new methods for treatment for this pathology, there are still several complications when using surgical methods for correction of eyesight and so far insurmountable limitations in the applications of modern approaches, such as gene therapy and genetic engineering. One of the promising approaches to the treatment of degenerative diseases of the retina may be an approach based on the application of regenerative capacities of its endogenous cells with high plasticity, in particular, of RPE cells and Müller glia. Currently, vertebrate RPE cells are of great interest as a source of new photoreceptors and other neurons in the degrading retina in vivo. In this regard, the possibilities of their direct reprogramming by genetic, epigenetic, and chemical methods and their combination are being investigated. This review focuses on research in gene-directed reprogramming of vertebrate RPE cells into retinal neurons, with detailed analysis of the genes used as the main reprogramming factors, comparative analysis, and extrapolation of experimental data from animals to humans. Also, this review covers studies on the application of alternative approaches to gene-directed reprogramming, such as chemical-mediated reprogramming with the use of cocktails of therapeutic low-molecular-weight compounds and microRNAs. In general, the research results indicate the complexity of the process for direct reprogramming of human RPE cells into retinal neurons. However, taking into account the results of direct reprogramming of vertebrate cells and the accessibility of human RPE cells for various vectors that deliver a variety of molecules to cells, such as transcription factors, chimeric endonucleases, recombinant proteins, and low-weight molecular compounds, the most optimal combination of factors for the successful conversion of human RPE cells to retinal neurons can be suggested.
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30
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García-García D, Locker M, Perron M. Update on Müller glia regenerative potential for retinal repair. Curr Opin Genet Dev 2020; 64:52-59. [PMID: 32619816 DOI: 10.1016/j.gde.2020.05.025] [Citation(s) in RCA: 22] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2020] [Revised: 05/08/2020] [Accepted: 05/24/2020] [Indexed: 12/20/2022]
Abstract
Retinal regeneration efficiency from Müller glia varies tremendously among vertebrate species, being extremely limited in mammals. Efforts towards the identification of molecular mechanisms underlying Müller cell proliferative and neurogenic potential should help finding strategies to awake them and ensure regeneration in mammals. We provide here an update on the most recent and original progresses made in the field. These include remarkable discoveries regarding (i) unprecedented cross-species comparison of Müller cell transcriptome using single-cell technologies, (ii) the identification of new strategies to promote both the proliferative and the neurogenic potential of mammalian Müller cells, (iii) the role of the epigenome in regulating Müller glia plasticity, (iv) miRNA-based regulatory mechanisms of Müller cell response to injury, and (v) the influence of inflammatory signals on the regenerative process.
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Affiliation(s)
- Diana García-García
- Université Paris-Saclay, CNRS, Retina France, Institut des Neurosciences Paris Saclay, Orsay, France
| | - Morgane Locker
- Université Paris-Saclay, CNRS, Retina France, Institut des Neurosciences Paris Saclay, Orsay, France
| | - Muriel Perron
- Université Paris-Saclay, CNRS, Retina France, Institut des Neurosciences Paris Saclay, Orsay, France.
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31
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Neuronal Reprogramming for Tissue Repair and Neuroregeneration. Int J Mol Sci 2020; 21:ijms21124273. [PMID: 32560072 PMCID: PMC7352898 DOI: 10.3390/ijms21124273] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/08/2020] [Revised: 06/08/2020] [Accepted: 06/08/2020] [Indexed: 02/07/2023] Open
Abstract
Stem cell and cell reprogramming technology represent a rapidly growing field in regenerative medicine. A number of novel neural reprogramming methods have been established, using pluripotent stem cells (PSCs) or direct reprogramming, to efficiently derive specific neuronal cell types for therapeutic applications. Both in vitro and in vivo cellular reprogramming provide diverse therapeutic pathways for modeling neurological diseases and injury repair. In particular, the retina has emerged as a promising target for clinical application of regenerative medicine. Herein, we review the potential of neuronal reprogramming to develop regenerative strategy, with a particular focus on treating retinal degenerative diseases and discuss future directions and challenges in the field.
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32
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Yuan R, Yang M, Fan W, Lan J, Zhou YG. Paired Immunoglobulin-like Receptor B Inhibition in Müller Cells Promotes Neurite Regeneration After Retinal Ganglion Cell Injury in vitro. Neurosci Bull 2020; 36:972-984. [PMID: 32445021 DOI: 10.1007/s12264-020-00510-w] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/22/2019] [Accepted: 01/31/2020] [Indexed: 02/07/2023] Open
Abstract
In the central nervous system (CNS), three types of myelin-associated inhibitors (MAIs) have major inhibitory effects on nerve regeneration. They include Nogo-A, myelin-associated glycoprotein, and oligodendrocyte-myelin glycoprotein. MAIs possess two co-receptors, Nogo receptor (NgR) and paired immunoglobulin-like receptor B (PirB). Previous studies have confirmed that the inhibition of NgR only results in a modest increase in regeneration in the CNS; however, the inhibitory effects of PirB with regard to nerve regeneration after binding to MAIs remain controversial. In this study, we demonstrated that PirB is expressed in primary cultures of retinal ganglion cells (RGCs), and the inhibitory effects of the three MAIs on the growth of RGC neurites are not significantly decreased after direct PirB knockdown using adenovirus PirB shRNA. Interestingly, we found that retinal Müller cells expressed PirB and that its knockdown enhanced the regeneration of co-cultured RGC neurites. PirB knockdown also activated the JAK/Stat3 signaling pathway in Müller cells and upregulated ciliary neurotrophic factor levels. These findings indicate that PirB plays a novel role in retinal Müller cells and that its action in these cells may indirectly affect the growth of RGC neurites. The results also reveal that PirB in Müller cells affects RGC neurite regeneration. Our findings provide a novel basis for the use of PirB as a target molecule to promote nerve regeneration.
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Affiliation(s)
- Rongdi Yuan
- Department of Ophthalmology, Xinqiao Hospital, Army Medical University, Chongqing, 400037, China.,The Molecular Biology Centre, State Key Laboratory of Trauma, Burn and Combined Injury, Research Institute of Surgery and Daping Hospital, Army Medical University, Chongqing, 400042, China
| | - Mei Yang
- Department of Ophthalmology, Xinqiao Hospital, Army Medical University, Chongqing, 400037, China
| | - Wei Fan
- Department of Ophthalmology, Xinqiao Hospital, Army Medical University, Chongqing, 400037, China
| | - Jian Lan
- Department of Ophthalmology, Xinqiao Hospital, Army Medical University, Chongqing, 400037, China
| | - Yuan-Guo Zhou
- The Molecular Biology Centre, State Key Laboratory of Trauma, Burn and Combined Injury, Research Institute of Surgery and Daping Hospital, Army Medical University, Chongqing, 400042, China.
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33
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El Wazan L, Urrutia-Cabrera D, Wong RCB. Using transcription factors for direct reprogramming of neurons in vitro. World J Stem Cells 2019; 11:431-444. [PMID: 31396370 PMCID: PMC6682505 DOI: 10.4252/wjsc.v11.i7.431] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 02/22/2019] [Revised: 06/07/2019] [Accepted: 06/27/2019] [Indexed: 02/06/2023] Open
Abstract
Cell therapy offers great promises in replacing the neurons lost due to neurodegenerative diseases or injuries. However, a key challenge is the cellular source for transplantation which is often limited by donor availability. Direct reprogramming provides an exciting avenue to generate specialized neuron subtypes in vitro, which have the potential to be used for autologous transplantation, as well as generation of patient-specific disease models in the lab for drug discovery and testing gene therapy. Here we present a detailed review on transcription factors that promote direct reprogramming of specific neuronal subtypes with particular focus on glutamatergic, GABAergic, dopaminergic, sensory and retinal neurons. We will discuss the developmental role of master transcriptional regulators and specification factors for neuronal subtypes, and summarize their use in promoting direct reprogramming into different neuronal subtypes. Furthermore, we will discuss up-and-coming technologies that advance the cell reprogramming field, including the use of computational prediction of reprogramming factors, opportunity of cellular reprogramming using small chemicals and microRNA, as well as the exciting potential for applying direct reprogramming in vivo as a novel approach to promote neuro-regeneration within the body. Finally, we will highlight the clinical potential of direct reprogramming and discuss the hurdles that need to be overcome for clinical translation.
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Affiliation(s)
- Layal El Wazan
- Cellular Reprogramming Unit, Centre for Eye Research Australia, Melbourne 3004, Australia
| | - Daniel Urrutia-Cabrera
- Cellular Reprogramming Unit, Centre for Eye Research Australia, Melbourne 3004, Australia
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