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Xu H, Cao L, Chen Y, Zhou C, Xu J, Zhang Z, Li X, Liu L, Lu J. Single-cell RNA sequencing reveals the heterogeneity and interactions of immune cells and Müller glia during zebrafish retina regeneration. Neural Regen Res 2025; 20:3635-3648. [PMID: 38934409 PMCID: PMC11974639 DOI: 10.4103/nrr.nrr-d-23-02083] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/26/2023] [Revised: 04/17/2024] [Accepted: 05/24/2024] [Indexed: 06/28/2024] Open
Abstract
JOURNAL/nrgr/04.03/01300535-202512000-00031/figure1/v/2025-01-31T122243Z/r/image-tiff Inflammation plays a crucial role in the regeneration of fish and avian retinas. However, how inflammation regulates Müller glia (MG) reprogramming remains unclear. Here, we used single-cell RNA sequencing to investigate the cell heterogeneity and interactions of MG and immune cells in the regenerating zebrafish retina. We first showed that two types of quiescent MG (resting MG1 and MG2) reside in the uninjured retina. Following retinal injury, resting MG1 transitioned into an activated state expressing known reprogramming genes, while resting MG2 gave rise to rod progenitors. We further showed that retinal microglia can be categorized into three subtypes (microglia-1, microglia-2, and proliferative) and pseudotime analysis demonstrated dynamic changes in microglial status following retinal injury. Analysis of cell-cell interactions indicated extensive crosstalk between immune cells and MG, with many interactions shared among different immune cell types. Finally, we showed that inflammation activated Jak1-Stat3 signaling in MG, promoting their transition from a resting to an activated state. Our study reveals the cell heterogeneity and crosstalk of immune cells and MG in zebrafish retinal repair, and may provide valuable insights into future mammalian retina regeneration.
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Affiliation(s)
- Hui Xu
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Lining Cao
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
| | - Yuxi Chen
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Cuiping Zhou
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Jie Xu
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Zhuolin Zhang
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Xiangyu Li
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Lihua Liu
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
| | - Jianfeng Lu
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
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Bhattacharya S, Deka J, Avallone T, Todd L. The neuroimmune interface in retinal regeneration. Prog Retin Eye Res 2025; 106:101361. [PMID: 40287050 DOI: 10.1016/j.preteyeres.2025.101361] [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: 02/28/2025] [Revised: 04/12/2025] [Accepted: 04/23/2025] [Indexed: 04/29/2025]
Abstract
Retinal neurodegeneration leads to irreversible blindness due to the mammalian nervous system's inability to regenerate lost neurons. Efforts to regenerate retina involve two main strategies: stimulating endogenous cells to reprogram into neurons or transplanting stem-cell derived neurons into the degenerated retina. However, both approaches must overcome a major barrier in getting new neurons to grow back down the optic nerve and connect to appropriate visual targets in environments that differ significantly from developmental conditions. While immune privilege has historically been associated with the central nervous system, an emerging literature highlights the active role of immune cells in shaping neurodegeneration and regeneration. This review explores the neuroimmune interface in retinal repair, dissecting how immune interactions influence glial reprogramming, transplantation outcomes, and axonal regeneration. By integrating insights from regenerative species with mammalian models, we highlight novel immunomodulatory strategies to optimize retinal regeneration.
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Affiliation(s)
- Sucheta Bhattacharya
- Department of Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, NY, 13210, USA; Department of Neuroscience and Physiology, SUNY Upstate Medical University, Syracuse, NY, 13210, USA
| | - Jugasmita Deka
- Department of Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, NY, 13210, USA; Department of Neuroscience and Physiology, SUNY Upstate Medical University, Syracuse, NY, 13210, USA
| | - Thomas Avallone
- Department of Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, NY, 13210, USA
| | - Levi Todd
- Department of Ophthalmology and Visual Sciences, SUNY Upstate Medical University, Syracuse, NY, 13210, USA; Department of Neuroscience and Physiology, SUNY Upstate Medical University, Syracuse, NY, 13210, USA.
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Norrie JL, Lupo MS, Little DR, Shirinifard A, Mishra A, Zhang Q, Geiger N, Putnam D, Djekidel N, Ramirez C, Xu B, Dundee JM, Yu J, Chen X, Dyer MA. Latent epigenetic programs in Müller glia contribute to stress and disease response in the retina. Dev Cell 2025; 60:1199-1216.e7. [PMID: 39753128 PMCID: PMC12014377 DOI: 10.1016/j.devcel.2024.12.014] [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: 10/31/2023] [Revised: 07/09/2024] [Accepted: 12/06/2024] [Indexed: 04/24/2025]
Abstract
Previous studies have demonstrated the dynamic changes in chromatin structure during retinal development correlate with changes in gene expression. However, those studies lack cellular resolution. Here, we integrate single-cell RNA sequencing (scRNA-seq) and single-cell assay for transposase-accessible chromatin sequencing (scATAC-seq) with bulk data to identify cell-type-specific changes in chromatin structure during human and murine development. Although promoter activity is correlated with chromatin accessibility, we discovered several hundred genes that were transcriptionally silent but had accessible chromatin at their promoters. Most of those silent/accessible gene promoters were in Müller glial cells, which function to maintain retinal homeostasis and respond to stress, injury, or disease. We refer to these as "pliancy genes" because they allow the Müller glia to rapidly change their gene expression and cellular state in response to retinal insults. The Müller glial cell pliancy program is established during development, and we demonstrate that pliancy genes are important for regulating inflammation in the murine retina in vivo.
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Affiliation(s)
- Jackie L Norrie
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Marybeth S Lupo
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Danielle R Little
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Abbas Shirinifard
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Akhilesh Mishra
- Departments of Computational Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Qiong Zhang
- Center for Applied Bioinformatics, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Natalie Geiger
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Daniel Putnam
- Departments of Computational Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Nadhir Djekidel
- Center for Applied Bioinformatics, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Cody Ramirez
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Beisi Xu
- Center for Applied Bioinformatics, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Jacob M Dundee
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Jiang Yu
- Departments of Computational Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Xiang Chen
- Departments of Computational Biology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA
| | - Michael A Dyer
- Departments of Developmental Neurobiology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
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Konar GJ, Vallone KT, Nguyen TD, Patton JG. Analysis of the senescence secretome during zebrafish retina regeneration. FRONTIERS IN AGING 2025; 6:1569422. [PMID: 40308558 PMCID: PMC12040975 DOI: 10.3389/fragi.2025.1569422] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 01/31/2025] [Accepted: 04/02/2025] [Indexed: 05/02/2025]
Abstract
Introduction Zebrafish possess the innate ability to regenerate any lost or damaged retinal cell type with Müller glia serving as resident stem cells. Recently, we discovered that this process is aided by a population of damage-induced senescent immune cells. As part of the Senescence Associated Secretory Phenotype (SASP), senescent cells secrete numerous factors that can play a role in the modulation of inflammation and remodeling of the retinal microenvironment during regeneration. However, the identity of specific SASP factors that drive initiation and progression of retina regeneration remains unclear. Materials and Methods We mined the SASP Atlas and publicly available RNAseq datasets to identify common, differentially expressed SASP factors after retina injury. These datasets included two distinct acute damage regimens, as well as two chronic, genetic models of retina degeneration. We identified overlapping factors between these models and used genetic knockdown experiments, qRT/PCR and immunohistochemical staining to test a role for one of these factors (npm1a). Results We discovered an overlapping set of 31 SASP-related regeneration factors across all data sets and damage paradigms. These factors are upregulated after damage with functions that span the innate immune system, autophagic processing, cell cycle regulation, and cellular stress responses. From among these, we show that depletion of Nucleophosmin 1 (npm1a) inhibits retina regeneration and decreases senescent cell detection after damage. Discussion Our data suggest that differential expression of SASP factors promotes initiation and progression of retina regeneration after both acute and chronic retinal damage. The existence of a common, overlapping set of 31 factors provides a group of novel therapeutic targets for retina regeneration studies.
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Affiliation(s)
| | | | | | - James G. Patton
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, United States
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Yin Z, Kang J, Cheng X, Gao H, Huo S, Xu H. Investigating Müller glia reprogramming in mice: a retrospective of the last decade, and a look to the future. Neural Regen Res 2025; 20:946-959. [PMID: 38989930 PMCID: PMC11438324 DOI: 10.4103/nrr.nrr-d-23-01612] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/25/2023] [Accepted: 02/05/2024] [Indexed: 07/12/2024] Open
Abstract
Müller glia, as prominent glial cells within the retina, plays a significant role in maintaining retinal homeostasis in both healthy and diseased states. In lower vertebrates like zebrafish, these cells assume responsibility for spontaneous retinal regeneration, wherein endogenous Müller glia undergo proliferation, transform into Müller glia-derived progenitor cells, and subsequently regenerate the entire retina with restored functionality. Conversely, Müller glia in the mouse and human retina exhibit limited neural reprogramming. Müller glia reprogramming is thus a promising strategy for treating neurodegenerative ocular disorders. Müller glia reprogramming in mice has been accomplished with remarkable success, through various technologies. Advancements in molecular, genetic, epigenetic, morphological, and physiological evaluations have made it easier to document and investigate the Müller glia programming process in mice. Nevertheless, there remain issues that hinder improving reprogramming efficiency and maturity. Thus, understanding the reprogramming mechanism is crucial toward exploring factors that will improve Müller glia reprogramming efficiency, and for developing novel Müller glia reprogramming strategies. This review describes recent progress in relatively successful Müller glia reprogramming strategies. It also provides a basis for developing new Müller glia reprogramming strategies in mice, including epigenetic remodeling, metabolic modulation, immune regulation, chemical small-molecules regulation, extracellular matrix remodeling, and cell-cell fusion, to achieve Müller glia reprogramming in mice.
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Affiliation(s)
- Zhiyuan Yin
- Key Lab of Visual Damage and Regeneration & Restoration of Chongqing, Southwest Eye Hospital, Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, China
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Xu J, Li Y, Li X, Tan X, Liu L, Cao L, Xu H. Microglia-Derived IL-6 Promotes Müller Glia Reprogramming and Proliferation in Zebrafish Retina Regeneration. Invest Ophthalmol Vis Sci 2025; 66:67. [PMID: 40266594 PMCID: PMC12025339 DOI: 10.1167/iovs.66.4.67] [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: 10/29/2024] [Accepted: 03/27/2025] [Indexed: 04/24/2025] Open
Abstract
Purpose Inflammation activates the Jak1-Stat3 signaling pathway in zebrafish Müller glia (MG), leading to their status transition and proliferation following retinal injury. However, the source of Stat3-activating molecules remains unclear. This study aims to explore the expression and function of a Stat3-activating cytokine IL-6 in zebrafish retina regeneration. Methods Mechanical retinal injury was induced in adult zebrafish by a needle-poke lesion. Single-cell RNA sequencing (scRNAseq) and PCR were used to determine gene expression. Microglia ablation was performed by using the mpeg1:nsfb-mcherry transgenic zebrafish. Morpholino oligonucleotides, a recombinant zebrafish IL-6 protein and drugs, were used to manipulate IL-6 or Stat3 signaling in the retina. The 5-Ethynyl-2'-deoxyuridine (EdU) labeling was used to evaluate MG proliferation and the formation of MG-derived progenitor cells (MGPCs). Neuronal regeneration in the retina was analyzed by lineage tracing and immunostaining. Results The scRNAseq reveals that IL-6 is mainly expressed by a subset of pro-inflammatory microglia in the injured retina. Loss- and gain-of-function experiments demonstrate that IL-6 signaling promotes MG proliferation and the formation of MGPCs following retinal injury. Additionally, IL-6 facilitates MG status transition by modulating Jak1-Stat3 signaling and the expression of regeneration-associated genes. Interestingly, IL-6 may also regulate MGPC formation via phase-dependent pro-inflammatory and anti-inflammatory mechanisms. Finally, IL-6 promotes the early differentiation of MGPCs and contributes to the regeneration of retinal neurons in the injured retina. Conclusions Our study unveils the critical role of microglia-derived IL-6 in zebrafish retina regeneration, with potential implications for mammalian MG reprogramming.
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Affiliation(s)
- Jie Xu
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Yi Li
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Xiangyu Li
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Xuan Tan
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
| | - Lihua Liu
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
| | - Lining Cao
- Shanghai YangZhi Rehabilitation Hospital (Shanghai Sunshine Rehabilitation Center), Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai, China
| | - Hui Xu
- Key Lab of Neuroregeneration of Jiangsu and Ministry of Education, Co-innovation Center of Neuroregeneration, NMPA Key Laboratory for Research and Evaluation of Tissue Engineering Technology Products, Nantong University, Nantong, Jiangsu Province, China
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Lee EJ, Kim M, Park S, Shim JH, Cho HJ, Park JA, Park K, Lee D, Kim JH, Jeong H, Matsuzaki F, Kim SY, Kim J, Yang H, Lee JS, Kim JW. Restoration of retinal regenerative potential of Müller glia by disrupting intercellular Prox1 transfer. Nat Commun 2025; 16:2928. [PMID: 40133314 PMCID: PMC11937340 DOI: 10.1038/s41467-025-58290-8] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/09/2024] [Accepted: 03/17/2025] [Indexed: 03/27/2025] Open
Abstract
Individuals with retinal degenerative diseases struggle to restore vision due to the inability to regenerate retinal cells. Unlike cold-blooded vertebrates, mammals lack Müller glia (MG)-mediated retinal regeneration, indicating the limited regenerative capacity of mammalian MG. Here, we identify prospero-related homeobox 1 (Prox1) as a key factor restricting this process. Prox1 accumulates in MG of degenerating human and mouse retinas but not in regenerating zebrafish. In mice, Prox1 in MG originates from neighboring retinal neurons via intercellular transfer. Blocking this transfer enables MG reprogramming into retinal progenitor cells in injured mouse retinas. Moreover, adeno-associated viral delivery of an anti-Prox1 antibody, which sequesters extracellular Prox1, promotes retinal neuron regeneration and delays vision loss in a retinitis pigmentosa model. These findings establish Prox1 as a barrier to MG-mediated regeneration and highlight anti-Prox1 therapy as a promising strategy for restoring retinal regeneration in mammals.
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Affiliation(s)
- Eun Jung Lee
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- Celliaz Ltd., Daejeon, South Korea
| | - Museong Kim
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Sooyeon Park
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- Celliaz Ltd., Daejeon, South Korea
| | | | - Hyun-Ju Cho
- Microbiome Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea
- KRIBB School, University of Science and Technology, Daejeon, South Korea
| | | | - Kihyun Park
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Dongeun Lee
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Jeong Hwan Kim
- Korea Bioinformation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea
| | - Haeun Jeong
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Fumio Matsuzaki
- Laboratory for Cell Asymmetry, RIKEN Centre for Biosystems Dynamics Research, Kobe, Hyogo, Japan
- Department of Aging Science and Medicine, Graduate School of Medicine, Kyoto University, Kyoto, Japan
| | - Seon-Young Kim
- Korea Bioinformation Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea
| | - Jaehoon Kim
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Hanseul Yang
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea
| | - Jeong-Soo Lee
- Microbiome Convergence Research Center, Korea Research Institute of Bioscience and Biotechnology, Daejeon, South Korea
- KRIBB School, University of Science and Technology, Daejeon, South Korea
| | - Jin Woo Kim
- Department of Biological Sciences, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea.
- KAIST Stem Cell Research Center, Korea Advanced Institute of Science and Technology (KAIST), Daejeon, South Korea.
- Celliaz Ltd., Daejeon, South Korea.
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Wu J, Xu M, Qin C, Guo Y, Wang Y, Wang Z, Li Q. Joint profiling of DNA methylomics and transcriptomic reveals roles of demethylation in regeneration of coelomocytes after evisceration in sea cucumber Apostichopus japonicus. COMPARATIVE BIOCHEMISTRY AND PHYSIOLOGY. PART D, GENOMICS & PROTEOMICS 2025; 55:101456. [PMID: 40015132 DOI: 10.1016/j.cbd.2025.101456] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 01/08/2025] [Revised: 02/21/2025] [Accepted: 02/22/2025] [Indexed: 03/01/2025]
Abstract
Generally, there is a large number of deaths for sea cucumber along with evisceration after which a weakly immune state occurs because of a poor resistance against a variety of pathogens and environmental stress. The regeneration of coelomocytes plays an important role in the remodeling of the immune defense system after evisceration with the decrease of methylation modification. In this study, the whole DNA methylation of coelomocytes is detected post evisceration in Apostichopus japonicus to explore the process of cell regeneration. Our results found that total methylation level reached a lowest point at 12 h (9.8 %), which was followed by increased at 24 h. The transcriptomic and DNA methylomic analysis indicated a total of 215 genes were identified by selecting the significant hypomethylation within the 2-kilobase region of the transcriptional start site upstream. The KEGG pathway enrichment analyses of the 215 genes showed that signal transduction, signaling molecules and interaction were significantly enriched. The genes were enriched in the top 20 signaling pathway, such as IGF1R, Notch2 and HSPA1s. Taken together, this study provides new clues for deciphering the coelomocytes regeneration after evisceration of A. japonicus by DNA demethylation.
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Affiliation(s)
- Jiong Wu
- Tianjin Key Lab of Aqua-ecology and Aquaculture, Fisheries College, Tianjin Agricultural University, Tianjin 300384, China; Key Laboratory of Efficient Utilization and Processing of Marine Fishery Resources of Hainan Province, Sanya Tropical Fisheries Research Institute, Sanya 572426, China
| | - Mingmei Xu
- Tianjin Key Lab of Aqua-ecology and Aquaculture, Fisheries College, Tianjin Agricultural University, Tianjin 300384, China
| | - Chuanxin Qin
- Key Laboratory of Efficient Utilization and Processing of Marine Fishery Resources of Hainan Province, Sanya Tropical Fisheries Research Institute, Sanya 572426, China
| | - Yu Guo
- Key Laboratory of Efficient Utilization and Processing of Marine Fishery Resources of Hainan Province, Sanya Tropical Fisheries Research Institute, Sanya 572426, China
| | - Yinan Wang
- Tianjin Key Lab of Aqua-ecology and Aquaculture, Fisheries College, Tianjin Agricultural University, Tianjin 300384, China
| | - Zhenhui Wang
- Yancheng Institute of Technology Department: College of Marine and Bioengineering, Yancheng 224051, China.
| | - Qiang Li
- Tianjin Key Lab of Aqua-ecology and Aquaculture, Fisheries College, Tianjin Agricultural University, Tianjin 300384, China.
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Suzuki R, Katada Y, Fujii M, Serizawa N, Negishi K, Kurihara T. Tropism of the AAV6.2 Vector in the Murine Retina. Int J Mol Sci 2025; 26:1580. [PMID: 40004046 PMCID: PMC11855373 DOI: 10.3390/ijms26041580] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/10/2024] [Revised: 02/05/2025] [Accepted: 02/06/2025] [Indexed: 02/27/2025] Open
Abstract
Retinitis pigmentosa (RP) is a progressive inherited retinal dystrophy (IRD) that primarily affects rod photoreceptor cells, leading to the degeneration of photoreceptors and the gradual loss of vision. While RP is one of the most studied IRDs, other neurodegenerative diseases affecting the retina and optic nerve, such as glaucoma, also involve common mechanisms of cellular stress and degeneration. Current therapeutic approaches under investigation include gene therapy, retina prosthesis, and neuroprotection. Among these approaches, gene therapy has shown promise, though challenges related to viral vector tropism and transduction efficiency persist. The adeno-associated virus (AAV) vector is commonly employed for gene delivery, but novel serotypes and engineered variants are being explored to improve specificity and efficacy. This study evaluates the gene transfer efficiency of the AAV6.2 vector following intravitreal injection into the murine retina. Male C57BL/6 mice (9 weeks old) were intravitreally injected with 1 µL of AAV2-CMV-EGFP, AAV6-CMV-EGFP, or AAV6.2-CMV-EGFP at a titer of 3.2 × 1012 vg/mL per eye. Retinal transduction was assessed using in vivo fluorescence imaging, flat-mount imaging, and immunohistochemistry. EGFP expression in retinal ganglion cells, Müller cells, amacrine cells, and bipolar cells was quantitatively analyzed. All three AAV serotypes effectively transduced retinal ganglion cells, but AAV6.2 exhibited enhanced transduction in Müller cells and other neuronal retinal cells, including bipolar and amacrine cells. AAV6.2 demonstrated more localized expression around retinal blood vessels compared to the diffuse expression observed with AAV2. Immunohistochemical analysis revealed that AAV6.2 had significantly higher transduction efficiency in Müller cells (p < 0.001) compared to AAV2 and AAV6. AAV6.2 shows superior transduction efficiency in Müller cells, positioning it as a promising vector for gene therapies targeting retinal degenerative diseases such as RP. Its ability to effectively transduce Müller cells suggests potential applications in neuroprotection and gene replacement therapies.
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Affiliation(s)
- Ryo Suzuki
- Laboratory of Photobiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-0016, Japan; (R.S.); (Y.K.); (M.F.); (N.S.)
| | - Yusaku Katada
- Laboratory of Photobiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-0016, Japan; (R.S.); (Y.K.); (M.F.); (N.S.)
- Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Momo Fujii
- Laboratory of Photobiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-0016, Japan; (R.S.); (Y.K.); (M.F.); (N.S.)
- Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Naho Serizawa
- Laboratory of Photobiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-0016, Japan; (R.S.); (Y.K.); (M.F.); (N.S.)
- Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Kazuno Negishi
- Department of Ophthalmology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-8582, Japan;
| | - Toshihide Kurihara
- Laboratory of Photobiology, Keio University School of Medicine, Shinjuku-ku, Tokyo 160-0016, Japan; (R.S.); (Y.K.); (M.F.); (N.S.)
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10
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Wohlschlegel J, Kierney F, Arakelian KL, Luxardi G, Suvarnpradip N, Hoffer D, Rieke F, Moshiri A, Reh TA. Stimulating the regenerative capacity of the human retina with proneural transcription factors in 3D cultures. Proc Natl Acad Sci U S A 2025; 122:e2417228122. [PMID: 39823300 PMCID: PMC11759899 DOI: 10.1073/pnas.2417228122] [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: 08/23/2024] [Accepted: 12/07/2024] [Indexed: 01/19/2025] Open
Abstract
Retinal diseases often lead to degeneration of specific retinal cell types with currently limited therapeutic options to replace the lost neurons. Previous studies have reported that overexpression of ASCL1 or combinations of proneural factors in Müller glia (MG) induce regeneration of functional neurons in the adult mouse retina. Recently, we applied the same strategy in dissociated cultures of fetal human MG and although we stimulated neurogenesis from MG, our effect in 2D cultures was modest and our analysis of newborn neurons was limited. In this study, we aimed to improve our MG reprogramming strategy in a more intact retinal environment. For this purpose, we used an in vitro culture system of human fetal retinal tissue and adult human postmortem retina. To stimulate reprogramming, we used lentiviral vectors to deliver constructs with a glial-specific promoter (HES1) driving ASCL1 alone or in combination with additional developmental transcription factors (TFs) such as ATOH1 and NEUROD1. Combining IHC, scRNA-seq, and electrophysiology, we show that human MG can generate new neurons even in adults. This work constitutes a key step toward a future clinical application of this regenerative medicine approach for retinal degenerative disorders.
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Affiliation(s)
| | - Faith Kierney
- Department of Biological Structure, University of Washington, Seattle, WA98125
| | - Kayla L. Arakelian
- Department of Biological Structure, University of Washington, Seattle, WA98125
| | - Guillaume Luxardi
- Department of Ophthalmology & Vision Science, University of California Davis School of Medicine, Sacramento, CA95616
| | - Naran Suvarnpradip
- Department of Ophthalmology & Vision Science, University of California Davis School of Medicine, Sacramento, CA95616
| | - Dawn Hoffer
- Department of Biological Structure, University of Washington, Seattle, WA98125
| | - Fred Rieke
- Department of Physiology and Biophysics, University of Washington, Seattle, WA98125
| | - Ala Moshiri
- Department of Ophthalmology & Vision Science, University of California Davis School of Medicine, Sacramento, CA95616
| | - Thomas A. Reh
- Department of Biological Structure, University of Washington, Seattle, WA98125
- Institute for Stem Cells and Regenerative Medicine, University of Washington, Seattle, WA98125
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11
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Nelson HM, Konar GJ, Patton JG. Isolation and Characterization of Extracellular Vesicles to Activate Retina Regeneration. Methods Mol Biol 2025; 2848:135-150. [PMID: 39240521 DOI: 10.1007/978-1-0716-4087-6_9] [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: 09/07/2024]
Abstract
Mammals do not possess the ability to spontaneously repair or regenerate damaged retinal tissue. In contrast to teleost fish which are capable of retina regeneration through the action of Müller glia, mammals undergo a process of reactive gliosis and scarring that inhibits replacement of lost neurons. Thus, it is important to discover novel methods for stimulating mammalian Müller glia to dedifferentiate and produce progenitor cells that can replace lost retinal neurons. Inducing an endogenous regenerative pathway mediated by Müller glia would provide an attractive alternative to stem cell injections or gene therapy approaches. Extracellular vesicles (EVs) are now recognized to serve as a novel form of cell-cell communication through the transfer of cargo from donor to recipient cells or by the activation of signaling cascades in recipient cells. EVs have been shown to promote proliferation and regeneration raising the possibility that delivery of EVs could be a viable treatment for visual disorders. Here, we provide protocols to isolate EVs for use in retina regeneration experiments.
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Affiliation(s)
- Hannah M Nelson
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Gregory J Konar
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - James G Patton
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA.
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12
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Hernández-Núñez I, Clark BS. Experimental Framework for Assessing Mouse Retinal Regeneration Through Single-Cell RNA-Sequencing. Methods Mol Biol 2025; 2848:117-134. [PMID: 39240520 DOI: 10.1007/978-1-0716-4087-6_8] [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: 09/07/2024]
Abstract
Retinal degenerative diseases including age-related macular degeneration and glaucoma are estimated to currently affect more than 14 million people in the United States, with an increased prevalence of retinal degenerations in aged individuals. An expanding aged population who are living longer forecasts an increased prevalence and economic burden of visual impairments. Improvements to visual health and treatment paradigms for progressive retinal degenerations slow vision loss. However, current treatments fail to remedy the root cause of visual impairments caused by retinal degenerations-loss of retinal neurons. Stimulation of retinal regeneration from endogenous cellular sources presents an exciting treatment avenue for replacement of lost retinal cells. In multiple species including zebrafish and Xenopus, Müller glial cells maintain a highly efficient regenerative ability to reconstitute lost cells throughout the organism's lifespan, highlighting potential therapeutic avenues for stimulation of retinal regeneration in humans. Here, we describe how the application of single-cell RNA-sequencing (scRNA-seq) has enhanced our understanding of Müller glial cell-derived retinal regeneration, including the characterization of gene regulatory networks that facilitate/inhibit regenerative responses. Additionally, we provide a validated experimental framework for cellular preparation of mouse retinal cells as input into scRNA-seq experiments, including insights into experimental design and analyses of resulting data.
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Affiliation(s)
- Ismael Hernández-Núñez
- John F Hardesty, MD Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, MO, USA
| | - Brian S Clark
- John F Hardesty, MD Department of Ophthalmology and Visual Sciences, Washington University School of Medicine, St. Louis, MO, USA.
- Department of Developmental Biology, Washington University School of Medicine, St. Louis, MO, USA.
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13
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Ceisel A, Emmerich K, McNamara G, Graziano G, Banerjee S, Reibman B, Saxena MT, Mumm JS. Automated In Vivo Phenotypic Screening Platform for Identifying Factors that Affect Cell Regeneration Kinetics. Methods Mol Biol 2025; 2848:217-247. [PMID: 39240526 DOI: 10.1007/978-1-0716-4087-6_14] [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: 09/07/2024]
Abstract
Various strategies for replacing retinal neurons lost in degenerative diseases are under investigation, including stimulating the endogenous regenerative capacity of Müller Glia (MG) as injury-inducible retinal stem cells. Inherently regenerative species, such as zebrafish, have provided key insights into mechanisms regulating MG dedifferentiation to a stem-like state and the proliferation of MG and MG-derived progenitor cells (MGPCs). Interestingly, promoting MG/MGPC proliferation is not sufficient for regeneration, yet mechanistic studies are often focused on this measure. To fully account for the regenerative process, and facilitate screens for factors regulating cell regeneration, an assay for quantifying cell replacement is required. Accordingly, we adapted an automated reporter-assisted phenotypic screening platform to quantify the pace of cellular regeneration kinetics following selective cell ablation in larval zebrafish. Here, we detail a method for using this approach to identify chemicals and genes that control the rate of retinal cell regeneration following selective retinal cell ablation.
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Affiliation(s)
- Anneliese Ceisel
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
- Cellular and Molecular Medicine Program, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Kevin Emmerich
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
- Department of Genetic Medicine, McKusick-Nathans Institute, Human Genetics Program, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - George McNamara
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Gianna Graziano
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Shreya Banerjee
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Barak Reibman
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Meera T Saxena
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Jeff S Mumm
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
- Cellular and Molecular Medicine Program, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
- Department of Genetic Medicine, McKusick-Nathans Institute, Human Genetics Program, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
- Department of Ophthalmology, Center for Nanomedicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
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14
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Kumar A, Kramer AC, Thummel R. Models of Photoreceptor Degeneration in Adult Zebrafish. Methods Mol Biol 2025; 2848:75-84. [PMID: 39240517 DOI: 10.1007/978-1-0716-4087-6_5] [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: 09/07/2024]
Abstract
Zebrafish maintain a remarkable ability to regenerate their neural retina following rapid and extensive loss of retinal neurons. This is mediated by Müller glial cells (MG), which re-enter the cell cycle to produce amplifying progenitor cells that eventually differentiate into the lost retinal neurons. For example, exposing adult albino zebrafish to intense light destroys large numbers of rod and cone photoreceptors, which are then restored by MG-mediated regeneration. Here, we describe an updated method for performing these acute phototoxic lesions to adult zebrafish retinas. Next, we contrast this method to a chronic, low light lesion model that results in a more muted and sustained damage to photoreceptors and does not trigger a MG-mediated regeneration response. Thus, these two methods can be used to compare and contrast the genetic and morphological changes associated with acute and chronic methods of photoreceptor degeneration.
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Affiliation(s)
- Arun Kumar
- Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine, Detroit, MI, USA
| | - Ashley C Kramer
- Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine, Detroit, MI, USA
| | - Ryan Thummel
- Department of Ophthalmology, Visual and Anatomical Sciences, Wayne State University School of Medicine, Detroit, MI, USA.
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15
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Rumford JE, Grieshaber A, Lewiston S, Reed JL, Long SS, Mitchell DM. Forced MyD88 signaling in microglia impacts the production and survival of regenerated retinal neurons. Front Cell Dev Biol 2024; 12:1495586. [PMID: 39633708 PMCID: PMC11614808 DOI: 10.3389/fcell.2024.1495586] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/12/2024] [Accepted: 11/07/2024] [Indexed: 12/07/2024] Open
Abstract
Inflammation and microglia appear to be key factors influencing the outcome of retinal regeneration following acute retinal damage. Despite such findings, direct connection of microglia-specific inflammatory factors as drivers of regenerative responses in the retina are still not defined, and intracellular pathways activated to stimulate such signals from microglia are currently unknown. We became interested in MyD88 regulation in microglia because transcriptomic datasets suggest myd88 could be regulated temporally in zebrafish microglia responding to damage in the central nervous system. MyD88 is an intracellular molecular adaptor that initiates signaling cascades downstream of several innate immune receptors, and probably most well-known for inducing gene expression of pro-inflammatory factors. Using zebrafish, which spontaneously regenerate retinal neurons after acute retinal damage, we studied the effects of overactivation of MyD88 signaling in microglia and macrophages on the Müller glia-mediated regenerative response. Our results indicate that increased MyD88 signaling in microglia/macrophages impacts the initial response of Müller glia entering a regenerative response after acute, neurotoxin-induced retinal damage to inner retinal neurons. In addition, increased MyD88 signaling in microglia/macrophages resulted in reduced survival of inner retinal neurons in regenerated retinas. This work supports the idea that temporal control of inflammatory signaling is a key component in the production of MG-derived progenitors yet further indicates that such control is important for differentiation and survival of regenerated neurons.
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Affiliation(s)
- Jordan E. Rumford
- Department of Biological Sciences, University of Idaho, Moscow, ID, United States
| | - Ailis Grieshaber
- Department of Biological Sciences, University of Idaho, Moscow, ID, United States
| | - Samantha Lewiston
- Department of Biological Sciences, University of Idaho, Moscow, ID, United States
| | - Jordan L. Reed
- Department of Computer Science, University of Idaho, Moscow, ID, United States
- Formerly North Idaho College, Coeur d’Alene, ID, United States
| | - Samuel S. Long
- Business and Computer Science Division, Lewis-Clark State College, Lewiston, ID, United States
| | - Diana M. Mitchell
- Department of Biological Sciences, University of Idaho, Moscow, ID, United States
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16
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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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17
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García-García D, Vidal-Gil L, Parain K, Lun J, Audic Y, Chesneau A, Siron L, Van Westendorp D, Lourdel S, Sánchez-Sáez X, Kazani D, Ricard J, Pottin S, Donval A, Bronchain O, Locker M, Roger JE, Borday C, Pla P, Bitard J, Perron M. Neuroinflammation as a cause of differential Müller cell regenerative responses to retinal injury. SCIENCE ADVANCES 2024; 10:eadp7916. [PMID: 39356769 PMCID: PMC11446274 DOI: 10.1126/sciadv.adp7916] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/11/2024] [Accepted: 08/28/2024] [Indexed: 10/04/2024]
Abstract
Unlike mammals, some nonmammalian species recruit Müller glia for retinal regeneration after injury. Identifying the underlying mechanisms may help to foresee regenerative medicine strategies. Using a Xenopus model of retinitis pigmentosa, we found that Müller cells actively proliferate upon photoreceptor degeneration in old tadpoles but not in younger ones. Differences in the inflammatory microenvironment emerged as an explanation for such stage dependency. Functional analyses revealed that enhancing neuroinflammation is sufficient to trigger Müller cell proliferation, not only in young tadpoles but also in mice. In addition, we showed that microglia are absolutely required for the response of mouse Müller cells to mitogenic factors while negatively affecting their neurogenic potential. However, both cell cycle reentry and neurogenic gene expression are allowed when applying sequential pro- and anti-inflammatory treatments. This reveals that inflammation benefits Müller glia proliferation in both regenerative and nonregenerative vertebrates and highlights the importance of sequential inflammatory modulation to create a regenerative permissive microenvironment.
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Affiliation(s)
- Diana García-García
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Lorena Vidal-Gil
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Karine Parain
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Jingxian Lun
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Yann Audic
- Univ Rennes, CNRS, IGDR (Institut de Genetique et Developpement de Rennes), Rennes, France
| | - Albert Chesneau
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Léa Siron
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Demi Van Westendorp
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Sophie Lourdel
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Xavier Sánchez-Sáez
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Despoina Kazani
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Julien Ricard
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Solène Pottin
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Alicia Donval
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Odile Bronchain
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Morgane Locker
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Jérôme E. Roger
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Caroline Borday
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Patrick Pla
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Juliette Bitard
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
| | - Muriel Perron
- Université Paris-Saclay, CNRS, Institut des Neurosciences Paris-Saclay, Saclay, France
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18
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Liu Y, Wang A, Chen C, Zhang Q, Shen Q, Zhang D, Xiao X, Chen S, Lian L, Le Z, Liu S, Liang T, Zheng Q, Xu P, Zou J. Microglial cGAS-STING signaling underlies glaucoma pathogenesis. Proc Natl Acad Sci U S A 2024; 121:e2409493121. [PMID: 39190350 PMCID: PMC11388346 DOI: 10.1073/pnas.2409493121] [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: 05/17/2024] [Accepted: 07/24/2024] [Indexed: 08/28/2024] Open
Abstract
Characterized by progressive degeneration of retinal ganglion cells (RGCs) and vision loss, glaucoma is the primary cause of irreversible blindness, incurable and affecting over 78 million patients. However, pathogenic mechanisms leading to glaucoma-induced RGC loss are incompletely understood. Unexpectedly, we found that cGAS-STING (2'3'-cyclic GMP-AMP-stimulator of interferon genes) signaling, which surveils displaced double-stranded DNA (dsDNA) in the cytosol and initiates innate immune responses, was robustly activated during glaucoma in retinal microglia in distinct murine models. Global or microglial deletion of STING markedly relieved glaucoma symptoms and protected RGC degeneration and vision loss, while mice bearing genetic cGAS-STING supersensitivity aggravated retinal neuroinflammation and RGC loss. Mechanistically, dsDNA from tissue injury activated microglial cGAS-STING signaling, causing deleterious macroglia reactivity in retinas by cytokine-mediated microglia-macroglia interactions, progressively driving apoptotic death of RGCs. Remarkably, preclinical investigations of targeting cGAS-STING signaling by intraocular injection of TBK1i or anti-IFNAR1 antibody prevented glaucoma-induced losses of RGCs and vision. Therefore, we unravel an essential role of cGAS-STING signaling underlying glaucoma pathogenesis and suggest promising therapeutic strategies for treating this devastating disease.
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Affiliation(s)
- Yutong Liu
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Ailian Wang
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Chen Chen
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Qian Zhang
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Qin Shen
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
| | - Dan Zhang
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
| | - Xueqi Xiao
- Eye Center of the Second Affiliated Hospital School of Medicine, Center for Genetic Medicine, Zhejiang University International Institute of Medicine, Hangzhou310029, China
| | - Shasha Chen
- College of Life and Environmental Science, Wenzhou University, Wenzhou325035, China
| | - Lili Lian
- National Clinical Research Center for Ocular Diseases, School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, Wenzhou325027, China
| | - Zhenmin Le
- National Clinical Research Center for Ocular Diseases, School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, Wenzhou325027, China
| | - Shengduo Liu
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Tingbo Liang
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
| | - Qinxiang Zheng
- National Clinical Research Center for Ocular Diseases, School of Ophthalmology and Optometry, Eye Hospital, Wenzhou Medical University, Wenzhou325027, China
| | - Pinglong Xu
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Institute of Intelligent Medicine, Zhejiang University-Hangzhou Global Scientific and Technological Innovation Center, Hangzhou310058, China
- Ministry of Education Laboratory of Biosystems Homeostasis and Protection, Zhejiang Provincial Key Laboratory for Cancer Molecular Cell Biology, Life Sciences Institute, Zhejiang University, Hangzhou310058, China
| | - Jian Zou
- Department of Hepatobiliary and Pancreatic Surgery and Zhejiang Provincial Key Laboratory of Pancreatic Disease, The First Affiliated Hospital, University School of Medicine, Zhejiang University, Hangzhou310058, China
- Eye Center of the Second Affiliated Hospital School of Medicine, Center for Genetic Medicine, Zhejiang University International Institute of Medicine, Hangzhou310029, China
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19
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Song P, Parsana D, Singh R, Pollock LM, Anand-Apte B, Perkins BD. Photoreceptor regeneration occurs normally in microglia-deficient irf8 mutant zebrafish following acute retinal damage. Sci Rep 2024; 14:20146. [PMID: 39209978 PMCID: PMC11362524 DOI: 10.1038/s41598-024-70859-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2024] [Accepted: 08/21/2024] [Indexed: 09/04/2024] Open
Abstract
Microglia are resident immune cells in the central nervous system, including the retina that surveil the environment for damage and infection. Following retinal damage, microglia undergo morphological changes, migrate to the site of damage, and express and secrete pro-inflammatory signals. In the zebrafish retina, inflammation induces the reprogramming and proliferation of Müller glia and the regeneration of neurons following damage or injury. Immunosuppression or pharmacological ablation of microglia reduce or abolish Müller glia proliferation. We evaluated the retinal architecture and retinal regeneration in adult zebrafish irf8 mutants, which have significantly depleted numbers of microglia. We show that irf8 mutants have normal retinal structure at 3 months post fertilization (mpf) and 6 mpf but fewer cone photoreceptors by 10 mpf. Surprisingly, light-induced photoreceptor ablation induced Müller glia proliferation in irf8 mutants and cone and rod photoreceptor regeneration. Light-damaged retinas from both wild-type and irf8 mutants show upregulated expression of mmp-9, il8, and tnfβ pro-inflammatory cytokines. Our data demonstrate that adult zebrafish irf8 mutants can regenerate normally following acute retinal injury. These findings suggest that microglia may not be essential for retinal regeneration in zebrafish and that other mechanisms can compensate for the reduction in microglia numbers.
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Affiliation(s)
- Ping Song
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Dhwani Parsana
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Rupesh Singh
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Lana M Pollock
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA
| | - Bela Anand-Apte
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA
- Department of Ophthalmology, Cleveland Clinic Lerner College of Medicine, Case Western Reserve University, Cleveland, OH, USA
- Department of Molecular Medicine, Cleveland Clinic Lerner College of Medicine, Case Western Reserve University, Cleveland, OH, USA
| | - Brian D Perkins
- Department of Ophthalmic Research, Cole Eye Institute, Cleveland Clinic, Cleveland, OH, USA.
- Department of Ophthalmology, Cleveland Clinic Lerner College of Medicine, Case Western Reserve University, Cleveland, OH, USA.
- Department of Molecular Medicine, Cleveland Clinic Lerner College of Medicine, Case Western Reserve University, Cleveland, OH, USA.
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20
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He M, Xia M, Yang Q, Chen X, Li H, Xia X. P-aminobenzoic acid promotes retinal regeneration through activation of Ascl1a in zebrafish. Neural Regen Res 2024; 19:1849-1856. [PMID: 38103253 PMCID: PMC10960302 DOI: 10.4103/1673-5374.389646] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2023] [Revised: 08/07/2023] [Accepted: 10/20/2023] [Indexed: 12/18/2023] Open
Abstract
JOURNAL/nrgr/04.03/01300535-202408000-00040/figure1/v/2023-12-16T180322Z/r/image-tiff The retina of zebrafish can regenerate completely after injury. Multiple studies have demonstrated that metabolic alterations occur during retinal damage; however to date no study has identified a link between metabolites and retinal regeneration of zebrafish. Here, we performed an unbiased metabolome sequencing in the N-methyl-D-aspartic acid-damaged retinas of zebrafish to demonstrate the metabolomic mechanism of retinal regeneration. Among the differentially-expressed metabolites, we found a significant decrease in p-aminobenzoic acid in the N-methyl-D-aspartic acid-damaged retinas of zebrafish. Then, we investigated the role of p-aminobenzoic acid in retinal regeneration in adult zebrafish. Importantly, p-aminobenzoic acid activated Achaetescute complex-like 1a expression, thereby promoting Müller glia reprogramming and division, as well as Müller glia-derived progenitor cell proliferation. Finally, we eliminated folic acid and inflammation as downstream effectors of PABA and demonstrated that PABA had little effect on Müller glia distribution. Taken together, these findings show that PABA contributes to retinal regeneration through activation of Achaetescute complex-like 1a expression in the N-methyl-D-aspartic acid-damaged retinas of zebrafish.
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Affiliation(s)
- Meihui He
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
| | - Mingfang Xia
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
| | - Qian Yang
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
| | - Xingyi Chen
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
| | - Haibo Li
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
| | - Xiaobo Xia
- Eye Center of Xiangya Hospital, Central South University, Changsha, Hunan Province, China
- Hunan Key Laboratory of Ophthalmology, Changsha, Hunan Province, China
- National Clinical Research Center for Geriatric Disorders, Xiangya Hospital, Central South University, Changsha, Hunan Province, China
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21
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Emmerich K, Hageter J, Hoang T, Lyu P, Sharrock AV, Ceisel A, Thierer J, Chunawala Z, Nimmagadda S, Palazzo I, Matthews F, Zhang L, White DT, Rodriguez C, Graziano G, Marcos P, May A, Mulligan T, Reibman B, Saxena MT, Ackerley DF, Qian J, Blackshaw S, Horstick E, Mumm JS. A large-scale CRISPR screen reveals context-specific genetic regulation of retinal ganglion cell regeneration. Development 2024; 151:dev202754. [PMID: 39007397 PMCID: PMC11361637 DOI: 10.1242/dev.202754] [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: 02/01/2024] [Accepted: 07/08/2024] [Indexed: 07/16/2024]
Abstract
Many genes are known to regulate retinal regeneration after widespread tissue damage. Conversely, genes controlling regeneration after limited cell loss, as per degenerative diseases, are undefined. As stem/progenitor cell responses scale to injury levels, understanding how the extent and specificity of cell loss impact regenerative processes is important. Here, transgenic zebrafish enabling selective retinal ganglion cell (RGC) ablation were used to identify genes that regulate RGC regeneration. A single cell multiomics-informed screen of 100 genes identified seven knockouts that inhibited and 11 that promoted RGC regeneration. Surprisingly, 35 out of 36 genes known and/or implicated as being required for regeneration after widespread retinal damage were not required for RGC regeneration. The loss of seven even enhanced regeneration kinetics, including the proneural factors neurog1, olig2 and ascl1a. Mechanistic analyses revealed that ascl1a disruption increased the propensity of progenitor cells to produce RGCs, i.e. increased 'fate bias'. These data demonstrate plasticity in the mechanism through which Müller glia convert to a stem-like state and context specificity in how genes function during regeneration. Increased understanding of how the regeneration of disease-relevant cell types is specifically controlled will support the development of disease-tailored regenerative therapeutics.
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Affiliation(s)
- Kevin Emmerich
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- McKusick-Nathans Institute and the Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - John Hageter
- Department of Biology, West Virginia University, Morgantown, WV 26505, USA
| | - Thanh Hoang
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI 48105, USA
- Department of Cell and Developmental Biology, University of Michigan School of Medicine, Ann Arbor, MI 48105, USA
| | - Pin Lyu
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Abigail V. Sharrock
- School of Biological Sciences, Victoria University of Wellington, Wellington 6012, New Zealand
| | - Anneliese Ceisel
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - James Thierer
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Zeeshaan Chunawala
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Saumya Nimmagadda
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Isabella Palazzo
- Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Frazer Matthews
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Liyun Zhang
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - David T. White
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Catalina Rodriguez
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Gianna Graziano
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Patrick Marcos
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Adam May
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Tim Mulligan
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Barak Reibman
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Meera T. Saxena
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - David F. Ackerley
- School of Biological Sciences, Victoria University of Wellington, Wellington 6012, New Zealand
| | - Jiang Qian
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Seth Blackshaw
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- McKusick-Nathans Institute and the Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Eric Horstick
- Department of Biology, West Virginia University, Morgantown, WV 26505, USA
- Department of Neuroscience, West Virginia University, Morgantown, WV 26506, USA
| | - Jeff S. Mumm
- Wilmer Eye Institute and the Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- McKusick-Nathans Institute and the Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Center for Nanomedicine, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
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22
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Le N, Vu TD, Palazzo I, Pulya R, Kim Y, Blackshaw S, Hoang T. Robust reprogramming of glia into neurons by inhibition of Notch signaling and nuclear factor I (NFI) factors in adult mammalian retina. SCIENCE ADVANCES 2024; 10:eadn2091. [PMID: 38996013 PMCID: PMC11244444 DOI: 10.1126/sciadv.adn2091] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 11/28/2023] [Accepted: 06/10/2024] [Indexed: 07/14/2024]
Abstract
Generation of neurons through direct reprogramming has emerged as a promising therapeutic approach for treating neurodegenerative diseases. In this study, we present an efficient method for reprogramming retinal glial cells into neurons. By suppressing Notch signaling by disrupting either Rbpj or Notch1/2, we induced mature Müller glial cells to reprogram into bipolar- and amacrine-like neurons. We demonstrate that Rbpj directly activates both Notch effector genes and genes specific to mature Müller glia while indirectly repressing expression of neurogenic basic helix-loop-helix (bHLH) factors. Combined loss of function of Rbpj and Nfia/b/x resulted in conversion of nearly all Müller glia to neurons. Last, inducing Müller glial proliferation by overexpression of dominant-active Yap promotes neurogenesis in both Rbpj- and Nfia/b/x/Rbpj-deficient Müller glia. These findings demonstrate that Notch signaling and NFI factors act in parallel to inhibit neurogenic competence in mammalian Müller glia and help clarify potential strategies for regenerative therapies aimed at treating retinal dystrophies.
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Affiliation(s)
- Nguyet Le
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Trieu-Duc Vu
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI 48105
- Michigan Neuroscience Institute, University of Michigan School of Medicine, Ann Arbor, MI 48105, USA
| | - Isabella Palazzo
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Ritvik Pulya
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Yehna Kim
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Seth Blackshaw
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
- Kavli Neuroscience Discovery Institute, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA
| | - Thanh Hoang
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI 48105
- Michigan Neuroscience Institute, University of Michigan School of Medicine, Ann Arbor, MI 48105, USA
- Department of Cell and Developmental Biology, University of Michigan School of Medicine, Ann Arbor, MI 48105, USA
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23
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Bludau O, Weber A, Bosak V, Kuscha V, Dietrich K, Hans S, Brand M. Inflammation is a critical factor for successful regeneration of the adult zebrafish retina in response to diffuse light lesion. Front Cell Dev Biol 2024; 12:1332347. [PMID: 39071801 PMCID: PMC11272569 DOI: 10.3389/fcell.2024.1332347] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/02/2023] [Accepted: 06/17/2024] [Indexed: 07/30/2024] Open
Abstract
Inflammation can lead to persistent and irreversible loss of retinal neurons and photoreceptors in mammalian vertebrates. In contrast, in the adult zebrafish brain, acute neural inflammation is both necessary and sufficient to stimulate regeneration of neurons. Here, we report on the critical, positive role of the immune system to support retina regeneration in adult zebrafish. After sterile ablation of photoreceptors by phototoxicity, we find rapid response of immune cells, especially monocytes/microglia and neutrophils, which returns to homeostatic levels within 14 days post lesion. Pharmacological or genetic impairment of the immune system results in a reduced Müller glia stem cell response, seen as decreased reactive proliferation, and a strikingly reduced number of regenerated cells from them, including photoreceptors. Conversely, injection of the immune stimulators flagellin, zymosan, or M-CSF into the vitreous of the eye, leads to a robust proliferation response and the upregulation of regeneration-associated marker genes in Müller glia. Our results suggest that neuroinflammation is a necessary and sufficient driver for retinal regeneration in the adult zebrafish retina.
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Affiliation(s)
- Oliver Bludau
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Anke Weber
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Viktoria Bosak
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Veronika Kuscha
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Kristin Dietrich
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Stefan Hans
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
| | - Michael Brand
- CRTD—Center for Regenerative Therapies, and PoL—Cluster of Excellence Physics of Life, Dresden, Germany
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24
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Lu C, Hyde DR. Cytokines IL-1β and IL-10 are required for Müller glia proliferation following light damage in the adult zebrafish retina. Front Cell Dev Biol 2024; 12:1406330. [PMID: 38938553 PMCID: PMC11208712 DOI: 10.3389/fcell.2024.1406330] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/24/2024] [Accepted: 05/16/2024] [Indexed: 06/29/2024] Open
Abstract
Zebrafish possess the ability to regenerate dying neurons in response to retinal injury, with both Müller glia and microglia playing integral roles in this response. Resident Müller glia respond to damage by reprogramming and undergoing an asymmetric cell division to generate a neuronal progenitor cell, which continues to proliferate and differentiate into the lost neurons. In contrast, microglia become reactive, phagocytose dying cells, and release inflammatory signals into the surrounding tissue following damage. In recent years, there has been increased attention on elucidating the role that microglia play in regulating retinal regeneration. Here we demonstrate that inflammatory cytokines are differentially expressed during retinal regeneration, with the expression of a subset of pro-inflammatory cytokine genes upregulated shortly after light damage and the expression of a different subset of cytokine genes subsequently increasing. We demonstrate that both cytokine IL-1β and IL-10 are essential for Müller glia proliferation in the light-damaged retina. While IL-1β is sufficient to induce Müller glia proliferation in an undamaged retina, expression of IL-10 in undamaged retinas only induces Müller glia to express gliotic markers. Together, these findings demonstrate the essential role of inflammatory cytokines IL-1β and IL-10 on Müller glia proliferation following light damage in adult zebrafish.
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Affiliation(s)
| | - David R. Hyde
- Department of Biological Sciences, Center for Stem Cells and Regenerative Medicine, and Center for Zebrafish Research, Galvin Life Sciences Building, University of Notre Dame, Notre Dame, IN, United States
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25
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Kelly LE, El-Hodiri HM, Crider A, Fischer AJ. Protein phosphatases regulate the formation of Müller glia-derived progenitor cells in the chick retina. Mol Cell Neurosci 2024; 129:103932. [PMID: 38679247 PMCID: PMC11362962 DOI: 10.1016/j.mcn.2024.103932] [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: 01/18/2024] [Revised: 03/26/2024] [Accepted: 04/18/2024] [Indexed: 05/01/2024] Open
Abstract
Different kinase-dependent cell signaling pathways are known to play important roles in glia-mediated neuroprotection and reprogramming of Müller glia (MG) into Müller glia-derived progenitor cells (MGPCs) in the retina. However, very little is known about the phosphatases that regulate kinase-dependent signaling in MG. Using single-cell RNA-sequencing (scRNA-seq) databases, we investigated patterns of expression of Dual Specificity Phosphatases (DUSP1/6) and other protein phosphatases in normal and damaged chick retinas. We found that DUSP1, DUSP6, PPP3CB, PPP3R1 and PPPM1A/B/D/E/G are widely expressed by many types of retinal neurons and are dynamically expressed by MG and MGPCs in retinas during the process of reprogramming. We find that inhibition of DUSP1/6 and PP2C phosphatases enhances the formation of proliferating MGPCs in damaged retinas and in retinas treated with insulin and FGF2 in the absence of damage. By contrast, inhibition of PP2B phosphatases suppressed the formation of proliferating MGPCs, but increased numbers of proliferating MGPCs in undamaged retinas treated with insulin and FGF2. In damaged retinas, inhibition of DUSP1/6 increased levels of pERK1/2 and cFos in MG whereas inhibition of PP2B's decreased levels of pStat3 and pS6 in MG. Analyses of scRNA-seq libraries identified numerous differentially activated gene modules in MG in damaged retinas versus MG in retinas treated with insulin+FGF2 suggesting significant differences in kinase-dependent signaling pathways that converge on the formation of MGPCs. Inhibition of phosphatases had no significant effects upon numbers of dying cells in damaged retinas. We conclude that the activity of different protein phosphatases acting through retinal neurons and MG "fine-tune" the cell signaling responses of MG in damaged retinas and during the reprogramming of MG into MGPCs.
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Affiliation(s)
- Lisa E Kelly
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH, USA
| | - Heithem M El-Hodiri
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH, USA
| | - Andrew Crider
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH, USA
| | - Andy J Fischer
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH, USA.
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26
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Zhang J, Yang SG, Zhou FQ. Glycogen synthase kinase 3 signaling in neural regeneration in vivo. J Mol Cell Biol 2024; 15:mjad075. [PMID: 38059848 PMCID: PMC11063957 DOI: 10.1093/jmcb/mjad075] [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: 06/15/2023] [Revised: 11/14/2023] [Accepted: 11/28/2023] [Indexed: 12/08/2023] Open
Abstract
Glycogen synthase kinase 3 (GSK3) signaling plays important and broad roles in regulating neural development in vitro and in vivo. Here, we reviewed recent findings of GSK3-regulated axon regeneration in vivo in both the peripheral and central nervous systems and discussed a few controversial findings in the field. Overall, current evidence indicates that GSK3β signaling serves as an important downstream mediator of the PI3K-AKT pathway to regulate axon regeneration in parallel with the mTORC1 pathway. Specifically, the mTORC1 pathway supports axon regeneration mainly through its role in regulating cap-dependent protein translation, whereas GSK3β signaling might be involved in regulating N6-methyladenosine mRNA methylation-mediated, cap-independent protein translation. In addition, GSK3 signaling also plays a key role in reshaping the neuronal transcriptomic landscape during neural regeneration. Finally, we proposed some research directions to further elucidate the molecular mechanisms underlying the regulatory function of GSK3 signaling and discover novel GSK3 signaling-related therapeutic targets. Together, we hope to provide an updated and insightful overview of how GSK3 signaling regulates neural regeneration in vivo.
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Affiliation(s)
- Jing Zhang
- Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China
| | - Shu-Guang Yang
- Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China
| | - Feng-Quan Zhou
- Sir Run Run Shaw Hospital, Zhejiang University School of Medicine, Hangzhou 310016, China
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27
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Hung JH, Tsai PH, Aala WJF, Chen CC, Chiou SH, Wong TW, Tsai KJ, Hsu SM, Wu LW. TIMP3/Wnt axis regulates gliosis of Müller glia. Biochim Biophys Acta Mol Basis Dis 2024; 1870:167087. [PMID: 38369214 DOI: 10.1016/j.bbadis.2024.167087] [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: 11/01/2023] [Revised: 02/11/2024] [Accepted: 02/15/2024] [Indexed: 02/20/2024]
Abstract
BACKGROUND Previous studies have confirmed the expression of tissue inhibitor of metalloproteinase-3 (TIMP3) in Müller glia (MG). However, the role of TIMP3 in MG remains unknown. METHODS A mouse model of laser-induced retinal damage and gliosis was generated using wild-type C57BL/6 mice. TIMP3 and associated proteins were detected using Western blotting and immunofluorescence microscopy. RNA sequencing (GSE132140) of mouse laser-induced gliosis was utilized for pathway analysis. TIMP3 overexpression was induced in human MG. Human vitreous samples were obtained from patients with proliferative diabetic retinopathy (PDR) and healthy controls for protein analysis. RESULTS TIMP3 levels increased in mouse eyes after laser damage. Morphology and spatial location of TIMP3 indicated its presence in MG. TIMP3-overexpressing MG showed increased cellular proliferation, migration, and cell nuclei size, suggesting TIMP3-induced gliosis for retinal repair. Glial fibrillary acidic protein (GFAP) and vimentin levels were elevated in TIMP3-overexpressing MG and laser-damaged mouse retinas. RNA sequencing and Western blotting suggested a role for β-catenin in mediating TIMP3 effects on the retina. Human vitreous samples from patients with PDR showed a positive correlation between TIMP3 and GFAP levels, both of which were elevated in patients with PDR. CONCLUSIONS TIMP3 is associated with MG gliosis to enhance the repair ability of damaged retinas and is mediated by the canonical Wnt/β-catenin. Changes in TIMP3 could potentially be used to control gliosis in a range of retinal diseases However, given the multifaceted nature of TIMP3, care must be taken when developing treatments that aim solely to boost the function of TIMP3. FUNDING National Cheng Kung University Hospital, Taiwan (NCKUH-10604009 and NCKUH-11202007); the Ministry of Science and Technology (MOST 110-2314-B-006-086-MY3).
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Affiliation(s)
- Jia-Horung Hung
- Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan; Department of Ophthalmology, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Ping-Hsing Tsai
- Department of Medical Research, Taipei Veterans General Hospital, Taipei, Taiwan
| | - Wilson Jr F Aala
- Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Chao-Chung Chen
- Department of Ophthalmology, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Shih-Hwa Chiou
- Department of Medical Research, Taipei Veterans General Hospital, Taipei, Taiwan; Institute of Pharmacology, School of Medicine, National Yang Ming Chiao Tung University, Taipei, Taiwan
| | - Tak-Wah Wong
- Department of Dermatology, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan; Department of Biochemistry and Molecular Biology, College of Medicine, National Cheng Kung University, Tainan, Taiwan; Center of Applied Nanomedicine, National Cheng Kung University, Tainan, Taiwan
| | - Kuen-Jer Tsai
- Institute of Clinical Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan
| | - Sheng-Min Hsu
- Department of Ophthalmology, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan, Taiwan.
| | - Li-Wha Wu
- Institute of Molecular Medicine, College of Medicine, National Cheng Kung University, Tainan, Taiwan; Department of Medical Laboratory Science and Biotechnology, Kaohsiung Medical University, Kaohsiung, Taiwan.
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28
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Du X, Butler AG, Chen HY. Cell-cell interaction in the pathogenesis of inherited retinal diseases. Front Cell Dev Biol 2024; 12:1332944. [PMID: 38500685 PMCID: PMC10944940 DOI: 10.3389/fcell.2024.1332944] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2023] [Accepted: 02/06/2024] [Indexed: 03/20/2024] Open
Abstract
The retina is part of the central nervous system specialized for vision. Inherited retinal diseases (IRD) are a group of clinically and genetically heterogenous disorders that lead to progressive vision impairment or blindness. Although each disorder is rare, IRD accumulatively cause blindness in up to 5.5 million individuals worldwide. Currently, the pathophysiological mechanisms of IRD are not fully understood and there are limited treatment options available. Most IRD are caused by degeneration of light-sensitive photoreceptors. Genetic mutations that abrogate the structure and/or function of photoreceptors lead to visual impairment followed by blindness caused by loss of photoreceptors. In healthy retina, photoreceptors structurally and functionally interact with retinal pigment epithelium (RPE) and Müller glia (MG) to maintain retinal homeostasis. Multiple IRD with photoreceptor degeneration as a major phenotype are caused by mutations of RPE- and/or MG-associated genes. Recent studies also reveal compromised MG and RPE caused by mutations in ubiquitously expressed ciliary genes. Therefore, photoreceptor degeneration could be a direct consequence of gene mutations and/or could be secondary to the dysfunction of their interaction partners in the retina. This review summarizes the mechanisms of photoreceptor-RPE/MG interaction in supporting retinal functions and discusses how the disruption of these processes could lead to photoreceptor degeneration, with an aim to provide a unique perspective of IRD pathogenesis and treatment paradigm. We will first describe the biology of retina and IRD and then discuss the interaction between photoreceptors and MG/RPE as well as their implications in disease pathogenesis. Finally, we will summarize the recent advances in IRD therapeutics targeting MG and/or RPE.
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Affiliation(s)
| | | | - Holly Y. Chen
- Department of Cell, Developmental and Integrative Biology, Heersink School of Medicine, University of Alabama at Birmingham, Birmingham, AL, United States
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29
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Konar GJ, Flickinger Z, Sharma S, Vallone KT, Lyon CE, Doshier C, Lingan A, Lyon W, Patton JG. Damage-Induced Senescent Immune Cells Regulate Regeneration of the Zebrafish Retina. AGING BIOLOGY 2024; 2:e20240021. [PMID: 39156966 PMCID: PMC11328971 DOI: 10.59368/agingbio.20240021] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 08/20/2024]
Abstract
Zebrafish spontaneously regenerate their retinas in response to damage through the action of Müller glia (MG). Even though MG are conserved in higher vertebrates, the capacity to regenerate retinal damage is lost. Recent work has focused on the regulation of inflammation during tissue regeneration, with temporal roles for macrophages and microglia. Senescent cells that have withdrawn from the cell cycle have mostly been implicated in aging but are still metabolically active, releasing a variety of signaling molecules as part of the senescence-associated secretory phenotype. Here, we discover that in response to retinal damage, a subset of cells expressing markers of microglia/macrophages also express markers of senescence. These cells display a temporal pattern of appearance and clearance during retina regeneration. Premature removal of senescent cells by senolytic treatment led to a decrease in proliferation and incomplete repair of the ganglion cell layer after N-methyl-D-aspartate damage. Our results demonstrate a role for modulation of senescent cell responses to balance inflammation, regeneration, plasticity, and repair as opposed to fibrosis and scarring.
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Affiliation(s)
- Gregory J. Konar
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Zachary Flickinger
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Shivani Sharma
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Kyle T. Vallone
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Charles E. Lyon
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Claire Doshier
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - Audrey Lingan
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - William Lyon
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
| | - James G. Patton
- Department of Biological Sciences, Vanderbilt University, Nashville, TN, USA
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30
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Fernández-Albarral JA, Ramírez AI, de Hoz R, Matamoros JA, Salobrar-García E, Elvira-Hurtado L, López-Cuenca I, Sánchez-Puebla L, Salazar JJ, Ramírez JM. Glaucoma: from pathogenic mechanisms to retinal glial cell response to damage. Front Cell Neurosci 2024; 18:1354569. [PMID: 38333055 PMCID: PMC10850296 DOI: 10.3389/fncel.2024.1354569] [Citation(s) in RCA: 23] [Impact Index Per Article: 23.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2023] [Accepted: 01/10/2024] [Indexed: 02/10/2024] Open
Abstract
Glaucoma is a neurodegenerative disease of the retina characterized by the irreversible loss of retinal ganglion cells (RGCs) leading to visual loss. Degeneration of RGCs and loss of their axons, as well as damage and remodeling of the lamina cribrosa are the main events in the pathogenesis of glaucoma. Different molecular pathways are involved in RGC death, which are triggered and exacerbated as a consequence of a number of risk factors such as elevated intraocular pressure (IOP), age, ocular biomechanics, or low ocular perfusion pressure. Increased IOP is one of the most important risk factors associated with this pathology and the only one for which treatment is currently available, nevertheless, on many cases the progression of the disease continues, despite IOP control. Thus, the IOP elevation is not the only trigger of glaucomatous damage, showing the evidence that other factors can induce RGCs death in this pathology, would be involved in the advance of glaucomatous neurodegeneration. The underlying mechanisms driving the neurodegenerative process in glaucoma include ischemia/hypoxia, mitochondrial dysfunction, oxidative stress and neuroinflammation. In glaucoma, like as other neurodegenerative disorders, the immune system is involved and immunoregulation is conducted mainly by glial cells, microglia, astrocytes, and Müller cells. The increase in IOP produces the activation of glial cells in the retinal tissue. Chronic activation of glial cells in glaucoma may provoke a proinflammatory state at the retinal level inducing blood retinal barrier disruption and RGCs death. The modulation of the immune response in glaucoma as well as the activation of glial cells constitute an interesting new approach in the treatment of glaucoma.
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Affiliation(s)
- Jose A. Fernández-Albarral
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
| | - Ana I. Ramírez
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - Rosa de Hoz
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - José A. Matamoros
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - Elena Salobrar-García
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - Lorena Elvira-Hurtado
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
| | - Inés López-Cuenca
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - Lidia Sánchez-Puebla
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, School of Medicine, Complutense University of Madrid, Madrid, Spain
| | - Juan J. Salazar
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, Faculty of Optics and Optometry, Complutense University of Madrid, Madrid, Spain
| | - José M. Ramírez
- Ramon Castroviejo Ophthalmological Research Institute, Complutense University of Madrid (UCM), Grupo UCM 920105, IdISSC, Madrid, Spain
- Department of Immunology, Ophthalmology and ENT, School of Medicine, Complutense University of Madrid, Madrid, Spain
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31
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Tempone MH, Borges-Martins VP, César F, Alexandrino-Mattos DP, de Figueiredo CS, Raony Í, dos Santos AA, Duarte-Silva AT, Dias MS, Freitas HR, de Araújo EG, Ribeiro-Resende VT, Cossenza M, P. Silva H, P. de Carvalho R, Ventura ALM, Calaza KC, Silveira MS, Kubrusly RCC, de Melo Reis RA. The Healthy and Diseased Retina Seen through Neuron-Glia Interactions. Int J Mol Sci 2024; 25:1120. [PMID: 38256192 PMCID: PMC10817105 DOI: 10.3390/ijms25021120] [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: 12/21/2023] [Revised: 01/10/2024] [Accepted: 01/12/2024] [Indexed: 01/24/2024] Open
Abstract
The retina is the sensory tissue responsible for the first stages of visual processing, with a conserved anatomy and functional architecture among vertebrates. To date, retinal eye diseases, such as diabetic retinopathy, age-related macular degeneration, retinitis pigmentosa, glaucoma, and others, affect nearly 170 million people worldwide, resulting in vision loss and blindness. To tackle retinal disorders, the developing retina has been explored as a versatile model to study intercellular signaling, as it presents a broad neurochemical repertoire that has been approached in the last decades in terms of signaling and diseases. Retina, dissociated and arranged as typical cultures, as mixed or neuron- and glia-enriched, and/or organized as neurospheres and/or as organoids, are valuable to understand both neuronal and glial compartments, which have contributed to revealing roles and mechanisms between transmitter systems as well as antioxidants, trophic factors, and extracellular matrix proteins. Overall, contributions in understanding neurogenesis, tissue development, differentiation, connectivity, plasticity, and cell death are widely described. A complete access to the genome of several vertebrates, as well as the recent transcriptome at the single cell level at different stages of development, also anticipates future advances in providing cues to target blinding diseases or retinal dysfunctions.
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Affiliation(s)
- Matheus H. Tempone
- Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.H.T.); (F.C.); (D.P.A.-M.); (V.T.R.-R.)
| | - Vladimir P. Borges-Martins
- Department of Physiology and Pharmacology, Biomedical Institute and Program of Neurosciences, Federal Fluminense University, Niterói 24020-150, Brazil; (V.P.B.-M.); (A.A.d.S.); (M.C.); (R.C.C.K.)
| | - Felipe César
- Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.H.T.); (F.C.); (D.P.A.-M.); (V.T.R.-R.)
| | - Dio Pablo Alexandrino-Mattos
- Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.H.T.); (F.C.); (D.P.A.-M.); (V.T.R.-R.)
| | - Camila S. de Figueiredo
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
| | - Ícaro Raony
- Institute of Medical Biochemistry Leopoldo de Meis, Federal University of Rio de Janeiro, Rio de Janeiro 21941-902, Brazil; (Í.R.); (H.R.F.)
| | - Aline Araujo dos Santos
- Department of Physiology and Pharmacology, Biomedical Institute and Program of Neurosciences, Federal Fluminense University, Niterói 24020-150, Brazil; (V.P.B.-M.); (A.A.d.S.); (M.C.); (R.C.C.K.)
| | - Aline Teixeira Duarte-Silva
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
| | - Mariana Santana Dias
- Laboratory of Gene Therapy and Viral Vectors, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.S.D.); (H.P.S.)
| | - Hércules Rezende Freitas
- Institute of Medical Biochemistry Leopoldo de Meis, Federal University of Rio de Janeiro, Rio de Janeiro 21941-902, Brazil; (Í.R.); (H.R.F.)
| | - Elisabeth G. de Araújo
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
- National Institute of Science and Technology on Neuroimmunomodulation—INCT-NIM, Oswaldo Cruz Institute, Oswaldo Cruz Foundation, Rio de Janeiro 21040-360, Brazil
| | - Victor Tulio Ribeiro-Resende
- Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.H.T.); (F.C.); (D.P.A.-M.); (V.T.R.-R.)
| | - Marcelo Cossenza
- Department of Physiology and Pharmacology, Biomedical Institute and Program of Neurosciences, Federal Fluminense University, Niterói 24020-150, Brazil; (V.P.B.-M.); (A.A.d.S.); (M.C.); (R.C.C.K.)
| | - Hilda P. Silva
- Laboratory of Gene Therapy and Viral Vectors, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.S.D.); (H.P.S.)
| | - Roberto P. de Carvalho
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
| | - Ana L. M. Ventura
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
| | - Karin C. Calaza
- Department of Neurobiology and Program of Neurosciences, Institute of Biology, Federal Fluminense University, Niterói 24020-141, Brazil; (C.S.d.F.); (A.T.D.-S.); (E.G.d.A.); (R.P.d.C.); (A.L.M.V.); (K.C.C.)
| | - Mariana S. Silveira
- Laboratory for Investigation in Neuroregeneration and Development, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil;
| | - Regina C. C. Kubrusly
- Department of Physiology and Pharmacology, Biomedical Institute and Program of Neurosciences, Federal Fluminense University, Niterói 24020-150, Brazil; (V.P.B.-M.); (A.A.d.S.); (M.C.); (R.C.C.K.)
| | - Ricardo A. de Melo Reis
- Laboratory of Neurochemistry, Institute of Biophysics Carlos Chagas Filho, Federal University of Rio de Janeiro, Rio de Janeiro 21949-000, Brazil; (M.H.T.); (F.C.); (D.P.A.-M.); (V.T.R.-R.)
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32
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Nomura-Komoike K, Nishino R, Fujieda H. Effects of different alkylating agents on photoreceptor degeneration and proliferative response of Müller glia. Sci Rep 2024; 14:61. [PMID: 38167441 PMCID: PMC10762013 DOI: 10.1038/s41598-023-50485-7] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/13/2023] [Accepted: 12/20/2023] [Indexed: 01/05/2024] Open
Abstract
Animal models for retinal degeneration are essential for elucidating its pathogenesis and developing new therapeutic strategies in humans. N-methyl-N-nitrosourea (MNU) has been extensively used to construct a photoreceptor-specific degeneration model, which has served to unveil the molecular process of photoreceptor degeneration as well as the mechanisms regulating the protective responses of remaining cells. Methyl methanesulphonate (MMS), also known to cause photoreceptor degeneration, is considered a good alternative to MNU due to its higher usability; however, detailed pathophysiological processes after MMS treatment remain uncharacterized. Here, we analyzed the time course of photoreceptor degeneration, Müller glial proliferation, and expression of secretory factors after MNU and MMS treatments in rats. While the timing of rod degeneration was similar between the treatments, we unexpectedly found that cones survived slightly longer after MMS treatment. Müller glia reentered the cell cycle at a similar timing after the two treatments; however, the G1/S transition occurred earlier after MMS treatment. Moreover, growth factors such as FGF2 and LIF were more highly upregulated in the MMS model. These data suggest that comparative analyses of the two injury models may be beneficial for understanding the complex regulatory mechanisms underlying the proliferative response of Müller glia.
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Affiliation(s)
- 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
| | - 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
| | - 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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33
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Lyu P, Iribarne M, Serjanov D, Zhai Y, Hoang T, Campbell LJ, Boyd P, Palazzo I, Nagashima M, Silva NJ, Hitchcock PF, Qian J, Hyde DR, Blackshaw S. Common and divergent gene regulatory networks control injury-induced and developmental neurogenesis in zebrafish retina. Nat Commun 2023; 14:8477. [PMID: 38123561 PMCID: PMC10733277 DOI: 10.1038/s41467-023-44142-w] [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: 08/24/2023] [Accepted: 12/01/2023] [Indexed: 12/23/2023] Open
Abstract
Following acute retinal damage, zebrafish possess the ability to regenerate all neuronal subtypes through Müller glia (MG) reprogramming and asymmetric cell division that produces a multipotent Müller glia-derived neuronal progenitor cell (MGPC). This raises three key questions. First, do MG reprogram to a developmental retinal progenitor cell (RPC) state? Second, to what extent does regeneration recapitulate retinal development? And finally, does loss of different retinal cell subtypes induce unique MG regeneration responses? We examined these questions by performing single-nuclear and single-cell RNA-Seq and ATAC-Seq in both developing and regenerating retinas. Here we show that injury induces MG to reprogram to a state similar to late-stage RPCs. However, there are major transcriptional differences between MGPCs and RPCs, as well as major transcriptional differences between activated MG and MGPCs when different retinal cell subtypes are damaged. Validation of candidate genes confirmed that loss of different subtypes induces differences in transcription factor gene expression and regeneration outcomes.
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Affiliation(s)
- Pin Lyu
- Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
| | - Maria Iribarne
- Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Stem Cells and Regenerative Medicine, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Zebrafish Research, University of Notre Dame, Notre Dame, IN, 46556, USA
| | - Dmitri Serjanov
- Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Stem Cells and Regenerative Medicine, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Zebrafish Research, University of Notre Dame, Notre Dame, IN, 46556, USA
| | - Yijie Zhai
- Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
| | - Thanh Hoang
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
- Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
- Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
| | - Leah J Campbell
- Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Stem Cells and Regenerative Medicine, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Zebrafish Research, University of Notre Dame, Notre Dame, IN, 46556, USA
| | - Patrick Boyd
- Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Stem Cells and Regenerative Medicine, University of Notre Dame, Notre Dame, IN, 46556, USA
- Center for Zebrafish Research, University of Notre Dame, Notre Dame, IN, 46556, USA
| | - Isabella Palazzo
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA
| | - Mikiko Nagashima
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI, 48105, USA
| | - Nicholas J Silva
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI, 48105, USA
| | - Peter F Hitchcock
- Department of Ophthalmology and Visual Sciences, University of Michigan School of Medicine, Ann Arbor, MI, 48105, USA
| | - Jiang Qian
- Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
| | - David R Hyde
- Department of Biological Sciences, University of Notre Dame, Notre Dame, IN, 46556, USA.
- Center for Stem Cells and Regenerative Medicine, University of Notre Dame, Notre Dame, IN, 46556, USA.
- Center for Zebrafish Research, University of Notre Dame, Notre Dame, IN, 46556, USA.
| | - Seth Blackshaw
- Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
- Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
- Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
- Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
- Kavli Neuroscience Discovery Institute, Johns Hopkins University School of Medicine, Baltimore, MD, 21287, USA.
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34
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Kelly LE, El-Hodiri HM, Crider A, Fischer AJ. Protein phosphatases regulate the formation of Müller glia-derived progenitor cells in the chick retina. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.12.11.570629. [PMID: 38168320 PMCID: PMC10760049 DOI: 10.1101/2023.12.11.570629] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 01/05/2024]
Abstract
Different kinase-dependent cell signaling pathways are known to play important roles in glia-mediated neuroprotection and reprogramming of Müller glia (MG) into Müller glia-derived progenitor cells (MGPCs) in the retina. However, very little is known about the phosphatases that regulate kinase-dependent signaling in MG. Using single-cell RNA-sequencing (scRNA-seq) databases, we investigated patterns of expression of Dual Specificity Phosphatases (DUSP1/6) and other protein phosphatases in normal and damaged chick retinas. We found that DUSP1, DUSP6, PPP3CB, PPP3R1 and PPPM1A/B/D/E/G are dynamically expressed by MG and MGPCs in retinas during the process of reprogramming. We find that inhibition of DUSP1/6 and PP2C phosphatases enhances the formation of proliferating MGPCs in damaged retinas and in retinas treated with insulin in FGF2 in the absence of damage. By contrast, inhibition of PP2B phosphatases suppressed the formation of proliferating MGPCs, but increased numbers of proliferating MGPCs in undamaged retinas treated with insulin and FGF2. In damaged retinas, inhibition of DUSP1/6 increased levels of pERK1/2 and cFos in MG whereas inhibition of PP2B's decreased levels of pStat3 and pS6 in MG. Analyses of scRNA-seq libraries identified numerous differentially activated gene modules in MG in damaged retinas versus MG in retinas treated with insulin+FGF2 suggesting significant differences in kinase-dependent signaling pathways that converge on the formation of MGPCs. Inhibition of phosphatases had no significant effects upon numbers of dying cells in damaged retinas. We conclude that the activity of different protein phosphatases "fine-tune" the cell signaling responses of MG in damaged retinas and during the reprogramming of MG into MGPCs.
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Affiliation(s)
- Lisa E. Kelly
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH
| | - Heithem M. El-Hodiri
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH
| | - Andrew Crider
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH
| | - Andy J. Fischer
- Department of Neuroscience, College of Medicine, The Ohio State University, Columbus, OH
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35
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Lo J, Mehta K, Dhillon A, Huang YK, Luo Z, Nam MH, Al Diri I, Chang KC. Therapeutic strategies for glaucoma and optic neuropathies. Mol Aspects Med 2023; 94:101219. [PMID: 37839232 PMCID: PMC10841486 DOI: 10.1016/j.mam.2023.101219] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/12/2023] [Revised: 10/02/2023] [Accepted: 10/09/2023] [Indexed: 10/17/2023]
Abstract
Glaucoma is a neurodegenerative eye disease that causes permanent vision impairment. The main pathological characteristics of glaucoma are retinal ganglion cell (RGC) loss and optic nerve degeneration. Glaucoma can be caused by elevated intraocular pressure (IOP), although some cases are congenital or occur in patients with normal IOP. Current glaucoma treatments rely on medicine and surgery to lower IOP, which only delays disease progression. First-line glaucoma medicines are supported by pharmacotherapy advancements such as Rho kinase inhibitors and innovative drug delivery systems. Glaucoma surgery has shifted to safer minimally invasive (or microinvasive) glaucoma surgery, but further trials are needed to validate long-term efficacy. Further, growing evidence shows that adeno-associated virus gene transduction and stem cell-based RGC replacement therapy hold potential to treat optic nerve fiber degeneration and glaucoma. However, better understanding of the regulatory mechanisms of RGC development is needed to provide insight into RGC differentiation from stem cells and help choose target genes for viral therapy. In this review, we overview current progress in RGC development research, optic nerve fiber regeneration, and human stem cell-derived RGC differentiation and transplantation. We also provide an outlook on perspectives and challenges in the field.
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Affiliation(s)
- Jung Lo
- Department of Ophthalmology, Kaohsiung Chang Gung Memorial Hospital and Chang Gung University College of Medicine, Kaohsiung, 83301, Taiwan
| | - Kamakshi Mehta
- Department of Ophthalmology, Louis J. Fox Center for Vision Restoration, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA
| | - Armaan Dhillon
- Sue Anschutz-Rodgers Eye Center and Department of Ophthalmology, School of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, 80045, USA
| | - Yu-Kai Huang
- Department of Neurosurgery, Kaohsiung Medical University Hospital, Kaohsiung, 80708, Taiwan; Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, 80708, Taiwan
| | - Ziming Luo
- Spencer Center for Vision Research, Byers Eye Institute, Stanford University School of Medicine, Palo Alto, CA, 94304, USA
| | - Mi-Hyun Nam
- Sue Anschutz-Rodgers Eye Center and Department of Ophthalmology, School of Medicine, University of Colorado, Anschutz Medical Campus, Aurora, CO, 80045, USA.
| | - Issam Al Diri
- Department of Ophthalmology, Louis J. Fox Center for Vision Restoration, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA.
| | - Kun-Che Chang
- Department of Ophthalmology, Louis J. Fox Center for Vision Restoration, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15219, USA; Department of Neurobiology, Center of Neuroscience, University of Pittsburgh School of Medicine, Pittsburgh, PA, 15213, USA; Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, 80708, Taiwan.
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36
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Celotto L, Rost F, Machate A, Bläsche J, Dahl A, Weber A, Hans S, Brand M. Single-cell RNA sequencing unravels the transcriptional network underlying zebrafish retina regeneration. eLife 2023; 12:RP86507. [PMID: 37988404 PMCID: PMC10662954 DOI: 10.7554/elife.86507] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2023] Open
Abstract
In the lesioned zebrafish retina, Müller glia produce multipotent retinal progenitors that generate all retinal neurons, replacing lost cell types. To study the molecular mechanisms linking Müller glia reactivity to progenitor production and neuronal differentiation, we used single-cell RNA sequencing of Müller glia, progenitors and regenerated progeny from uninjured and light-lesioned retinae. We discover an injury-induced Müller glia differentiation trajectory that leads into a cell population with a hybrid identity expressing marker genes of Müller glia and progenitors. A glial self-renewal and a neurogenic trajectory depart from the hybrid cell population. We further observe that neurogenic progenitors progressively differentiate to generate retinal ganglion cells first and bipolar cells last, similar to the events observed during retinal development. Our work provides a comprehensive description of Müller glia and progenitor transcriptional changes and fate decisions in the regenerating retina, which are key to tailor cell differentiation and replacement therapies for retinal dystrophies in humans.
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Affiliation(s)
- Laura Celotto
- Technische Universität Dresden, CRTD - Center for Regenerative Therapies Dresden, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Fabian Rost
- Technische Universität Dresden, DRESDEN-Concept Genome Center, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Anja Machate
- Technische Universität Dresden, CRTD - Center for Regenerative Therapies Dresden, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Juliane Bläsche
- Technische Universität Dresden, DRESDEN-Concept Genome Center, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Andreas Dahl
- Technische Universität Dresden, DRESDEN-Concept Genome Center, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Anke Weber
- Technische Universität Dresden, CRTD - Center for Regenerative Therapies Dresden, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Stefan Hans
- Technische Universität Dresden, CRTD - Center for Regenerative Therapies Dresden, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
| | - Michael Brand
- Technische Universität Dresden, CRTD - Center for Regenerative Therapies Dresden, Center for Molecular and Cellular Bioengineering (CMCB), FetscherstraßeDresdenGermany
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Le N, Vu TD, Palazzo I, Pulya R, Kim Y, Blackshaw S, Hoang T. Robust reprogramming of glia into neurons by inhibition of Notch signaling and NFI factors in adult mammalian retina. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.10.29.560483. [PMID: 37961663 PMCID: PMC10634926 DOI: 10.1101/2023.10.29.560483] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/15/2023]
Abstract
Generation of neurons through direct reprogramming has emerged as a promising therapeutic approach for neurodegenerative diseases. Despite successful applications in vitro , in vivo implementation has been hampered by low efficiency. In this study, we present a highly efficient strategy for reprogramming retinal glial cells into neurons by simultaneously inhibiting key negative regulators. By suppressing Notch signaling through the removal of its central mediator Rbpj, we induced mature Müller glial cells to reprogram into bipolar and amacrine neurons in uninjured adult mouse retinas, and observed that this effect was further enhanced by retinal injury. We found that specific loss of function of Notch1 and Notch2 receptors in Müller glia mimicked the effect of Rbpj deletion on Müller glia-derived neurogenesis. Integrated analysis of multiome (scRNA- and scATAC-seq) and CUT&Tag data revealed that Rbpj directly activates Notch effector genes and genes specific to mature Müller glia while also indirectly represses the expression of neurogenic bHLH factors. Furthermore, we found that combined loss of function of Rbpj and Nfia/b/x resulted in a robust conversion of nearly all Müller glia to neurons. Finally, we demonstrated that inducing Müller glial proliferation by AAV (adeno-associated virus)-mediated overexpression of dominant- active Yap supports efficient levels of Müller glia-derived neurogenesis in both Rbpj - and Nfia/b/x/Rbpj - deficient Müller glia. These findings demonstrate that, much like in zebrafish, Notch signaling actively represses neurogenic competence in mammalian Müller glia, and suggest that inhibition of Notch signaling and Nfia/b/x in combination with overexpression of activated Yap could serve as an effective component of regenerative therapies for degenerative retinal diseases.
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Abstract
The neural retina of mammals, like most of the rest of the central nervous system, does not regenerate new neurons after they are lost through damage or disease. The ability of nonmammalian vertebrates, like fish and amphibians, is remarkable, and lessons learned over the last 20 years have revealed some of the mechanisms underlying this potential. This knowledge has recently been applied to mammals to develop methods that can stimulate regeneration in mice. In this review, we highlight the progress in this area, and propose a "wish list" of how the clinical implementation of regenerative strategies could be applicable to various human retinal diseases.
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Affiliation(s)
- Marina Pavlou
- Department of Biological Structure, University of Washington School of Medicine, Institute of Stem Cells and Regenerative Medicine, Seattle, Washington 98195, USA
| | - Thomas A Reh
- Department of Biological Structure, University of Washington School of Medicine, Institute of Stem Cells and Regenerative Medicine, Seattle, Washington 98195, USA
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Norrie JL, Lupo M, Shirinifard A, Djekidel N, Ramirez C, Xu B, Dundee JM, Dyer MA. Latent Epigenetic Programs in Müller Glia Contribute to Stress, Injury, and Disease Response in the Retina. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.10.15.562396. [PMID: 37905050 PMCID: PMC10614790 DOI: 10.1101/2023.10.15.562396] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/02/2023]
Abstract
Previous studies have demonstrated the dynamic changes in chromatin structure during retinal development that correlate with changes in gene expression. However, a major limitation of those prior studies was the lack of cellular resolution. Here, we integrate single-cell (sc) RNA-seq and scATAC-seq with bulk retinal data sets to identify cell type-specific changes in the chromatin structure during development. Although most genes' promoter activity is strongly correlated with chromatin accessibility, we discovered several hundred genes that were transcriptionally silent but had accessible chromatin at their promoters. Most of those silent/accessible gene promoters were in the Müller glial cells. The Müller cells are radial glia of the retina and perform a variety of essential functions to maintain retinal homeostasis and respond to stress, injury, or disease. The silent/accessible genes in Müller glia are enriched in pathways related to inflammation, angiogenesis, and other types of cell-cell signaling and were rapidly activated when we tested 15 different physiologically relevant conditions to mimic retinal stress, injury, or disease in human and murine retinae. We refer to these as "pliancy genes" because they allow the Müller glia to rapidly change their gene expression and cellular state in response to different types of retinal insults. The Müller glial cell pliancy program is established during development, and we demonstrate that pliancy genes are necessary and sufficient for regulating inflammation in the murine retina in vivo. In zebrafish, Müller glia can de-differentiate and form retinal progenitor cells that replace lost neurons. The pro-inflammatory pliancy gene cascade is not activated in zebrafish Müller glia following injury, and we propose a model in which species-specific pliancy programs underly the differential response to retinal damage in species that can regenerate retinal neurons (zebrafish) versus those that cannot (humans and mice).
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Emmerich K, Walker SL, Wang G, White DT, Ceisel A, Wang F, Teng Y, Chunawala Z, Graziano G, Nimmagadda S, Saxena MT, Qian J, Mumm JS. Transcriptomic comparison of two selective retinal cell ablation paradigms in zebrafish reveals shared and cell-specific regenerative responses. PLoS Genet 2023; 19:e1010905. [PMID: 37819938 PMCID: PMC10593236 DOI: 10.1371/journal.pgen.1010905] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/09/2023] [Revised: 10/23/2023] [Accepted: 08/07/2023] [Indexed: 10/13/2023] Open
Abstract
Retinal Müller glia (MG) can act as stem-like cells to generate new neurons in both zebrafish and mice. In zebrafish, retinal regeneration is innate and robust, resulting in the replacement of lost neurons and restoration of visual function. In mice, exogenous stimulation of MG is required to reveal a dormant and, to date, limited regenerative capacity. Zebrafish studies have been key in revealing factors that promote regenerative responses in the mammalian eye. Increased understanding of how the regenerative potential of MG is regulated in zebrafish may therefore aid efforts to promote retinal repair therapeutically. Developmental signaling pathways are known to coordinate regeneration following widespread retinal cell loss. In contrast, less is known about how regeneration is regulated in the context of retinal degenerative disease, i.e., following the loss of specific retinal cell types. To address this knowledge gap, we compared transcriptomic responses underlying regeneration following targeted loss of rod photoreceptors or bipolar cells. In total, 2,531 differentially expressed genes (DEGs) were identified, with the majority being paradigm specific, including during early MG activation phases, suggesting the nature of the injury/cell loss informs the regenerative process from initiation onward. For example, early modulation of Notch signaling was implicated in the rod but not bipolar cell ablation paradigm and components of JAK/STAT signaling were implicated in both paradigms. To examine candidate gene roles in rod cell regeneration, including several immune-related factors, CRISPR/Cas9 was used to create G0 mutant larvae (i.e., "crispants"). Rod cell regeneration was inhibited in stat3 crispants, while mutating stat5a/b, c7b and txn accelerated rod regeneration kinetics. These data support emerging evidence that discrete responses follow from selective retinal cell loss and that the immune system plays a key role in regulating "fate-biased" regenerative processes.
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Affiliation(s)
- Kevin Emmerich
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
- McKusick-Nathans Institute of the Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, Maryland, United States of America
| | - Steven L. Walker
- Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta University, Augusta, Georgia, United States of America
| | - Guohua Wang
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - David T. White
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Anneliese Ceisel
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Fang Wang
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Yong Teng
- Department of Hematology and Medical Oncology, Winship Cancer Institute, Emory University, Atlanta, Georgia, United States of America
| | - Zeeshaan Chunawala
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Gianna Graziano
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Saumya Nimmagadda
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Meera T. Saxena
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Jiang Qian
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
| | - Jeff S. Mumm
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, Maryland, United States of America
- McKusick-Nathans Institute of the Department of Genetic Medicine, Johns Hopkins School of Medicine, Baltimore, Maryland, United States of America
- Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta University, Augusta, Georgia, United States of America
- Solomon H. Snyder Department of Neuroscience, Johns Hopkins University, Baltimore, Maryland, United States of America
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Zhang X, Leavey P, Appel H, Makrides N, Blackshaw S. Molecular mechanisms controlling vertebrate retinal patterning, neurogenesis, and cell fate specification. Trends Genet 2023; 39:736-757. [PMID: 37423870 PMCID: PMC10529299 DOI: 10.1016/j.tig.2023.06.002] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2023] [Revised: 06/06/2023] [Accepted: 06/07/2023] [Indexed: 07/11/2023]
Abstract
This review covers recent advances in understanding the molecular mechanisms controlling neurogenesis and specification of the developing retina, with a focus on insights obtained from comparative single cell multiomic analysis. We discuss recent advances in understanding the mechanisms by which extrinsic factors trigger transcriptional changes that spatially pattern the optic cup (OC) and control the initiation and progression of retinal neurogenesis. We also discuss progress in unraveling the core evolutionarily conserved gene regulatory networks (GRNs) that specify early- and late-state retinal progenitor cells (RPCs) and neurogenic progenitors and that control the final steps in determining cell identity. Finally, we discuss findings that provide insight into regulation of species-specific aspects of retinal patterning and neurogenesis, including consideration of key outstanding questions in the field.
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Affiliation(s)
- Xin Zhang
- Department of Ophthalmology, Columbia University School of Medicine, New York, NY, USA; Department of Pathology and Cell Biology, Columbia University School of Medicine, New York, NY, USA.
| | - Patrick Leavey
- Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Haley Appel
- Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA
| | - Neoklis Makrides
- Department of Ophthalmology, Columbia University School of Medicine, New York, NY, USA
| | - Seth Blackshaw
- Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Psychiatry and Behavioral Science, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Ophthalmology, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, MD, USA; Kavli Neuroscience Discovery Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
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Blackshaw S, Lyu P, Zhai Y, Qian J, Iribarne M, Serjanov D, Campbell L, Boyd P, Hyde D, Palazzo I, Hoang T, Nagashima M, Silva N, Hitchcock P. Common and divergent gene regulatory networks control injury-induced and developmental neurogenesis in zebrafish retina. RESEARCH SQUARE 2023:rs.3.rs-3294233. [PMID: 37790324 PMCID: PMC10543505 DOI: 10.21203/rs.3.rs-3294233/v1] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 10/05/2023]
Abstract
Following acute retinal damage, zebrafish possess the ability to regenerate all neuronal subtypes. This regeneration requires Müller glia (MG) to reprogram and divide asymmetrically to produce a multipotent Müller glia-derived neuronal progenitor cell (MGPC). This raises three key questions. First, does loss of different retinal cell subtypes induce unique MG regeneration responses? Second, do MG reprogram to a developmental retinal progenitor cell state? And finally, to what extent does regeneration recapitulate retinal development? We examined these questions by performing single-nuclear and single-cell RNA-Seq and ATAC-Seq in both developing and regenerating retinas. While MG reprogram to a state similar to late-stage retinal progenitors in developing retinas, there are transcriptional differences between reprogrammed MG/MGPCs and late progenitors, as well as reprogrammed MG in outer and inner retinal damage models. Validation of candidate genes confirmed that loss of different subtypes induces differences in transcription factor gene expression and regeneration outcomes. This work identifies major differences between gene regulatory networks activated following the selective loss of different subtypes of retina neurons, as well as between retinal regeneration and development.
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Affiliation(s)
| | | | - Yijie Zhai
- Johns Hopkins University School of Medicine
| | - Jiang Qian
- Johns Hopkins University School of Medicine
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Yasuda T, Nakazawa T, Hirakawa K, Matsumoto I, Nagata K, Mori S, Igarashi K, Sagara H, Oda S, Mitani H. Retinal regeneration after injury induced by gamma-ray irradiation during early embryogenesis in medaka, Oryzias latipes. Int J Radiat Biol 2023; 100:131-138. [PMID: 37555698 DOI: 10.1080/09553002.2023.2242932] [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: 09/11/2022] [Revised: 07/06/2023] [Accepted: 07/21/2023] [Indexed: 08/10/2023]
Abstract
PURPOSE Zebrafish, a small fish model, exhibits a multipotent ability for retinal regeneration after damage throughout its lifetime. Compared with zebrafish, birds and mammals exhibit such a regenerative capacity only during the embryonic period, and this capacity decreases with age. In medaka, another small fish model that has also been used extensively in biological research, the retina's inner nuclear layer (INL) failed to regenerate after injury in the hatchling at eight days postfertilization (dpf). We characterized the regenerative process of the embryonic retina when the retinal injury occurred during the early embryonic period in medaka. METHODS We employed a 10 Gy dose of gamma-ray irradiation to initiate retinal injury in medaka embryos at 3 dpf and performed histopathological analyses up to 21 dpf. RESULTS One day after irradiation, numerous apoptotic neurons were observed in the INL; however, these neurons were rarely observed in the ciliary marginal zone and the photoreceptor layer. Numerous pyknotic cells were clustered in the irradiated retina until two days after irradiation. These disappeared four days after irradiation, but the abnormal bridging structures between the INL and ganglion cell layer (GCL) were present until 11 days after irradiation, and the neural layers were completely regenerated 18 days after irradiation. After gamma-ray irradiation, the spindle-like Müller glial cells in the INL became rounder but did not lose their ability to express SOX2. CONCLUSIONS Irradiated retina at 3 dpf of medaka embryos could be completely regenerated at 18 days after irradiation (21 dpf), although the abnormal layer structures bridging the INL and GCL were transiently formed in the retinas of all the irradiated embryos. Four days after irradiation, embryonic medaka Müller glia were reduced in number but maintained SOX2 expression as in nonirradiated embryos. This finding contrasts with previous reports that 8 dpf medaka larvae could not fully regenerate damaged retinas because of loss of SOX2 expression.
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Affiliation(s)
- Takako Yasuda
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
- Department of Chemical and Biological Sciences, Japan Women's University, Tokyo, Japan
| | - Takuya Nakazawa
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
| | - Kei Hirakawa
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
| | - Ikumi Matsumoto
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
| | - Kento Nagata
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
- Department of Radiation Effects Research, Institute for Radiological Science, National Institutes for Quantum Science and Technology, Chiba, Japan
| | - Shunta Mori
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
| | - Kento Igarashi
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
- Department of Applied Pharmacology, Kagoshima University, Kagoshima, Japan
| | - Hiroshi Sagara
- Medical Proteomics Laboratory, Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Shoji Oda
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
| | - Hiroshi Mitani
- Department of Integrated Biosciences, Graduate School of Frontier Science, The University of Tokyo, Kashiwa, Japan
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Lyu P, Iribarne M, Serjanov D, Zhai Y, Hoang T, Campbell LJ, Boyd P, Palazzo I, Nagashima M, Silva NJ, HItchcock PF, Qian J, Hyde DR, Blackshaw S. Common and divergent gene regulatory networks control injury-induced and developmental neurogenesis in zebrafish retina. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2023:2023.08.08.552451. [PMID: 37609307 PMCID: PMC10441373 DOI: 10.1101/2023.08.08.552451] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 08/24/2023]
Abstract
Following acute retinal damage, zebrafish possess the ability to regenerate all neuronal subtypes. This regeneration requires Müller glia (MG) to reprogram and divide asymmetrically to produce a multipotent Müller glia-derived neuronal progenitor cell (MGPC). This raises three key questions. First, does loss of different retinal cell subtypes induce unique MG regeneration responses? Second, do MG reprogram to a developmental retinal progenitor cell state? And finally, to what extent does regeneration recapitulate retinal development? We examined these questions by performing single-nuclear and single-cell RNA-Seq and ATAC-Seq in both developing and regenerating retinas. While MG reprogram to a state similar to late-stage retinal progenitors in developing retinas, there are transcriptional differences between reprogrammed MG/MGPCs and late progenitors, as well as reprogrammed MG in outer and inner retinal damage models. Validation of candidate genes confirmed that loss of different subtypes induces differences in transcription factor gene expression and regeneration outcomes. This work identifies major differences between gene regulatory networks activated following the selective loss of different subtypes of retina neurons, as well as between retinal regeneration and development.
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45
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Xiao X, Liao Z, Zou J. Genetic and epigenetic regulators of retinal Müller glial cell reprogramming. ADVANCES IN OPHTHALMOLOGY PRACTICE AND RESEARCH 2023; 3:126-133. [PMID: 37846362 PMCID: PMC10577857 DOI: 10.1016/j.aopr.2023.05.004] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 04/13/2023] [Revised: 05/18/2023] [Accepted: 05/29/2023] [Indexed: 10/18/2023]
Abstract
Background Retinal diseases characterized with irreversible loss of retinal nerve cells, such as optic atrophy and retinal degeneration, are the main causes of blindness. Current treatments for these diseases are very limited. An emerging treatment strategy is to induce the reprogramming of Müller glial cells to generate new retinal nerve cells, which could potentially restore vision. Main text Müller glial cells are the predominant glial cells in retinae and play multiple roles to maintain retinal homeostasis. In lower vertebrates, such as in zebrafish, Müller glial cells can undergo cell reprogramming to regenerate new retinal neurons in response to various damage factors, while in mammals, this ability is limited. Interestingly, with proper treatments, Müller glial cells can display the potential for regeneration of retinal neurons in mammalian retinae. Recent studies have revealed that dozens of genetic and epigenetic regulators play a vital role in inducing the reprogramming of Müller glial cells in vivo. This review summarizes these critical regulators for Müller glial cell reprogramming and highlights their differences between zebrafish and mammals. Conclusions A number of factors have been identified as the important regulators in Müller glial cell reprogramming. The early response of Müller glial cells upon acute retinal injury, such as the regulation in the exit from quiescent state, the initiation of reactive gliosis, and the re-entry of cell cycle of Müller glial cells, displays significant difference between mouse and zebrafish, which may be mediated by the diverse regulation of Notch and TGFβ (transforming growth factor-β) isoforms and different chromatin accessibility.
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Affiliation(s)
- Xueqi Xiao
- Zhejiang Provincial Key Laboratory for Water Environment and Marine Biological Resources Protection, College of Life and Environmental Science, Wenzhou University, Wenzhou, China
| | - Zhiyong Liao
- Zhejiang Provincial Key Laboratory for Water Environment and Marine Biological Resources Protection, College of Life and Environmental Science, Wenzhou University, Wenzhou, China
| | - Jian Zou
- Department of Ophthalmology, The Second Affiliated Hospital of Zhejiang University School of Medicine, Hangzhou, China
- The Institute of Translational Medicine, Zhejiang University, Hangzhou, China
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Tomczak W, Winkler-Lach W, Tomczyk-Socha M, Misiuk-Hojło M. Advancements in Ocular Regenerative Therapies. BIOLOGY 2023; 12:biology12050737. [PMID: 37237549 DOI: 10.3390/biology12050737] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/07/2023] [Revised: 05/12/2023] [Accepted: 05/16/2023] [Indexed: 05/28/2023]
Abstract
The use of stem cells (SCs) has emerged as a promising avenue in ophthalmology, offering potential therapeutic solutions for various vision impairments and degenerative eye diseases. SCs possess the unique ability to self-renew and differentiate into specialised cell types, making them valuable tools for repairing damaged tissues and restoring visual function. Stem cell-based therapies hold significant potential for addressing conditions such as age-related macular degeneration (AMD), retinitis pigmentosa (RP), corneal disorders, and optic nerve damage. Therefore, researchers have explored different sources of stem cells, including embryonic stem cells (ESC), induced pluripotent stem cells (iPSCs), and adult stem cells, for ocular tissue regeneration. Preclinical studies and early-phase clinical trials have demonstrated promising outcomes, with some patients experiencing improved vision following stem cell-based interventions. However, several challenges remain, including optimising the differentiation protocols, ensuring transplanted cells' safety and long-term viability, and developing effective delivery methods. The field of stem cell research in ophthalmology witnesses a constant influx of new reports and discoveries. To effectively navigate these tons of information, it becomes crucial to summarise and systematise these findings periodically. In light of recent discoveries, this paper demonstrates the potential applications of stem cells in ophthalmology, focusing on their use in various eye tissues, including the cornea, retina, conjunctiva, iris, trabecular meshwork, lens, ciliary body, sclera, and orbital fat.
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Affiliation(s)
| | | | | | - Marta Misiuk-Hojło
- Department of Ophthalmology, Wroclaw Medical University, 50556 Wroclaw, Poland
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Emmerich K, White DT, Kambhampati SP, Casado GL, Fu TM, Chunawala Z, Sahoo A, Nimmagadda S, Krishnan N, Saxena MT, Walker SL, Betzig E, Kannan RM, Mumm JS. Nanoparticle-based targeting of microglia improves the neural regeneration enhancing effects of immunosuppression in the zebrafish retina. Commun Biol 2023; 6:534. [PMID: 37202450 PMCID: PMC10193316 DOI: 10.1038/s42003-023-04898-9] [Citation(s) in RCA: 6] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2021] [Accepted: 05/02/2023] [Indexed: 05/20/2023] Open
Abstract
Retinal Müller glia function as injury-induced stem-like cells in zebrafish but not mammals. However, insights gleaned from zebrafish have been applied to stimulate nascent regenerative responses in the mammalian retina. For instance, microglia/macrophages regulate Müller glia stem cell activity in the chick, zebrafish, and mouse. We previously showed that post-injury immunosuppression by the glucocorticoid dexamethasone accelerated retinal regeneration kinetics in zebrafish. Similarly, microglia ablation enhances regenerative outcomes in the mouse retina. Targeted immunomodulation of microglia reactivity may therefore enhance the regenerative potential of Müller glia for therapeutic purposes. Here, we investigated potential mechanisms by which post-injury dexamethasone accelerates retinal regeneration kinetics, and the effects of dendrimer-based targeting of dexamethasone to reactive microglia. Intravital time-lapse imaging revealed that post-injury dexamethasone inhibited microglia reactivity. The dendrimer-conjugated formulation: (1) decreased dexamethasone-associated systemic toxicity, (2) targeted dexamethasone to reactive microglia, and (3) improved the regeneration enhancing effects of immunosuppression by increasing stem/progenitor proliferation rates. Lastly, we show that the gene rnf2 is required for the enhanced regeneration effect of D-Dex. These data support the use of dendrimer-based targeting of reactive immune cells to reduce toxicity and enhance the regeneration promoting effects of immunosuppressants in the retina.
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Affiliation(s)
- Kevin Emmerich
- McKusick-Nathans Institute of the Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - David T White
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Siva P Kambhampati
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
- The Center for Nanomedicine, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Grace L Casado
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Tian-Ming Fu
- Janelia Farms Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA
- Department of Electrical and Computer Engineering and Princeton Bioengineering Initiative, Princeton University, Princeton, NJ, USA
| | - Zeeshaan Chunawala
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Arpan Sahoo
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Saumya Nimmagadda
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Nimisha Krishnan
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Meera T Saxena
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA
| | - Steven L Walker
- School of Biomedical Sciences, The Chinese University of Hong Kong, Hong Kong, China
| | - Eric Betzig
- Janelia Farms Research Campus, Howard Hughes Medical Institute, Ashburn, VA, USA.
| | - Rangaramanujam M Kannan
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
- The Center for Nanomedicine, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
| | - Jeff S Mumm
- McKusick-Nathans Institute of the Department of Genetic Medicine, Johns Hopkins University School of Medicine, Baltimore, MD, USA.
- Department of Ophthalmology, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
- The Center for Nanomedicine, Wilmer Eye Institute, Johns Hopkins University, Baltimore, MD, USA.
- Solomon H Snyder Department of Neuroscience, Johns Hopkins University, Baltimore, MD, USA.
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Zhang Y, Yang X, Deng X, Yang S, Li Q, Xie Z, Hong L, Cao M, Yi G, Fu M. Single-cell transcriptomics-based multidisease analysis revealing the molecular dynamics of retinal neurovascular units under inflammatory and hypoxic conditions. Exp Neurol 2023; 362:114345. [PMID: 36736650 DOI: 10.1016/j.expneurol.2023.114345] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/10/2022] [Revised: 12/27/2022] [Accepted: 01/29/2023] [Indexed: 02/05/2023]
Abstract
The retinal neurovascular unit (NVU) is paramount to maintaining the homeostasis of the retina and determines the progression of various diseases, including diabetic retinopathy (DR), glaucoma, and retinopathy of prematurity (ROP). Although some studies have investigated these diseases, a combined analysis of disease-wide etiology in the NUV at the single-cell level is lacking. Herein, we constructed an atlas of the NVU under inflammatory and hypoxic conditions by integrating single-cell transcriptome data from retinas from wild-type, AireKO, and NdpKO mice. Based on the heterogeneity of the NVU structure and transcriptome diversity under normal and pathological conditions, we discovered two subpopulations of Müller cells: Aqp4hi and Aqp4lo cells. Specifically, Aqp4lo cells expresses phototransduction genes and represent a special type of Müller cell distinct from Aqp4hi cells, classical Müller cells. AireKO mice exhibit experimental autoimmune uveitis (EAU) with severe damage to the NVU structure, mainly degeneration of Aqp4hi cells. NdpKO mice exhibited familial exudative vitreoretinopathy (FEVR), with damage to the endothelial barrier, endothelial cell tight junction destruction and basement membrane thickening, accompanied by the reactive secretion of proangiogenic factors by Aqp4hi cells. In both EAU and FEVR, Aqp4hi cells are a key factor leading to NVU damage, and the mechanism by which they are generated is regulated by different transcription factors. By studying the pattern of immune cell infiltration in AireKO mice, we constructed a regulatory loop of "inflammatory cells/NVU - monocytes - APCs - Ifng+ T cells", providing a new target for blocking the inflammatory cascade. Our elucidation of the cell-specific molecular changes, cell-cell interactions and transcriptional mechanisms of the retinal NVU provides new insights to support the development of multipurpose drugs to block or even reverse NVU damage.
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Affiliation(s)
- Yuxi Zhang
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Xiongyi Yang
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Xiaoqing Deng
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Siyu Yang
- Department of Ophthalmology, The Seventh Affiliated Hospital, Sun Yat-Sen University, Shenzhen, PR China
| | - Qiumo Li
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Zhuohang Xie
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Libing Hong
- Zhujiang Hospital, Southern Medical University, Guangzhou, PR China; The Second Clinical School, Southern Medical University, Guangzhou, Guangdong, PR China
| | - Mingzhe Cao
- Department of Ophthalmology, The Seventh Affiliated Hospital, Sun Yat-Sen University, Shenzhen, PR China.
| | - Guoguo Yi
- Department of Ophthalmology, The Sixth Affiliated Hospital, Sun Yat-Sen University, No. 26, Erheng Road, Yuancun, Tianhe, Guangzhou, Guangdong, PR China.
| | - Min Fu
- Department of Ophthalmology, Zhujiang Hospital, Southern Medical University, Guangzhou, Guangdong, PR China.
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An Overview towards Zebrafish Larvae as a Model for Ocular Diseases. Int J Mol Sci 2023; 24:ijms24065387. [PMID: 36982479 PMCID: PMC10048880 DOI: 10.3390/ijms24065387] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2023] [Revised: 02/03/2023] [Accepted: 02/06/2023] [Indexed: 03/14/2023] Open
Abstract
Despite the obvious morphological differences in the visual system, zebrafish share a similar architecture and components of the same embryonic origin as humans. The zebrafish retina has the same layered structure and cell types with similar metabolic and phototransduction support as humans, and is functional 72 h after fertilization, allowing tests of visual function to be performed. The zebrafish genomic database supports genetic mapping studies as well as gene editing, both of which are useful in the ophthalmological field. It is possible to model ocular disorders in zebrafish, as well as inherited retinal diseases or congenital or acquired malformations. Several approaches allow the evaluation of local pathological processes derived from systemic disorders, such as chemical exposure to produce retinal hypoxia or glucose exposure to produce hyperglycemia, mimicking retinopathy of prematurity or diabetic retinopathy, respectively. The pathogenesis of ocular infections, autoimmune diseases, or aging can also be assessed in zebrafish larvae, and the preserved cellular and molecular immune mechanisms can be assessed. Finally, the zebrafish model for the study of the pathologies of the visual system complements certain deficiencies in experimental models of mammals since the regeneration of the zebrafish retina is a valuable tool for the study of degenerative processes and the discovery of new drugs and therapies.
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50
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Chen XD, Liu HL, Li S, Hu KB, Wu QY, Liao P, Wang HY, Long ZY, Lu XM, Wang YT. The latest role of nerve-specific splicing factor PTBP1 in the transdifferentiation of glial cells into neurons. WILEY INTERDISCIPLINARY REVIEWS. RNA 2023; 14:e1740. [PMID: 35574699 DOI: 10.1002/wrna.1740] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/21/2022] [Revised: 04/16/2022] [Accepted: 04/21/2022] [Indexed: 11/07/2022]
Abstract
Central nervous system injury diseases can cause the loss of many neurons, and it is difficult to regenerate. The field of regenerative medicine believes that supplementing the missing neurons may be an ideal method for nerve injury repair. Recent studies have found that down-regulation of polypyrimidine tract binding protein 1 (PTBP1) expression can make glial cells transdifferentiate into different types of neurons, which is expected to be an alternative therapy to restore neuronal function. This article summarized the research progress on the structure and biological function of the PTBP family, the mutual regulation of PTBP1 and PTBP2, their role in neurogenesis, and the latest research progress in targeting PTBP1 to mediate the transdifferentiation of glial cells into neurons, which may provide some new strategies and new ideas for the future treatment of central nervous system injury and neurodegenerative diseases. This article is categorized under: RNA Processing > Splicing Regulation/Alternative Splicing.
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Affiliation(s)
- Xing-Dong Chen
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China.,State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China
| | - Hui-Lin Liu
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China
| | - Sen Li
- State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China
| | - Kai-Bin Hu
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China
| | - Qing-Yun Wu
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China
| | - Ping Liao
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China
| | - Hai-Yan Wang
- State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China
| | - Zai-Yun Long
- State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China
| | - Xiu-Min Lu
- College of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing, China
| | - Yong-Tang Wang
- State Key Laboratory of Trauma, Burns and Combined Injury, Daping Hospital, Army Medical University, Chongqing, China
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