Supplement to EY021482 to carry out a screen for retinal regeneration using CRISPR-Cas9 gene activation.

补充 EY021482，利用 CRISPR-Cas9 基因激活进行视网膜再生筛选。

基本信息

批准号：
9313143
负责人：
THOMAS A REH
金额：
$ 6.33万
依托单位：
UNIVERSITY OF WASHINGTON
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-04-01 至 2018-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9313143
关键词：
Affect Age related macular degeneration Amphibia Area Azacitidine Biochemical Biological Assay Blindness Brain CRISPR/Cas technology Cell Cycle Cells ChIP-seq Chromatin Complete Blindness DNA DNA Methylation Degenerative Disorder Development Differentiated Gene Disease Electrophysiology (science)Epigenetic Process Fishes Funding Gene Activation Gene Expression Genes Genetic Glaucoma Goals Histologic Homologous Gene Human Impairment In Vitro Inherited Injury Ischemia Lead Mammals Mediating Modeling Modification Morphology Muller&apos s cell Mus Natural regeneration Nervous system structure Neuroglia Neurons Neurotransmitters Patients Phenotype Photoreceptors Process Production Proliferating Regenerative response Retina Retinal Retinal Degeneration Retinal Diseases Retinitis Pigmentosa Sensory Source Stem cell transplant Testing Tetanus Helper Peptide Transgenic Mice Vertebrates Viral Visual Visual impairment advanced disease demethylation differential expression effective therapy experimental study follow-up genome-wide genome-wide analysis histone modification in vivo in vivo regeneration novel strategies photoreceptor degeneration prevent progenitor programs promoter public health relevance regenerative repaired response retinal damage retinal neuron retinal progenitor cell retinal regeneration transcription factor treatment strategy

项目摘要

DESCRIPTION (provided by applicant): Like other areas of the nervous system, the retina is subject to many acquired and inherited neuronal degenerative diseases. Since the retina provides the input for all visual sensory information to the brain, the loss of cells results in viual impairment and potentially complete blindness. Many retinal degenerative diseases affect only a subset of the retinal cells, although, frequently in more advanced disease, loss and reorganization of the entire retina can occur. In mammals, there is very limited regeneration of the degenerated cells; however, in fish, new neurons of all types regenerate from M¿ller glia following retinal damage and they are functionally integrated into the existing circuitry. Nevertheless, M¿ller glia, the cellular source for regeneration, is present in all vertebrate retins. In the proposal we submitted three years ago, we hypothesized that regeneration from mammalian M¿ller glia was limited because they fail to express a key proneural transcription factor, Ascl1, after injury. We proposed to test this hypothesis by virally-mediated expression of Ascl1 in mouse M¿ller glia. In the two years of funding, we have tested the hypothesis, and found that viral expression of Ascl1 is sufficient to activate a neurogenic program in mouse M¿ller glia, both in dissociated cultures and in the intact retina. The reprogrammed M¿ller glia generates cells that resemble neurons in morphology, gene expression and their responses to neurotransmitters. In the next funding period, we propose to further optimize this reprogramming process, using other transcription factors and epigenetic modifiers, and then to test whether Ascl1-reprogrammed Muller glia can provide a source for regeneration in vivo in a newly developed line of transgenic mice.

描述（由适用提供）：与神经系统的其他区域一样，视网膜受到许多获得和遗传的神经元退行性疾病的约束。由于视网膜为大脑提供了所有视觉感觉信息的输入，因此细胞的损失会导致视觉障碍和潜在的完全失明。许多视网膜退行性疾病仅影响视网膜细胞的一部分，尽管在哺乳动物中整个视网膜的疾病损失和重组中经常经常影响，但退化的细胞的再生非常有限。然而，在鱼类中，所有类型的新神经元在残留损伤后从m ller胶质中再生，并且在功能上将其整合到现有电路中。然而，在所有脊椎动物视网膜中都存在于再生的细胞来源M ller Glia。在我们三年前提交的提案中，我们假设哺乳动物的再生是有限的，因为它们在受伤后未能表达关键的主权转录因子ASCL1。我们提出了通过小鼠胶质神经胶质中ASCL1的病毒介导的ASCL1表达来检验这一假设。在两年的资金中，我们已经检验了该假设，发现ASCL1的病毒表达足以激活小鼠麦粒中的神经源程序，无论是在解离培养物和完整的视网膜中。重编程的M ller Glia产生的细胞类似于形态学，基因表达及其对神经递质的反应。在下一个资金期间，我们建议使用其他转录因子和表观遗传修饰符进一步优化此重编程过程，然后测试ASCL1编程的Muller Glia是否可以在新开发的转基因小鼠系列中提供体内再生的来源。