Regulating cell-cell interactions to improve donor retinal ganglion cell integration

调节细胞间相互作用以改善供体视网膜神经节细胞整合

基本信息

批准号：
10559535
负责人：
Jonathan R Soucy
金额：
$ 7.22万
依托单位：
SCHEPENS EYE RESEARCH INSTITUTE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-01 至 2025-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10559535
关键词：
Affect Animals Biological Models Blindness CDK5 gene Cell Adhesion Molecules Cell Communication Cell Separation Cell Surface Receptors Cell Survival Cell Transplantation Cells Cellular Morphology Cellular biology Cerebrum Colcemid Cues Cytoskeleton Development Disease Dose Down Syndrome Down Syndrome Cell Adhesion Molecule Electrophysiology (science)Event Ganglion Cell Layer Glaucoma Harvest Image Immunohistochemistry Investigation Isogenic transplantation Lead Life Expectancy Masks Mediating Modality Molecular Mus Muscle fasciculation Natural regeneration Neurodegenerative Disorders Neurons Optic Nerve Outcome Peptide Hydrolases Persons Pharmaceutical Preparations Population Positioning Attribute Process Reproducibility Research Proposals Retina Retinal Degeneration Retinal Ganglion Cells Role Series Signal Transduction Small Interfering RNA Spinal Surface System Transplantation Trisomy United States Vertebral column age related cell motility cell replacement therapy economic impact experimental study gain of function improved induced pluripotent stem cell inhibitor intravitreal injection loss of function loss of function mutation migration mosaic mosaic pattern multi-electrode arrays neural neural circuit neuron loss neuronal cell body neuronal replacement optic nerve disorder preservation prevent receptor expression repaired retina transplantation retinal damage retinal neuron retinogenesis sight restoration small molecule socioeconomics stem cell differentiation stem cells synaptogenesis transcriptome

项目摘要

Project Summary An estimated 3 million people are affected by glaucoma in the United States, and increasing life expectancy exacerbates the disease’s socio-economic impact. Glaucoma and other optic neuropathies lead to permanent damage of the optic nerve and loss of retinal ganglion cells (RGCs). No therapies are currently available to mitigate irreversible vision loss. The feasibility of cell replacement therapy was recently demonstrated using RGCs isolated from a developing retina. Furthermore, we have shown that it is possible to achieve robust and reproducible transplants with stem cell-derived RGCs. While our donor RGCs survived in host retinas following transplantation, cell survival does not equate to the restoration of vision, and poor structural and functional integration remains a significant challenge for successful RGC replacement. One of the key molecular features limiting donor RGC integration into the existing circuitry is likely to be the vestigial homophilic molecular cues that guide somatic spacing and dendritic arborization during development. Down Syndrome Cell Adhesion Molecule (DSCAM) has been identified as a key molecular cue that mediates neuronal self-avoidance to prevent fasciculation and preserve mosaic spacing in the retina during development. We hypothesize that these same mechanisms govern the integration of transplanted RGCs and that homophilic molecular cues, including DSCAM, limit donor RGC migration towards their natural connecting points within the retina. Therefore, this proposal aims to investigate DSCAM in the context of RGC transplantation to understand how self-avoidance mechanisms contribute to neural circuit development and repair. Using RGC transplantation into the retina as a model system, we will determine if DSCAM-mediated self- avoidance mechanisms are dose-mediated, rely on transcellular interactions, and function similarly irrespective of neural migration. To investigate the need for transcellular DSCAM expression for RGC self-avoidance, we will conduct a series of transplantation experiments using gain- and lose-of-function (GOF and LOF) mice. Expression of DSCAM by mouse stem cell-derived RGC will be suppressed with siRNA before intravitreal injections. Similarly, to investigate if DSCAM regulates donor RGC spacing independent of the mode of migration, we will suppress DSCAM in host and donor RGCs while temporarily destabilizing the donor RGC’s cytoskeleton to alter their migratory modality between somal translocation and multipolar migration. Live imaging and quantitative immunohistochemistry will be used to assess donor cell morphology and distribution in the retina. Anterograde tracing and retinal explants cultured on multielectrode arrays will be used to evaluate synapse formation with host bipolar cells. Altogether, this mechanistic approach would significantly impact the development of cell replacement therapy for glaucoma and other neurodegenerative diseases.

项目概要据估计，美国有 300 万人受到青光眼的影响，并且预期寿命不断延长青光眼和其他视神经病变会导致永久性的目前尚无治疗方法可用于治疗视神经损伤和视网膜神经节细胞 (RGC) 损失。最近使用细胞替代疗法证明了减轻不可逆视力丧失的可行性。此外，我们已经证明，从发育中的视网膜中分离出 RGC 是有可能实现稳健且稳定的。使用干细胞衍生的 RGC 进行可重复移植，而我们的供体 RGC 在宿主视网膜中存活下来。移植后，细胞存活不等于视力恢复，结构和功能较差集成仍然是成功替代 RGC 的一个重大挑战。限制供体 RGC 整合到现有电路中的关键分子特征之一可能是在发育过程中指导体细胞间距和树突分枝的残余同质分子线索。唐氏综合症细胞粘附分子 (DSCAM) 已被确定为介导唐氏综合症细胞粘附分子的关键分子线索神经元自我回避，以防止发育过程中的束震并保留视网膜中的马赛克间距。我们着迷的是，这些相同的机制控制着移植的 RGC 的整合，并且同质分子线索，包括 DSCAM，限制供体 RGC 向其自然迁移因此，本提案旨在研究 RGC 背景下的 DSCAM。移植以了解自我回避机制如何促进神经回路的发育和维修。使用 RGC 移植到视网膜作为模型系统，我们将确定 DSCAM 是否介导自我调节回避机制是剂量介导的，依赖于跨细胞相互作用，并且无论何种情况都具有相似的功能为了研究 RGC 自我回避的跨细胞 DSCAM 表达的必要性，我们将使用功能获得和功能丧失（GOF 和 LOF）小鼠进行一系列移植实验。玻璃体内注射前，小鼠干细胞来源的 RGC 的 DSCAM 表达将被 siRNA 抑制类似地，研究 DSCAM 是否独立于注射模式调节供体 RGC 间距。迁移时，我们将抑制宿主和供体 RGC 中的 DSCAM，同时暂时破坏供体 RGC 的稳定性细胞骨架改变其在体易位和多极迁移之间的迁移方式。免疫组织化学将用于评估定量供体细胞的形态和分布顺行追踪和在多电极阵列上培养的视网膜外植体将用于评估总而言之，这种机制方法将显着影响宿主双极细胞的突触形成。开发针对青光眼和其他神经退行性疾病的细胞替代疗法。