Mesenchymal stem cell extracellular vesicles for ischemic retinal damage

间充质干细胞胞外囊泡治疗缺血性视网膜损伤

• 批准号: 10843511
• 负责人: STEVEN ROTH
• 金额: $ 5.16万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-30 至 2026-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10843511
• 关键词: Acute; Address; Adjuvant; Affect; Alteplase; Animal Model; Anti-Inflammatory Agents; Antiinflammatory Effect; Apoptosis; Apoptotic; Attenuated; Basic Science; Biology; Biomimetics; Blindness; Blood-Retinal Barrier; Cardiac Surgery procedures; Case Series; Cell Death; Cell secretion; Cells; Central Retinal Artery Occlusion; Characteristics; Cytokine Gene; Cytoprotection; Dependence; Dermal; Diabetic Retinopathy; Disease; Economic Burden; Elderly; Emergency Situation; Engineering; Enrollment; Event; Eye; Face; Filler; Generations; Growth; Hyperbaric Oxygen; Hypoxia; Iatrogenesis; Immune response; In Vitro; Incidence; Inflammation; Inflammatory; Injections; Interruption; Intervention; Intravenous; Ischemia; Knowledge; Laboratories; Long-Term Care; Massage; Mediating; Mesenchymal Stem Cells; Methodology; MicroRNAs; Microglia; Modeling; Molecular; Muller' s cell; Natural regeneration; Neurons; Paracentesis; Pathway interactions; Patients; Permeability; Physiologic Intraocular Pressure; Play; Property; Public Health; Publishing; Reporting; Retina; Retinal Diseases; Retinal Ganglion Cells; Rodent Model; Role; Specificity; Stroke; System; Testing; Therapeutic; Translating; Vascular Diseases; Vascular Endothelial Cell; Vision; antagonist; anterior chamber; cell injury; cerebrovascular; clinical translation; combat; disability burden; effectiveness evaluation; experimental study; extracellular vesicles; falls; functional disability; functional mimics; gain of function; human disease; human model; immunoregulation; in vivo; innovation; ischemic injury; knock-down; loss of function; model development; nanovesicle; neuroinflammation; neuroprotection; novel; overexpression; precision medicine; preconditioning; prevent; protective effect; randomized, clinical trials; repaired; response; restoration; retinal damage; retinal ischemia; retinal neuron; stem cells; stroke model; success; thrombolysis; tool; transcriptome sequencing; translational medicine; vascular injury

Central retinal artery occlusion (CRAO) is an ophthalmological emergency with few proven therapies. Stem cell-based retinal cell replacement is a highly encouraging approach to achieve retinal neuroprotection and to save vision in retinal diseases. However, with limitations including few cells integrated, adverse immune re- sponses, and aberrant growth, an alternative cell-free approach is required. EVs are nano-vesicular bodies that, when endocytosed by target cells, trigger specific responses. Here, the microRNA (miRNA) cargo of the EVs plays a key role. This proposal targets restoration of retinal function using engineered MSC-EVs containing function-specific miRNA. Compared to MSCs, their EVs are non-immunogenic, non-tumorigenic, and modifiable for specific delivery modes. These characteristics render them ideal biomimetic agents fitting precision-based medicine. Our studies indicate that EVs can rescue retinal cells that have been acutely subjected to hypoxia or ischemia, the key mechanism that starts cells dying in CRAO. We also found that hypoxic preconditioning of MSCs resulted in EVs (H-EVs) with enhanced cytoprotective properties including anti-apoptosis and anti-inflam- mation. A number of miRNAs overexpressed in the H-EVs have cytoprotective properties in retinal cells. Our central hypothesis is that targeted EV-specific expression of miR-424/other key miRNAs in MSC-EVs will re-capitulate the anti-apoptosis and anti-inflammatory actions of H-EVs. We designate such EVs as Func- tionally Engineered EVs (FEEs). To facilitate clinical translation of MSC-EV therapy, we have identified key fun- damental knowledge gaps: (1) The relationship between EV miRNA and its anti-apoptotic properties; (2) EV miRNA and its role in anti-inflammatory actions of MSC-EVs in retina; and (3) Can MSC-EVs be enhanced for targeted functionality by engineering their miRNA cargo? In Aim 1 we will produce FEEs overexpressing miR- 424 (FEE-424) and 146b (FEE-146b). We will evaluate the mechanisms of action of the FEEs, and their candi- dacy for generation of FEEs in retinal ganglion cells, microglia, Muller cells, and retinal vascular endothelial cells using loss and gain of function studies in models of simulated ischemia in vitro. These results will serve as a proof-of-principle model for development of FEEs for amelioration of cell damage in the retina. In Aim 2, FEEs containing miR-424 and -146b will be used to test specific targeting of anti-apoptotic and inflammatory mecha- nisms in a rodent model of CRAO. Overall, the proposed studies are expected to provide transformative results whereby MSC-EVs are modified and delivered for retinal protective action after the ischemic event to treat CRAO. Innovations are cell-free therapy of retinal diseases, EV miR-mediated application-specificity, and direct determination of the impact of EVs on specific cells involved in retinal ischemic injury. Translational significance is the high likelihood of impacting novel molecular therapy. Underlying basic research significance is that the studies will enable vertical advancement of the field by determining mechanisms of actions of EVs in the retina.

视网膜中央动脉阻塞（CRAO）是一种眼科急症，目前几乎没有经过验证的治疗方法。干 基于细胞的视网膜细胞替代是一种非常令人鼓舞的方法，可实现视网膜神经保护和 挽救视网膜疾病的视力。然而，由于细胞整合较少等限制，不利的免疫重新 反应和异常生长，需要一种替代的无细胞方法。 EV 是纳米囊泡体， 当被靶细胞内吞时，触发特定反应。此处，EV 的 microRNA (miRNA) 货物 发挥着关键作用。该提案的目标是使用工程化的 MSC-EV 恢复视网膜功能，其中包含 功能特异性 miRNA。与 MSC 相比，它们的 EV 不具有免疫原性、非致瘤性且可修饰 对于特定的交付模式。这些特性使它们成为基于精确拟合的理想仿生剂 药品。我们的研究表明，EV 可以拯救严重缺氧或严重缺氧的视网膜细胞。 缺血是 CRAO 中细胞死亡的关键机制。我们还发现，低氧预处理 MSC 产生的 EV（H-EV）具有增强的细胞保护特性，包括抗凋亡和抗炎作用 化。 H-EV 中过表达的许多 miRNA 在视网膜细胞中具有细胞保护特性。 我们的中心假设是 MSC-EV 中 miR-424/其他关键 miRNA 的靶向 EV 特异性表达 将重现 H-EV 的抗凋亡和抗炎作用。我们将此类 EV 指定为 Func- 工程电动汽车（FEE）。为了促进 MSC-EV 疗法的临床转化，我们确定了关键的功能 基础知识空白：（1）EV miRNA与其抗凋亡特性的关系； (2) 电动车 miRNA及其在MSC-EVs视网膜抗炎作用中的作用； (3) MSC-EV 能否增强 通过设计 miRNA 货物来实现目标功能？在目标 1 中，我们将生产过表达 miR-的 FEE 424 (FEE-424) 和 146b (FEE-146b)。我们将评估 FEE 的作用机制及其候选者 dacy 用于在视网膜神经节细胞、小胶质细胞、Muller 细胞和视网膜血管内皮细胞中生成 FEE 在体外模拟缺血模型中使用功能丧失和获得的研究。这些结果将作为 开发用于改善视网膜细胞损伤的 FEE 的原理验证模型。在目标 2 中，费用 含有miR-424和-146b的药物将用于测试抗凋亡和炎症机制的特异性靶向 CRAO 啮齿动物模型中的 Nisms。总体而言，拟议的研究预计将提供变革性的结果 由此，MSC-EV 被修改并在缺血事件后提供视网膜保护作用，以治疗 CRAO。 创新包括视网膜疾病的无细胞疗法、EV miR介导的应用特异性以及直接治疗 确定 EV 对参与视网膜缺血性损伤的特定细胞的影响。翻译意义 影响新型分子疗法的可能性很高。根本性的基础研究意义在于 研究将通过确定 EV 在视网膜中的作用机制来实现该领域的垂直发展。