VRC: Engineered extracellular vesicles for mild TBI-induced retinal injury

VRC：工程细胞外囊泡治疗轻度 TBI 引起的视网膜损伤

• 批准号: 10688145
• 负责人: STEVEN ROTH
• 金额: $ 31.76万
• 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: 2022-09-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10688145
• 关键词: Address; Adjuvant; Affect; Anti-Inflammatory Agents; Antiinflammatory Effect; Apoptosis; Athletic Injuries; Attenuated; Biology; Blindness; Cell Death; Cell Line; Cells; Cessation of life; Cytoprotection; Data; Diabetic Retinopathy; Engineering; Exposure to; Eye; Functional disorder; Genes; Glutamates; Goals; Health; Hypoxia; In Vitro; Inflammation; Inflammation Mediators; Inflammatory; Injury; Interruption; Knowledge; Laboratories; Left; Lentivirus Vector; Mesenchymal Stem Cells; Methodology; MicroRNAs; Microglia; Muller' s cell; Natural regeneration; Neurodegenerative Disorders; Neuronal Dysfunction; Parents; Pathway interactions; Play; Precision therapeutics; Public Health; Publishing; Retina; Retinal Diseases; Retinal Ganglion Cells; Rodent; Rodent Model; Role; Specificity; Sports; Strategic Planning; Stress; Testing; Therapeutic; Translating; Trauma; Traumatic Brain Injury; Tropism; United States; Visual; antagonist; automobile accident; clinically relevant; combat; cranium; cytokine; disability; exosome; experimental study; extracellular vesicles; functional mimics; glial activation; human disease; human model; immunoregulation; improved; in vivo; innovation; knock-down; loss of function; mild traumatic brain injury; mouse model; neuroinflammation; neuron loss; novel; overexpression; precision medicine; preconditioning; prevent; repaired; response; retinal damage; retinal neuron; stem cells; stroke model; transcriptome sequencing; translational medicine; visual dysfunction

Traumatic brain injury (TBI) is the leading cause of vision loss in sports injury, and automobile accidents in the United States, for which there currently is no treatment. Our long-term objective is to develop safe, innovative strategies to combat vision loss from TBI. Mild TBI (mTBI), even without direct trauma to the eye, results in retinal damage via microglia and Müller cell-driven inflammation, and dysfunction from loss of retinal ganglion cells (RGCs). Mesenchymal stem cell (MSC) extracellular vesicles (EVs) are rapidly emerging as a stem cell alternative that promote immunomodulation, repair, and regeneration. MSC EVs injected into the vitreous in rodents demonstrated tropism for RGCs, Müller cells, and microglia, and triggered specific cellular responses, with micro RNA (miRNA) from the EVs playing a key role. Here we capitalize upon our published and preliminary data demonstrating that MSC EVs attenuate microglial activation and RGC death in vitro. Anti-apoptosis and anti-inflammatory effects of MSC EVs were further boosted by “supercharging” the EVs using hypoxic preconditioning of the parent MSCs, yielding “H-EVs.” MSC cell lines were genetically altered to stably overexpress microRNAs in EVs (i.e., Functionally-Engineered EVs, or FEEs), and FEEs mimicked the anti-apoptosis and anti-inflammatory action of H-EVs. This proposal targets interrupting retinal damage from mild TBI with MSC-EVs containing function-specific miRNA to attenuate neuro-inflammation and accompanying neuronal dysfunction and cell death. Our central hypothesis is that targeted EV-specific expression of key miRNAs ameliorates microglia and Müller cell-driven inflammation, and loss of RGCs and subsequent visual dysfunction from mTBI. Aim 1 engineers EVs mimicking functionality of H-EVs to attenuate microglia and Müller cell activation and retinal neuron death in vitro. We hypothesize that FEEs enhance anti-inflammatory and anti-apoptosis effects of MSC-EVs and mimic H-EVs. Two of the highest expressed miRs in H-EVs will be studied as FEEs in comparison to MSC EVs, H-EVs, and controls in activated retinal microglia, and RGCs and Müller cells exposed to glutamate. Identifying mechanisms of action of FEEs and H-EVs will provide a novel pathway to therapeutic precision medicine for visual dysfunction in mTBI. Aim 2 determines in vivo anti-inflammatory and anti-apoptosis actions of FEEs in a clinically-relevant mouse model of mTBI. We will test the hypothesis that FEEs administered into the vitreous rescue the retina when given after mTBI. Retinal function, apoptosis, glutamate levels, and inflammatory mediators post mTBI using 50 psi blast to the cranium will be quantitatively compared, and cell-specific effects identified in groups treated with H- EVs, FEEs, MSC EVs, and controls. The proposed studies are expected to provide new results with MSC-EVs modified for protective action to treat the retinal cells affected by TBI, and provide novel potential therapy for other neurodegenerative disorders.

创伤性脑损伤 (TBI) 是导致运动损伤和汽车事故中视力丧失的主要原因 美国目前尚无治疗方法。我们的长期目标是开发安全、创新的 对抗 TBI 引起的视力丧失的策略。轻度 TBI (mTBI)，即使没有直接伤害眼睛，也会导致视网膜损伤 小胶质细胞和穆勒细胞驱动的炎症造成的损伤，以及视网膜神经节细胞损失造成的功能障碍 （RGC）。间充质干细胞 (MSC) 细胞外囊泡 (EV) 作为一种干细胞正在迅速崛起 促进免疫调节、修复和再生的替代方案。 MSC EVs 注入玻璃体 啮齿动物表现出对 RGC、Müller 细胞和小胶质细胞的趋向性，并引发特定的细胞反应， EV 中的微小 RNA (miRNA) 发挥着关键作用。 在这里，我们利用已发表的初步数据证明 MSC EV 可以减弱小胶质细胞 体外激活和 RGC 死亡。 MSC EVs的抗凋亡和抗炎作用进一步增强 通过使用母体 MSC 的缺氧预处理对 EV 进行“增压”，产生“H-EV”。间充质干细胞系 经过基因改造，可在 EV 中稳定过度表达 microRNA（即功能工程 EV 或 FEE）， FEE 模仿了 H-EV 的抗凋亡和抗炎作用。该提案的目标是中断 使用含有功能特异性 miRNA 的 MSC-EV 减轻轻度 TBI 引起的视网膜损伤，以减轻神经炎症 以及伴随的神经元功能障碍和细胞死亡。我们的中心假设是针对电动汽车特定 关键 miRNA 的表达可改善小胶质细胞和 Müller 细胞驱动的炎症以及 RGC 和 RGC 的损失 mTBI 导致的后续视觉功能障碍。 Aim 1 工程师设计了模仿 H-EV 功能的 EV，以减弱小胶质细胞和 Müller 细胞的激活， 体外视网膜神经元死亡。我们假设 FEE 增强了抗炎和抗凋亡作用 MSC-EV 和模拟 H-EV。 H-EV 中表达最高的两个 miR 将作为 FEE 进行研究进行比较 MSC EV、H-EV 以及暴露于谷氨酸的活化视网膜小胶质细胞、RGC 和 Müller 细胞中的对照。 确定 FEE 和 H-EV 的作用机制将为实现治疗精准度提供新途径 治疗 mTBI 视觉功能障碍的药物。 目标 2 确定 FEE 在临床相关小鼠体内的抗炎和抗凋亡作用 mTBI 模型。我们将检验以下假设：将 FEE 注入玻璃体可挽救视网膜 mTBI 后。 mTBI 后使用 50 psi 的视网膜功能、细胞凋亡、谷氨酸水平和炎症介质 将定量比较颅骨爆炸，并确定 H-治疗组中的细胞特异性效应 EV、FEE、MSC EV 和控件。 拟议的研究预计将提供经过修改的 MSC-EV 的新结果，以起到保护作用 治疗受 TBI 影响的视网膜细胞，并为其他神经退行性疾病提供新的潜在疗法。