Neural Stem Cell-derived EVs for Improving Aged Brain Function

项目摘要 该项目的主要目标是开发一种非侵入性的细胞外水泡(EV)疗法来维持 老年人的认知和记忆功能较好。该项目建议检查以下项目的有效性 人诱导多能干细胞(HiPSC)来源的EVS的鼻腔给药 神经干细胞(HNSCs)用于改善衰老模型小鼠的认知和记忆功能。RNA序列 蛋白质组学研究证实了miRNAs和具有神经保护作用的蛋白质的存在， HNSC-EVS的抗炎和神经生成特性。初步研究表明，IN 给予hNSC-EVS会导致它们被所有脑区的小胶质细胞和神经元摄取，并导致 改善中年小鼠的认知功能，与减轻神经炎症有关， 一磷酸腺苷活化蛋白激酶(AMPK)活化，雷帕霉素的哺乳动物靶点 (MTOR)抑制，增强自噬，并改善神经发生。因此，该计划的理由是 提出的方法是hNSC-Evs的miRNA和蛋白质载体将激活有益的信号转导。 在老年大脑中的靶细胞，如小胶质细胞、神经元和神经干细胞中的通路，这反过来， 会减少神经炎症，增强神经元功能和神经再生，从而改善 认知和记忆功能。该项目将测试假设，在hNSC-EVS的管理中 中年早期或晚期在老年时保持较好的认知功能，在老年时使用hNSC-EV会保持较好的认知功能 年龄会逆转认知功能障碍。此外，预计hNSC的功能改进-- EV对老年脑的治疗将涉及通过抑制NLRP3来调节激活的小胶质细胞 炎性小体，AMPK激活，线粒体功能改善，mTOR抑制，增强自噬， 改善神经发生，减少突触丢失。利用老鼠的衰老模型，这个项目将首先 研究在中年早期或中年晚期使用EVS预防认知功能障碍和 老年记忆障碍(特定目标1)。在特定目标2，老年使用电动汽车的效果 为了逆转认知和记忆障碍，将对其进行测量。然后，为了了解小胶质细胞的作用 在hNSC-EV介导的认知功能改善中的调制，小胶质细胞耗竭时的影响 EV给药将在中年和老年小鼠中进行检查(特定目标3)。由于小胶质细胞 衰老小鼠的耗竭改善了认知功能，hNSC-EV治疗的任何相加效应 小胶质细胞耗竭的影响表明hNSC-EVS也作用于其他类型的神经细胞或途径 调节治疗效果。大脑的两个区域，即海马体和内侧前额叶皮质 (MPFC)，将在雄性和雌性老年小鼠身上进行分析。拟议的研究与以下方面高度相关 预防和治疗老年相关的认知和记忆障碍。

Project Summary This project's primary goal is to develop a non-invasive, extracellular vesicle (EV) therapy for maintaining better cognitive and memory function in old age. The project proposes to examine the effectiveness of intranasal (IN) administration of EVs generated from human induced pluripotent stem cell (hiPSC)-derived neural stem cells (hNSCs) for improving cognitive and memory function in a mouse model of aging. RNA-seq and proteomic studies have confirmed the presence of miRNAs and proteins having neuroprotective, antiinflammatory, and neurogenic properties in hNSC-EVs. Preliminary studies have suggested that IN administration of hNSC-EVs results in their uptake by microglia and neurons in all brain regions and leads to improved cognitive function in middle-aged mice, in association with diminished neuroinflammation, adenosine monophosphate-activated protein kinase (AMPK) activation, mammalian target of rapamycin (mTOR) inhibition, enhanced autophagy, and improved neurogenesis. Therefore, the rationale for the proposed approach is that the miRNA and protein cargo of hNSC-EVs would activate beneficial signaling pathways in target cells such as microglia, neurons, and neural stem cells in the aged brain, which, in turn, would reduce neuroinflammation, enhance neuronal function and neurogenesis, and thereby improve cognitive and memory function. The project will test the hypothesis that IN administration of hNSC-EVs in early or late middle age will maintain better cognitive function in old age, and hNSC-EV administration in old age will reverse cognitive dysfunction. Furthermore, it is envisaged that functional improvements with hNSC- EV treatment in the aged brain will involve modulation of activated microglia with inhibition of NLRP3 inflammasomes, AMPK activation, improved mitochondrial function, mTOR inhibition, enhanced autophagy, improved neurogenesis, and reduced synapse loss. Using a mouse model of aging, this project will first investigate the effectiveness of EVs administered in the early or late middle age to prevent cognitive and memory dysfunction in old age (Specific Aim 1). In Specific Aim 2, the effect of EVs administered in old age for reversing cognitive and memory dysfunction will be measured. Then, to understand the role of microglial modulation in hNSC-EV-mediated improved cognitive function, the effects of microglia depletion at the time of EV administration will be examined in both middle-aged and aged mice (Specific Aim 3). Since microglia depletion in the aged mice improves cognitive function, any additive effects of hNSC-EV treatment over the effects of microglia depletion would suggest that hNSC-EVs also act on other neural cell types or pathways to mediate therapeutic benefits. Two regions of the brain, the hippocampus, and the medial prefrontal cortex (mPFC), will be analyzed in male and female aged mice. The proposed studies are highly relevant for preventing and treating age-related cognitive and memory impairments in old age.