Investigating a role for eNAMPT-containing extracellular vesicles in mitigating age- and Alzheimer Disease-related cognitive decline

项目总结/摘要 认知功能的持续侵蚀是衰老的标志，在阿尔茨海默病中明显加重 （AD）。这些缺陷没有有效的治疗方法。烟酰胺腺嘌呤二核苷酸（NAD+）的消耗和 在衰老和AD中NAD+依赖性沉默调节蛋白活性的降低已经被充分证明。恢复NAD+或 激活沉默调节蛋白SIRT 1已经诱导了包括认知衰退在内的衰老症状的减轻。然而，在这方面， 目前的NAD+补充策略是非特异性的，SIRT 1激活作为抗衰老的成功 治疗可能需要组织特异性激活和伴随的NAD+恢复。我们已经表明 以前，烟酰胺磷酸核糖基转移酶（NAMPT），主要 在哺乳动物NAD+生物合成途径中，NAD+被包含在细胞外囊泡（EV）中并分泌到血浆中。 用含有细胞外NAMPT的血浆衍生EV（eNAMPT）治疗老年小鼠， 寿命和健康寿命。我们的初步数据表明EV治疗可以改善认知功能障碍 并在24个月大的小鼠中挽救海马CA 1突触计数的年龄相关性减少。电动汽车也是 靶向大脑，可以增加培养神经元中的NAD+。然而，组织特异性EV靶向 机制以及EV治疗在AD相关认知功能下降背景下的意义仍然存在 未知本研究将检验EV-包含的eNAMPT挽救与年龄相关的认知障碍的假设。 通过激活SIRT 1在关键神经元群体中的功能障碍，并利用这一途径可以减轻AD， 相关的认知能力下降在目标1中，将利用细胞培养衍生的EV来测试eNAMPT的必要性 在体外和体内对EV的靶向和摄取方面。在目标2中， 将评价通过eNAMPT介导的SIRT 1激活进行的EV治疗。中将该确定 通过EV治疗（有或无EV-含eNAMPT）和（有或无海马 通过病毒递送shRNA敲低Sirt 1。目标3将测试电动汽车在治疗认知障碍方面的有效性。 在AD小鼠模型中的缺陷和病理进展。因此，这项研究有可能 阐明了eNAMPT在促进EV的细胞靶向和EV在治疗中的功效方面的新作用。 挽救与年龄和AD相关的认知功能障碍。该项目还可以建立包含eNAMPT的EV 作为一种可行的生物制剂来治疗衰老和AD中的认知衰退。本研究金提案将 NAD+生物学和老龄化专业知识的赞助商新今井博士与综合培训的行为 神经科学、AD模型、高级突触分析、EV工程和实验室指导， 领导这种培训将进一步增强智力刺激和高度协作 圣路易斯华盛顿大学的环境。因此，该奖学金将构成理想的准备 申请人的职业生涯作为一个学术实验室的未来领导者。

PROJECT SUMMARY/ABSTRACT The steady erosion of cognitive function is a hallmark of aging and markedly exacerbated in Alzheimer Disease (AD). These deficits have no effective treatments. Depletion of nicotinamide adenine dinucleotide (NAD+) and reduction of NAD+-dependent sirtuin activity in aging and AD have been well documented. Restoring NAD+ or activating the sirtuin SIRT1 has induced abatement of aging symptoms including cognitive decline. However, current NAD+ replenishment strategies are non-specific and the success of SIRT1 activation as an anti-aging therapeutic may require tissue-specific activation and the concomitant restoration of NAD+. We have shown previously that nicotinamide phosphoribosyltransferase (NAMPT), the rate-limiting enzyme in the major mammalian NAD+ biosynthesis pathway, is contained in extracellular vesicles (EVs) and secreted to plasma. Treatment of aged mice with plasma-derived EVs containing extracellular NAMPT (eNAMPT) extends mouse lifespan and healthspan. Our preliminary data suggest that EV treatment can ameliorate cognitive dysfunction and rescue age-related decreases in hippocampal CA1 synapse counts in 24-month-old mice. EVs are also targeted to the brain and can increase NAD+ in cultured neurons. However, the tissue-specific EV targeting mechanism as well as the implications for EV treatment in the context of AD-related cognitive decline remain unknown. This study will test the hypothesis that EV-contained eNAMPT rescues age-related cognitive dysfunction via activating SIRT1 in key neuronal populations and that leveraging this pathway can lessen AD- associated cognitive decline. In Aim 1, cell culture-derived EVs will be utilized to test the necessity of eNAMPT in the targeting and uptake of EVs both in vitro and in vivo. In Aim 2, the possibility that the cognitive benefits of EV treatment occur through eNAMPT-mediated activation of SIRT1 will be evaluated. This determination will be accomplished via EV treatment with and without EV-contained eNAMPT and with and without hippocampal knockdown of Sirt1 by viral delivery of shRNA. Aim 3 will test the effectiveness of EVs in treating cognitive deficits and pathological progression in a mouse model of AD. Therefore, this study has the potential to elucidate a novel role for eNAMPT in facilitating both the cellular targeting of EVs and the efficacy of EVs in rescuing age- and AD-related cognitive dysfunction. This project may also establish eNAMPT-containing EVs as a viable biologic to treat cognitive deterioration in aging and AD. This fellowship proposal integrates the NAD+ biology and aging expertise of sponsor Dr. Shin Imai with comprehensive training in behavioral neuroscience, AD models, advanced synaptic analysis, EV engineering, and laboratory mentorship and leadership. This training will be further empowered by the intellectually stimulating and highly collaborative environment of Washington University in St. Louis. Thus, this fellowship will constitute ideal preparation for the applicant’s career as the future leader of an academic laboratory.