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

项目摘要/摘要 认知功能的稳步下降是衰老的一个标志，在阿尔茨海默病中明显加剧 (Ad)。这些赤字没有有效的治疗方法。烟酰胺腺嘌呤二核苷酸(NAD+)和 NAD+依赖的sirtuin活性在衰老和AD中的降低已经被很好地记录下来。恢复NAD+或 激活sirtuin SIRT1可以缓解衰老症状，包括认知能力下降。然而， 目前的NAD+补充策略是非特异性的，SIRT1的成功激活是一种抗衰老 治疗可能需要组织特异性激活和伴随的NAD+的恢复。我们已经展示了 以前认为烟酰胺磷酸核糖转移酶(NAMPT)是主要的限速酶 哺乳动物NAD+生物合成途径存在于细胞外小泡(EVS)中，并分泌到血浆中。 含胞外NAMPT的血浆源性EVS(ENAMPT)治疗老龄小鼠 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 并挽救24月龄小鼠海马区CA1突触计数与年龄相关的减少。电动汽车也是 靶向于大脑，可以增加培养神经元中的NAD+。然而，组织特异性EV靶向 阿尔茨海默病相关认知功能减退的机制及其对EV治疗的影响仍然存在 未知。这项研究将检验含有EV的eNAMPT拯救年龄相关认知的假设 通过激活关键神经元群体中的SIRT1而导致的功能障碍，以及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EVS来测试eNAMPT的必要性 在体外和体内靶向和摄取EVS。在目标2中，认知益处的可能性 EV治疗是通过eNAMPT介导的SIRT1激活进行的，将进行评估。这一决心将 通过使用和不使用包含EV的eNAMPT以及使用和不使用海马体的EV治疗来完成 通过病毒传递shRNA来敲除Sirt1。目标3将测试EVS在治疗认知方面的有效性 阿尔茨海默病小鼠模型的缺陷和病理进展。因此，这项研究有可能 阐明eNAMPT在促进EVS的细胞靶向和EVS的疗效方面的新作用 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的电动汽车 作为一种可行的生物疗法来治疗衰老和阿尔茨海默病的认知衰退。这项奖学金提案整合了 NAD+生物学和老龄化专业知识赞助商Shin Imai博士接受了全面的行为培训 神经科学、AD模型、高级突触分析、电动汽车工程和实验室指导以及 领导力。这一培训将进一步得到智力刺激和高度协作的支持 圣路易斯华盛顿大学的环境。因此，这一奖学金将成为 申请者的职业生涯是未来学术实验室的领导者。

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.