New Roles for sAPP in Neuroprotection and Neurogenesis

描述（由申请人提供）：淀粉样前体蛋白（APR）及其可溶性分泌片段（sAPPa）（一种正常的α-分泌酶裂解产物）的神经保护作用已在培养系统中针对广谱神经元损伤得到证实。在某些过表达APP的转基因（Tg）小鼠模型中观察到的几种保护作用也被推断为sAPPa。这些行动的机制，但是，很少研究。因此，本申请的目的是研究sAPPa功能的分子机制，并评估其对减轻小鼠AD模型中的神经病理学的有益作用。在初步研究中，我们证明了缺乏APP的小鼠或细胞与增加的应激激酶CDK 5活性和升高的tau磷酸化相关，这可以通过外源性sAPPa来恢复。sAPPa在NMDA诱导的神经元死亡中具有强的抗凋亡活性，其通过涉及Bcl-2的途径与IGF协同作用。重要的是，我们在年轻的成年APP-/-小鼠中发现了神经源性缺陷，并在各种细胞/组织培养物中鉴定了sAPPa的神经营养功能，其被EGF协同作用。慢性输注 结果表明，脑室下区的神经元祖细胞含有sAPPa在体内的主要结合位点。sAPPa优先结合到细胞表面上的脂质富集的微区。综上所述，我们假设sAPPa在预防神经元变性和死亡方面起着广泛的神经保护作用，而更重要的是Quinalv在产前和产后阶段的神经发生中起着至关重要的作用。以适当的剂量和时间在CNS中进行sAPPa治疗将改善在小鼠AD模型中发展的神经病理学。因此，我们在目的1中提出评估sAPPa在Tau Tg小鼠模型中减少神经病理学的作用。我们将使用tau突变体（R406 W）转基因小鼠测试sAPPa治疗是否可以减少tau磷酸化和NFT形成，提供兴奋保护，并防止记忆丧失。目的2将评估sAPPa在APP+和APP/APLP 2双敲除小鼠中神经发生中的潜在作用。我们将检查这些小鼠中的神经发生水平是否与它们的死亡率相关，以及sAPPa的慢性输注是否可以刺激这些小鼠中神经元祖细胞/干细胞的增殖并挽救神经发生损伤。目的3是了解sAPPa生物学功能的分子和细胞机制，并鉴定sAPPa相关的膜蛋白。分子和细胞生物学方法，包括膜蛋白与结合的生物素化sAPPa的交联、脂筏分离，结合蛋白质组学技术，将用于分离和鉴定sAPPa的推定膜受体。我们的研究将揭示APP/sAPPa的新的生理功能以及潜在的分子机制，并为未来sAPPa的潜在治疗窗口提供帮助。

DESCRIPTION (provided by applicant): The neuroprotective roles for amyloid precursor protein (APR) and its soluble secreted fragment (sAPPa), a normal a-secretase cleavage product, have been demonstrated in culture systems against a broad spectrum of neuronal insults. Several protective roles observed in certain transgenic (Tg) mouse models overexpressing APP have also been inferred to sAPPa. The mechanisms underlying these actions, are however, scarcely studied. The objectives of this application are thus to investigate the molecular mechanisms of sAPPa's functions and to evaluate its beneficial effects on alleviating neuropathologies in mouse AD models. In preliminary studies, we demonstrated that mice or cells lacking APP are associated with increased stress kinase CDK5 activity and elevated tau phosphorylation, which can be restored by exogenous sAPPa. sAPPa possesses strong anti-apoptotic activity in NMDA-induced neuronal death which synergizes with IGF via a pathway involving Bcl-2. Importantly, we found neurogenic defect in young adult APP"'" mice and identified a neurotrophic function of sAPPa in various cell/tissue cultures, which is synergized by EGF. Chronic infusion of into adult mice reveals that the neuronal progenitor cells in the subventricular zone contain major binding sites for sAPPa in vivo. sAPPa preferentially binds to the lipid-enriched microdomains on the cell surface. Taken together, we hypothesize that sAPPa plays broad neuroprotective roles in preventing neuronal degeneration and death, and more intriQuinalv, a crucial role in neurogenesis at both the prenatal and postnatal stages. sAPPa treatment in CNS at a proper dosage and timing will improve neuropathologies developed in mouse AD models. We therefore propose in Aim 1 to assess the roles of sAPPa in reducing neuropathologies in Tau Tg mouse model. We will test whether sAPPa treatment can reduce tau-phosphorylation and NFT formation, provide excitoprotection, and prevent memory loss using tau mutant (R406W) transgenic mice. Aim 2 will assess the potential oles of sAPPa in neurogenesis in APP"'* and APP/APLP2 double knockout mice. We will examine whether neurogenesis levels in these mice associate with their mortality, and whether chronic i9nfusion of sAPPa can stimulate proliferation of neuronal progenitor/stem cells in these mice and rescue neurogenesis impairment. Aim 3 is to understand the molecular and cellular mechanism underlying the biological functions of sAPPa and to identify sAPPa-associated membrane protein(s). Molecular and cell biological approaches including cross linking of membrane proteins with bound biotinylated sAPPa, lipid raft isolation, in combination with proteomic technologies, will be used to isolate and identify putative membrane-receptor(s) for sAPPa. Our study will reveal novel physiological functions of APP/sAPPa as well as the underlying molecular mechanisms, and should shed light on potential therapeutic windows for sAPPa in the future.