Restorative gene therapy on hippocampal spino-dendritic abnormalities: a focus on Alzheimer’s disease

摘要/摘要 目前还没有有效的治疗方法来预防、延迟或逆转记忆缺陷和 阿尔茨海默病相关的神经病理改变--特别是树突损伤和突触丢失 疾病(AD)。如果不尽快得到有效的治疗，数以百万计的老年人将继续遭受 这种毁灭性的疾病。因此，迫切需要研究涉及到的新分子/机制 为了挽救认知功能而维持和再生树突及其棘突。 长期目标是开发一种有效的治疗方法来预防或阻止AD。这样做的总体目标是 应用是确定最近发现的一种磷蛋白CRMP3(折叠反应)的潜力 中介蛋白3)，以改善两种不同小鼠的树突异常和/或挽救记忆缺陷 具有类似AD特征的模型。3xTg-AD小鼠是最常用的AD小鼠模型之一。 它们含有3种突变(APP、PSEN1和MAPT)，导致斑块和缠结的积累， 树突和突触结构异常，以及严重的认知障碍。CRMP3-/-小鼠展示 营养不良的海马树突和严重损害的突触可塑性(LTP)。重要的是，过度表达 在体外，海马神经元中的CRMP3蛋白显著促进树突的形成。中环 假设在体内过度表达CRMP3将防止进一步的树突状细胞退化和/或唤起 在其他退化的海马神经元中，树突的再生和棘突的重塑，从而保存 或者恢复记忆功能。这一假设是在强大的初步数据的基础上提出的 在申请者的实验室生产的。这一假设将通过追求两个具体目标来检验：1) 确定体内CRMP3过表达在多大程度上增强了海马树突状细胞的复杂性 以及CRMP3-/-和3xTg-AD小鼠的脊柱结构；以及2)确定体内过度表达的程度 CRMP3基因缺失影响CRMP3-/-和3xTg-AD小鼠的海马区依赖记忆功能。在第一个目标下， 将使用腺相关病毒载体血清型9(AAV9)基因治疗CRMP3在 两种类AD小鼠模型的海马区及其对AD相关棘树突异常的影响 将会被检查。在第二个目标下，CRMP3基因治疗可以在多大程度上改善上下文 空间记忆功能将使用两种行为范式(情景恐惧条件反射和 Morris水迷宫)。这种方法是创新的，因为CRMP3是一种鲜为人知的蛋白质，具有巨大的 修复和再生营养不良和受损的棘突和树突的潜力。拟议的研究是 这一发现意义重大，因为它有望提高人们对一种蛋白质的认识，这种蛋白质具有强大的减速、停滞、 甚至逆转可能是深度痴呆症先兆的早期AD过程，从而显著提高质量 生活的一部分。最终，这些发现有可能进入临床试验，作为AD的治疗方法。

SUMMARY/ABSTRACT There are currently no effective treatments to prevent, delay, or reverse the memory deficits and neuropathological alterations—specifically dendritic damage and synaptic loss—associated with Alzheimer’s disease (AD). Without an effective treatment soon, many more millions of older adults will continue to suffer from this devastating disease. Thus, there is a critical need to investigate novel molecules/mechanisms involved in the maintenance and regeneration of dendrites and their spines for the purpose of rescuing cognitive functions. The long-term goal is to develop an effective treatment to prevent or halt AD. The overall objective of this application is to determine the potential of a recently identified phosphoprotein, CRMP3 (collapsing response mediator protein 3), to ameliorate dendritic abnormalities and/or rescue memory deficits in two distinct mouse models with AD-like characteristics. 3xTg-AD mice are one of the most commonly used mouse models of AD. They harbor 3 mutations (APP, PSEN1, and MAPT) that result in the accumulation of plaques and tangles, abnormalities in dendritic and synaptic structures, and significant cognitive impairments. CRMP3-/- mice exhibit dystrophic hippocampal dendrites and profoundly impaired synaptic plasticity (LTP). Importantly, overexpressing the CRMP3 protein in hippocampal neurons, in vitro, dramatically enhances dendritic formation. The central hypothesis is that over-expressing CRMP3, in vivo, will prevent further dendritic degeneration and/or evoke dendritic regrowth and remodeling of spines in otherwise degenerating hippocampal neurons, thus preserving or rescuing memory function. This hypothesis has been formulated on the basis of strong preliminary data produced in the applicants’ laboratories. This hypothesis will be tested by pursuing two specific aims: 1) Determine the extent to which in vivo over-expression of CRMP3 enhances hippocampal dendritic complexity and spine structure in CRMP3-/- and 3xTg-AD mice; and 2) Determine the extent to which in vivo over-expression of CRMP3 affects hippocampal-dependent memory function in CRMP3-/- and 3xTg-AD mice. Under the first aim, adeno-associated viral vector serotype 9 (AAV9) gene therapy will be used to overexpress CRMP3 in the hippocampus of these two AD-like mouse models and its effects on AD-associated spino-dendritic abnormalities will be examined. Under the second aim, the extent to which CRMP3 gene therapy can ameliorate contextual and spatial memory function will be measured using two behavioral paradigms (contextual fear conditioning and Morris water maze). The approach is innovative because CRMP3 is a little-known protein with tremendous potential to repair and regenerate dystrophic and damaged spines and dendrites. The proposed research is significant because it is expected to advance knowledge about a protein with the robust capacity to slow, halt, or even reverse early AD processes that may be precursors to deep dementia, thus profoundly improving quality of life. Ultimately, such findings have the potential to move to clinical trials as a treatment for AD.