miR-194对Neuritin的靶向调控及其在阿尔兹海默病中的作用及机制研究

The study showed that the expression of Neuritin (NRN1) was decreased in nerve injury (including AD), and NRN1 protein treatment have obvious repairment. However, the molecular mechanism about NRN1 reduction in AD was unclear. We hypothesized that some miRNAs in AD maybe downregulated NRN1 expression to affect the generation and development of AD. We have screened candidate miRNAs that maybe targeted NRN1 in AD, and preliminary results showed the correlation between miRNAs and NRN1 in AD and the inhibiton of miR-194 on NRN1. This study will make clear the association of the level of miR-194 or NRN1 and the ability of learning and memory in AD transgenic mice. And then we will investigate the role and mechanism of miR-194 in the generation and development of AD by up-regulation of miR-194 and down-regulation of NRN1. Finally, we will treated AD mice or AD cell model with miR - 194 specific inhibitors and NRN1 recombinant protein to observe the improvement in AD pathological feature, the synaptic plasticity and mice behavior. We will give an aggregate analysis about the feasibility of miR - 194 and NRN1 as the diagnostic and therapeutic targets in AD, which will provide the foundation for further clinical application research.

研究发现在神经损伤时（包括AD）Neuritin(NRN1)表达降低，而给予NRN1蛋白有明显修复作用。然而，关于AD中NRN1表达降低的分子机制尚不明确。我们推测：AD中某些miRNA可能下调NRN1表达而影响AD的发生发展。我们前期筛选了AD中可能靶向NRN1的候选miRNA，基本明确在AD中它们与NRN1的相关性及miR-194对NRN1的抑制作用。本研究拟首先明确miR-194和NRN1的表达与转基因AD鼠学习记忆障碍的相关性；进一步通过上调miR-194和下调NRN1，探讨miR-194在AD发生发展中作用及机制；最后给予miR-194抑制剂及NRN1重组蛋白，观察对AD鼠及细胞的AD病理特征、突触可塑性及行为学的改善作用，明确miR-194抑制剂和NRN1蛋白在AD治疗中的有效性，综合分析miR-194和NRN1作为AD诊断和治疗靶点的可行性，为进一步临床应用研究提供实验依据。

Neuritin在神经发育和可塑性中发挥重要作用。研究发现，阿尔茨海默病(AD)小鼠中Neuritin表达降低，上调Neuritin水平可减轻AD鼠的突触损伤。然而，关于AD中Neuritin的表达降低分子机制，目前尚未明确。本研究首先利用生物信息学手段，预测分析AD中靶向Neuritin的miRNA，初步筛选出候选miRNA。进一步在AD模型鼠中，分别检测了候选miRNA和Neuritin的表达，分析了二者的相关性；确定了目标miRNA（miR-182/188/199a），并明确了目标miRNA对Neuritin的靶向抑制作用。使用脑立体定位注射目标miRNA——miRNA-182/188/199a拮抗剂（AM）干预AD模型鼠，从行为学、突触可塑性和AD病理学变化评价了AM对AD模型鼠的影响，发现AM促进AD鼠突触的传递和突触基础传递效率，促使树突棘发育成熟，改善突触可塑性；同时，能够改善AD鼠海马中AD病理学表征（减少Aβ斑块的形成，降低tau蛋白的磷酸化水平），并提高了AD小鼠的学习记忆能力。在AD细胞模型中，AM促进AD细胞模型中与细胞形态可塑性相关指标的变化，抑制了细胞凋亡；并检测了AD模型鼠发展的不同阶段相关信号通路的变化，以及AD细胞模型给予AM干预后，相关信号通路分子指标的变化，揭示目标miRNA抑制剂通过上调Neuritin、抑制Notch信号通路，进而发挥作用的分子机制。