NEDL1动物模型的神经生物学功能及机制的在体研究

The comorbidity is common in the nervous system diseases in children which tells these nervous system diseases have the same pathogenic pathway. Neural precursor cell-expressed developmentally down-regulated gene 4 (NEDD4)-like ubiquitin ligase 1 (NEDL1,also known as HECW1)expresses in the nervous system preferentially. In our previous study, we have found NEDL1 can inhibit mTOR signal pathway; in the zebra fish with NEDL1 knockdown, brain cells happened necrosis which can be inverted by rescue experiment. So, it is can be concluded that NEDL1 is the necessary protein for the occurrence and development of center nervous system. In this research, we will investigate the changes of molecule and biochemistry information transfer in the neural circuit of related brain region by a combination of in vitro and in vivo approaches including cell biology, electroneurophysiology and rescue experiment based on the NEDL1 knockout/knockdown animal and mTOR signal pathway, to confirm what kind of role that NEDL1 plays in the formation and disfunction of neurons and synapses; meanwhile, we will deliberate the contribution and mechanism of the related gene and mutant in the formation and function of neuron circuit by neuroethologic and electrophysiologic methods and analysis of molecule, synapse, cell and circuit based on the autism spectrum disorder and epilepsy mice model with NEDL1 knockout. This study will help us figure out the potential pathogenesy in autism spectrum disorder and epilepsy, and provide new research directions, logic proofs and new targets in searching drug treatment of nervous system diseases in children.

儿童常见的神经系统疾病常有共患现象，提示它们存在共同致病通路。泛素连接酶NEDL1 mRNA优先表达于神经组织，我们研究证明NEDL1(又称HECW1)能抑制mTOR通路的激活，NEDL1敲低的斑马鱼显示脑组织细胞特异性死亡，予拯救实验后，其导致的脑细胞死亡表型消失，提示NEDL1是脑神经系统发生所必须蛋白。本课题将在前期研究基础上，利用NEDL1敲除动物模型，以mTOR通路为切入点，采用细胞生物学、神经生理学等方法，检测NEDL1敲除动物有关脑区神经环路中的分子及生化信息传递变化，证实NEDL1在神经元及突触形态和功能异常中的作用；通过建立NEDL1敲除鼠的癫痫及孤独症模型, 结合神经行为学及电生理研究，对其分子、突触、细胞、环路各层次进行相关分析，揭示有关基因及突变体在神经元发育环路的形成和功能中的作用及机理，为临床上常见儿童神经系统疾病的治疗提供新的研究方向、理论依据以及药物治疗靶点

NEDL1主要在脑中高表达，并且NEDL1基因敲低的斑马鱼脑组织细胞特异性死亡，给予拯救实验后，导致的脑细胞死亡表型消失，提示NEDL1在神经系统发育中起重要作用。本课题从两方面NEDL1的的脑功能: (1)利用NEDL1基因敲除小鼠模型，从组织病理、行为学、电生理、代谢等多个方面分析NEDL1的作用，(2) 建立NEDL1基因敲除鼠的癫痫模型，研究NEDL1与癫痫的关系。. 1.NEDL1基因敲除鼠和野生鼠比较：成年前NEDL1基因敲除鼠脑中胶质细胞数量较野生型鼠明显增加。42天的NEDL1基因敲除鼠表现出明显的刻板行为，而社会交往能力、重复行为、学习能力及焦虑状态方面无明显差异。代谢组及转录组发现NEDL1功能与氨基酸、蛋白合成，肿瘤形成、神经系统兴奋性及脑发育相关。. 2.NEDL1基因敲除鼠癫痫模型：成功构建NEDL1基因敲除鼠匹鲁卡品癫痫模型，NEDL1基因敲除组小鼠癫痫发作严重程度及死亡率低于野生鼠组，并且在电生理、组织病理以及行为学方面KO组较WT组改变轻微，NEDL1基因敲除可以降低癫痫的严重程度。. 3.NEDL1与mTOR通路：NEDL1基因敲除鼠癫痫发作严重程度轻微可能与维持mTOR通路稳定相关。. 共发表论文29篇，在国内杂志发表19篇，国外杂志发表10篇，培养了2名博士，3名硕士研究生。