miR-30a在颅脑创伤后的表达及其在继发性脑损伤中的作用

The secondary brain injury following traumatic brain injury (TBI) results in tremendous neuronal death and apoptosis. The voltage-gated sodium channels (VGSCs) take part in the excitotoxic effect, which is one of the most important mechanisms of the secondary brain injury. We previously reported that the expression of Nav1.3, one of the VGSCs which mainly located in central nervous system (CNS), upregulated significantly at the early stage post-TBI, and the alterations in expression of Nav1.3 were correlated with severity of TBI. Blockage of the upregulation of Nav1.3 improved outcomes after severe TBI. We now has certificated that miR-30a was the miRNA which inhibiting mRNA translation or promoting mRNA degradation of Nav1.3. So we assume that miR-30a takes part in the excitotoxic process post-TBI and regulation of the expression of miR-30a may improve the outcomes of secondary brain injury. In this project, we try to validate this hypothesis in three parts: 1.investigate the expression of miR-30a in the rats suffered from brain fluid percussion injury and make cofirm that miR-30a takes part in the process of secondary brain injury. 2.regulate the expression of Nav1.3 by intracerebroventricular injection of miR-30a overrepresentation virus in order to observe the neuroprotective effect in the secondary brain injury. 3.explore the neurons physical status (especially the influence of sodium current bursting cluster in neurons and the intra-neuronal sodium concentration), after the alteration of expression of miR-30a in the cell stretch injury model. From all the results above, we can gain a better understand of the role of miR-30a in the secondary brain injury following TBI, and find out an effective and characteristic neuroprotective method post-TBI.

颅脑创伤（TBI）后的继发性脑损伤导致大量神经元死亡和凋亡，钠通道参与的兴奋性毒性作用是继发性脑损伤的最重要机制之一。前期研究发现TBI后钠通道α亚单位Nav1.3表达上调，其上调水平与TBI程度呈相关性，阻断其表达上调可改善TBI预后，同时证实miR-30a抑制Nav1.3表达。我们推测miR-30a参与TBI后兴奋性毒性过程，调控其表达可改善继发性脑损伤。本项目拟从三个方面验证该假说：1.通过大鼠脑液压伤模型研究TBI后miR-30a表达的时空变化确定其参与继发性脑损伤；2.通过脑室内注入miR-30a过表达病毒调控Nav1.3表达水平，确定miR-30a表达变化对继发性脑损伤的神经保护作用；3.通过细胞牵张性损伤模型确定miR-30a表达改变对神经元钠电流簇状发放特性和细胞内钠离子浓度影响。上述研究结果将初步阐明miR-30a参与继发性脑损伤的分子机制，提供有效且特异的神经保护方法。

颅脑创伤（TBI）是社会引起高死残率的严重疾病之一，既往研究发现TBI导致miRNA水平变化，并可作为TBI的可能生物标志之一。前期工作发现Nav1.3是TBI后继发性脑损伤重要的基因，并找到miR-30a是调控Nav1.3的上游miRNA。本项目主要是通过研究大鼠颅脑创伤后miR-30a表达情况，检测了其在不同损伤程度下血清、脑脊液和伤侧海马中的表达水平，并将表达水平与神经功能情况相联系，评估两者的关系，但未能有效发现miR-30a变化水平的规律性。进一步证实在TBI急性期体内阻滞Nav1.3的表达上调可以减轻因TBI引发的继发性脑损伤，实验中通过向脑室内注入针对Nav1.3反义寡核苷酸，使海马组织中Nav1.3表达下降，并使脑细胞水肿减轻和降低凋亡细胞数，同时学习记忆功能有改善。课题研究中扩大对大鼠海马miRNA表达水平进行检测，共发现82种miRNA表达水平在TBI后发生改变，其中17种miRNA表达水平上调，而65种miRNA表达水平下降。进一步对82种miRNA针对的特异生理过程和神经信号通路的基因网络进行分析，并对既往研究结果感兴趣的Nav1.3起作用的miRNA进一步分析，发现miR-3562和miR-361-3p位于该网络中其核心作用，相应的结果已整理准备投稿。后期将对此两种miRNA再进一步研究，对TBI的治疗可能作为诊断性生物标志物。