锂剂通过miR-30靶向Atg家族基因对脊髓损伤后细胞自噬的机制研究

The neuroprotective effect of lithium on spinal cord injury (SCI) has already been demonstrated, but its mechanism of autophagy is unknown. We found that lithium could protect the SCI rats by up-regulating the autophagy-related genes (Atg), and it could make the expression of miR-30 in SCI rats lower. It is predicted that miR-30 is targeting Atg genes by bioinformatics, suggesting that miR-30 regulates autophagy by targeting the Atg genes to mediate the role of lithium on the repair of SCI. In order to validate this hypothesis, the SD rats and the neuroma SH-SY5Y cell lines are selected as the research objects. The rat and isolated cell models are established by modified Allen method and hydrogen peroxide injury method respectively. The miR-30 and Atg family genes are identified by molecular biology techniques, and the targeting relationship of miR-30 and Atg genes is determined as well. The molecular, cellular, behavioral and imaging methods are used to further investigate the effects of miR-30 and lithium on autophagy in SCI. The aim of this study is to investigate the mechanism of autophagy that miR-30 regulates Atg family genes, and to reveal the new mechanism of repairing SCI through lithium by autophagy, and to provide theoretical basis for the comprehensive treatment of drugs based on lithium.

锂剂对脊髓损伤(SCI)的神经保护作用已被证实，但其细胞自噬作用机制不明。我们研究发现，锂剂可以通过上调细胞自噬相关基因（Atg）对SCI大鼠产生保护作用，并且能使SCI大鼠中miR-30低表达；生物信息学预测miR-30靶向Atg基因家族，提示miR-30靶向Atg基因来调节自噬作用从而介导锂剂对SCI的修复作用。为了验证这一假设，本课题拟以SD大鼠和神经瘤SH-SY5Y细胞株为研究对象，采用改良Allen法和过氧化氢损伤法分别建立大鼠和离体细胞SCI模型，通过分子生物学技术明确锂剂治疗后miR-30和Atg家族基因表达定位，并确定二者的靶向关系，通过分子、细胞、行为学和影像学检测方法进一步对miR-30以及锂剂在SCI修复中自噬作用机制进行研究。本课题旨在认识miR-30调控Atg基因的自噬机制，揭示锂剂通过自噬作用修复SCI的新机理，为SCI后以锂剂为基础的药物综合治疗提供理论依据。

脊髓损伤（spinal cord injury, SCI）是一系列内外因素导致中枢神经损伤的严重致残性疾病。锂剂作为治疗双相情感障碍的主要药物已有100多年的历史，研究证实锂剂对神经元有保护作用，但其具体机制不明。项目负责人在此项目资助下，发现了脊髓损伤后锂剂起到神经保护的相关信号通路，并从凋亡、炎症、自噬、内质网应激、NLRP3炎性小体及铁死亡等多角度，采用动物模型实验和相应的细胞水平实验，全方面阐释锂剂通过miRNA、长链非编码RNA等分子在脊髓损伤中的作用。在脊髓损伤凋亡和炎症方面，揭示了锂剂通过长链非编码RNA BDNF-AS/miR-9-5p组成的调控通路；在脊髓损伤自噬方面，揭示了长链非编码RNA MALAT1/miR-105/ULK1和长链非编码RNA MALAT1/miR-105/Atg7组成的调控通路；在脊髓损伤内质网方面，发现脊髓损伤后可以激活内质网应激，而锂剂可以通过内质网应激来调控细胞自噬和减轻细胞凋亡，这一过程是通过激活IRE1 和 PERK/eIF2α信号通路完成的；在脊髓损伤焦亡过程中，我们发现脊髓损伤后出现新型炎症过程焦亡，激活NLRP3炎症小体，锂剂能够通过Nrf2/HO-1信号通路减轻细胞氧化应激、缓解焦亡反应，从而起到神经保护作用；在脊髓损伤铁死亡方面，揭示了miR-672-3p通过靶向新型铁死亡标志蛋白AIFM2/FSP1调控细胞铁死亡机制。通过深入探讨锂剂在脊髓损伤中的分子机理，能够为锂剂早日应用于脊髓损伤患者提供理论依据。