CircRNA-ITCH/microRNA-214/caspase1轴介导小胶质细胞焦亡参与脊髓损伤的机制研究

Neuroinflammation is an important factor leading to secondary spinal cord injury, and pyroptosis of microglia is an important part of neuroinflammation after spinal cord injury. Previous studies reported that pyroptosis of microglia occurs after spinal cord injury. Howerver, the mechanism remains undefined. In our preliminary studies, we have obtained the exciting results showing that microglia cells in the injured area occured pyroptosis after spinal cord injury. Both animal and cell experiments confirmed that the expression of circRNA-ITCH/ microRNA-214/caspase1 axis changed significantly after spinal cord injury. We postulate that circRNA-ITCH/microRNA-214/caspase1 axis mediates pyroptosis of microglia cells, participates in neuroinflammation and regulates the pathophysiological process of spinal cord injury. In the current proposal, we will further explore the underlying mechanisms in both in vitro and in vivo models of spinal cord injury. The overall hypothesis is that circRNA-ITCH promotes the expression of caspase1 by acting as a competitive sponge for microRNA-214 and mediates pyroptosis of microglia cells in the pathophysiological process of spinal cord injury. A combination of state-of-the-art approaches including situ hybridization, gene knockout and immunohistochemistry method will be used to test this hypothesis. This project will analyze the regulatory mechanism of neuroinflammation after spinal cord injury, which can provide experimental basis for the treatment of clinical spinal cord injury.

神经炎症是导致继发性脊髓损伤的重要因素，小胶质细胞焦亡是介导脊髓损伤后神经炎症的重要部分，研究显示脊髓损伤后小胶质细胞发生焦亡，但机制不清。我们前期工作发现，脊髓损伤后，损伤处小胶质细胞焦亡，而且动物实验和细胞实验均证实脊髓损伤后circRNA-ITCH/microRNA-214/ caspase1轴的表达发生了明显的变化。由此推测，脊髓损伤后circRNA-ITCH/microRNA-214/caspase1轴通过介导小胶质细胞焦亡参与神经炎症，调控脊髓损伤的病理生理过程。本研究基于动物和细胞模型，采用荧光原位杂交、基因敲除、免疫组化等技术，证明circRNA-ITCH通过作为microRNA-214的竞争性海绵促进caspase1表达，介导小胶质细胞焦亡参与脊髓损伤的病理生理过程。本研究分析脊髓损伤后神经炎症的调节机制，可为临床上脊髓损伤的治疗提供实验依据。

神经炎症是导致继发性脊髓损伤的重要因素，小胶质细胞焦亡是介导脊髓损伤后神经炎症的重要部分，研究显示脊髓损伤后小胶质细胞发生焦亡，但机制不清。本项目我们从两个方面进行相关的研究：(1) 基于动物和细胞模型，研究脊髓损伤后，小胶质细胞焦亡情况和circRNA0000381表达情况；(2)研究脊髓损伤后circRNA0000381和小胶质细胞焦亡之间的关系，进一步明确circRNA0000381在脊髓损伤中的作用。研究结果显示：1)在动物模型和细胞模型中均证实小胶质细胞中相关焦亡基因显著上调，同时circRNA0000381的表达也显著上调。2)细胞实验证实抑制circ0000381的表达后，小胶质细胞相关焦亡基因表达显著下调。说明circ0000381对脊髓损伤后小胶质细胞的焦亡有调控作用。此外，课题进行过程中也取得以下新的研究发现：1)发现利鲁唑具有神经保护作用，能够促进脊髓损伤后神经功能恢复，诱导小胶质细胞由促炎性向抗炎性表型极化，同时利鲁唑促进了NF200和MBP蛋白的表达，说明利鲁唑有可能促进轴突生长，并防止脱髓鞘的发生。2)揭示了利鲁唑通过对GSK-3β/CRMP-2信号通路的调控，促进大鼠脊髓损伤后神经轴突生长，加强脊髓损伤后的神经保护作用。3)阐明了利鲁唑治疗能够促进大鼠长期神经功能恢复，利鲁唑治疗的大鼠，GAP43蛋白表达增加，进行电生理实验发现N1峰潜伏期缩短，N1-p1振幅增大。说明脊髓损伤后早期应用利鲁唑，对长期功能恢复至关重要，因此这可能是关键治疗方法中有前景的候选治疗方法。总之，本项目的研究结果将为脊髓损伤后神经功能恢复治疗提供新思路。