外源性miRNA-210对大鼠急性脊髓损伤后血管生成的作用及其机制研究

Acute spinal cord injury (SCI) is a serious neurotrauma with high probability of limbs paralysis, in which the secondary injury is aggravated by posttraumatic ischemia. MiR-210 is commonly induced in hypoxia condition to promote angiogenesis as an upstream controlling gene. Previous study indicated the miR-210 differential expression in the rat model of SCI, which implied its regulatory potential in the posttraumatic angiogenesis of the SCI model. However, further research is needed. In this study, the exogenous miR-210 will be intended to treat rat with acute spinal cord injury by the way of miR-210 continuous intrathecal injection within the ALZET osmotic pump, and the recovery of the neural function will be examined. In this proposed study, an new appraisal method based on the 3D vascular morphometry using the high-resolution synchrotron radiation phase contrast imaging (PCI) technique will be applied to the rat SCI model for the first time, to evaluate the pro-angiogenic effects after the miR-210 administration. Besides, the proliferation, apoptosis and survival of the spinal endothelial cells will also be studied using the typical molecular biotechnology followed by quantitative analysis. Lastly, the HIF-1 α/VEGF and Dll4/Notch pro-angiogenec pathway activation level, as well as the downstream target gene expression, are to be evaluated to analyze the miR-210 upstream regulatory role, and proceed to reveal the relationship between the miR-210 treatment and the posttraumatic angiogenesis after rat SCI, all of which shall explore the relative vascular protective mechanism by the MiR-210 treatment in the rat SCI model.

急性脊髓损伤（SCI）是一类严重的运动系统创伤，创伤后血供障碍是SCI继发性加重的重要因素。miR-210是低氧状态下促血管生成的重要调控基因，我们的前期研究显示其在急性SCI前后有显著差异表达，提示其在SCI后血管生成过程中具有重要的调控功能，但具体作用方式及机制有待进一步研究。本项目拟采用外源性miR-210治疗大鼠急性SCI，首次利用高分辨率同步辐射相衬成像技术在无需血管造影的条件下充分显示脊髓三维微血管网络，以此评估miR-210对SCI后血管生成的形态计量学影响；同时检测miR-210对脊髓内皮细胞的增殖和凋亡、下游靶基因表达以及HIF-1α/VEGF和Dll4/Notch促血管生成通路活化的影响。综上研究以明确miR-210在急性SCI后血管生成过程中的作用及相关机制，进而从结构-功能角度阐明miR-210治疗与脊髓功能修复的关系，为其应用于SCI治疗提供临床前科学依据。

急性脊髓损伤（ASCI）高致瘫性、神经功能恢复难度大，仍然是目前神经创伤领域的研究热点和难点。本课题以大鼠Allen’s打击ASCI模型为基础，系列的研究了miR-210在改善脊髓损伤后缺血、减少神经细胞凋亡、减轻神经炎症，促进神经功能恢复方面的积极作用及相关机制。主要研究内容及结果：①miR-210在大鼠ASCI后1、3、7和14天呈时间依赖性差异表达下调；②持续硬脊膜鞘内泵入给药方式可有效提高伤段组织内miR-210的表达水平，促进残存神经细胞存活，改善后肢运动功能恢复；③miR-210有效改善大鼠ASCI后伤段脊髓血管的3D形态构筑，提高灌注容量，改善血管分枝、长度和密度等，同时增加大鼠ASCI后伤段脊髓内皮细胞的增殖，促进创伤后血管再生；④miR-210在ASCI后下调了Ephrin A3和PTP-1B mRNA的表达水平，上调VEGF表达水平；⑤miR-210有效减少伤段组织细胞凋亡，减轻损伤早期神经炎症反应。本研究证实：外援性局部泵入miR-210以减轻大鼠ASCI后脊髓组织中的细胞凋亡和神经炎症，增加伤段脊髓血管再生对于改善局部缺血、增加血供灌注，以及促进组织细胞存活、提高后肢运动功能恢复具有积极有效的作用，该课题为外源性miR-210干预治疗ASCI及相关缺血性脊髓疾病治疗的临床前疗效评估提供了实验科学依据。在该课题经费支持下，共培养或协助培养博士毕业生3名，硕士毕业生1名，中青年骨干医师1名，衍生课题1项。课题负责人及团队成员共发表中、英文论文8篇，其中SCI收录6篇，中文统计源期刊 2篇，参与相关的国内、外重要学术会议5次。