音猬因子调控线粒体能量代谢在脑梗死后神经可塑性中的作用及机制

The most important point for the treatment of ischemic cerebral infarction is how to promote nerve repair. The basis of nerve repair after ischemic cerebral infarction is neuroplasticity including axonal sprouting and synaptic remodeling. A better understanding of neuroplasticity after ischemic stroke will be crucial to the development of pharmacologic and rehabilitative therapies for ischemic cerebral injury. Recent studies have indicated that a secreted glycoprotein factor-Shh might mediate neuroplasticity after ischemic cerebral infarction. It has been found that mitochondrial energy metabolism plays a vital role in regulating neuroplasticity after ischemic cerebral infarction. Our previous study has showed that Shh promoted neurite outgrowth of cortical neuron under oxidative stress possibly through protecting mitochondrial functions, elevating the cellular ATP levels and ameliorating mitochondrial complex II activities. However, the effect and the potent mechanism of Shh on mitochondrial energy metabolism in neuroplasticity after ischemic cerebral infarction have not yet been reported. Our preliminary experimental results showed that knocking down Shh expression seriously restricted the nerve repair after ischemic cerebral infarction which companied by aggravating mitochondrial damage and inhibiting axonal sprouting. Hereby, we hypothesized that Shh may mediate neuroplasticity after ischemic cerebral infarction by improving mitochondrial energy metabolism. In this project, the primary cortical neurons culture and focal cerebral cortex infarction model were used. The neural anterograde tracer (BDA) staining, molecular biology, gene chip and immunoprecipitation technique were used to assess the effect of Shh on mitochondrial energy metabolism in neuroplasticity after ischemic stroke at the integral, cell, molecular, protein and gene level. These findings may provide new targets and new strategies for the treatment of ischemic cerebral infarction.

脑梗死治疗的难点是如何促进神经修复。神经可塑性是脑梗死后神经修复的基础。研究表明音猬因子（Sonic Hedghog, Shh）可介导脑梗死后神经可塑性。已有研究发现线粒体能量代谢是调节脑梗死后神经可塑性的关键环节。我们前期研究表明Shh可通过改善线粒体能量代谢促进皮层神经元神经突起的生长。但关于Shh对脑梗死后神经可塑性与线粒体能量代谢的作用及机制尚未见报道。在预实验中我们发现敲低Shh表达可通过加重脑梗死后线粒体损伤和抑制轴突再生阻碍神经修复。因此我们推测Shh可通过改善线粒体能量代谢介导脑梗死后神经可塑性。本研究拟建立皮层神经元培养和脑皮层梗死小鼠模型，通过敲低Shh表达，应用神经顺行示踪剂（BDA）染色、分子生物学、基因芯片和免疫共沉淀等技术，从整体、细胞、分子、蛋白和基因水平上验证我们的假说。本项目的实施可为脑梗死的治疗提供新靶点和新策略。

脑梗死治疗的难点是如何促进神经修复。神经可塑性是脑梗死后神经修复的基础。研究表明Sonic Hedghog（Shh）可介导脑梗死后神经可塑性。已有研究发现线粒体能量代谢是调节脑梗死后神经可塑性的关键环节。本课题应用体外细胞皮层神经元细胞实验中发现抑制Shh可抑制神经元可塑性，并发现沉默Shh可使皮层神经元细胞 MMP降低并且ATP生成下降。体内脑梗死动物实验发现：敲低Shh表达可加重脑梗死后线粒体损伤并且可抑制轴突再生，加重脑梗死后神经功能损伤。应用基因芯片筛选出脑梗死小鼠脑皮质组织中差异表达 microRNA 17个，其中表达上调8个，表达下调9个。进一步筛选出microRNA-210，运用q-PCR技术验证其在脑梗死后脑皮质组织中表达上调。Shh可能通过上调microRNA-210逆转能量生成障碍对脑梗死发挥治疗作用。该研究有望为改善脑梗死后神经修复提供一个潜在的关键性靶点，为研发脑梗死治疗的有效药物提供实验依据和线索，为促进脑梗死患者神经功能恢复治疗提供理论依据，具有很大的社会和经济效益及广泛的应用前景。