脑卒中后的神经再生修复是当今神经科学的重大难题，通过干预神经干细胞（NSCs）促进其增殖分化重建神经通路，具有良好应用前景。本项目前期已经证明，肾脑复元合剂能发挥显著的脑缺血后神经保护效应，并促进NSCs的增殖和神经分化；为进一步明确其干预NSCs的分子机制，我们采用高通量基因测序和PCR技术，证实该方药能显著提高海马区miR-25表达；miR-25是促NSCs增殖分化的上游分子，通过生物信息学技术我们发现miR-25可抑制TGF-β1/Smad通路，并很可能进一步靶向下游Hes-1基因干预NSCs增殖分化。因此，我们提出假说：肾脑复元合剂调控“miR-25↑—TGF-β1/Smad↓—Hes-1↓—NSCs增殖分化↑”信号轴，促进脑缺血后神经新生。本项目将采用miRNA转染、慢病毒构建等手段，从干预miRNA信号轴调控NSCs增殖分化的角度，探究肾脑复元合剂对脑缺血损伤的神经保护机制。

Neuroregeneration after stroke is a major problem in neuroscience today. It’s promising to have good application by interfering with neural stem cells (NSCs) to promote their proliferation and differentiation and restore neural network. In the preliminary study of the project, it has been proved that Shennao Fuyuan Mixture have significant neuroprotective effects after cerebral ischemia and promote the migration and neural differentiation of NSCs. Then, we use high-throughput gene sequencing and PCR technology to further study the molecular mechanism of its intervention in NSCs. It was confirmed that the prescription can significantly increase the expression of miR-25 in hippocampus while miR-25 is an upstream molecule that promotes the proliferation and differentiation of NSCs. Using bioinformatics technology, we found that miR-25 can inhibit TGF-β1/Smad pathway, and further affect the proliferation and differentiation of NSCs by targeting Hes-1 gene probably. Therefore, we propose the hypothesis: Shennao Fuyuan Mixture regulates the signal axis of "miR-25↑-TGF-β1/Smad↓-Hes-1↓-NSCs proliferation and differentiation" and promotes neuronal regeneration after cerebral ischemia. This project is based on the perspective of interfering with miRNA signal axis to regulate the proliferation and differentiation of NSCs. In the experiment, miRNA transfection and lentiviral construction will be used to explore the neuroprotective mechanism of Shennao Fuyuan Mixture on cerebral ischemic injury.