【摘要】 目的:探讨海马外泌体中miR-219a-1-3p对神经干细胞(neural stem cells, NSCs)向神经元分化的影响。方法:切割穹窿海马伞,构建去神经支配海马大鼠模型,分离提取切割组和正常组海马外泌体,与SD大鼠海马NSCs共培养,利用实时定量聚合酶链式反应(real-time polymerase chain reaction, Real-time PCR)、Western Blot和免疫荧光技术检测外泌体在NSCs向神经元分化中的作用;通过RNA-seq方法检测外泌体中表达变化的miRNA,利用Real-time PCR对差异表达的miR-219a-1-3p进行验证,并检测其在成年SD大鼠各组织以及NSCs、星形胶质细胞和神经元中的表达水平。利用miR-219a-1-3p mimic转染NSCs,Real-time PCR、Western Blot和细胞免疫荧光检测miR-219a-1-3p对NSCs向神经元分化的影响。结果:切割组海马组织外泌体可促进NSCs向神经元分化;海马组织外泌体中差异表达的miR-219a-1-3p与RNA-seq结果一致;miR-219a-1-3p在端脑和海马中呈现优势表达;miR-219a-1-3p在NSCs中表达较高,而在星形胶质细胞中表达较低;过表达miR-219a-1-3p可抑制NSCs向神经元分化。结论:海马外泌体促进NSCs向神经元分化可能与miR-219a-1-3p下调有关。更多还原

【Abstract】 Objective: To explore the effect of miR-219 a-1-3 p in the hippocampus exosomes on the differentiation of neural stem cells(NSCs) into neurons. Methods: After fimbria-fornix transection, denervated and untreated hippocampal exosomes were separated and extracted. Then the rat hippocampal exosomes were co-cultured with SD rat hippocampal NSCs. The effect of hippocampal exosomes on the differentiation of NSCs into neurons was detected by real-time polymerase chain reaction(Realtime PCR), Western Blot and immunofluorescence assay. Utilizing high-throughput RNA sequencing technology, we identified a differentially expressed miR-219 a-1-3 p, and further detected the expression of miR-219 a-1-3 p in various tissues, as well as NSCs, astrocytes and neurons of adult SD rats. After transfected with miR-219 a-1-3 p mimic, the effect of miR-219 a-1-3 p on the differentiation of NSCs into neurons was detected by Real-time PCR, Western Blot and immunofluorescence assay.Results: Exosomes of denervated hippocampus promoted the differentiation of NSCs into neurons. Differentially expressed miR-219 a-1-3 p was consistent with the trends observed using RNA sequencing. miR-219 a-1-3 p showed dominant expression in telencephalon and hippocampus, and its expression was higher in NSCs and lower in astrocytes. Overexpression of miR-219 a-1-3 p inhibited NSCs differentiation into neurons. Conclusion: The promotion of exosomes of denervated hippocampus to the differentiation of NSCs into neurons may be related to the down-regulation of miR-219 a-1-3 p.更多还原