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目的基于微小核糖核酸(miRNA)-信使核糖核酸(mRNA)调控网络探讨重复经颅磁刺激(rTMS)促进缺血性脑卒中(IS)神经修复的潜在分子机制。方法从基因表达综合数据库(GEO)下载rTMS干预7 d后的IS大鼠皮质miRNA表达谱。使用R软件分析差异miRNAs
通过在线数据库预测其对应的下游mRNAs
构建miRNA-mRNA调控网络
筛选关键miRNAs。对靶基因进行功能富集分析
构建靶基因所编码的蛋白质相互作用网络
识别网络中的核心mRNAs。选取无特定病原体动物(SPF)级雄性Sprague-Dawley大鼠24只
按随机数字表法随机分为假手术组、模型组和磁刺激组
每组8只大鼠。模型组和磁刺激组制备IS大鼠模型
仅磁刺激组接受连续7 d的rTMS干预
其余2组不进行干预。采用改良神经功能评分(mNSS)于造模前和造模成功1 d、3 d、7 d后评价3组大鼠的神经功能缺损程度;于造模成功8 d后深度麻醉大鼠
取脑组织行苏木精-伊红和尼氏染色观察组织缺失情况和神经元形态
利用实时荧光定量聚合酶链反应验证差异基因在3组大鼠缺血侧皮质中的表达趋势。结果共筛选出167个差异miRNAs和预测到25个mRNAs。富集分析表明
这些基因与细胞迁移的正向调节、神经营养因子受体信号通路的正向调节等生物学过程有关
涉及腺苷酸激活蛋白激酶、神经营养因子通路、大鼠肉瘤等信号通路。通过构建miRNA-mRNA调控网络
识别出miR-206-3p、miR-378a-3p、miR-107-3p、miR-92a-3p和miR-29b-3p共5个关键miRNAs;通过蛋白互作网络分析筛选出4个核心mRNAs
分别为整合素亚基β1、水通道蛋白4、脑源性神经生长因子(BDNF)、溶质载体家族2成员4。造模成功7 d后
磁刺激组的mNSS评分显著低于模型组同时间点(P<0.05)。与模型组相比
磁刺激大鼠右侧皮质的miR-206-3p表达明显降低(P<0.05)
BDNF表达则显著增高(P<0.05)。结论 miR-206-3p-BDNF调控网络和相关作用途径在rTMS促进IS大鼠神经修复过程中发挥重要的作用。
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