七氟烷麻醉大鼠海马组织特异microRNA的差异表达及靶标预测

Differential expression and target prediction of microRNA in hippocampus in Sevoflurane-anesthetized rats

目的定量检测和分析七氟烷麻醉SD大鼠海马miR-133a、miR-96、miR-183的表达,并结合生物信息学,分析预测差异表达miRNA的调控靶标基因。方法 54只SD雄性大鼠以6 L/min通气量不同浓度七氟烷(1.5%、2.5%、3.5%)麻醉4 h,苏醒后不同时间(0 h、4 h、2 d和10 d,每组6只)提取大鼠海马miRNA,并反转录为cDNA,用RT-qPCR方法对miRNA进行定量分析。使用3种生物信息学预测软件对miR-133a、miR-96、miR-183进行靶标分析。结果 miR-133a、miR-96、miR-183在麻醉后大鼠海马中均有表达,但表达差异不一致,与未麻醉的大鼠相比,miR-133a差异最显著,麻醉后0 h、4 h、2 d和10 d分别下调2、1、2.5倍和1倍。信息学分析表明,mirSV score在1.0以上得分有12个靶标基因,其中3个靶基因与记忆障碍有关,分别为溶质载体家庭6成员1(Slc6a1)、腺苷酸环化酶(Adcyap1)和蛋白磷酸酶催化亚基(Ppp2c),它们可能与七氟烷麻醉导致的记忆减退相关。结论 miR-133a可能参与七氟烷致学习记忆下降相关,它可能通过多个基因参与对麻醉后记忆障碍的调控。

Objective To quantitatively analyze the differential expression of miR-133a,miR-96,miR-183 in hippocampus of Sevoflurane-anesthetized rats,combined with the bioinformatics predicte to the potential targets of the differential expressed miRNAs. Methods 54 male SD-rats were anesthezied for 4 h by the use of 6 L/min ventilatory capacity different concentrations of Sevoflurane including 1.5%,2.5% and 3.5%. Rats hippocampus was harvested after different aweaking time（0 h,4 h,2 d and 10 d）. miRNAs were extracted from hippocampus and reverse transcript into cDNA. Then,RT-qPCR was used to detect and analyze the expression of these miRNAs. Three bioformatics softwares including Target,PicTar and Micro were utilized to predict the miRNA target. Results Three miRNA including miR-133a,miR-9 and miR-183 were all expressed in hippocampus of Sevoflurane-anesthetized rats. However,their differential values were inconsistent. Of three miNRAs from Sevoflurane-anesthetized rats,miR-133a was most differentially expressed compared with unanesthetized-rats,and had 2 fold differential down-regulation for 0 h aweaking,1 fold for 4 h aweaking,2.5 fold for 2 d aweaking and 1 fold for 10 d after anesthetization,respectively. The analysis of bioinformatics indicated there were 12 targets for miR-133a. Among these targets,Slc6a1,Adcyap1 and Ppp2c were possibly associated with memory impairment induced by Sevoflurane. Conslusion miR-133a may be implicated in memory impairment,which may be regulated by several genes together.