化学合成小干扰RNA对大鼠脑内AQP4蛋白沉默效果验证与筛选

摘要

目的：针对大鼠脑内AQP4蛋白使用化学合成法制备数条小干扰RNA链（siRNA），对其中两条链进行沉默效果验证，并筛选出效果显著的小干扰RNA链。方法根据大鼠的AQP4mRNA序列选择2个不同的靶点，采用化学合成的方法分别合成2条siRNA，并同时合成阴性对照siRNA。雄性成年Wistar大鼠18只，随机分为注射阴性对照液组、注射725链组和注射1006链组，并分别经侧脑室注射相应药物。6 h后将其全部处死，利用免疫组织化学法和原位杂交法分别检测脑内AQP4蛋白和mRNA的表达水平。结果给药6 h后，注射725链siRNA组大鼠脑内AQP4在蛋白质和基因两个水平均显著低于注射阴性对照液组和注射1006链组（P＜0．05）。结论脑室注射725链siRNA 可以在6 h 内有效沉默大鼠脑内AQP4的表达。%Objective Prepared several small interfering RNAs (siRNA)for AQP4 protein in rat brain by chemical synthe-sis.The silencing effect of two chains were verified and selected the one which was more effective.Methods Select two dif-ferent target sequences of the rat according to the AQP4 mRNA.Two siRNAs were synthesised by chemical synthesis,mean-while negative siRNA was synthesised as control.18 male adult Wistar rats were randomly divided into negative siRNA in-jection group,725 line injection group and 1006 line injection group.Corresponding drugs were injected into the left ventri-cle respectively.All animals were executed after 6 hours.The protein expression of AQP4 was detected by immunohisto-chemical staining and the AQP4mRNA expression was detected by in situ hybridization.Results 6 hours after injection,the expression of AQP4 protein and AQP4 mRNA of brain in the 725 line injection group were significantly lower than those of the 1006 line injection group and the negative siRNA injection group (P<0.05).Conclusion 725 line injection could ef-fectively silence the expression of AQP4 in rat brain in 6 hours.