抑制CYP4A11基因表达的siRNAs优选及对血管收缩相关因子的影响  

Optimization of siRNAs in Inhibiting the Expression of CYP4A11 Gene and Effects on Related Factors Affecting Blood Vessel Contraction

作　　者：李晋[1,2] 胡静[3] 李健[1] 石磊[1] 赵树进[1,2] 

机构地区：[1]广州军区广州总医院药剂科,中国广东广州510006 [2]华南理工大学生物科学与工程学院,中国广东广州510006 [3]广州军区广州总医院肾脏内科,中国广东广州510010

出　　处：《生命科学研究》2017年第1期41-45,54,共6页Life Science Research

基　　金：广州市科技计划项目(201509010012)

摘　　要：为了考察人细胞中细胞色素P450 4A11(CYP4A11)表达受抑制之后是否会影响血管相关收缩因子的表达,在线设计多个抑制细胞中代谢酶编码基因CYP4A11表达的siRNAs并通过BLAST验证;选择效果好的3对siRNAs合成并转染至EA.hy926细胞株,通过荧光染色和定量多聚酶链反应(quantitative polumerase chain reaction,qPCR)初步检测后,再采用表达量相对更高的MG63细胞系进行验证,发现25 nmol/L的siRNA2抑制效果最好;随后使用25 nmol/L siRNA2抑制CYP4A11在EA.hy926细胞株中的表达,并通过qPCR与Western-blot检测血管收缩相关因子的表达情况,发现促血管收缩相关因子内皮素1受体(endothelin 1 receptor,ET1R)、血管紧张素1型受体(angiotensin type 1 receptor,AT1R)表达下调(P<0.05),而内皮型一氧化氮合酶(endothelial nitric oxide synthase,eNOS)的表达显著上调(P<0.001)。因此,siRNA2能通过显著抑制EA.hy926细胞中CYP4A11的表达,调节血管紧张的相关指标,有促血管松弛作用的趋势。To investigate the relationship between the suppressed expression of the metabolic enzyme CYP4A11 in the human cells and the vasoconstriction-related factors, gene silencing techniques were used to suppress CYP4A 11 expression in cells after siRNAs were online designed and verified by BLAST. Three siRNAs were synthesized and transferred into the EA.hy926 cells. After detection by florescence expression and qPCR, and verification by transfection of MG63 cells which could express higher amount of CYP4A11, siRNA2 at a dose of 25 nmol/L proved to have the best inhibitory effect on CYP4A 11. When 25 nmol/L siRNA2 was used to inhibit the expression of CYP4A 11 in EA.hy926 cells, the expressions of vasoconstric- tion-related factors ET1R and AT1R decreased significantly （P〈0.05） and eNOS was upregulated obviously （P〈0.001）, as detected by qPCR and Western-blot. Therefore, siRNA2 can significantly inhibit the expres- sion of CYP4A 11 in EA.hy926 cells, and regulate the related factors that affect the tension in blood vessels,promoting blood vessel relaxation.

关 键 词：CYP4A11 小干扰RNA(si RNA) 血管收缩因子 内皮素1受体(ET1R) 血管紧张素1型受体(AT1R) 

分 类 号：Q955[生物学—动物学]