RNAi干扰大鼠VSMCs钟基因Per2对Dbp及AT_1受体基因表达的影响  

The Influence on the Expression of Clock gene and AGTR1 after the Silence of Rat Per2 gene by RNA Interference in Vascular Smooth Muscles Cells

作　　者：王翔凌[1] 孙宁玲[2] 马丽萍[3] 郭晓夏[2] 姜娟[2] 何湘君[3] WANG Xiang-ling;SUN Ning-ling;MA Li-ping;GUO Xiao-xia;JIANG Juan;HE Xiang-jun(： Department of Cardiology, Beijing Hospital of The Ministry of Health, Beijing 100730, China;：Department of Cardiology, The People＇s Hospital of Peking University, Beijing 100044, China;Central Lab, The People＇s Hospital of Peking University, Beijing 100044, China.)

机构地区：[1]卫生部北京医院心内科,北京市100730 [2]北京大学人民医院心内科,北京市100044 [3]北京大学人民医院中心实验室,北京市100044

出　　处：《中国分子心脏病学杂志》2018年第3期2462-2467,共6页Molecular Cardiology of China

基　　金：国家自然科学基金面上项目(81670274)

摘　　要：目的应用RNAi技术干扰大鼠A7r5血管平滑肌细胞(vascular smooth muscle cells,VSMCs)Per2基因并检测VSMCs时钟基因Bmal1、输出基因Dbp及AT1受体基因(AGTR1)m RNA表达的变化,初步探讨时钟基因、输出基因及AGTR1间的相关性。方法应用已筛选携带大鼠Per2基因干扰序列的慢病毒载体转染大鼠血管平滑肌A7r5细胞,通过RT-PCR方法检测时钟基因Per2及Bmal1、钟控基因Dbp和AGTR1 m RNA水平及节律变化。结果慢病毒载体转染A7r5细胞后,RT-PCR方法检测钟基因Per2的m RNA峰值时相后移,表达节律异常;钟基因Bmal1的m RNA表达峰值增强,表达节律未受影响;下游输出基因Dbp的m RNA表达明显受抑,表达节律异常;AGTR1的m RNA表达后期明显下降,提示其变化延迟于Per2变化。结论通过RNAi技术沉默大鼠血管平滑肌细胞中时钟基因Per2表达,使钟基因Bmal1反向增强,明显抑制输出基因Dbp的表达,影响其节律,可能造成AGTR1的异常延后表达。Objective To make the rat Per2 gene silence in vascular smooth muscle cells by using RNA interference sequence and evaluate the influence on the mRNA expression of the Bmall, Dbp and AGTR1 gene. The results may help us to figure out the probable relationship with gene Per2 and AGTR1. Methods RNA inference sequences, which has been proved to depress the expression of the rat Per2 gene, was transducted in rat A7r5 VSMCs by Lentivirus vector. Then, the influence on the mRNA expression of Per2, Bmall, Dbp and AGTRI gene will be detected by real-time PCR. Results The real-time PCR analysis confirmed that the levels of Per2 and Dbp mRNA were reduced significantly as compared with the negative control group, but the levels of Bmall were increased. And the rhythm of Dbp and Per2 were abnormal. The levels of AGTR1 were significantly reduced at CT20. These results may imply that the variation of AGTR1 is the influence of Per2 that delay to the reduction of Per2. Conclusion The rat lentivirus vectors with Per2-specific shRNA can make the expression of Per2 gene depress in A7r5 VSMCs. The influence of this silence is to decrease the expression of Dbp and interfere the rhythm of Dbp. Otherwise, the variation of the AGTR1 expression at last may be described by the influence of Per2 on the downstream gene.

关 键 词：RNA干扰 时钟基因 血管平滑肌细胞 AT1受体基因 

分 类 号：R363[医药卫生—病理学]