摘要
目的观察天麻钩藤饮对AngⅡ诱导的大鼠胸主动脉血管平滑肌细胞A7r5中时钟基因(Per2、Bmal1、Clock)、AT1R基因及Src、p-Src蛋白昼夜节律表达的影响,探讨天麻钩藤饮通过AT1R信号途径调节时钟基因昼夜节律变化的可能机制。方法以AngⅡ诱导A7r5细胞建立时钟基因昼夜节律变化模型,将细胞模型分为AngⅡ诱导组、天麻钩藤饮组。AngⅡ诱导组以终浓度为100 nmol·L^(-1)的AngⅡ诱导24 h,天麻钩藤饮组以终浓度为100 nmol·L^(-1)的AngⅡ和200 mg·L^(-1)的天麻钩藤饮处理24 h,以加样时间为ZT0,每隔4小时收集细胞,提取蛋白和RNA,采用Q-PCR方法检测各时间点A7r5细胞中的时钟基因(Per2、Bmal1、Clock)和AT1R基因表达水平,Western Blot检测各时间点A7r5细胞中Src、p-Src蛋白表达水平。结果 AngⅡ作用于A7r5细胞后,时钟基因Bmal1和Per2表达中值升高,峰相位前移,呈现出与正常细胞不同的昼低夜高的反杓型节律变化;AT1R基因表达水平显著升高、Src、p-Src蛋白表达亦呈现出昼低夜高的反杓型节律变化;与AngⅡ诱导组相比,天麻钩藤饮组A7r5细胞Bmal1、Per2基因m RNA表达中值降低,呈现昼高夜低的杓型节律变化,AT1R基因及Src、p-Src蛋白的表达水平下降,呈现类似空白对照组的昼夜节律变化。结论天麻钩藤饮能够通过AT1R信号通路调节AngⅡ诱导的A7r5细胞时钟基因昼夜节律改变。
Objective To observe the effect of Tianma Gouteng Decoction on regulating the circadian rhythm ofclock genes（Per2,Bmal1,Clock）,AT1 R gene and the Src,p-Src protein of the vascular smooth muscle cells（A7 r5 cell）induced by angiotensin Ⅱ（AngⅡ）,and investigate the mechanism of the circadian rhythm variations ofclock gene by Tianma Gouteng Decoction through angiotensin Ⅱreceptor signaling pathway. Methods The circadianrhythm model of A7 r5 cells induced by AngⅡ was established. The cells were divided into AngⅡinduction group,Tianma Gouteng Decoction group. The cells in AngⅡinduction group were cultured with AngⅡ（100 nmol·L^-1）for24 hours,and the cells in Tianma Gouteng Decoction group were treated with AngⅡ（100 nmol·L^-1）and Tianma Gouteng Decoction（200 mg·L^-1）for 24 hours. The drugs were added into cell culture from ZT0,then cells werecollected every four hours; protein and RNA of the cells were extracted. Expression levels of the clock genesincluding Per2,Bmal1,Clock and AT1 R were examine by Q-PCR,and protein expression levels of Src,p-Src atdifferent time points were detected by Western Blot. Results Our findings demonstrated that the means expression levels of Per2 and Bmal1 gene increased in V7 r5 cell induced by Ang Ⅱ. The acrophase shifted shifted forward,which presented an anti-dipper type rhythm that low at day time and high at night. Gene expression level of AT1 Rincreased significantly, as well as the Src and p-Src proteins expression levels. Compared with Ang Ⅱ inductiongroup,the means of Bmal1 and Per2 gene expression of Tianma Gouteng Decoction group decreased,which showeda dipper type of high day and low night; the gene expression of AT1 R and protein expression of Src, p-Srcdecreased as well, representing a circadian rhythm variation similar to the control group. Conclusions Tianma Gouteng Decoction regulates the circadian rhythm of clock genes in A7 r5 cells induced by AngⅡthrough regulationof AT1 R pathway.
作者
杨柳
曾武
张振服
余柱立
江章瑜
叶然
温文浩
刘昌辉
张荣
杨蕾
YANG Liu;ZENG Wu;ZHANG Zhenfu;YU Zhuli;JIANG Zhangyu;YE Ran;WEN Wenhao;LIU Changhui;ZHANG Rong;YANG Lei(Institute of Clinical Pahrmacology, Guangzhou University of Chinese Medicine, Guangzhou 510405 Guangdong, China)
出处
《中药新药与临床药理》
CAS
CSCD
北大核心
2018年第3期307-312,共6页
Traditional Chinese Drug Research and Clinical Pharmacology
基金
国家自然科学基金面上项目(81173182
81773969)
国家自然科学基金青年科学基金项目(30801462)
广东省自然科学基金项目(S2012010010195)
广州中医药大学青年英才项目
广州中医药大学高水平建设项目(A1-AFD018161Z1515
A1-AFD018161Z1518)
广州市中医药防治脑病研究重点实验室资助项目(201805010005)
关键词
天麻钩藤饮
时钟基因
AT1R信号通路
昼夜节律
Tianma Gouteng Decoction
clock gene
AT1 receptor signaling pathway
circadian rhythm
