中国药理学通报Issue(2)：289-293,5.DOI:10.3969/j.issn.1001-1978.2015.02.028
基于报告基因检测的PXR、FXR和LXRα激动剂高通量筛选模型的建立
Establishment of cell models for PXR, FXR and LXRαagonists high-throughput screening based on reporter gene assay
摘要
Abstract
Aim To develop an in vitro high throughput drug screening system based on reporter gene assay for identification of novel compounds with PXR, FXR and LXRα agonist activity. Methods The expressions of exogenous PXR, FXR and LXRαgene in HEK293, HepG2 and LS174T cells were examined by Real-Time quantity PCR. pSG5-hPXR and pGL3-XREM-CYP3A4, pEGFP-N3-hFXR and EcRE-TK-Luc, pCMX-FLAG-hLXRα and pGL3-XREM-CYP3A4 were cotransfected into cells and the optimal ratio of three plasmids was determined. The dose-response relationship between the positive drug and the fold induction was determined. The specificity of the model was ex-amined, and the repeatability was also determined by Z′ value. Results ① The PXR, FXR and LXRα mRNA expression in HEK293 cell is low among three different cells. ②reporter gene vector and expression plasmid ratio of 1∶ 1, 2∶ 1 and 2∶ 1 were proved to be suitable for highest relative luciferase activity for PXR, FXR or LXRα agonist screening model. ③ The relative luciferase activity was induced by Rif, CDCA or T0901317 in a dose-dependent manner. ④Only Rif, CDCA or T0901317 could significantly increase the relative luciferase activity in PXR,FXR or LXRα agonist screening model, no effect of other nuclear re-ceptors agonist was observed, and the values of Z′-factor for PXR, FXR and LXRαagonist screening model were 0. 58, 0. 66 and 0. 63, respectively. Conclusion An in vitro PXR, FXR and LXRα agonist high-throughput screening models are devel-oped with acceptable specificity and repeatability, and the mod-els can be used to screen PXR, FXR and LXRα agonist.关键词
核受体/报告基因/高通量筛选/PXR/FXR/LXRαKey words
nuclear receptor/reporter gene/high-throughput screening/PXR/FXR/LXRα分类
医药卫生引用本文复制引用
庄嘉琅,曾行,钟国平,金晶,苟晓丽,毕惠嫦,黄民..基于报告基因检测的PXR、FXR和LXRα激动剂高通量筛选模型的建立[J].中国药理学通报,2015,(2):289-293,5.基金项目
广东省新药设计与评价重点实验室开放基金( No 2011 A060901014-009) （ No 2011 A060901014-009）
