人孕烷X受体介导的UGT1A1报告基因模型的建立及初步应用

 目的 建立基于人孕烷X受体（human pregnane X receptor，hPXR）的UGT1A1的报告基因模型，研究中药提取物通过人孕烷X受体途径对UGT1A1的潜在诱导能力。方法 以人基因组DNA为模板扩增UGT1A1的远端和近端启动子，连接到pGL3载体上，构建pGL3-PXRE重组质粒，并与人孕烷X受体表达质粒共转染HepG2细胞，从而建立报告基因模型。运用该模型考察了9种常见中药对人孕烷X受体的激活作用，即对UGT1A1的潜在诱导作用。结果 将UGT1A1启动子片段克隆到pGL3载体上，构建了pGL3-PXRE重组质粒，成功构建了报告基因模型，并考察了多种常见中药的提取物通过人孕烷X受体途径对UGT1A1的诱导作用。结论 成功地建立了基于人孕烷X受体的UGT1A1的报告基因模型，为人孕烷X受体激活剂的体外筛选提供了一种有效的方法。

Establishment of hPXR Mediated Reporter Gene Model for UGT1A1 and Its Application

OBJECTIVE To develop hPXR mediated reporter gene model for UGT1A1， and use it as an in vitro model for determination of induction activity toward UGT1A1 of various commonly used traditional Chinese medicines (TCMs) extracts. METHODS The distal and proximal promoters of UGT1A1 were amplified from human genetic DNA， and were cloned into pGL3 vector as pGL3-PXRE plasmid， which was then cotransfected into HepG2 cells with hPXR expression plasmid to establish the reporter gene model. Then the model was applied to determine the induction activity toward UGT1A1 of various commonly used TCMs extracts. RESULTS The promoters of UGT1A1 were successfully cloned into pGL3 vector to form pGL3-PXRE recombinant plasmid， and the reporter gene model was successfully established. Nine kinds of TCMs extracts were investigated， and three were found to activate hPXR and therefore have the potential to induce UGT1A1. CONCLUSION The model established is an effective method for determination of activators of hPXR and potential inducers of UGT1A1 in vitro.