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The present study have established an hPXR mediated reporter gene assay for UGTIAI, and it was used as an in vitro model to evaluate the induction activity of various commonly used traditional Chinese medicines (TCMs) extract and their constituents toward UGT1A1. And their effects on expression of UGT1A1 RNA and protein were then investigated by real-time PCR and western-blotting.Objective To develop an hPXR mediated reporter gene assay for UGT1A1, and use it as an in vitro model to evaluate the induction activity of various compounds toward UGTIAI.Methods The distal promoter and proximal promoter of UGTIAI were amplified from human genetic DNA, and then cloned into pGL3 vector to form pGL3-PXRE plasmid. The pGL3-PXRE plasmid was then cotransfected into HepG2 cells with hPXR expression plasmid pcDNA3.1-hPXR and reference plasmid pRL-TK to establish a reporter gene model. The model was applied to determine the induction activity of various commonly used TCMs extract and their constituents toward UGTIAI. Then real-time PCR and western-blotting as used to validate the induction effect of those compounds by investigating the change of UGTIAI RNA and protein level. Results The promoters of UGT1A1 were successfully cloned into pGL3 vector to form pGL3-PXRE recombinant plasmid, and the hPXR mediated reporter gene model was successfully established. Induction effects of 9 TCMs extracts and 114 constituents were investigated. Three TCMs extract including Schisandra sphenanthera, Atractylodes and Ligusticum chuanxiong extracts and 20 compounds including chrysin, luteolin, tanshinoneⅠ, tanshinoneⅡA, cryptotanshinone, schisandrin A, schisandrin B, schisandrin C, schisantherin A, schizandrol A, ginkgolide A, ginkgolide B, ginkgolic acid(17:1), piperine, rhynchophylline, vindoline, rutaecarpine, hyperincin, artemether, oridonin were found to activate hPXR and therefore have the potential to induce UGT1A1. Real-time PCR and western-blotting were used to validate the induction activity of those compounds, and the results showed that chrysin, tanshinoneⅠ, tanshinoneⅡA, cryptotanshinone, hyperincin and rutaecarpine could induce the RNA and protein expression of UGT1A1.Conclusion The model established is an effective method to determine the activator of hPXR and potential inducer of UGT1A1 in vitro. Several compounds were found to have the potential to induce UGT1A1. Therefore, more attention should be paid on the coadministration of these constituents of TCMs with the drugs metabolized by UGT1A1, to avoid the undesired drug-drug interactions.
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