Abstract
Ginkgo biloba extract activates pregnane X receptor (PXR), but how this occurs is not known. Therefore, we investigated the mechanism of PXR activation by the extract and the role of five individual terpene trilactones in the activation. In a cell-based reporter gene assay, G. biloba extract activated human PXR (hPXR), and at a concentration present in the extract, ginkgolide A, but not ginkgolide B, ginkgolide C, ginkgolide J, or bilobalide was partially responsible for the increase in hPXR activity of the extract. Likewise, in cultured human hepatocytes, only ginkgolide A contributed to the increase in hPXR target gene expression (CYP3A4 mRNA and CYP3A-mediated testosterone 6β-hydroxylation). The extract, but none of the terpene trilactones, bound to hPXR ligand-binding domain, as analyzed by a time-resolved fluorescence resonance energy transfer competitive binding assay. Only the extract and ginkgolide A recruited steroid receptor coactivator-1, as determined by a mammalian two-hybrid assay. Compared with hPXR, rat PXR (rPXR) was activated to a lesser extent by G. biloba extract. Similar to hPXR, only ginkgolide A contributed to rPXR activation by the extract. In contrast to the effect of G. biloba extract on PXR function, it did not affect hPXR expression. Overall, the main conclusions are that G. biloba extract is an hPXR agonist, and among the five terpene trilactones investigated, only ginkgolide A contributes to the actions of the extract. Our findings provide insights into the biological and chemical mechanisms of hPXR activation by G. biloba extract.
Footnotes
This work was supported by the Canadian Institutes of Health Research [Grant MOP-84581]; a major equipment grant from the Dawson Endowment Fund in Pharmaceutical Sciences; and a Senior Scholar Award from the Michael Smith Foundation for Health Research (to T.K.H.C.).
Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
doi:10.1124/jpet.110.172338.
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ABBREVIATIONS:
- PXR
- pregnane X receptor
- SRC-1
- steroid receptor coactivator-1
- hPXR
- human pregnane X receptor
- rPXR
- rat pregnane X receptor
- SR12813
- tetraethyl 2-(3,5-di-t-butyl-4-hydroxyphenyl)ethenyl-1,1-bisphosphonate
- PCN
- pregnenolone 16α-carbonitrile
- TO901317
- N-(2,2,2-trifluoro-ethyl)-N-[4-(2,2,2-trifluoro-1-hydroxy-1-trifluoromethyl-ethyl)-phenyl] benzenesulfonamide
- DMSO
- dimethyl sulfoxide
- HBSS
- Hanks' balanced salt solution
- LDH
- lactate dehydrogenase
- PCR
- polymerase chain reaction
- TR-FRET
- time-resolved fluorescence resonance energy transfer
- HPRT
- human hypoxanthine phosphoribosyltransferase 1
- MEM
- minimum essential medium.
- Received July 1, 2010.
- Accepted August 24, 2010.
- Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics
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