Cytochrome P450 3A represents the most abundant P450 subfamily in human liver, and is responsible for the metabolism of >50 % of drugs on the market today. An area of intense study has been the xenobiotic regulation of CYP3A gene expression: this phenomenon potentially leads to altered pharmacokinetics/pharmacodynamics of substrates and adverse drug reactions. The use of hepatoma cell lines is common in drug metabolism studies; HuH7 is a human hepatoma cell line increasingly being used in both academia and the pharmaceutical industry. However, little is known regarding the expression profile of these cells. Recent data have made it increasingly clear that the gene expression profile of a cell system, and its alteration in response to external stimuli, is important in controlling expression of cytochrome P450 3As. Using quantitative PCR, transcript levels of CYP3A subfamily members and transcription factors implicated in their regulation were obtained in human adult and foetal RNA samples. These levels were then compared to those obtained from both primary human hepatocytes and HuH7 cells, showing that in general, primary or hepatoma cells show a distinct profile compared to either adult or foetal samples, with the major variables between the cell types being ratios of the transcription factors COUP-TF1 and RXRalpha to PXR. Whereas exposure to the CYP3A transcriptional activators PCN, rifampicin, dexamethasone and phenobarbital produced induction in primary human hepatocytes, no transcriptional activation was observed in HuH7 cells, suggesting that regulatory pathways were disrupted. Alterations in the gene expression profile, particularly the levels of COUP-TFl and PXR, and in the chromatin conformation surrounding the CYP3A gene cluster, were implicated to underlie this disruption. In conclusion, I have demonstrated that there are a number of levels of control of CYP3A gene expression, with both transcription factor expression levels and chromatin status having an impact on transcriptional activation of CYP3A genes in human hepatocytes.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:412044 |
| Date | January 2004 |
| Creators | Phillips, Anna Louise |
| Publisher | University of Surrey |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://epubs.surrey.ac.uk/842667/ |
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