During infection or inflammatory stress, the capacity of the liver to metabolize drugs is impaired due to a reduction in the hepatic content of cytochrome P450 (P450). This can lead to elevated serum levels, and exaggerated toxicity, of clinically important drugs. There is evidence that the cytokines interleukin-1 (IL1) and interleukin-6 (IL6), as well as interferons and glucocorticoids, all may contribute as blood-borne mediators of this effect on the liver. These agents decrease total hepatic P450 content and the levels of the protein and mRNA products of certain P450 genes. In addition to their putative roles in the inflammatory response, cytokines and interferons are under investigation for potential clinical applications, while glucocorticoid drugs are widely used in medical practice. Therefore, it is important to determine which of these agents affect P450 gene expression, what P450 gene products are affected, and how these effects are attained. The goals of this project are to characterize the suppression of P450 genes by these agents in vivo, and to understand the mechanisms by which their suppressive effects are achieved. The relative contributions of IL1, IL6, interferons and glucocorticoids will be assessed by investigating their effects on a number of constitutively expressed P450 proteins and mRNAs, as well as on specific P450 gene transcription, in rat livers in vivo and in cultured rat hepatocytes. Thus, we will determine whether the suppression of P450 enzymes by the above agents are achieved by transcriptional, posttranscriptional, or posttranslational mechanisms. P450 enzymes that are expressed in both humans and rats, and P450s that are expressed only in rats, will be studied to allow a critical evaluation of the applicability of our results to humans. An Il1 receptor antagonist protein will be used to delineate the role of Il1 in P450 suppression in vivo and in vitro. The cellular signalling pathways by which the inflammatory mediators suppress P450 expression will be investigated in hepatocyte cultures, using known agonists and inhibitors of signalling pathway components. The DNA sequences necessary for inflammatory suppression of P450 genes will be sought by studying the effects of deleting potential regulatory regions on transcription of cloned fragments of the P450IIC11 and P450IIC12 genes in extracts of rat liver nuclei. Lastly, the nuclear proteins that bind to these regions, and whose binding is modulated by inflammation, will be identified and purified.
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