Feed-forward Regulation of Bile Acid Detoxification by CYP3A4

Abstract

Bile acids are potentially toxic end products of cholesterol metabolism and their concentrations must be tightly regulated. Homeostasis is maintained by both feed-forward regulation and feedback regulation. We used humanized transgenic mice incorporating 13 kb of the 5 regulatory flanking sequence of CYP3A4 linked to a lacZ reporter gene to explore the in vivo relationship between bile acids and physiological adaptive CYP3A gene regulation in acute cholestasis after bile duct ligation (BDL). Male transgenic mice were subjected to BDL or sham surgery prior to sacrifice on days 3, 6, and 10, and others were injected with intraperitoneal lithocholic acid (LCA) or vehicle alone. BDL resulted in marked hepatic activation of the CYP3A4/lacZ transgene in pericentral hepatocytes, with an 80-fold increase in transgene activation by day 10. Individual bile acids were quantified by liquid chromatography/mass spectrometry. Serum 6␤-hydroxylated bile acids were increased following BDL, confirming the physiological relevance of endogenous Cyp3a induction to bile acid detoxification. Although concentrations of conjugated primary bile acids increased after BDL, there was no increase in LCA, a putative PXR ligand, indicating that this cannot be the only endogenous bile acid mediating this protective response. Moreover, in LCA-treated animals, 5-bromo-4-chloro-3-indolyl-␤-D-galactopyranoside staining showed hepatic activation of the CYP3A4 transgene only on the liver capsular surface, and minimal parenchymal induction, despite significant liver injury. This study demonstrates that CYP3A up-regulation is a significant in vivo adaptive response to cholestasis. However, this up-regulation is not dependent on increases in circulating LCA and the role of other bile acids as regulatory molecules requires further exploration.

Benefit of farnesoid X receptor inhibition in obstructive cholestasis

“…This injury has been attributed to the loss of various detoxification mechanisms, because concentrations of bile acids are not increased in these mice (13). This study shows that concurrent deletion of another nuclear receptor is able to reduce this liver injury and explore mechanisms for this protection.…”

“…Mice were anesthetized and aseptically subjected to BDL or sham operation as described in ref. 13, then killed by exsanguination under anesthesia 6 days after surgery. Blood samples (35 l) were collected into heparinized capillary tubes from the tail vein of mice before the initial surgical procedure on day 0 and repeated on days 3 and 6 before killing.…”

“…2A), with increases in all major primary bile acids and their respective metabolites in WT mice as reported in ref. 13. In PXRKO mice, a relative reduction in concentration of the hydroxylated bile acid, tauro-␤-muricholic acid (T␤-MCA), was observed (Fig.…”

“…Bile duct ligation (BDL) is an established animal model of cholestasis, with complete biliary obstruction and accumulation of multiple primary bile acids in liver and serum (13). Changes in serum lipids and increases in circulating endotoxin also parallel those that occur in humans with cholestasis (9).…”

Regulatory network of lipid-sensing nuclear receptors: roles for CAR, PXR, LXR, and FXR

“…FXR-mediated induction of SHP by bile acids decreases the transcriptional activity of PXR [94]. Moreover, in mice with transgenically incorporated human CYP3A4 5'-flanking region linked to a reporter gene, increase in CYP3A4-driven reporter gene expression is not primarily dependent on the levels of circulating lithocholic acid, the primary bile acid-ligand of mouse PXR [95]. FXR activates CYP3A4 drug-responsive enhancer elements and might thus directly increase CYP3A levels [51,83].…”