Feed-forward regulation of bile acid detoxification by CYP3A4: studies in humanized transgenic mice.
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 6beta-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-beta-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.
Stedman
Catherine
C
Department of Clinical Pharmacology, University of Sydney, New South Wales 2145, Australia.
Robertson
Graham
G
Coulter
Sally
S
Liddle
Christopher
C
D000818:Animals; D001483:Base Sequence; D002779:Cholestasis / Q000201:enzymology / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism*; D017931:DNA Primers; D006801:Humans; D008658:Inactivation, Metabolic; D007763:Lac Operon; D008095:Lithocholic Acid / Q000008:administration & dosage / Q000097:blood / Q000378:metabolism* / Q000652:urine; D008099:Liver / Q000378:metabolism / Q000503:physiopathology; D008297:Male; D051379:Mice; D008822:Mice, Transgenic; D009929:Organ Size
D017931:DNA Primers; D008095:Lithocholic Acid; D003577:Cytochrome P-450 Enzyme System; C104464:CYP3A protein, human; D051544:Cytochrome P-450 CYP3A; C510163:CYP3A4 protein, human