Function and regulation of intestinal p450 enzymes: Insights from conditional cytochrome p450 reductase knockout mouse models

The overall goal of this thesis work was to investigate the function and regulation of intestinal P450 enzymes (P450s). The central hypotheses are that 1) small intestinal (SI) P450s play an important role in the first-pass clearance of oral drugs and in bioactivation of chemicals that cause intestinal toxicity, 2) endogenous regulatory compounds produced by intestinal P450s are important for protection against chemical-induced colon inflammation, and 3) there is a cross talk between the liver and the SI that helps to maintain the overall xenobiotic-metabolism capacity of the digestive tract. To test these hypotheses, our specific aims are to determine the roles of SI P450s 1) in modulating the bioavailability of oral lovastatin (LVS); 2) in diclofenac (DCF)-induced SI toxicity; and 3) in protection against dextran sodium sulfate (DSS)-induced colon inflammation; and to examine 4) the impact of liver-specific loss of the P450 reductase (CPR) on the expression and function of SI P450s. In Chapter Two, an intestinal epithelium-specific Cpr-null (IE-Cpr-null) mouse model was used to determine the role of SI P450s in the first-pass metabolism of LVS. The results showed that the bioavailability of oral LVS was 15% in IE-Cpr-null mice, compared to 5% in wild-type (WT) mice. These findings provided definitive evidence that SI P450s play an essential role in modulating the bioavailability of oral LVS. In Chapter Three, the SI P450-mediated bioactivation of DCF was found suppressed in IE-Cpr-null mice. Reductions in rates of reactive metabolites formation and amounts of DCF protein adducts formed were also observed, which may contribute to the resistance of IE-Cpr-null mice to DCF-induced toxicity in the SI. Grapefruit juice (GFJ) treatment was found to ameliorate DCF-induced ulcer formation in WT, but not in IE-Cpr-null mice, via inhibition of SI P450-mediated formation of DCF reactive metabolites. These findings demonstrated that SI P450s play a critical role in DCF-induced toxicity in the SI. In Chapter Four, IE-Cpr-null mice were found to be more sensitive than WT mice to DSS-induced colitis, possibly due to reduced local corticosterone (CC) biosynthesis in the colon. Co-treatment of DSS-exposed mice with deoxycorticosterone (DOC), a precursor of CC biosynthesis, abolished the hypersensitivity of IE-Cpr-null mice to DSS-induced colitis. These results suggest that intestinal microsomal P450 enzymes play a critical role in protecting colon from DSS-induced inflammation, possibly through increased local CC synthesis in response to DSS challenge. In Chapter Five, the liver-specific Cpr-null (LCN) mouse model was used to examine the impact of hepatic P450/CPR deficiency on the expression and function of SI P450s. The results indicated that the loss of hepatic P450/CPR activities in the LCN mice led to compensatory increases in SI P450 expression and capacity of first-pass metabolism of oral LVS. The fibroblast factor 15 (FGF15), which is produced in the ileum and known to regulate bile acid synthesis in the liver, was found to play a direct role in the regulation of SI P450 expression. Overall, these findings are important for improvements in drug bioavailability, for identification of sites of potential drug-drug interactions, and for prevention against xenobiotic induced toxicity in the intestine.