High-Fat Diet Feeding Alters Expression of Hepatic Drug-Metabolizing Enzymes in Mice

Abstract

Medical conditions accompanying obesity often require drug therapy, but whether and how obesity alters the expression of drug-metabolizing enzymes and thus drug pharmacokinetics is poorly defined. Previous studies have shown that high-fat diet (HFD) feeding and subsequent obesity in mice lead to altered expression of transcriptional regulators for cytochrome P450 CYP2D6, including hepatocyte nuclear factor 4α (HNF4α, a transcriptional activator of CYP2D6) and small heterodimer partner (SHP, a transcriptional repressor of CYP2D6). The objective of this study was to examine whether diet-induced obesity alters CYP2D6 expression by modulating HNF4α and SHP expression. Male CYP2D6-humanized transgenic (Tg-CYP2D6) mice were fed with HFD or matching control diet for 18 weeks. Hepatic mRNA expression of CYP2D6 decreased to a small extent in the HFD group (by 31%), but the differences in CYP2D6 protein and activity levels in hepatic S9 fractions were found insignificant between the groups. Although hepatic SHP expression did not differ between the groups, HNF4α mRNA and protein levels decreased by ∼30% in the HFD group. Among major mouse endogenous cytochrome P450 genes, Cyp1a2 and Cyp2c37 showed significant decreases in the HFD group, whereas Cyp2e1 expression did not differ between groups. Cyp2b10 and Cyp3a11 expression was higher in the HFD group, with corresponding 2.9-fold increases in hepatic CYP3A activities in HFD-fed mice. Together, these results suggest that obesity has minimal effects on CYP2D6-mediated drug metabolism, although it modulates the expression of mouse endogenous P450s in a gene-specific manner.

Fig. 1.

Expression and activity of HNF4α and CYP2D6 in HFD-fed Tg-CYP2D6 mice. Liver tissues were collected from Tg-CYP2D6 mice after 18 weeks of HFD or matching control (Ctrl) diet feeding. (A) Relative Hnf4α mRNA levels (left) determined by qRT-PCR were normalized by that of control group. Relative HNF4α protein levels (right) were normalized against β-actin band intensities. (B) Hepatic mRNA levels of representative HNF4α target genes were determined by qRT-PCR and normalized by that of control group. (C) Hepatic mRNA level of CYP2D6 was normalized by that of control group. (D) CYP2D6 protein level in hepatic S9 fraction (right) was determined using Western blot normalized against β-actin band intensities. (E) CYP2D6 activity in mouse liver S9 fraction was measured using debrisoquine as a substrate. CYP2D6 activity is presented as 4-hydroxydebrisoquine formation rate per mg S9 protein. (F) Total CYP2D6 activity per liver was the product of CYP2D6 activity per mg S9 protein and total S9 protein level per liver. N = 6–8 per group for qRT-PCR and activity assay. Data shown are mean ± S.D. NS, not significant, *p < 0.05, **p < 0.01, ***p < 0.001.

Fig. 2.

Mouse endogenous P450 expression upon HFD feeding in Tg-CYP2D6 mice. Liver tissues were collected from mice after 18 weeks of HFD or matching control (Ctrl) diet feeding. (A) Hepatic mRNA levels of drug-metabolizing P450s were measured using qRT-PCR and normalized to those of the control group. (B) Mouse CYP3A activity in liver S9 fraction was measured using triazolam as a substrate. CYP3A activity is presented as 1-hydroxytriazolam formation rate. (C) Mouse CYP3A activity correlated with Cyp3a11 relative mRNA level (Pearson correlation efficiency r = 0.81, p < 0.001). (D) Hepatic mRNA levels of Pxr and Car were determined using qRT-PCR and normalized by those of the control group. N = 7–8 per group. Data shown are mean ± S.D. NS, not significant, *p < 0.05, **p < 0.01.