High-Fat Diets Alter the Modulatory Effects of Xenobiotics on Cytochrome P450 Activities

Abstract

Cytochrome P450 monooxygenase (P450) enzymes metabolize critical endogenous chemicals and oxidize nearly all xenobiotics. Dysregulated P450 activities lead to altered capacity for drug metabolism and cellular stress. The effects of mixed exposures on P450 expression and activity are variable and elusive. A high-fat diet (HFD) is a common exposure that results in obesity and associated pathologies including hepatotoxicity. Herein, we report the effects of cigarette smoke on P450 activities of normal weight and HFD induced obese mice. Activity-based protein profiling results indicate that HFD mice had significantly decreased P450 activity, likely instigated by proinflammatory chemicals, and that P450 enzymes involved in detoxification, xenobiotic metabolism, and bile acid synthesis were effected by HFD and smoke interaction. Smoking increased activity of all lung P450 and coexposure to diet effected P450 2s1. We need to expand our understanding of common exposures coupled to altered P450 metabolism to enhance the safety and efficacy of therapeutic drug dosing.

Figure 1.

ABPP of P450 activity due to individual or concomitant exposures. Mice were exposed to HFD and/or CSE, and liver and lungs harvested. Microsomes were isolated from tissue homogenates and incubated with activity-based probes (ABP). Following P450 labeling, click chemistry was used to append azido biotin, and the protein-probe-biotin complexes were enriched on streptavidin resin. ABP-targeted enzymes were trypsin digested on-resin and the resulting peptides were analyzed by quantitative high-resolution LC-MS.

Figure 2.

Mean body weight (A), and mean serum glucose (B) at necropsy (± SE, n = 8). After 11 weeks on a SD or a HFD, the HFD mice had significantly increased (p<0.05) body weight and serum glucose concentration compared to the SD counterparts.

Figure 3.

A-E. Active P450 profiles for mice fed standard diet (SD) or high-fat diet (HFD), and exposed to active or passive cigarette smoke (ACSE and PCSE respectively), as determined by ABPP. A) liver and B) lung heatmap columns represent each exposure group, and each row maps to a functionally active P450 as determined by probe labeling and measurement by LC-MS and the log2 transformed mean (n=8) relative abundance. These liver data can be found in Dataset S1, and lung data can be found in Dataset S2. Each P450 is grouped by primary substrate (fatty acid, xenobiotic, steroid) or “orphan” if the enzyme function is still unclassified. Arrows indicate key functions and products. C) liver data, D) lung data with significantly different (p<0.05) relative abundance values that were increased or decreased compared to the control group (NoCSE-SD). Primary substrate(s) are included next to P450 in bar graph: fatty acid (FA), xenobiotic (X), steroid (S), or orphan (O). E) 2-way ANOVA with Bonferonni correction performed to test the significance of the diet and smoke interaction on liver P450 and lung P450 ABPP activity. Graph only includes P450 with significant differences (p<0.05).

Figure 4.

Bile acid synthesis and regulation. A) Hepatic metabolism of cholesterol to bile acids. Key enzymes involved in bile acid synthesis, metabolites that function as nuclear receptor ligands (outlined in pink), corresponding nuclear receptor (pink text) and their regulatory effects. B) Once conjugated, primary bile acids reach the intestinal tract via gallbladder, the microbiome transforms BA prior to enterohepatic cycling or excretion. C) Heatmap view of HFD and CSE effect on P450 7a1 and 8b1 (details in Figure 3A with the exception that data presented here is Z-scaled). D) List of pathologies regulated by BA. Abbreviations: BAAT; BSH, bile acid hydrolase; BACS, bile acid CoA synthase; BAAT, bile acid CoA–amino acid N-acetyltransferase; FXR, Farnesoid X receptor; LXR, Liver X receptor; IBD, Irritable Bowel Syndrome; T2DM, Type 2 Diabetes Mellitus.

Figure 5.

Stress induced eicosanoid synthesis. General overview of stress-induced activation of phospholipase A₂ (PLA₂) to release arachidonic acid from the cell membrane, and the varied metabolic pathways where arachidonic acid is converted to eicosanoids including: Cyclooxygenase-1 and −2 (COX1/2), 5-, 8-, 12-, and 15-lipoxygenases (5-, 8-, 12-, 15-LOX), and cytochrome P450 epoxyhydrolase and ω-hydroxylase (green text). Eicosanoid species (black text) and proposed receptors (pink text). Abbreviations: ALX, COX, cyclooxygenase; P450, cytochrome P450; EET, epoxyeicosatrienoic acid; GPR, G protein-coupled receptor; HETE, hydroxyeicosatetraenoic acid; HX, hepoxilin; LOX, lipoxygenase; LX, lipoxin; PLA2, phospholipase A2; PPAR, peroxisome proliferator-activated receptor; TRPA1, transient receptor potential ankyrin 1; TRPV1, transient receptor potential vanilloid 1; and TX, thromboxane.