ABSTRACT:Members of the cytochrome P450 (P450) enzyme families CYP1, CYP2, and CYP3 are responsible for the metabolism of approximately 75% of all clinically relevant drugs. With the increased prevalence of nonalcoholic fatty liver disease (NAFLD), it is likely that patients with this disease represent an emerging population at significant risk for alterations in these important drug-metabolizing enzymes. The purpose of this study was to determine whether three progressive stages of human NALFD alter hepatic P450 expression and activity. Microsomes isolated from human liver samples diagnosed as normal, n ؍ 20; steatosis, n ؍ 11; nonalcoholic steatohepatitis (NASH) (fatty liver), n ؍ 10; and NASH (no longer fatty), n ؍ 11 were analyzed for P450 mRNA, protein, and enzyme activity. Microsomal CYP1A2, CYP2D6, and CYP2E1 mRNA levels were decreased with NAFLD progression, whereas CYP2A6, CYP2B6, and CYP2C9 mRNA expression increased. Microsomal protein expression of CYP1A2, CYP2C19, CYP2D6, CYP2E1, and CYP3A4 tended to decrease with NAFLD progression. Likewise, functional activity assays revealed decreasing trends in CYP1A2 (p ؍ 0.001) and CYP2C19 (p ؍ 0.05) enzymatic activity with increasing NAFLD severity. In contrast, activity of CYP2A6 (p ؍ 0.001) and CYP2C9 (diclofenac, p ؍ 0.0001; tolbutamide, p ؍ 0.004) was significantly increased with NAFLD progression. Increased expression of proinflammatory cytokines tumor necrosis factor ␣ and interleukin 1 was observed and may be responsible for observed decreases in respective P450 activity. Furthermore, elevated CYP2C9 activity during NAFLD progression correlated with elevated hypoxia-induced factor 1␣ expression in the later stages of NAFLD. These results suggest that significant and novel changes occur in hepatic P450 activity during progressive stages of NAFLD.
ABSTRACT:Nonalcoholic fatty liver disease (NAFLD) is characterized by a series of pathological changes that range from simple fatty liver to nonalcoholic steatohepatitis (NASH). The objective of this study is to describe changes in global gene expression associated with the progression of human NAFLD. This study is focused on the expression levels of genes responsible for the absorption, distribution, metabolism, and elimination (ADME) of drugs. Differential gene expression between three clinically defined pathological groups-normal, steatosis, and NASH-was analyzed. Genome-wide mRNA levels in samples of human liver tissue were assayed with Affymetrix GeneChip Human 1.0ST arrays. A total of 11,633 genes exhibited altered expression out of 33,252 genes at a 5% false discovery rate. Most gene expression changes occurred in the progression from steatosis to NASH. Principal component analysis revealed that hepatic disease status was the major determinant of differential ADME gene expression rather than age or sex of sample donors. Among the 515 drug transporters and 258 drug-metabolizing enzymes (DMEs) examined, uptake transporters but not efflux transporters or DMEs were significantly over-represented in the number of genes downregulated. These results suggest that uptake transporter genes are coordinately targeted for down-regulation at the global level during the pathological development of NASH and that these patients may have decreased drug uptake capacity. This coordinated regulation of uptake transporter genes is indicative of a hepatoprotective mechanism acting to prevent accumulation of toxic intermediates in disease-compromised hepatocytes.
Farnesoid X receptor (FXR) is a master regulator of bile acid homeostasis through transcriptional regulation of genes involved in bile acid synthesis and cellular membrane transport. Impairment of bile acid efflux due to cholangiopathies results in chronic cholestasis leading to abnormal elevation of intrahepatic and systemic bile acid levels. Obeticholic acid (OCA) is a potent and selective FXR agonist that is 100‐fold more potent than the endogenous ligand chenodeoxycholic acid (CDCA). The effects of OCA on genes involved in bile acid homeostasis were investigated using sandwich‐cultured human hepatocytes. Gene expression was determined by measuring mRNA levels. OCA dose‐dependently increased fibroblast growth factor‐19 (FGF‐19) and small heterodimer partner (SHP) which, in turn, suppress mRNA levels of cholesterol 7‐alpha‐hydroxylase (CYP7A1), the rate‐limiting enzyme for de novo synthesis of bile acids. Consistent with CYP7A1 suppression, total bile acid content was decreased by OCA (1 μmol/L) to 42.7 ± 20.5% relative to control. In addition to suppressing de novo bile acids synthesis, OCA significantly increased the mRNA levels of transporters involved in bile acid homeostasis. The bile salt excretory pump (BSEP), a canalicular efflux transporter, increased by 6.4 ± 0.8‐fold, and the basolateral efflux heterodimer transporters, organic solute transporter α (OST α) and OST β increased by 6.4 ± 0.2‐fold and 42.9 ± 7.9‐fold, respectively. The upregulation of BSEP and OST α and OST β, by OCA reduced the intracellular concentrations of d8‐TCA, a model bile acid, to 39.6 ± 8.9% relative to control. These data demonstrate that OCA does suppress bile acid synthesis and reduce hepatocellular bile acid levels, supporting the use of OCA to treat bile acid‐induced toxicity observed in cholestatic diseases.
Non-alcoholic steatohepatitis (NASH) is a disease that compromises hepatic function and the capacity to metabolize numerous drugs. Aryl hydrocarbon receptor (AhR), constitutive androstane receptor (CAR), pregnane X receptor (PXR), peroxisome proliferator-activated receptor alpha (PPARalpha), and nuclear factor-E2 related factor 2 (Nrf2) are xenobiotic activated transcription factors that regulate induction of a number of drug metabolizing enzymes (DMEs). The purpose of the current study was to determine whether experimental NASH alters the xenobiotic activation of these transcription factors and induction of downstream DME targets Cyp1A1, Cyp2B10, Cyp3A11, Cyp4A14 and NAD(P)H:quinone oxidoreductase 1 (Nqo1), respectively. Mice fed normal rodent chow or methionine-choline-deficient (MCD) diet for 8 weeks were then treated with microsomal enzyme inducers beta-naphoflavone (BNF), 1,4-bis-[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP), pregnenolone-16alpha-carbonitrile (PCN), clofibrate (CFB) or oltipraz (OPZ), known activators of AhR, CAR, PXR, PPARalpha and Nrf2, respectively. Results of this study show that (1) Hepatic PXR mRNA levels were significantly increased (1.4-fold) in mice fed MCD diet, while AhR, CAR, PPARalpha and Nrf2 were not affected. (2) The MCD diet did not alter hepatic inducibility of Cyp1A1, Cyp2B10, Cyp3A11 mRNA levels by their respective microsomal inducers. (3) Constitutive levels of Cyp4A14 mRNA were significantly increased in mice fed the MCD diet, yet further induction by clofibrate was not observed. (4) Hepatic Nqo1 mRNA levels were significantly increased by the MCD diet; however, additional induction of Nqo1 was still achievable following treatment with the Nrf2 activator OPZ.
Cytochrome P450 monoxygenases (P450) are responsible for the metabolism and detoxification of many therapeutics. Induction of P450 enzymes by drugs can lead to potentially harmful drug‐drug interactions and alterations in drug therapy efficacy. Drug induction of P450 enzymes has been shown to involve the constitutive androstane receptor (CAR) and the pregnane X receptor (PXR). Recently, we reported that the nuclear receptor CAR mediated phenytoin (DPH) induction of the murine Cyp2b10 and Cyp2c29 genes. Herein, we report the discovery of a second DPH inducible murine CYP2C, Cyp2c37. Quantitative RT‐PCR and immunoblot data demonstrate that hepatic CYP2C37 mRNA and protein amounts are increased by DPH. We have identified a putative CAR response element (CAR‐RE) at ‐2.8kb from the start of translation of the Cyp2c37 gene. Mutation of this CAR‐RE in Cyp2c37 luciferase promoter constructs demonstrated that it is necessary for mCAR constitutive transactivation and that induction is governed by a transcriptional regulation mechanism. Furthermore in CAR‐null mice, quantitative RT‐PCR results indicate that the induction of CYP2C37 mRNA by DPH is abolished. These results suggest that DPH induction of the Cyp2c37 gene, like the Cyp2b10 and Cyp2c29 genes, is regulated by mCAR. However, in CAR‐null mice induction of CYP3A11 mRNA is not abolished suggesting that both CAR and PXR can mediate DPH induction of P450 genes, depending on the gene promoter. This research was supported by the intramural division of the NIEHS.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.