Elsevier

Biochemical Pharmacology

Volume 93, Issue 1, 1 January 2015, Pages 92-103
Biochemical Pharmacology

Sex-dependent regulation of hepatic CYP3A by growth hormone: Roles of HNF6, C/EBPα, and RXRα

https://doi.org/10.1016/j.bcp.2014.10.010Get rights and content

Abstract

Sex-based differences in the pharmacological profiles of many drugs are due in part to the female-predominant expression of CYP3A4, which is the most important CYP isoform responsible for drug metabolism. Transcription factors trigger the sexually dimorphic expression of drug-metabolizing enzymes in response to sex-dependent growth hormone (GH) secretion. We investigated the roles of HNF6, C/EBPα, and RXRα in the regulation of human female-predominant CYP3A4, mouse female-specific CYP3A41, and rat male-specific CYP3A2 expression by GH secretion patterns using HepG2 cells, growth hormone receptor (GHR) knockout mice as well as rat models of orchiectomy and hypophysectomy. The constitutive expression of HNF6 and RXRα was GH-dependent, and GHR deficiency decreased HNF6/C/EBPα complex levels and increased HNF6/RXRα complex levels. Feminine GH secretion induced the binding of HNF6 and C/EBPα to the CYP3A4 and Cyp3a41 promoters and HNF6/C/EBPα complex levels was more efficiently compared with masculine pattern. Additionally, a greater inhibition of the binding of RXRα to the CYP3A4 and Cyp3a41 promoters and HNF6/RXRα complex levels was observed by feminine GH secretion, but less inhibition was observed by masculine pattern. The binding of HNF6, C/EBPα, and RXRα to the CYP3A2 promoter was not directly regulated by androgens. RXRα completely abolished the synergistic activation of the CYP3A4, Cyp3a41, and CYP3A2 promoters by HNF6 and C/EBPα. The results demonstrate that sex-dependent GH secretion patterns affect the expressions and interactions of HNF6, C/EBPα, and RXRα as well as their binding to CYP3A genes. RXRα mediates the sex-dependent influence of GH on CYP3A expression as an important signalling molecule.

Introduction

The sexually dimorphic expression of drug-metabolizing enzymes is one of the primary causes of sex-based differences in the pharmacological profiles of many drugs and the prevalence of liver diseases. The cytochrome P450 (CYP) superfamily is principally responsible for the biotransformation of clinical drugs, toxins, and endogenous compounds [1]. The more rapid clearance of different drugs in women compared with men has been reported, which is due in part to the female-predominant expression of CYP3A4, the most important CYP isoform responsible for drug metabolism [2], [3], [4]. CYP3A4 levels in women vary from approximately 25 to 200% above the observed levels in men [5], [6], [7], [8].

Sexually dimorphic growth hormone (GH) secretion is considered to be a major factor in establishing and maintaining sexual dimorphism in hepatic gene transcription, including that of CYP isoforms [3], [9], [10], [11]. The characteristic GH secretory profiles of all mammals include constant secretion in females and pulsatile secretion in males [3], [10], [12]. A clinical study revealed that continuous GH administration induces CYP3A4 activity in humans [13]. Both CYP3A4 protein and mRNA levels can be increased by the constant GH treatment in primary human hepatocytes but are suppressed by the pulsatile GH treatment [12], [14].

GH signalling is initiated by the binding of GH to the extracellular domain of the growth hormone receptor (GHR), which recruits and/or activates multiple intracellular signalling molecules [3], [15]. Hepatocyte nuclear factors (HNFs) and CCAAT/enhancer-binding protein (C/EBP) α have been considered to be the key transcription factors that collaborate with intracellular signalling molecules to mediate the sex-dependent effects of GH on liver gene expression [16]. Although HNF6 and C/EBPα have been reported to be involved in the regulation of CYP3A4 expression [17], [18], the regulatory mechanism controlled by GH secretion is still largely unknown. Retinoid X receptor (RXR) α is an obligate partner for multiple nuclear receptors. RXRα expression was not found to be subject to sexual dimorphism based on microarray-derived data from mouse livers [19]; however, RXRα binding to chromatin was shown to be enriched in 1945 genes and 1214 genes in male and female mouse livers, respectively, including hepatic gender-specific genes [20].

Hypophysectomy and growth hormone replacement indicated that the mRNA level of mouse female-specific CYP3A41 was dependent on the feminine growth hormone profile [21]. However, the rat male-specific CYP3A2 has been shown to be down-regulated by the replacement of the pulsatile GH secretion in the male hypophysectomized rats [22], [23]. We investigated the roles of HNF6, C/EBPα, and RXRα in the regulation of human female-predominant CYP3A4, mouse female-specific CYP3A41, and rat male-specific CYP3A2 expression in response to GH secretion patterns. This work adds to current knowledge of the sex-associated roles of individual transcription factors in the regulation of liver gene expression.

Section snippets

Animals and treatment

Adult GHR knockout mice [24] and their wild-type counterparts (C57/Black) were kindly provided by Professor Michael J. Waters and were bred at the Centre for Animal Experiments Laboratory of Wuhan University. The mice were genotyped using DNA extracted from their tails. Adult male Wistar rats (250–300 g, Certificate No. SCXK2008-0005) were supplied by the Experimental Animal Centre (Hubei, China). The rats were subjected to one of the following operational procedures: bilateral orchiectomy,

Effects of GH secretion patterns on CYP3A4, HNF6, RXRα, and C/EBPαlevels in HepG2 cells

CYP3A4 mRNA and protein levels increased following the constant (feminine) GH administration but not the pulsatile (masculine) GH administration (Fig. 1A, E). The constant GH secretion at a physiological concentration (2 ng/mL) increased HNF6 mRNA and protein levels more efficiently compared with the pulsatile GH secretion (Fig. 1B, F). In contrast, the pulsatile GH secretion induced RXRα mRNA and protein levels more efficiently compared with the constant GH secretion (Fig. 1C, G). Both the

Discussion

This study demonstrated the involvement of HNF6, RXRα, and C/EBPα in the regulation of human female-predominant CYP3A4, rat male-specific CYP3A2, and mouse female-specific CYP3A41 levels by sexually dimorphic GH secretion. We showed that (i) constitutive expression of HNF6 and RXRα was GH-dependent; (ii) feminine GH secretion induced the binding of HNF6 and C/EBPα to the CYP3A4 and Cyp3a41 promoters and HNF6/C/EBPα complex levels was more efficiently compared with masculine pattern; (iii) the

Conflict of interest

The authors have no conflicts of interest.

Acknowledgments

This study was supported by the Program for New Century Excellent Talents in University (NCET-11-0399) and the Natural Science Foundation of China (No. 81173122, 30973582, and 30960334). We thank Professor Michael J. Waters (Institute for Molecular Bioscience, University of Queensland, Australia) for providing GHR-deficient mice and the members of the Animal Care Committee of Wuhan University for their helpfulness and assistance with animal care.

References (39)

  • J. Yue et al.

    Protective effects of thiopronin against isoniazid-induced hepatotoxicity in rats

    Toxicology

    (2009)
  • Y. Bing et al.

    Reduction of thyroid hormones triggers down-regulation of hepatic CYP2B through nuclear receptors CAR and TR in a rat model of acute stroke

    Biochem Pharmacol

    (2014)
  • V. Ribeiro et al.

    Cloning and characterization of a novel CYP3A1 allelic variant: analysis of CYP3A1 and CYP3A2 sex-hormone-dependent expression reveals that the CYP3A2 gene is regulated by testosterone

    Arch Biochem Biophys

    (1992)
  • C.H. Johnson et al.

    Xenobiotic metabolomics: major impact on the metabolome

    Annu Rev Pharmacol Toxicol

    (2012)
  • M. Chen et al.

    Sex differences in CYP3A activity using intravenous and oral midazolam

    Clin Pharmacol Ther

    (2006)
  • D.J. Waxman et al.

    Growth hormone regulation of sex-dependent liver gene expression

    Mol Endocrinol

    (2006)
  • M.M. Cotreau et al.

    The influence of age and sex on the clearance of cytochrome P450 3A substrates

    Clin Pharmacokinet

    (2005)
  • M. Schirmer et al.

    Sex-dependent genetic markers of CYP3A4 expression and activity in human liver microsomes

    Pharmacogenomics

    (2007)
  • V. Wauthier et al.

    Intrinsic sex differences in the early growth hormone responsiveness of sex-specific genes in mouse liver

    Mol Endocrinol

    (2010)
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