Non-Clinical Drug Safety, Boehringer Ingelheim Pharmaceuticals, Ridgefield, Connecticut (P.P., W.W.K.) and Department of Pharmaceutical Sciences, School of Pharmacy, University of Connecticut, Storrs, Connecticut (P.P., X.-b.Z.).
Non-Clinical Drug Safety, Boehringer Ingelheim Pharmaceuticals, Ridgefield, Connecticut (P.P., W.W.K.) and Department of Pharmaceutical Sciences, School of Pharmacy, University of Connecticut, Storrs, Connecticut (P.P., X.-b.Z.)
Drug Metab Dispos. 2020 Dec;48(12):1321-1329. doi: 10.1124/dmd.120.000195. Epub 2020 Oct 19.
Obesity and nonalcoholic fatty liver disease (NAFLD) affect expression and function of cytochrome P450 genes (P450s). The increased expression of inflammatory cytokines is a major driver of the downregulation of P450 expression in NAFLD. Decrease in P450 expression could potentially lead to drug-drug interaction, inefficient pharmacological effect of a drug, or hepatotoxicity. An epigenetic modifier, histone 3 lysine 9 methyl transferase enzyme (G9a), known to increase histone 3 lysine 9 methylation, is downregulated in diet-induced obesity animal models. In a liver-specific G9a knockout animal model, expression of P450s was downregulated. Currently, the role of G9a in regulation of P450s in steatosis is unknown. Our hypothesis is that in steatosis G9a plays a role in downregulation of P450 expression. In this study, we used HepaRG cells to induce steatosis using a combination of free fatty acids oleic acid and palmitic acid. The G9a was knocked down and overexpressed using small interfering RNA and adenovirus mediated approaches, respectively. Knockdown and overexpression of G9a in the absence of steatosis decreased and increased expression of nuclear receptors constitutive androstane receptor (CAR), pregnane X receptor, small heterodimer partner, and CYP2B6, 2E1, 2C8, 2C9, and 3A4, respectively. In steatotic conditions, overexpression of G9a prevented fatty acid mediated decreased expression of CAR, CYP2C19, 2C8, 7A1, and 3A4. Our current study suggests that G9a might serve as a key regulator of P450 expression at both the basal level and in early steatotic conditions. Single nucleotide polymorphism of G9a leading to loss/gain of function could lead to the poor metabolizer or ultrarapid metabolizer phenotypes. SIGNIFICANCE STATEMENT: The current study demonstrates that histone modification enzyme G9a is involved in the regulation of expression of nuclear receptors constitutive androstane receptor, pregnane X receptor, and small heterodimer partner as well as drug-metabolizing cytochrome P450s (P450s) at basal conditions and in fatty acid induced cellular model of steatosis. Histone 3 lysine 9 methylation should be considered together with histone 3 lysine 4 and histone 3 lysine 27 methylation as the epigenetic mechanisms controlling gene expression of P450s.
肥胖和非酒精性脂肪性肝病（NAFLD）影响细胞色素 P450 基因（P450s）的表达和功能。炎症细胞因子的增加是 P450 表达在 NAFLD 中下调的主要驱动因素。P450 表达的减少可能导致药物-药物相互作用、药物的药理作用效率降低或肝毒性。组蛋白 3 赖氨酸 9 甲基转移酶酶（G9a）是一种表观遗传修饰剂，已知其增加组蛋白 3 赖氨酸 9 的甲基化，在饮食诱导的肥胖动物模型中下调。在肝特异性 G9a 敲除动物模型中，P450 的表达下调。目前，G9a 在调节脂肪变性中的 P450 作用尚不清楚。我们的假设是，在脂肪变性中，G9a 在下调 P450 表达中起作用。在这项研究中，我们使用 HepaRG 细胞，通过使用游离脂肪酸油酸和棕榈酸的组合来诱导脂肪变性。使用小干扰 RNA 和腺病毒介导的方法分别敲低和过表达 G9a。在没有脂肪变性的情况下，G9a 的敲低和过表达分别降低和增加了核受体细胞色素 P450 诱导的雄激素受体（CAR）、孕烷 X 受体、小异二聚体伴侣和 CYP2B6、2E1、2C8、2C9 和 3A4 的表达。在脂肪变性条件下，G9a 的过表达可防止脂肪酸介导的 CAR、CYP2C19、2C8、7A1 和 3A4 表达降低。我们目前的研究表明，G9a 可能是基础水平和早期脂肪变性条件下 P450 表达的关键调节剂。G9a 的单核苷酸多态性导致功能丧失/获得，可能导致代谢不良或超快代谢表型。意义陈述：本研究表明，组蛋白修饰酶 G9a 参与调节核受体细胞色素 P450 诱导的雄激素受体、孕烷 X 受体和小异二聚体伴侣以及药物代谢细胞色素 P450（P450s）在基础条件下和脂肪酸诱导的脂肪变性细胞模型中的表达。组蛋白 3 赖氨酸 9 甲基化应与组蛋白 3 赖氨酸 4 和组蛋白 3 赖氨酸 27 甲基化一起考虑，作为控制 P450 基因表达的表观遗传机制。
