Department of Pharmacology and Toxicology, Faculty of Pharmacy in Hradec Kralove, Charles University, Akademika Heyrovskeho 1203, Hradec Kralove, 500 05, Czech Republic.
Department of Biophysics and Physical Chemistry, Faculty of Pharmacy in Hradec Kralove, Charles University, Akademika Heyrovskeho 1203, Hradec Kralove, 500 05, Czech Republic.
Arch Toxicol. 2022 Jan;96(1):195-210. doi: 10.1007/s00204-021-03177-y. Epub 2021 Oct 23.
The pregnane X receptor (PXR) is a ligand-activated nuclear receptor controlling hepatocyte expression of numerous genes. Although expression changes in xenobiotic-metabolizing, lipogenic, gluconeogenic and bile acid synthetic genes have been described after PXR activation, the temporal dynamics of their expression is largely unknown. Recently, 3D spheroids of primary human hepatocytes (PHHs) have been characterized as the most phenotypically relevant hepatocyte model. We used 3D PHHs to assess time-dependent expression profiles of 12 prototypic PXR-controlled genes in the time course of 168 h of rifampicin treatment (1 or 10 µM). We observed a similar bell-shaped time-induction pattern for xenobiotic-handling genes (CYP3A4, CYP2C9, CYP2B6, and MDR1). However, we observed either biphasic profiles for genes involved in endogenous metabolism (FASN, GLUT2, G6PC, PCK1, and CYP7A1), a decrease for SHP or oscillation for PDK4 and PXR. The rifampicin concentration determined the expression profiles for some genes. Moreover, we calculated half-lives of CYP3A4 and CYP2C9 mRNA under induced or basal conditions and we used a mathematical model to describe PXR-mediated regulation of CYP3A4 expression employing 3D PHHs. The study shows the importance of long-term time-expression profiling of PXR target genes in phenotypically stable 3D PHHs and provides insight into PXR function in liver beyond our knowledge from conventional 2D in vitro models.
pregnane X 受体 (PXR) 是一种配体激活的核受体，控制着肝细胞中许多基因的表达。虽然在 PXR 激活后，已经描述了外源性代谢物、脂生成、糖异生和胆汁酸合成基因的表达变化，但它们的表达时间动态在很大程度上是未知的。最近，原代人肝细胞 (PHH) 的 3D 球体已被表征为最具表型相关性的肝细胞模型。我们使用 3D PHH 来评估 rifampicin 治疗 168 小时过程中 12 个典型 PXR 控制基因的时间依赖性表达谱（1 或 10 μM）。我们观察到外源性处理基因（CYP3A4、CYP2C9、CYP2B6 和 MDR1）的诱导时间呈相似的钟形时间诱导模式。然而，我们观察到内源性代谢基因（FASN、GLUT2、G6PC、PCK1 和 CYP7A1）的双相谱，SHP 减少，PDK4 和 PXR 振荡。利福平浓度决定了一些基因的表达谱。此外，我们计算了诱导或基础条件下 CYP3A4 和 CYP2C9 mRNA 的半衰期，并使用数学模型来描述 3D PHH 中 PXR 介导的 CYP3A4 表达调控。该研究表明，在表型稳定的 3D PHH 中，对 PXR 靶基因进行长期时间表达谱分析的重要性，并提供了对 PXR 功能的深入了解，超出了我们从传统 2D 体外模型中获得的知识。
