人胎儿心室心肌细胞RL-14细胞系是研究药物代谢酶及其相关花生四烯酸代谢产物的一个有前景的模型。

Human fetal ventricular cardiomyocyte, RL-14 cell line, is a promising model to study drug metabolizing enzymes and their associated arachidonic acid metabolites.

作者信息

Maayah Zaid H, Elshenawy Osama H, Althurwi Hassan N, Abdelhamid Ghada, El-Kadi Ayman O S

机构信息

Faculty of Pharmacy & Pharmaceutical Sciences, University of Alberta, Edmonton, Canada.

出版信息

J Pharmacol Toxicol Methods. 2015 Jan-Feb;71:33-41. doi: 10.1016/j.vascn.2014.11.005. Epub 2014 Nov 29.

DOI:10.1016/j.vascn.2014.11.005

PMID:25454080

Abstract

INTRODUCTION

RL-14 cells, human fetal ventricular cardiomyocytes, are a commercially available cell line that has been established from non-proliferating primary cultures derived from human fetal heart tissue. However, the expression of different drug metabolizing enzymes (DMEs) in RL-14 cells has not been elucidated yet. Therefore, the main objectives of the current work were to investigate the capacity of RL-14 cells to express different cytochrome P450 (CYP) isoenzymes and correlate this expression to primary cardiomyocytes.

METHODS

The expression of CYP isoenzymes was determined at mRNA, protein and catalytic activity levels using real time-PCR, Western blot analysis and liquid chromatography-electron spray ionization-mass spectrometry (LC-ESI-MS), respectively.

RESULTS

Our results showed that RL-14 cells constitutively express CYP ω-hydroxylases, CYP1A, 1B, 4A and 4F; CYP epoxygenases, CYP2B, 2C and 2J; in addition to soluble epoxide hydrolayse (EPHX2) at mRNA and protein levels. The basal expression of CYP ω-hydroxylases, epoxygenases and EPHX2 was supported by the ability of RL-14 cells to convert arachidonic acid to its biologically active metabolites, 20-hydroxyeicosatetraenoic acids (20-HETEs), 14,15-epoxyeicosatrienoic acids (14,15-EET), 11,12-EET, 8,9-EET, 5,6-EET, 14,15-dihydroxyeicosatrienoic acid (14,15-DHET), 11,12-DHET, 8,9-DHET and 5,6-DHET. Furthermore, RL-14 cells express CYP epoxygenases and ω-hydroxylase at comparable levels to those expressed in adult and fetal human primary cardiomyocytes cells implying the importance of RL-14 cells as a model for studying DMEs in vitro. Lastly, different CYP families were induced in RL-14 cells using 2,3,7,8-tetrachlorodibenzo-p-dioxin and fenofibrate at mRNA and protein levels.

DISCUSSION

The current study provides the first evidence that RL-14 cells express CYP isoenzymes at comparable levels to those expressed in the primary cells and thus offers a unique in vitro model to study DMEs in the heart.

摘要

引言

RL-14细胞，即人胎儿心室心肌细胞，是一种可从人胎儿心脏组织的非增殖原代培养物中建立的商业化细胞系。然而，RL-14细胞中不同药物代谢酶（DME）的表达尚未阐明。因此，当前工作的主要目标是研究RL-14细胞表达不同细胞色素P450（CYP）同工酶的能力，并将这种表达与原代心肌细胞进行关联。

方法

分别使用实时定量PCR、蛋白质免疫印迹分析和液相色谱 - 电喷雾电离质谱（LC-ESI-MS）在mRNA、蛋白质和催化活性水平上测定CYP同工酶的表达。

结果

我们的结果表明，RL-14细胞组成性表达CYP ω-羟化酶、CYP1A、1B、4A和4F；CYP环氧化酶、CYP2B、2C和2J；此外，在mRNA和蛋白质水平上还表达可溶性环氧化物水解酶（EPHX2）。RL-14细胞将花生四烯酸转化为其生物活性代谢产物20-羟基二十碳四烯酸（20-HETEs）、14,15-环氧二十碳三烯酸（14,15-EET）、11,12-EET、8,9-EET、5,6-EET、14,15-二羟基二十碳三烯酸（14,15-DHET）、11,12-DHET、8,9-DHET和5,6-DHET的能力支持了CYP ω-羟化酶、环氧化酶和EPHX2的基础表达。此外，RL-14细胞中CYP环氧化酶和ω-羟化酶的表达水平与成人和胎儿人原代心肌细胞中的表达水平相当，这意味着RL-14细胞作为体外研究DMEs模型的重要性。最后，使用2,3,7,8-四氯二苯并 - 对二恶英和非诺贝特在mRNA和蛋白质水平上诱导RL-14细胞中不同的CYP家族。