Helal Sara A, El-Sherbeni Ahmed A, El-Kadi Ayman O S
Faculty of Pharmacy and Pharmaceutical Sciences, University of Alberta, Edmonton, AB, Canada.
Department of Biochemistry, Faculty of Pharmacy, Tanta University, Tanta, Egypt.
Front Pharmacol. 2024 Aug 12;15:1438567. doi: 10.3389/fphar.2024.1438567. eCollection 2024.
R/S enantiomers of 11-hydroxyeicosatertraenoic acid (11-HETE) are formed from arachidonic acid by enzymatic and non-enzymatic pathways. 11-HETE is predominately formed by the cytochrome P450 1B1 (CYP1B1). The role of CYP1B1 in the development of cardiovascular diseases is well established.
This study aimed to assess the cellular hypertrophic effect of 11-HETE enantiomers in human RL-14 cardiomyocyte cell line and to examine their association with CYP1B1 levels.
Human fetal ventricular cardiomyocyte, RL-14 cells, were treated with 20 µM (R) or (S) 11-HETE for 24 h. Thereafter, cellular hypertrophic markers and cell size were then determined using real-time polymerase chain reaction (RT-PCR) and phase-contrast imaging, respectively. The mRNA and protein levels of selected CYPs were determined using RT-PCR and Western blot, respectively. In addition, we examined the effect of (R) and (S) 11-HETE on CYP1B1 catalytic activity using human recombinant CYP1B1 and human liver microsomes.
Both (R) and (S) 11-HETE induced cellular hypertrophic markers and cell surface area in RL-14 cells. Both enantiomers significantly upregulated CYP1B1, CYP1A1, CYP4F2, and CYP4A11 at the mRNA and protein levels, however, the effect of the S-enantiomer was more pronounced. Furthermore, 11(S)-HETE increased the mRNA and protein levels of CYP2J and CYP4F2, whereas 11(R)-HETE increased only CYP4F2. Only 11(S)-HETE significantly increased the catalytic activity of CYP1B1 in recombinant human CYP1B1, suggesting allosteric activation in an enantioselective manner.
Our study provides the first evidence that 11-HETE can induce cellular hypertrophy in RL-14 cells via the increase in CYP1B1 mRNA, protein, and activity levels.
11-羟基二十碳四烯酸（11-HETE）的R/S对映体通过酶促和非酶促途径由花生四烯酸形成。11-HETE主要由细胞色素P450 1B1（CYP1B1）形成。CYP1B1在心血管疾病发展中的作用已得到充分证实。
本研究旨在评估11-HETE对映体在人RL-14心肌细胞系中的细胞肥大效应，并研究它们与CYP1B1水平的关系。
用20µM（R）或（S）11-HETE处理人胎儿心室心肌细胞RL-14细胞24小时。此后，分别使用实时聚合酶链反应（RT-PCR）和相差成像法测定细胞肥大标志物和细胞大小。分别使用RT-PCR和蛋白质印迹法测定所选细胞色素P450（CYPs）的mRNA和蛋白质水平。此外，我们使用人重组CYP1B1和人肝微粒体检测了（R）和（S）11-HETE对CYP1B1催化活性的影响。
（R）和（S）11-HETE均诱导RL-14细胞中的细胞肥大标志物和细胞表面积增加。两种对映体均在mRNA和蛋白质水平上显著上调CYP1B1、CYP1A1、CYP4F2和CYP4A11，然而，S-对映体的作用更明显。此外，11（S）-HETE增加了CYP2J和CYP4F2的mRNA和蛋白质水平，而11（R）-HETE仅增加了CYP4F2。只有11（S）-HETE显著增加了重组人CYP1B1中CYP1B1的催化活性，表明其以对映体选择性方式进行变构激活。
我们的研究提供了首个证据，即11-HETE可通过增加CYP1B1的mRNA、蛋白质和活性水平诱导RL-14细胞肥大。
