雌二醇通过膜结合雌激素受体/磷酸肌醇 3-激酶/蛋白激酶 B/环磷腺苷反应元件结合蛋白信号通路上调 L 型钙通道。

Estradiol up-regulates L-type Ca channels via membrane-bound estrogen receptor/phosphoinositide-3-kinase/Akt/cAMP response element-binding protein signaling pathway.

机构信息

Hubei Key Laboratory of Drug Target Research and Pharmacodynamic Evaluation, Department of Pharmacology, School of Basic Medicine, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei, China.

Falk Library of Health Sciences, University of Pittsburgh, Pittsburgh, Pennsylvania.

出版信息

Heart Rhythm. 2018 May;15(5):741-749. doi: 10.1016/j.hrthm.2018.01.019. Epub 2018 Jan 9.

DOI:10.1016/j.hrthm.2018.01.019

PMID:29330129

Abstract

BACKGROUND

In long QT syndrome type 2, women are more prone than men to the lethal arrhythmia torsades de pointes. We previously reported that 17β-estradiol (E2) up-regulates L-type Ca channels and current (I) (∼30%) in rabbit ventricular myocytes by a classic genomic mechanism mediated by estrogen receptor-α (ERα). In long QT syndrome type 2 (I blockade or bradycardia), the higher Ca influx via I causes Ca overload, spontaneous sarcoplasmic reticulum Ca release, and reactivation of I that triggers early afterdepolarizations and torsades de pointes.

OBJECTIVE

The purpose of this study was to investigate the molecular mechanisms whereby E2 up-regulates I, which are poorly understood.

METHODS

H9C2 and rat myocytes were incubated with E2 ± ER antagonist, or inhibitors of downstream transcription factors, for 24 hours, followed by western blots of Cav1.2α1C and voltage-clamp measurements of I.

RESULTS

Incubation of H9C2 cells with E2 (10-100 nM) increased I density and Cav1.2α1C expression, which were suppressed by the ER antagonist ICI182,780 (1 μM). Enhanced I and Cav1.2α1C expression by E2 was suppressed by inhibitors of phosphoinositide-3-kinase (Pi3K) (30 μM LY294002; P <.05) and Akt (5 μM MK2206) but not of mitogen-activated protein kinase (5 μM U0126) or protein kinase A (1 μM KT5720). E2 incubation increased p-CREB via the Pi3K/Akt pathway, reached a peak in 20 minutes (3-fold), and leveled off to 1.5-fold 24 hours later. Furthermore, a CREB decoy oligonucleotide inhibited E2-induced Cav1.2α1C expression, whereas membrane-impermeable E2 (E2-bovine serum albumin) was equally effective at Cav1.2α1C up-regulation as E2.

CONCLUSION

Estradiol up-regulates Cav1.2α1C and I via plasma membrane ER and by activating Pi3K, Akt, and CREB signaling. The promoter regions of the CACNA1C gene (human-rabbit-rat) contain adjacent/overlapping binding sites for p-CREB and ERα, which suggests a synergistic regulation by these pathways.

摘要

背景

在长 QT 综合征 2 型中，女性比男性更容易发生致命性心律失常尖端扭转型室性心动过速。我们之前报道过，17β-雌二醇（E2）通过经典的雌激素受体-α（ERα）介导的基因组机制上调兔心室肌细胞中的 L 型钙通道和电流（I）（约 30%）。在长 QT 综合征 2 型（I 阻滞或心动过缓）中，通过 I 的更高钙内流导致钙超载、自发性肌浆网钙释放和 I 的再激活，从而引发早期后除极和尖端扭转型室性心动过速。

目的

本研究旨在探讨 E2 上调 I 的分子机制，目前对此知之甚少。

方法

用 E2 ± ER 拮抗剂或下游转录因子抑制剂孵育 H9C2 和大鼠心肌细胞 24 小时，然后进行 Cav1.2α1C 的 Western blot 和 I 的电压钳测量。

结果

用 10-100 nM 的 E2 孵育 H9C2 细胞可增加 I 密度和 Cav1.2α1C 的表达，这一作用被 ER 拮抗剂 ICI182,780（1 μM）抑制。E2 增强 I 和 Cav1.2α1C 的表达被 PI3K（30 μM LY294002；P <.05）和 Akt（5 μM MK2206）抑制剂抑制，但不被丝裂原激活蛋白激酶（5 μM U0126）或蛋白激酶 A（1 μM KT5720）抑制剂抑制。E2 孵育通过 PI3K/Akt 通路增加 p-CREB，在 20 分钟时达到峰值（增加 3 倍），24 小时后达到 1.5 倍。此外，CREB 封闭寡核苷酸抑制了 E2 诱导的 Cav1.2α1C 表达，而不能透过细胞膜的 E2（E2-牛血清白蛋白）在 Cav1.2α1C 上调方面与 E2 同样有效。