Department of Cardiology, First AffiliatedHospital of Wenzhou Medical University, Wenzhou 325100, China.
World J Emerg Med. 2014;5(2):122-7. doi: 10.5847/wjem.j.issn.1920-8642.2014.02.008.
Retinoid X receptor (RXR) plays a central role in the regulation of intracellular receptor signaling pathways. The activation of RXR has protective effect on H2O2-induced apoptosis of H9c2 ventricular cells in rats. But the protective effect and mechanism of activating RXR in cardiomyocytes against hypoxia/reoxygenation (H/R)-induced oxidative iniury are still unclear.
The model of H/R injury was established through hypoxia for 2 hours and reoxygenation for 4 hours in H9c2 cardiomyocytes of rats. 9-cis-retinoic acid (9-cis RA) was obtained as an RXR agonist, and HX531 as an RXR antagonist. Cultured cardiomyocytes were randomly divided into four groups: sham group, H/R group, H/R+9-cis RA -pretreated group (100 nmol/L 9-cis RA), and H/R+9-cis RA+HX531-pretreated group (2.5 μmol/L HX531). The cell viability was measured by MTT, apoptosis rate of cardiomyocytes by flow cytometry analysis, and mitochondrial membrane potential (ΔΨm) by JC-1 fluorescent probe, and protein expressions of Bcl-2, Bax and cleaved caspase-9 with Western blotting. All measurement data were expressed as mean±standard deviation, and analyzed using one-way ANOVA and the Dunnett test. Differences were considered significant when P was <0.05.
Pretreatment with RXR agonist enhanced cell viability, reduced apoptosis ratio, and stabled ΔΨm. Dot blotting experiments showed that under H/R stress conditions, Bcl-2 protein level decreased, while Bax and cleaved caspase-9 were increased. 9-cis RA administration before H/R stress prevented these effects, but the protective effects of activating RXR on cardiomyocytes against H/R induced oxidative injury were abolished when pretreated with RXR pan-antagonist HX531.
The activation of RXR has protective effects against H/R injury in H9c2 cardiomyocytes of rats through attenuating signaling pathway of mitochondria apoptosis.
视黄酸 X 受体 (RXR) 在调节细胞内受体信号通路中发挥核心作用。RXR 的激活对大鼠 H9c2 心室细胞中 H2O2 诱导的凋亡具有保护作用。但是，激活 RXR 对心肌细胞缺氧/复氧（H/R）诱导的氧化损伤的保护作用及其机制尚不清楚。
通过在大鼠 H9c2 心肌细胞中缺氧 2 小时和复氧 4 小时建立 H/R 损伤模型。9-顺式视黄酸（9-cis RA）作为 RXR 激动剂，HX531 作为 RXR 拮抗剂。将培养的心肌细胞随机分为四组：假手术组、H/R 组、H/R+9-cis RA 预处理组（100 nmol/L 9-cis RA）和 H/R+9-cis RA+HX531 预处理组（2.5 μmol/L HX531）。通过 MTT 测量细胞活力，通过流式细胞术分析心肌细胞凋亡率，通过 JC-1 荧光探针测量线粒体膜电位（ΔΨm），并用 Western blot 测量 Bcl-2、Bax 和 cleaved caspase-9 的蛋白表达。所有测量数据均表示为均数±标准差，采用单因素方差分析和 Dunnett 检验进行分析。当 P<0.05 时，认为差异具有统计学意义。
RXR 激动剂预处理可增强细胞活力、降低凋亡率和稳定 ΔΨm。斑点印迹实验显示，在 H/R 应激条件下，Bcl-2 蛋白水平降低，而 Bax 和 cleaved caspase-9 增加。H/R 应激前给予 9-cis RA 预处理可防止这些作用，但用 RXR 泛拮抗剂 HX531 预处理时，激活 RXR 对心肌细胞对抗 H/R 诱导的氧化损伤的保护作用被消除。
通过减弱线粒体凋亡信号通路，激活 RXR 对大鼠 H9c2 心肌细胞的 H/R 损伤具有保护作用。
