Yang Bicheng, Huang Yanrui, Zhang Haifeng, Huang Yan, Zhou Huanjiao Jenny, Young Lawrence, Xiao Haipeng, Min Wang
The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong 510080, China; The Vascular Biology and Therapeutics Program, Yale University School of Medicine, New Haven, CT 06519, USA.
The Vascular Biology and Therapeutics Program, Yale University School of Medicine, New Haven, CT 06519, USA.
J Mol Cell Cardiol. 2020 Jan;138:291-303. doi: 10.1016/j.yjmcc.2019.10.009. Epub 2019 Nov 18.
Sick sinus syndrome (SSS) is associated with loss of HCN4 (hyperpolarization-activated cyclic nucleotide-gated potassium channel 4) function in the cardiac conduction system. The underlying mechanism for SSS remains elusive. This study is to investigate how mitochondrial oxidative stress induces HCN4 downregulation associated with in sick sinus syndrome.
Trx2 mice were crossed with α-myosin heavy chain (α-Mhc)-Cre and Hcn4-CreER deleter mice to generate Trx2 deletion mice in the whole heart (Trx2cKO) and in the conduction system (Trx2ccsKO), respectively. Echocardiography was applied to measure hemodynamics and heart rhythm. Histological analyses, gene profiling and chromatin immunoprecipitation were performed to define the mechanism by which thioredoxin-2 (Trx2) regulates HCN4 expression and cardiac function. Trx2cKO mice displayed dilated cardiomyopathy, low heart rate, and atrial ventricular block (AVB) phenotypes. Immunofluorescence revealed that HCN4 expression was specifically reduced within the sinoatrial node in Trx2cKO mice. Interestingly, Trx2ccsKO mice displayed low heart rate and AVB without dilated cardiomyopathy. Both mRNA and protein levels of HCN4 were reduced in the sinoatrial node, suggesting transcriptional HCN4 regulation upon Trx2 deletion. ChIP indicated that the binding of MEF2 to the HCN4 enhancer was not altered by Trx2 deletion; however, histone 3 acetylation at the MEF2 binding site was decreased, and expression of histone deacetylase 4 (HDAC4) was elevated following Trx2 deletion. Moreover, HDAC4 binding to the HCN4 enhancer was mediated by MEF2. Mitochondrial ROS were increased by Trx2 deletion and importantly, mitochondria-specific ROS scavenger MitoTEMPO suppressed HDAC4 elevation, HCN4 reduction, and sinus bradycardia in Trx2ccsKO mice.
In the conduction system, Trx2 is critical for maintaining HCN4-mediated normal heart rate. Loss of Trx2 reduces HCN4 expression via a mitochondrial ROS-HDAC4-MEF2C pathway and subsequently induces sick sinus syndrome in mice.
病态窦房结综合征（SSS）与心脏传导系统中HCN4（超极化激活的环核苷酸门控钾通道4）功能丧失有关。SSS的潜在机制仍不清楚。本研究旨在探讨线粒体氧化应激如何诱导与病态窦房结综合征相关的HCN4下调。
将Trx2小鼠与α-肌球蛋白重链（α-Mhc）-Cre和Hcn4-CreER敲除小鼠杂交，分别在全心（Trx2cKO）和传导系统（Trx2ccsKO）中产生Trx2缺失小鼠。应用超声心动图测量血流动力学和心律。进行组织学分析、基因谱分析和染色质免疫沉淀，以确定硫氧还蛋白-2（Trx2）调节HCN4表达和心脏功能的机制。Trx2cKO小鼠表现出扩张型心肌病、心率降低和房室传导阻滞（AVB）表型。免疫荧光显示，Trx2cKO小鼠窦房结内HCN4表达特异性降低。有趣的是，Trx2ccsKO小鼠表现出心率降低和AVB，但无扩张型心肌病。窦房结中HCN4的mRNA和蛋白水平均降低，提示Trx2缺失后HCN4的转录调控。染色质免疫沉淀表明，Trx2缺失不改变MEF2与HCN4增强子的结合；然而，Trx2缺失后，MEF2结合位点处的组蛋白3乙酰化降低，组蛋白去乙酰化酶4（HDAC4）的表达升高。此外，HDAC4与HCN4增强子的结合由MEF2介导。Trx2缺失增加了线粒体活性氧，重要的是，线粒体特异性活性氧清除剂MitoTEMPO抑制了Trx2ccsKO小鼠中HDAC4升高、HCN4降低和窦性心动过缓。
在传导系统中，Trx2对维持HCN4介导的正常心率至关重要。Trx2缺失通过线粒体活性氧-HDAC4-MEF2C途径降低HCN4表达，随后在小鼠中诱导病态窦房结综合征。
