Kao Yu-Hsun, Hsu Jung-Chieh, Chen Yao-Chang, Lin Yung-Kuo, Lkhagva Baigalmaa, Chen Shih-Ann, Chen Yi-Jen
Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan; Department of Medical Education and Research, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan.
Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan.
Int J Cardiol. 2016 May 1;210:85-92. doi: 10.1016/j.ijcard.2016.02.091. Epub 2016 Feb 17.
ZFHX3 plays an important role in the genesis of atrial fibrillation. However, the atrial electrophysiological effects of ZFHX3 are not clear. This study sought to investigate roles of ZFHX3 in atrial electrophysiology and calcium homeostasis by using HL-1 atrial myocytes knocked-down with ZFHX3.
Patch clamp, confocal fluorescence microscopy and Western blot were used to study electrical activity, ionic currents, calcium homeostasis and protein expressions in stable ZFHX3 shRNA cells.
As compared to control, ZFHX3 shRNA cells with 28% decline of ZFHX3 protein had a larger sarcoplasmic reticulum Ca(2+) content by 62%, Ca(2+) transient by 20%, and calcium leak by 75%. ZFHX3 shRNA cells (n=35) had shorter action potential duration (APD) at 50% (14.7 ± 0.9 versus 20.3 ± 1.4 ms, P<0.005), and 20% (6.1 ± 0.3 versus 8.3 ± 0.8 ms, P<0.005) repolarization than control cells (n=30). ZFHX3 shRNA cells (n=10) had larger amplitudes of isoproterenol (1 μM)-induced delayed after depolarization (14.1 ± 0.9 versus 7.2 ± 0.2 mV, P<0.05) than control cells (n=10). Besides, acetylcholine (3 μM) shortened APD at 90% repolarization to a greater extent (19 ± 4% versus 7 ± 2%, P<0.01) in ZFHX3 shRNA cells (n=11) than in control cells (n=12). In addition, ZFHX3 shRNA cells had increased expressions of SERCA2a, ryanodine receptor, Kv1.4, Kv1.5 and Kir3.4. Moreover, ZFHX3 shRNA cells had a larger SERCA2a activity, ultra-rapid delayed rectifier potassium currents, transient outward currents and acetylcholine-sensitive potassium currents.
ZFHX3 knock-down in atrial myocytes dysregulated calcium homeostasis and increased atrial arrhythmogenesis, which may contribute to the occurrence of AF.
锌指同源盒蛋白3（ZFHX3）在房颤的发生中起重要作用。然而，ZFHX3对心房电生理的影响尚不清楚。本研究旨在通过使用ZFHX3基因敲低的HL-1心房肌细胞来研究ZFHX3在心房电生理和钙稳态中的作用。
采用膜片钳、共聚焦荧光显微镜和蛋白质印迹法研究稳定转染ZFHX3短发夹RNA（shRNA）细胞的电活动、离子电流、钙稳态和蛋白质表达。
与对照组相比，ZFHX3蛋白下降28%的ZFHX3 shRNA细胞肌浆网Ca²⁺含量增加62%，Ca²⁺瞬变增加20%，钙泄漏增加75%。ZFHX3 shRNA细胞（n = 35）在复极化50%时动作电位时程（APD）较短（14.7±0.9对20.3±1.4毫秒，P<0.005），在复极化20%时也较短（6.1±0.3对8.3±0.8毫秒，P<0.005），而对照组细胞（n = 30）较长。ZFHX3 shRNA细胞（n = 10）异丙肾上腺素（1μM）诱导的延迟后除极幅度（14.1±0.9对7.2±0.2毫伏，P<0.05）大于对照组细胞（n = 10）。此外，乙酰胆碱（3μM）使ZFHX3 shRNA细胞（n = 11）在复极化90%时APD缩短的程度更大（19±4%对7±2%，P<0.01），大于对照组细胞（n = 12）。此外，ZFHX3 shRNA细胞中肌浆网Ca²⁺-ATP酶2a（SERCA2a）、兰尼碱受体、电压门控钾通道1.4（Kv1.4）、电压门控钾通道1.5（Kv1.5）和内向整流钾通道3.4（Kir3.4）的表达增加。而且，ZFHX3 shRNA细胞具有更大的SERCA2a活性、超快速延迟整流钾电流、瞬时外向电流和乙酰胆碱敏感性钾电流。
心房肌细胞中ZFHX3基因敲低导致钙稳态失调并增加心房心律失常的发生，这可能有助于房颤的发生。
