He J H, Yang L, Xia G L, Deng N, Yang Y Y, Tian Y, Fu Z N, Huang Y Q
Guizhou Provincial People's Hospital, Guiyang 550002, China.
Zhonghua Yi Xue Za Zhi. 2018 Nov 6;98(41):3345-3349. doi: 10.3760/cma.j.issn.0376-2491.2018.41.011.
To investigate the effects of calcineurin gene silencing on the remodeling of transient outward potassium current (Ito) ionic channel and action potential duration (APD) in phenylephrine (PE)-induced hypertrophic ventricular myocytes from neonatal rats. The ventricular myocytes of 1-day-old Sprague-Dawley rats were isolated and cultured for 48 h. RNA interference mediated by adenovirus carrying short hairpin RNA was used to knock down the gene which encodes the beta subtype of calcineurin A subunit (CnAβ) and the cells were divided into 4 groups as Ad-null group, Ad-null+ PE group, Ad-CnAβshRNA1(A1) group and A1+ PE group, and then cultured for 48 h. The gene expression of Kv4.2 was assayed by real-time reverse transcriptase-polymerase chain reaction. The protein expressions of CnAβ and Kv4.2 were assayed by Western blot test. Whole cell patch clamp technique was used to record Ito and action potential. Treatment of the neonatal rat ventricular myocytes with PE induced the cell hypertrophy, up-regulated the protein expression of CnAβ, attenuated the gene and protein expressions of Kv4.2 and the Ito current density, and prolonged APD. Silencing of CnAβ in the neonatal rat ventricular myocytes using Ad-CnAβshRNA1 inhibited the aforementioned ability of PE significantly. CnAβ gene silencing inhibits the remodeling of transient outward potassium current ionic channel and change of APD in PE-induced hypertrophic ventricular myocytes from neonatal rats.
探讨钙调神经磷酸酶基因沉默对苯肾上腺素（PE）诱导的新生大鼠肥厚性心室肌细胞瞬时外向钾电流（Ito）离子通道重塑及动作电位时程（APD）的影响。分离1日龄Sprague-Dawley大鼠的心室肌细胞并培养48小时。利用携带短发夹RNA的腺病毒介导的RNA干扰来敲低编码钙调神经磷酸酶A亚基β亚型（CnAβ）的基因，将细胞分为4组，即Ad-空载体组、Ad-空载体+PE组、Ad-CnAβshRNA1（A1）组和A1+PE组，然后再培养48小时。通过实时逆转录聚合酶链反应检测Kv4.2的基因表达。通过蛋白质免疫印迹法检测CnAβ和Kv4.2的蛋白表达。采用全细胞膜片钳技术记录Ito和动作电位。用PE处理新生大鼠心室肌细胞可诱导细胞肥大，上调CnAβ的蛋白表达，减弱Kv4.2的基因和蛋白表达以及Ito电流密度，并延长APD。使用Ad-CnAβshRNA1使新生大鼠心室肌细胞中的CnAβ沉默可显著抑制PE的上述作用。CnAβ基因沉默可抑制PE诱导的新生大鼠肥厚性心室肌细胞中瞬时外向钾电流离子通道的重塑及APD的改变。
