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转基因联合抑制小鼠心脏CaMKII与Ik1的初步研究

黄芸, 戴淼, 杜以梅, 姚雨峰, 张家明, 苏冠华, 舒砚文, 崔天盆, 杜心灵, 李景东

华中科技大学1附属协和医院老年病科；附属协和医院心内科；生命科学院；武汉市第一医院检验中心；附属协和医院心外科，武汉 430022

摘要

本文旨在建立转基因联合抑制心脏钙/钙调素依赖性蛋白激酶II (Ca2+/calmodulin-dependent protein kinase II, CaMKII)及内向整流性钾电流(Ik1)小鼠模型，初步探讨该联合抑制在基础和应激状态下对小鼠心脏电生理、结构与功能重构的影响。将动物分为4组：野生组(WT)，转基因CaMKII抑制组(AC3-I)，转基因Ik1抑制组(Kir2.1-AAA)，转基因联合抑制组(AC3-I+ Kir2.1-AAA)。记录各组动物在基础状态下及注射异丙肾上腺素后的心电图；同时分别对各组动物进行心脏超声检查，以了解心脏重构情况；在酶解法分离单个左室心肌细胞后，采用全细胞膜片钳法记录心肌细胞Ik1和瞬间外向钾电流(Ito)。实验结果显示：在基础状态下，心电图显示各组动物之间的心率、PR间期和QRS间期均无显著差别；注射异丙基肾上腺素后，上述心电图指标和小鼠发生室性心律失常的情况在各组间也无显著差别；4组小鼠间M型(Motion-mode)和二维超声检测的各项指标无显著差异；全细胞膜片钳实验结果表明，AC3-I组动物心肌细胞的Ik1电流密度最大(P < 0.01)，WT组的次之(P < 0.01)，其他两组较WT组显著降低(P < 0.01)；同时，AC3-I组Ito电流密度显著高于其他三组(P < 0.01)，而其他三组间无明显差异。上述结果提示：转基因联合抑制CaMKII和Ik1可使CaMKII抑制的小鼠Ik1下调，在基础状态下不影响心脏的结构和功能变化，在注射异丙基肾上腺素后心律失常的发生亦无明显差别。本结果为进一步探讨联合抑制CaMKII与Ik1在各种心脏疾病状态下能否更有效地拮抗心肌的不利重构打下了基础。

关键词： 钙/钙调素依赖性蛋白激酶II; 内向整流性钾电流; 心脏重构; 心律失常

分类号：R331

[Combined transgenic inhibition of CaMKII and Ik1 on cardiac remodeling.] [Article in Chinese]

HUANG Yun, DAI Miao, DU Yi-Mei, YAO Yu-Feng, ZHANG Jia-Ming, SU Guan-Hua, SHU Yan-Wen, CUI Tian-Pen, DU Xin-Ling, LI Jing-Dong

Department of Gerontology； Department of Cardiology, Union Hospital； Life Science College； Laboratory Medicine Center, the First Hospital of Wuhan； Department of Cardiac Surgery of Union Hospital, Huazhong University of Science and Technology, Wuhan 430022, China

Abstract

This study was aimed to establish an experimental mouse model of combined transgenic inhibition of both multifunctional Ca2+/calmodulin-dependent protein kinase II (CaMKII) and inward rectifier potassium current (Ik1), and to observe whether the specific inhibition of both CaMKII and Ik1 can bring about any effects on cardiac remodeling. Mice were divided into 4 groups: wild type (WT), CaMKII inhibited (AC3-I), Ik1 inhibited (Kir2.1-AAA) and combined inhibition of both CaMKII and Ik1 (AC3-I+Kir2.1-AAA). Mice in each group received electrocardiogram (ECG) and echocardiography examination. ECG in the condition of isoproterenol (ISO) injection was also checked. The whole cell patch clamp technique was used to measure Ik1 and the transient outward potassium current (Ito) from enzymatically isolated myocytes of left ventricle. In the condition of basal status, no significant changes of heart rate, PR interval and QRS interval were observed. No mouse showed ventricular arrhythmias in all of the 4 groups. After ISO injection, each group presented no significant ventricular arrhythmias either. The indexes measured by M-mode (motion-mode) and two-dimensional echocardiography had no significant differences among the four groups. Ik1 in AC3-I group was significantly higher than those in other three groups (P < 0.01) because of the results brought about by CaMKII inhibition. Among the latter three groups, both Kir2.1-AAA group and AC3-I+Kir2.1-AAA group had a significant reduced Ik1 compared with that of WT group, which was due to the Ik1 inhibition (P < 0.01). Ito in AC3-I group was higher than that of the other three groups (P < 0.01), but there were no significant differences in Ito among WT, Kir2.1-AAA and AC3-I+Kir2.1-AAA groups. Thus, combined transgenic myocardial CaMKII and Ik1 inhibition eliminated the up-regulation of Ik1 in CaMKII inhibited mice, and had no effects on cardiac remodeling including heart structure and function as well as arrhythmias at the basic and ISO conditions. The results of this study may provide a basis for the further investigation of combined inhibition of CaMKII and Ik1 in pathogenic cardiac remodeling.

Key words: CaMKII; Ik1; arrhythmias; cardiac remodeling

收稿日期：2014-10-11　　录用日期：2015-01-02

通讯作者：李景东　　E-mail: jingdong-li@mail.hust.edu.cn

引用本文：

黄芸, 戴淼, 杜以梅, 姚雨峰, 张家明, 苏冠华, 舒砚文, 崔天盆, 杜心灵, 李景东. 转基因联合抑制小鼠心脏CaMKII与Ik1的初步研究[J]. 生理学报 2015; 67 (2): 201-206.

HUANG Yun, DAI Miao, DU Yi-Mei, YAO Yu-Feng, ZHANG Jia-Ming, SU Guan-Hua, SHU Yan-Wen, CUI Tian-Pen, DU Xin-Ling, LI Jing-Dong. [Combined transgenic inhibition of CaMKII and Ik1 on cardiac remodeling.] [Article in Chinese]. Acta Physiol Sin 2015; 67 (2): 201-206 (in Chinese with English abstract).