小电导钙激活钾通道SK2在容量超负荷心力衰竭大鼠窦房结表达的改变

Changes of small conductance calcium-activated potassium channel SK2 expression in sinoatrial node of heart failure rats induced by volume-overload

目的观察小电导钙激活钾通道SK2通道在心力衰竭(HF)大鼠窦房结组织中表达的改变。方法以腹主动脉-下腔静脉穿刺造瘘术建立容量超负荷心力衰竭大鼠模型(n=7),假手术对照组(SO)大鼠切开腹腔,穿刺腹主动脉,但不造成腹主动脉-下腔静脉之间的瘘(n=7)。于术后8周通过超声心动图测定左心室射血分数(left ventricular ejection fraction,LVEF)、左心室短轴缩短率(left ventricular fraction shortening,LVFS)、左心室收缩末内径(left ventricular end-systolic diameter,LVEDs)、左心室舒张末内径(left ventrieular end-diastolic diameter,LVEDd),并于取材时测定心脏质量/体质量(heart weight/body weight),评价心脏结构与功能改变。分离大鼠窦房结细胞以免疫荧光染色法观察SK2通道蛋白的表达,同时取材窦房结组织,提取组织蛋白,行Western blot测定比较心衰大鼠窦房结组织SK2通道表达的改变情况。结果超声心动图结果显示,心力衰竭大鼠心脏收缩功能与舒张功能明显减低[LVEDd(mm):SO 5.91±0.36 vs HF 11.15±0.91,P<0.01;LVEDs(mm):SO 2.89±0.28 vs HF 7.89±0.70,P<0.01;LVEF(%):SO 87.10±1.63 vs HF 62.10±2.57,P<0.01;LVFS(%):SO51.27±2.03 vs HF 30.05±1.66,P<0.01],心脏质量/体质量明显增大(SO 244.19±36.87 vs HF 593.21±28.72,P<0.01)。单细胞免疫荧光染色结果示SK2通道表达于两组大鼠窦房结细胞膜上,Western blot定量分析结果示心力衰竭组大鼠窦房结组织SK2通道蛋白表达水平较假手术组明显减低(0.22±0.10 vs 0.31±0.16,P<0.05)。结论 SK2通道在心力衰竭大鼠窦房结组织中表达明显下调。

Objective To investigate the changes of SK2 channel expression in sinoatrial node of the heart failure（ HF） rats induced by volume-overload. Methods The volume-overload HF rats were induced by aortocaval fistula（ ACF,n = 7）,and the sham-operated rats were regarded as control group（ SO,n = 7）. The abdominal cavity was opened to puncture abdominal aorta without making a fistula between abdominal aorta and inferior vena cava in SO rats. The echocardiogram indexes such as left ventricular end-systolic diameter（ LVEDs）,left ventricular end-diastolic diameter（ LVEDd）,left ventricular fraction shortening（ LVFS）,left ventricular end-systolic diameter（ LVEDs） were measured at week 8 after operation,and the ratio of heart weight（ HW） to body weight（ BW） was caculated.Then the sinoatrial node cells were isolated and collected to observe the expression of the SK2 channel protein by immunofluorescent staining. Meanwhile,sinoatrial node tissues of rats were isolated to determine the change of SK2 channel protein expression by Western blot. Results The data of echocardiogram showed that the systolic and diastolic function in HF rats significantly declined [LVEDd（ mm） ： SO 5. 91 ± 0. 36 vs HF 11. 15 ± 0. 91,P 〈0. 01; LVEDs（ mm） ： SO 2. 89 ± 0. 28 vs HF 7. 89 ± 0. 70,P 〈0. 01; LVEF（ %） ： SO87. 10 ± 1. 63 vs HF 62. 10 ± 2. 57,P〈 0. 01; LVFS（ %） ： SO 51. 27 ± 2. 03 vs HF 30. 05 ± 1. 66,P〈 0. 01） ],and the ratio of HW to BW significantly increased（ SO 244. 19 ± 36. 87 vs HF 593. 21 ± 28. 72,P 〈0. 01）. The results of immunofluorescent staining indicated that the SK2 channels were located on membrane of single sinoatrial node cell. The Western blot results demonstrated that the protein expression of SK2 was down-regulated significantly in sinoatrial node of HF rats compared with SO rats（ 0. 22 ± 0. 10 vs 0. 31 ±0. 16,P 〈0. 01）. Conclusion The expression of SK2 channel protein is significantly down-regulated in sinoatrial node of HF rats.