肌浆网钙ATP酶基因转导对慢性心力衰竭犬心肌蛋白质组影响的初步研究  

Overexpression of sarcoplasmic reticulum calcium ATPase induced hemodynamic and proteomic changes in a dog model of heart failure

作　　者：付治卿[1] 李小鹰[1] 刘秀华[2] 孙胜[2] 刘涛[3] 米亚非[1] 周声安[1] 叶卫华[4] 王青松[5] 

机构地区：[1]解放军总医院老年心内科,北京100853 [2]解放军总医院病现生理研究室,北京100853 [3]解放军总医院老年超声科,北京100853 [4]解放军总医院心外科,北京100853 [5]北京大学生命科学院

出　　处：《中华心血管病杂志》2008年第3期260-265,共6页Chinese Journal of Cardiology

基　　金：国家自然科学基金资助项目（30470716）;国家重点基础研究发展规划资助项目（G2000056906）

摘　　要：目的分析心肌肌浆网Ca^2＋-ATP酶（sarcoplasmic reticulum Ca^2＋ ATPase 2a，SERCA2a）基因转导对慢性心力衰竭（HF）犬心肌蛋白质组的影响，探讨SERCA2a基因转导改善心功能的机制。方法快速右心室起搏建立HF犬模型并随机分为HF组、HF＋绿色荧光蛋白（enhanced green fluorescent protein，EGFP）组、HF＋SERCA2a组。后两组分别向心肌内注射携带EGFP和SERCA2a基因的rAAV载体。于基因转导30d时停止起搏后进行超声心动图和血流动力学检查并制备心室肌双向电泳蛋白样品和心肌双向电泳图谱，图像分析软件分析蛋白表达差异点，MALDI-TOF-MS数据库搜索鉴定蛋白质。结果基因转导30d时，HF＋SERCA2a组犬的症状、超声心动图和血流动力学指标与HF＋EGFP组相比有显著好转（P〈0．05）；与对照组相比差异无统计学意义（P〉0．05）。挑选SERCA2a基因转导后表达量发生明显改变的10个蛋白点进行分析，经质谱鉴定分别为心肌收缩相关蛋白、线粒体能量代谢酶类和应激相关蛋白。结论以rAAV为载体介导SERCA2a基因转导能够改善HF犬心脏的收缩和舒张功能，其可能的机制是恢复了心肌收缩相关蛋白正常表型或正常表达量，增加了心肌能量的产生，改变了应激相关蛋白的表达。Objective Overexpression of SERCA2a could improve cardiac function in human and experimental heart failure （HF） models. We observed the proteomics changes pest SERCA2a overexpression in a pacing induced HF model in dogs. Methods Beagles were divided into four groups： control group, HF group （230 beats/min for 4 weeks）, HF ± EGFP group （myocardial injection of 1 × 10^12v. g recombinant adeno-assoeiated virus carrying enhanced green fluorescent protein gene, rAAV2/1-EGFP） and HF ± SERCA2a group （ myocardial injection of 1 ×10^12v. g recombinant adeno-assoeiated virus carrying SERCA2a gene, rAAV2/1-SERCA2a）. Thirty days after gene transduction, left ventricular systolic and diastolic functions were measured by echoeardiography and invasive hemodynamics in all animals. By use of 2- dimensional gel electrophoresis （ 2-DE）, - 500 distinct protein spots were detected in myoeardium of all animals. Protein spots observed to be altered between failing and SERCA2a overexpressed hearts were subjected to tryptic peptide mass fingerprinting for identification by MALDI-TOF mass spectrometry in combination with LC/MS/MS analysis. Results At 30 day after gene transfer, HF signs were significantly reduced, cardiac function [ LVSP： （ 214.72 ± 31.74） mm Hg （ 1 mm Hg = 0. 133 kPa） vs. （ 139.32 ± 36.79）mm Hg, ±dp/dtmax：（6779.43 ±217.58） mm Hg/s vs. （2746.85 ±931.23） mm Hg/s and -dp/ dtmmax：（ -4341.42 ± 322.02） mm Hg/s vs. （ -2531.14 ± 616.15） mm Hg/s, LVEDP： （21.86 ± 6.95） mm Hg vs. （59.78 ± 6.92） mm Hg] significantly improved in HF ± SERCA2a dogs than those in HF ± EGFP group（ all P 〈0.05 ） and parameters were comparable between HF ± SERCA2a and control groups. We identified alterations in the expression level of more than 10 proteins in myoeardium. These protein changes were observed mainly in two subeellular compartments： the cardiac contractile apparatus and metabolism/energetics. Conclusion These results showed that overexpression of SERCA2a

关 键 词：心力衰竭 充血性 肌浆网 基因疗法 心肌收缩 能量代谢 

分 类 号：R686[医药卫生—骨科学]