重组腺相关病毒为载体的心肌肌浆网钙离子ATP酶2a基因转导治疗慢性充血性心力衰竭:短期及中期作用(英文)  

Short-and middle-period therapeutic effects of recombinant adeno-associated viral gene transfer of sarcoplasmic reticulum Ca^(2+) ATPase on chronic congestive heart failure rats

作　　者：惠海鹏[1] 李小鹰[1] 刘秀华[2] 鲁晓春[1] 杨伟[3] 孙胜[2] 

机构地区：[1]解放军总医院老年心内科,北京市100853 [2]解放军总医院病理生理研究室,北京市100853 [3]北京大学生命科学院,北京市100871

出　　处：《中国组织工程研究与临床康复》2007年第29期5840-5845,共6页Journal of Clinical Rehabilitative Tissue Engineering Research

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(九七三)资助项目(G2000056906)~~

摘　　要：背景:慢性充血性心力衰竭(CHF)时,心肌肌浆网钙离子ATP酶2a(sarcoplasmic reticulum Ca2+ ATPase,SERCA2a)的蛋白表达和活性均明显降低,这导致钙离子的调控异常和心脏收缩和舒张功能不全。目的:观察腺相关病毒为载体的SERCA2a基因转导对CHF大鼠的短期和中长期治疗作用,探讨其治疗心力衰竭的多种可能的机制。设计:随机对照设计。单位:解放军总医院老年心内科和病理生理研究室。方法:将雄性SD大鼠随机分为,假手术组,CHF组,CHF+GFP组和CHF+SERCA2a组4组,每组又分为基因转导10,30d2个时间点,后3组采用腹主动脉缩窄术建立CHF大鼠模型,手术后18～20周,造模成功的大鼠进行经腹心包腔内注射,CHF组注射500μL无菌生理盐水,CHF+GFP组和CHF+SERCA2a组分别注射等量携带GFP基因和SERCA2a基因的重组腺相关病毒。主要观察指标:①于基因转导后10,30d,检测大鼠的血流动力学参数,应用WesternBlot方法检测SERCA2a蛋白表达;改良的Jones法测定SERCA2a活性。②应用蛋白质组学研究技术分析比较基因转导30d组中CHF和CHF+SERCA2a亚组大鼠的心肌蛋白质组表达的差异。③应用电泳分离和定量的方法检测基因转导30d组的各亚组大鼠心肌肌球蛋白重链亚型的表达情况。结果:①SERCA2a蛋白表达和活性:CHF组和CHF+GFP组低于假手术组。基因转导10d后,CHF+SERCA2a组与CHF组比较,SERCA2a蛋白表达增加80.7%,活性增加89.9%,但低于假手术组水平;基因转导30d后,CHF+SERCA2a组SERCA2a蛋白表达和活性已经达到假手术组大鼠水平。②心脏功能:基因转导10d后,CHF+SERCA2a组血流动力学指标中左室舒张末压力明显下降,其他指标明显升高,但未完全达到对照大鼠的心功能水平;基因转导30d后心脏收缩和舒张功能进一步改善,各项指标与假手术组比较差异不显著。③心肌蛋白质组的研究发现,基因转导30d后,CHF+SERCA2a组多种能量代谢的酶表达明显增加。④凝�BACKGROUND： in chronic congestive heart failure （CHF）, both protein expression and activity of sarcoplasmic reticulum （SR） Ca^2＋ ATPase （SERCA2a） are decreased, which results in abnormal regulation of calcium ion and systole and diastole dysfunction. OBJECTIVE：To observe the short- and middle-term therapeutic effects of adeno-associated viral gene transfer of SERCA2a in treatment of CHF in rats, and to investigate the possible mechanisms. DESIGN ： Randomized-controlled design SETTING ： Department of Geriatric Cardiology and Department of Pathophysiology, General Hospital of Chinese PLA. METHODS：Male Sprague-Dawley rats, were randomized into 4 groups： sham-operation group, CHF group, CHF＋ green fluorescent protein （GFP） group and CHF＋SERCA2a group. Gene transfer 10 and 30 days two time points were set in each group. CHF rat models were established by coarctation of abdominal aorta. At postoperative 18 to 20 weeks, the successful rats in CHF group were intraperitoneally injected with 500 μL aseptic normal saline, and those in the CHF＋ GFP group and CHF＋ SERCA2a group were injected with the same dose of Adeno-associated viral gene with GFP and SERCA2a gene respectively. MAIN OUTCOME MEASURES： ① At 10 and 30 days after gene transfer, hemodynamic parameters, the SERCA2a protein expression was analyzed by Western Blot, and SERCA2a activity was carded out by modified Jones method. ②The difference of proteome of hearts was detected by expression proteomics in between CHF＋SERCA2a group and CHF group at 30 days.③Electrophoretic separation and quantitation of cardiac myosin heavy chain（MHC） isoforms were done at 30 days. RESULTS： ①In CHF group and CHF＋GFP group, SERCA2a expression and activity were lower than those in sham-operation group. At 10 days after gene transfer, SERCA2a expression was increased by 80.7% and still lower than that of sham-operation group, and activity was also increased by 89.9%, but still lower than that of sham-operation group. And at 30 da

关 键 词：慢性心力衰竭 肌浆网离子ATP酶2a 腺相关病毒 基因疗法 

分 类 号：R541.6[医药卫生—心血管疾病]