心肌肌浆网Ca2+-ATP酶基因转导改善慢性心力衰竭犬心功能的研究

目的： 探讨以重组腺相关病毒(rAAV)为载体的心肌肌浆网Ca2+-ATP酶(SERCA2a)基因转导对慢性心力衰竭犬心功能的影响。方法: 选取成年比格犬17只，随机分为对照组4只和心力衰竭组11只。快速右心室起搏建立慢性心力衰竭犬模型并随机分为心力衰竭组4只、心力衰竭＋绿色荧光蛋白（EGFP）组4只、心力衰竭＋SERCA2a组5只(其中1只在开胸后死亡)。接受基因导入的心力衰竭犬行开胸术，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。结果: 基因转导30 d时，心力衰竭＋SERCA2a 组犬的症状及超声心动图指标与心力衰竭＋EGFP组相比有显著好转（P<0.05）；与对照组相比无显著差别。血流动力学监测发现，转导SERCA2a的犬LVSP、+dp/dtmax和-dp/dtmax明显升高，平均值较EGFP组分别增加54.12％［(214.72±31.74)mmHg vs (139.32±36.79)mmHg］、146.81％［(6 779.43±217.58)mmHg/s vs (2 746.85±931.23)mmHg/s］和71.52％［(-4 341.42±322.02)mmHg/s vs (-2 531.14±616.15 mmHg/s)］，LVEDP则降低了63.43％［(21.86±6.95)mmHg vs (59.78±6.92)mmHg］，所有参数与对照组相比无显著差异。心力衰竭＋EGFP组犬心肌冰冻切片在激光共聚焦显微镜下可见弥漫绿色荧光。结论: 以rAAV为载体介导SERCA2a基因转导能够改善慢性心力衰竭犬心脏的收缩和舒张功能，是1种有前景的治疗慢性心力衰竭的方法。

Restoration of cardiac function in failing beagle hearts by recombinant adeno-associated viral gene transfer of sarcoplasmic reticulum Ca2+-ATPase

AIM: Abnormal Ca2+ homeostasis is one basic cause of heart failure. Studies have recently shown that overexpression of sarcoplasmic reticulum (SR) Ca2+-ATPase (SERCA2a) by adenoviral/adeno-associated viral gene transfer restores contractile function ex vivo and in murine or rabbit models. We therefore hypothesized that an increase in SERC A2a protein will improve cardiac function in a pacing-induced big animal model of heart failure.METHODS: 17 beagles were randomized into control group (CG, n=4) and chronic heart failure group (n=11). Four weeks after right ventricular rapid pacing (230 beats/min), 11 beagles all got heart failure (documented by >29.3% decrease in ejection fraction). 4 of 11 were used as heart failure group (HF, n=4). 9 HF beagles were randomized to receive either a recombinant adeno-associated viral carrying the SERCA2a gene (HF+SERC A2a, n=5) or the reporter gene enhanced green fluorescent protein (HF+EGFP, n=4) by thoracotomy. All HF beagles paced by 180 beats/min in order to maintain failing state. Thirty days after infection, parameters of systolic and diastolic function were measured by doppler echocardiography and hemodynamic monitor in all beagles.RESULTS: At 30 days after gene transfer, symptoms of HF+SERCA2a dogs improved. Echocardiogram parameters were superior to those in HF+EGFP group (P<0.05). Cardiac hemodynamic parameters of HF+SERCA2a dogs strikingly improved: LVSP, +dp/dtmax and -dp/dtmax increased, mean value increased respectively 54.12％［(214.72±31.74) mmHg vs (139.32±36.79) mmHg］, 146.81％［(6 779.43±217.58) mmHg/s vs (2 746.85±931.2) mmHg/s］ and 71.52％［(-4 341.42±322.02) mmHg/s vs (-2 531.14±616.15) mmHg/s］; LVEDP lowered 63.43％［(21.86±6.95) mmHg vs (59.78±6.92) mmHg］ compared with the dogs in HF+EGFP group. No significant difference in all parameters compared with those of control group was observed. Under laser confocal microscopy, widespread green fluorescence was observed in the myocardial frozen section of dogs in HF+EGFP group. CONCLUSION: These results support the hypothesis that overexpression of SERCA2a improves cardiac function in big animal model of chronic heart failure. The study demonstrates that gene transfer of SERCA2a into cardiac with recombinant adeno-associated viral vector is a prospective therapy methods.