Svensson E C, Marshall D J, Woodard K, Lin H, Jiang F, Chu L, Leiden J M
Departments of Medicine and Pathology, University of Chicago, Chicago, IL 60637, USA.
Circulation. 1999 Jan 19;99(2):201-5. doi: 10.1161/01.cir.99.2.201.
The delivery of recombinant genes to cardiomyocytes holds promise for the treatment of a variety of cardiovascular diseases. Previous gene transfer approaches that used direct injection of plasmid DNA or replication-defective adenovirus vectors have been limited by low transduction frequencies and transient transgene expression due to immune responses, respectively. In this report, we have tested the feasibility of using intramyocardial injection or intracoronary infusions of recombinant adeno-associated virus (rAAV) vectors to program transgene expression in murine cardiomyocytes in vivo.
We constructed an rAAV containing the LacZ gene under the transcriptional control of the cytomegalovirus (CMV) promoter (AAVCMV-LacZ). We then injected 1x10(8) infectious units (IU) of this virus into the left ventricular myocardium of adult CD-1 mice. Control hearts were injected with the AdCMV-LacZ adenovirus vector. Hearts harvested 2, 4, and 8 weeks after AAVCMV-LacZ injection demonstrated stable beta-galactosidase (beta-gal) expression in large numbers of cardiomyocytes without evidence of myocardial inflammation or myocyte necrosis. In contrast, the AdCMV-LacZ-injected hearts displayed transient beta-gal expression, which was undetectable by 4 weeks after injection. Explanted C57BL/6 mouse hearts were also perfused via the coronary arteries with 1.5x10(9) IU of AAVCMV-LacZ and assayed 2, 4, and 8 weeks later for beta-gal expression. beta-Gal expression was detected in <1% of cardiomyocytes at 2 weeks after perfusion but was detected in up to 50% of cardiomyocytes 4 to 8 weeks after perfusion.
Direct intramyocardial injection or coronary artery perfusion with rAAV vectors can be used to program stable transgene expression in cardiomyocytes in vivo. rAAV appears to represent a useful vector for the delivery of therapeutic genes to the myocardium.
将重组基因导入心肌细胞有望用于治疗多种心血管疾病。以往使用直接注射质粒DNA或复制缺陷型腺病毒载体的基因转移方法，分别受到转导频率低和因免疫反应导致转基因表达短暂的限制。在本报告中，我们测试了通过心肌内注射或冠状动脉输注重组腺相关病毒（rAAV）载体在体内对小鼠心肌细胞进行转基因表达编程的可行性。
我们构建了一种在巨细胞病毒（CMV）启动子转录控制下含有LacZ基因的rAAV（AAVCMV-LacZ）。然后将1×10⁸感染单位（IU）的这种病毒注射到成年CD-1小鼠的左心室心肌中。对照心脏注射AdCMV-LacZ腺病毒载体。在注射AAVCMV-LacZ后2、4和8周收获的心脏显示，大量心肌细胞中β-半乳糖苷酶（β-gal）表达稳定，没有心肌炎症或心肌细胞坏死的迹象。相比之下，注射AdCMV-LacZ的心脏显示β-gal表达短暂，注射后4周就检测不到了。将取出的C57BL/6小鼠心脏也通过冠状动脉灌注1.5×10⁹IU的AAVCMV-LacZ，并在灌注后2、4和8周检测β-gal表达。灌注后2周，在<1%的心肌细胞中检测到β-gal表达，但在灌注后4至8周，高达50%的心肌细胞中检测到了β-gal表达。
直接心肌内注射或冠状动脉灌注rAAV载体可用于在体内对心肌细胞进行稳定的转基因表达编程。rAAV似乎是一种将治疗性基因递送至心肌的有用载体。
