Robust cardiomyocyte-specific gene expression following systemic injection of AAV: in vivo gene delivery follows a Poisson distribution.
Robust cardiomyocyte-specific gene expression following systemic injection of AAV: in vivo gene delivery follows a Poisson distribution.
复制标题
作者：
新分离的AAV血清型容易穿过内皮屏障，以在整个身体中提供有效的转基因递送。然而，在大多数实验研究和基因治疗方案中，组织特异性表达是优选的。以前的努力限制基因表达的心肌往往依赖于直接注射到心肌或冠状动脉内灌注。在这里，我们报告了一个AAV载体系统采用心肌肌钙蛋白T启动子（cTnT）。使用荧光素酶和eGFP，在对1周龄小鼠全身施用后测试使用AAV血清型衣壳：AAV-1、2、6、8或9的心脏报告基因表达的效率和特异性。荧光素酶测定显示cTnT启动子与每种AAV血清型衣壳组合工作以提供心肌细胞特异性基因表达，但AAV-9紧随其后，其次是AAV-8是最有效的。与下一个最高的组织（肝脏）相比，来自cTnT启动子的AAV 9介导的基因表达在心脏中高640倍。eGFP荧光显示使用AAV-9在每只小鼠仅3.15×1010个病毒颗粒的剂量下的转导效率为96%。此外，在逐个细胞的基础上测量的心肌细胞eGFP荧光的强度显示，心脏中AAV介导的基因表达可以被建模为泊松分布;每个细胞平均需要近两个载体基因组才能达到85%的转导效率。
Newly-isolated serotypes of AAV readily cross the endothelial barrier to provide efficient transgene delivery throughout the body. However, tissue-specific expression is preferred in most experimental studies and gene therapy protocols. Previous efforts to restrict gene expression to the myocardium often relied on direct injection into heart muscle or intracoronary perfusion. Here, we report an AAV vector system employing the cardiac troponin T promoter (cTnT). Using luciferase and eGFP, the efficiency and specificity of cardiac reporter gene expression using AAV serotype capsids: AAV-1, 2, 6, 8 or 9 were tested after systemic administration to 1 week old mice. Luciferase assays showed that the cTnT promoter worked in combination with each of the AAV serotype capsids to provide cardiomyocyte-specific gene expression, but AAV-9 followed closely by AAV-8 was the most efficient. AAV9-mediated gene expression from the cTnT promoter was 640-fold greater in the heart compared to the next highest tissue (liver). eGFP fluorescence indicated a transduction efficiency of 96% using AAV-9 at a dose of only 3.15×1010 viral particles per mouse. Moreover, the intensity of cardiomyocyte eGFP fluorescence measured on a cell-by-cell basis revealed that AAV-mediated gene expression in the heart can be modeled as a Poisson distribution; requiring an average of nearly two vector genomes per cell to attain an 85% transduction efficiency.
登录
查看更多内容
影响因子：
12.4
作者：
Prasad, Konkal-Matt R.;Xu, Yaqin;French, Brent A.
通讯作者：
French, Brent A.
影响因子：
20.1
作者：
Pacak, Christina A.;Mah, Cathryn S.;Byrne, Barry J.
通讯作者：
Byrne, Barry J.
影响因子：
14.9
作者：
Jordan, M;Schallhorn, A;Wurm, FM
通讯作者：
Wurm, FM
影响因子：
5.1
作者：
Barker, SD;Dmitriev, IP;Hemminki, A
通讯作者：
Hemminki, A
DOI：
10.1073/pnas.182412299
发表时间：
2002-09-03
影响因子：
11.1
作者：
Gao, GP;Alvira, MR;Wilson, JM
通讯作者：
Wilson, JM