人内皮型一氧化氮合酶在心肌细胞和内皮细胞中的表达。

Expression of human eNOS in cardiac and endothelial cells.

作者信息

Wang Xiuqing, Cade Robert, Sun Zhongjie

机构信息

Department of Medicine, University of Florida, Gainesville, FL, USA.

出版信息

Methods Mol Med. 2005;112:91-107. doi: 10.1007/978-1-59259-879-3_7.

DOI:10.1007/978-1-59259-879-3_7

PMID:16010013

Abstract

In this chapter we provide a detailed technique-focused protocol for expression of human endothelial nitric oxide synthase (eNOS) gene delivered by replication-deficient adenovirus. It includes construction of recombinant plasmid with human eNOS gene full-length cDNA (heNOS), packaging of adenovirus with heNOS (Adv-heNOS) in 293 cells, confirmation of Adv-heNOS, amplification of high-titer stockAdv-heNOS, determination of the Adv-heNOS titer, expression of Adv-heNOS in the human abdominal aorta endothelial (HAAE1) cell line and rat heart myoblast cell line (H9C2), and measurement of human eNOS activity and NO production.

摘要

在本章中，我们提供了一个详细的、以技术为重点的方案，用于表达由复制缺陷型腺病毒递送的人内皮型一氧化氮合酶（eNOS）基因。该方案包括用人eNOS基因全长cDNA（heNOS）构建重组质粒，在293细胞中用heNOS包装腺病毒（Adv-heNOS），确认Adv-heNOS，扩增高滴度储备Adv-heNOS，测定Adv-heNOS滴度，在人腹主动脉内皮（HAAE1）细胞系和大鼠心脏成肌细胞系（H9C2）中表达Adv-heNOS，以及测量人eNOS活性和NO生成量。