Zhang G, Vargo D, Budker V, Armstrong N, Knechtle S, Wolff J A
Department of Pediatrics, University of Wisconsin--Madison, 53705, USA.
Hum Gene Ther. 1997 Oct 10;8(15):1763-72. doi: 10.1089/hum.1997.8.15-1763.
A variety of reporter genes within plasmid constructs were injected into the afferent and efferent vessels of the liver in mice, rats, and dogs. Efficient plasmid expression was obtained following delivery via the portal vein, the hepatic vein, and the bile duct. The use of hyperosmotic injection solutions and occlusion of the blood outflow from the liver substantially increased the expression levels. Combining these surgical approaches with improved plasmid vectors enabled uncommonly high levels of foreign gene expression in which over 15 microg of luciferase protein/liver was produced in mice and over 50 microg in rats. Equally high levels of beta-galactosidase (beta-Gal) expression were obtained, in that over 5% of the hepatocytes had intense blue staining. Expression of luciferase or beta-Gal was evenly distributed in hepatocytes throughout the entire liver when either of the three routes were injected. Peri-acinar hepatocytes were preferentially transfected when the portal vein was injected in rats. These levels of foreign gene expression are among the highest levels obtained with nonviral vectors. Repetitive plasmid administration through the bile duct led to successive events of foreign gene expression. The integration of these findings into laboratory and clinical protocols is discussed.
将质粒构建体中的多种报告基因注射到小鼠、大鼠和狗的肝脏传入和传出血管中。通过门静脉、肝静脉和胆管给药后可获得高效的质粒表达。使用高渗注射溶液和阻断肝脏的血液流出可显著提高表达水平。将这些手术方法与改良的质粒载体相结合，能够实现异常高水平的外源基因表达，其中小鼠肝脏产生的荧光素酶蛋白超过15微克，大鼠超过50微克。β-半乳糖苷酶（β-Gal）也获得了同样高的表达水平，超过5%的肝细胞有强烈的蓝色染色。当通过三种途径中的任何一种进行注射时，荧光素酶或β-Gal的表达在整个肝脏的肝细胞中均匀分布。在大鼠中经门静脉注射时，腺泡周围肝细胞优先被转染。这些外源基因表达水平是使用非病毒载体获得的最高水平之一。通过胆管重复给予质粒导致外源基因表达的连续事件。本文讨论了将这些研究结果整合到实验室和临床方案中的情况。
