Beckett Travis, Bonneau Laura, Howard Alan, Blanchard James, Borda Juan, Weiner Daniel J, Wang Lili, Gao Guang Ping, Kolls Jay K, Bohm Rudolf, Liggitt Denny, Weiss Daniel J
Pulmonary and Critical Care, Vermont Lung Center, University of Vermont College of Medicine, Burlington, VT 05405, USA.
Hum Gene Ther Methods. 2012 Apr;23(2):98-110. doi: 10.1089/hgtb.2012.014. Epub 2012 May 8.
Use of perfluorochemical liquids during intratracheal vector administration enhances recombinant adenovirus and adeno-associated virus (AAV)-mediated lung epithelial gene expression. We hypothesized that inhalation of nebulized perfluorochemical vapor would also enhance epithelial gene expression after subsequent intratracheal vector administration. Freely breathing adult C57BL/6 mice were exposed for selected times to nebulized perflubron or sterile saline in a sealed Plexiglas chamber. Recombinant adenoviral vector was administered by transtracheal puncture at selected times afterward and mice were killed 3 days after vector administration to assess transgene expression. Mice tolerated the nebulized perflubron without obvious ill effects. Vector administration 6 hr after nebulized perflubron exposure resulted in an average 540% increase in gene expression in airway and alveolar epithelium, compared with that with vector alone or saline plus vector control (p<0.05). However, vector administration 1 hr, 1 day, or 3 days after perflubron exposure was not different from either nebulized saline with vector or vector alone and a 60-min exposure to nebulized perflubron is required. In parallel pilot studies in macaques, inhalation of nebulized perflubron enhanced recombinant AAV2/5 vector expression throughout the lung. Serial chest radiographs, bronchoalveolar lavages, and results of complete blood counts and serum biochemistries demonstrated no obvious adverse effects of nebulized perflubron. Further, one macaque receiving nebulized perflubron only was monitored for 1 year with no obvious adverse effects of exposure. These results demonstrate that inhalation of nebulized perflubron, a simple, clinically more feasible technique than intratracheal administration of liquid perflubron, safely enhances lung gene expression.
在气管内载体给药过程中使用全氟化学液体可增强重组腺病毒和腺相关病毒（AAV）介导的肺上皮基因表达。我们推测，雾化全氟化学蒸汽吸入在随后的气管内载体给药后也会增强上皮基因表达。将自由呼吸的成年C57BL/6小鼠在密封的有机玻璃箱中暴露于雾化的全氟辛烷或无菌盐水中特定时间。之后在选定时间通过经气管穿刺给予重组腺病毒载体，并在载体给药后3天处死小鼠以评估转基因表达。小鼠耐受雾化的全氟辛烷，无明显不良影响。与单独使用载体或盐水加载体对照相比，雾化全氟辛烷暴露6小时后给予载体导致气道和肺泡上皮中的基因表达平均增加540%（p<0.05）。然而，全氟辛烷暴露1小时、1天或3天后给予载体与雾化盐水加载体或单独载体给药无差异，并且需要60分钟的雾化全氟辛烷暴露。在猕猴的平行试点研究中，雾化全氟辛烷吸入增强了重组AAV2/5载体在整个肺部的表达。系列胸部X线片、支气管肺泡灌洗以及全血细胞计数和血清生化结果显示雾化全氟辛烷无明显不良反应。此外，对仅接受雾化全氟辛烷的一只猕猴进行了1年的监测，暴露无明显不良反应。这些结果表明雾化全氟辛烷吸入是一种比气管内给予液体全氟辛烷更简单、临床上更可行的技术，可安全地增强肺基因表达。
