715. Noninvasive Bioluminescence Imaging of Gene Expression Following Intracerebral Gene Delivery of AAV and Adenovirus into the Mouse Brain

Abstract

Top of pageAbstract At present, the most methods to evaluate in vivo gene expression are limited to the use of tissue obtained from sacrificed animals. To develop a simple, non-invasive and quantitative method for the detection of gene expression in the mouse central nervous system (CNS) while the animal is living, we evaluated a method of repetitively tracking in vivo gene expression of firefly luciferase (Luc) using a Cooled Charged Couple Detector (CCCD) camera. To evaluate the sensitivity of the bioluminescence imaging system, the initial experiment was performed by stereotactic injection of different doses of recombinant E1-deleted type 5 adenovirus expressing the Luc, Ad5.Luc, into the striatum of anesthetized adult C57BL/6 mice. The real-time in vivo gene expression of luciferase was monitored by CCCD camera at day 2, 7, 14, 28, and day 60 post gene transfer. The results indicated that mice treated with middle dose of Ad5.Luc (4|[times]|107 VP), a bioluminescence signal was detected at the site of injection as early as 2 days post injection. In contrast, the mice treated with higher dose of adenovirus (4|[times]|108 VP), the expression level reached its peak at day 7 and declined after day14. There was no detectable signal with the lowest dose of adenovirus (4|[times]|106 VP) at any of the time point post injection. To evaluated two recombinant adeno-associated virus (AAV) serotypes (AAV2/2, and AAV 2/5) with the bioluminescence imaging system in the mouse CNS, a subsequent experiment was performed using the similar protocol, stereotactic-guide injected 5|[times]|109 VP of AAV.Luc in 4 ul of PBS into the striatum of C57BL/6 mice. The results showed comparable expression of Luc in the mice treated with both AAV 2/2 and AAV 2/5 on day 2 post gene transfer. Between day 7 to day 28, the luciferase imaging in the mice treated with AAV 2/5 was two to five fold higher than in the mice treated with AAV 2/2. Gene expression of both AAV 2/2 and AAV2/5 have been persistent over two months in the live mice. Our results demonstrate that real-time in vivo bioluminescence imaging is a promising method to measure gene expression in the CNS of mice.