High efficiency gene transfer to skeletal muscle inadult mouse for Duchenne muscular dystrophy using adenovirus helper cells.

我们已经开发了一种策略，以实现一个有效的基因转导E1-删除腺病毒的骨骼肌成年小鼠（>2个月大）。使用来自mdx小鼠的原代成肌细胞开发携带5型腺病毒E1基因的新生产细胞（E32细胞）。用携带编码lacZ基因的E1缺失腺病毒感染新的辅助细胞和293细胞，并注射到成年mdx小鼠的骨骼肌中。在成年mdx小鼠的骨骼肌中检测到许多lacZ阳性纤维，这些小鼠在注射后5天被处死。用293细胞和E32细胞进行基因转导的效率分别是成肌细胞介导的基因转导的6.2倍和3.6倍。我们假设腺病毒感染的293细胞释放某种因子，破坏成熟肌纤维对病毒感染的屏障，从而增加转导。总之，使用293细胞和E32细胞的基因转导效率高于成肌细胞介导的基因转移和直接病毒注射。更重要的是，这些辅助细胞使成年骨骼肌的有效基因转导成为可能。

We have developed a strategy for achieving an efficient gene transduction of E1-deleted adenovirus to skeletal muscle in adult mice (>2 months old). New producer cells carrying the E1 gene of adenovirus type 5 (E32 cell) were developed using primary myoblasts from mdx mouse. The new helper cells and 293 cells were infected with E1-deleted adenovirus carrying the gene-encoding lacZ and were injected into skeletal muscle of adult mdx mice. Many lacZ-positive fibers were detected in the skeletal muscle of adult mdx mice, which were sacrificed at 5 days post-injectiorL This new method was compared with regular myoblast-mediated gene transfer and direct injection of adenovirus. The efficiency of gene transduction, using 293 cells and E32 cells, were 6.2 times and 3.6 times as high as myoblast-mediated gene transfer, respectively. We hypothesize that the adenovirus-infected 293 cells release some type of factor that breaks down the mature myofibers' barrier to viral infection, thereby increasing transduction. In conclusion, the efficiencies of gene transduction using 293 cells and E32 cells were higher than myoblast-mediated gene transfer and direct viral injection. More importantly, those helper cells enabled efficient gene transduction of adult skeletal muscle.