Figure 2.

ΔH2 mini-dystrophin expression from two independent sets of trans-splicing AAV vectors. The EDL and the TA muscles of 2-month-old mdx mice were co-infected with 1 × 10¹⁰ vector genomes (vg) (for the EDL muscle) and 2 × 10¹⁰ vg (for the TA muscle) particles of each pair of trans-splicing viruses. Single vector infection (donor or acceptor alone) was also included in the cohort study (5 × 10⁹ vg particles for the EDL muscle and 1 × 10¹⁰ vg particles for the TA muscle). Mini-dystrophin expression was evaluated 1 month later. (a) Schematic diagram of ΔH2 mini-dystrophin trans-splicing vectors (not drawn to scale). The donor virus carries the cytomegalovirus (CMV) promoter, the 5′ portion of the ΔH2 minigene and the endogenous splicing donor signal (from intron 60 and 63, respectively). The acceptor virus carries the endogenous splicing acceptor signal from the same intron (60 and 63, respectively), the remaining minigene and the polyA sequence. The viruses are accordingly named after the respective intron. (b) Western blot analysis of mini-dystrophin expression in muscle extracts with monoclonal N-terminal (specific for human dystrophin) and C-terminal antibodies, respectively. Each lane is from one infected TA muscle. Different samples were used for the N-terminal and C-terminal immunoblots. (c) Quantification of ΔH2 minigene expression by immunofluorescence staining with a human dystrophin–specific antibody. Left panels are the representative photomicrographs from the TA muscles infected with the indicated set of trans-splicing vectors. Scale bar, 200 μm. Right panel represents mini-dystrophin-positive myofibers per muscle section. n = 3–7 samples for each group. *, the difference between two sets of trans-splicing vectors was statistically significant.