Figure 4.

Restoration of Dystrophin in Tibialis Anterior Muscles of mdx Mice Aged 4–5 Weeks 2 Weeks after Intramuscular Injection

Muscles treated with PMOE23 only were used as controls; all other samples were from muscles treated with 5 μg polymer formulated with 2 μg PMOE23 or 2 μg Z8-PMO in 40 μL saline. (A) Dystrophin was detected by immunohistochemistry with rabbit polyclonal antibody P7 against dystrophin. Blue nuclear staining with 4,6-diamidino-2-phenylindole (original magnification, ×200; scale bar, 200 μm). (B) The percentage of dystrophin-positive fibers. The number of dystrophin-positive fibers was counted in a single cross-section (n = 5, two-tailed Student’s t test, *p ≤ 0.05 compared with PMO). (C) Detection of exon 23 skipping by RT-PCR. Total RNA of 100 ng from each sample was used for amplification of dystrophin mRNA from exon 20 to exon 26. The upper bands (1,093 bp) correspond to the normal mRNA, and the lower bands (880 bp) correspond to the mRNA with exon E23 skipped. (D) Western blots demonstrate the expression of dystrophin protein from treated mdx mice in comparison with C57BL/6 and untreated mdx mice (10 μg of total protein was loaded for PMO/modified PMO and control mdx samples; 10 μg for the wild-type (WT) C57 control also). Dys, dystrophin detected with monoclonal antibody Dys 1. α-Actin used as the loading control.