Fig. 1

Expression of iNOS in DMD muscle. (A) Western blot analysis of human quadriceps muscle extract for iNOS. Proteins were separated in reducing conditions by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS) and transferred to nitrocellulose membrane. The blot was incubated with an antibody specific for iNOS and developed as described in Methods. Prestained molecular standards were shown on the left. Lane 1, positive control: mouse macrophage lysate prepared from the RAW 264.7 cell line, which were stimulated with IFNγ and LPS for 12 hr; lanes 3, 5, samples from patients with DMD; lane 7, patient with BMD; lanes 2, 4, 6, samples from control normal subjects. All lanes contain 45 µg of total protein. No specific signal was detected on the muscle samples from control normal subjects whereas a signal was observed on the muscle samples from patients with DMD/BMD. (B) Transverse serial cryostat sections (10-µm thick) of a deltoid muscle from control (left column) and DMD (right column) patients stained by HE (upper row) and immunostained by an antibody anti-iNOS (lower row). Muscle fibers from control patient did not exhibit immunoreactivity for iNOS whereas a cytoplasmic staining for iNOS was observed in DMD muscle fibers. No immunostaining of connective tissue or vessels was seen in DMD. (C) NADPH-d reactivity in the muscle of control and DMD patients. Transverse serial cryostat sections (10-µm thick) of a deltoid muscle from control (left column) and DMD (right column) patients stained by HE (upper row) and for NADPH-d reactivity (lower row). NADPH-d reactivity was lower in control patients compared to DMD patients. In DMD patients, NADPH-d reactivity was predominantly found in the sarcoplasm where it showed a spotty distribution. No reactivity was shown in the connective tissue. Some vessels exhibited NADPH-reactivity. Bar: 80 µm.