The Effects of a NAD(P)H Oxidase Inhibition on Matrix Metalloproteinases and TIMP-1 in the MDX Diaphragm

Duchenne muscular dystrophy (DMD) is characterized by devastating muscle degeneration that includes oxidative stress, loss of contractile tissue, muscle weakness and increased fibrosis in respiratory muscles (e.g., diaphragm). The mdx mice diaphragm undergoes a progressive degeneration similar to that occurring in patients with DMD. We showed that apocynin, a NAD(P)H oxidase inhibitor, protects against reduction in diaphragm mass, oxidative capacity, and apoptosis. We hypothesized that apocynin (1.5mmol/L per day) would attenuate extramyocyte space and collagen content by ameliorating matrix metalloproteinases (e.g., MMP-2, MMP-9), tissue inhibitors of metalloproteinases (e.g., TIMP-1) and transforming growth factor-β (TGF-β) in the mdx diaphragm. Eight to nine week old mdx mice and age-matched C57BL wild-types were divided into 4 groups: wild-type controls + water (WW, n=7); wild-type controls + apocynin (WA, n=7); mdx mice + water (MW, n=7), and mdx mice + apocynin (MA, n=7). After 8 days of treatment, the diaphragm was extracted. Both MMP-2 (-22.6%) and MMP-9 (-27.8%) were lower in MW than WW. TIMP-1 (+61.3 %) and TGF-beta levels (+39.4%) were higher in MW than WW. MMP-2 (-21.8%) and TIMP-1 (-8.5%) levels were significantly decreased with apocynin in the mdx diaphragm. Our findings indicate cell protection of apocynin against fibrosis in the mdx diaphragm by regulating the protein levels of MMP-2/9 and TIMP-1