Nitrosative stress elicited by nNOSµ delocalization inhibits muscle force in dystrophin‐null mice

Abstract

The mechanism of force reduction is not completely understood in Duchenne muscular dystrophy (DMD), a dystrophin-deficient lethal disease. Nitric oxide regulates muscle force. Interestingly, neuronal nitric oxide synthase μ (nNOSμ), a major source of muscle nitric oxide, is lost from the sarcolemma in DMD muscle. We hypothesize that nNOSμ delocalization contributes to force reduction in DMD. To test this hypothesis, we generated dystrophin/nNOSμ double knockout mice. Genetic elimination of nNOSμ significantly enhanced force in dystrophin-null mice. Pharmacological inhibition of nNOS yielded similar results. To further test our hypothesis, we studied δ-sarcoglycan-null mice, a model of limb-girdle muscular dystrophy. These mice had minimal sarcolemmal nNOSμ delocalization and muscle force was less compromised. Annihilation of nNOSμ did not improve their force either. To determine whether nNOSμ delocalization itself inhibited force, we corrected muscle disease in dystrophin-null mice with micro-dystrophins that either restored or did not restore sarcolemmal nNOSμ. Similar muscle force was obtained irrespective of nNOSμ localization. Additional studies suggest that nNOSμ delocalization selectively inhibits muscle force in dystrophin-null mice via nitrosative stress. In summary, we have demonstrated for the first time that nitrosative stress elicited by nNOSμ delocalization is an important mechanism underlying force loss in DMD.

nNOS regulation of skeletal muscle fatigue and exercise performance

“…While nNOSμ deficiency does not impact muscle strength in nNOS knockouts, the loss of cytosolic nNOSμ expression increases muscle strength in dystrophin-deficient mdx skeletal muscle (Li et al 2011a). This result suggests a negative gainof-function for cytoplasmic nNOSμ in a dystrophic environment.…”

“…nNOSμ-mediated RyR1 hypernitrosylation was observed in mdx mice and correlated with muscle weakness (Bellinger et al 2009;Li et al 2011a;Li et al 2011b). nNOSμ depletion restored RyR1 nitrosylation to basal levels and enhanced muscle strength (Li et al 2011a), suggesting that aberrant activation of cytosolic nNOSμ was leading to excessive nitrosylation of RyR1 and, consequently, suppressing muscle strength. RyR1 hypernitrosylation was also observed in mdx mice (Bellinger et al 2009).…”

“…However, in human muscle approximately 75% of nNOSμ is associated with the sarcolemmal DGC, suggesting species-specific differences in nNOSμ distribution (Chang et al 1996). A pool of nNOSμ is associated with the ryanodine receptor 1 Ca 2+ release channel (RyR1) at the sarcoplasmic reticulum (SR) Ca 2+ store where it can S-nitrosylate and regulate RyR1 channel activity (Xu et al 1999;Stamler and Meissner 2001;Salanova et al 2008;Li et al 2011a, Figure 1). nNOSμ association and regulation of the ryanodine receptor is also seen in the heart (Gonzalez et al 2007).…”

A Simplified Immune Suppression Scheme Leads to Persistent Micro-dystrophin Expression in Duchenne Muscular Dystrophy Dogs

“…The proviral cis plasmid (YL196) for the R16-17/H3/DC lDys vector was based on the canine dystrophin gene. This microgene is modeled according to a previously described R16-17/DC micro-dystrophin gene (Lai et al, 2009;Banks et al, 2010;Li et al, 2011). It contains the dystrophin N-terminal domain, hinge 1, spectrin-like repeat 1, 16 and 17 (R1, R16, and R17), hinge 3, R24, hinge 4, and the cysteine-rich (CR) domain.…”

Molecular dissection of dystrophin identifies the docking site for nNOS

“…Thus, DMD muscles that lack functional dystrophin, and subsequently have mis-localized nNOS, become susceptible to both mechanical damage and exacerbating blood-flow restrictions (called functional ischemia). Although the mislocalization of nNOS in DMD was described in 1995, the importance of this phenomenon was not fully appreciated until more recently, and the Duan laboratory has led the charge (2,7,(11)(12)(13)(14). This most recent report provides the most detailed characterization yet of the structural elements in dystrophin required for proper nNOS localization and function.…”