Résumé :
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The development of more efficacious treatments for Duchenne and related muscular dystrophies would be facilitated by an improved understanding of the mechanism which promotes enhanced nuclear p65 activation in dystrophic skeletal muscle. To accomplish this objective, the status of the classical pathway was examined in cytosolic, nuclear, and whole cell extracts from adult nondystrophic and mdx costal diaphragm. Although absolute p65 activation was increased several fold in nuclear extracts from mdx diaphragm (Trans AM ELISA based assay), the proportion of activated p65 in the nuclear compartment (nuclear/nuclear + cytosolic) was identical in nondystrophic and mdx muscle preparations. Consistent with this observation, cytosolic and whole cell densitometric levels (GAPDH loading control) of I?B-? were not decreased, but were significantly increased in the mdx diaphragm. The proportion of cytosolic phosphorylated I?B-? (Pi- I?B-?/ total I?B-?) was the same in nondystrophic and mdx diaphragm. Whole cell extracts from mdx muscle exhibited significant increases in the expression of both p65 and p50, and increases in the proportion of phosphorylated p65 (Pi – p65/ total p65). The expression of I?B kinase (IKK) ?, ?, and ?, the proportion of phosphorylated IKK? and the expression of SUMO-1 were also significantly increased in mdx muscle. To examine the potential role of the proteasome in regulating cytosolic I?B-?, preparations were exposed to the inhibitor MG132. This treatment to decrease I?B-? degradation significantly increased cytosolic I?B-? levels in nondystrophic, but had no significant effects on cytosolic I?B-? in mdx diaphragm. These results indicate that baseline reductions in cytosolic I?B-? do not occur in dystrophic skeletal muscle, and that increases in the expression of each of the signaling components of the classical NF?B pathway contribute to the increases in nuclear p65 activation. (Supported by AFM, Charley’s Fund, Strategic Research Grant from ATSU)
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