Agents Which Inhibit NF‐κB Signaling Block Spontaneous Contractile Activity and Negatively Influence Survival of Developing Myotubes

Abstract

Inhibiting the NF-κB signaling pathway provides morphological and functional benefits for the mdx mouse, a model for Duchenne muscular dystrophy characterized by chronic elevations in the nuclear expression of p65, the transactivating component of the NF-κB complex. The purpose of this study was to examine p65 expression in nondystrophic and mdx myotubes using confocal immunofluorescence, and determine whether inhibitors of the NF-κB pathway alter myotube development. Primary cultures of nondystrophic and mdx myotubes had identical levels of nuclear and cytosolic p65 expression and exhibited equivalent responses to TNF-α, thus excluding the hypothesis that the lack of dystrophin is sufficient to induce increases in NF-κB signaling. The NF-κB inhibitors pyrrolidine dithiocarbamate (PDTC) and sulfasalazine decreased spontaneous contractile activity and reduced myotube viability in a dose- and time-dependent manner. Similarly, a vivo-morpholino designed to block translation of murine p65 (m-p65tb-vivomorph1) rapidly abolished spontaneous contractile activity, reduced p65 expression measured by confocal immunofluorescence, and induced cell death in primary cultures of nondystrophic and mdx myotubes. Similar effects on p65 immunofluorescence and cell viability were observed following m-p65tb-vivomorph1 exposure to spontaneously inactive C2C12 myotubes, while exposure to a control scrambled vivo morpholino had no effect. These results indicate a direct role of the NF-κB pathway in myotube development and identify a potential therapeutic limitation to the use of NF-κB inhibitors in treating Duchenne and related muscular dystrophies. J. Cell. Physiol. 231: 788-797, 2016. © 2015 Wiley Periodicals, Inc.

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“…ST2825 is a specific inhibitor of myeloid differentiation factor 88 (MyD88), which inhibits the dimerization of MyD88, and prevents the activation and transduction of the TLR signal, thereby inhibiting the TLR signaling pathway ( 27 ). PDTC is an inhibitor of the NF-κB signaling pathway, preventing degradation of the inhibitor of NF-κB subunit α (IκB), activation of NF-κB and translocation of NF-κB to the nucleus ( 28 ). MβCD has a strong affinity for cholesterol, and is able to remove it from cells, and dysregulate lipid raft integrity and function ( 29 ).…”