Recent studies suggest that the expression and activity of the histone acetyltransferase p300 are upregulated in catabolic muscle allowing for acetylation of cellular proteins. The function of transcription factors is influenced by posttranslational modifications, including acetylation. It is not known if transcription factors involved in the regulation of muscle mass are acetylated in atrophying muscle. We determined cellular levels of acetylated C/EBPβ, C/EBPδ, FOXO1, FOXO3a, and NF-kB/p65 in dexamethasone-treated L6 muscle cells, a commonly used in vitro model of muscle wasting. The role of p300 in dexamethasone-induced transcription factor acetylation and myotube atrophy was examined by transfecting muscle cells with p300 siRNA. Treatment of L6 myotubes with dexamethasone resulted in increased cellular levels of acetylated C/EBPβ and δ, FOXO1 and 3a, and p65. Downregulation of p300 with p300 siRNA reduced acetylation of transcription factors and decreased dexamethasone-induced myotube atrophy and expression of the ubiquitin ligase MuRF1. The results suggest that several muscle wasting-related transcription factors are acetylated supporting the concept that posttranslational modifications of proteins regulating gene transcription may be involved in the loss of muscle mass. The results also suggest that acetylation of the transcription factors is at least in part regulated by p300 and plays a role in glucocorticoid-induced muscle atrophy. Targeting molecules that regulate acetylation of transcription factors may help reduce the impact of muscle wasting.

The purpose of this study was to compare the effects of various regimens of pyridostigmine administration on the contractile strength of skeletal muscle. Rats were exposed to pyridostigmine according to 3 dosing schedules: 2 mg/kg ip daily, 5 mg/kg X d by sc infusion, and 25 mg/kg X d by sc infusion. After 1, 4, 10, and 20 d of exposure, measurements were made of muscle tension during tetanic stimulation, and of muscle mass, erythrocyte acetylcholinesterase activity, and body weight. Pyridostigmine produced a dose-dependent decrement in the contracture generated during tetanic stimulation. Peak effect was observed after 4 d of exposure but remained depressed after 20 d. The magnitude of the decrement correlated with the frequency of the tetanic stimulation (from 20 to 100 Hz). Muscle tension at the end of the tetanic stimulus was affected to a greater extent than the initial tension. The 25-mg/kg infusion of pyridostigmine significantly depressed erythrocyte acetylcholinesterase activity throughout the study and also decreased body weight on d 1-4. No change in muscle mass was observed in any treatment group. These results show that pyridostigmine exposure results in decrements in skeletal muscle contracture that are dose-dependent, frequency-dependent, and time-dependent. The effect is probably not the result of muscle wasting and does not correlate well with erythrocyte acetylcholinesterase inhibition. The results are consistent with effects of pyridostigmine, both presynaptically and postsynaptically, at the neuromuscular junction that affect neurotransmitter release and receptor responsiveness.

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