Butyrate epigenetically modifies satellite cells

Histone deacetylase (HDAC) inhibitors have proven to be effective at enhancing the SC program, but how they alter their epigenetic landscape remains undetermined. Our objective was to determine how an HDAC inhibitor, butyrate, promotes myogenic differentiation. Neonatal piglets were fed a standard milk replacer or one supplemented with 0.5% spray dried Tributyrin (TB), a butyrate pro-drug. Satellite cells from TB supplemented piglets showed a decrease in the expression of EZH2 (enhancer of zeste homolog 2), the methyltransferase subunit of the PRC2 (polycomb repressive complex 2). SC ChIP-Seq analysis revealed that TB treatment resulted in a global reduction of the H3K27me3 repressive chromatin mark, specifically around those sites associated with genes involved in myogenic differentiation, negative regulation of cell proliferation, and micro-RNAs normally upregulated during differentiation. SC were transfected with siRNA targeting EZH2, treated with 0.5mM butyrate, or both to determine if this was the primary mechanism through which butyrate alters SC myogenesis. Treatment with butyrate significantly reduced the expression of genes associated with satellite cell proliferation 24h after treatment, while EZH2 ablation had no effect on the expression of these transcription factors. Immunostaining revealed that EZH2 depletion resulted in an increase in differentiating SC, but not myotube hypertrophy. These results indicate that while EZH2 ablation may force myogenic differentiation, butyrate may operate through a parallel mechanism to enhance the myogenic program. These findings support the notion that butyrate may be used to enhance muscle growth and could prove useful at treating myopathic conditions.