Molecular Mechanism of Sphingosine-1-phosphate Action in Duchenne Muscular Dystrophy

Abstract

Duchenne Muscular Dystrophy (DMD) is a lethal muscle wasting disease. In Drosophila, a genetic increase of the bioactive lipid Sphingosine-1-Phosphate (S1P) and oral delivery of 2-Acetyl-5-tetrahydroxybutyl imidazole (THI), an S1P lyase inhibitor, suppresses dystrophic muscle degeneration. In dystrophic mice, up regulation of S1P by THI increases muscle force, fiber size and reduces fibrosis as well as fat deposition. S1P can exert extracellular and intracellular mode of function, acting as a ligand for mammalian S1P receptors and as a repressor for HDAC1 and HDAC2 activities. Since Drosophila lacks orthologues for S1P receptors and because muscular dystrophies correlate with increased HDAC2 protein levels, we tested whether the common molecular mechanism for the S1P effect in muscle involves HDAC inhibition. Here, we show that total nuclear HDAC activity, HDAC2 protein levels and HDAC2 activity are increased in several mdx muscle types. Importantly, THI-treatment significantly increased nuclear S1P and decreased HDAC activity in injured mdx TA muscles. In un-injured mdx mice, a one-month THI treatment similarly increased muscle nuclear S1P levels, decreased HDAC activity and furthermore, increased H3K9, K18 and H2BK12 acetylation. In addition, we observed an increased expression of HDAC2 target genes. These effects correlate with reduced white blood cell number and diaphragm fibrosis in addition to an increased normal Titin pattern in the sarcomeres. We also demonstrate a significant decrease in gene expression of inflammation-related genes and an increase in metabolic genes; especially genes involved in fatty acid metabolism in THI-treated adductor muscles by using a microarray-based gene expression analysis. Finally, we show an increase in cellular S1P levels and a functional increase in mitochondrial activity, especially in fatty acid oxidation after THI treatment of differentiating C2C12 cells. This result supports our proposal that THI treatment is beneficial for the energy metabolism of mdx muscles.