A21 Assessment of satellite progenitor cell differentiation in hd skeletal muscle in vitro

Abstract

Background Intracellular calcium (Ca2+) transients regulate all aspects of cell function, including excitability, motility, exocytosis and gene expression. Abnormal Ca2+ signalling has been previously linked with Huntington’s disease (HD) neuropathology, however, how and to what extent it affects muscle tissue is unknown. Ghrelin is the gut-peptide hormone that induces intracellular Ca2+ mobilization by binding to ghrelin receptors in the skeletal muscle. Ghrelin may potentially modify HD muscle phenotype, thus be of relevance for HD treatment. Aim Our main goal is to establish a novel analytical platform to characterize intracellular Ca2+ dyshomeostasis and to assess the effect of ghrelin on Ca2+ dynamics in myofibers. Methods To assess intracellular Ca2+ transients in R6/2 mouse muscle cells and in clinical HD and control myoblasts in vitro, we isolate satellite cells from R6/2 and WT gastrocnemius muscle at postnatal days 7-9 and treat them with ghrelin for 3 days at the proliferation stage. Following 7 days of differentiation, we assess myofiber diameter, and gene expression by mRNA and protein levels of muscle-specific targets in response to ghrelin treatment. Results Compared to WT satellite cells, R6/2 satellite cells exhibit reduced myofiber diameter in vitro. MyoD (a protein which plays a crucial role in muscle differentiation) level is significantly decreased in R6/2 mice. Ghrelin treatment promotes myofiber formation, as ghrelin-treated satellite cells from R6/2 gastrocnemius muscle exhibit increased myofiber diameter and MyoD protein levels compared to the control group. Conclusion Our preliminary data suggest that treatment with ghrelin delays muscle atrophy in R6/2 mice at early stages of HD disease/phenotype. To understand its role in Ca2+ homeostasis, we will analyze intracellular Ca2+ signal patterns and correlate this with the morphological shape descriptors in satellite cells from both R6/2 mice and HD patients.