Abstract
Backgroundγ-cytoplasmic (γ-cyto) actin levels are elevated in dystrophin-deficient mdx mouse skeletal muscle. The purpose of this study was to determine whether further elevation of γ-cyto actin levels improve or exacerbate the dystrophic phenotype of mdx mice.MethodsWe transgenically overexpressed γ-cyto actin, specifically in skeletal muscle of mdx mice (mdx-TG), and compared skeletal muscle pathology and force-generating capacity between mdx and mdx-TG mice at different ages. We investigated the mechanism by which γ-cyto actin provides protection from force loss by studying the role of calcium channels and stretch-activated channels in isolated skeletal muscles and muscle fibers. Analysis of variance or independent t-tests were used to detect statistical differences between groups.ResultsLevels of γ-cyto actin in mdx-TG skeletal muscle were elevated 200-fold compared to mdx skeletal muscle and incorporated into thin filaments. Overexpression of γ-cyto actin had little effect on most parameters of mdx muscle pathology. However, γ-cyto actin provided statistically significant protection against force loss during eccentric contractions. Store-operated calcium entry across the sarcolemma did not differ between mdx fibers compared to wild-type fibers. Additionally, the omission of extracellular calcium or the addition of streptomycin to block stretch-activated channels did not improve the force-generating capacity of isolated extensor digitorum longus muscles from mdx mice during eccentric contractions.ConclusionsThe data presented in this study indicate that upregulation of γ-cyto actin in dystrophic skeletal muscle can attenuate force loss during eccentric contractions and that the mechanism is independent of activation of stretch-activated channels and the accumulation of extracellular calcium.
Highlights
Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disease caused by mutations in the dystrophin gene
We previously reported that g-cyto actin levels are elevated fivefold in dystrophin-deficient skeletal muscle [23,25,26] and that muscle-specific ablation of g-cyto actin does not exacerbate the dystrophic condition [26]
Levels of g-cyto actin in mdx-TG skeletal muscle SDS extracts were analyzed alongside a purified g-cyto actin calibration curve on SDS-PAGE gels and blotted with g-cyto actin-specific antibodies
Summary
Duchenne muscular dystrophy (DMD) is a severe muscle-wasting disease caused by mutations in the dystrophin gene. Dystrophin localizes primarily to costameres, where it links the cortical actin cytoskeleton to the sarcolemma and the extracellular matrix [1]. It is part of the dystrophin-glycoprotein complex (DGC), a large oligomeric complex of proteins thought primarily to Dystrophin binds to filamentous actin with high affinity via two distinct actin-binding domains: an N-terminal tandem calponin homology domain and a region of basic spectrin-like repeats in the middle rod domain [1]. Despite restoration of DGC members at the sarcolemma, at least one actin-binding domain is essential for partial or complete rescue of the dystrophic phenotype in transgenic mdx mice [4,5,6]
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