Duchenne muscular dystrophy is driven by defective membrane repair and annexin-A2 dysregulation in skeletal muscle

Abstract

BackgroundDuchenne muscular dystrophy (DMD) is caused by mutations in the DMD gene, which encodes dystrophin in skeletal muscle cells. Although the role of dystrophin as a structural protein is well known, the cellular processes underlying myofiber degeneration are still not fully understood. Despite advances from studies in murine models, these models do not fully replicate the human pathology. MethodsWe investigated sarcolemmal integrity, membrane repair capacity, and annexin protein expression in DMD patient muscle biopsies and human skeletal muscle cell lines using immunohistochemistry, both shear stress-based and laser irradiation injury assays, western blotting, and live-cell imaging of GFP-tagged annexins. ResultsWe identified defective membrane repair in DMD skeletal muscle cells, independent of increased membrane fragility, by evaluating resealing capacity in control and DMD derived-patient cell lines using both a shear stress assay (N = l2, p < 0.000l) and a laser irradiation assay (N = 3, p < 0.000l). Analyses performed on human DMD muscle biopsies (N = l0) further confirmed this defect, demonstrating massive intracellular IgG uptake (p < 0.000l) together with altered annexin expression profiles. While mechanical stress induces the upregulation of annexin A5 (ANXA5, p < 0.0l) and A6 (ANXA6, p < 0.05) in healthy skeletal muscle cells - suggesting an adaptive response to membrane damage, given the annexin familys central role in membrane repair - we observed dysregulated expression patterns of these proteins in DMD cells. Notably, ANXAl (p < 0.05) and ANXA2 (p < 0.0l) were not only significantly overexpressed but also aberrantly localized to the extracellular space, a putative consequence of defective membrane repair. Since extracellular ANXA2 has been associated with adipocyte accumulation in the muscle tissue of patients with dysferlinopathy, a similar pathological mechanism may be at play in DMD. ConclusionsOur findings propose that ANXA2 contributes to muscle degeneration in DMD and highlight it as a potential therapeutic target to prevent adipogenesis and muscle loss.