Effects of prednisolone on the dystrophin-associated proteins in the blood–brain barrier and skeletal muscle of dystrophic mdx mice

Abstract

The mdx mouse, the most widely used animal model of Duchenne muscular dystrophy (DMD), develops a seriously impaired blood-brain barrier (BBB). As glucocorticoids are used clinically to delay the progression of DMD, we evaluated the effects of chronic treatment with a-methyl-prednisolone (PDN) on the expression of structural proteins and markers in the brain and skeletal muscle of the mdx mouse. We analyzed the immunocytochemical and biochemical expression of four BBB markers, including endothelial ZO-1 and occludin, desmin in pericytes, and glial fibrillary acidic protein (GFAP) in glial cells, and the expression of the short dystrophin isoform Dp 71, the dystrophin-associated proteins (DAPs), and aquaporin-4 (AQP4) and a-b dystroglycan (DG) in the brain. We evaluated the BBB integrity of mdx and PDN-treated mdx mice by means of intravascular injection of horseradish peroxidase (HRP). The expression of DAPs was also assessed in gastrocnemius muscles and correlated with utrophin expression, and laminin content was measured in the muscle and brain. PDN treatment induced a significant increase in the mRNA and protein content of the BBB markers; a reduction in the phosphorylation of occludin in the brain and of AQP4/b DG in both tissues; an increase of Dp71 protein content; and an increase of both mRNA and protein levels of the AQP4/a-b DG complex. The latter was associated with enhanced laminin and utrophin in the muscle. The HRP assay demonstrated functional restoration of the BBB in the PDN-treated mdx mice. Specifically, mdx mice showed extensive perivascular labeling due to escape of the marker, while HRP was exclusively intravascular in the PDN-treated mice and the controls. These data illustrate for the first time that PDN reverses the BBB alterations in the mdx mouse and re-establishes the proper expression and phosphorylation of b-DG in both the BBB and skeletal muscle. Further, PDN partially protects against muscle damage. The reduction in AQP4 and occludin phosphorylation, coupled with their anchoring to glial and endothelial membranes in PDN-treated mice, suggests that the drug may target the glial and endothelial cells. Our results suggest a novel mechanism for PDN action on cerebral and muscular function, restoring the link between DAPs and the extracellular matrix, most likely through protein kinase inactivation.

Galectin-1 Protein Therapy Prevents Pathology and Improves Muscle Function in the mdx Mouse Model of Duchenne Muscular Dystrophy

“…These include neuronal loss, neurofibrillary tangles and changes in the blood-brain barrier. Recent studies show prednisone treatments are able to alleviate blood-brain barrier fragility in mdx mice 45 . Our results show rMsGal-1 could be detected in the brain of mdx mice after treatment indicating the exogenous protein was able to cross from the blood compartment into the brain parenchyma.…”

Therapies that are available and under development for Duchenne muscular dystrophy: What about lung function?

“…Basic research studies conducted in animal models of Duchenne muscular dystrophy have shown that prednisone enhances alpha‐7 integrin and laminin alpha 2 expressions in cultured myogenic cells . Increasing functional glucosylation of α‐dystroglycan, promoting the coupling of dystrophin‐associated proteins with extracellular matrix (glial and endothelial membranes) and inducing the expression of genes coding for proteins which mediate myofiber repair (eg, annexins A1 and A6) provide additional pathophysiological background for the effects of steroids . Furthermore, recent research has identified a potential role of L‐Arg/NO in the pathogenesis of this disorder, since its impaired function results in endothelial dysfunction, affects negatively the perfusion of contracting muscles and leads to ischemia and hypoxia.…”

“…At present, therapy with steroids (deflazacort and prednisone) is the only pharmacological treatment that has proved to have a beneficial effect on muscle strength, prolonging ambulation, and preventing scoliosis . Animal‐based research has demonstrated that the principle underlying mechanism of the steroids effect on muscle tissue includes targeting molecular pathways involved in the process of sarcolemmal recovery after injury, as well as in the link between dystrophin‐associated proteins and the extracellular matrix . From a clinical perspective, although long‐term benefits of glucocorticoid therapy have not yet been widely investigated, there is evidence that their use is associated with a decline in the rate of the loss of clinically significant motor milestones and decreased risk of death …”

“…9 Animal-based research has demonstrated that the principle underlying mechanism of the steroids effect on muscle tissue includes targeting molecular pathways involved in the process of sarcolemmal recovery after injury, as well as in the link between dystrophin-associated proteins and the extracellular matrix. 10,11 From a clinical perspective, although long-term benefits of glucocorticoid therapy have not yet been widely investigated, there is evidence that their use is associated with a decline in the rate of the loss of clinically significant motor milestones and decreased risk of death. 12 On the other hand, the impact of steroids on lung function has not been studied systematically, while any respiratory benefits from treatment with steroids may be confounded by concurrent adverse effects (eg, weight gain and osteoporosis).…”

Clinical aspects of patients with sarcoglycanopathies under steroids therapy

“…Steroid treatment with prednisone or deflazacort has been shown to slow disease progression in Duchenne muscular dystrophy (DMD) effective to delay restrictive respiratory insufficiency, scoliosis and in a lesser extent cardiac involvement in patients with this disease 8,[12][13][14] . The mechanisms of this improvement are not completely clear but steroids may have the potential benefits of inhibiting muscle proteolysis, stabilizing fiber membrane, reducing intracellular calcium concentration, and stimulating myoblast proliferation [15][16][17] . Albeit controversial, steroid therapy has also been sporadically used to treat other forms of muscular dystrophy.…”