vendex and Muscular-Dystrophy--Animal

Src tyrosine kinase (TK), a redox-sensitive protein overexpressed in dystrophin-deficient muscles, can contribute to damaging signaling by phosphorylation and degradation of β-dystroglycan (β-DG). We performed a proof-of-concept preclinical study to validate this hypothesis and the benefit-safety ratio of a pharmacological inhibition of Src-TK in Duchenne muscular dystrophy (DMD). Src-TK inhibitors PP2 and dasatinib were administered for 5 weeks to treadmill-exercised mdx mice. The outcome was evaluated in vivo and ex vivo on functional, histological and biochemical disease-related parameters. Considering the importance to maintain a proper myogenic program, the potential cytotoxic effects of both compounds, as well as their cytoprotection against oxidative stress-induced damage, was also assessed in C2C12 cells. In line with the hypothesis, both compounds restored the level of β-DG and reduced its phosphorylated form without changing basal expression of genes of interest, corroborating a mechanism at post-translational level. The histological profile of gastrocnemius muscle was slightly improved as well as the level of plasma biomarkers. However, amelioration of in vivo and ex vivo functional parameters was modest, with PP2 being more effective than dasatinib. Both compounds reached appreciable levels in skeletal muscle and liver, supporting proper animal exposure. Dasatinib exerted a greater concentration-dependent cytotoxic effect on C2C12 cells than the more selective PP2, while being less protective against H

Topics: Animals; Cell Line; Cell Survival; Dasatinib; Dystroglycans; Liver; Male; Mice, Inbred mdx; Muscle Fatigue; Muscle Strength; Muscle, Skeletal; Muscular Dystrophy, Animal; Muscular Dystrophy, Duchenne; Protein Kinase Inhibitors; Pyrimidines; Reproducibility of Results; src-Family Kinases; Torque

Topics: Adaptation, Physiological; Adenylate Kinase; Animals; Diaphragm; Disease Models, Animal; Male; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Inbred mdx; Muscle Contraction; Muscle Strength; Muscle Weakness; Muscle, Skeletal; Muscular Dystrophy, Animal; Muscular Dystrophy, Duchenne; Physical Conditioning, Animal; Torque; Up-Regulation

Secondary dystroglycanopathies are a subset of muscular dystrophy caused by abnormal glycosylation of α-dystroglycan (αDG). Loss of αDG functional glycosylation prevents it from binding to laminin and other extracellular matrix receptors, causing muscular dystrophy. Mutations in a number of genes, including FKTN (fukutin), disrupt αDG glycosylation.. We analyzed conditional Fktn knockout (Fktn KO) muscle for levels of mTOR signaling pathway proteins by Western blot. Two cohorts of Myf5-cre/Fktn KO mice were treated with the mammalian target of rapamycin (mTOR) inhibitor rapamycin (RAPA) for 4 weeks and evaluated for changes in functional and histopathological features.. Muscle from 17- to 25-week-old fukutin-deficient mice has activated mTOR signaling. However, in tamoxifen-inducible Fktn KO mice, factors related to Akt/mTOR signaling were unchanged before the onset of dystrophic pathology, suggesting that Akt/mTOR signaling pathway abnormalities occur after the onset of disease pathology and are not causative in early dystroglycanopathy development. To determine any pharmacological benefit of targeting mTOR signaling, we administered RAPA daily for 4 weeks to Myf5/Fktn KO mice to inhibit mTORC1. RAPA treatment reduced fibrosis, inflammation, activity-induced damage, and central nucleation, and increased muscle fiber size in Myf5/Fktn KO mice compared to controls. RAPA-treated KO mice also produced significantly higher torque at the conclusion of dosing.. These findings validate a misregulation of mTOR signaling in dystrophic dystroglycanopathy skeletal muscle and suggest that such signaling molecules may be relevant targets to delay and/or reduce disease burden in dystrophic patients.

Topics: Animals; Biomechanical Phenomena; Disease Models, Animal; Down-Regulation; Dystroglycans; Electric Stimulation; Female; Genetic Predisposition to Disease; Glycosylation; Male; Mice, Knockout; Muscle Contraction; Muscle Strength; Muscle, Skeletal; Muscular Dystrophy, Animal; Myogenic Regulatory Factor 5; Phenotype; Protein Kinase Inhibitors; Protein Processing, Post-Translational; Proteins; Proto-Oncogene Proteins c-akt; Signal Transduction; Sirolimus; Time Factors; TOR Serine-Threonine Kinases; Torque; Transferases