sirolimus and Muscular-Dystrophy--Animal

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

A-type lamins, generated from the LMNA gene by differential splicing, are type V intermediate filament proteins that polymerize to form part of the nuclear lamina, and are of considerable medical interest because missense mutations in LMNA give rise to a wide range of dystrophic and progeroid syndromes. Among these are dilated cardiomyopathy and two forms of muscular dystrophy (limb-girdle and Emery-Dreifuss), which are modeled in lmna (-/-) mice and mice engineered to express human disease mutations. Our recent study demonstrates that cardiac and skeletal muscle pathology in lmna (-/-) mice can be attributed to elevated MTORC1 signaling leading to impairment of autophagic flux. An accompanying paper from another laboratory shows similar impairments in mice engineered to express the LMNA H222P associated with dilated cardiomyopathy in humans and also in left ventricular tissue from human subjects. MTORC1 inhibition with rapalogs restores autophagic flux and improves cardiac function in both mouse models, and extends survival in the lmna (-/-) mice. These findings elaborate a potential treatment option for dilated cardiomyopathy and muscular dystrophy associated with LMNA mutation and supplement growing evidence linking impaired autophagy to human disease.

Topics: Animals; Autophagy; Cardiomyopathy, Dilated; Disease Models, Animal; Humans; Lamin Type A; Mechanistic Target of Rapamycin Complex 1; Mice; Mice, Knockout; Multiprotein Complexes; Muscular Dystrophy, Animal; Proteins; Signal Transduction; Sirolimus; TOR Serine-Threonine Kinases; Up-Regulation

Mammalian target of rapamycin (mTOR) is a key regulator of cell growth that associates with raptor and rictor to form the mTOR complex 1 (mTORC1) and mTORC2, respectively. Raptor is required for oxidative muscle integrity, whereas rictor is dispensable. In this study, we show that muscle-specific inactivation of mTOR leads to severe myopathy, resulting in premature death. mTOR-deficient muscles display metabolic changes similar to those observed in muscles lacking raptor, including impaired oxidative metabolism, altered mitochondrial regulation, and glycogen accumulation associated with protein kinase B/Akt hyperactivation. In addition, mTOR-deficient muscles exhibit increased basal glucose uptake, whereas whole body glucose homeostasis is essentially maintained. Importantly, loss of mTOR exacerbates the myopathic features in both slow oxidative and fast glycolytic muscles. Moreover, mTOR but not raptor and rictor deficiency leads to reduced muscle dystrophin content. We provide evidence that mTOR controls dystrophin transcription in a cell-autonomous, rapamycin-resistant, and kinase-independent manner. Collectively, our results demonstrate that mTOR acts mainly via mTORC1, whereas regulation of dystrophin is raptor and rictor independent.

Topics: Adaptor Proteins, Signal Transducing; Age Factors; Animals; Carrier Proteins; Cells, Cultured; Dystrophin; Electroporation; Energy Metabolism; Enzyme Activation; Female; Glucose; Glycogen; Mice; Mice, Inbred C57BL; Mice, Knockout; Mitochondria, Muscle; Muscle Contraction; Muscle, Skeletal; Muscular Dystrophy, Animal; Mutation; Oxidation-Reduction; Phosphotransferases (Alcohol Group Acceptor); Proto-Oncogene Proteins c-akt; Rapamycin-Insensitive Companion of mTOR Protein; Rats; Regulatory-Associated Protein of mTOR; Severity of Illness Index; Sirolimus; TOR Serine-Threonine Kinases; Transduction, Genetic; Utrophin

Topics: Animals; Azathioprine; Cyclosporine; Graft Survival; Immunosuppression Therapy; Immunosuppressive Agents; Methylprednisolone; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Muscles; Muscular Dystrophy, Animal; Mycophenolic Acid; Polyenes; Sirolimus; Transplantation, Homologous