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Proteomic profiling of liver tissue from the mdx-4cv mouse model of Duchenne muscular dystrophy.

ABSTRACT: Background:Duchenne muscular dystrophy is a highly complex multi-system disease caused by primary abnormalities in the membrane cytoskeletal protein dystrophin. Besides progressive skeletal muscle degeneration, this neuromuscular disorder is also associated with pathophysiological perturbations in many other organs including the liver. To determine potential proteome-wide alterations in liver tissue, we have used a comparative and mass spectrometry-based approach to study the dystrophic mdx-4cv mouse model of dystrophinopathy. Methods:The comparative proteomic profiling of mdx-4cv versus wild type liver extracts was carried out with an Orbitrap Fusion Tribrid mass spectrometer. The distribution of identified liver proteins within protein families and potential protein interaction patterns were analysed by systems bioinformatics. Key findings on fatty acid binding proteins were confirmed by immunoblot analysis and immunofluorescence microscopy. Results:The proteomic analysis revealed changes in a variety of protein families, affecting especially fatty acid, carbohydrate and amino acid metabolism, biotransformation, the cellular stress response and ion handling in the mdx-4cv liver. Drastically increased protein species were identified as fatty acid binding protein FABP5, ferritin and calumenin. Decreased liver proteins included phosphoglycerate kinase, apolipoprotein and perilipin. The drastic change in FABP5 was independently verified by immunoblotting and immunofluorescence microscopy. Conclusions:The proteomic results presented here indicate that the intricate and multifaceted pathogenesis of the mdx-4cv model of dystrophinopathy is associated with secondary alterations in the liver affecting especially fatty acid transportation. Since FABP5 levels were also shown to be elevated in serum from dystrophic mice, this protein might be a useful indicator for monitoring liver changes in X-linked muscular dystrophy.

SUBMITTER: Murphy S

PROVIDER: S-EPMC6205794 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Proteomic profiling of liver tissue from the <i>mdx</i>-<i>4cv</i> mouse model of Duchenne muscular dystrophy.

Murphy Sandra S Zweyer Margit M Henry Michael M Meleady Paula P Mundegar Rustam R RR Swandulla Dieter D Ohlendieck Kay K

Clinical proteomics 20181029

<h4>Background</h4>Duchenne muscular dystrophy is a highly complex multi-system disease caused by primary abnormalities in the membrane cytoskeletal protein dystrophin. Besides progressive skeletal muscle degeneration, this neuromuscular disorder is also associated with pathophysiological perturbations in many other organs including the liver. To determine potential proteome-wide alterations in liver tissue, we have used a comparative and mass spectrometry-based approach to study the dystrophic ...[more]

PMID: 30386187

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Project description:BACKGROUND:X-linked muscular dystrophy is a primary disease of the neuromuscular system. Primary abnormalities in the Dmd gene result in the absence of the full-length isoform of the membrane cytoskeletal protein dystrophin. Besides progressive skeletal muscle wasting and cardio-respiratory complications, developmental cognitive deficits and behavioural abnormalities are clinical features of Duchenne muscular dystrophy. In order to better understand the mechanisms that underlie impaired brain functions in Duchenne patients, we have carried out a proteomic analysis of total brain extracts from the mdx-4cv mouse model of dystrophinopathy. RESULTS:The comparative proteomic profiling of the mdx-4cv brain revealed a significant increase in 39 proteins and a decrease in 7 proteins. Interesting brain tissue-associated proteins with an increased concentration in the mdx-4cv animal model were represented by the glial fibrillary acidic protein GFAP, the neuronal Ca(2+)-binding protein calretinin, annexin AnxA5, vimentin, the neuron-specific enzyme ubiquitin carboxyl-terminal hydrolase isozyme L1, the dendritic spine protein drebrin, the cytomatrix protein bassoon of the nerve terminal active zone, and the synapse-associated protein SAP97. Decreased proteins were identified as the nervous system-specific proteins syntaxin-1B and syntaxin-binding protein 1, as well as the plasma membrane Ca(2+)-transporting ATPase PMCA2 that is mostly found in the brain cortex. The differential expression patterns of GFAP, vimentin, PMCA2 and AnxA5 were confirmed by immunoblotting. Increased GFAP levels were also verified by immunofluorescence microscopy. CONCLUSIONS:The large number of mass spectrometrically identified proteins with an altered abundance suggests complex changes in the mdx-4cv brain proteome. Increased levels of the glial fibrillary acidic protein, an intermediate filament component that is uniquely associated with astrocytes in the central nervous system, imply neurodegeneration-associated astrogliosis. The up-regulation of annexin and vimentin probably represent compensatory mechanisms involved in membrane repair and cytoskeletal stabilization in the absence of brain dystrophin. Differential alterations in the Ca(2+)-binding protein calretinin and the Ca(2+)-pumping protein PMCA2 suggest altered Ca(2+)-handling mechanisms in the Dp427-deficient brain. In addition, the proteomic findings demonstrated metabolic adaptations and functional changes in the central nervous system from the dystrophic phenotype. Candidate proteins can now be evaluated for their suitability as proteomic biomarkers and their potential in predictive, diagnostic, prognostic and/or therapy-monitoring approaches to treat brain abnormalities in dystrophinopathies.

Project description:Duchenne muscular dystrophy is due to genetic abnormalities in the dystrophin gene and represents one of the most frequent genetic childhood diseases. In the X-linked muscular dystrophy (mdx) mouse model of dystrophinopathy, different subtypes of skeletal muscles are affected to a varying degree albeit the same single base substitution within exon 23 of the dystrophin gene. Thus, to determine potential muscle subtype-specific differences in secondary alterations due to a deficiency in dystrophin, in this study, we carried out a comparative histological and proteomic survey of mdx muscles. We intentionally included the skeletal muscles that are often used for studying the pathomechanism of muscular dystrophy. Histological examinations revealed a significantly higher degree of central nucleation in the soleus and extensor digitorum longus muscles compared with the flexor digitorum brevis and interosseus muscles. Muscular hypertrophy of 20-25% was likewise only observed in the soleus and extensor digitorum longus muscles from mdx mice, but not in the flexor digitorum brevis and interosseus muscles. For proteomic analysis, muscle protein extracts were separated by fluorescence two-dimensional (2D) gel electrophoresis. Proteins with a significant change in their expression were identified by mass spectrometry. Proteomic profiling established an altered abundance of 24, 17, 19 and 5 protein species in the dystrophin-deficient soleus, extensor digitorum longus, flexor digitorum brevis and interosseus muscle, respectively. The key proteomic findings were verified by immunoblot analysis. The identified proteins are involved in the contraction-relaxation cycle, metabolite transport, muscle metabolism and the cellular stress response. Thus, histological and proteomic profiling of muscle subtypes from mdx mice indicated that distinct skeletal muscles are differentially affected by the loss of the membrane cytoskeletal protein, dystrophin. Varying degrees of perturbed protein expression patterns in the muscle subtypes from mdx mice may be due to dissimilar downstream events, including differences in muscle structure or compensatory mechanisms that counteract pathophysiological processes. The interosseus muscle from mdx mice possibly represents a naturally protected phenotype.

Dataset Information

Proteomic profiling of liver tissue from the mdx-4cv mouse model of Duchenne muscular dystrophy.

Publications

Proteomic profiling of liver tissue from the <i>mdx</i>-<i>4cv</i> mouse model of Duchenne muscular dystrophy.

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