Mass Spectrometry-Based Identification of Muscle-Associated and Muscle-Derived Proteomic Biomarkers of Dystrophinopathies

Abstract

The optimization of large-scale screening procedures of pathological specimens by genomic, proteomic and metabolic methods has drastically increased the bioanalytical capability for swiftly identifying novel biomarkers of inherited disorders, such as neuromuscular diseases. X-linked muscular dystrophy represents the most frequently inherited muscle disease and is characterized by primary abnormalities in the membrane cytoskeletal protein dystrophin. Mass spectrometry-based proteomics has been widely employed for the systematic analysis of dystrophin-deficient muscle tissues, using patient samples and animal models of dystrophinopathy. Both, gel-based methods and label-free mass spectrometric techniques have been applied in comparative analyses and established a large number of altered proteins that are associated with muscle contraction, energy metabolism, ion homeostasis, cellular signaling, the cytoskeleton, the extracellular matrix and the cellular stress response. Although these new indicators of muscular dystrophy have increased our general understanding of the molecular pathogenesis of dystrophinopathy, their application as new diagnostic or prognostic biomarkers would require the undesirable usage of invasive methodology. Hence, to reduce the need for diagnostic muscle biopsy procedures, more recent efforts have focused on the proteomic screening of suitable body fluids, such as plasma, serum or urine, for the identification of changed concentration levels of muscle-derived peptides, protein fragments or intact proteins. The occurrence of muscular dystrophy-related protein species in biofluids will be extremely helpful for the future development of cost-effective and non-invasive diagnostic procedures. Novel biomarker signatures of dystrophinopathies will be indispensible for the swift evaluation of innovative therapeutic approaches, such as exon skipping, codon-read-through or stem cell therapy.

Simultaneous Pathoproteomic Evaluation of the Dystrophin-Glycoprotein Complex and Secondary Changes in the mdx-4cv Mouse Model of Duchenne Muscular Dystrophy

“…This type of organelle biochemistry usually enables the exploration of low abundance proteins in the hidden proteome [ 20 , 21 , 22 ], which may otherwise be masked by the highly abundant actomyosin assembly in conventional studies of crude muscle homogenates [ 19 ]. Since low copy number proteins may represent important biomarkers of muscular dystrophy [ 68 ], the systematic profiling of the deep proteome may improve our detailed understanding of its pathogenesis [ 69 ]. The success of this subcellular fractionation technique was evaluated by gel electrophoresis and immunoblotting.…”

The Extracellular Matrix Complexome from Skeletal Muscle

“…In the case of X-linked muscular dystrophy, a large number of proteomic studies have surveyed secondary changes down-stream of the primary abnormality in the membrane cytoskeletal protein dystrophin [126]. Muscular dystrophy-related changes affect energy metabolism, cellular signalling, the excitation-contraction-relaxation cycle, the stress response, the cytoskeletal network and the ECM [127]. The recent proteomic profiling of established genetic animal models of dystrophinopathy has revealed a drastic increase in various collagens, proteoglycans and matricellular proteins [83][84][85][86]128,129].…”

Biomarkeridentifizierung und Anwendung bei der Muskeldystrophie Typ Duchenne

“…Bioanalytische Analysen mit einem global ausgerichteten Forschungsansatz haben mithilfe der Flüssigkeitschromatografie, der 2D-Gelelektrophorese und der Massenspektrometrie diskrete Veränderungen bei der zeitlichen Abfolge der Proteinexpression bei der Muskeldystrophie ermittelt (5). Proteomanalysen des erkrankten Muskelgewebes mit Hochdurchsatz basieren meist auf Proteinpräparationen aus groben Gewebeextrakten, werden aber auch vermehrt mit verschiedenen subzellulären Fraktionen mit geringerer Probenkomplexität durchgeführt (6).…”