Proteome Profiling of the Dystrophic mdx Mice Diaphragm

Abstract

Mdx mice with a spontaneous mutation in exon 23 of the Dmd gene represent the most common model to investigate the pathophysiology of Duchenne muscular dystrophy (DMD). The disease, caused by the lack of functional dystrophin, is characterized by irreversible impairment of muscle functions, with the diaphragm affected earlier and more severely than other skeletal muscles. We applied a label-free (LF) method and the more thorough tandem mass tag (TMT)-based method to analyze differentially expressed proteins in the diaphragm of 6-week-old mdx mice. The comparison of both methods revealed 88 commonly changed proteins. A more in-depth analysis of the TMT-based method showed 953 significantly changed proteins, with 867 increased and 86 decreased in dystrophic animals (q-value < 0.05, fold-change threshold: 1.5). Consequently, several dysregulated processes were demonstrated, including the immune response, fibrosis, translation, and programmed cell death. Interestingly, in the dystrophic diaphragm, we found a significant decrease in the expression of enzymes generating hydrogen sulfide (H2S), suggesting that alterations in the metabolism of this gaseous mediator could modulate DMD progression, which could be a potential target for pharmacological intervention.

Cardioprotective Effects of Hydrogen Sulfide and Its Potential Therapeutic Implications in the Amelioration of Duchenne Muscular Dystrophy Cardiomyopathy

“…Of note, when we performed a proteomic analysis of the diaphragm of younger, 5-6-week-old animals, a lower abundance of CTH was found. This effect was confirmed through real-time and Western blot analyses, and additionally, we detected the decreased mRNA of Cbs and Mpst, as well as a reduced MPST protein level [156]. In line with these results, both dystrophic Caenorhabditis elegans (dys-1(eg33) mutant) [188,189] and mouse models [162,187,190] of DMD have demonstrated the usefulness of H 2 S donors in attenuating the severity of the disease.…”

“…During DMD progression, increased levels of fibrotic markers, like periostin (POSTN) and/or osteopontin (OPN), have been noted. A high abundance of POSTN, the nonstructural matricellular protein that functions upstream and downstream of TGF-β, was found in the dystrophic diaphragm [156,157] as well as in the heart [158]. Additionally, it was strongly induced in other models of cardiac impairment, such as myocardial infarction and left ventricular pressure overload [159,160].…”

Changes to the Autophagy-Related Muscle Proteome Following Short-Term Treatment with Ectoine in the Duchenne Muscular Dystrophy Mouse Model MDX

“…An important autophagy substrate selector is LC3, of which altered activation has been reported in MDX (Table 1) [40][41][42][43][44][45][46][47]. We observed significantly increased LC3II/I ratios in EDL muscle of 26weeks-old MDX compared to age-matched healthy controls.…”

How Can Proteomics Help to Elucidate the Pathophysiological Crosstalk in Muscular Dystrophy and Associated Multi-System Dysfunction?

“…Proteins with a significant change in abundance were identified in a variety of skeletal muscle types [ 264 ]. The most prominent and reproducibly identified proteins include the adenylate kinase isoform AK1, annexins, small heat shock proteins, desmin, vimentin, tubulins, collagens, calsequestrin, B-type lamin, myoferlin, dysferlin, ferritin, carbonic anhydrase isoform CA3, the fatty acid-binding protein FABP3 and various contractile proteins [ 134 , 148 , 180 , 232 , 266 , 267 , 268 , 269 , 270 , 271 , 272 , 273 , 274 , 275 , 276 , 277 , 278 , 279 , 280 , 281 , 282 , 283 , 284 , 285 , 286 , 287 , 288 , 289 , 290 , 291 , 292 ]. Independent verification analyses using comparative immunoblotting, enzyme assays, histochemistry and immunofluorescence microscopy were employed to confirm proteome-wide changes in dystrophic skeletal muscles [ 172 ].…”