Project description:Duchenne muscular dystrophy (DMD) is an X-linked recessive disease caused by deleterious mutations in the DMD gene, rendering non-functional forms or complete absence of the protein dystrophin. Eccentric contraction-induced force loss is the most robust and reproducible phenotype of dystrophin-deficient skeletal muscle, yet the molecular mechanisms underlying force loss remain obscure. To this end, we utilized the mdx mouse model of DMD, which displays extreme sensitivity to eccentric contractions. An existing mouse line from our lab that overexpresses cytoplasmic gamma-actin specifically in skeletal muscle (mdx/Actg1-TG) was shown to significantly protect mdx muscle against contraction-induced force loss. To understand the mechanism behind this protection, we performed iTRAQ proteomics on mdx/Actg1-TG tibialis anterior (TA) muscle versus non-transgenic littermate controls to identify differentially-expressed proteins that may afford protection upon gamma-actin overexpression.
Project description:Despite over 3,000 articles published on dystrophin in the last 15 years, the reasons underlying the progression of the human disease, differential muscle involvement, and disparate phenotypes in different species are not understood. The present experiment employed a screen of 12,488 mRNAs in 16-wk-old mouse mdx muscle at a time when the skeletal muscle is avoiding severe dystrophic pathophysiology, despite the absence of a functional dystrophin protein. A number of transcripts whose levels differed between the mdx and human Duchenne muscular dystrophy were noted. A fourfold decrease in myostatin mRNA in the mdx muscle was noted. Differential upregulation of actin-related protein 2/3 (subunit 4), beta-thymosin, calponin, mast cell chymase, and guanidinoacetate methyltransferase mRNA in the more benign mdx was also observed. Transcripts for oxidative and glycolytic enzymes in mdx muscle were not downregulated. These discrepancies could provide candidates for salvage pathways that maintain skeletal muscle integrity in the absence of a functional dystrophin protein in mdx skeletal muscle. Keywords: other
Project description:In this study, Pax7-Cre mediated inactivation of Sirt6 in mdx mice resulted in profound improvement of the mdx phenotype at the functional level. To study the underlying molecular mechanisms we performed RNA-seq of muscles from control, mdx and Sirt6mKO/mdx mice.
Project description:In this study, Pax7-Cre mediated inactivation of Sirt6 in mdx mice resulted in profound improvement of the mdx phenotype at the functional level. To study the underlying molecular mechanisms we performed RNA-seq of MuSCs from control, mdx and Sirt6mKO/mdx mice.
Project description:In this study, in order to minimize the genetic variability of muscle samples we have used two approaches. First, we have analyzed the gene expression profile from a single skeletal muscle, the medial gastrocnemius (MG), and not from a pool of different muscles which could have different expression profiles. Second, we have performed the temporal gene expression profiling by extracting the MG muscles of the same individual from the both legs at two different times to minimize the inter-individual genetic variability. The MG muscle represent an excellent candidate for biopsy due to its easy accessible by surgery and also because its biopsy is well tolerated by the animals allowing us to perform another later biopsy in the other leg to obtain two MG samples of the same individual at two different times. Moreover, MG muscle is composed of approximately 20-30% type I (red) fibers and 70-80% type II (white) fibers {Ariano MA, 1973}, {Simard C, 1988}, {Zhan WZ, 1992} and therefore is more representative of skeletal muscle tissue in general than a muscle composed exclusively by red or white fibers.<br><br><br><br>The transcript expression profiles in MG muscles from mdx and wild-type mice were analyzed at 3 weeks, 1.5 months and 3 months of life by using the 430 2.0 gene chips from Affymetrix (n=3 for each condition). The differentially expressed transcripts which showed differences ?1.5-fold were obtained by performing three different comparisons: 1) genes differentially expressed in mdx compared with controls at each point in time (additional file 1); 2) temporal analysis of the genes differentially expressed in mdx mice between the three points in time also compared with the variations in control mice (additional file 2); and 3) temporal analysis of the genes differentially expressed in control mice between the three points in time also compared with the variations in mdx mice (additional file 3). The first comparison that we performed, by comparing the gene expression between mdx and control mice at every point in time, was similar to that performed in previous longitudinal studies {Porter JD, 2003}, {Rouger K, 2002}, {Turk R, 2005}. However, the other two comparisons were directed to elucidate the genes that are varying throughout the period of time analyzed in every mice strain, and therefore we obtained on the one hand the genes that vary in mdx mice but not in wild-type, and on the other hand the genes that vary in control animals but remain unchanged in mdx mice between the times analyzed. To present the results in a more comprehensive form, all the genes were classified in seven different categories: Cell adhesion & extracellular matrix; Proteolysis; Muscle structure & regeneration; Inflammation & immune response; Cell signaling & cell communication; Metabolism; and Others/unknown. The resulting genes from our study were classified in their functional categories using information from Affymetrix (www.affymetrix.com) and from the Gene Ontology database accessible in the Jackson Laboratory Mouse Genome Informatics website (www.informatics.jax.org).<br><br>