The super-healing MRL strain promotes muscle growth in muscular dystrophy through a regenerative extracellular matrix
Joseph G. O’Brien, Alexander B. Willis, Ashlee M. Long, Jason Kwon, GaHyun Lee, Frank W. Li, Patrick G.T. Page, Andy H. Vo, Michele Hadhazy, Melissa J. Spencer, Rachelle H. Crosbie, Alexis R. Demonbreun, Elizabeth M. McNally
Joseph G. O’Brien, Alexander B. Willis, Ashlee M. Long, Jason Kwon, GaHyun Lee, Frank W. Li, Patrick G.T. Page, Andy H. Vo, Michele Hadhazy, Melissa J. Spencer, Rachelle H. Crosbie, Alexis R. Demonbreun, Elizabeth M. McNally
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Research Article Muscle biology Stem cells

The super-healing MRL strain promotes muscle growth in muscular dystrophy through a regenerative extracellular matrix

Abstract

The Murphy Roths Large (MRL) mouse strain has “super-healing” properties that enhance recovery from injury. In mice, the DBA/2J strain intensifies many aspects of muscular dystrophy, so we evaluated the ability of the MRL strain to suppress muscular dystrophy in the Sgcg-null mouse model of limb girdle muscular dystrophy. A comparative analysis of Sgcg-null mice in the DBA/2J versus MRL strains showed greater myofiber regeneration, with reduced structural degradation of muscle in the MRL strain. Transcriptomic profiling of dystrophic muscle indicated strain-dependent expression of extracellular matrix (ECM) and TGF-β signaling genes. To investigate the MRL ECM, cellular components were removed from dystrophic muscle sections to generate decellularized myoscaffolds. Decellularized myoscaffolds from dystrophic mice in the protective MRL strain had significantly less deposition of collagen and matrix-bound TGF-β1 and TGF-β3 throughout the matrix. Dystrophic myoscaffolds from the MRL background, but not the DBA/2J background, were enriched in myokines like IGF-1 and IL-6. C2C12 myoblasts seeded onto decellularized matrices from Sgcg–/– MRL and Sgcg–/– DBA/2J muscles showed the MRL background induced greater myoblast differentiation compared with dystrophic DBA/2J myoscaffolds. Thus, the MRL background imparts its effect through a highly regenerative ECM, which is active even in muscular dystrophy.

Authors

Joseph G. O’Brien, Alexander B. Willis, Ashlee M. Long, Jason Kwon, GaHyun Lee, Frank W. Li, Patrick G.T. Page, Andy H. Vo, Michele Hadhazy, Melissa J. Spencer, Rachelle H. Crosbie, Alexis R. Demonbreun, Elizabeth M. McNally

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Figure 3

TGF-β gene expression and signaling were downregulated in Sgcg-MRL skeletal muscle compared with Sgcg-D2 muscle.

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TGF-β gene expression and signaling were downregulated in Sgcg-MRL skele...
RNA sequencing was performed on quadriceps muscles from 16-week-old female mice in biological triplicate. Gene expression was normalized to counts per million (CPM). (A) Comparison of Sgcg-MRL versus Sgcg-D2 transcriptomic profiles showed muscle development and differentiation genes to be highly enriched the Sgcg-MRL background (top). In contrast, immune response, extracellular matrix, and TGF-β pathways were highly enriched in the Sgcg-D2 cohort. (B) Clustered heatmap shows reduction of extracellular matrix genes in Sgcg-MRL muscle. (C) Clustered heatmap shows reduction in TGF-β pathway genes in Sgcg-MRL muscle. (D) Comparative log-scale analysis showed substantially higher average expression of Tgfb1 in the Sgcg-D2 cohort (WT-MRL 4.86, Sgcg-MRL 21.3, WT-D2 5.73, and Sgcg-D2 55.9 CPM). (E) Immunoblotting of quadriceps muscles from Sgcg-D2 and Sgcg-MRL mice showed reduced phosphorylated SMAD3 (p-SMAD3) and total SMAD3 in Sgcg-MRL. (F) The ratio of p-SMAD3 to total SMAD3 was quantified and indicated TGF-β signaling was decreased in the MRL background. Data are presented as mean ± SD. *P < 0.05; **P < 0.01; ***P < 0.001; ****P < 0.0001 by 2-way ANOVA (D) or 2-tailed Student’s t test (F).