J Clin Neurol.  2016 Jul;12(3):351-360. 10.3988/jcn.2016.12.3.351.

Activation of the wnt/β-Catenin Signaling Pathway in Polymyositis, Dermatomyositis and Duchenne Muscular Dystrophy

• Liu F
• Liang Z
• Xu J
• Li W
• Zhao D
• Zhao Y
• Yan C

• KMID: 2354124
• DOI: http://doi.org/10.3988/jcn.2016.12.3.351

Abstract

BACKGROUND AND PURPOSE
The wnt/β-catenin signaling pathway plays a critical role in embryonic development and adult-tissue homeostasis. Recent investigations implicate the importance of wnt/β-catenin signaling in normal wound healing and its sustained activation being associated with fibrogenesis. We investigated the immunolocalization and activation of wnt/β-catenin in polymyositis (PM), dermatomyositis (DM), and Duchenne muscular dystrophy (DMD).
METHODS
Immunofluorescence staining and Western blot analysis of β-catenin were performed in muscle specimens from 6 PM, 8 DM, and 6 DMD subjects. The β-catenin/Tcf4 DNA-binding activity in muscle was studied using an electrophoretic mobility shift assay (EMSA), and serum wnt/β-catenin/Tcf transcriptional activity was measured using a luciferase reporter gene assay.
RESULTS
Immunoreactivity for β-catenin was found in the cytoplasm and nuclei of muscle fibers in PM, DM, and DMD. The protein level of β-catenin was elevated, and EMSA analysis confirmed the activation of wnt/β-catenin signaling. The transcriptional activities of β-catenin/Tcf in the circulation were increased in patients with PM, DM, and DMD, especially in those with interstitial lung disease, and these transcriptional activities decreased when PM or DM patients exhibited obvious clinical improvements.
CONCLUSIONS
Our findings indicate that wnt/β-catenin signaling is activated in PM, DM, and DMD. Its activation in muscle tissue and the circulation may play a role in modulating muscle regeneration and be at least partly involved in the process of muscle and pulmonary fibrosis.

Keyword

β-catenin; dermatomyositis; polymyositis; duchenne muscular dystrophy; muscle biopsy

MeSH Terms

Blotting, Western
Cytoplasm
Dermatomyositis*
Electrophoretic Mobility Shift Assay
Embryonic Development
Female
Fluorescent Antibody Technique
Genes, Reporter
Homeostasis
Humans
Luciferases
Lung Diseases, Interstitial
Muscular Dystrophy, Duchenne*
Polymyositis*
Pregnancy
Pulmonary Fibrosis
Regeneration
Wound Healing
Luciferases

Figure

• 

  Fig. 1 Immunofluorescence staining of β-catenin in skeletal muscle of control subjects (Aa–c) and PM (Ad–f), DM (Ag–i), DMD (Aj–l), and NM (Am–o) patients. Western blot results and semiquantitative analysis of β-catenin and GSK-3β in skeletal muscle of PM, DM, DMD, and NM patients and controls (B–D) (data represent mean±SD values from three separate experiments. *p≤0.01). DM: dermatomyositis, DMD: duchenne muscular dystrophy, GSK-3β: glycogen synthase kinase 3-β, NM: nemaline myopathy, PM: polymyositis.

• 

  Fig. 2 Western blot results (A) and semiquantitative analysis (B) of GSK-3β phosphorylation in skeletal muscle of PM, DM, DMD patients and normal controls (data represent mean±SD values from three separate experiments. *p≤0.01 between PM patients and controls, *p≤0.01). DM: dermatomyositis, DMD: duchenne muscular dystrophy, GSK-3β: glycogen synthase kinase 3-β.

• 

  Fig. 3 Results of EMSA for nuclear extracts from skeletal muscle of PM, DM, DMD, NM patients and normal controls (data represent mean±SD values from three separate experiments. *p≤0.01). DM: dermatomyositis, DMD: duchenne muscular dystrophy, EMSA: electrophoretic mobility shift assay, NM: nemaline myopathy, PM: polymyositis.

• 

  Fig. 4 Relative luciferase activities of the wnt/β-catenin/Tcf pathway reporter constructs 8xTopFlash in HEK293 cells treated with DMEM containing 20% serum from PM, DM, DMD, and NM patients and normal controls (A and B). C: Comparison of relative serum β-catenin/Tcf4 transcriptional activities in PM and DM patients with or without ILD. D: Comparison of relative serum β-catenin/Tcf4 transcriptional activities in PM and DM patients before and after treatment (data represent mean±SD values from three separate experiments. *p≤0.01). DM: dermatomyositis, DMEM: Dulbecco's modified eagle medium, DMD: duchenne muscular dystrophy, PM: polymyositis.

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