Important Role of Heparan Sulfate in Skeletal Muscles

[Speaker] Takeo Yoshikawa:1
[Co-author] Asuka Mogi:1, Yamato Miura:1, Takuro Matsuzawa:1, Kazuhiko Yanai:1
1:Pharmacology, Tohoku University Graduate School of Medicine, Japan

Heparan sulfate (HS) is a sulfated linear polysaccharide at the cell surface and in the extracellular matrix. HS plays an important role in various physiological processes such as development and angiogenesis. Recent studies revealed that lack of HS induced autism-like behaviors and chondrosarcoma, indicating the pathophysiological involvement of HS in various disorders. Previous studies showed the existence of HS in skeletal muscles, although the roles of HS in these tissues remain unclear.
First, we examined the role of HS in the differentiation of muscle cells using C2C12 cells, a mouse myoblast cell line. CRISPR/CAS9 technology was used to delete Ext1, which encodes a heparan sulfate synthase, to generate a stable HS-deleted C2C12 cell line. HS deletion dramatically impaired myoblast differentiation but not proliferation, demonstrating the essential role of HS in myoblast differentiation. In order to confirm the importance of HS in skeletal muscle in vivo, we created skeletal muscle-specific Ext1 knockout mice by Cre-loxP system (cKO). Body weight was decreased in cKO and 2-year survival rate was significantly lower in cKO. Righting reflex was sluggished and locomotor activity in home cages was reduced in cKO, indicating HS depletion impaired motor activity. Muscle weakness of cKO was also confirmed in treadmill tests and wire hang tests. HE staining of skeletal muscles revealed that the cross sectional areas of quadriceps femoris, gastrocnemius and soleus muscles were significantly smaller in cKO. Electromicroscopic observation showed that myofibrils were thinner in cKO. Finally, we examined muscle differentiation after muscle injury by BaCl2 injection to tibialis anterior muscle (TA). Myosin heavy chain expression, one of the marker proteins for muscle differentiation, was significantly decreased in cKO TA. These results demonstrate that HS plays an important role in skeletal muscle, especially in differentiation. Furthermore, HS might be a target for treatment strategies of muscle atrophy such as aging, sarcopenia and muscle dystrophy.

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