In this paper, we show for the first time that impaired myotube formation on type I collagen scaffolds can be enhanced by modification with laminin and entactin, two extracellular proteins from the satellite cell niche. This offers a promising approach for regenerative medicine approaches to heal skeletal muscle.
In skeletal muscle, the stem cell niche is important for controlling the quiescent, proliferation and differentiation states of satellite cells, which are key for skeletal muscle regeneration after wounding. It has been shown that type I collagen, often used as 3D-scaffolds for regenerative medicine purposes, impairs myoblast differentiation. This is most likely due to the absence of specific extracellular matrix proteins providing attachment sites for myoblasts and/or myotubes. In this study we investigated the differentiation capacity of primary murine myoblasts on type I collagen films either untreated or modified with elastin, laminin, type IV collagen, laminin/entactin complex, combinations thereof, and Matrigel as a positive control. Additionally, increased reactive oxygen species (ROS) and ROCK signaling might also be involved. To measure ROS levels with live-cell microscopy, fibronectin-coated glass coverslips were additionally coated with type I collagen and Matrigel onto which myoblasts were differentiated. On type I collagen-coated coverslips, myotube formation was impaired while ROS levels were increased. However, anti-oxidant treatment did not enhance myotube formation. ROCK inhibition, which generally improve cellular attachment to uncoated surfaces or type I collagen, enhanced myoblast attachment to type I collagen-coated coverslips and -films, but slightly enhanced myotube formation. Only modification of type I collagen films by Matrigel and a combination of laminin/entactin significantly improved myotube formation. Our results indicate that type I collagen scaffolds can be modified by satellite cell niche factors of which specifically laminin and entactin enhanced myotube formation. This offers a promising approach for regenerative medicine purposes to heal skeletal muscle wounds.
The above study was a collaboration of the three workgroups of the Dept. of Biochemistry: Membrane Biochemistry, Matrix Biochemistry and Biochemistry of Integrated Systems.
Impaired primary mouse myotube formation on crosslinked type I collagen films is enhanced by laminin and entactin
Sander Grefte, Merel Adjobo-Hermans, Elly Versteeg, Werner Koopman, Willeke Daamen (photo up)
Acta Biomater. S1742-7061-30186-0, 2015
Sander Grefte Merel Adjobo Elly Versteeg Werner Koopman
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