N-acetylglucosamine facilitates coordinated flow-like movement of myoblasts, forming a foundation for efficient...

Skeletal muscle comprises 30-40% of a mammal's body mass, maintaining its integrity through efficient muscle fiber regeneration, which involves myoblast differentiation into myotubes. Previously, we reported that N-acetylglucosamine (GlcNAc) promotes myogenesis in C2C12 cells, although the underlying processes remained unclear. GlcNAc's activated form, UDP-GlcNAc, is critical for the biosynthesis of highly branched (N-acetyllactosamine-rich) N-linked oligosaccharides, which are recognized by galectin-3 (Gal-3), a protein that facilitates dynamic cell-cell and cell-matrix interactions and modulating the motility dynamics of membrane-associated proteins.

In this study, we used primary myoblasts from both wild-type and Gal-3 null (Gal-3KO) mice, observing myotube formation through long-term live-cell imaging and single-cell tracking to reveal the dynamic process that occurred during the myotube formation.

We found that GlcNAc enhances myoblast fusion in a dose-dependent manner, and that the addition of Gal-3 with GlcNAc leads to the formation of larger myotubes. Gal-3KO myoblasts exhibited a reduced capacity for myotube formation-a deficiency that was rectified by supplementing with GlcNAc and Gal-3. Our results highlight the role of Gal-3 interaction with oligosaccharides, whose synthesis is promoted by GlcNAc in facilitating myotube formation. Single-cell tracking revealed that GlcNAc and Gal-3 increase myoblast motility, leading to a faster, coordinated, flow-like movement-a collective behavior, along which myotubes form through cell fusion. Interestingly, myoblasts contributing to myotube formation were pre-positioned along the eventual shape of the myotubes before this flow-like movement was fully established. These myoblasts moved along the flow, paused, and even moved against it, suggesting that both coordinated flow and initial spatial positioning contribute to myoblast alignment along the axis of future myotubes.

Our findings suggest that GlcNAc, in conjunction with Gal-3, enhances myotube formation by fostering an environment conducive to myoblast positioning, establishing optimal coordinated flow-like movement, and facilitating fusion. This suggests potential therapeutic applications of GlcNAc in muscle repair and muscle disorders.