Heparin-modified aligned collagen scaffolds enhance in vitro myogenesis

Biomaterial-based skeletal muscle tissue engineering approaches have largely focused on mimicking the 3D aligned architecture of native muscle, which is critical for guiding myotube formation and force transmission. In contrast, fewer studies incorporate glycosaminoglycan (GAG)-mediated biochemical cues despite their known role in regulating myogenesis and growth factor sequestration. In this study, we develop aligned collagen-GAG (CG) scaffolds using directional freeze-drying and systematically vary GAG content by incorporating GAGs of increasing sulfation levels (hyaluronic acid, chondroitin sulfate, and heparin). While all scaffold variants support myoblast adhesion, metabolic activity, and myotube alignment, heparin-modified collagen scaffolds significantly enhance myoblast metabolic activity and myogenic differentiation as measured by myosin heavy chain (MHC) expression and myotube size. We additionally show that heparin-modified scaffolds sequester and retain significantly higher levels of insulin-like growth factor-1 (IGF-1), a potent promoter of myogenesis, compared to other scaffold groups. Together, these results highlight the importance of optimizing GAG content in CG scaffolds for targeted applications and underscore the promise of heparin-modified CG scaffolds as a material platform for skeletal muscle tissue engineering.