Myotube/Adipocyte Powder-Enriched Alginate-Zein Hydrogels Support Myotube Alignment for 3D Myoblast Culture

Recent advances in cultured-meat research emphasize the development of edible scaffolds that promote myogenic differentiation. Nonetheless, many materials provide only structural support and do not replicate native muscle or serve as alternatives to muscle-adipocyte co-culture, highlighting the need for cytocompatible, tissue-specific scaffolds. This study aimed to develop a composite alginate-zein (Algi/zein) hydrogel enriched with myotube (MP) and adipocyte (AP) powders to provide a structural, biochemical, and potentially cultured-meat hydrogel. Algi/zein hydrogels enriched with myotube (MP) and adipocyte (AP) powders were fabricated and evaluated for structural, cellular, and biochemical properties using C2C12 myoblasts cultured in 2D and 3D environments. Metabolite profiling was performed to evaluate the biochemical features. MP/AP incorporation generated extra cellular matrix (ECM)-like microstructures and significantly enhanced myotube alignment in Algi/zein scaffolds compared with MP/AP-free controls, increasing the proportion of axially aligned fibers by up to ~6-fold at a 1:1 AP:MP ratio. Organized myosin expression was observed, while metabolomic profiling indicated partial biochemical similarity to beef. Incorporating MP and AP into Algi/zein hydrogels enhanced myotube alignment and showed partial structural and biochemical similarity to native muscle tissue.