A 3D-printable gelatin/alginate/ε-poly-l-lysine hydrogel scaffold to enable porcine muscle stem cells expansion and differentiation for cultured meat development.

The mass proliferation of seed cells and imitation of meat structures remain challenging for cell-cultured meat production. With excellent biocompatibility, high water content and porosity, hydrogels are frequently-studied materials for anchorage-dependent cell scaffolds in biotechnology applications. Herein, a scaffold based on gelatin/alginate/ε-Poly-l-lysine (GAL) hydrogel is developed for skeletal muscle cells, which has a great prospect in cell-cultured meat production. In this work, the hydrogel GAL-4:1, composed of gelatin (5 %, w/v), alginate (5 %, w/v) and ε-Poly-l-lysine (molar ratio vs. alginate: 4:1) is selected as cell scaffold based on Young's modulus of 11.29 ± 1.94 kPa, satisfactory shear-thinning property and suitable porous organized structure. The commercially available C2C12 mouse skeletal myoblasts and porcine muscle stem cells (PMuSCs), are cultured in the 3D-printed scaffold. The cells show strong ability of attachment, proliferation and differentiation after induction, showing high biocompatibility. Furthermore, the cellular bioprinting is performed with GAL-4:1 hydrogel and freshly extracted PMuSCs. The extracted PMuSCs exhibit high viability and display early myogenesis (desmin) on the 3D scaffold, suggesting the great potential of GAL hydrogel as 3D cellular constructs scaffolds. Overall, we develop a novel GAL hydrogel as a 3D-printed bioactive platform for cultured meat research.