RESUMEN
The integration of intramuscular fat-or marbling-into cultured meat will be critical for meat texture, mouthfeel, flavor, and thus consumer appeal. However, culturing muscle tissue with marbling is challenging since myocytes and adipocytes have different media and scaffold requirements for optimal growth and differentiation. Here, we present an approach to engineer multicomponent tissue using myogenic and adipogenic microtissues. The key innovation in our approach is the engineering of myogenic and adipogenic microtissues using scaffolds with customized physical properties; we use these microtissues as building blocks that spontaneously adhere to produce multicomponent tissue, or marbled cultured meat. Myocytes are grown and differentiated on gelatin nanofiber scaffolds with aligned topology that mimic the aligned structure of skeletal muscle and promotes the formation of myotubes in both primary rabbit skeletal muscle and murine C2C12 cells. Pre-adipocytes are cultured and differentiated on edible gelatin microbead scaffolds, which are customized to have a physiologically-relevant stiffness, and promote lipid accumulation in both primary rabbit and murine 3T3-L1 pre-adipocytes. After harvesting and stacking the individual myogenic and adipogenic microtissues, we find that the resultant multicomponent tissues adhere into intact structures within 6-12 h in culture. The resultant multicomponent 3D tissue constructs show behavior of a solid material with a Young's modulus of â¼ 2 ± 0.4 kPa and an ultimate tensile strength of â¼ 23 ± 7 kPa without the use of additional crosslinkers. Using this approach, we generate marbled cultured meat with â¼ mm to â¼ cm thickness, which has a protein content of â¼ 4 ± 2 g/100 g that is comparable to a conventionally produced Wagyu steak with a protein content of â¼ 9 ± 4 g/100 g. We show the translatability of this layer-by-layer assembly approach for microtissues across primary rabbit cells, murine cell lines, as well as for gelatin and plant-based scaffolds, which demonstrates a strategy to generate edible marbled meats derived from different species and scaffold materials.