Scalable production of muscle and adipose cell-laden microtissues using edible macroporous microcarriers for 3D printing of cultured fish fillets

Cellular agriculture is a novel platform for addressing the issues of protein scarcity, environmental pressures, and food safety. However, expanding seed cells at a large scale remains a prerequisite for achieving industrial cultured meat production. We here propose the production of large-pore-sized edible porous microcarriers (EPMs) by using NaCl to precisely control ice crystal growth during cryogenic crosslinking. Muscle satellite cells (SCs) and adipose-derived stem cells (ASCs) from large yellow croakers exhibit remarkable adhesion, proliferation, and differentiation on gelatin-based EPMs. Following consecutive expansion, SCs and ASCs densities reach 6.25 × 10⁵ and 5.77 × 10⁵ cells/mL, leading to a 499-fold and 461-fold increase in cell numbers, respectively. Subsequently, the mature microtissues are incorporated into a bioink, thereby enabling successful three-dimensional printing of cultured fish fillets with sensory properties similar to native fish fillets. EPM-based cell expansion and food structuring techniques are promising in facilitating large-scale cultured fish meat production.