Edible bird’s nests (EBN)—the nests of swiftlet birds harvested from the wild— are high-end healthcare food in East Asia, while their excessive harvesting poses increasing ecological, environmental, and food safety concerns. Here, we report for the first time a tissue-engineering (TE) approach for fabricating EBNs substitutes by integrating the technologies of three-dimensional (3D) printing and live cell culture. The engineered products, tissue-engineered edible bird’s nests (TeeBN), comprise two layers. The first is a feeding layer that encapsulates epithelial cells in 3D-printed biocompatible gelation scaffolds. These cells secrete bioactive ingredients, e.g., sialic acid and epidermal growth factors (EGF), recapitulating the natural production of these substances by birds. The second is a receiving layer, consisting of foodgrade natural polymers, e.g., polysaccharides, which mimics the building blocks of natural EBNs while biologically stabilizing the factors released from the feeding layer. In vitro characterizations demonstrate that the feeding layer facilitates 3D cell growth and functions, and the receiving layer (as the end product) contains the necessary nutrients expected from natural EBNs—while without harmful substances commonly detected in natural EBNs. Further, in vivo metabolomics studies in mice indicate that TeeBN showed a similar profile of serum metabolites as natural EBN, reflecting comparable nutritional effects. In summary, we innovatively developed a tissue engineering-based substitute for EBNs with comparable metabolic functions and minimized safety risks, opening a new avenue for producing delicacy food from laboratorial cell culture with 3D printing technology.