Context. In times where human kind is facing serious challenges due to global warming, scarcity of natural resources and inequality, to continue producing food, especially meat, as it is now done, does not seem sustainable. For people that want to continue enjoying their delicious piece of steak, the manner in which meat is sourced will need to be redesigned. One of the disruptive initiatives in this field is in vitro cultured meat. For the cultivation of muscle tissue, one needs cells, chemical factors and the appropriate biomaterials
that function as a scaffold. Meatable B.V. is a food-technology startup that is a pioneer in the cultivation of steak-like meat using induced pluripotent stem cells. Aims. The aim of the study is to optimise the extracellular matrix (i.e. scaffold) mimicry of natural biomaterials to serve as a scaffold for 3D skeletal muscle tissue engineering, using iPSCs. The supplementary aim of this study is to determine the feasibility of the optimised scaffold as a proof of concept for cultured meat. Methods. The proof of concept model (Model A) was created in collaboration with researchers from the Loughborough University who demonstrated a scalable model for 3D human skeletal muscle tissue engineering. Their strategy was to use a hard plastic mould to confine and provide initial support for the viscoelastic hydrogel (matrigel and rat tail collagen) encapsulated with cells (C2C12 myoblasts). For the optimisation of the extracellular matrix for iPSCs for the production of cultured meat, this model was copied and repeated with different edible scaffold alternatives. The best alternative biomaterial (Model B) was further optimised in large scale
tissue moulds and compared to the proof of concept model and meat. Lastly, Model C was created by seeding iPSCs in Model B. The qualitative and quantitative comparisons were based on different analytical parameters.
Results. The hydrogel scaffolds of both models A and B were highly comparable in terms of permeability, scaffold compaction by cell activity and cell alignment. The improved model (B) resulted in a higher cell proliferation as seen by cell development and cell density. The stiffness of model A was half the stiffness of Model B, and the stiffness of Model B was more comparable with the higher
stiffness of beef steak (factor 5 difference). Nevertheless, when the improved model was seeded with pre-differentiated iPSCs (Model C) instead of C2C12 myoblasts, the results were clearly less successful than the first models and were not comparable with beef steak. Conclusions. This research describes the successful improvements of an existing 3D in-vitro skeletal tissue engineering method, as prototype model for cultured meat, that was for the first time seeded with iPSCs. Model B should be set as the basis model to optimise for the culture of iPSCs. The bovine collagen model with C2C12s was not comparable to steak, but a longer culture period could result in more comparable tissue. The main reason for the limitations of Model C was the maturation of iPSCs, which could be improved in different ways. In addition, there remain certain requirements to be met for the hydrogel scaold, mainly connected with current developments for recombinant animal-free materials. It can still be concluded that the less successful Model C was a step in the right direction to becoming a feasible model for the culture of in vitro meat.