Mechano-biomimetic hydrogel 3D cell cultivation as a strategy to improve mammalian cell protein expression

Eukaryotic expression systems are frequently employed for the production of recombinant proteins as therapeutics as well as research tools. Among which mammalian cell protein expression approach is the most powerful one, which can express complex proteins or genetic engineered biological drugs, such as PD-1. However, the high expense, which partially derives from its low protein yielding efficiency, limited the further application of such approach in large scale production of target proteins. To address this issue, we proposed a novel technique to promote the protein production efficiency of mammal cells without using conventional genetic engineered approaches. By placing 293T cells in a hydrogel 3D cell culture platform and adjusting the stress relaxation of the matrix hydrogel, cells formed multicellular spheroids by self-organization. In particular, the multicellular spheroids have a significantly enhanced ability to transiently express multiple proteins (SHH–N, PD-1 and PDL-1). We also examined in detail the mechanism underlying this phenomenon, and found that the reorganization of cytoskeleton during spheroids formation enhances the translation process of protein by recruiting ribosomes. Overall, this finding provides a novel approach for subsequent improvement of large-scale mammalian protein expression cell systems.