Cell-cell and cell-matrix interactions play a crucial role in the complex formation of entire tissues and tissue substructures to achieve a specific spatial cell organization and tissue function. For the creation of bioartificial tissues, the recapitulation of these processes in vitro is essential. Precisely controlling the assembly and disassembly of different cell types presents a major challenge. Cells embedded in a synthetic scaffold, such as polymers, also referred to as hybrid living materials (HLMs), present a promising new approach to generate multicellular entities that recapitulate tissue structures and function.

Here, we present our strategies to graft polymers from the cell surfaces of yeast and mammalian cells with the goal of generating autonomously formed, stimuli-responsive extracellular matrices (ECM). Controlled radical polymerizations, such as atom transfer radical polymerization (ATRP) and reversible addition–fragmentation chain-transfer polymerization (RAFT), including photo- and enzyme-catalyzed variants, have shown high biocompatibility with different cell types  and are used here with water - soluble monomers to form a polymeric ECM. To ensure that the polymer adheres to the cell surface, it is anchored either to lectins (Concanavalin A), which then interact with the cell membrane via glycan binding or directly grafted from the cell surface by covalent initiator attachment through N-hydroxy succinimide (NHS) esters. In addition, fluorescent tags are incorporated through co-monomers or click chemistry for in situ visualization and matrix customization.

Furthermore, we started to implement those strategies for mammalian cells  to improve the self-organization of one or multiple cell types into 3D structures. For this purpose, we generated mammalian cell lines expressing different outer membrane-localized horseradish peroxidase (mHRP) fusion proteins. mHRP expression is tightly controlled via the TetOn system. Tetracycline-induced expression, membrane localization, and mHRP activity were successfully confirmed for those cell lines by immunofluorescent stainings and measuring the catalytic activity of mHRP. We further optimized the concentrations of different components of the bioATRP reaction, such as the NHS-BIB initiator, DMSO, Sodium ascorbate (NaAsc), and Poly(ethyleneglycol) methylether methacrylate (PEGMA), to adapt the polymerization protocol for mammalian cells. In addition, we also analyzed whether the toxic effect of high NaAsc concentrations can be countered by adding Sodium pyruvate.