Engineering Three-Dimensional Cellular Organization by Regulating Bound Water-Mediated Cell–Substrate Interactions for Disease Modeling

Recently, organoid culture technologies have advanced rapidly as a method for constructing biomimetic models for studying tissue and organ physiology. However, reproducibly constructing complex organoid architectures using synthetic polymers, which are potential alternatives to conventional natural polymer gels derived from animal sources, remains challenging. Controlling cell-substrate interactions is crucial for regulating reproducible three-dimensional (3D) cell adhesion behaviors. In our previous study, we demonstrated that interactions between cells and substrates can be modulated by altering the hydrated water content of polymer coatings using poly-(2-methoxyethyl acrylate) (PMEA) derivatives. Based on these findings, we hypothesized that further increasing the hydrated water content of the coating polymer may induce a transition in the cell adhesion behavior from two-dimensional (2D) to 3D configurations. In this study, we focused on PMEA derivatives and synthesized a series of block copolymers with varying ethylene glycol (EG) side-chain lengths to control the bound water content on the substrate surface. Using atomic force microscopy-based single-cell force spectroscopy, we quantitatively demonstrated that increasing the bound water content of the surface polymer weakens cell-substrate interactions. This, in turn, enhances the relative contribution of cell-cell interactions and promotes 3D cell adhesion. Furthermore, we successfully used these polymer substrates to establish an in vitro model of metabolic dysfunction-associated steatohepatitis (MASH). This model successfully reproduced pathological features such as lipid accumulation and elevated expression of inflammatory cytokines interleukin-6 (IL-6) and interleukin-1β (IL-1β), thereby demonstrating their utility for drug screening applications. Our results present a novel strategy for guiding 3D cellular organization by modulating the hydrated water content of polymer coatings. These findings also suggested that PMEA derivatives are promising materials for 3D organoid engineering and disease modeling platforms.