Solid tumors exhibit elevated stiffness and growth-induced solid stress, imposing significant physical confinement on individual cells. Despite these compressive forces, tumor cells continue to proliferate, driving overall tumor expansion. What biophysical and biochemical mechanisms enable cells to grow and divide under such restrictive mechanical conditions? To address this question, we use a multicellular breast cancer spheroid model to mimic solid tumor growth, culturing spheroids either on top of hydrogels with minimal constraint or embedded within hydrogels of tunable stiffness and viscoelasticity to impose varying degrees of confinement. We quantified stress within spheroids by incorporating polyacrylamide microbeads and tracking their deformation over time. Substantial solid stress in the kilopascal range was observed and further increased in tumor spheroids under enhanced confinement. Upon release, spheroids exhibited rapid stress relaxation with only a slight increase in overall volume, suggesting that cells were pressurized, potentially through elevated intracellular osmotic pressure. Ion channel perturbation experiments indicated that regulating ion flux can modulate stress build-up. Concurrently, confocal microscopy measurements revealed a progressive decrease in individual cell size. Preliminary results from biosensors and fluorescence microscopy suggest a potential increase in macromolecular crowding within proliferating spheroids. Taken together, these observations point to a model in which cells under confinement may elevate internal osmotic pressure by tuning both ions and macromolecules.
Zhu et al. (Sun,) studied this question.