Dense HeLa cell monolayers remain liquid-like despite strong crowding

Abstract: Collective dynamics in dense cell monolayers are governed by the interplay between crowding and cellular motility. Although increasing density can slow cellular motion and promote glass-like behaviour, the dynamical state of dense HeLa monolayers remains unclear. Here, we combine in vitro time-lapse imaging of HeLa cell monolayers with simulations of a deformable active-cell model to examine how cell density and motility regulate collective relaxation. Within the experimentally accessible density and time ranges, untreated HeLa monolayers remain liquid-like: structural relaxation progressively slows down with increasing density but remains observable throughout the investigated range. Under low-nutrient conditions, cell motility is strongly reduced, and structural relaxation becomes substantially slower. To elucidate the mechanisms underlying these experimental observations, we further performed simulations using a deformable-cell model. The model qualitatively reproduces the density-dependent increase in structural relaxation time and further shows that reducing self-propulsion promotes long-lived caging dynamics at high packing fractions. These results show that dense HeLa monolayers can sustain slow, heterogeneous, yet relaxing collective dynamics under untreated conditions, and indicate that persistent cellular motility is an important factor in maintaining structural relaxation at high density, which may provide insight into the metastatic potential of cancer cells.