Novel growth regime of MDCK II model tissues on soft substrates

It is well established that MDCK II cells grow in circular colonies that densify until contact inhibition takes place. Here, we show that this behavior is only typical for colonies developing on hard substrates and report a new growth phase of MDCK II cells on soft gels. At the onset, the new phase is characterized by small, three-dimensional droplets of cells attached to the substrate. When the contact area between the agglomerate and the substrate becomes sufficiently large, a very dense monolayer nucleates in the center of the colony. This monolayer, surrounded by a belt of three-dimensionally packed cells, has a well-defined structure, independent of time and cluster size, as well as a density that is twice the steady-state density found on hard substrates. To release stress in such dense packing, extrusions of viable cells take place several days after seeding. The extruded cells create second-generation clusters, as evidenced by an archipelago of aggregates found in a vicinity of mother colonies, which points to a mechanically regulated migratory behavior.