Collective Mechanical Behavior of Epithelial Cells - The Impact of Micro-Wounding

For epithelia, which are lining organs or blood vessels, the mechanical integrity of a whole cell sheet is indispensable. The question arises, how neighboring cells mechanically work together to build a functioning cell sheet. In order to address this question, epithelial cells were monitored during layer formation and critical culture conditions such as temperature to characterize the cell sheet behavior exposed to different environmental cues. To disturb cell sheet homeostasis, single cells were mechanically manipulated with a new micro-wounding technique which finally leads to a complete cell failure and subsequently a single-cell wound. Via optical microscopy the closure process was followed and an efficient purse string mechanism was found for MDCK II cells. The closure was accompanied by an increase in mechanical pretension, which was detected with atomic force microscopy. The increase in pretension was also found in cells which are not in direct contact with the wound but separated by up to three cell widths from the wounding site. This indicates that also small disturbances such as micro-wounding elicit a coordinated response in cell sheets, most probably mediated by cell junctions. Since the paradigm organ for wounding is the skin, keratinocyte cells were used for which a genetically modified knock out, lacking the entire keratin network, exists. In these knock out cells, junction localization and actin architecture was altered. Functional studies showed that differences in cell-cell coupling and barrier function, measured with electric cell-substrate impedance sensing, occur. Cells lacking the keratin network also showed a fragile layer integrity and an increase in wound size indicating an impaired response to micro-wounding. These changes were attributed to an altered mechanical homeostasis in individual cells and subsequently also the cell sheet. In conclusion, this new approach of micro-wounding demonstrated that even single-cell wounds induce a collective mechanical response in epithelial cell ensembles and that wellcontrolled single-cell mechanical homeostasis is crucial for proper cell sheet function