Pneumatic equiaxial compression device for mechanical manipulation of epithelial cell packing and physiology

It is well established that mechanical cues, e.g., tensile- compressive- or shear forces, are important co-regulators of cell and tissue physiology. To understand the mechanistic effects these cues have on cells, technologies allowing precise mechanical manipulation of the studied cells are required. As the significance of cell density i.e., packing on cellular behavior is beginning to unravel, we sought to design an equiaxial cell compression device based on our previously published cell stretching system. We focused on improving the suitability for microscopy and the user-friendliness of the system. By introducing a hinge structure to the substrate stretch generating vacuum chamber, we managed to decrease the z-displacement of the cell culture substrate, thus reducing the focal plane drift. The vacuum battery, the mini-incubator, as well as the custom-made vacuum pressure controller make the experimental setup more flexible and portable. Furthermore, we improved the efficiency and repeatability of manufacture of the device by designing a mold that can be used to cast the body of the device. We also compared several different silicone membranes, and chose SILPURAN® due to its best microscopy imaging properties. Here, we show that the device can produce a maximum 8.5% radial pre-strain which leads to a 15% equiaxial areal compression as the pre-strain is released. When tested with epithelial cells, upon compression, we saw a decrease in cell cross-sectional area and an increase in cell layer height. Additionally, before compression the cells had two distinct cell populations with different cross-sectional areas that merged into a more uniform population due to compression. In addition to these morphological changes, we detected an alteration in the nucleo-cytoplasmic distribution of YAP1, suggesting that the cellular packing is enough to induce mechanical signaling in the epithelium.