During mitosis, adherent cells round up, by increasing the tension of the contractile actomyosin cortex while increasing the internal hydrostatic pressure. In the simple scenario of a liquid cell interior, the surface tension is related to the local curvature and the hydrostatic pressure difference by Laplace's law. However, verification of this scenario for cells requires accurate measurements of cell shape. Here, we use wedged micro-cantilevers to uniaxially confine single cells and determine confinement forces while concurrently determining cell shape using confocal microscopy. We fit experimentally measured confined cell shapes to shapes obeying Laplace's law with uniform surface tension and find quantitative agreement. Geometrical parameters derived from fitting the cell shape and the measured force were used to calculate hydrostatic pressure excess and surface tension of cells. We find that HeLa cells increase their internal hydrostatic pressure excess and surface tension from ≈ 40 Pa and 0.2 mNm⁻¹ during interphase to ≈ 400 Pa and 1.6 mNm⁻¹ during metaphase. The method introduced provides a means to determine internal pressure excess and surface tension of rounded cells accurately and with minimal cellular perturbation and should be applicable to characterize the mechanical properties of various cellular systems.

Mammalian cells regulate adhesion by expressing and regulating a diverse array of cell adhesion molecules on their cell surfaces. Since different cell types express distinct sets of cell adhesion molecules, substrate-specific adhesion is cell type- and condition-dependent. Single-cell force spectroscopy is used to quantify the contribution of cell adhesion molecules to adhesion of cells to specific substrates at both the cell and single molecule level. However, the low throughput of single-cell adhesion experiments greatly limits the number of substrates that can be examined. In order to overcome this limitation, segmented polydimethylsiloxane (PDMS) masks were developed, allowing the measurement of cell adhesion to multiple substrates. To verify the utility of the masks, the adhesion of four different cell lines, HeLa (Kyoto), prostate cancer (PC), mouse kidney fibroblast and MDCK, to three extracellular matrix proteins, fibronectin, collagen I and laminin 332, was examined. The adhesion of each cell line to different matrix proteins was found to be distinct; no two cell lines adhered equally to each of the proteins. The PDMS masks improved the throughput limitation of single-cell force spectroscopy and allowed for experiments that previously were not feasible. Since the masks are economical and versatile, they can aid in the improvement of various assays.

Eph receptor (Eph) and ephrin signaling can play central roles in prostate cancer and other cancer types. Exposed to ephrin-A1 PC3 prostate cancer cells alter adhesion to extracellular matrix (ECM) proteins. However, whether PC3 cells increase or reduce adhesion and by which mechanisms they change adhesion to the ECM remains to be characterized. Here, we assay how ephrin-A1 stimulates PC3 cells to adhere to ECM proteins using single-cell force spectroscopy. We find that PC3 cells binding to immobilized ephrin-A1 but not to solubilized ephrin-A1 specifically strengthen adhesion to collagen I. This Eph-ephrin-A1 signaling, which we suppose is based on mechanotransduction, stimulates β₁-subunit containing integrin adhesion via the protein kinase Akt and the guanine nucleotide-exchange factor cytohesin. Inhibiting the small GTPases, Rap1 or Rac1, generally lowered adhesion of PC3 prostate cancer cells. Our finding suggests a mechanism by which PC3 prostate cancer cells exposed to ephrins crosstalk to β₁-integrins and preferably metastasize in bone, a collagen I rich tissue.

Mammalian cells regulate adhesion by expressing and regulating a diverse array of cell adhesion molecules on their cell surfaces. Since different cell types express distinct sets of cell adhesion molecules, substrate-specific adhesion is cell type- and condition-dependent. Single-cell force spectroscopy is used to quantify the contribution of cell adhesion molecules to adhesion of cells to specific substrates at both the cell and single molecule level. However, the low throughput of single-cell adhesion experiments greatly limits the number of substrates that can be examined. In order to overcome this limitation, segmented polydimethylsiloxane (PDMS) masks were developed, allowing the measurement of cell adhesion to multiple substrates. To verify the utility of the masks, the adhesion of four different cell lines, HeLa (Kyoto), prostate cancer (PC), mouse kidney fibroblast and MDCK, to three extracellular matrix proteins, fibronectin, collagen I and laminin 332, was examined. The adhesion of each cell line to different matrix proteins was found to be distinct; no two cell lines adhered equally to each of the proteins. The PDMS masks improved the throughput limitation of single-cell force spectroscopy and allowed for experiments that previously were not feasible. Since the masks are economical and versatile, they can aid in the improvement of various assays.