The cell volume changes induced by hypotonic electrolyte and sucrose solutions were studied in Chinese-hamster-ovary epithelial cells. The effects in the solutions with osmolarities between 32 and 315 mosM/L and distilled water were analyzed using bright-field and fluorescence confocal microscopy. The changes of the cell volume, accompanied by the detachment of cells, the formation of blebs, and the occurrence of almost spherical vesicle-like cells (“cell-vesicles”), showed significant differences in the long-time responses of the cells in the electrolyte solutions compared with the sucrose-containing solutions. A theoretical model based on different permeabilities of ions and sucrose molecules and on the action of Na⁺/K⁺-ATPase pumps is applied. It is consistent with the observed temporal behavior of the cells’ volume and the occurrence of tension-induced membrane ruptures and explains lower long-time responses of the cells in the sucrose solutions.

Cell based carriers are increasingly recognized as a good system for cargo delivery to cells. One of the reasons is their biocompatibility and low toxicity compared to artificial systems. Giant plasma membrane vesicles (GPMV) derive from the cell plasma membrane. Thus they offer the closest approximation to it, which makes them good candidates for potential drug delivery systems. To evaluate the applicability of GPMVs as carriers, we analyzed their basic biophysical properties to test their robustness in the face of changeable physiological conditions, as well as their ability to translocate across the membrane into cells. GPMVs formed from human umbilical vein endothelial cells (HUVEC) sustain a drastic osmotic challenge (50–500 mOsmoL/kg) unlike giant unilamelar vesicles (GUVs). In hyper-osmotic solutions the average volume decreases and membrane invaginations form, while in the hypo-osmolar buffer the volume of GPMVs increases and these changes were not reversible. The membranes of flaccid GPMVs started to wrinkle unevenly giving rise to buds after exposure to lipopolysaccharide (LPS). The shape changes in GUVs are reversible in contrast to GPMVs after LPS removal. GPMVs exposed to fluorescent LPS exhibited a signal that remained visible in some GPMVs even after LPS removal, which was never the case with GUVs. Calcein penetrated both into GUVs and GPMVs, however after the removal from the bulk solution some of the GPMVs still exhibited very bright signal, while in GUVs only a weak fluorescent signal was detected. We could also see that practically all GPMVs incorporated dextran initially, but after the dextran solution was changed with the initial non-fluorescent solution it remained only in 20% of them. The majority of HUVEC cells displayed a fluorescent signal after the incubation with GPMVs that contained fluorescently labeled dextran. Our findings indicate that GPMVs behave quite differently from artificially made giant phospholipid vesicles and the changes induced by the different treatments we subjected them to are not reversible. We also demonstrate that different substances can be both loaded into them and delivered into cells, so GPMVs may be of potential use as cargo/therapy delivery systems.

Cytoskeleton-disrupting drugs can have different effects on cell mechanics at different deformation scales. We therefore applied two complementary indentation techniques to study the effects of two actin-disrupting drugs on cellular stiffness of human umbilical vein endothelial cells. Optical tweezers were used to probe the cortical stiffness at small deformations, and atomic force microscopy was used to probe the bulk cell stiffness at larger deformations. The first drug studied was CK-869, which is an inhibitor of the actin branching complex Arp2/3, and has not been analysed yet in terms of mechanical effects. A significant decrease in cell stiffness upon treatment with CK-869 was measured with both techniques, which implies that actin branching is important for cell mechanics at small and large deformations. The second drug studied was jasplakinolide, for which ambiguous effects on cell mechanics have been reported. In line with previous studies, we found that jasplakinolide caused significant cell stiffening at large deformations but slight cell softening under small deformations. This result implies that jasplakinolide has different effects on different levels of actin organization.

Keratin intermediate filaments are the principal structural element of epithelial cells. Their importance in providing bulk cellular stiffness is well recognized, but their role in the mechanics of cell cortex is less understood. In this study, we therefore compared the cortical stiffness of three keratinocyte lines: primary wild type cells (NHEK2), immortalized wild type cells (NEB1) and immortalized mutant cells (KEB7). The cortical stiffness was measured by lateral indentation of cells with AOD-steered optical tweezers without employing any moving mechanical elements. The method was validated on fixed cells and Cytochalasin-D treated cells to ensure that the observed variations in stiffness within a single cell line were not a consequence of low measurement precision. The measurements of the cortical stiffness showed that primary wild type cells were significantly stiffer than immortalized wild type cells, which was also detected in previous studies of bulk elasticity. In addition, a small difference between the mutant and the wild type cells was detected, showing that mutation of keratin impacts also the cell cortex. Thus, our results indicate that the role of keratins in cortical stiffness is not negligible and call for further investigation of the mechanical interactions between keratins and elements of the cell cortex.