(607f) Biomechanics Study of Endothelial Cellular Membranes Under Hypo-Osmotic Challenge

Cell volume regulation is a fundamental property of all mammalian cells. Numerous signaling pathways are known to be activated by cell swelling and to contribute to cell volume homeostasis. Cellular biomechanics and membrane tension have long been proposed to couple cell swelling to signaling pathways, however, the impact of swelling on these parameters has yet to be fully elucidated. In this study, we utilize atomic force microscopy (AFM) under isotonic and hypo-osmotic conditions to measure the mechanical properties of human aortic endothelial membranes. From AFM force/displacement curves, we obtain estimates of (i) membrane stiffness, which reflects the stiffness of the sub-membrane cytoskeleton complex, and (ii) the force required for membrane tether formation, reflecting membrane tension and membrane-cytoskeleton attachment.

We find that hypo-osmotic swelling results in significant stiffening of the membrane region of endothelial cells, without a change in membrane tension/membrane-cytoskeleton attachment. Furthermore, depolymerization of F-actin in the cytoskeleton, which, as expected, results in a dramatic decrease in the cellular elastic modulus of both the membrane and the deeper cytoskeleton, indicating a collapse of the cytoskeleton scaffold, does not abrogate swelling-induced stiffening of the membrane. Instead, this stiffening is enhanced. We propose that the hypo-osmotically induced membrane stiffening should be attributed to an increase in hydrostatic pressure that results from an influx of solutes and water into the cells. Most importantly, our results suggest that increased hydrostatic pressure, rather than changes in membrane tension, could be responsible for activating volume-sensitive mechanisms in hypo-osmotically swollen cells.