Single Cell Mechanotransduction and Its Modulation Analyzed by Atomic Force Microscope Indentation

AbstractThe skeleton adapts to its mechanical usage, although at the cellular level, the distribution and magnitude of strains generated and their detection are ill-understood. The magnitude and nature of the strains to which cells respond were investigated using an atomic force microscope (AFM) as a microindentor. A confocal microscope linked to the setup enabled analysis of cellular responses. Two different cell response pathways were identified: one, consequent upon contact, depended on activation of stretch-activated ion channels; the second, following stress relaxation, required an intact microtubular cytoskeleton. The cellular responses could be modulated by selectively disrupting cytoskeletal components thought to be involved in the transduction of mechanical stimuli. The F-actin cytoskeleton was not required for responses to mechanical strain, whereas the microtubular and vimentin networks were. Treatments that reduced membrane tension, or its transmission, selectively reduced contact reactions. Immunostaining of the cell cytoskeleton was used to interpret the results of the cytoskeletal disruption studies. We provide an estimate of the cellular strain magnitude needed to elicit intracellular calcium responses and propose a model that links single cell responses to whole bone adaptation. This technique may help to understand adaptation to mechanical usage in other organs