Detection of cellular traction forces by microfabricated silicon force sensors

Paper i proceeding, 2004

To investigate mechanical cell-substrate interactions, we have microfabricated force-sensitive cell adhesion substrates by micropatterning silicon wafers. Sub-cellular forces are measured by imaging independent deflections of vertical micro-cantilevers in the 2D plane of the surface. These substrates model surfaces of different elasticity and porosity, as well as quantify direction and magnitude of sub-cellular forces. (Petronis et al, J. Micromech. Microeng, 2003:13, 900) Silicon microchips have been patterned using photolithography and deep reactive ion etching to produce substrates with embedded cantilevers with varying spatial arrays, dimensions, and therefore sensitivity. Specifically, the organisation of the substrates allows multiple sensors to be placed under each cell at any given time-point, and multiple measurements made for individual cells. SEM and lateral force microscopy have been used to characterize these cantilevers and determine their spring constants. Time-lapse microscopy in differential interference-contrast (DIG) mode with an onstage incubator and immersible objective has been used to observe the behaviour of various large cells at low-seeding densities on untreated substrates. These included human vascular endothelial cells (EC), smooth muscle cells (SMC), fibroblasts, as well as the smaller adult-derived hippocampal progenitor (AHP) cells. In particular, forces at the substrate surface associated with cell morphology during attachment and migration (in the presence or absence of contact guidance) were investigated. Analysis of image sequences with custom written macros has resulted in real time mapping of cantilever deflections by cells, and their force-vectors. The nanoNewton range forces are greatest in the spreading cell periphery; and the leading or trailing edges in migrating cells, where they are parallel to the direction of movement. With live fluorescent membrane labels we can simultaneously image cell and substrate in two separate channels. These studies are currently in progress and the results will be presented. The measurements are made in real-time, without the artefacts commonly associated with fixation. Ultimately, the aim is to use cells with enhanced fluorescent protein modified cytoskeleton components such as tubulin, actin and vinculin to correlate cytoskeleton organization with mechanics of cell-substrate interactions.