Validation Of A Novel Experimental And Computational Methodology To Measure Intercellular Forces During Tissue Morphogenesis

During three-dimensional tissue development, cell-cell confinement, cell-cell traction forces, and the resulting intracellular tension play a key role in regulating cell proliferation differentiation, migration, and apoptosis. Yet, the molecular mechanisms that underlie the biochemical response of living cells to these mechanical forces are largely elusive, in part due to the lack of suitable methods to measure accurately the intercellular forces in three-dimensional, developing living tissues with high spatial and temporal resolution. We have developed a novel experimental and computational methodology to measure the spatial and temporal evolution of the intercellular forces during tissue development while simultaneously monitoring the dynamics of the resulting proliferation, differentiation, and migration of the cells within the tissue using a small, slender, elastic PDMS microrod. The microfabricated rod is functionalized and embedded in the tissue as we track its shape deformations over time. As tissue forms around the rod, the cells in the developing tissue exert stresses producing bending deformations along the length of the microrod. By tracking these deformations we can backtrack a solution to the primary forces exerted by the cells. Preliminary experimental results on wild type HaCat cells show that the method has enough sensitivity to measure single cell-to-cell forces and to evaluate the dynamics of wound healing processes. A validation strategy for this method is presented and tested in 1D and 2D scenarios. Traction forces are obtained solving the classical solution of the elastostatic equation from the deformations obtained from fluorescent beads on polyacrylamide substrates. Direct measurements of the intercellular stresses are then compared with calculations of the monolayer stresses obtained solving the Kirchhoff-Love thin plate theory equations.