Cell Morphology and Substrate Ligand Density Determines Adhesion Strength and Remodelling Under Dynamic Shear

Abstract

Cells adhere to substrates through dynamic mechanosensitive focal adhesion complexes. These complexes are connected mechanically and through biochemical signalling pathways to the underlying cytoskeleton. Adhesion strength of cells to substrates depends on the cellular shape and area, distribution and integrin density, and the focal adhesion composition. Fluid shear-based experiments to quantify the critical shear stress required for cell detachment provide a quantitative measure of cell-substrate interactions. We used a custom microscope-mountable fluid shear device to quantify the adhesion strength of NIH3T3 cells when exposed to increasing shear stress at different rates (1, 5, and 10 minutes) ranging from 0-8 Pa. Cells were allowed to attach to glass petri-dishes coated with either fibronectin (10, 20, 40, 80, 200 μm/ ml) or collagen-1 (10, 20, 40, 80, 200 μm/ ml) prior to the experiment. The number of cells adhered to the petri-dish decreased with increasing shear stress in a sigmoidal profile. The critical shear stress for 50% cell attachment increased with ligand density and with higher shear exposure duration for both fibronectin and collagen-1 coated substrates. The mean spread area increased linearly (p<0.05) from 1371.6±516.9 for 10 μg/ml fibronectin coated substrates by ∼1.8 times as compared to the corresponding 200μg/ml group. Collagen-1 treated substrates showed a similar response with two-fold increase. There were no differences in cell areas between the two ligand groups at low concentrations; the differences were apparent at higher concentrations alone. These results will be correlated with in changes to actin and vinculin remodelling and organization at. The mean tractions will be quantified using regularized-FTTC following shear-induced cytoskeletal remodelling. A better understanding of cell adhesion strength under mechanical cues may help identify highly metastatic cells which may aid in faster diagnosis and cell sorting.