Interaction of cells with fibronectin fibres

Cell motility is a delicate balance between adhesion and detachment. Identifying key proteins involved and understanding the mechanisms employed to find this balance will provide new insight into the means by which motility can be controlled. Mechanical force plays a key regulatory role in biological cells and therefore to understand how cells move and adhere ultimately relies on how forces are generated and propagated or in essence, how the cell interacts with the surface. The stress on fibronectin fibres may be the deciding factor in determining the attachment of the cell to the matrix and the adhesion of the matrix to the substrate. Studies geared toward understanding fibronectin structure, organization, and binding affinity under mechanical stretching are providing information crucial to the understanding of the effect of mechanical forces on cell function.;Movement of fibronectin fibres composed of modified protomers in living cells is clearly observed and provides information about matrix structure, organization, and movement. The structure and organization of fibronectin on the surface must reflect both how it is laid down to aid in adhesion, and the mechanism by which cells move and cause strain on the matrix. A static analysis of fibronectin structures and their patterns provides insight into how they are produced and how they are reorganized. A model is proposed to account for fibronectin organization during cell motility and the role that membrane tension may play. To test the prediction of the model that stretching fibronectin reduces integrin-binding activity, an AFM compatible tool is designed to apply a mechanical force to fibronectin while monitoring the change in intermolecular interactions that result.;The force required to rupture the interaction between an integrin mimic and fibronectin is determined to be approximately 100 pN. After stretching fibronectin, a trend toward fewer rupture events characterized by smaller pull-off forces in each force curve is observed which implies a decrease in potential binding sites available to the integrin mimic and possibly weaker interactions. More data are needed to make firmer conclusions, but the experimental method employed here is exceptionally promising.;Orientation dependent binding experiments must be tested on fibronectin fibres laid down by the cells. A combination NSOM, AFM and confocal microscope which will identify fibronectin fibres by fluorescence and perform force measurements at precise locations, is developed and tested to achieve this goal. The combination experiment is possible but it is clear that technical improvements are required in the design of NSOM probes and the reduction of forces applied to the cells.*.;*This dissertation is a compound document (contains both a paper copy and a CD as part of the dissertation). The CD requires the following system requirements: Windows MediaPlayer or RealPlayer.