The circulating neutrophil has three major tasks that it must accomplish before it can carry out its final task of phagocytosis. First, it must deform and flow through the capillaries often while in a stimulated state. Second, it must recognize sites of inflammation in the post-capillary venules and rapidly attach to these sites by rolling and then firmly adhering to the endothelial cells that line the walls of these venules. Finally, it must migrate through the vessel wall and into the surrounding tissue by a continuous process of adhesion and detachment. The fundamental mechanical processes that underlie each of these tasks are not well understood and will be the focus of the three specific aims of this proposal. The first specific aim is to study the deformation and flow of neutrophils into capillaries when the cells have been stimulated by a sac chemoattractant or cytokine or priming agent, either separately or together. The mechanical sensitivity of our experimental system will allow us to study low levels of these stimulating agents when no obvious morphological changes have occurred The second aim is to study adhesion and detachment of the unstimulated and stimulated neutrophil from artificial surfaces and from endothelial cells and other neutrophils. Again, the mechanical sensitivity of our system will allow us to approach this aim from the most fundamental level by first determining the forces and lifetimes of individual chemical bonds. Larger bond forces from multiple bonds will be the sum of the basic canonical forces sustained by the single bonds.
The third aim will be to characterize the pressures, contraction forces and rates of the fundamental polymerization and depolymerization processes that ate responsible for the motion of the cell. In all cases the emphasis is on the underlying mechanical characteristics that govern the life of the neutrophil. Only by understanding these fundamental mechanical processes in vitro can we explain the cell's complex behavior in vivo. Experiments will be done with a glass micropipet, which serves as a fundamental, in-vitro model of a capillary vessel. Driving pressures, forces and chemical environments can be precisely controlled and deformations and velocities can be measured. This permits the mechanical characteristics of the cell, such as cortical tension, viscosity, stiffening due to activation, adhesion due to chemical bond formation, detachment due to the failure of chemical bonds and polymerization due to mechanical and chemical stimulation, to be precisely measured.
Showing the most recent 10 out of 42 publications
