|Smith, C Wayne|
Submitted to: Meeting Abstract
Publication Type: Other
Publication Acceptance Date: 3/1/2003
Publication Date: 3/1/2003
Citation: GONZALEZ, A., GOBIN, A., DEMOU, Z., WEST, J., SMITH, W.C., MCINTIRE, L.V. TISSUE ENGINEERING INVESTIGATION OF LEUKOCYTE ADHESION AND MIGRATION. MEETING ABSTRACT. 2003. Interpretive Summary: None required for an abstract.
Technical Abstract: The classical model of cell locomotion suggests that successful invasion and migration is dependent on the biochemical degradation of the extracellular matrix (ECM). Many properties of the ECM regulate the cascade of events involved in leukocyte migration. Neutrophil migration in extravascular tissue has been correlated to protease release resulting in degradation of ECM components, although the details of extravasation and migration remain unclear. By simulating the many properties of the ECM in relation to ECM/cell interaction, we propose to evaluate in vitro neutrophil adhesion, protease release and cellular migration using 2-and 3-dimensional biomimetic systems. A biomimetic hydrogel will provide mechanical support, adhesion ligands, proteolytic susceptibility, chemotactic stimuli and growth factors that induce signaling pathways within a cell. Such a system can be engineered using polyethylene-glycol (PEG) as a synthetic polymer base. Side chain modification techniques, as well as the formation of polymer-peptide hybrids, allow us to control cell adhesion, polymer degradation, and hydrogel stiffness in such a way as to mimic the biochemical and biomechanical properties of the ECM in a synthetic material. Specifically, adhesion-promoting peptide sequences, like RGDS (Arg-Gly-Asp-Ser), can be covalently grafted to PEG to provide a means by which cell motility as a function of substrate adhesivity may be investigated. Copolymers with inserted degradable peptide sequences, like those that serve as targets for elastase and collagenase, are prepared as a means of observing proteolytic dependent migration. By inserting integrin specific peptides into the PEG backbone, we are able to dissect the requirement of specific integrin activation for cell movement and ECM degradation. This will allow for direct testing of hypotheses concerning the molecular mechanisms of neutrophil migration in tissue. Classic 2-D static adhesion systems, along with the biomimetic hydrogels, are used to evaluate and quantify cell adhesion, cell spreading, and protease release. A 3-D cell tracking system will be implemented for the quantification of migratory parameters. We will use computer automated scanning optical microscopy to capture real-time images of neutrophil migration through the synthetic ECM. Imaging languages will be used to perform automatic image acquisition, image analysis, cell trajectory reconstruction, and data analysis. The 3-D coordinates obtained from image analysis allow us to reconstruct the trajectory of individual cells. This real-time cell trajectory reconstruction will be used to predict parameters including the average cell waiting times, cell state transition probabilities, and the steady-state motion probabilities. Using the polymer-peptide hybridized biomimetic hydrogel in conjunction with the 2-D adhesion chamber and an automated 3-D cell tracking system, the mechanisms of leukocyte adhesion and migration can be simulated and quantified.