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https://hdl.handle.net/2142/87143
Description
Title
Visualizing Cell Migration in Situ
Author(s)
Knight, Brian Lee
Issue Date
2000
Doctoral Committee Chair(s)
Howard Gelberg
Department of Study
Veterinary Clinical Medicine
Discipline
Veterinary Clinical Medicine
Degree Granting Institution
University of Illinois at Urbana-Champaign
Degree Name
Ph.D.
Degree Level
Dissertation
Keyword(s)
Biology, Veterinary Science
Language
eng
Abstract
Cell migration has been studied extensively by manipulating and observing cells bathed in putative chemotactic or chemokinetic agents on planar substrates. This system is attractive since the cells and their molecular components can be readily imaged and matrix and soluble parameters easily manipulated. However, migration in culture appears to differ dramatically from the complex, but poorly defined, environment in which cells migrate in vivo. The result is that cells in culture often do not show highly directed migratory behavior and express adhesive and signaling components that might not be present in vivo. As a more appropriate environment in which address mechanisms underlying migration, I have studied the pathways, morphologies, and molecular organization and dynamics of adhesion related molecules in muscle precursor cells migrating from the somite into the forelimb using embryo slices. New technologies were developed to stabilize slices within the microscopic field and to apply surface support along cut margins of the slice to prevent exuberant outgrowth from the slice. Muscle precursors, in somites, initiate migration synchronously and migrate in broad, rather than highly defined, regions. Bursts of directed migration are followed by periods of meandering and protrusiveness. Introducing cDNAs encoding GFP-fusion proteins demonstrate that alpha-actinin resides in linear, punctate structures; the alpha5 integrin, while not generally well organized, resides in some small, focal complex-like concentrations and in vesicles, which can localize at the base of protrusions; and paxillin is not organized detectably. The muscle precursors migrating in situ form unusually large, long-lived protrusions that are polarized in the direction of migration, which are not seen in the same cells migrating out of the slices. GFP-Rac fusion proteins demonstrate that constitutively activated Rac, in contrast to wild type Rac, localizes continuously around the cell surface and promotes random, rather than directed, protrusive activity and migration. I show the feasibility of observing migration and cellular and molecular organizations at high temporal and spatial resolution in situ using tissue slices along with a readily accessible imaging system. Migration from the somite to the wing bud is discontinuous and not highly stereotyped. The cellular events that comprise migration in situ differ from those in dissociated cell culture.
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