Cell-matrix interactions : collagen-GAG scaffold fabrication, characterization, and measurement of cell migratory and contractile behavior via confocal microscopy

• Main Author: Harley, Brendan A. (Brendan Andrew), 1978-
• Other Authors: Lorna J. Gibson.
• Format: Others
• Language: English
• Published: Massachusetts Institute of Technology 2008
• Subjects: Mechanical Engineering.
• Online Access: http://hdl.handle.net/1721.1/35678

Thesis (Sc. D.)--Massachusetts Institute of Technology, Dept. of Mechanical Engineering, 2006. === Includes bibliographical references (v. 2, leaves 371-393). === Three-dimensional, collagen scaffolds are an analog of the extracellular matrix and are used for many tissue engineering applications. While material and microstructural properties significantly affect overall scaffold bioactivity, the specific influence of construct mechanical properties, composition, and pore microstructure is unknown. In this thesis, experimental and theoretical approaches are employed to systematically examine the independent effect of extracellular features on cell behavior within a series of standardized, well-characterized, collagen-glycosaminoglycan (CG) scaffolds, providing valuable information for designing biomaterials with improved physiological relevance. This thesis also aims to provide experimental and theoretical approaches appropriate for characterizing and describing a wide range of porous biomaterials and for quantifying the effect of extracellular cues on cell behavior within these biomaterials. CG scaffolds are fabricated via freeze drying. Novel thermal processing conditions were developed to produce two homologous series of uniform, mechanically isotropic CG scaffolds, one with varying pore size and constant stiffness and the other with constant pore size and varying stiffness. === (cont.) The thermal processing conditions and the resultant scaffold microstructure have been modeled using an isothermal coarsening heat transfer model within a conductive mold with interface resistance, allowing fabrication of future scaffolds with engineered microstructures. The mechanical properties, specific surface area, and permeability of the scaffolds have been experimentally measured and theoretically described using a cellular solids framework appropriate for modeling many porous biomaterials. This thesis research has produced a standardized series of CG scaffolds appropriate for quantitative in vitro cell behavior assays. An experimental methodology for measuring cell-generated contractile forces and cell motility is described and implemented; the independent effect of scaffold pore size and stiffness on the magnitude and kinetics of cell motility within the scaffolds was determined via confocal microscopy. Slight changes in the extracellular environment appreciably influence cell behavior. Significant effects of cell density, cell type, scaffold microstructure, and scaffold stiffness were observed: cell migration speed increased with decreasing pore size or increasing cell density and increased asymptotically with scaffold stiffness. An improved measurement of the contraction force generated by single dermal fibroblasts (Fc = 26 + 13 nN) within the CG scaffold has also been made. === by Brendan A. Harley. === Sc.D.