Rebecca L. Carrier
Daniel D. Burkey, Shashi Krishna Murthy, David E. (David Edward) Budil
Date of Award
Doctor of Philosophy
Department or Academic Unit
College of Engineering. Department of Chemical Engineering.
chemical engineering, biomaterials, chemical vapor deposition, scaffolds, tissue engineering
Tissue scaffolds, Extracellular matrix, Chemical vapor deposition
Molecular, cellular, and tissue engineering
Replication of native tissue extracellular matrix (ECM) using CVD or microfabrication techniques enables creation of cell culture scaffolds with precisely biomimetic, irregular, and multi-scale native tissue topography. Topological cues have been shown to be very important in controlling cell phenotype in literature. However little has been done to investigate the effect of topography on cell behavior using exact replicas of native ECM, which often involves multi-scale and irregular topographies. Hence, we proposed a novel, alternative approach to creating cell culture scaffolds with exact replication of ECM topography by using actual native tissues as molds or templates from which to produce synthetic scaffolds with micron and sub micron scale topography that exactly mirrors that of the original tissue. The fabrication of the primary biomimetic replica includes creation of a rigid, inorganic mold using either PDMS, silica or parylene. Chemical vapor deposition, a technique capable of coating surface with complex nano and micron scale topography was used to produce silica and parylene films. CVD of parylene was able to replicate the micron and sub micron topography of the intestinal ECM creating a rigid, negative primary replica. Next, a biocompatible positive secondary replica was development using CVD pHEMA and PDMS. CVD of the biocompatible polymer pHEMA enables control of the degree of cross-linking during the CVD process by adjusting the vapor flowrates of monomer and crosslinker. The degree of cross-linking of the pHEMA films measured using XPS oxygen elemental analysis. Results suggested that higher flowrates of EGDA crosslinker during deposition caused higher cross-linking of pHEMA. SEM images of pHEMA coated intestinal tissue demonstrated preservation of topographical details at the micron to sub-micron scale, and XPS elemental analysis confirmed the conformal pHEMA coating on the surface of intestinal tissue. Caco-2 cells were cultured on cross-linked pHEMA films, and it was found that cell growth, attachment and viability on the CVD pHEMA were comparable to tissue culture treated polystyrene. It was discovered that higher cross-linked pHEMA has higher cell attachment, due to the balancing of the hydrophobic and hydrophilic properties in the films. To replicate and separate pHEMA from the primary parylene mold, thicker pHEMA films are required to be able to separate from parylene mold, which were not achieved with our system. To form a secondary thick, easy to separate replica, poly dimethyl siloxane (PDMS) was implemented. SEM images comparing bare intestinal membrane with the negative parylene replica and the positive PDMS mold indicated replication of intestinal basement membrane features at the hundreds of micron scale and down to the single micron scale in the positive PDMS mold.
Courtney Allyn Pfluger
Pfluger, Courtney Allyn, "Biomimetic replication of intestinal basement membrane topography" (2011). Chemical Engineering Dissertations. Paper 10. http://hdl.handle.net/2047/d20001077
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