Systematic Investigation of Hydrogel Material Properties on Cell Responses for Vocal Fold and Vascular Graft Tissue Engineering

摘要

The research presented here deals with synthetic materials for application intissue engineering, primarily poly(ethylene glycol) (PEG) and poly(dimethyl siloxane)star(PDMS)star. Tissue engineering seeks to repair or replace damaged tissue throughimplantation of cell encapsulated in an artificial scaffold. Cell differentiation andextracellular matrix (ECM) deposition can be influenced through a wide variety of invitro culture techniques including biochemical stimuli, cell-cell interactions, mechanicalconditioning and scaffold physical properties. In order to systematically optimize invitro conditions for tissue engineering experiments, the individual effects of thesedifferent components must be studied. PEG hydrogels are a suitable scaffold for thisbecause of their biocompatibility and biological "blank slate" nature.This dissertation presents data investigating: the effects of glycosaminoglycans(GAGs) as biochemical stimuli on pig vocal fold fibroblasts (PVFfs); the effects ofmechanical conditioning and cell-cell interactions on smooth muscle cells (SMCs); andthe effects of scaffold physical properties on SMCs. Results show that GAGs influence PVFf behavior and are an important component in scaffold design. Hyaluronic acid (HA) formulations showed similar production in collagen I and III as well as reducedlevels of smooth muscle a-actin (SMa-actin), while chondroitin sulfate (CSC) andheparin sulfate showed enriched collagen III environments with enhanced expression ofSMa-actin.A physiological flow system was developed to give comprehensive control overin vitro mechanical conditioning on TEVGs. Experiments performed on SMCs involvedcreating multi-layered TEVGs to mimic natural vascular tissue. Constructs subjected tomechanical conditioning with an endothelial cell (EC) layer showed enhancedexpression of SMC differentiation markers calponin h1 and myocardin and enhanceddeposition of elastin. Consistent with other studies, EC presence diminished overallcollagen production and collagen I, specifically.Novel PDMSstar-PEG hydrogels were studied to investigate the effects ofinorganic content on mesenchymal stem cell differentiation for use in TEVGs. Resultsagree with previous observations showing that a ratio of 5:95 PDMSstar: PEG by weightenhances SMC differentiation markers; however, statistically significant conclusionscould not be made. By studying and optimizing in vitro culture conditions includingscaffold properties, mechanical conditioning and multi-layered cell-cell interactions,TEVGs can be designed to maximize SMC differentiation and ECM production.