A composite bioscaffold was constructed by encapsulating human decellularized adipose tissue (DAT) within a photopolymerized polysaccharide hydrogel towards the goal of forming an injectable scaffold for adipose tissue engineering. Methacrylated glycol chitosan (MGC) and methacrylated chondroitin sulphate (MCS) were investigated as the hydrogel base materials with varying DAT concentrations. Glycol chitosan and chondroitin sulphate were converted to photopolymerizable prepolymers through graft methacrylation using glycidyl methacrylate and methacrylate anhydride respectively to achieve a degree of substitution (DOS) of 15% and 16%, respectively. MGC and MCS gels containing 0, 3 and 5 w/v% cryo-milled DAT were fabricated and characterized by measuring sol content, equilibrium water content and compressive mechanical properties (n=4, n=3). An increase in stiffness and a decrease in sol and water contents were observed in the gels with higher DAT concentration, suggesting that the DAT was acting as a filler material that contributed to the crosslinking reaction. In vitro studies were conducted with primary human adipose-derived stem cells (ASCs) encapsulated in the DAT-polymer constructs to assess cellular viability (n=3, N=3) as well as adipogenic differentiation, quantitatively via glycerol-3-phosphate dehydrogenase (GPDH) enzyme activity (n=3, N=3) and qualitatively through end-point RT-PCR analysis of key adipogenic genes (LPL, PPARγ, and CEPBα) (n=2, N=3) and intracellular lipid staining (n=3, N=3). Incorporating the DAT with MGC or MCS hydrogels enhanced cell viability as compared to the MGC and MCS scaffolds alone, with the MCS + 5 w/v% DAT scaffold having the highest overall cell viability and total cell number. The addition of the DAT in the MGC and MCS scaffold groups enhanced ASC adipogenesis as measured by an increase in GPDH levels, adipogenic gene expression and intracellular lipid accumulation characteristic of adipocytes. The highest GPDH levels were observed in the induced MCS with 5 w/v% DAT scaffolds, as compared to all other scaffold groups and tissue culture controls. The GPDH activity in this group increased by almost three times between 3 and 14 days, consistent with the progression of differentiation. The results indicated that the MCS-based scaffolds incorporating the DAT promoted cell viability and adipogenesis, demonstrating great promise as composite scaffolds for soft tissue regeneration. / Thesis (Master, Chemical Engineering) -- Queen's University, 2011-12-23 15:12:37.772
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:OKQ.1974/6965 |
| Date | 18 January 2012 |
| Creators | CHEUNG, HOI KI |
| Contributors | Queen's University (Kingston, Ont.). Theses (Queen's University (Kingston, Ont.)) |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English, English |
| Detected Language | English |
| Type | Thesis |
| Rights | This publication is made available by the authority of the copyright owner solely for the purpose of private study and research and may not be copied or reproduced except as permitted by the copyright laws without written authority from the copyright owner. |
| Relation | Canadian theses |
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