archives@tulane.edu / Hydrogels serve as three-dimensional scaffolds whose composition can be customized to allow the attachment and proliferation of several different cell types. Decellularized tissue-derived scaffolds are considered close replicates of the tissue microenvironment. Decellularized adipose tissue (DAT) hydrogel has proven to be a useful tool for tissue engineering applications in pre-clinical models. The first aim of the present study was to characterize the biochemical composition of DAT hydrogel. The DAT hydrogel was prepared by processing adipose tissue acquired from three female human donors, and subsequently quantitatively analyzed using liquid chromatography-mass spectrometry (LC-MS). The enriched and depleted proteins were determined in DAT hydrogel and further analyzed by gene ontology (GO) analysis. Extracellular matrix proteins were found to be enriched, while cellular proteins were depleted relative to native adipose tissue. Furthermore, GO analysis identified that the enriched proteins could affect various biological processes via the regulation of a range of cellular pathways. The second aim was focused on the analysis of the effect of adipose-derived stromal/stem cells (ASCs) and DAT hydrogel interaction on cell morphology, proliferation, differentiation, and hydrogel microstructure. The ASCs seeded in DAT hydrogel remained viable and displayed proliferation. The adipogenic and osteogenic differentiation of ASCs seeded in DAT hydrogel was confirmed by marker gene expression and histochemical staining. Moreover, ASC attachment and differentiation altered the fibril arrangement, which indicated remodeling of the DAT hydrogel. The third aim was to analyze the regenerative potential of DAT hydrogel in a critical-sized mouse femoral defect model. The DAT hydrogel alone, or its composites with ASCs, osteo-induced ASCs (OIASC), and hydroxyapatite were tested for the ability to mediate repair of the femoral defect. The data indicated that DAT hydrogel promoted bone regeneration alone, while the regeneration was enhanced in the presence of OIASCs and hydroxyapatite. In summary, the current findings confirm that DAT hydrogel is a cytocompatible and bio-active scaffold, with potential utility as an off-the-shelf product for tissue engineering applications. In future, the analysis of DAT hydrogel using a wider range of donors representing different body mass index, age, gender, and ethnicity will provide a more comprehensive characterization. / 0 / Omair A. Mohiuddin

In vivo, adipose tissue demonstrates only a limited capacity for self-repair, and the long-term treatment of subcutaneous defects remains an unresolved clinical problem. With the goal of regenerating healthy tissues, many tissue-engineering strategies have pointed to the potential of implementing three-dimensional (3-D), cell-seeded scaffolds for soft tissue augmentation and wound healing. In particular, microcarriers have shown promise as both cell expansion substrates and injectable cell-delivery vehicles for these applications. However, limited research has investigated the engineering of tissue-specific microcarriers, designed to closely mimic the native extracellular matrix (ECM) composition. In this work, methods were developed to fabricate microcarriers from decellularized adipose tissue (DAT) via non-cytotoxic protocols. Characterization by microscopy confirmed the efficacy of the fabrication protocols in producing stable beads, as well as the production of a microporous surface topography. The mean bead diameter was 934 ± 51 μm, while the porosity was measured to be 29 ± 4 % using liquid displacement. Stability and swelling behavior over 4 weeks indicated that the DAT-based microcarriers were effectively stabilized with the non-cytotoxic photochemical crosslinking agent rose bengal, with only low levels of protein release measured within a simulated physiological environment. In cell-based studies, the DAT-based microcarriers successfully supported the proliferation and adipogenic differentiation of human adipose-derived stem cells (hASCs) in a dynamic spinner flask system, with a more favorable response observed in terms of adhesion, proliferation, and adipogenesis on the DAT-based microcarriers relative to gelatin control beads. More specifically, dynamically-cultured hASCs on DAT-based microcarriers demonstrated greater lipid loading, as well as higher glycerol-3-phosphate dehydrogenase (GPDH) activity, a key enzyme involved in triacylglycerol biosynthesis, at 7 days and 14 days in culture in an inductive medium. Overall, the results indicated that the DAT-based microcarriers provided a uniquely supportive environment for adipogenesis. Established microcarrier sterility and injectability further support the broad potential of these tissue-specific microcarriers as a novel, adipogenic, clinically-translatable strategy for soft tissue engineering. / Thesis (Master, Chemical Engineering) -- Queen's University, 2010-12-01 14:28:14.628