Thesis (M.S.)--New Jersey Institute of Technology, Dept. of Biomedical Engineering, 2003. / HFT20030804. Includes bibliographical references (p. 75-76). Also available via the World Wide Web.

published_or_final_version / Orthopaedics and Traumatology / Doctoral / Doctor of Philosophy

Formation of an intact, continuous and biological interface with proper zonal organization between mechanically dissimilar tissues is a key challenge in complex tissue engineering. The presence of a stable interface between soft and hard tissues is important. In particular, the presence of the osteochondral interface can prevent mechanical failure by reducing the shear stress across it. It also prevents vascularization and subsequent mineralization of the uncalcified cartilage, thus maintaining the normal tissue function. In this study, we demonstrated that with the use of mesenchymal stem cells, the collagen scaffold and the microencapsulation technology, an osteochondral interface with a zone of calcified cartilage could be generated in vitro in 5 weeks. Specifically, by placing an undifferentiated mesenchymal stem cell-collagen gel between an upper cartilage-like part and a lower bone-like part, cells in the middle layer were able to remodel the collagen gel into an interface similar to that found in vivo. Hypertrophic chondrocytes populated this in vitro generated interface, secreting GAGs, collagen type II and X, and calcium phosphates. Vertically running collagen fibers were found in this interface as well. We also demonstrated the importance of culture medium together with an appropriate configuration for interface formation. In particular, only with the use of both the chondrogenic medium and the three-layer configuration could we generate the osteochondral interface in vitro. Finally we conducted a pilot animal study on the efficacy of cartilage repair using constructs with a pre-formed osteochondral interface and demonstrated that cartilage re-surfacing was successful in only one month. Hyaline-like cartilage with a continuous tidemark was regenerated. This observed phenomenon could be maintained up to 3 months. Results of this study contribute to the development of better cartilage repair in future. / published_or_final_version / Mechanical Engineering / Doctoral / Doctor of Philosophy

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No description available.

In vitro grown cartilage is functionally inferior to native tissue, and improvements in its quality should be attempted so it can be used therapeutically. In these studies we investigated the effects of cell shape on tissue quality through alteration of substrate geometry and application of mechanical stimuli. Articular chondrocytes were isolated and cultured on the surface Ti-6Al-4V substrates with various geometries. When cultured on fully porous titanium alloy substrates, chondrocyte spreading was enhanced over those grown on substrates with solid bases. Chondrocytes which remained round did not synthesize significant amounts of matrix and were thus unable to form cartilaginous tissue. In contrast, chondrocytes which were directed to spread to a limited amount, resulting in a polygonal morphology, accumulated significantly more matrix molecules and in time formed cartilage-like tissue. Computational fluid dynamics analyses demonstrated that cells on fully porous substrates experience time-dependent shear stresses that differ from those experienced by cells on substrates with solid bases where media flow-through is restricted. Integrin-blocking experiments revealed that integrins are important regulators of cell shape, and appeared to influence the accumulation of collagen and proteoglycans by chondrocytes. Furthermore, compressive mechanical stimulation induced a rapid, transient increase in chondrocyte spreading by 10 minutes, followed by a retraction to pre-stimulated size within 6 hours. This has been shown to be associated with increased accumulation of newly synthesized proteoglycans. Blocking the α5β1 integrin, or its β1 subunit, inhibited cell spreading and resulted in a partial inhibition of compression-induced increases in matrix accumulation, thereby substantiating the role of β1 integrins in this process. These results suggest that both fluid induced shear forces and compressive forces regulate chondrocyte matrix accumulation by altering cell morphology, which is mediated by integrins. Identifying the molecular mechanisms that influence chondrocyte shape and thus tissue formation may ultimately lead to the development of a tissue that more closely resembles native articular cartilage.

Whole-organ perfusion decellularization was performed with rat hearts on a modified chromatography apparatus. Analysis of the flow properties and effluent material over time provided insights into the decellularization process, and allowed non-destructive testing of perfused cardiac tissue. Decellularized matrices were stored for up to 1 year at -80°C and then conditioned to remove residual detergent and cryoprotectant. Tissue was reseeded with canine blood outgrowth endothelial cells (BOECs) and cultured in an autoclavable closed-circuit bubble-free reactor. The entire process was considered in the context of eventual scale-up in equipment design, the use of disposable components, and extracellular matrix (ECM) product storage. Tissue patch substrates for cell growth were studied for cytotoxic effects towards process development. Decellularization protocols were compared. Extracellular matrix derived coatings and gels were investigated as process assays and potential cell delivery vehicles. Peracetic acid and UV disinfection were tested. Micronized ECM carriers were developed for scalable culture, with considerations to carrier morphology, cell attachment, and egress. Micronized ECM carriers were tested with a novel in vitro assay to simulate the support of adherent cells for gene-modified cell therapy.

Thesis (Ph.D.)--Drexel University, 2007. / Includes abstract and vita. Includes bibliographical references (leaves 204-213).

Universiẗat, Diss.--Jena, 2008.

Zugl.: Dresden, Techn. Universiẗat, Diss., 2008.