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ENGINEERING OF CLINICAL-SCALE, IN VITRO VASCULARIZED BONE TISSUE FOR IMPLANTATION
Title
ENGINEERING OF CLINICAL-SCALE, IN VITRO VASCULARIZED BONE TISSUE FOR IMPLANTATION
Creator
Gandhi, Jarel K.
Date
2016, 2016-05
Description
Tissue engineering has been a rapidly expanding field dedicated to regeneration of tissue. The field has focused on application through...
Show moreTissue engineering has been a rapidly expanding field dedicated to regeneration of tissue. The field has focused on application through combinations of 3 key components: cells, signals, and scaffolds. One ambitious combination of all three is the desire to engineer functional tissues in vitro to meet the clinical-demand of organ replacement. While major advances have been made, a critical obstacle that has yet to be overcome is the need to grow large volumes of complex 3D tissue. In this proposal, this issue is addresed in two ways: the use of a perfusion bioreactor system to culture 3D scaffolds to enhance mass transport, and engineering of a vascular network withing the scaffold for rapid perfusion once implanted in vivo. This thesis aims to address both aspects for bone tissue engineering by engineering pre-vascularized, mineralizing scaffolds that can be scaled up to clinically-relevant volumes by using a tubular perfusion bioreactor system (TPS). To address this, 3 aims were addressed. First, 3D culture of endothelial colony forming cells (ECFCs), a clinically-relevant cell population, was demonstrated utilizing fibrin gels within the TPS. The TPS allowed for viable culture of ECFCs within fibrin bead scaffold up to 1 week without a reduction in cell amount or genomic quality of the cells. Second, a co-culture model of angiogenesis utilizing ECFCs and mesenchymal stem cells (MSCs) was demonstrated to reproducibly form pre-formed vessel networks within a mineralizing fibrin scaffold. Data shows that MSC suspension concentration and fibrinogen concentration modulate the angiogenic response. Mineralization is demonstrated without the use of osteogenic media utilizing shear stress within the TPS. Finally, functionality of the pre-formed vessels is demonstrated following implantation to a SCID mouse model. Engineered human vessels showed anastasmosis to the host vasculature, with evidence of interconnected host and human vessel networks as well as formation of hybrid vessels. Additionally, evidence of mineralization within the scaffolds is maintained in TPS-cultured samples. In demonstrating these aims, future work should focus on fortifying the scaffold material to enable addressing implantation and persistence of clinically-relevant tissue volumes. In conclusion, pre-vascularization within bioreactor-cultured scaffolds represents a promising solution for future tissue engineering application.
Ph.D. in Biomedical Engineering, May 2016
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BIOMATERIAL SYSTEMS WITH PERSISTENT GROWTH FACTOR GRADIENTS IN VIVO FOR TISSUE ENGINEERING APPLICATIONS
Title
BIOMATERIAL SYSTEMS WITH PERSISTENT GROWTH FACTOR GRADIENTS IN VIVO FOR TISSUE ENGINEERING APPLICATIONS
Creator
Akar, Banu
Date
2016, 2016-12
Description
Tissue engineering aims to develop strategies for the replacement of damaged, injured or missing tissues with biologically compatible...
Show moreTissue engineering aims to develop strategies for the replacement of damaged, injured or missing tissues with biologically compatible substitutes such as bioengineered tissues. However, generating tissues of su cient volume for clinical application requires the formation of stable and extensive vasculature within the tissue constructs. The overall goal of this work is to enhance vascularization using a gradient biomaterial system and apply this research to engineering vascularized bone of clinical size. First, a method was developed to create persistent growth factor gradients with an adjustable gradient magnitude in vivo. This method generated persistent gradients of platelet-derived growth factor (PDGF-BB) within brin/poly (ethylene glycol) (PEG) sca olds. The presence of a growth factor gradient within the system was veri ed in vivo using near-infrared imaging. Also, a computational model was developed to investigate gradient characteristics within the system. Gradient properties can be controlled by varying the degradation rate of the gradient layer components or dose of PDGF-BB delivered. The angiogenic potential of gradient sca olds was tested in rodents using a subcutaneous implantation model. The depth of tissue invasion and density of blood vessels formed in response to the biomaterial increased with dose of the growth factor. The gradient biomaterial system allows formation of persistent gradients that can be in uenced by biomaterial characteristics, and enhances vascularization. Therefore, this biomaterial system can be used for tissue engineering applications. Second, the brin/PEG-based sca olds were modi ed to be degradable via hydrolysis and to include bioactive ceramic particles (hydroxyapatite and -tri-calcium phosphate). Characteristics of the hydrogel ceramic composites were investigated in vitro and in vivo. The presence of ceramic particles extended degradation time of thehydrogels in vitro and in vivo. Hydrogel ceramic composites were tested in a rodent cranial defect model and enhanced bone tissue regeneration. Third, strategies developed from the previous studies were combined to prepare ceramic supplemented gradient sca olds for bone tissue engineering applications. A gradient layer was applied to the hydrogel-ceramic composites and bone tissue response was evaluated in a periosteum guided large animal model. Ceramic supplemented gradient sca olds augmented vascularization and bone regeneration in vivo. In conclusion, a biomaterial system with persistent growth factor gradients was developed and enhanced vascularization and bone regeneration in vivo. This system holds a great potential for tissue engineering applications.
Ph.D. in Biomedical Engineering, December 2016
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