Thesis (Ph.D.)--Virginia Commonwealth University, 2010. / Prepared for: Dept. of Pathology. Title from title-page of electronic thesis. Bibliography: leaves 139-149.

The current model of complete separation between different industries, buildings and the public, hampers growth and sustainable development within the city. If connections can be identified between several related programmatic elements, can these work together to share and build common resources that benefit all parties involved? The showgrounds in Pretoria West currently exist as a void in the urban fabric, A large part of its infrastructure built for, and is now mostly only used for the annual Jacaranda Show that will now longer take place there. The role of the agricultural show within the city, however, remains a meaningful part of how the public interacts with the agricultural industry. In order to prevent the loss of this relationship and the heritage of the showgrounds, how can a new development around the existing Champion ring retain this role as well as build on the concept of connecting related industries and the public? The main aim of this dissertation is to investigate how architecture can address the interface between the livestock industry, research, education and the public, so that the collaboration between such programs will result in a reduced environmental impact and be of mutual benefit. / Mini Dissertation (MArchProf)--University of Pretoria, 2018. / Architecture / MArch(Prof) / Unrestricted

The mainstay of treatment for injuries to the anterior cruciate ligament (ACL) is with a tendon graft harvested from elsewhere in the knee. This procedure, known as ACL reconstruction (ACLR), has excellent reported outcomes in terms of restoring the gross stability of the knee. However there are reported rates of graft failure in the pediatric population.1 ACL reconstruction also does not provide full protection from developing premature osteoarthritis, which is often seen in patients after ACL injury. A new method must be found in order to find a solution to these adverse outcomes from the current standard treatment of ACL injuries. The Bridge-enhanced anterior cruciate ligament repair (BEAR) technique is a new innovation on primary repair of a torn ACL. It avoids the requirement for autograft harvesting and use of an allograft by combining a primary suture repair with an extraceullular matrix scaffold (the BEAR scaffold). This scaffold is placed between the two torn ends of the ACL to help facilitate natural healing of the ligament in order to restore the intrinsic functions of the ACL to protect the knee from trauma and instability. This ability to repair the native ACL maintains the proper anatomy of its insertion points, avoids donor site morbidity, and helps to prevent early osteoarthritis by maintaining innate proprioceptive functions of the ACL. Our study will prove the BRIDGE repair to be a promising step in the direction of preserving the native ACL.

The goal of this work is to explore the built context of our food system as a manifestation of a set of social and environmental conditions that are antithetical to the long-term health and survival of human life on this planet. The specific focus of this work is the small-scale, integrated farm. The farm is but one piece of the puzzle of how we eat and resides within the larger context of storage, distribution, economy, culture etc. Using precedents, both past and present, and through design explorations this work seeks to develop a positive course forward that will enable humanity to reconnect with its food source. We have the potential and impetus to rebuild and to heal our local resilience, food security, and egalitarian access to fresh, healthy food. Arguably, these goals have coinciding and connected paths within other aspects of our cultural and human needs – housing, manufacturing, healthcare, etc. The essential questions to be answered are: What does a healthy food system look like? How can this be designed to integrate into and support diverse and positive communities? What infrastructure is necessary to support the type of endeavor that creates healthy food, feeds a culture, and heals the damaged soil that is the basis of our sustenance. It is clear that industrial agriculture, the source of nearly all food consumed by Americans, is not this model. Appropriate food systems will vary by culture, climate, economy, settlement patterns, and the like. This work focuses on the condition of the Northeast region of the United States.

The aim of the thesis is to propose active damping as a potential control strategy for chatter instability in machine tools. The regenerative process theory explains chatter as a closed loop interaction between the structural dynamics and the cutting process. This is considered to be the most dominant reason behind machine tool chatter although other instability causing mechanisms exist. The stability lobe diagram provides a quantitative idea of the limits of stable machining in terms of two physical parameters: the width of contact between tool and the workpiece, called the width of cut and the speed of rotation of the spindle. It is found that the minimum value of the stability limit is proportional to the structural damping ratio for turning operations. This important finding provides the motivation of influencing the structural dynamics by active damping to enhance stability limits of a machining operation. A direct implementation of active damping in an industrial environment may be difficult. So an intermediate step of testing the strategy in a laboratory setup, without conducting real cutting is proposed. Two mechatronic "Hardware in the Loop" simulators for chatter in turning and milling are presented, which simulate regenerative chatter experimentally without conducting real cutting tests. A simple cantilever beam, representing the MDOF dynamics of the machine tool structure constitutes the basic hardware part and the cutting process is simulated in real time on a DSP board. The values of the cutting parameters such as spindle speed and the axial width of cut can be changed on the DSP board and the closed loop interaction between the structure and the cutting process can be led to instability. The demonstrators are then used as test beds to investigate the efficiency of active damping, as a potential chatter stabilization strategy. Active damping is easy to implement, robust and does not require a very detailed model of the structure for proper functioning, provided a collocated sensor and actuator configuration is followed. The idea of active damping is currently being implemented in the industry in various metal cutting machines as part of the European Union funded SMARTOOL project (www.smartool.org), intended to propose smart chatter control technologies in machining operations.

The aim of this study was to determine the trends in protocol, success rates, and consistency in follow up of revascularization procedures in a controlled environment. Patients of the Virginia Commonwealth University School of Dentistry were identified who were offered revascularization therapy as a treatment option on immature permanent teeth from January 1, 2010 to May 31, 2017. A total of 77 patients and 78 teeth were evaluated for revascularization therapy. For patients accepting treatment, records were reviewed for outcome assessment and consistency of follow up. A total of 30 patients (31 teeth) were treated following revascularization protocols, with only 20 patients (21 teeth) returning for follow up. Six of the 21 teeth needed some form of additional therapy due to patients remaining symptomatic, however 15/21 exhibited varying levels of success. Recall rate was 67.7%. With a success rate of 71.4%, revascularization therapy should continue to be considered for all patients with teeth having necrotic pulps and immature root apices. However, changes to recall protocols need to be improved in order to better monitor the status of teeth that undergo revascularization therapy.

Das Ziel dieser Arbeit war die Entwicklung eines vaskularisierten, autologen Implantats zur Behandlung von schweren Verletzungen der Trachea im Umfeld der guten Herstellungspraxis. Die Matrix besteht aus einem circa 14 cm langen Stück porcinen, azellularisierten Dünndarm und BioVaSc (Biological Vascularized Scaffold) genannt wird. Dieses wird dann mit isolierten und kultivierten Zellen des Patienten besiedelt und reift für zwei Wochen in einem speziell hierfür entwickelten Bioreaktorsystem. Danach erfolgt die Analyse bzw. die Implantation in den Patienten. Nach der Präparation und Überprüfung der Qualität, erfolgte die Azellularisierung der BioVaSc zur Entfernung der porcinen Zellen und der enzymatische Abbau der DNS, unter Erhalt des natürlichen Gefäßsystems. Hierfür ist Natriumdesoxycholat verwendet worden, wobei Rückstände davon das Ansiedeln der autologen Zellen negativ beeinflussen könnten. Deshalb wurde ein Test etabliert, mit dessen Hilfe, das Auswaschen der Azellularisierungsdetergenz bis zur Sterilisation nachweisbar war. Des Weiteren könnten in der BioVaSc natürlicherweise enthaltene Endotoxine Immunreaktionen im späteren Empfänger auslösen. Die gesetzlichen Grenzwerte konnten durch Modifikationen des Protokolls, unter Berücksichtigung der guten Herstellungspraxis, erreicht werden. Weiterhin konnte histologisch eine weitgehende DNS- und Zellfreiheit nachgewiesen werden, in der quantitativen Analyse ergab sich eine Abreicherung von 97% im Vergleich zum Ausgangsmaterial. Zur Bestimmung der funktionellen Stabilität der azellularisierten Matrix wurde die maximal tolerable Zugspannung bestimmt. Zur Besiedlung der Gefäße der Matrix wurden mikrovaskuläre Endothelzellen und für das Lumen Fibroblasten und Skelettmuskelzellen verwendet. Die Protokolle zur Isolation und Kultur sind hierzu unter den Bedingungen der guten Herstellungspraxis etabliert, optimiert und mit, soweit möglich, zertifizierten Reagenzien durchgeführt worden. Zur genauen Charakterisierung der Zellen wurden diese immunhistologisch über vier Passagen analysiert, wobei sich je nach Zelltyp und Differenzierungsstadium unterschiedliche Expressionsmuster ergaben. Zur Herstellung des autologen Implantats wurden zunächst die mikrovaskulären Endothelzellen in das vorhandene Gefäßsystem der BioVaSc eingebracht und dann für sieben Tage im etablierten Bioreaktorsystem kultiviert. Danach erfolgte die Besiedlung des Lumens mit Skelettmuskelzellen und Fibroblasten und die weitere siebentägige Kultur im Bioreaktorsystem. Die Besiedlung des Gefäßsystems musste optimiert werden, um sowohl die Besiedlungsdichte zu steigern als auch die Effizienz zu erhöhen. Das Lumen konnte mit der etablierten Methode vollständig besiedelt werden. Nach vierzehntägiger Kultur im Bioreaktorsystem erfolgte die Kontrolle der Zellvitalität, wobei sowohl in den Gefäßstrukturen als auch im Lumen der BioVaSc vitale Zellen nachweisbar waren. Histologische Analysen zeigten, dass die mikrovaskulären Endothelzellen in den verbliebenen vaskulären Strukturen CD31 und den vWF exprimieren. Wohingegen die histologische Unterscheidung zwischen Fibroblasten und Skelettmuskelzellen nicht möglich ist. Zusätzlich wurde die BioVaSc mit upcyte mvEC der Firma Medicyte besiedelt. Nach der vierzehntägigen Kultur im Bioreaktorsystem waren die Zellen sowohl in den Gefäßstrukturen als auch im Lumen und im Bindegewebe vital nachweisbar. In der histologischen Analyse konnte die Ausbildung von CD31, eNOS und vWF nachgewiesen werden. Des Weiteren wurde die Matrix mit mesenchymalen Stammzellen besiedelt, um zu analysieren, ob die Scherkräfte die Ausbildung endothelialer Marker stimulieren können. Nach vierzehntägiger Kultur konnte in den histologischen Analysen keine Ausbildung von CD31 oder dem vWF gefunden, allerdings vitale Zellen nachgewiesen werden. / In this work, a vascularized implant for the treatment for tracheal defects was developed according to GMP standards. For this purpose, a part of porcine small intestine was prepared, decellularized and sterilized. The remaining matrix, trademarked BioVaSc “Biological, Vascularized Scaffold”, was colonized with isolated and cultured cells from the patient and then matured for two weeks in a bioreactor system. Finally, the prepared for implantation autologous implant was extensively characterized. After the integrity check of the vessel system the decellularization process was started, which is performed by removing the porcine cells with sodium desoxycholat and enzymatic degradation of the residual DNA. As traces of sodium desoxycholat could negatively affect the seeding of the autologous cells, a test was established to demonstrate the depletion of sodium desoxycholat to acceptable traces in the final matrix preparation. Furthermore, the porcine starting material for the BioVaSc contains endotoxins, which could trigger immune reactions in the recipient if not efficiently removed. The legal limit for endotoxine levels in pharmaceutical products could be achieved through modifications of the protocol. In order to establish a GMP compliant process, specially certified chemicals were used wherever possible. The protocol was optimized until histological analysis showed only few residual cells and DNA residues. The quantitative DNA analysis revealed a decrease of 97 % of the initial DNA content. To determine storage stability, a tensile test to check elasticity of the BioVaSc was established. To colonize the matrix, autologous microvascular endothelial cells, fibroblasts and skeletal muscle cells were used. The protocols were established and optimized under GMP conditions and, wherever possible, certified reagents were used. For accurate characterization of these cells, immunohistology analyses were performed at each of the four passages for all cell types. For the final manufacturing of the autologous implant, microvascular endothelial cells were introduced into the vascular system of the BioVaSc and were cultured for seven days in a custom made bioreactor system under defined shear stress conditions resembling the human blood pressure. This was followed by culturing of skeletal muscle cells and fibroblasts in the lumen of the gut, followed by an additional seven-day culture period. Colonization of the vascular system had to be optimized in order to increase the population density as well as the efficiency of reseeding. The lumen was fully populated with fibroblasts and skeletal muscle cells by the established protocol. However, the discrimination between fibroblasts and skeletal muscle cells with normal histology was difficult because no fitting antibody was available. After a two-week culture in the custom made bioreactor system the analysis showed vital cells in the vascular structures and in the lumen of the BioVaSc. Further histological analysis were performed. In order to explore alternative cell sources, the BioVaSc was reseeded with upcyte mvEC. These transfected cells are highly proliferative and show typical endothelial markers. After fourteen days of culture in the bioreactor system, cells could be detected in vascular structures, lumen and in connective tissue. Live / dead staining and MTT identified vital cells within vascular structures. The histological analysis revealed expression of CD31, eNOS and vWF. Furthermore, the matrix was reseeded with mesenchymal stem cells; to test if shear stress triggers differentiation into endothelial like cells. This was checked through displaying the corresponding endothelial markers in histological analyses. After fourteen days of culture in the bioreactor system, histological analyzes show no expression of CD31 or vWF factor. Vital cells could be detected.

This is a report about modern theories of sustainability and the progression of regenerative design. I utilize a framework of sustainability put forth in 2001 by Simon Guy and Graham Farmer to analyze the content of the Beyond LEED Symposium held in January, 2012 on the campus of the University of Texas at Austin. Using the six logics of sustainability proposed by Guy and Farmer; eco-technic, eco-centric, eco-aesthetic, eco-cultural, eco-medical, and eco-social, I examine the results of the symposium and determine which of these logics is being employed in the development of a new, regenerative design paradigm. I will also examine whether or not the Guy and Farmer framework of sustainability, and its contained logics, represent an incomplete definition of contemporary theories of sustainability. The results of this study and of the Beyond LEED Symposium suggest a change in how we think about sustainability and regenerative design. / text

Skeletal system plays a crucial role in our life. Skeletal diseases and disorders unlike cancer, are not fatal, but affect the quality of our life. Cell-based therapeutic strategies to generate targeted desired cell types for repair or replacement of damaged skeletal tissues are ideal regenerative medicines. Because of the heterogeneous cell types generated from embryonic and mesenchymal stem cells, the ability of progenitor population to differentiate into a target cell type appear to be a better alternative for tissue regeneration. Osteo-chondroprogenitors uniquely co-expressing Sox9 and Runx2 with dual differentiation potential to become chondrocytes and osteoblasts is a progenitor cell which is suitable for cell based therapy of bone disease. Therefore, developing effective strategies to generate sufficient quantities of osteo-chondroprogenitors are essential. Toward this, we took advantage of two lineage conversion approaches. The first strategy was to interrogate the ability of osteoblasts to be reprogrammed into induced pluripotent stem (iPS) cells and another one was to use defined transcription factors to induce chondrocyte lineage from skin fibroblasts. The selection of osteoblasts is based on the fact that it is originally derived from osteo-chondroprogenitor lineage and the stochastic events of iPS induction might revert osteoblasts first to their progenitor state before becoming pluripotent. The second approach is based on a previous report using three transcription factors (Sox9, Klf4 and c-Myc) to reprogramme skin fibroblasts into chondrocyte lineage. Our aim is to examine whether osteo-chondroprogenitors would be formed during the two reprogramming processes using Sox9-EGFP knock-in mice as a reporter. We reasoned that osteoblasts can be reprogrammed into iPS cells by four Yamanaka’s factors with pluripotency as shown by their ability to form teratomas and contribute to chimeric embryos. However base on the limitation of selector marker of osteo-chondroprogenitor we still cannot capture this progenitor during iPS reprogramming. And because of the pluripotency potential, pluripotent reprogramming approach also brings high risk of teratoma formation. Therefore our second objective was performed to examine whether osteo-chondroprogenitors would be formed during lineage reprogramming. Transient appearance of Sox9-EGFP/Runx2+ve cells was observed in the intermediate stage of over 14 days of chondrocyte lineage induction from skin fibroblasts by Sox9, klf4 and c-Myc. Cells expressing Sox9-EGFP/Runx2+ve showed typical molecular markers of osteo-chondroprogenitors. In vitro and in vivo differentiation assays demonstrated that Sox9-EGFP/Runx2+ve cells can differentiate predominantly into osteoblasts and chondrocytes. Taken together our data indicate that cells with osteo-chondrogenic potential could be generated by defined transcription factors-mediated reprogramming processes. / published_or_final_version / Biochemistry / Doctoral / Doctor of Philosophy

The fungal metabolites vinaxanthone and xanthofulvin possess the remarkable ability to restore motor function in animal models of complete spinal cord transection making them the most promising small molecules for the development of spinal cord injury (SCI) therapeutics. A concise nine-step total synthesis of vinaxanthone was accomplished utilizing a biomimetic dimerization of the putative precursor 5,6-dehydropolivione and the first reported synthesis of xanthofulvin was achieved in 15-steps highlighted by an unprecedented enaminone O-to-C carboxyl transfer to forge key carbon-carbon bonds. Both natural products were also identified as positive allosteric modulators of the G-protein coupled receptor (GPCR), GPR91, thus elucidating their modes of action accounting for their regenerative capabilities. Furthermore, a unique ynone coupling reaction was developed in order to access various vinaxanthone analogs for structure activity relationship (SAR) studies. This resulted in the preparation of a small molecule library of 25 vinaxanthone analogs that demonstrated pronounced neuronal regeneration within laser axotomy assays performed in vivo on C. elegans. The plant derived natural product eupalinilide E has been found to promote the ex vivo expansion of hematopoietic stem and progenitor cells (HSPCs) which have the potential to improve the success of medical procedures such as bone marrow transplants. In light of its promising applications, unknown mechanism of action, and scarcity in nature the total synthesis of eupalinilide E was undertaken. Efforts culminated in the first enantioselective total synthesis of the natural product in 20-steps, which showcases a Favorskii rearrangement, borylative enyne cyclization, aldehyde-ene ring closure, and a dual allylic oxidation. / text