Thesis (M. S.)--Biomedical Engineering, Georgia Institute of Technology, 2008. / Committee Chair: Boyan, Barbara; Committee Member: Bonassar, Lawrence; Committee Member: Sambanis, Anthanassios; Committee Member: Schwartz, Zvi; Committee Member: Wick, Timothy.

Thesis (Ph. D.)--University of Missouri-Columbia, 2007. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Vita. "December 2007" Includes bibliographical references.

Habilitation-Thesis--Cologne. / Includes bibliographical references (p. 45-48) and index.

Thesis (Ph. D.)--Mechanical Engineering, Georgia Institute of Technology, 2008. / Barbara D. Boyan, Committee Member ; Ravi Bellamkonda, Committee Member ; Joseph Le Doux, Committee Member ; Andres J. Garcia, Committee Member ; Marc E. Levenston, Committee Chair.

<p>The ends of long bones that articulate with respect to one another are lined with a crucial connective tissue called articular cartilage. This tissue plays an essential biomechanical function in synovial joints, as it serves to both dissipate load and lubricate articulating surfaces. Osteoarthritis is a painful and debilitating disease that drives the deterioration of articular cartilage. Like many chronic diseases, pro-inflammatory cytokines feature prominently in the onset and progression of osteoarthritis. Because cartilage lacks physiologic features critical for regeneration and self-repair, the development of effective strategies to create functional cartilage tissue substitutes remains a priority for the fields of tissue engineering and regenerative medicine. The overall objectives of this dissertation are to (1) develop a bioactive scaffold capable of mediating cell differentiation and formation of extracellular matrix that recapitulates native cartilage tissue and (2) to produce stem cells specifically tailored at the scale of the genome with the ability to resist inflammatory cues that normally lead to degeneration and pain. </p><p>Engineered replacements for musculoskeletal tissues generally require extensive ex vivo manipulation of stem cells to achieve controlled differentiation and phenotypic stability. By immobilizing lentivirus driving the expression of transforming growth factor-β3 to a highly structured, three dimensionally woven tissue engineering scaffold, we developed a technique for producing cell-instructive scaffolds that control human mesenchymal stem cell differentiation and possess biomechanical properties approximating those of native tissues. This work represents an important advance, as it establishes a method for generating constructs capable of restoring biological and mechanical function that may circumvent the need for ex vivo conditioning of engineered tissue substitutes.</p><p>Any functional cartilage tissue substitute must tolerate the inflammation intrinsic to an arthritic joint. Recently emerging tools from synthetic biology and genome engineering facilitate an unprecedented ability to modify how cells respond to their microenvironments. We exploited these developments to engineer cells that can evade signaling of the pro-inflammatory cytokine interleukin-1 (IL-1). Our study provides proof-of-principle evidence that cartilage derived from such engineered stem cells are resistant to IL-1-mediated degradation. </p><p>Extending on this work, we developed a synthetic biology strategy to further customize stem cells to combat inflammatory cues. We commandeered the highly responsive endogenous locus of the chemokine (C-C motif) ligand 2 gene in pluripotent stem cells to impart self-regulated, feedback-controlled production of biologic therapy. We demonstrated that repurposing of degradative signaling pathways induced by IL-1 and tumor necrosis factor toward transient production of cytokine antagonists enabled engineered cartilage tissue to withstand the action of inflammatory cytokines and to serve as a cell-based, auto-regulated drug delivery system.</p><p>In this work, we combine principles from synthetic biology, gene therapy, and functional tissue engineering to develop methods for generating constructs with biomimetic molecular and mechanical features of articular cartilage while precisely defining how cells respond to dysfunction in the body’s finely-tuned inflammatory systems. Moreover, our strategy for customizing intrinsic cellular signaling pathways in therapeutic stem cell populations opens innovative possibilities for controlled drug delivery to native tissues, which may provide safer and more effective treatments applicable to a wide variety of chronic diseases and may transform the landscape of regenerative medicine.</p> / Dissertation

Orientador: Bruno Watanabe Minto / Banca: Luis Gustavo Gosuen Gonçalves Dias / Banca: Paulo César Jark / Resumo: A cartilagem articular possui capacidade de reparação limitada, aumentado a predisposição ao desenvolvimento de alterações degenerativas, muitas vezes irreversíveis. Diversas formas de tratamento, cirúrgicas ou conservativas, são descritas, entretanto a terapêutica da osteoartrite continua sendo grande desafio ao médico veterinário. Neste contexto, a pesquisa envolvendo células-tronco mesenquimais destaca-se na busca de melhorias e avanços na reparação da cartilagem articular. Objetivou-se, no presente projeto, comparar a regeneração cartilaginosa da articulação coxofemoral de coelhos, com e sem o transplante de células-tronco mesenquimais autólogas, por meio de exames radiográficos e histopatológicos. Dois grupos, com 15 animais da espécie leporina cada, foram submetidos à indução química de osteoartrite com solução de colagenase 2% na articulação coxofemoral direita. No Grupo 1 (Células-tronco) realizou-se a aplicação intra-articular de células-tronco mesenquimais autólogas, enquanto que, o Grupo 2 (Controle) foi constituído por animais submetidos à aplicação intra-articular de solução salina estéril. Foram realizadas avaliações radiográficas e histopatológicas aos 30, 60 e 90 dias após a aplicação. Os resultados histológicos deste ensaio indicam que células-tronco mesenquimais (Grupo 1) melhoraram discretamente a qualidade do tecido de reparo, de acordo com os critérios da escala semi-quantitativa ICRS 1 ("International Cartilage Repair Society"). O Grupo 1 (Células-Tronco... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The articular cartilage has limited repair capacity, leading to an increased risk for degenerative changes, potentially irreversible. Several treatments, surgical or not, are described, however osteoarthritis remains a major challenge for the veterinarian. In this context, research involving mesenchymal stem cells stands out. The aim of this study was to compare cartilage regeneration of the hip in rabbits, with and without the transplantation of autologous mesenchymal stem cells. Radiographic and histopathological evaluation were used. Thirty rabbits were submitted to chemical induction of osteoarthritis with a 2% colagenase in the right hip. They were divided into 2 groups of 15 animals each: Group 1 (intra-articular application of autologous mesenchymal stem cells) and Group 2 (control - intra-articular application of sterile saline solution). Radiographic and histopathological evaluations were performed at 30, 60 and 90 days after application. The mesenchymal stem cells group (Group 1) showed slight improvement of the quality of the repair tissue, according to the semi-quantitative scale criteria ICRS 1 (International Cartilage Repair Society). The Group 1 (Stem Cells) showed superiority in relation to Group 2, specially in the parameters joint surface, extracellular matrix and cellular distribution. / Mestre

Made available in DSpace on 2014-06-11T19:31:09Z (GMT). No. of bitstreams: 0 Previous issue date: 2011-04-25Bitstream added on 2014-06-13T19:47:15Z : No. of bitstreams: 1 meirelles_vm_dr_jabo.pdf: 1134100 bytes, checksum: b4602b4a9ef93b3deb00fc8b50e5af14 (MD5) / A osteoartrose (OA) do joelho é uma doença de caráter inflamatório e degenerativo que provoca a destruição da cartilagem articular e leva à deformidade da articulação. A luxação patelar é um problema ortopédico comum na clínica de pequenos animais. A cirurgia não impede a progressão da OA. O laser de baixa intensidade (LBI) tem sido utilizado para acelerar processos de reparação tecidual, por aumentar o fluxo sangüíneo, possuir ação antiinflamatória, antiedematosa, analgésica e estimular o metabolismo celular. Este trabalho estudou a influência da laserterapia em dois espectros eletromagnéticos, vermelho e infravermelho, na reparação cartilagínea, diferenciando histologicamente o tipo de cartilagem formada durante a reparação articular. Foram utilizados 36 coelhos machos raça Norfolk divididos em 3 grupos com 12 animais cada: G1 (controle, trocleoplastia sem tratamento pós-operatório), G2 (trocleoplastia + irradiação laser 670 nm) e G3 (trocleoplastia + irradiação laser 904 nm). Os animais dos grupos G2 e G3 foram irradiados diariamente, com intervalo de 24 horas, durante 10 dias utilizando um laser In-Ga-Al com comprimento de onda (λ) de 670 e um laser de As-Ga com λ de 904 nm respectivamente, na dose de 2 J por ponto totalizando 4 pontos. Os grupos G2 e G3 apresentaram recuperação funcional mais rápida que G1. Histologicamente, G3 apresentou um aspecto osteocondral mais regular, com formação de cartilagem hialina enquanto em G1 e G2 formou-se fibrocartilagem / Stifle osteoarthrosis is an inflammatory and degenerative disease that causes destruction of the articular cartilage and leads to deformity in the articulation. Patellar luxation is a common orthopedic problem in the clinic of small animals. The surgery does not avoid the progression of the osteoarthrosis. Low intensity laser has been used to improve degenerated processes for it increases blood flow, it has antiinflammatory, antiedematous, analgesic effects and stimulates cellular metabolism. This work studied the influence of laser therapy of two electromagnetic spectra, red and infrared, for repairing cartilage, histologically telling apart the kind of cartilage formed during articular regeneration. We use 36 male rabbits bred Norfolk divided into 3 groups with 12 animals each: G1 (control, trochleoplasty without treatment post-operatory), G2 (trochleoplasty + laser therapy 670 nm) and G3 (trochleoplasty + laser therapy 904 nm). The animals from G2 and G3 were irradiated daily with an interval of 24 hours over 10 days using a laser Ga-Al-In with wavelength (λ) of 670 and a Ga-As laser with λ 904 nm respectively. At a dose of 2 J per point total of 4 points. The groups G2 and G3 showed faster functional recovery that G1. Histologically, G3 had a most regular osteochondral aspect, with hyaline cartilage formation while in G1 and G2 formed fibrocartilage

Orientador : Vitor Baranauskas / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Engenharia Eletrica e de Computação / Made available in DSpace on 2018-07-29T02:00:52Z (GMT). No. of bitstreams: 1 Guerino_MarceloRenato_D.pdf: 16707408 bytes, checksum: f510ea4730bed4c37a782d27bb313981 (MD5) Previous issue date: 2001 / Resumo: Este trabalho apresenta os efeitos da radiação 1aserno processo de cura da inflamação induzida por injeção intra-articular de adjuvante de Freund incompleto na cartilagem articular do joelho de cobaias. A radiação dos animais foi feita a partir do quinto dia da indução, com doses de 0,5, 7,0 e 50 J/cm2 (laser de HeNe) e de 1,0, 5,0 e 25,0 J/cm2 (1aser de GaAs). As cartilagens foram analisadas 7, 14 e 21 dias após a indução, através da Microscopia Eletrônica de Varredura (MEV), Microscopia de Força Atômica (MFA), teor de hidroxiprolina e Testes Mecânicos (rigidez). A MEV mostrou evidências de melhor organização na reparação da cartilagem articular dos animais tratados com o 1asers de HeNe e GaAs em relação aos animais inflamados sem tratamento. As doses mais eficientes foram: 7,0 J/cm2(ReNe) e: 1,0 J/cm2 (GaAs). O tratamento de HeNe, na MFA pôde identificaruma melhor organização das moléculas de colágeno, formação de feixes mais espessos, melhor direcionalidadee maior agrupamento das fibrilas que indicamuma atuação positiva do laser sobre o colágeno. Em relação ao teor de hidroxiprolina, o 1aserde HeNe pareceu ter efeito excitatório nas doses maiores e inibitório nas doses menores. Os maiores aumentos no teor de hidroxiprolina foram encontrados com a dose de 50,0 J/cm2 (HeNe). Entretanto o laser de GaAs produziu efeito inibidor no processo de sintese de colágeno em quase todas etapas. O teste de indentação mecânica demonstrou uma tendência a aumento da rigidez pelos lasers quando comparados aos animais inflamados sem tratamento. A dose mais adequada foi de 7,0 J/cm2com o laser de HeNe / Abstract: This work presents the effects of 1aserirradiationon the curing process of inf1ammationinduced in the articular cartilage of Gufuea Pigs's knee. The int1ammationwas induced by intra-articular injection of incomplete Freund's Adjuvant. Irradiation of the animaIswas undertaken irom the fifth day of induction, with doses of 0.5, 7.0 and 50 J/crrl-(HeNe) and of 1.0, 5.0 and 25.0 J/cm2 (GaAs). The cartilages were analysed at 7, 14 and 21 days after induction, Scanning Electron Microscopy (SEM), Atomic Force Microscopy (AFM), concentration of Hidroxyprolyne and Mechanical Tests (rigidity). SEM showed evidence of better organization in the repair of the articular cartilage ofthe animals treated with the HeNe and GaAs 1asersin relation to the untreated animals. The most efficient doses were: 7.0 J/cm2 (HeNe) and 1.0 J/cm2 (GaAs). Following the laser treatment of ReNe, AFM better organization of the molecules of collagen, fonnation of thicker bunches, better direction and larger grouping ofthe fibrils,which indicate the positive effect of the 1aser on the collagen. In relation to the hidroxyprolyne concentration, the HeNe laser seemed to have a stirnulating effect great doses and an inlu'bitory effect in smaller doses. The largest increases in the hydroxiprolyne concentration were found with the dose of 50.0 J/crrl- (HeNe). However, the laser of GaAs produced an inhibitory effect the process of synthesis of collagen in almost alI of stages. The mechanical indentation test, showed a tendency of increasing rigidity of the treated tissue, compared with the animaIs inflamed without treatment. The most effective dose was of 7.0 J/crrl- (HeNe) / Doutorado / Engenharia Eletrica / Doutor em Engenharia Elétrica

Disorders affecting articular cartilage are amongst the most common problems in orthopaedics. Osteoarthritis, the end stage of the disease of articular cartilage, reduces the quality of life for tens of millions of people in the world, and has a profound impact on the economics of industrialized countries. Despite progress in articular cartilage research, the problem is still far from being defeated. Various models e.g. in vitro cartilage explants and in vivo animal models, have been established for cartilage research, but each has its own limitations. Thus, a novel ex vivo isolated joint organ culture model was developed. Bovine metatarsophalangeal joints were chosen as a suitable synovial joint because it consists of a hinge-type joint that is similar to the human knee joint, and has a large cartilage surface that provides enough space for multiple sampling in the same joint. The joints were isolated aseptically and placed into culture media. The viability of chondrocytes, glycosaminoglycan (GAG) content of cartilage matrix, cartilage morphology and water content of matrix were evaluated under different culture conditions, i.e. static, static with flowing media, and dynamic with different durations of the movement period. The model was used to investigate the effect on the sharp scalpel cartilage injury of adding serum to the culture medium by culturing the whole joint explants in serum-supplied or serum-free media. The feasibility of investigating the early phases of chondrocyte implantation in this model was also studied: circular holes of 2.5 mm diameter were created by making a pilot hole with a 2.0 mm drill followed by using a fresh 2.5 mm biopsy punch. Allogeneic isolated chondrocytes at different passages were aggregated as cell pellets and implanted in the holes to evaluate their integration ability and the response from the recipient cartilage. Results from the static model showed that, after 28 days culture, the chondrocytes were still alive with 66.5%, 80.9% and 46.9% viability in the superficial, middle and deep zones, respectively. The GAG content of the static model decreased 19.2% after the first week of culture and then lost another 15.0% during the third week. Paradoxically, at end of the 4th week the GAG level rebounded to some extent and increased 19.0% relative to the previous week. Interestingly, the cell viability of all three zones improved if the culture fluid was flowing as seen with the experiments carried out with stirred media or dynamic movement of the articular surfaces. (e.g. for the stirred media after 28 days of culture the chondrocyte viability was 80.6%, 92.4% and 70.4% for the superficial, middle and deep zones respectively.) The GAG content was maintained at a constant level in the contact area of the dynamic model, but decreased as in the media-stirred model and non-contact area of the dynamic model to a similar extent to that observed with the static model. In the injury model, the GAG content fell approximately 10.8% straight after the scalpel cut, but no further loss was observed if the joint was cultured in the serum-supplied media. In contrast, if the injured joint was cultured in the serum-free media, the GAG content continued to fall week by week and finally dropped by 41.7% at the end of the 4th week. In the chondrocyte implantation model, the majority of the host chondrocytes around the circular defect were alive (78.5 % viability). Viewed from the surface, the dead cells were all within 20 μm from the cut edge. The implanted chondrocytes, which were aggregated as cell pellets, began to transform their shapes and spread to the surrounding surface of the recipient cartilage, but did not appear to integrate with the host tissue during the first 2 weeks of culture. The results supported the validity of this ex vivo joint model and demonstrated that the chondrocytes subjected to flow of the media or dynamic loads survived well over a 4 week period. Of importance was the finding that there was no measured loss of the matrix GAG content when the joints were under dynamic load compared to all of the non-loaded conditions. This whole joint model could be of value in providing a more natural and controllable platform where research involving the normal processes or pathologic mechanisms of articular cartilage can be investigated, as well as the early response to newly developed pharmacological agents and cartilage tissue engineering constructs.

Doctor of Philosophy / Department of Animal Sciences and Industry / Jim L. Nelssen / Four experiments using 350 pigs were conducted to determine the effects of dietary nutrients on the incidence of osteochondrosis (OC) and to evaluate the use of biomarkers to predict its occurrence in growing-finishing pigs. The first experiment was conducted to evaluate the potential of dietary ingredients with known functions in cartilage and bone metabolism on incidence of OC in pigs (PIC 327 × 1050, initially 39 kg). Results suggest that pigs fed high levels of added copper and manganese, silicon, methionine and threonine, or proline and glycine had reduced OC severity scores. A second experiment evaluated other dietary ingredients that may impact OC as well as with or without ractopamine HCl (PIC 327 × 1050, initially 47 kg). Results suggest that feeding a combination of added methionine, manganese, proline, and glycine or added silicon can numerically reduce OC severity scores compared to pigs fed a standard corn-soybean meal based diet. Feeding ractopamine HCl did not affect the incidence or severity of OC. A third experiment was conducted to evaluate the effect of dietary lysine concentration with or without the addition of high methionine, manganese, and copper in a 3 × 2 factorial arrangement in growing-finishing pigs (PIC 327 × 1050, initially 41 kg). Results suggest that increasing dietary lysine concentrations increased the severity of OC. Furthermore, feeding additional methionine, manganese, and copper reduced OC severity scores. A fourth experiment was conducted to determine the usefulness of ten different biomarkers of cartilage and bone metabolism to predict the incidence of OC and the correlation of individual markers with the severity of OC. Results suggest that measuring serum C-propeptide of type II collagen (CPII) will predict animals with OC, as gilts with a two-fold increase in CPII are 97 times more likely to have OC. As well, serum collagen type II carboxy-terminal 3/4 long fragment (C2C) explained 49% of the variation in OC severity scores. The results of these experiments suggest that feeding added copper, manganese, methionine, silicon, or proline and glycine may reduce OC severity scores in pigs, while measuring biomarkers CPII and C2C can aid in determining animals with OC.