O objetivo deste trabalho foi avaliar a membrana de celulose bacteriana associada a células mesenquimais, que foram inoculadas juntamente com derivado do ácido hialurônico, hylano G-F 20, em patelas lesadas de suínos. Foram utilizados 12 suínos (24 joelhos) submetidos à raspagem das superfícies articulares das patelas até osso esponjoso; joelhos esquerdos sem tratamento, grupo controle (GC). Joelhos direitos, grupo tratado (GT), tiveram membranas de celulose bacteriana fixadas às superfícies de suas patelas, sendo que as mesmas foram preparadas com inoculação de células mesenquimais, ósseas e cartilaginosas, provenientes das raspagens das superfícies, lavadas em solução fisiológica 0,9%, filtradas e diluídas em 5 ml de solução fisiológica à 0,9% e posteriormente acrescentados 2 ml de hylano G-F 20. Colhidos 5 ml dessa papa que teve contido o volume de 16,35 x \'10 POT.8\' células na sua totalidade, e foram infiltrados na membrana de celulose revestindo a patela, obedecendo a distribuição de 3,27 x \'10 POT.8\' por 1,2 \'CM POT.3\'. Após 2, 4 e 6 meses os animais foram sacrificados para avaliações macroscópicas e histológicas. As superfícies patelares apresentaram aspecto de neoformação de cartilagem aos dois meses; essas membranas evoluíram para tecido fibroso de altíssima resistência, com densidade de fibras conjuntivas aumentando proporcionalmente ao tempo de cirurgia, quatro ou seis meses conforme as células do infiltrado tiveram alterações morfológicas para fibroblastos Ao mesmo tempo, parte do volume dessas células sofreu processo de degeneração e morte celular, semelhante ao mecanismo de apoptose celular. Dessa forma, ofereceu substratos para as células remanescentes, colaborando para a neoformação desse tecido conjuntivo. Não ocorreram processos inflamatórios ou agressão às cartilagens em contigüidade à membrana. Pelo contrário, em relação ao grupo controle não tratado, a membrana de celulose bacteriana utilizada segundo a metodologia aplicada, atuou com uma barreira protetora para agressões maiores à cartilagem da fossa intercondilar do fêmur que manteve a integridade de seus contornos articulares. Desta forma, segundo a metodologia utilizada, concluímos que a membrana de celulose bacteriana apresentou biocompatibilidade para uso articular, hábil para agregar condrócitos e fibroblastos. / The goal of this work was to evaluate the bacterial cellulose membrane associated with mesenchymal cells, which were inoculated with hyaluronic acid derivative, G-F 20 hylano in patelas injured pigs. 12 pigs were used (24 knees) submitted to the scraping of the articular surfaces of spongy bone patelas up; knees left without treatment, group controller (GC). Knees rights, treated group (GT), bacterial cellulose membranes had fixed on their surfaces patelas, being that they were prepared with inoculation of mesenchymal cells, bone and cartilage from sweeps of surfaces, washed in 0.9% saline solution, filtered and diluted in 5 ml of saline solution to 0.9% and subsequently added 2 ml hylano G-F 20. Collected 5 ml that Pope had contained the volume 16.35 x \'10 POT.8\' cells in its entirety, and were infiltrated in the membrane of cellulose coating the patella, obeying distribution 3.27 x \'10 POT.8\' per 1.2 \'CM POT.3\'. After 2, 4 and 6 months the animals were sacrificed to macroscopic and histological evaluations. The surfaces patellar components submitted aspect of neoplasm of cartilage to two months; These membranes evolved into fibrous tissue resistance of high density fibrous tears increasing in proportion to the time of surgery, four or six months as had infiltrated cells for morphological changes while fibroblasts, part of the volume of these cells suffered process of degeneration and cell death, similar to the mechanism of cellular apoptosis. This way, offered substrates for the remaining cells, collaborating for this connective tissue neoplasm. There were no inflammatory processes or aggression on cartilage in adjacency to the membrane. On the contrary, compared to the untreated control group, the bacterial cellulose membrane used according to the methodology applied, served with a protective barrier for larger aggressions of cartilage of the femur intercondilar trench that maintained the integrity of its articular contours. This way, according to the methodology, we found that the bacterial cellulose membrane biocompatibility presented for use articulate, skilled to aggregate condrócitos and fibroblasts.

Articular cartilage damage and degeneration is a siginficant clinical problem which no technique has been able to adequately and reliably repair or regenerate. Recent research has investigated the use of cell-based therapies to treat focal cartilage lesions. In clinical practice proliferated autologous chondrocytes are used and clinical trials are investigating the use of mesenchymal stem cells. The aim of this thesis was to assess aspects of current cell-based therapy and to investigate the potential of perivascular stem cells for articular cartilage repair. The phenotype of expanded matrix-applied autologous chondrocytes utilised in current cell therapies was confirmed using immunocytochemistry and polymerase chain reaction (PCR) expression of hyaluronan and proteoglycan link protein 1 (HAPLN1), transcription factor sox-9 (SOX9) and aggrecan (ACAN). Quantitative real-time PCR demonstrated that they were down-regulated for expression of COL2A1, SOX9 and ACAN but up-regulated for COL1A1 compared to unproliferated chondrocytes. Confocal laser-scanning microscopy (CLSM) demonstrated a significant decrease in cell viability and density when the membranes were subjected to levels of trauma similar to those that could be experienced during surgery. Hyperosmolar solutions did not confer a chondroprotective effect. Pericytes and adventitial cells, collectively termed perivascular stem cells (PSCs), from the infra-patellar fat pad were identified using immunohistochemistry and isolated using enzymatic digestion and fluorescence-activated cell sorting (FACS). Cell identity was ascertained using PCR, FACS and mesenchymal differentiation (osteogenesis, adipogenesis and chondrogenesis). Quantitative real-time PCR analysis of micromass cultures indicated that PSCs displayed increased chondrogenic potential compared to mesenchymal stem cells. An ovine model of perivascular stem cells was developed and a pilot study using three sheep was undertaken to confirm the viability of the model. Autologous ovine PSCs were isolated and re-implanted into articular cartilage defects. Green fluorescent protein transfected cells were identified in the cartilage defect four weeks following re-implantation using CLSM. This thesis has examined aspects of matrix-applied autologous chondrocyte implantation for cell based cartilage repair and has identified a new source of prospectively identified and purified stem cells that have demonstrated increased chondrogenic potential compared to mesenchymal stem cells, which are commonly used in clinical research. The methods to identify and purify ovine perivascular stem cells were developed to investigate the use of autologous PSCs and to track the cells following implantation.

Vários estudos demonstraram que o estrógeno exerce um papel fundamental na manutenção da estrutura da cartilagem articular, e que a deficiência em mulheres podem causar um aumento na incidência de degeneração articular. Sabe-se também que a atividade física é capaz de trazer benefícios para este tipo de tecido, reduzindo os riscos de lesão articular. Contudo, existem poucos os trabalhos que relacionam estes dois fatores: deficiência estrogenica e atividade física regular. O objetivo deste trabalho é verificar os efeitos da deficiência estrogênica sobre a cartilagem articular, suas implicações, e se a atividade física regular e moderada é capaz de reverter este processo natural de degeneração articular. Foram utilizadas 12 ratas Wistar adultas, divididas em quatro grupos de três animais, de modo aleatório e sistematizado, sendo eles: ratas controle sedentárias (RCS), ratas controle treinadas (RCT), ovariectomizadas sedentárias (ROS) e ovariectomizadas treinadas (ROT). Cada grupo foi analisado pelas técnicas de microscopia de luz e microscopia eletrônica de transmissão. Os resultados foram obtidos a partir do método estereológico. Os valores foram estatisticamente significantes em relação a deficiencia estrogenica para os parametros: peso, condicionamento físico, densidade numérica de condrócitos, número total de condrócitos, densidade de volume dos condrócitos, densidade de volume da matriz extracelular, volume total de matriz extracelular e densidade de volume de colágeno. Em relação aos efeitos concomitantes da atividade física e da deficiência estrogênica, os resultados foram significativamente diferentes em relação a densidade numérica, volume total da cartilagem, volume médio dos condrócitos e densidade de volume de colágeno. Estes dados nos levam a concluir que a diminuição dos níveis de estrógeno afeta profundamente a cartilagem articular da cabeça do fêmur, enquanto a atividade fisica contínua é benéfica para esta cartilagem, incluseve quando há deficiência estrogênica. / Several studies show that estrogen has an important function in maintaining the structure of articular cartilage and that deficiency in women can cause an increase in the incidence of joint degeneration. It is also known that physical activity can bring benefits to this type of tissue, reducing the risk of joint damage. However, there are few works that relate these two factors: continuous physical activity and estrogen deficiency. The aim of this study is to verify the effects of estrogen deficiency on joint cartilage, its implications, and if frequent and moderate physical activity is able to reverse the natural process of joint degeneration. A total of 12 adult female rats were divided into four groups of three animals in a random and systematic way, as follows: sedentary control rats (RCS), trained control rats (RCT), ovariectomized sedentary rats (ROS) and trained ovariectomized rats (ROT). Each group was analyzed by light microscopy and transmission electron microscopy. Results were obtained from the stereological method. The values were statistically significant in relation to estrogen deficiency for the parameters: weight, physical condition, numerical density of chondrocytes, total number of chondrocytes, chondrocyte volume density, volume density of extracellular matrix, the total volume of the extracellular matrix and volume density of collagen. Regarding the concomitant effects of physical activity and estrogen deficiency, the results were significantly different in numerical density, total volume of cartilage, mean volume of chondrocytes and volume density of collagen. These data take us to conclude that the decrease in estrogen levels profoundly affects the articular cartilage of the femoral head, while the continuous physical activity is beneficial to this cartilage, also in estrogen deficiency.

Vários estudos demonstram que a atividade física é capaz de trazer benefícios para as cartilagens articulares. Estes manifestam-se reduzindo os riscos de lesão articular, prevenindo aderências na articulação, aumentando e mantendo sua mobilidade e combatendo a obesidade, grande fator de risco para o desenvolvimento de lesões articulares. Por outro lado, sabe-se que o envelhecimento provoca alterações deletérias em vários tecidos, inclusive nas cartilagens articulares. O objetivo deste trabalho foi verificar se a atividade física regular promove alterações no processo de envelhecimento da cartilagem articular da cabeça do fêmur de ratos Wistar corredores. Para este estudo foram utilizados 24 ratos Wistar corredores, os quais foram divididos em três grupos de oito animais cada: Sedentários (S) - ratos adultos sedentários; Treinados (T) - ratos adultos treinados; e Controle (C) - ratos adultos jovens. As cartilagens articulares das cabeças femorais foram analisadas por dois métodos: microscopia de luz (três de cada grupo) e microscopia eletrônica de varredura (cinco de cada grupo). Os resultados em microscopia de luz foram obtidos a partir do método esteriológico. Os valores não foram estatisticamente diferentes em relação à idade e ao treinamento para os parâmetros: volume total da cartilagem, número total de condrócitos, densidade de volume de matriz e volume total de matriz (P=0,05). Os dados foram significantes em \"border line\" para idade nos parâmetros densidade do volume celular e volume celular indireto (P=0,058) e para treinamento no parâmetro densidade celular (P=0,058). À microscopia eletrônica de varredura foi possível observar que no grupo S a superfície articular da cabeça femoral apresentava-se lisa e com áreas de erosões bem evidentes, enquanto que, no grupo T e C, haviam nítidas ondulações, sem presença de erosões. Estes dados nos levam a concluir que um programa de atividade física constante durante o envelhecimento é benéfico para a cartilagem articular da cabeça do fêmur, pois sugere que a cartilagem dos animais do grupo S foi mais susceptível a lesões do que as do grupo T e C, e que a atividade física promoveu a estimulação dos condrócitos, aumentando, assim, a resistência da cartilagem às modificações deletérias do envelhecimento. / Several studies has demonstrate that the physical activity can bring benefits for the articular cartilage. These show a reduction in articular lesion risks, preventing the adherences in the articulation, increasing and maintaining its mobility and combating the obesity, which is a great risk factor for the development of articulate lesions. On the other hand, it is known that the aging provokes harmful alterations in several tissues, including articular cartilage. The objective of this work was to verify if the regular physical activity promotes alterations in the process of aging of the articular cartilage of the femoral head of Wistar runner rats. For this research were used 24 Wistar runner rats. The animals were divided in three groups containing eight animals each one: Sedentary (S) - adult sedentary rats; (T) - adult trained rats; and Control (C) - young adult rats. The articular cartilages of the femoral heads were analyzed by two methods: light microscopy (three each group) and scanning electron microscopy (five each group). The results in light microscopy were obtained using stereological method and there were not statistically significant differences in relation to age and training for the parameters: total volume of the cartilage, total number of chondrocytes, volume density of extra cellular matrix and total volume of extra cellular matrix (P=0,05). The data were statistically significant in \"border line\" for age in the parameters chondrocyte volume density and mean chondrocyte volume (P=0,058) and for training in the parameter cellular density (P=0,058). By scanning electron microscopy was possible to observe that in S group, the surface of articular cartilage of the femoral head came flat and with areas of evident erosions, while, in T group, there were clear undulations, without the presence of erosions. These data take us to conclude that a program of constant physical activity during the aging promotes benefits for the articular cartilage of the femoral head by suggests that the cartilage of the animals of the S group was more susceptible to lesions than the ones of the T and C group and that the physical activity promoted the stimulation of the chondrocytes, increasing the resistance of the cartilage to the harmful modifications of the aging.

O resveratrol (RES), um composto natural, é encontrado em diversas plantas, mas pode ser observado em altas concentrações na uva e nos vinhos tintos, apresentando propriedades antioxidantes e condroprotetoras capazes de proteger a cartilagem articular, no entanto, os efeitos do resveratrol frente à subnutrição ainda são desconhecidos. Assim, o presente estudo teve por objetivo avaliar os efeitos da suplementação com resveratrol sobre o tecido cartilagíneo do joelho de ratos submetidos a uma subnutrição protéica. Para tanto, foram formados 6 grupos experimentais de animais subnutridos e nutridos nos períodos pré e pós-natal com 21 e 60 dias de vida (Grupos S21 e N21; e S60 e N60, respectivamente). Dois grupos de 60 dias, um submetido à recuperação protéica e outro suplementado com resveratrol (ambos a partir do 22º dia) formaram, respectivamente os grupos RN60 e RES60. Após o desmame, foram mensurados os parâmetros metabólicos de ingestão hídrica e alimentar, bem como os de excreção de fezes e urina. As amostras da articulação do joelho foram processadas com técnicas rotineiras de histologia e submetidas a colorações que permitam evidenciar os componentes cartilagíneo e de colágeno. Os aspectos tridimensionais e ultraestruturais da cartilagem articular também foram analisados. Para as avaliações relativas aos dados metabólicos foi utilizada a Análise de Variância - ANOVA com dois fatores (idade e dieta) e quando necessário aplicou-se pós-teste de Bonferroni. Para as demais variáveis empregou-se a Análise de Variância - ANOVA com dois fatores (idade e dieta) e a Análise de Variância ANOVA com um fator e, quando necessário, aplicaram-se comparações múltiplas pelo método de Tukey. Os nossos resultados mostram que a subnutrição é capaz de modificar as propriedades biomecânicas das cartilagens articulares do fêmur e da tíbia, causando atraso no seu desenvolvimento. Quanto às dietas aqui empregadas, mostram que a renutrição protéica melhora os efeitos da subnutrição sobre a cartilagem articular, mas não recupera em comparação ao grupo controle, enquanto que, a renutrição suplementada com resveratrol, reestabelece os parâmetros analisados. Conclui-se que o resveratrol auxiliou na recuperação dos efeitos deletérios da subnutrição, melhorando os aspectos metabólicos, estruturais e ultraestruturais da cartilagem articular do fêmur e da tíbia em fases precoces do desenvolvimento. / Resveratrol (RES), a naturally occurring compound is found in several plants, but can be observed at high concentrations in grapes and red wines having antioxidant and chondroprotective properties and able to protect the articular cartilage, however, the effects of resveratrol front malnutrition are still unknown. Thus, this study aimed to evaluate the effects of supplementation with resveratrol on the cartilage tissue of the knee of rats submitted to protein malnutrition. Thus, we formed six experimental groups of malnourished and nourished animals pre- and postnatal 21 and 60 days (Groups S21 and N21, and N60 and S60, respectively). Two groups of 60 days, one submitted to protein recovery and other supplemented with resveratrol (both from the 22nd day) formed respectively the RN60 and RES60 groups. After weaning, metabolic parameters of water and food intake and the excretion of faeces and urine were measured. The knee joint specimens were processed for routine histology staining techniques and submitted to which emphasize the collagen and cartilage components. Three-dimensional and ultrastructural aspects of articular cartilage were also analyzed. For assessments of the metabolic data was used analysis of variance - ANOVA with two factors (age and diet) and when necessary applied Bonferroni post-test. For the other variables we used analysis of variance - ANOVA with two factors (age and diet) and analysis of variance - ANOVA with one factor and, when necessary, we applied multiple comparisons by the Tukey method. Our results show that malnutrition is able to modify the biomechanical properties of articular cartilage of the femur and tibia, causing delays in their development. The diets used here, show that the protein renutrition improves the effects of malnutrition on the articular cartilage, but does not recover as compared to the control group, while renutrition supplemented with resveratrol, restores the parameters analyzed. We conclude that resveratrol helped him overcome the deleterious effects of malnutrition, improving metabolic, structural and ultrastructural aspects of the articular cartilage of the femur and tibia in the early stages of development.

Articular cartilage represents a primary target for tissue engineering strategies as it does not functionally regenerate within the joint. Many tissue engineering approaches have focused on the in vitro generation of neo-cartilage using chondrocyte-seeded scaffolds. Several studies have reported the morphological appearance of native cartilage, although its functional competence has not been demonstrated. Accordingly, mechanical conditioning has often been introduced to enhance biosynthetic activity of chondrocytes within 3D constructs. However although this strategy has significantly up-regulated proteoglycan synthesis, its effects on the synthesis of the other major solid constituent, type II collagen, has been modest. Analyses of normal joint activities reveal that cartilage is subjected to shear superimposed on uniaxial compression. This complex mechanical state has motivated the design of a biaxial loading system intended for use in vitro to stimulated bovine chondrocytes seeded in agarose constructs. This necessitated the redesign of the construct from cylindrical morphology to accommodate shear loading. The experimental approach was complemented with the development of computational models, which permitted prediction of both cell distortion under biaxial loading regimens and nutrient diffusion within the 3D constructs. An initial study established the profile of proteoglycan and collagen synthesis in free swelling cultures up to day 12. The introduction of dynamic compression (15% strain, 1 Hz for 48 h) enhanced proteoglycan synthesis significantly. In addition, when dynamic shear (10%, 1 Hz) was superimposed on dynamic compression, total collagen synthesis was also up-regulated, within 3 days of culture, without compromising proteoglycan synthesis. Histological analysis revealed marked collagen deposition around individual chondrocytes. However, a significant proportion (50%) of collagen was released into the culture medium, suggesting that it was not fully processed. The overall biosynthetic activity was enhanced more when the biaxial stimulation was applied in a continuous mode as opposed to intermittent loading. The present work offers the potential for a more effective preconditioning of cell-seeded constructs with functional integrity intended for use to resolve defects in joint cartilage.

Skeletal development begins when mesenchymal stem cells migrate, condensate and differentiate into chondrocytes. The chondrocytes differentiate in one of two ways. Either the cells form the cartilaginous template for endochondral ossification or they form the articular cartilage which express proteoglycan 4. The underlying mechanisms for articular cartilage formation are poorly understood. The purpose of this study was to assess the effect of different fractions of synoviocyte-conditioned medium on chondrocyte differentiation. We show evidence that Synovial-like fluid contains a protein which promotes chondrocytes to express proteoglycan 4, thus promoting articular cartilage formation. The synovial-like fluid was fractionized by size exclusion chromatography and reversed phase chromatography and thus, with that method, this manuscript lays the foundations for further research to identify the putative factor. Because of this study, we are now closer in identifying the proteins that promote articular cartilage formation.

Articular cartilage is a hydrated soft tissue with a fibrous solid matrix characterized by high porosity and low permeability. It is the bearing material of diarthrodial joints, permitting motion and transmitting loads with extraordinarily low friction. This function may be disrupted pathologically by osteoarthritis, a disease where cartilage becomes weakened and eroded. Osteoarthritis creates pain during normal activities like walking or grasping, thus diminishing quality of life. The disease affects nine percent of Americans and is one of the leading causes of disability worldwide. There is presently no cure or prevention for osteoarthritis, only palliative treatments designed to help patients manage pain and regain mobility. New such treatments are developed in part by advancing the science of cartilage mechanics, structure and function, and this dissertation presents novel contributions toward this effort: Chapters 2, 3, and 4 enhance our knowledge of the structure-function relationships critical to our understanding of cartilage friction and load support. Whereas most prior theoretical and experimental studies have focused on the analysis of small cylindrical explants, or idealized joint geometries such as cylindrical or spherical articular layers, these chapters describe novel investigations performed on whole articular layers of the shoulder and knee joints. Insights from these investigations have a direct impact on our formulation of design objectives in cartilage tissue engineering, whose purpose is to grow constructs that reproduce the functional properties of native cartilage. The studies presented in this chapter are critical to ongoing tissue engineering studies in our laboratory, which has pioneered the development of anatomically sized cartilage constructs. Finally, Chapter 5 describes the development of a novel clinical treatment for thumb osteoarthritis that uses bent osteochondral allografts (living bone and cartilage from human donors) to replace the eroded thumb trapezial articular layer with a healthy and thick articular layer from another joint such as the knee. This highly promising treatment strategy overcomes the limitation of size mismatch between donor and recipient which had relegated osteochondral allograft surgery to a niche treatment. Like other fibrous tissues, cartilage exhibits tension-compression nonlinearity, meaning it can be 100 times stiffer in tension than in compression. Tension-compression nonlinearity allows compressive physiologic joint loads to be supported by tensile stress within the collagen fibers and elevated fluid pressure, effectively shielding the solid matrix from compressive load. According to theory, fluid load support derives directly from tension-compression nonlinearity. Fluid load support is also a dominant mechanism of cartilage lubrication. Because cartilage is 80 to 90% water, most of the contact traction on the porous cartilage surface takes the form of hydrostatic fluid pressure. Friction forces only occur upon solid-on-solid contact, so cartilage friction is nearly negligible, even for joint contact forces that may routinely exceed three or four times the body’s total weight. The dependence of friction on fluid load support is demonstrated by experiments that simultaneously measure interstitial fluid pressure and friction - a transient rise in friction occurs as pressure subsides and fluid drains from the tissue. These structure-function relationships have been identified over decades of research, mostly through small cartilage explant studies, which have supported hypothesized mechanisms under non-physiologic conditions. Therefore, in situ studies utilizing intact, naturally-congruent articular surfaces under physiologic loading conditions would significantly extend and validate these principles. For example, friction may rise nearly 100-fold after only 1 hour in cartilage explant experiments, yet there is no evidence that normal daily activities spanning 16 hours or more lead to cartilage damage. Can fluid load support sustain low friction under these relatively harsh conditions? To date, no study has examined this question, so Chapter 2 of this work addresses the hypothesis that the friction coefficient of diarthrodial joints can remain low over a full day of loading at physiologic speeds and load magnitudes. Another question that may be uniquely addressed by an in situ analysis is: What is the complete state of stress within naturally-congruent cartilage layers? A primary hypothesis for the initiation and progression of osteoarthritis is that the state of stress within articular cartilage may exceed a threshold beyond which the tissue is unable to repair itself. Since the complete stress tensor within a material is immeasurable, techniques such as finite element analysis must be used to examine the state of stress. Additionally, a theoretical framework such as mixture theory may be used to examine the stresses in the fluid and solid constituents of the tissue separately, making it possible to test theories of solid matrix damage. Chapter 3 of this work uses this strategy to examine the hypothesis that physiologic solid matrix stresses within anatomically-shaped, biphasic, tension-compression nonlinear cartilage layers are primarily tensile, despite the fact that the articular layers are loaded in compression. The proteoglycan content of articular cartilage gives the tissue an osmotic swelling pressure that is resisted by tensile stresses in the collagen fibrils, even in the absence of external loads. This charge effect may be additionally incorporated into a mixture theory finite element analysis to examine the role of osmotic swelling on the solid matrix stresses in a physiologic, in situ analysis. This capability has only been developed recently and is explored for the first time in Chapter 4. The final part of this work translates basic cartilage science into a clinical therapy for thumb joint osteoarthritis, a common site for this disease. The current gold-standard treatment for thumb joint osteoarthritis replaces the trapezium bone with a soft-tissue tendon autograft, relieving pain but significantly weakening hand strength. Living osteochondral allograft transplantation may provide a relatively straightforward treatment alternative, though this procedure has not been used for the thumb due to the inadequate availability of suitable allografts. The ideal allograft would have a relatively thick articular layer to provide sufficient compliance for promoting joint congruence with the mating metacarpal surface, and surface curvatures that match the saddle-shaped anatomy of the distal trapezial articular surface to reproduce the normal joint motions. A potential solution that would provide suitable trapezium osteochondral allografts for patients involves precisely machining and bending allografts from a lower extremity joint with thicker cartilage, such as the distal femoral surface of the knee, to match the shape and curvature of the trapezium. Such bent osteochondral allografts would provide all the desired benefits of the ideal arthroplasty. Chapter 5 outlines the development of this novel technology, including proof of concept and feasibility demonstrations, business strategy and market analysis.

A doença degenerativa da cartilagem articular, ou osteoartrose, pode ser causada por diversos fatores, dentre eles o envelhecimento. Com o aumento da longevidade no Brasil, a prevalência da osteoartrose vai aumentar com o aumento progressivo da idade média da população nas próximas décadas. O estresse do dia a dia, com o avanço da vida moderna, são acompanhados pela má alimentação, sendo essa caracterizada por uma subnutrição ou até uma nutrição excessiva. Utilizando ratos Wistar, como modelo experimental, o objetivo deste trabalho é verificar se a alimentação com baixo nível calórico ameniza ou acelera as alterações na estrutura da cartilagem articular das epífises distal do fêmur e proximal da tíbia, causadas pelo envelhecimento. Para tanto foram utilizadas 15 ratos, separados em três grupos, com 5 animais cada: C- animais de 06 meses alimentados com dieta normal (A); SR- animais de 18 meses alimentados dieta normal (A); RC- animais de 18 meses submetidos à restrição calórica de 31% alimentados com dieta B. A avaliação foi realizada pela microscopia de luz em cortes histológicos corados pela Hematoxilina-Eosina, Picrossírius e por imunohistoquímica com marcação do colágeno tipo II. Foram feitas medidas da espessura das zonas da cartilagem articular, contado o número de condrócitos por área, determinados os volumes dos núcleos dos condrócitos, a morfometria do colágeno, a identificação dos tipos de colágeno e a análise imunohistoquímica. Os resultados mostram que os animais do grupo RC ganharam massa corpórea de maneira significativamente mais lenta que os do SR a partir dos 8 meses. Os animais do SR apresentaram na tíbia, aumento da espessura total bicondilar provocado pelo aumento da zona profunda, aumento do número de condrócitos no côndilo lateral, manutenção do volume dos núcleos e da proporção volumétrica de colágeno, marcação mais heterogenea do colágeno tipo II no côndilo lateral, predominância desse tipo de fibras na zona média do côndilo medial. No fêmur os animais do SR mostraram diminuição da espessura total bicondilar provocada pela diminuição da zona profunda, sendo menor no côndilo medial, diminuição do número de condrócitos causado por diminuição da zona profunda, sendo menor no côndilo medial, diminuição do volume total dos núcleos dos condrócitos no côndilo medial, diminuição da proporção volumétrica do colágeno bicondilar, marcação heterogênea do colágeno tipo II, predomínio desse tipo de fibras nas zonas média e profunda bicondilar. Os animais do RC apresentaram na tíbia, manutenção da espessura, aumento do número de condrócitos bicondilar, manutenção do volume nuclear, aumento da densidade de colágeno, marcação menos heterogênea do colágeno tipo II no côndilo lateral, predominância desse tipo de fibras nas zonas média e profunda do côndilo lateral. No fêmur, os animais do RC mostraram aumento da espessura total pelo aumento da zona profunda no côndilo medial, aumento do número de condrócitos pelo aumento da zona profunda, sendo maior no côndilo medial, aumento do volume total dos núcleos dos condrócitos bicondilar, aumento da proporção volumétrica de fibras colágenas no côndilo medial, marcação heterogênea do colágeno tipo II e predomínio desse tipo de fibras nas três zonas da cartilagem. Dessa maneira, concluímos que os efeitos do envelhecimento e da restrição calórica são diferentes nas cartilagens articulares da tíbia e do fêmur. As alterações naturais do envelhecimento são amenizadas pela restrição calórica na cartilagem articular do fêmur, porém na cartilagem articular da tíbia, essas alterações são compatíveis com início de doença degnerativa sendo intensificadas pela restrição com sinias de avanço de osteoartrose. / A degenerative disease of articular cartilage, or osteoarthritis, can be caused by several factors, including aging. With increased longevity in Brazil, the prevalence of osteoarthritis will increase with the progressive increase of the average age of the population in the coming decades. The stress of everyday life, with the advance of modern life, are accompanied by poor diet, this being characterized by malnutrition or even excessive nutrition. Using Wistar rats as experimental model, the aim of this work is to verify if feeding with low calorie softens or accelerates changes in the structure of articular cartilage of the epiphysis of the distal femur and proximal tibia, caused by aging. For this purpose were used 15 rats divided into three groups with five animals each: C, 6 month animals fed with normal diet (A), SR-18 month animals fed with normal diet (A), RC-18 month animals submitted to calorie restriction of 31% fed with diet B. The evaluation was performed by light microscopy in histological sections stained with Hematoxylin-Eosin, Picrossirius and by immunohistochemistry with tagging of type II collagen. Measurements were made of thick zones of articular cartilage, counted the number of chondrocytes per area, the volumes of certain nuclei of chondrocytes, collagen morphometry, the identification of the types of collagen and immunohistochemical analysis. The results show that the RC group animals gained body mass significantly slower than the SR from 8 months. The SR animals showed increasing of total thickness bicondylar of the tibia caused by the increase in the deep zone, increasing of the number of chondrocytes in the lateral condyle, maintaining the volume of the cores and the volumetric proportion of collagen, the most heterogeneous labeling of type II collagen in the lateral condyle, predominance of such fibers in the middle zone of the medial condyle. In the femur the SR animals showed decreased of total bicondylar thickness caused by the decrease of deep zone, being lower in the medial condyle, decreasing the number of chondrocytes caused by decreased deep zone, being lower in the medial condyle, decreased total volume of nuclei chondrocytes in the medial condyle, decreased volumetric proportion of collagen bicondylar, marking heterogeneous collagen type II predominance of such fibers in the middle and deep zones bicondylar. The RC animals showed in the tibia, maintaining thickness, increasing the number of chondrocytes bicondylar, maintenance of nuclear volume, increased collagen density, less heterogeneous marking of type II collagen in the lateral condyle, the predominance of this type of fiber areas and average deep lateral condyle. In the femur, the animals RC showed increased of total thickness caused by increasing of the medial condyle deep zone, increasing the number of chondrocytes by increasing the deep zone, being greater in the medial condyle, increasing the total volume of the nuclei of chondrocytes bicondylar, increased volumetric proportion of collagen fibers in the medial condyle, heterogeneous marking of type II collagen and prevalence of this type of fiber in the three zones of cartilage. Thus, we conclude that the effects of aging and caloric restriction are different in the articular cartilage of the tibia and femur. The natural changes of aging are attenuated by caloric restriction in the articular cartilage of the femur, but in the articular cartilage of the tibia, these changes are consistent with early degenerative disease being intensified by restriction with signs of advanced osteoarthritis.

A incidência de osteoartose, ou degeneração da cartilagem articular aumenta na pós-menopausa, condição esta relacionada à deficiência de estrógeno. Por outro lado, tem sido demonstrado que a atividade física regular e moderada tem efeitos benéficos sobre o sistema esquelético, em qualquer condição, mas especialmente na menopausa. Utilizando ratos Wistar, como modelo experimental, os objetivos deste trabalho são: a) verificar se a ooforectomia produz alterações na estrutura da cartilagem articular da epífise proximal da tíbia e b) verificar se a realização de exercícios físicos aeróbicos tem efeito sobre estas alterações. Para a realização deste estudo foram utilizadas 24 ratas com 6 meses de idade, da linhagem Wistar, distribuídas em 3 grupos de oito ratas cada: GC-Ratas com seis meses de idade, não submetidas à ooforectomia nem atividade física; GOS-Ratas com seis meses de idade, que sofreram ooforectomia bilateral, mas que não fizeram atividade física e, GOT-Ratas com seis meses de idade que foram submetidas à ooforectomia bilateral mais atividade física (corrida em esteira) durante 3 meses. Todos os animais foram eutanasiados com 9 meses de idade. A avaliação foi realizada através de microscopia de luz em cortes histológicos corados pela Hematoxilina-Eosina e pelo Picrossírius e estudos à microscopia eletrônica de varredura. À microscopia de luz, foram feitas medidas da espessura das zonas da cartilagem articular, contado o número de condrócitos por área, determinados os volumes dos núcleos dos condrócitos e a densidade de volume das fibras colágenas e à microscopia eletrônica de varredura foi analisada a superfície da cartilagem nos três grupos. Os dados quantitativos foram comparados estatisticamente pelo ANOVA e teste de Tukey. Os resultados mostraram que houve um aumento de peso nos animais do GOS e GOT em relação ao GC. Quanto ao treinamento, os animais do GOT tiveram melhor rendimento nos TEMs. Quanto à espessura da cartilagem observamos um aumento da espessura da zona superficial no côndilo medial da tíbia no GOS em relação ao GC. O número de condrócitos por campo foi alterado apenas no GOS, que mostrou um aumento de 34% em relação ao GC, o que, ao que parece, se deveu ao aumento do número de condrócitos da camada profunda. O volume nuclear dos condrócitos da zona superficial foi menor no GOS que no GC. O mesmo ocorreu na zona média do côndilo medial. Quanto aos valores da cartilagem da epífise proximal como um todo, observamos que o volume nuclear dos condrocitos do GOS e também do GOT foram menores do que do GC. Para o colágeno observamos um aumento da densidade de volume do colágeno no GOS e no GOT em relação ao GC. A microscopia eletrônica revelou a presença de lesões degenerativas semelhantes tanto para o GOS, quanto para o GOT. Podemos concluir que a depressão dos níveis de estrógeno acarreta alterações importantes na cartilagem articular da epífise proximal da tíbia tanto no componente celular, quanto na matriz e que a realização de exercícios físicos, isoladamente, parece não influenciar essas alterações. / The incidence of osteoarthritis increase in women in the post-menopausal period. This condition is relationed with the estrogen deficiency. Otherwise, it\'s being demonstrated that regular or moderate physical activity have beneficial effects on the skeletal system, in any condition, but especially in the menopause. Using Wistar rats, as an experimental model, the aims of this work are: a) to check if the ovariectomy produces alterations in the articular cartilage structure of the tibial proximal epiphysis and b) To check if aerobic physical exercises have effects on this alterations. To achieve these goals it was used 24 Wistar rats of 6 months of age, distributed in 3 groups of eight rats each: Control group- Rats with six months of age without ovariectomy and physical activity; GOS- Rats with six months of age with ovariectomy bilateral and without physical activity and, GOT- Rats with six months of age with ovariectomy bilateral and physical activity during 3 months. Al rats were euthanasied with 9 months of age. The results were obtained by using light microscopy to observe the histological sections stained with Hematoxylin and Eosin, and Picrosirius stain and the Scanning Electron Microscope. For light microscopy, were measured the thickness of cartilage, counted the number of chondrocytes, determined the volumes nuclei of chondrocytes and the density of volume of the collagens fibers. For the Scanning Electron Microscope, was analyzed the cartilage surface in the three groups. The data were statistically compared for the ANOVA and Tukey´s test. The results showed that there was an increase in the weight in the animals of GOS and of GOT. In relation to training, the animals of GOT obtained the best income in the TEMs. In relation to the thickness of the articular cartilage it was observed a decrease in the superficial layer of the medial condyle in the GOS and GOT. The number of chondrocytes was altered only in the GOS, that obtained an increase of the 34% on GC in the deep layer. The nuclear volume of chondrocytes in the superficial layer was higher in the GC than the GOS .The same occurred in the medium layer when the two tibial condyles, lateral and medial were assessed separately, but when the values of the epiphysial cartilage was considered as a whole, we found that the volume nuclei of chondrocytes in the GC was higher than the GOS and the GOT. For the collagen, the values of density of volume of the collagen fibers was higher in the GOS and in the GOT than the GC. The Scanning Electron microscopy showed the presence of degenerative lesions in both GOS and GOT. We can conclude that the depression levels of estrogen produced important changes in the tibial cartilage and that the physical activity doesn\'t have effects on these changes.