Orientador: Silvana Martinez Baraldi Artoni / Banca: Isabel Cristina Boleli / Banca: Cristiane Soares da Silva Araújo / Resumo: Este estudo teve como objetivo avaliar os efeitos do sulfato de glucosamina, de condroitina e da associação de ambos no sistema locomotor e no desempenho de frangos de corte de 1 a 42 dias de idade. Foram utilizados 900 pintos de corte machos de um dia, da linhagem Cobb-500®, distribuídos em delineamento inteiramente casualizado, com seis tratamentos e cinco repetições de 30 aves cada. Os tratamentos utilizados foram: T1 - controle negativo, T2 - 25% sulfato de glucosamina e 75% condroitina, T3 - 50% sulfato de glucosamina e 50% condroitina, T4 - 75% sulfato de glucosamina e 25% condroitina, T5 - 100% sulfato de 100% glucosamina e T6 - 100% sulfato de condroitina. As rações experimentais para fase de criação foram formuladas à base de milho e farelo de soja, seguindo-se as recomendações de Rostagno et al. (2011), para as respectivas fases de criação: inicial (1-21 dias) e crescimento (22-42 dias). Foram avaliados os parâmetros de desempenho zootécnico, rendimento de carcaça e partes, densitometria óssea dos tibiotarsos, histoquímica das cartilagens das tíbias localizadas nas epífises proximais e morfometria hepática, analisadas aos 21 e 42 dias. As análises estatísticas dos dados foram realizadas pela análise de variância com o auxílio do procedimento General Linear Model do SAS (2002) e em caso de significância estatística, as médias foram comparadas pelo teste de Tukey, ao nível de 5% de probabilidade. Os diferentes níveis de sulfato de glucosamina e de condroitina não determinaram efeito significativo sobre os parâmetros de desempenho das aves, rendimento de carcaça e partes e morfometria hepática. Entretanto, foi observado que o tratamento com o nível de 100% de sulfato de glucosamina proporcionou maior número de condrócitos nas cartilagens articulares de aves com 21 e 42 dias de idade. Para os parâmetros de densitometria, o nível de 100% de ... / Abstract: This study aimed to evaluate the effects of glucosamine sulfate, chondroitin and combination of both in the locomotor system and the performance of broiler chickens during 1-42 days of age. It was 900 Cobb - 500 ® line chicks, in a completely randomized design with six treatments and five replicates of 30 birds each design. The treatments were: T1 - negative control, T2 - 25 % glucosamine sulfate and 75% chondroitin, T3 - 50 % glucosamine sulfate and 50% chondroitin, T4 - 75 % glucosamine sulfate and 25 % chondroitin, T5 - 100 % of glucosamine sulfate and T6 - 100% chondroitin sulfate. The experimental diets for creation phase were formulated based on corn and soybean meal, following the recommendations of Rostagno et al. (2011), for the respective phases of creation: initial (1-21 days ) and grower ( 22-42 days). Parameters of growth performance, carcass and parts, bone densitometry of tibiotarsos, staining of cartilage located in the proximal tibia epiphysis and liver morphology, analyzed at 21 and 42 days were evaluated. Statistical analyzes of the data were performed by analysis of variance method with the aid of the General Linear Model of SAS  (2002 ) and in case of statistical significance, the means were compared by Tukey test at 5 % probability. The different levels of glucosamine and chondroitin sulfate did not cause significant effect on the parameters of broiler performance, carcass yield and liver parts and morphometry. However, it was observed that the treatment with the level of 100% glucosamine sulfate yielded a greater number of chondrocytes in articular cartilage in birds 21 and 42 days of age. For the parameters of densitometry, the level of 100 % of glucosamine sulfate provided greater bone mineral density at 21 days, but at 42 days of age, the level of 100 % of chondroitin sulfate provided higher income densitometry, outperforming the locomotors system / Mestre

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.

A subnutrição tem sido ao longo dos anos um dos maiores problemas de saúde pública do mundo, que acomete principalmente crianças de países em desenvolvimento. Estima-se que em Moçambique na África Oriental, cerca de 40% das crianças são acometidas pela desnutrição crônica que resulta dentre outras afecções em baixa estatura e diminuição da capacidade física. Este estudo visa avaliar a estrutura óssea de ratos wistar submetidos à dieta básica da população de Moçambique (DM) reproduzida em laboratório. Para isso a DM foi oferecida aos animais em fase pré e pós-natal durante 42 dias. Para alguns desses animais foi fornecida a DM acrescida de 20% de caseína (DMC) subsequentemente os grupos subnutrido de Moçambique (SM) e nutrido de Moçambique (NM). Foi formado também um grupo denominado renutrido (RM) composto por animais que receberão a DMC a partir do 22&ordm; dia e até completarem 42 dias de vida. Após o desmame foram mensurados os parâmetros metabólicos da ingestão alimentar e hídrica, excreção de urina e fezes. A tíbia e o úmero do lado direito foram macerados quimicamente a fim de verificar as medidas ósseas. Esses ossos do lado esquerdo foram processados com técnicas rotineiras histológicas e corados de forma a evidenciar as estruturas cartilagineas. Os dados obtidos foram submetidos a análise de variância paramétrica seguida por comparações múltiplas pelo método de Tukey, com nível de significância p&lt;0,05. Os resultados apontam que a Subnutrição é capaz de modificar a espessura da CA e da LE e os componentes de sua MEC. Embora a inserção de caseína na dieta dos animais SM, tenha reestabelecido parâmetros da osteometria, na LE e na CA a melhora apresentada, não foi o suficiente para atingir seus parâmetros normais apresentados no grupo NM / Undernutrition has been over the years one of the major public health problems worldwide, which affects mainly children in developing countries. It is estimated that in Mozambique in East Africa, about 40% of children are affected by chronic undernutrition resulting from other conditions in stature and diminished physical capacity. This study aims to evaluate the bone structure of Wistar rats submitted to the staple diet of the population of Mozambique (DM) reproduced in the laboratory. To this DM was given to the animals in pre and postnatal 42 days. For some of these animals was provided to DM plus 20% casein (DMC) subsequently the undernourished groups of Mozambique (SM) and nursed Mozambique (NM). Also it has formed a group called renutrido (RM) consists of animals that receive the DMC from the 22nd day until completing 42 days of life. After weaning metabolic parameters were measured food and water intake, urine output and feces. The tibia and the humerus on the right side were chemically macerated in order to verify the bone measurements. These bones on the left side were processed with routine histological techniques, and stained to show the form cartilage structures. The data were subjected to parametric analysis of variance followed by multiple comparisons by the Tukey method, with significance level of p <0.05. The results show that the Malnutrition is able to modify the thickness of the CA and LE and the components of its MEC. Although the inclusion of casein in the diet of animals SM, has reestablished osteometria parameters in LE and CA improvement presented, was not enough to achieve their normal parameters presented in NM group

Traumatic injuries lead to articular cartilage lesion formation and result in the development of osteoarthritis. Recent research suggests that the early stage of mechanical injuries involve cell death (apoptosis and necrosis) and inflammation. In this thesis, we focus on building mathematical models to investigate the biological mechanism involving chondrocyte death and inflammatory responses in the process of cartilage degeneration. Chapter 1 describes the structure of articular cartilage, the process of carti- lage degeneration, and reviews of existing mathematical models. Chapter 2 presents a delay-diffusion-reaction model of cartilage lesion formation under cyclic loading. Computational methods were used to simulate the impact of varying loading stresses and erythropoietin levels. The model is parameterized with experimental results, and is therefore clinically relevant. Due to numerical limitations using delay differential equations, a new model is presented using tools for population dynamics. Chapter 3 presents an age and space-structured model of articular cartilage lesion formation un- der a single blunt impact. Age structure is introduced to represent the time delay in cytokine synthesis and cell transition. Numerical simulations produce similar tempo- ral and spatial patterns to our experimental data. In chapter 4, we extend our model under the cyclic loading setting. Chapter 5 builds a spatio-temporal model adapted from the former models, and investigates the distribution of model parameters using experimental data and statistical methods. Chapter 6 concludes.

Research in the biomedical sciences is incredibly diverse and often involves the interaction of specialists in a variety of fields. In particular, quantitative, mathematical, and computational methods are increasingly playing significant roles in studying problems arising in biomedical science. This is particularly exciting for mathematical modeling as the complexity of biological systems poses new challenges to modelers and leads to interesting mathematical problems. On the other hand mathematical modeling can provide considerable insight to laboratory and clinical researchers. In this thesis we develop mathematical representations for three biological processes that are of current interest in biomedical science. A deeper understanding of these processes is desirable not only from the standpoint of basic science, but also because of the connections these processes have with certain diseases. The processes we consider are collective cell motility, bone remodeling, and injury response in articular cartilage. Our goals are to develop mathematical representations of these processes that can provide a conceptual framework for understanding the processes at a fundamental level, that make rigorous the intuition biological researchers have developed about these processes, and that help to translate theoretical and experimental work into information that can be used in clinical settings where the primary concern is in treating diseases associated with the process.

While mechanical loading is essential for articular cartilage homeostasis, it also plays a central role in the etiology of osteoarthritis. The mechanotransduction events underlying these dual effects, however, remain unclear. Previously, we have shown that lethal amounts of reactive oxygen species (ROS) were liberated from mitochondrial complex 1 in response to a mechanical insult. The sensitivity of this response to an actin polymerase inhibitor, cytochalasin B, indicated a link between ROS release and cytoskeletal distortion caused by excessive compressive strain. It did not, however, rule out the possibility that ROS may also mediate the beneficial effects of normal stresses that induce lower tissue strains required for proper homeostasis. If this possibility is true, one would expect the amount of ROS released in loaded cartilage to be positively correlated with the level of strain, and ROS should only reach lethal levels under super-physiological deformations. To test this hypothesis, full cartilage tissue strains were measured in cartilage explants subjected to static normal stresses of 0, 0.1, 0.25, 0.5, and1.0 MPa. After compression, the percentage of ROS-producing cells was measured using the oxidation-sensitive fluorescent probe, dihydroethidium, and confocal microscopy. In support of our theory, the percentage of fluorescing cells increased linearly with increasing strains (0-75%, r2 = 0.8, p < 0.05). Additionally, hydrostatic stress, which causes minimal tissue strain, induced minimal ROS release. In terms of cell viability, cartilage explants compressed with strains >40% experienced substantial cell death, while explants with strains

Articular cartilage is a complex soft tissue covering the end of moving bones in joints which provide pressure load distribution over the joint surface and smooth lubrication with little friction for establishing movement. Articular cartilage has an intrinsically limited capacity for self-repair when injured due to the lack of nerve and blood supply. Considered that injured cartilage is left untreated, it is likely to undergo progressive cartilage degeneration without pain which may lead to posttraumatic osteoarthritis. Therefore functional and physiologic restoration of injured cartilage back to a normal condition has long been in demand, yet current available repairing methods in clinics have met with limited success. Mechanically applied loads to articular cartilage is necessary for chondrocytes, cartilage cells, since they are responsible for cartilage matrix turnover by synthesizing extracellular matrix (ECM) molecules in response to bio- chemical and mechanical changes in ECM. Ultrasound has emerged as an anabolic stimulator over the past few decades and a number of studies have proven that ultrasound therapy is beneficial for cartilage repair by synthesizing cartilage ECM components such as type II collagen and proteoglycan. Ultrasound therapy has also proven its potential for the attenuation of progressive cartilage degradation and induction of chondrogenic differentiation of mesenchymal stem cells. The use of ultrasound as an anabolic stimulator would be valuable with respect to cartilage repair since ultrasound as a form of mechanical energy can be non-invasively transferred into a human body. However, understanding the underlying mechanisms has been slow and the mechanisms have been roughly classified into thermal and non-thermal effects. Biologically detailed underlying mechanisms have not been sufficiently studied. That might be the reason why the application of ultrasound as a therapeutic tool has been limitedly available in clinics. In this study, mechanism involved biophysical effects of low intensity ultrasound has been studied for cartilage regeneration. First of all, the effect of ultrasound therapy as a mechanical stimulator on chondrogenic progenitor cell homing toward injured sites in cartilage was investigated with underlying biologic mechanisms. And the feasibility of ultrasound therapy for reactive oxygen species production mediated cartilage energy modulation was evaluated. There have been extensive preclinical studies about the effects of microbubble mediated ultrasound therapy on the targeted drugs or gene delivery into tissues of interest. Mechanical shock waves are released during ultrasound mediated microbubble destruction and the waves facilitate drug delivery into target tissues through transient blood vessel disruption. However, the clinical use of this technique has been limited through vascular system. In this study, the effects of microbubble mediated low intensity ultrasound therapy on directly delivered mechanical shock waves and controlled drug release were investigated. In conclusion, low intensity ultrasound therapy accelerates the homing of chondrogeic progenitor cells toward injured sites in cartilage via triggering mechanotransductive cell signaling pathways. This may result in speed up the return to normal cellularity and cartilage integrity by accelerating cartilage matrix repair. Low intensity ultrasound therapy was investigated as an energy modulator for chondrocytes via reactive oxygen species production in articular cartilage; however, little effects of ultrasound therapy driven cartilage energy modulation were found. The strong relationship between microbubbles mediated low intensity ultrasound therapy and the controlled release of drugs and mechanical shock waves was found. This strongly suggests that low intensity ultrasound therapy can play a role as a non-invasive controller for the release of drugs and lethal shock waves upon request.

[Truncated abstract] Autologous Chondrocyte Implantation (ACI) has shown early clinical success as a repair procedure to address focal articular cartilage defects in the knee, and involves isolating and culturing a patient's own chondrocytes in vitro and re-implantation of those cells into the cartilage defect. Over time, repair tissue can develop and remodel into hyaline-like cartilage. A progressive partial weight bearing (PWB) program becomes the critical factor in applying protection and progressive stimulation of the implanted cells, to promote best chondrocyte differentiation and development, without overloading the graft. The aim of this thesis was to investigate whether patients could replicate this theoretical load bearing model to possibly render the best quality tissue development. In addition, this proposed external load progression is only a means to loading the articular surface. Several factors, including those that may result from pathology, have the potential to influence gait patterns, and therefore, articular loading. The association between increasing external loads (ground reaction forces - GRF) and knee joint kinetics during partial and full weight bearing gait was, therefore, investigated in the ACI patient group, as was the contribution of other gait variables to these knee joint kinetics which may be modified by the clinician. Finally, current weight bearing (WB) protocols have been based on early ACI surgical techniques. With advancement in the surgical procedure and ongoing clinical experience, we employed a randomised controlled clinical trial to assess the effectiveness of an 'accelerated' load bearing program, compared with the traditionally 'conservative' post-operative protocol. ... Although similar spatio-temporal, knee kinematic and external loading parameters were observed between the traditional and accelerated rehabilitation groups, the accelerated group was 'more comparable' to the controls in their external knee adduction and flexion moments, where the traditional group had lower knee moments. Knee moments greatly affect knee articular loading, and large adduction moments have been related to poor clinical outcomes after surgery. Therefore, the return of normal levels may be ideal for graft stimulation, however, may overload the immature chondrocytes. Acceleration of the intensive rehabilitation program will enable the patient to return to normal activities earlier, whilst reducing time and expenses associated with the rehabilitative process, and may enhance long-term tissue development. However, continued follow-up is required to determine if there are any detrimental effects that may emerge as a result of the accelerated load bearing program, and assess the recovery of normal gait patterns and whether longer term graft outcomes are affected by the recovery time course of normal gait function, and/or abnormal loading mechanics in gait. Furthermore, analysis at all levels of PWB is needed to identify a more complete set of variables attributing to the magnitude of external knee joint kinetics and, therefore, knee articular loading, while the influence muscle activation patterns may have on articular loading needs to be investigated. This becomes critical when you consider loads experienced by the articular surface throughout the early post-operative period following ACI may be important to short- and long-term graft development.

Thesis (M.S.) -- Mississippi State University. Department of Agricultural and Biological Engineering. / Title from title screen. Includes bibliographical references.

This thesis demonstrated the effects of exogenous stimuli on engineered articular cartilage constructs and elucidated mechanisms underlying the responses to these agents. In particular, a series of studies detailed the effects of chondroitinase-ABC (C-ABC), hyaluronic acid (HA), and TGF-β1 on the biochemical and biomechanical properties of self-assembled articular cartilage. Work with C-ABC showed that this catabolic agent can be employed to improve the tensile properties of constructs. When constructs were cultured for 6 weeks, treating with C-ABC at 2 weeks enhanced the tensile stiffness. Furthermore, treating at 2 and 4 weeks synergistically increased tensile properties and allowed compressive stiffness to recover to control levels. Another study showed that combining C-ABC and TGF-β1 synergistically enhanced the biochemical and biomechanical properties of neotissue. Microarray analysis demonstrated that TGF-β1 increased MAPK signaling in self-assembled neocartilage whereas C-ABC had minimal effects on gene expression. SEM analysis showed that C-ABC increased collagen fibril diameter and fibril density, indicating that C-ABC potentially acts via a biophysical mechanism. Constructs treated with C-ABC and TGF-β1 also showed stability and maturation in vivo , exhibiting a tensile stiffness of 3.15±0.47 MPa compared to a pre-implantation stiffness of 1.95±0.62 MPa. To assess the response to HA application, studies were conducted to optimize HA administration and examine its effects in conjunction with TGF-β1. Applying HA increased the compressive stiffness 1-fold and increased GAG content by 35%, with these improvements depending on HA molecular weight, application commencement time, and concentration. Microarray and PCR analyses showed that HA also influenced genetic signaling, up-regulating multiple genes associated with the TGF-β1 pathway. In addition to genetic effects, the enhanced GAG retention due to HA treatment could increase the fixed charge density of the matrix and thereby increase resistance to compressive loading. Additive effects were observed when HA was applied in conjunction with TGF-β1, with the combined treatment increasing compressive stiffness and GAG content by 150% and 65%, respectively. In general, results demonstrated mechanisms underlying C-ABC, HA, and TGF-β1 treatments and showed how these agents can be applied to improve cartilage regeneration efforts.