Orientador: Ibsen Bellini Coimbra / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciencias Medicas / Made available in DSpace on 2018-08-07T22:55:15Z (GMT). No. of bitstreams: 1 Andrade_AndreLuisLugnanide_M.pdf: 2450375 bytes, checksum: f33edb219ba25e56f663f1256a99bbeb (MD5) Previous issue date: 2006 / Resumo: Introdução: Os condrócitos da cartilagem articular vivem em um ambiente com baixa concentração de oxigênio. Nestas condições, a proteína do fator induzido por hipóxia (HIF-1a) mantém-se estável e ativa genes que são fundamentais na homeostase do oxigênio. A expressão do HIF-1a aumenta, em joelhos com osteoartrite (OA), principalmente nas áreas mais afetadas pela degeneração. Os condrócitos são capazes de produzir mediadores inflamatórios, como a interleucina-1 (IL-1) e o fator de necrose tumoral a (TNF-a), que estimulam a produção de prostaglandinas, metaloproteinases e óxido nítrico e relacionam-se com o início e com a progressão da osteoartrite. Os antiinflamatórios são drogas freqüentemente utilizadas no tratamento sintomático da OA. Material e Método: condrócitos humanos de joelhos osteoartríticos cultivados em suspensão e em condições normais de oxigênio foram divididos em quatro grupos: 1) controle, 2) estimulados com IL-1 ou TNF-a, 3) estimulados com meloxicam ou parecoxibe e 4) estimulados com meloxicam ou parecoxibe associados a IL-1 ou TNF-a. Os grupos foram submetidos à extração de RNA (ácido ribonucléico) e de proteína nuclear. O RNA foi convertido em cDNA, sendo então realizada a reação de PCR em tempo real para verificar a expressão do HIF-1a. As proteínas nucleares foram extraídas, quantificadas e analisadas pela técnica de Western Blotting. Resultados: Foi detectada a expressão de HIF-1a e cDNA de HIF-1a em todos os grupos de condrócitos cultivados em suspensão em tensões normais de oxigênio, não havendo diferenças significativas entre os grupos. Discussão: a meia-vida do HIF-1a é extremamente curta em normóxia e marcadamente prolongada em hipóxia, por isso muitos pesquisadores acreditam não ser possível a detecção da proteína do HIF-1a em condrócitos cultivados em condições normais de oxigênio. Neste estudo foi possível constatar a expressão do HIF-1a em normóxia, possivelmente devido ao modelo de cultura utilizado. O estímulo com IL-1, TNF-a e inibidores da COX-2 não alterou a expressão de HIF-1a. Condrócitos oriundos de articulações osteoartríticas avançadas poderiam apresentar resistência à ação das citocinas / Abstract: Introduction: The chondrocytes of joint surface live in low concentration of oxygen environment. In this condition, the hypoxia inducible factor 1 a (HIF-1a) becomes stable and regulates the expression of genes that are important for oxygen homeostasis. The expression of HIF-1a mRNA is augmented in chondrocytes from osteoarthritic knees, especially in more degenerated areas. Chondrocytes are capable of producing inflammatory mediators, such as interleukin 1 (IL-1) and tumoral necrosis factor a (TNF-a), that stimulate the production of prostaglandin, metalloproteinases and nitric oxide, correlated with the onset and progression of osteoarthritis. Antiinflammatory drugs are frequently used in the treatment of symptoms of osteoarthritis. Material e Methods: human chondrocytes from osteoarthritic knees were cultivated in suspension and in normal tension of oxygen. The cells were divided in 4 groups: control, stimulated with IL-1 or TNF-a, stimulated with meloxicam or parecoxib and the last one stimulated with meloxicam or parecoxib and IL-1 or TNF-a. Nuclear protein and RNA were extracted from these cells. cDNA was synthesized from RNA and real time PCR was performed with this product in order to determine HIF-1a expression. Nuclear protein was analyzed using the Western-Blotting method. Results: HIF-1a and HIF-1a mRNA was detectable in all cell groups, and there was not a statistic significant difference between them. Discussion: As half live of HIF-1a is extremely short when in normoxic and greater in hypoxic conditions, many researchers believe it is not possible to detect this protein in chondrocytes cultivated in normoxic environment. Our results presented expression of HIF-1a in normal oxygen tensions, probably due to the fact that chondrocytes were cultivated in suspension. As chondrocytes were obtained from advanced osteoarthritic knees and in such conditions the cells can be more resistant to the action of cytokines, this could explain why IL-1, TNF-a and antiinflamatory did not result in modification of HIF-1a / Mestrado / Clinica Medica / Mestre em Clinica Medica

Orientador: Célia Marisa Rizzatti Barbosa / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Odontologia de Piracicaba / Made available in DSpace on 2018-08-18T16:04:07Z (GMT). No. of bitstreams: 1 FariasNeto_Arcelino_D.pdf: 1895628 bytes, checksum: 3d9e789620307fdbeaffe939bf75437e (MD5) Previous issue date: 2011 / Resumo: A cartilagem articular do côndilo mandibular é responsável pelo crescimento ósseo endocondral durante o desenvolvimento mandibular. Ela depende do funcionamento adequado da articulação temporomandibular (ATM) para sua diferenciação e maturação. Trabalhos demonstram que a manipulação funcional da mandíbula foi capaz de alterar a dinâmica fisiológica dessa cartilagem. Nesse sentido, a protrusão diminuiria a ação de cargas sobre o côndilo mandibular, estimulando o crescimento endocondral, e de forma inversa, a retrusão aumentaria a pressão sobre a cartilagem, inibindo o crescimento. Essas técnicas têm sido utilizadas com relativo sucesso na ortopedia facial com o intuito de corrigir discrepâncias maxilo-mandibulares. Entretanto, alguns quadros patológicos presentes nas ATMs podem alterar o seu desenvolvimento normal. Um dos fatores etiológicos que pode ser associado à presença de alterações no côndilo mandibular é a oclusão dental. A hipótese formulada é de que a presença de instabilidade ortopédica causada por um fator oclusal durante a fase de desenvolvimento pode levar à deficiência do crescimento mandibular e alterações intra-articulares. Assim, este trabalho teve por objetivo avaliar, em modelo animal, alterações da oclusão dental sobre o crescimento mandibular e tecidos intra-articulares. O estudo foi randomizado e cego. Foram utilizadas 40 ratas Wistar com 5 semanas de idade divididas aleatoriamente em 4 grupos com o mesmo número de animais: controle, com interferência oclusal, com ausência dos molares inferiores unilateral e com ausência dos molares inferiores bilateral. Os animais foram acompanhados por 8 semanas, período que correspondeu a sua fase de maturação óssea. Após esse período, os animais foram sacrificados e realizou-se tomografia computadorizada de feixe cônico (Cone beam) de suas cabeças para construção de protótipos de biomodelos, sobre os quais foram mensurados o comprimento da mandíbula, a altura do ramo mandibular e distância intercondilar. Em seguida, as articulações temporomandibulares foram cuidadosamente preparadas para análise imunohistoquímica dos níveis de colágeno tipo II, Fator de Crescimento Endotelial Vascular, e Interleucina 1? na cartilagem condilar. Os dados foram submetidos a análise estatística através do Software SPSS versão 17.0. As médias entre os grupos foram comparadas através do One-way Anova, enquanto as diferenças entre os lados da mandíbula foram avaliadas através do teste t de Student (?=0.05). A partir da análise dos resultados, observou-se que alterações oclusais podem afetar o desenvolvimento do osso mandibular, bem como alterar a expressão de Colágeno tipo II, Fator de Crescimento Endotelial Vascular e Interleucina 1? na cartilagem condilar. Diante do exposto, conclui-se que a oclusão dentária é capaz de interferir na dinâmica dos tecidos intra-articulares, sendo um fator importante durante o desenvolvimento craniofacial / Abstract: The condylar cartilage regulates the endochondral ossification during mandibular development. Mechanical stimulus in the temporomandibular joint (TMJ) plays an important role in cell proliferation and differentiation of mandibular condyle. Studies have shown that functional mandibular displacement can affect TMJ cartilage dynamics. Mandibular advancement induces profound metabolic changes in the condyle and enhances growth. In contrast, mandibular retraction reduces growth. The overall picture emerging from the data is that unloading of the condyle increases growth, while loading reduces it. Therefore, dental occlusion could be one of the factors associated with the alteration of the TMJ growth. The hypothesis is that orthopedic instability caused by occlusal factors present during TMJ development can affect mandibular growth and intra-articular tissue. Thus, the purpose of this study was to evaluate the influence of dental occlusion on mandibular growth and intra-articular tissue in Wistar rats. The study was randomized and blinded. Forty 5 weeks old female Wistar rats composed the sample. The animals were randomly allocated to four groups with the same number of rats: (1) control, (2) occlusal appliance for functional posterior displacement of the mandible, (3) unilateral mandibular tooth extraction, (4) bilateral mandibular tooth extraction. The rats were sacrificed after 8 weeks, when they had achieved skeletal maturity. Immediately after death, the heads were fixed in 10% paraformaldehyde, and cone beam CT scan images were taken using the Classic I-CAT (Imaging Sciences International, Hatfield, PA, USA). The 3-dimensional images of rats' skulls were exported in multifile Digital Imaging and Communications in Medicine (DICOM) format, and acrylic rapid-prototyped templates of the mandibles were constructed for measurement of mandibular growth. Immunostaining was used for the detection of type II collagen, vascular endothelial growth factor (VEGF) and interleukin-1?. The data were processed with SPSS software (V 17.0 for Windows, SPSS Inc, Chicago, IL, USA). Differences among the groups were analyzed by one-way ANOVA (Tukey test as post-hoc test), while differences between sides were analyzed by non-paired Student's t test. Shapiro-Wilk and Levene tests were used to observe normality and variance homogeneity, respectively. Confidence level was set at 5%. The results of this study showed that dental occlusion is an important factor for the integrity of intra-articular tissues and to the healthy craniofacial development, emphasizing the importance of early treatment to normalize occlusion and create appropriate conditions for normal craniofacial development / Doutorado / Protese Dental / Doutor em Clínica Odontológica

The immune system is composed of innate and adaptive mechanisms. Innate immune responses are significantly modulated by immunomodulatory factors that act through the induction of specific patterns of cytokine production in responding cells. Human leukocytes have been shown to respond to substance(s) present in acid extracts of commercial shark cartilage (SC). Shark cartilage is a food supplement taken by consumers as a prophylaxis and for the treatment of conditions ranging from arthritis to cancer. No reliable scientific evidence in the literature supports the alleged usefulness of shark cartilage supplements, but their use remains popular. Cartilage extracts exhibit immunomodulatory properties by inducing various inflammatory, Th1-type cytokines and potent chemokines in human peripheral blood leukocytes (HPBL) in vitro. The objectives of the study were to (1) to determine the nature of the active component(s), (2) to define the scope of cellular response to SC extract, and (3) to elucidate the molecular mechanisms underlying bioactivity. Results showed that there are at least two cytokine-inducing components which are acid stable. One anionic component has been identified as a small (14-21 kDa) glycoprotein with at least 40% carbohydrate content. Shark cartilage stimulated HPBL to produce cytokines resembling an inflammatory, Th1 polarized response. Leukocyte-specific responses consist of both initial cytokine responses to SC directly (i.e., TNF-a) and secondary responses such as the IFN-γ response by lymphocytes following initial SC stimulation. Response of RAW cells, a murine macrophage cell line, indicated that TNF-α could be induced in macrophages of another mammalian species in the absence of other cell types. The results suggest that the human monocyte/macrophage is most likely to be the initial responding cell to SC stimulation. Stimulation of cells appears to engage at least one ligand-receptor interaction with TLR 4, although the role of TLR 2 cannot be ruled out. Initial activation is likely followed by the activation of the JNK and p38 MAPK signal transduction pathways resulting in activation, release, and translocation of transcription factor nuclear factor κB (Nf-kB). This dissertation research study represents the first in-depth study into characterizing the bioactive component(s) of commercial shark cartilage responsible for its immunomodulating properties and defining cellular responses at the molecular level.

In Canada, one in five people suffers from arthritis, of which the most common type is osteoarthritis (OA). OA is a group of joint diseases that cause pain and loss of range of motion and for which there is currently no cure. OA can be caused by numerous factors such as aging, genetics, environmental elements, and abnormal joint biomechanics (e.g., injury, obesity). These diseases are degenerative and lead to the progressive breakdown of joint cartilage, as well as changes in the underlying bone and other tissues of the joint over a period of years to decades. Articular cartilage incorporates a single type of resident cells, termed chondrocyte cells. These cells are entrapped within a dense extracellular matrix that limits their ability to proliferate and migrate to a site of injury, while the absence of blood vessels in the cartilage, amongst other factors, hinders the ability of progenitor cells to reach the site of injury, contributing to a limited capacity for intrinsic regeneration of the damaged tissue following an injury. As such, efforts to develop tissue engineering strategies that combine a biomaterial with bioactive signals to induce cells with the chondrogenic potential to regenerate tissue have been pursued actively. In this thesis, we investigate the potential of one such cartilage tissue engineering approach, whereby chondrocytes are encapsulated with alginate hydrogels incorporating inorganic polyphosphate (polyP), a promising chondrogenic signal. The driving hypothesis of the work was that polyanionic polyP would crosslink within the alginate hydrogel meshwork by ionic bonds with the multivalent cations used to form the hydrogel. Initial efforts focussed on optimizing the sterile chondrocyte encapsulation protocol for alginate beads, chondrocyte culture conditions to reduce proliferation – a response that is associated with dedifferentiation and a pathological state – and protocol for the incorporation of polyP in alginate bead when using calcium as a cationic crosslinker. We observed that polyP release from the calcium-alginate bead exhibited an important burst release to nearly 80% of the initial polyP loading within 24 hours of incubation in the culture medium. Increasing the alginate concentration led to approximately a 2.5-fold increase in polyP retention following the burst release. Subsequent incubation showed a more controlled release for at least 1 week. Efforts to reduce hydrogel swelling and increase its stability by substituting Ca2+ by Sr2+ as a crosslinker did not reduce the release rate during the burst release phase, nor did it increase the polyP retention following this initial stage. Other divalent cations including Mg2+ and Co2+, and pre-loading the polyP-alginate solution with a small concentration of Ca2+ did not impact the release profile either. Chondrocytes encapsulated in calcium- and strontium-alginate beads showed decreased DNA content and increased sulfated glycosaminoglycan accumulation at 1 week when polyP was incorporated in the beads compared to controls without polyP; however, this effect was lost at longer time points. These results suggest that this new material may find applications as a vehicle for the short-term delivery of polyP in joints and other tissues. Further efforts to improve the polyP release profile from alginate beads lead to promising results with the use of polyethylenimine (PEI) as a cationic tethering molecule between polyP and alginate. This thesis aims to generate novel biomaterials that can be used to stimulate cartilage tissue regeneration and to eventually develop a treatment strategy for OA. The work presented here will serve as a basis for continued efforts to ensure the prolonged retention of exogenous polyP into the joint.

No description available.

This thesis deals with the study of the effect of the constituents of the model synovial fluid on the frictional properties of articular cartilage. The influence of constituents, concentration, speed and load is observed. Experiments were performed on a commercial tribometer at configuration pin-on-plate. Several types of lubricants containing synovial fluid constituents have been selected for the experiments. Lubricants were prepared at two concentrations, the concentration of healthy individuals and at a concentration typical of for osteoarthritic patients. Speeds 5 and 10 mm/s and 5 and 10 N loads were used for all experiments. It is shown that when using only lubricant containing proteins, no difference in the coefficient of friction is observed and the effect of concentration is also not observed. The addition of hyaluronic acid has a synergistic effect with -globulin, however in the case of lubricants containing albumin, the effect is opposite. After the addition of phospholipids, no significant effect on friction is observed in -globulin containing lubricants. No significant effect of the composition and concentration of the lubricants is observed with the load change.

Articular cartilage is a highly organized, anisotropic tissue lining the ends of bones within synovial joints. Composed primarily of water, collagens, proteoglycans and chondrocytes which synergistically give rise to the tissue's mechanical and tribological properties. Fluid pressurization and resistance to fluid flow within the porous extracellular matrix of cartilage, coupled with the low hydraulic permeability of the tissue endow the tissue with a viscoelastic response to loading and aid to reduce the coefficient of friction between articulating surfaces, with the pressurized fluid supporting 95% of applied loads. Experiencing millions of articulations throughout an average lifetime, articular cartilage possesses distinct biotribological properties. These require effective lubrication, mediated by the synergistic interaction between fluid and boundary lubricants, to provide a low coefficient of friction and prevent wear at the cartilage surface. Osteoarthritis is the progressive deterioration of articular cartilage and synovial joint structure and function, leading to softer and wear prone tissue on account of altered biochemical composition of the extracellular matrix. Plain radiography remains the most accessible tool and the current standard of care to visualize musculoskeletal diseases and injuries (e.g., osteoarthritis), but cannot directly visualize soft tissues or cartilage, and diagnoses are based solely on boney changes, which occur in the later stages of the disease. Coupled with no way to quantitatively assess tissue health prior to irreversible deterioration, there remains no cure for osteoarthritis. Integral to OA pathology are concomitant changes in the biochemical composition of synovial fluid that result in deterioration of rheological properties, contributing to increased cartilage wear. To address both the lack of quantitative diagnosis methods and lack of chondroprotective therapies, this dissertation presents a dual faceted approach to quantitatively image articular cartilage health, coupled with lubrication strategies to improve cartilage lubrication, and preserve cartilage tissue. This dissertation describes the synthesis of tantalum oxide nanoparticles of varying surface charges for use as contrast agents for rapid, minimally invasive, non-destructive, and quantitative contrast-enhanced computed tomography to assess both the biochemical content and biomechanical integrity of articular cartilage. Ex vivo contrast enhanced computed tomography attenuation using the nanoparticle contrast agent reveals correlations between attenuation and the mechanical and biochemical properties of the tissue. The lubrication strategy described within this dissertation involves introducing a rolling ball element between two surfaces to reduce friction. In this strategy, either single, globular macromolecules or nanoparticles are employed as ball bearings between articulating surfaces to reduce friction when asperities on the surfaces are in direct contact. Rheological characterization and construction of classical Stribeck curves using the lubricant formulations reveal that introducing the rolling element reduces the coefficient of friction during boundary lubrication, while leaving the rheological properties of the base fluid intact. Ex vivo cartilage mechanical testing involving shear deformation under varying speeds and loads reveal improved biotribological performance compared to pure synovial fluid or saline.

First, an in vitro growth experiment was performed to test the hypothesis that applying dynamic unconfined compression during culture produces het- erogeneous remodeling in newborn bovine articular cartilage explants. Het- erogeneous measures of cartilage microstructure were obtained by biochemical assays and quantified polarized light microscopy. Significant differences were measured between the GAG content in the inner and outer portions of the sam- ples stimulated with dynamic unconfined compression. The COL fiber network was found to be more highly aligned in the inner portion of the sample than in the peripheral region. Next, a poroelastic finite element model with a remodeling subroutine was developed to test the hypothesis that the magnitude of relative interstitial fluid velocity and maximum principle strain stimulate GAG and COL fiber network remodeling, respectively, in articular cartilage during culture with dynamic unconfined compression. The GAG remodeling law was successful in predicting the heterogeneous changes in GAG content. The collagen remodeling law was not successful in predicting the changes in the COL network microstructural orientation, suggesting another mechanical cue is responsible for stimulating the remodeling of the COL fiber network.

Joint disorders significantly affect quality of life and present unique challenges for tissue engineering. In the craniofacial space, and especially for the temporomandibular joint (TMJ), there is an unmet need for anatomically precise and mechanically robust cartilage and bone tissues to recapitulate native function. Current surgical reconstruction methods, whether using autologous or synthetic options, suffer from imprecision, comorbidities, complications, and frequently require subsequent operations. Furthermore, many craniofacial graft efforts have focused on improving bone without addressing cartilage, which is essential to proper TMJ function. Thus, there is a compelling need to engineer a human-sized, biologically and anatomically matched cartilage-bone TMJ replacement. This dissertation demonstrates the ability to generate such a graft with native-like properties in a human-sized large animal model by focusing on two aims: (i) establish methods to fabricate and culture anatomically specific, autologous cartilage-bone grafts (Aim 1), and (ii) show improvement of graft performance after six months implantation in vivo compared to previous methods, controls, and native tissue (Aim 2). Using Yucatan mini-pigs as a human-sized model, the ramus-condyle unit (RCU), a geometrically intricate portion of the mandible and primary load bearing section of the TMJ, was targeted for reconstruction. Scaffolds were created using computer tomography (CT) image-guided micromilling of decellularized bone matrix, then infused with autologous adipose-derived chondrogenic and osteogenic progenitors. These biological constructs were then cultured in vitro in a novel dual-perfusion bioreactor before in vivo implantation. Similar in vitro culture of representative constructs done in parallel demonstrated cell attachment and some differentiation. After six months implantation, the dual cartilage-bone RCU grafts maintained their predefined anatomical structure and regenerated full-thickness, stratified, and mechanically robust cartilage over the underlying bone, to a significantly greater extent than either bone-only grafts or acellular scaffolds, and showed remarkable similarity to native tissue. Furthermore, tracking of implanted cells enabled additional insights into the progression of cartilage and bone regeneration. The methods and results established in this dissertation form a promising basis for the next evolution in engineering full-sized, patient-specific, and biologically and mechanically robust TMJ replacements.

L’arthrose et l’arthrite rhumatoïde sont deux pathologies articulaires caractérisées par la dégradation du cartilage articulaire, subséquente à la production de divers médiateurs inflammatoires. Le traitement de ces pathologies se limite généralement à soulager le patient des épisodes douloureux et inflammatoires et à améliorer sa qualité de vie. Dans le cas de l’arthrose, peu de traitements permettent d’enrayer significativement l’évolution de la dégradation du cartilage et donc de la maladie. Par contre, l’arthrite rhumatoïde peut être efficacement ralentie grâce à l’administration de certaines molécules. Néanmoins, ces traitements n’ont généralement montré qu’une efficacité à court-terme, requérant une administration fréquente. L’objectif de ce travail repose donc sur l’élaboration de nouvelles options thérapeutiques permettant de réduire la fréquence d’administration ainsi que les effets indésirables des traitements actuels. La délivrance de molécules en intra-articulaire associée à une libération prolongée offre l’avantage d’exposer les sites directement impliqués dans l’évolution de la maladie à une ou plusieurs molécules efficaces contre l’inflammation et la douleur, et aidant à la régénération du cartilage, durant plusieurs semaines, voire des mois.<p>Des microparticules de PLGA chargées en clonidine ou en bétaméthasone ont donc été optimisées afin d’obtenir des efficacités d’encapsulation appréciables (clonidine HCl :EE ≈ 20% ;dipropionate de bétaméthasone :EE ≈ 70%), une taille adaptée à l’administration intra-articulaire (12 – 38 µm) et une libération de la molécule s’échelonnant sur 5 à 8 semaines. La libération prolongée de la clonidine implique des mécanismes de diffusion de la molécule ainsi que de dégradation/érosion du polymère. Au vu de l’absence de réaction inflammatoire, les microparticules développées sont correctement tolérées par les chondrocytes, synoviocytes, PBMC et neutrophiles, principales cellules impliquées dans les mécanismes inflammatoires de l’arthrose et de l’arthrite rhumatoïde. L’évaluation de l’efficacité anti-inflammatoire des microparticules vides et chargées en clonidine ou en bétaméthasone via l’étude de l’expression et de la sécrétion de différents médiateurs de l’inflammation a permis d’aboutir à plusieurs conclusions :(i) les microparticules vides sont associées à un effet anti-inflammatoire, (ii) les microparticules chargées en clonidine n’ont pas montré d’activité anti-inflammatoire propre pouvant être attribuée à la clonidine, et (iii) les microparticules de bétaméthasone ont confirmé l’effet anti-inflammatoire de la bétaméthasone. Enfin, l’étude de la toxicité des principes actifs et microparticules vides ou chargées a montré une toxicité significative de la clonidine sur les synoviocytes. Néanmoins, l’encapsulation des principes actifs dans les microparticules de PLGA a permis d’éliminer cette toxicité, protégeant donc efficacement les cellules articulaires.<p>Les microparticules développées permettent alors d’envisager l’encapsulation d’autres molécules anti-inflammatoires ou une combinaison de molécules ayant des effets complémentaires (anti-inflammatoire et antidouleur). L’utilisation de la clonidine dans ces indications devra être réévaluée en étudiant de façon approfondie son efficacité dans la douleur. / Both osteoarthritis and rheumatoid arthritis are articular diseases characterized by the degeneration of the joint cartilage, resulting from the production of various inflammatory mediators. The current treatment of these diseases is restricted to alleviate the painful and inflammatory episodes of the patients and to improve its quality of life. In osteoarthritic patients, few treatments allow to significantly stop the evolution of the degradation of the cartilage and, consequently, the disease. In rheumatoid arthritis, the evolution can be slowed down following the administration of some drugs. Nevertheless, these treatments are often associated to a short-term efficacy. The objective of this work is to develop new therapeutic options that allow to reduce the frequency of administration and the side effects of the current treatments. The intraarticular delivery combined to controlled-release presents the advantage to expose the sites directly involved in the evolution of the disease to one or more molecules effective to relieve the pain, inflammation and to help the regeneration of the cartilage.<p>Clonidine or betamethasone-loaded PLGA microparticles were optimized to reach suitable encapsulation efficiencies (clonidine HCl: EE ≈ 20%; betamethasone dipropionate: EE ≈ 70%), an appropriate size for an intraarticular delivery (12 – 38 µm) and a controlled-release of the molecule over 5 to 8 weeks. The release of clonidine implies mechanisms of diffusion and degradation/erosion of the polymer. Given the absence of an inflammatory reaction, the developed microparticles were properly tolerated by the chondrocytes, synoviocytes, PMBC and neutrophils, which are the main cells involved in the inflammatory reaction of osteoarthritis and rheumatoid arthritis. The assessment of the anti-inflammatory efficacy of the drug-free and drug-loaded microparticles through the evaluation of the expression and the secretion of various inflammatory mediators allowed to draw several conclusions: (i) drug-free microparticles were associated to an anti-inflammatory effect, (ii) clonidine-loaded microparticles did not show any anti-inflammatory activity that could be attributed to clonidine, and (iii) betamethasone- loaded microparticles confirmed the anti-inflammatory effect of betamethasone. Finally, the evaluation of the toxicity of the drugs and microparticles showed a significant toxicity of clonidine against synoviocytes. Nevertheless, the encapsulation of the drugs in PLGA microparticles induced the suppression of this toxicity, protecting in this way the articular cells. <p>Entrapping other anti-inflammatory molecules or a combination of molecules with complementary effects (anti-inflammatory and anti-nociceptive drugs) in the PLGA microparticles developed has to be considered. Moreover, the use of clonidine in these indications has to be reassessed by a thorough study of its anti-nociceptive potential.<p><p> / Doctorat en Sciences biomédicales et pharmaceutiques / info:eu-repo/semantics/nonPublished