<b>Background and objectives.</b> Knee osteoarthritis (KOA) is a leading cause of disability in the developed world. Additionally, it is possibly linked with premature mortality. Low-grade inflammation is associated with a high risk of non-traumatic KOA incidence but also with metabolic syndrome. Knee injury is a major risk factor for KOA and is associated with an inflammatory response. Heterogeneity of both the symptoms and progression makes early identification of individuals at risk difficult. The aim of this thesis was to examine the diagnostic value of selected biomarkers to identify microscopic synovitis, examine their predictive value for KOA development and any association with excess premature mortality. <b>Methods.</b> Four serum biomarkers, COMP, resistin, C3M and CRPM, all linked with both knee pain and synovial or systemic inflammation, were selected and an additional sonographic marker was tested. Data from two cohorts has been utilised for the purpose of this thesis: the Oxford Knee Injury Cohort, a prospective study of NHS patients with recent ACL and/or meniscal injuries, and the Chingford women's study, a community-based prospective cohort with over 23 years of follow-up. <b>Results.</b> Cross-sectionally, sonographic markers correspond well with microscopically defined post-traumatic synovitis. Two biomarkers, COMP and resistin, were associated with development of incident radiographic KOA (RKOA), while two others, C3M and CRPM, predicted development of painful RKOA independently to age and BMI. High C3M levels and presence of painful RKOA were associated with premature mortality. Knee pain alone, especially in presence of RKOA was an independent predictor of mortality, but RKOA without pain was not. <b>Conclusion.</b> These results support the use of the studied biomarkers in complex predictive models for development of KOA and associated premature mortality. Taking the competing risk of death into account is an important consideration in this field.

Orientador: Maria Helena Andrade Santana / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Química / Made available in DSpace on 2018-08-17T16:54:08Z (GMT). No. of bitstreams: 1 Shimojo_AndreaArrudaMartins_M.pdf: 3448066 bytes, checksum: 3fb61b807d37d04fb162230b89e8166b (MD5) Previous issue date: 2011 / Resumo: Neste trabalho foram estudadas as reações de modificação química do ácido hialurônico (AH) com divinilsulfona (DVS) para preparação de géis reticulados e com carbodiimidas para preparação de fosfolipídios derivatizados. A modificação com DVS envolve os grupos hidroxílicos do AH, enquanto a modificação mediada por carbodiimida envolve os grupos carboxílicos do AH e o grupo amina do fosfolipídio. A extensão da reação de reticulação do AH com divinilsulfona (DVS) foi estudada em relação à razão mássica AH/DVS e concentração de NaOH na mistura reacional. O grau de modificação dos géis foi correlacionado com a reologia e a razão de intumescimento (RI). As reações de reticulação foram realizadas com AH de origem bacteriana de massa molar média da ordem de 105 Da. O gel plano preparado com concentrações de AH e NaCl de 3g/100g e em pH alcalino foi submetido a alto cisalhamento para produção de partículas. A reologia desses géis foi estudada em relação ao diâmetro médio das partículas e razão mássica gel/fluido. Um gel com propriedades reológicas semelhantes ao fluido sinovial foi preparado com partículas entre 75 e 100 µm e razão mássica gel/fluido (AH 1g/100g) de 75:25, o qual foi avaliado quanto à sua citotoxicidade em ensaios in vitro. Os resultados mostraram que a concentração de DVS é diretamente proporcional à dureza e à viscoelasticidade do gel, mas inversamente proporcional a RI. Os géis mostraram comportamento não Newtoniano pseudoplástico em toda a faixa de taxa de cisalhamento estudada. Nas medidas oscilatórias, foi observado comportamento característico de géis, com o módulo elástico maior que o viscoso (G'>G"), ambos independentes da frequência. Baseado nos parâmetros reológicos, os géis foram classificados em covalentes e fracos. A alcalinidade da mistura reacional foi inversamente proporcional à viscosidade do meio e à dureza do gel. O tamanho de partícula foi diretamente proporcional à dureza do gel e à viscosidade. O aumento da porcentagem da fase fluida (solução de AH 1% não reticulado) causou redução em G' e G" e nas forças de extrusão. O gel analisado in vitro não apresentou efeitos citotóxicos. O conjugado AH-fosfolipídio foi preparado através da ativação do AH com 1-ethyl-3-[3-dimethyl)aminopropyl] carbodiimide hydrochloride (EDCA) em pH 4,0 e adição do fosfolipídio 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) em pH 8,6. Os conjugados obtidos foram separados por ultrafiltração e caracterizados por cromatografia em camada delgada, ressonância magnética nuclear de próton, teor de fosfato e AH. A capacidade de incorporação do conjugado em bicamada lipídica foi avaliada através da preparação de lipossomas compostos de lecitina de ovo e DPPE, contendo 5, 10 e 15% do conjugado. Os resultados obtidos indicaram modificação do tamanho, polidispersidade e das propriedades de superfície dos lipossomas preparados com incorporação do conjugado AH DPPE. Esses resultados contribuem para o desenvolvimento de produtos à base de AH, para aplicações nas áreas médica, farmacêutica e de cosméticos / Abstract: In this work, it was studied the chemical modification of hyaluronic acid (HA) with divinyl sulfone (DVS) for the preparation of cross-linked hydrogels and with carbodiimides for the preparation of derivatized phospholipids. The modification with DVS involves the hydroxyl groups of HA, while the carbodiimidemediated modification involves the carboxylic groups of HA and the amino group of the phospholipid. The cross-linking reaction of HA with DVS has been studied regarding the HA/DVS mass ratio and NaOH concentration in the reaction mixture. The modification degree of the gels was correlated with their rheological properties and swelling ratios (SR). The crosslinking reactions were performed with bacterial hyaluronic acid of molecular weight around 105 Da. The hydrogels plans prepared with concentrations of HA and NaCl of 3g/100g and at alkaline pH were subjected to high shear to produce particles. The rheology of these hydrogels was studied in relation to the average particle size and gel/fluid mass ratio. A gel with rheological properties similar to synovial fluid was prepared with particles between 75 and 100 micrometres and gel/fluid mass ratio (HA 1g/100g) of 75:25. The cytotoxicity of this gel was evaluated through tests in vitro. The results showed that the DVS concentration was directly proportional to the hardness and viscoelasticity of the hydrogel, but it was inversely proportional to the SR. The hydrogels showed non-Newtonian pseudoplastic behavior in the range of shear rate studied. In the oscillatory measurements, the cross-linked HA hydrogels exhibited characteristic behavior of gels, with the elastic modulus higher than the viscous modulus (G' > G"), both frequency independent. Furthermore, based on the rheological parameters, the gels were classified as covalent and weak. The alkalinity of the reaction mixture was inversely proportional to the viscosity of the medium and to the hardness of the hydrogel. The particle size was directly proportional to the hardness and viscosity of the hydrogel. The increase in the percentage of fluid phase caused a reduction in G' and G" and the extrusion forces. The hydrogel analised in vitro showed no cytotoxic effects. The HA-phospholipid conjugate was prepared through the activation of HA with 1-ethyl-3-[3-dimethyl aminopropyl] carbodiimide hydrochloride (EDCA) at pH 4.0 and addition of the phospholipid 1,2-dipalmitoyl-sn-glycero-3-phosphoethanolamine (DPPE) at pH 8.6. The conjugates were separated by ultrafiltration and characterized by thin layer chromatography, proton nuclear magnetic resonance, phosphate and HA content. The incorporation ability of the conjugate into the lipid bilayer was evaluated through the preparation of liposomes composed of egg lecithin and DPPE containing 5, 10 and 15% of the conjugate. The liposomes were characterized by size, polydispersity and zeta potential, and compared with the control (no conjugate). The results indicated modification of the size, polydispersity and surface properties of liposome-conjugates HA-DPPE. These results contribute to the development of HA-based products for applications in medical, pharmaceutical and cosmetic industries / Mestrado / Desenvolvimento de Processos Biotecnologicos / Mestre em Engenharia Química