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Publication
Nanopartículas de colagénio para terapias da cartilagem baseadas em recursos marinhos
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Abstract(s)
A cartilagem articular é constituída por matriz extracelular e condrócitos, sendo
ancorada no osso subcondral. O seu principal problema é a limitada capacidade de reparação
relacionada com a sua falta de vascularização. Embora tenham sido testadas diversas
metodologias para o tratamento da degeneração da cartilagem, a sua incompleta eficácia, bem
como complicações associadas, torna necessário e urgente o desenvolvimento de novas técnicas
para essa finalidade. Assim, a nanotecnologia apresenta-se como ferramenta com potencial
aplicação no tratamento da cartilagem, nomeadamente pelo uso de nanopartículas capazes de
transportar moléculas bioativas. Sendo o colagénio o principal constituinte da matriz
extracelular cartilaginosa, o uso de colagénio proveniente de várias fontes, como organismos
marinhos, tem sido investigado para regeneração cartilaginosa, dada a sua potencial
biocompatibilidade.
Nesta tese de Mestrado são apresentados resultados preliminares relativamente à
preparação de nanopartículas de colagénio de alforreca e tubarão azul visando sua futura
aplicação como transportadores de fatores de crescimento para engenharia de tecido
cartilaginoso. A preparação dessas nanopartículas foi otimizada, sendo estas caracterizadas por
Espalhamento Dinâmico da Luz, medição de potencial Zeta e Microscopia Eletrónica de
Transmissão. Posteriormente, realizaram-se ensaios de viabilidade e diferenciação celular com
células estaminais mesenquimais humanas após exposição às nanopartículas de colagénio de
ambas fontes. A diferenciação celular foi estudada por métodos histoquímicos e avaliação da
expressão génica usando a técnica de RT-qPCR.
As partículas sintetizadas com colagénio de alforreca e tubarão azul apresentaram
diâmetros hidrodinâmicos de, respetivamente, 453 ± 23 nm e 634 ± 18 nm. No entanto, na
análise feita às partículas de colagénio de tubarão por microscopia de transmissão (amostras
desidratadas), verificou-se que estas tinham forma elíptica e dimensões na gama 200-300 nm
(eixo maior). Ambos os tipos de nanopartículas apresentaram potencial Zeta negativo e boa
citocompatibilidade. Por outro lado, a presença das nanopartículas pareceu não afetar a
diferenciação condroblástica das hMSCs.
Articular cartilage consists of an extracellular matrix and chondrocytes, anchored in the subchondral bone. Its main problem is the limited repair capacity due to the lack of vascularization. Although several methodologies for the treatment of cartilage degeneration have been tested, their incomplete efficacy, as well as the complications associated, make it necessary and urgent to develop new techniques for this purpose. Therefore, nanotechnology presents itself as a tool with potential application in the treatment of cartilage, namely by the use of nanoparticles capable of transporting bioactive molecules. As collagen is the main constituent of cartilaginous extracellular matrix, the use of collagen from various sources, including marine organisms, has been investigated for cartilaginous regeneration, given its potential biocompatibility. In this Master thesis, preliminary results are presented regarding the preparation of jellyfish and blue shark collagen nanoparticles having in view their possible application as vehicles for the transport of growth factors in cartilage tissue engineering. The preparation of these nanoparticles has been optimized and they were characterized by Dynamic Light Scattering, Zeta potential measurement and Transmission Electron Microscopy. Subsequently, cell viability and differentiation assays were performed using human mesenchymal stem cells after exposure to jellyfish and blue shark collagen nanoparticles. Cell differentiation was studied by histochemical methods and gene expression using RT-qPCR. The particles synthesized with jellyfish and blue shark collagen had hydrodynamic diameters of 453 ± 189 nm and 634 ± 81 nm, respectively. However, transmission microscopy analysis of shark collagen particles (dehydrated samples) showed that they were elliptical in shape and dimensions in the range 200-300 nm (long axis). Both types of nanoparticles showed negative Zeta potential and good cytocompatibility. On the other hand, the presence of nanoparticles did not seem to affect the chondroblastic differentiation of hMSCs.
Articular cartilage consists of an extracellular matrix and chondrocytes, anchored in the subchondral bone. Its main problem is the limited repair capacity due to the lack of vascularization. Although several methodologies for the treatment of cartilage degeneration have been tested, their incomplete efficacy, as well as the complications associated, make it necessary and urgent to develop new techniques for this purpose. Therefore, nanotechnology presents itself as a tool with potential application in the treatment of cartilage, namely by the use of nanoparticles capable of transporting bioactive molecules. As collagen is the main constituent of cartilaginous extracellular matrix, the use of collagen from various sources, including marine organisms, has been investigated for cartilaginous regeneration, given its potential biocompatibility. In this Master thesis, preliminary results are presented regarding the preparation of jellyfish and blue shark collagen nanoparticles having in view their possible application as vehicles for the transport of growth factors in cartilage tissue engineering. The preparation of these nanoparticles has been optimized and they were characterized by Dynamic Light Scattering, Zeta potential measurement and Transmission Electron Microscopy. Subsequently, cell viability and differentiation assays were performed using human mesenchymal stem cells after exposure to jellyfish and blue shark collagen nanoparticles. Cell differentiation was studied by histochemical methods and gene expression using RT-qPCR. The particles synthesized with jellyfish and blue shark collagen had hydrodynamic diameters of 453 ± 189 nm and 634 ± 81 nm, respectively. However, transmission microscopy analysis of shark collagen particles (dehydrated samples) showed that they were elliptical in shape and dimensions in the range 200-300 nm (long axis). Both types of nanoparticles showed negative Zeta potential and good cytocompatibility. On the other hand, the presence of nanoparticles did not seem to affect the chondroblastic differentiation of hMSCs.
Description
Keywords
Nanopartículas Colagénio Cartilagem Regeneração Nanoparticles Collagen Cartilage Regeneration Bioquímica Aplicada . Faculdade de Ciências Exatas e da Engenharia