Avaliação de novas propostas em arcabouços tridimencionais (3D) para cultura de células-tronco mesenquinas e condogênese
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Data
2009-02-19
Autores
Moroz, Andrei[UNESP]
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Editor
Universidade Estadual Paulista (Unesp)
Resumo
Levando-se em consideração avanços tecnológicos na área médica e o impacto dos programas de saúde que determinaram curvas de longevidade cada vez maiores e taxas de natalidade cada vez menores, o novo desafio do gestor público são as conseqüências: o envelhecimento e a vida como uma “doença crônica”. Entre os principais desafios está a abordagem das doenças crônico-degenerativas que determinam o aumento de lesões cartilaginosas articulares. A débil capacidade de regeneração e as limitações das alternativas de tratamento fazem as técnicas derivadas da biotecnologia, como o transplante autólogo de condrócitos (TAC) e o uso de células-tronco, o foco das investigações. O TAC requer coleta de material cartilaginoso de área sadia, podendo causar nova lesão; no entanto, pode-se evitar este perigo com o uso de células-tronco. As células-tronco mesenquimais, adultas, podem se diferenciar em condrócitos mediante o uso de meio de cultura específico em consonância a um arcabouço 3D, mas muitos problemas como evitar a calcificação e estimular a condrogênese em meio favorável constituem o desafio para os pesquisadores na atualidade. 1) produzir anticorpo monoclonal específico a CTMs de coelho para monitorá-las, 2) determinar o volume ideal de coleta de medula óssea para microencapsulação e condrogênese, 3) realizar a microencapsulação das CTMs em novos arcabouços: BIOGEL3D e BACTCELL3D, comparando seu desempenho com o modelo clássico em alginato. Foram utilizados 25 coelhos Nova Zelândia sendo divididos em diferentes grupos em função do volume de MO coletado: G1 = 6mL, G2 = 9mL, G3 = 12mL, G4 = 15mL e G5 = volume ideal de coleta determinado pelos indicadores dos outros grupos. O material coletado foi diluído 1:2 em RPMI 1640 com 3.000U de heparina sódica. Após a contagem celular, as amostras foram submetidas a separação em gradiente...
Technological advances in the medical area combined with the impact of health programs that enhance longevity, together with lower natality rates created new challenges to the public manager such as aging and life as a “chronic disease”. Among the major problems are the chronic degenerative diseases that increase articular lesions. The limited regeneration capabilities and the limitations of actual treatment alternatives made biotechnology derived techniques the focus of investigations. The autologous chondrocyte transplantation (ACT) requires a small biopsy of health cartilage, which can lead to a new lesion. However, the use of stem cells can avoid this possibility. Mesenchymal stem cells (MSCs) can differentiate into chondrocytes by using a specific culture medium together with a 3D scaffold, but some questions such as the risks of calcification remain as key factors to researchers. 1) to produce a monoclonal antibody that recognizes MSCs in order to characterize them, 2) to determine the optimal bone marrow collection volume for cell microencapsulation and chondrogenesis and 3) to microencapsulate MSCs in two novel scaffolds: BIOGEL3D and BACTCELL3D, comparing them with sodium alginate. 25 New Zealand rabbits were divided into 5 groups related to bone marrow collection volume: G1 = 6mL, G2 = 9mL, G3 = 12mL, G4 = 15mL and G5 = optimal volume determined by the study. The collected material was diluted in RPMI1640 medium 1:2, with 3000U sodium heparin. After cell count and viability assessment the samples were submitted to density gradient centrifugation in order to isolate the lymphomononuclear (LMN) fraction. These cells were seeded to obtain and expand the MSCs in DMEM Knockout® (InvitrogenTM) supplemented with antibiotic/antimycotic, Lglutamine, essential aminoacids, non essential aminoacids and fetal bovine serum (all from InvitrogenTM). The cells were cultivated in 5% CO2 ...(Complete abstract click electronic access below)
Technological advances in the medical area combined with the impact of health programs that enhance longevity, together with lower natality rates created new challenges to the public manager such as aging and life as a “chronic disease”. Among the major problems are the chronic degenerative diseases that increase articular lesions. The limited regeneration capabilities and the limitations of actual treatment alternatives made biotechnology derived techniques the focus of investigations. The autologous chondrocyte transplantation (ACT) requires a small biopsy of health cartilage, which can lead to a new lesion. However, the use of stem cells can avoid this possibility. Mesenchymal stem cells (MSCs) can differentiate into chondrocytes by using a specific culture medium together with a 3D scaffold, but some questions such as the risks of calcification remain as key factors to researchers. 1) to produce a monoclonal antibody that recognizes MSCs in order to characterize them, 2) to determine the optimal bone marrow collection volume for cell microencapsulation and chondrogenesis and 3) to microencapsulate MSCs in two novel scaffolds: BIOGEL3D and BACTCELL3D, comparing them with sodium alginate. 25 New Zealand rabbits were divided into 5 groups related to bone marrow collection volume: G1 = 6mL, G2 = 9mL, G3 = 12mL, G4 = 15mL and G5 = optimal volume determined by the study. The collected material was diluted in RPMI1640 medium 1:2, with 3000U sodium heparin. After cell count and viability assessment the samples were submitted to density gradient centrifugation in order to isolate the lymphomononuclear (LMN) fraction. These cells were seeded to obtain and expand the MSCs in DMEM Knockout® (InvitrogenTM) supplemented with antibiotic/antimycotic, Lglutamine, essential aminoacids, non essential aminoacids and fetal bovine serum (all from InvitrogenTM). The cells were cultivated in 5% CO2 ...(Complete abstract click electronic access below)
Descrição
Palavras-chave
Envelhecimento - Aspectos biológicos, Biologia celular, Clonagem, Scaffolds 3D, Mesenchymal stem cells, Chondrogenesis
Como citar
MOROZ, Andrei. Avaliação de novas propostas em arcabouços tridimencionais (3D) para cultura de células-tronco mesenquinas e condogênese. 2009. 78 f. Dissertação (mestrado) - Universidade Estadual Paulista, Faculdade de Medicina de Botucatu, 2009.