Isolamento e Caracterização de Células Estaminais do Líquido Amniótico
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2009
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As células estaminais (CE) são células indiferenciadas, não especializadas, capazes de se dividirem e auto-renovarem, originando novas células estaminais. Possuem a
capacidade de se diferenciarem em diferentes linhagens celulares, em condições
biológicas apropriadas.
As células estaminais fetais (CEF) são consideradas uma nova classe de CE, tendo sido posicionadas a nível evolutivo e operacional entre as células estaminais embrionárias(CEE) e as células estaminais adultas (CEA), uma vez que exibem características de pluripotência e multipotência associadas, respectivamente, aos dois tipos celulares referidos.
As CEF integram, entre outras, células estaminais do líquido amniótico - CELA.
Neste trabalho procedeu-se ao isolamento das CELA. Comprovou-se a sua capacidade
de expansão e demonstrou-se a estabilidade citogenética, verificando-se a
manutenção de um cariótipo normal ao longo do processo de expansão celular. Para
além disso caracterizou-se fenotipicamente as CELA em cultura. Estas células
expressam marcadores celulares característicos das CEE, como o POU (Oct-4), c-Kit e SCF, responsáveis pela manutenção de um estado de stemness, caracterizado, por sua
vez pelo estado indiferenciado e pluripotente destas células. Para além destes, as CELA também expressam marcadores celulares característicos de células estaminais mesênquimais (CEM), como o CD29, CD44, CD73, CD90 e CD105, e expressão de HNF-
4a e Nestina.
A expressão de marcadores característicos de CEE e CEA justifica a presença de CE no
líquido amniótico com origem embrionária nas três linhas germinativas, o que explica a
capacidade de diferenciação destas células nos vários tipos celulares.
As CELA são então consideradas uma população celular com um elevado potencial no
que diz respeito à terapia celular e, portanto, determinantes na evolução da
engenharia de tecidos e medicina regenerativa.
Stem cells are undifferentiated cells that retain the ability to self-renew and proliferate in order to provide progenitor cells that can differentiate into specialized cells under appropriated biological conditions. Fetal stem cells are considered a new class of stem cells, developmentally and operationally located between embryonic stem cells and adult stem cells, sharing and exhibiting features of pluripotency and multipotency associated, respectively, to both cellular kinds. Human amniotic fluid stem cells (hAFSCs) are included in the fetal stem cell class’s. In this work we were able to isolate hAFSCs, and we prove that this kind of stem cells are capable of expand extensively, maintaining a normal karyotipe during all the expansion process. RT-PCR analyses showed that this kind of stem cell express some cellular markers characteristic of embryonic stem cells, as POU (Oct-4), c-Kit and SCF, wich are responsible for the maintenance of a state of stemness, characterized for the undifferentiated and pluripotent stem cells; and express HNF-4α and Nestin, wich is an indicator of the differentiation potential of this stem cells. The results of flow citometry analyses showed that this kind of cells also express some cellular markers characteristic of mesenchymal stem cells, as CD29, CD44, CD73, CD90 and CD105. The expression of cellular markers adjusted to both cellular kinds (embryonic and adult stem cells) explains the presence of stem cells in amniotic fluid originated from the three germ cell lines, as well as their differentiation potential into multiple cellular kinds. hAFSCs are then considered a cellular population extremely important in reconstitution therapy and tissue and organ reparation, determinant in the evolution of tissue engineering and regenerative medicine.
Stem cells are undifferentiated cells that retain the ability to self-renew and proliferate in order to provide progenitor cells that can differentiate into specialized cells under appropriated biological conditions. Fetal stem cells are considered a new class of stem cells, developmentally and operationally located between embryonic stem cells and adult stem cells, sharing and exhibiting features of pluripotency and multipotency associated, respectively, to both cellular kinds. Human amniotic fluid stem cells (hAFSCs) are included in the fetal stem cell class’s. In this work we were able to isolate hAFSCs, and we prove that this kind of stem cells are capable of expand extensively, maintaining a normal karyotipe during all the expansion process. RT-PCR analyses showed that this kind of stem cell express some cellular markers characteristic of embryonic stem cells, as POU (Oct-4), c-Kit and SCF, wich are responsible for the maintenance of a state of stemness, characterized for the undifferentiated and pluripotent stem cells; and express HNF-4α and Nestin, wich is an indicator of the differentiation potential of this stem cells. The results of flow citometry analyses showed that this kind of cells also express some cellular markers characteristic of mesenchymal stem cells, as CD29, CD44, CD73, CD90 and CD105. The expression of cellular markers adjusted to both cellular kinds (embryonic and adult stem cells) explains the presence of stem cells in amniotic fluid originated from the three germ cell lines, as well as their differentiation potential into multiple cellular kinds. hAFSCs are then considered a cellular population extremely important in reconstitution therapy and tissue and organ reparation, determinant in the evolution of tissue engineering and regenerative medicine.
Descrição
Dissertação de Mestrado em Genética Molecular Comparativa e Tecnológica