Isolamento, cultivo e caracterização de células-tronco CD133+ de glioblastoma humano

OBJECTIVE: To establish the method of isolation and culture of human glioblastoma neurospheres, and the purification of their stem cells, followed by the process of obtaining tumor subspheres, immunophenotypically characterizing this clonogenic set. METHODS: Through the processing of glioblastoma samples (n=3), the following strategy of action was adopted: (i) establish primary culture of glioblastoma; (ii) isolation and culture of tumor neurospheres; (iii) purify cells that initiate tumors (CD133+) by magnetic separation system (MACS); (iv) obtain tumor subspheres; (v) study the expression of the markers nestin, CD133, and GFAP. RESULTS: The study successfully described the process of isolation and culture of glioblastoma subspheres, which consist of a number of clonogenic cells immunophenotypically characterized as neural, which are able to initiate tumor formation. CONCLUSION: These findings may contribute to a better understanding of the process of gliomagenesis.OBJETIVO: Estabelecer o método de isolamento e cultivo das neuroesferas de glioblastoma humano, bem como purificação de suas células-tronco, seguido do processo de obtenção de subesferas tumorais, caracterizando imunofenotipicamente esse conjunto clonogênico. MÉTODOS: Por meio do processamento de amostras de glioblastomas (n=3), cumpriu-se a seguinte estratégia de ação: (i) estabelecimento da cultura primária de glioblastoma; (ii) isolamento e cultura de neuroesferas tumorais; (iii) purificação das células que iniciam os tumores (CD133+) por sistema de separação magnética (MACS); (iv) obtenção subesferas tumorais; (v) estudo da expressão de marcadores GFAP, CD133 e nestina. RESULTADOS: Este estudo descreveu com sucesso o processo de isolamento e cultivo de subesferas de glioblastoma, as quais são constituídas por um conjunto clonogênico de células caracterizadas imunofenotipicamente como neurais, capazes de iniciar a formação tumoral. CONCLUSÃO: Estes achados poderão contribuir para a compreensão do processo de gliomagênese.Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Hospital Israelita Albert Einstein Instituto do CérebroHospital Israelita Albert Einstein Centro de Pesquisa ExperimentalHospital Israelita Albert Einstein Faculdade de EnfermagemUniversidade Federal de São Paulo (UNIFESP) Department of Neurology and NeurosurgeryHospital Israelita Albert Einstein Center for Neuro-oncologyHospital Israelita Albert EinsteinUNIFESP, Department of Neurology and NeurosurgerySciEL

Isolation, cultivation and characterization of CD133+ stem cells from human glioblastoma

OBJECTIVE: To establish the method of isolation and culture of human glioblastoma neurospheres, and the purification of their stem cells, followed by the process of obtaining tumor subspheres, immunophenotypically characterizing this clonogenic set. METHODS: Through the processing of glioblastoma samples (n=3), the following strategy of action was adopted: (i) establish primary culture of glioblastoma; (ii) isolation and culture of tumor neurospheres; (iii) purify cells that initiate tumors (CD133+) by magnetic separation system (MACS); (iv) obtain tumor subspheres; (v) study the expression of the markers nestin, CD133, and GFAP. RESULTS: The study successfully described the process of isolation and culture of glioblastoma subspheres, which consist of a number of clonogenic cells immunophenotypically characterized as neural, which are able to initiate tumor formation. CONCLUSION: These findings may contribute to a better understanding of the process of gliomagenesis