RESUMO
Recent studies have found that cellular self-renewal capacity in brain cancer is heterogeneous, with only stem-like cells having this property. A link between adult stem cells and cancer stem cells remains, however, to be shown. Here, we describe the emergence of cancer stem-like cells from in vitro cultured brain stem cells. Adult rat subventricular zone (SVZ) stem cells transformed into tumorigenic cell lines after expansion in vitro. These cell lines maintained characteristic features of stem-like cells expressing Nestin, Musashi-1 and CD133, but continued to proliferate upon differentiation induction. Karyotyping detected multiple acquired chromosomal aberrations, and syngeneic transplantation into the brain of adult rats resulted in malignant tumor formation. Tumors revealed streak necrosis and displayed a neural as well as an undifferentiated phenotype. Deficient downregulation of platelet-derived growth factor (PDGF) receptor alpha was identified as candidate mechanism for tumor cell proliferation, and its knockdown by siRNA resulted in a reduction of cell growth. Our data point to adult brain precursor cells to be transformed in malignancies. Furthermore, in vitro expansion of adult neural stem cells, which will be mandatory for therapeutic strategies in neurological disorders, also harbors the risk for amplifying precursor cells with acquired genetic abnormalities and induction of malignant tumors after transplantation.
Assuntos
Células-Tronco Adultas/patologia , Neoplasias Encefálicas/patologia , Transformação Celular Neoplásica/patologia , Células-Tronco Neoplásicas/patologia , Neurônios/patologia , Animais , Western Blotting , Encéfalo/citologia , Neoplasias Encefálicas/genética , Transplante de Tecido Encefálico/efeitos adversos , Diferenciação Celular/fisiologia , Transformação Celular Neoplásica/genética , Aberrações Cromossômicas , Imuno-Histoquímica , RNA Interferente Pequeno , Ratos , Ratos Wistar , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/genética , Receptor alfa de Fator de Crescimento Derivado de Plaquetas/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , TransfecçãoRESUMO
Neural stem cells (NSCs) from the adult central nervous system are currently being investigated for their potential use in autologous cell replacement strategies. High expansion rates of NSCs in culture are crucial for the generation of a sufficient amount of cells needed for transplantation. Here, we describe efficient growth of adult NSCs in Neurobasal medium containing B27 supplement under clonal and low-density conditions in the absence of serum or conditioned medium. Expansion of up to 15-fold within 1 week was achieved on low-density NSC cultures derived from the lateral ventricle wall, the hippocampal formation, and the spinal cord of adult rats. A 27% single-cell cloning efficiency in Neurobasal/B27 combination further demonstrates its growth-promoting ability. Multipotency and nontumorgenicity of NSCs were retained despite the high rate of culture expansion. In addition, increased cell survival was obtained when Accutase, instead of trypsin, was used for enzymatic dissociation of NSC cultures. This work provides an important step toward the development of standardized protocols for highly efficient in vitro expansion of NSCs from the adult central nervous system to move more closely to the clinical use of NSCs.
