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RB mutation and RAS overexpression induce resistance to NK cell-mediated cytotoxicity in glioma cells.

Orozco-Morales M, Sánchez-García FJ, Golán-Cancela I, Hernández-Pedro N, Costoya JA, de la Cruz VP, Moreno-Jiménez S, Sotelo J, Pineda B - Cancer Cell Int. (2015)

Bottom Line: All these cells were characterized in terms of Rb and Ras gene expression, morphology, proliferative capacity, expression of MHC I, Rae1δ, and Rae1αβγδε, mult1, H60a, H60b, H60c, as ligands for NK cell receptors, and their susceptibility to NK cell-mediated cytotoxicity.Our results show that transformation of astrocytes (Rb loss, Ras overexpression, or both) induced phenotypical and functional changes associated with resistance to NK cell-mediated cytotoxicity.Abstract available from the publisher.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de inmunorregulación, Escuela Nacional de Ciencias Biologicas, Instituto Politecnico Nacional, Mexico, DF Mexico ; Molecular Oncology Laboratory MOL, CIMUS; IDIS Departamento de Fisioloxia, Universidade de Santiago de Compostela, Av de Barcelona s/n 15782, Santiago de Compostela, Spain ; Neuroimmunology and Neuro-Oncology Unit, Instituto Nacional de Neurología y Neurocirugía, Insurgentes sur 3877, 14269 Mexico City, Mexico.

ABSTRACT
Several theories aim to explain the malignant transformation of cells, including the mutation of tumor suppressors and proto-oncogenes. Deletion of Rb (a tumor suppressor), overexpression of mutated Ras (a proto-oncogene), or both, are sufficient for in vitro gliomagenesis, and these genetic traits are associated with their proliferative capacity. An emerging hallmark of cancer is the ability of tumor cells to evade the immune system. Whether specific mutations are related with this, remains to be analyzed. To address this issue, three transformed glioma cell lines were obtained (Rb(-/-), Ras(V12), and Rb(-/-)/Ras(V12)) by in vitro retroviral transformation of astrocytes, as previously reported. In addition, Ras(V12) and Rb(-/-)/Ras(V12) transformed cells were injected into SCID mice and after tumor growth two stable glioma cell lines were derived. All these cells were characterized in terms of Rb and Ras gene expression, morphology, proliferative capacity, expression of MHC I, Rae1δ, and Rae1αβγδε, mult1, H60a, H60b, H60c, as ligands for NK cell receptors, and their susceptibility to NK cell-mediated cytotoxicity. Our results show that transformation of astrocytes (Rb loss, Ras overexpression, or both) induced phenotypical and functional changes associated with resistance to NK cell-mediated cytotoxicity. Moreover, the transfer of cell lines of transformed astrocytes into SCID mice increased resistance to NK cell-mediated cytotoxicity, thus suggesting that specific changes in a tumor suppressor (Rb) and a proto-oncogene (Ras) are enough to confer resistance to NK cell-mediated cytotoxicity in glioma cells and therefore provide some insight into the ability of tumor cells to evade immune responses.

No MeSH data available.


Related in: MedlinePlus

Characterization of in vitro transformed astrocytes. (a) Morphological changes of astrocytes stained with violet crystal, (b) expression of GFAP and GFP in transformed astrocytes, by immunofluorescence, (c) expression of pRb, p53, p-p53, RasV12 and p-H2AX, by Western blot with specific antibodies, (d) cell senescence, as assessed by the percentage of SA-β-galactosidase positive cells, (e) cell proliferation rate, as assessed by violet crystal violet uptake. All images are representative of at least three independent experiments
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Fig1: Characterization of in vitro transformed astrocytes. (a) Morphological changes of astrocytes stained with violet crystal, (b) expression of GFAP and GFP in transformed astrocytes, by immunofluorescence, (c) expression of pRb, p53, p-p53, RasV12 and p-H2AX, by Western blot with specific antibodies, (d) cell senescence, as assessed by the percentage of SA-β-galactosidase positive cells, (e) cell proliferation rate, as assessed by violet crystal violet uptake. All images are representative of at least three independent experiments

Mentions: No significant cell senescence, as assessed by SA-β-gal expression was observed in any of the transformed astrocytes, and the maximal proliferation rate was observed in the cRb−/−/RasV12 cells. All these results, shown in Fig. 1, were very similar to the reported by Seoane et al. [12], in addition to confirming previous data, the cells that were specifically derived for this work were characterized.Fig. 1


RB mutation and RAS overexpression induce resistance to NK cell-mediated cytotoxicity in glioma cells.

Orozco-Morales M, Sánchez-García FJ, Golán-Cancela I, Hernández-Pedro N, Costoya JA, de la Cruz VP, Moreno-Jiménez S, Sotelo J, Pineda B - Cancer Cell Int. (2015)

Characterization of in vitro transformed astrocytes. (a) Morphological changes of astrocytes stained with violet crystal, (b) expression of GFAP and GFP in transformed astrocytes, by immunofluorescence, (c) expression of pRb, p53, p-p53, RasV12 and p-H2AX, by Western blot with specific antibodies, (d) cell senescence, as assessed by the percentage of SA-β-galactosidase positive cells, (e) cell proliferation rate, as assessed by violet crystal violet uptake. All images are representative of at least three independent experiments
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4491266&req=5

Fig1: Characterization of in vitro transformed astrocytes. (a) Morphological changes of astrocytes stained with violet crystal, (b) expression of GFAP and GFP in transformed astrocytes, by immunofluorescence, (c) expression of pRb, p53, p-p53, RasV12 and p-H2AX, by Western blot with specific antibodies, (d) cell senescence, as assessed by the percentage of SA-β-galactosidase positive cells, (e) cell proliferation rate, as assessed by violet crystal violet uptake. All images are representative of at least three independent experiments
Mentions: No significant cell senescence, as assessed by SA-β-gal expression was observed in any of the transformed astrocytes, and the maximal proliferation rate was observed in the cRb−/−/RasV12 cells. All these results, shown in Fig. 1, were very similar to the reported by Seoane et al. [12], in addition to confirming previous data, the cells that were specifically derived for this work were characterized.Fig. 1

Bottom Line: All these cells were characterized in terms of Rb and Ras gene expression, morphology, proliferative capacity, expression of MHC I, Rae1δ, and Rae1αβγδε, mult1, H60a, H60b, H60c, as ligands for NK cell receptors, and their susceptibility to NK cell-mediated cytotoxicity.Our results show that transformation of astrocytes (Rb loss, Ras overexpression, or both) induced phenotypical and functional changes associated with resistance to NK cell-mediated cytotoxicity.Abstract available from the publisher.

View Article: PubMed Central - PubMed

Affiliation: Laboratorio de inmunorregulación, Escuela Nacional de Ciencias Biologicas, Instituto Politecnico Nacional, Mexico, DF Mexico ; Molecular Oncology Laboratory MOL, CIMUS; IDIS Departamento de Fisioloxia, Universidade de Santiago de Compostela, Av de Barcelona s/n 15782, Santiago de Compostela, Spain ; Neuroimmunology and Neuro-Oncology Unit, Instituto Nacional de Neurología y Neurocirugía, Insurgentes sur 3877, 14269 Mexico City, Mexico.

ABSTRACT
Several theories aim to explain the malignant transformation of cells, including the mutation of tumor suppressors and proto-oncogenes. Deletion of Rb (a tumor suppressor), overexpression of mutated Ras (a proto-oncogene), or both, are sufficient for in vitro gliomagenesis, and these genetic traits are associated with their proliferative capacity. An emerging hallmark of cancer is the ability of tumor cells to evade the immune system. Whether specific mutations are related with this, remains to be analyzed. To address this issue, three transformed glioma cell lines were obtained (Rb(-/-), Ras(V12), and Rb(-/-)/Ras(V12)) by in vitro retroviral transformation of astrocytes, as previously reported. In addition, Ras(V12) and Rb(-/-)/Ras(V12) transformed cells were injected into SCID mice and after tumor growth two stable glioma cell lines were derived. All these cells were characterized in terms of Rb and Ras gene expression, morphology, proliferative capacity, expression of MHC I, Rae1δ, and Rae1αβγδε, mult1, H60a, H60b, H60c, as ligands for NK cell receptors, and their susceptibility to NK cell-mediated cytotoxicity. Our results show that transformation of astrocytes (Rb loss, Ras overexpression, or both) induced phenotypical and functional changes associated with resistance to NK cell-mediated cytotoxicity. Moreover, the transfer of cell lines of transformed astrocytes into SCID mice increased resistance to NK cell-mediated cytotoxicity, thus suggesting that specific changes in a tumor suppressor (Rb) and a proto-oncogene (Ras) are enough to confer resistance to NK cell-mediated cytotoxicity in glioma cells and therefore provide some insight into the ability of tumor cells to evade immune responses.

No MeSH data available.


Related in: MedlinePlus