New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas
Background. The incidence of brain gliomas firmly occupies a leading position among all central nervous system tumors – 40–50 % of the cases detected, more than half of them are glioblastoma. Existing cell lines and cultivation methods do not reflect all the features of the three-dimensional (3D) or...
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ABV-press
2019-12-01
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| Series: | Успехи молекулярной онкологии |
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| Online Access: | https://umo.abvpress.ru/jour/article/view/243 |
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| author | Yu. A. Khochenkova I. G. Dyrda Yu. S. Machkova E. Sh. Solomko T. A. Sidorova D. A. Khochenkov E. A. Avilova |
| author_facet | Yu. A. Khochenkova I. G. Dyrda Yu. S. Machkova E. Sh. Solomko T. A. Sidorova D. A. Khochenkov E. A. Avilova |
| author_sort | Yu. A. Khochenkova |
| collection | DOAJ |
| description | Background. The incidence of brain gliomas firmly occupies a leading position among all central nervous system tumors – 40–50 % of the cases detected, more than half of them are glioblastoma. Existing cell lines and cultivation methods do not reflect all the features of the three-dimensional (3D) organization of native glioblastoma. The use of temozolomide leads to the development of drug resistance and acute relapse, followed by a poor clinical outcome. The development of resistance is largely associated with the presence of tumor stem cells in the population and intratumoral heterogeneity. Obtaining 3D cultures from the primary material will allow us to save the stem cell pool and tumor-specific features.The study objective. Get a 3D model based on primary cell cultures, which allows you to save a heterogeneous population and the original phenotype of tumor cells.Materials and methods. We used U-87MG human glioma cells and GBM002 primary cell culture obtained from surgical material with a confirmed diagnosis of glioblastoma. Neurospheres were obtained from cell lines, the growth of which was monitored using the InCell Analyzer 6000 automatic cell analysis system. Flow cytometry was used to determine the CD133+ cell content. The expression of the receptor tyrosine kinases VEGFR1, VEGFR2 (endothelial growth factor type 1 and 2 receptors), FGFR2 (fibroblast growth factor receptor type 2) and the hypoxia marker HIF-1α (hypoxia inducible factor, 1α) in the neurospheres was evaluated using confocal microscopy.Results. GBM002 glioblastoma cells isolated from the surgical material formed neurospheres, while the number of CD133+ cells increased from 1–2 to 16–19 % compared with two-dimensional cultures. During long-term cultivation of cells with non-cytotoxic doses of temozolomide, it was found that such cells form smaller neurospheres compared to control cells. It was shown that the expression of receptor tyrosine kinases during cultivation of GBM002 glioblastoma cells in neurospheres differs from that in two-dimensional cultures. We found that in neurospheres, the expression of FGFR2 and VEGFR1, is significantly increased.Conclusion. 3D cultivation of primary cultures allows one to obtain a more heterogeneous population of tumor cells that reflects the spatial heterogeneity of cells, increase the pool of stem cells and recreate hypoxia conditions inside the brain micro-tumors. |
| format | Article |
| id | doaj-art-71b239b4d09340f9bb783e3ceaf9f178 |
| institution | Kabale University |
| issn | 2313-805X 2413-3787 |
| language | Russian |
| publishDate | 2019-12-01 |
| publisher | ABV-press |
| record_format | Article |
| series | Успехи молекулярной онкологии |
| spelling | doaj-art-71b239b4d09340f9bb783e3ceaf9f1782025-08-20T04:00:14ZrusABV-pressУспехи молекулярной онкологии2313-805X2413-37872019-12-0164697410.17650/2313-805X-2019-6-4-69-74175New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomasYu. A. Khochenkova0I. G. Dyrda1Yu. S. Machkova2E. Sh. Solomko3T. A. Sidorova4D. A. Khochenkov5E. A. Avilova6N.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of RussiaS.P. Botkin City Clinical HospitalN.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of RussiaN.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of RussiaN.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of RussiaN.N. Blokhin National Medical Research Center of Oncology, Ministry of Health of Russia; Togliatti State UniversityGE HealthcareBackground. The incidence of brain gliomas firmly occupies a leading position among all central nervous system tumors – 40–50 % of the cases detected, more than half of them are glioblastoma. Existing cell lines and cultivation methods do not reflect all the features of the three-dimensional (3D) organization of native glioblastoma. The use of temozolomide leads to the development of drug resistance and acute relapse, followed by a poor clinical outcome. The development of resistance is largely associated with the presence of tumor stem cells in the population and intratumoral heterogeneity. Obtaining 3D cultures from the primary material will allow us to save the stem cell pool and tumor-specific features.The study objective. Get a 3D model based on primary cell cultures, which allows you to save a heterogeneous population and the original phenotype of tumor cells.Materials and methods. We used U-87MG human glioma cells and GBM002 primary cell culture obtained from surgical material with a confirmed diagnosis of glioblastoma. Neurospheres were obtained from cell lines, the growth of which was monitored using the InCell Analyzer 6000 automatic cell analysis system. Flow cytometry was used to determine the CD133+ cell content. The expression of the receptor tyrosine kinases VEGFR1, VEGFR2 (endothelial growth factor type 1 and 2 receptors), FGFR2 (fibroblast growth factor receptor type 2) and the hypoxia marker HIF-1α (hypoxia inducible factor, 1α) in the neurospheres was evaluated using confocal microscopy.Results. GBM002 glioblastoma cells isolated from the surgical material formed neurospheres, while the number of CD133+ cells increased from 1–2 to 16–19 % compared with two-dimensional cultures. During long-term cultivation of cells with non-cytotoxic doses of temozolomide, it was found that such cells form smaller neurospheres compared to control cells. It was shown that the expression of receptor tyrosine kinases during cultivation of GBM002 glioblastoma cells in neurospheres differs from that in two-dimensional cultures. We found that in neurospheres, the expression of FGFR2 and VEGFR1, is significantly increased.Conclusion. 3D cultivation of primary cultures allows one to obtain a more heterogeneous population of tumor cells that reflects the spatial heterogeneity of cells, increase the pool of stem cells and recreate hypoxia conditions inside the brain micro-tumors.https://umo.abvpress.ru/jour/article/view/243glioblastoma3d culturereceptor tyrosine kinasestemozolomide |
| spellingShingle | Yu. A. Khochenkova I. G. Dyrda Yu. S. Machkova E. Sh. Solomko T. A. Sidorova D. A. Khochenkov E. A. Avilova New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas Успехи молекулярной онкологии glioblastoma 3d culture receptor tyrosine kinases temozolomide |
| title | New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas |
| title_full | New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas |
| title_fullStr | New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas |
| title_full_unstemmed | New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas |
| title_short | New approaches in 3D modeling of in vitro growth of primary cultures of malignant gliomas |
| title_sort | new approaches in 3d modeling of in vitro growth of primary cultures of malignant gliomas |
| topic | glioblastoma 3d culture receptor tyrosine kinases temozolomide |
| url | https://umo.abvpress.ru/jour/article/view/243 |
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