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Establishment and Comprehensive Molecular Characterization of an Immortalized Glioblastoma Cell Line from a Brazilian Patient.

ABSTRACT: Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults, with few effective treatment strategies. The research on the development of new treatments is often constrained by the limitations of preclinical models, which fail to accurately replicate the disease's essential characteristics. Herein, we describe the obtention, molecular, and functional characterization of the GBM33 cell line. This cell line belongs to the GBM class according to the World Health Organization 2021 Classification of Central Nervous System Tumors, identified by methylation profiling. GBM33 expresses the astrocytic marker GFAP, as well as markers of neuronal origin commonly expressed in GBM cells, such as βIII-tubulin and neurofilament. Functional assays demonstrated an increased growth rate when compared to the U87 commercial cell line and a similar sensitivity to temozolamide. GBM33 cells retained response to serum starvation, with reduced growth and diminished activation of the Akt signaling pathway. Unlike LN-18 and LN-229 commercial cell lines, GBM33 is able to produce primary cilia upon serum starvation. In summary, the successful establishment and comprehensive characterization of this GBM cell line provide researchers with invaluable tools for studying GBM biology, identifying novel therapeutic targets, and evaluating the efficacy of potential treatments.

SUBMITTER: da Silva FF

PROVIDER: S-EPMC10649167 | biostudies-literature | 2023 Nov

REPOSITORIES: biostudies-literature

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Establishment and Comprehensive Molecular Characterization of an Immortalized Glioblastoma Cell Line from a Brazilian Patient.

da Silva Fernanda F FF Lupinacci Fernanda C S FCS Elias Bruno D S BDS Beserra Adriano O AO Sanematsu Paulo P Roffe Martin M Kulikowski Leslie D LD Costa Felipe D'almeida FD Santos Tiago G TG Hajj Glaucia N M GNM

International journal of molecular sciences 20231101 21

Glioblastoma (GBM) is the most common and aggressive primary brain tumor in adults, with few effective treatment strategies. The research on the development of new treatments is often constrained by the limitations of preclinical models, which fail to accurately replicate the disease's essential characteristics. Herein, we describe the obtention, molecular, and functional characterization of the GBM33 cell line. This cell line belongs to the GBM class according to the World Health Organization 2 ...[more]

PMID: 37958846

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Project description:Background and objectiveGingival keratinocytes are used in model systems to investigate the interaction between periodontal bacteria and the epithelium in the initial stages of the periodontal disease process. Primary gingival epithelial cells (GECs) have a finite lifespan in culture before they enter senescence and cease to replicate, while epithelial cells immortalized with viral proteins can exhibit chromosomal rearrangements. The aim of this study was to generate a telomerase immortalized human gingival epithelial cell line and compare its in vitro behaviour to that of human GECs.Material and methodsHuman primary GECs were immortalized with a bmi1/hTERT combination to prevent cell cycle triggers of senescence and telomere shortening. The resultant cell-line, telomerase immortalized gingival keratinocytes (TIGKs), were compared to GECs for cell morphology, karyotype, growth and cytokeratin expression, and further characterized for replicative lifespan, expression of toll-like receptors and invasion by P. gingivalis.ResultsTIGKs showed morphologies, karyotype, proliferation rates and expression of characteristic cytokeratin proteins comparable to GECs. TIGKs underwent 36 passages without signs of senescence and expressed transcripts for toll-like receptors 1-6, 8 and 9. A subpopulation of cells underwent stratification after extended time in culture. The cytokeratin profiles of TIGK monolayers were consistent with basal cells. When allowed to stratify, cytokeratin profiles of TIGKs were consistent with suprabasal cells of the junctional epithelium. Further, TIGKs were comparable to GECs in previously reported levels and kinetics of invasion by wild-type P. gingivalis and an invasion defective ΔserB mutant.ConclusionResults confirm bmi1/hTERT immortalization of primary GECs generated a robust cell line with similar characteristics to the parental cell type. TIGKs represent a valuable model system for the study of oral bacteria interactions with host gingival cells.

Project description:Glioblastoma multiforme is a highly aggressive malignant primary brain tumor in humans, with poor prognosis. ANGM5 has been established from a cerebral glioblastoma multiforme in a 72-years-old Iraqi man who underwent surgery for an intracranial tumor in 2005. The morphology, growth kinetics, karyotype, immunocytochemistry and angiogenesis factors expression profile were studied. ANGM5 has been grown continuously for more than 200 serial passages in culture for the last 13 years. The cultured cells are elongated or multipolar in shape and the population doubling time is 28 hours. The karyotype is complex, and the chromosomal number varied between 39-114. ANGM5 is resistant to Temozolomide, cisplatin, vincristine and etoposide. This chemoresistance could be explained by the overexpression of the breast cancer resistance protein (BCRP) and MGMT. Immunocytochemistry analysis of glial markers demonstrated that cells are positive for glial fibrillary acidic protein (GFAP), and negative for nestin and calbindin. Protein microarray analysis showed high production of tissue inhibitor of metalloproteinase 2 (TIMP2) as well as other factors that are important for the invasiveness and aggressiveness of glioblastoma. ANGM5 is a useful addition to the cell lines currently available for study of the pathobiology and chemoresistance properties of glioblastoma multiforme and antitumor drug discovery. Angiogenesis signaling is being evaluated in new human Iraqi glioblastoma multiforme cell line. Here, we measured angiogenesis factors released by cancer cells in vitro. We find that glioblastoma up-regulates many proteins and metabolites during the logarithmic phase, suggesting initiation of their endo-vesiculo-membrane system. Importantly, GM-CSF and G-CSF as they increase angiogenesis through promoting endothelial cell function. Also, b-FGF which promote tumor growth and proliferation. Moreover, it showed up-regulation to inflammatory, and extracellular matrix proteins. These factors will help proliferating glioblastoma cells to overcome stress conditions, such as cytotoxic chemotherapy, serum deprivation, hypoxia in vitro and in vivo. These findings support other evidence of chemoresistance ability of this cell line named AMGN5 as it found to resist several types of conventional chemotherapies.

Dataset Information

Establishment and Comprehensive Molecular Characterization of an Immortalized Glioblastoma Cell Line from a Brazilian Patient.

Publications

Establishment and Comprehensive Molecular Characterization of an Immortalized Glioblastoma Cell Line from a Brazilian Patient.

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