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Establishment and Biological Characterization of a Panel of Glioblastoma Multiforme (GBM) and GBM Variant Oncosphere Cell Lines.

Binder ZA, Wilson KM, Salmasi V, Orr BA, Eberhart CG, Siu IM, Lim M, Weingart JD, Quinones-Hinojosa A, Bettegowda C, Kassam AB, Olivi A, Brem H, Riggins GJ, Gallia GL - PLoS ONE (2016)

Bottom Line: When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity.Multipotency was confirmed using in vitro differentiation.These oncosphere cell lines will expand the resources available for preclinical study.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, MD, United States of America.

ABSTRACT

Objective: Human tumor cell lines form the basis of the majority of present day laboratory cancer research. These models are vital to studying the molecular biology of tumors and preclinical testing of new therapies. When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity. Using a modified neural stem cell culture technique, here we report the characterization of GBM cell lines including GBM variants.

Methods: Tumor tissue samples were obtained intra-operatively and cultured in neural stem cell conditions containing growth factors. Tumor lines were characterized in vitro using differentiation assays followed by immunostaining for lineage-specific markers. In vivo tumor formation was assayed by orthotopic injection in nude mice. Genetic uniqueness was confirmed via short tandem repeat (STR) DNA profiling.

Results: Thirteen oncosphere lines derived from GBM and GBM variants, including a GBM with PNET features and a GBM with oligodendroglioma component, were established. All unique lines showed distinct genetic profiles by STR profiling. The lines assayed demonstrated a range of in vitro growth rates. Multipotency was confirmed using in vitro differentiation. Tumor formation demonstrated histologic features consistent with high grade gliomas, including invasion, necrosis, abnormal vascularization, and high mitotic rate. Xenografts derived from the GBM variants maintained histopathological features of the primary tumors.

Conclusions: We have generated and characterized GBM suspension lines derived from patients with GBMs and GBM variants. These oncosphere cell lines will expand the resources available for preclinical study.

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Related in: MedlinePlus

Outcome of short tandem repeat DNA profiling of the established cell lines showed the unique profile of each line.Twenty markers were assayed and an overlap of 16 or more markers (80%) was considered an indication of cross-contamination. HeLa and K562 were included both as quality controls and to show lack of contamination from these ubiquitous cell lines.
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pone.0150271.g002: Outcome of short tandem repeat DNA profiling of the established cell lines showed the unique profile of each line.Twenty markers were assayed and an overlap of 16 or more markers (80%) was considered an indication of cross-contamination. HeLa and K562 were included both as quality controls and to show lack of contamination from these ubiquitous cell lines.

Mentions: To ensure cell line independence, we performed STR profiling (Fig 2). An overlap of 80% was considered non-coincidental and suggestive of a shared primary source [23]. With the exception of JHU-1016A and JHU-1016B, none of the lines showed significant genetic overlap. JHU-1016A and JHU-1016B were established from the same tumor specimen and were, as anticipated, identical with respect to their STR profiles. Among the remaining lines, the overlap was minimal, ranging from 0 to 40%. STR profile markers are shown in S1 Table. The cell lines’ STR profiles were also compared to 15 established astrocytoma cell lines from ATCC. Overlap ranged from 17 to 61%, below the 80% threshold and confirming the genetically unique nature of each new cell line established.


Establishment and Biological Characterization of a Panel of Glioblastoma Multiforme (GBM) and GBM Variant Oncosphere Cell Lines.

Binder ZA, Wilson KM, Salmasi V, Orr BA, Eberhart CG, Siu IM, Lim M, Weingart JD, Quinones-Hinojosa A, Bettegowda C, Kassam AB, Olivi A, Brem H, Riggins GJ, Gallia GL - PLoS ONE (2016)

Outcome of short tandem repeat DNA profiling of the established cell lines showed the unique profile of each line.Twenty markers were assayed and an overlap of 16 or more markers (80%) was considered an indication of cross-contamination. HeLa and K562 were included both as quality controls and to show lack of contamination from these ubiquitous cell lines.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4814135&req=5

pone.0150271.g002: Outcome of short tandem repeat DNA profiling of the established cell lines showed the unique profile of each line.Twenty markers were assayed and an overlap of 16 or more markers (80%) was considered an indication of cross-contamination. HeLa and K562 were included both as quality controls and to show lack of contamination from these ubiquitous cell lines.
Mentions: To ensure cell line independence, we performed STR profiling (Fig 2). An overlap of 80% was considered non-coincidental and suggestive of a shared primary source [23]. With the exception of JHU-1016A and JHU-1016B, none of the lines showed significant genetic overlap. JHU-1016A and JHU-1016B were established from the same tumor specimen and were, as anticipated, identical with respect to their STR profiles. Among the remaining lines, the overlap was minimal, ranging from 0 to 40%. STR profile markers are shown in S1 Table. The cell lines’ STR profiles were also compared to 15 established astrocytoma cell lines from ATCC. Overlap ranged from 17 to 61%, below the 80% threshold and confirming the genetically unique nature of each new cell line established.

Bottom Line: When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity.Multipotency was confirmed using in vitro differentiation.These oncosphere cell lines will expand the resources available for preclinical study.

View Article: PubMed Central - PubMed

Affiliation: Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, MD, United States of America.

ABSTRACT

Objective: Human tumor cell lines form the basis of the majority of present day laboratory cancer research. These models are vital to studying the molecular biology of tumors and preclinical testing of new therapies. When compared to traditional adherent cell lines, suspension cell lines recapitulate the genetic profiles and histologic features of glioblastoma multiforme (GBM) with higher fidelity. Using a modified neural stem cell culture technique, here we report the characterization of GBM cell lines including GBM variants.

Methods: Tumor tissue samples were obtained intra-operatively and cultured in neural stem cell conditions containing growth factors. Tumor lines were characterized in vitro using differentiation assays followed by immunostaining for lineage-specific markers. In vivo tumor formation was assayed by orthotopic injection in nude mice. Genetic uniqueness was confirmed via short tandem repeat (STR) DNA profiling.

Results: Thirteen oncosphere lines derived from GBM and GBM variants, including a GBM with PNET features and a GBM with oligodendroglioma component, were established. All unique lines showed distinct genetic profiles by STR profiling. The lines assayed demonstrated a range of in vitro growth rates. Multipotency was confirmed using in vitro differentiation. Tumor formation demonstrated histologic features consistent with high grade gliomas, including invasion, necrosis, abnormal vascularization, and high mitotic rate. Xenografts derived from the GBM variants maintained histopathological features of the primary tumors.

Conclusions: We have generated and characterized GBM suspension lines derived from patients with GBMs and GBM variants. These oncosphere cell lines will expand the resources available for preclinical study.

Show MeSH
Related in: MedlinePlus