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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

H&E stains of orthotopic tumors formed in athymic mice, showing hallmark characteristics of GBMs.(A and B) All of the lines showed diffuse invasion of normal parenchymal. The hallmark features of glioblastoma were identified in the brains of mice injected with oncosphere lines, including increased mitotic activity (C, arrows), necrosis (D, designated “N”), and vascular proliferation (E, arrows). The oncosphere lines also demonstrated other histologic features specific to invasive gliomas, including spread through white matter tracts (F), neuronal satellitosis (G), and subventricular tumor formation (H). Original magnification for panel A was 25x; for F and H, 50x; for B and D, 100x; and for C, E, and G, 200x.
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pone.0150271.g004: H&E stains of orthotopic tumors formed in athymic mice, showing hallmark characteristics of GBMs.(A and B) All of the lines showed diffuse invasion of normal parenchymal. The hallmark features of glioblastoma were identified in the brains of mice injected with oncosphere lines, including increased mitotic activity (C, arrows), necrosis (D, designated “N”), and vascular proliferation (E, arrows). The oncosphere lines also demonstrated other histologic features specific to invasive gliomas, including spread through white matter tracts (F), neuronal satellitosis (G), and subventricular tumor formation (H). Original magnification for panel A was 25x; for F and H, 50x; for B and D, 100x; and for C, E, and G, 200x.

Mentions: Pathological evaluation showed some heterogeneity between the lines. All the xenografts contained cells with angulated or elongated nuclei and hyperchromasia, characteristic of astrocytomas. All of the xenografts generated showed diffuse invasion into brain parenchyma (Fig 4A and 4B). Furthermore, the standard histologic criteria used to diagnose GBM were identified among the different lines, including increased mitotic activity (Fig 4C), necrosis (Fig 4D), and vascular proliferation (Fig 4E). While significant mitotic activity was seen in all cases, the presence of vascular proliferation and necrosis was variable. The invasive growth was very prominent in many of the lines, including invasion of the corpus callosum and other white matter tracts (Fig 4F). This invasion is characteristic of infiltrating gliomas but often absent in adherent models [8, 9]. Some xenografts demonstrated other features specific to invasive glial neoplasms including neuronal satellitosis (Fig 4G) and subventricular tumor formation (Fig 4H). Taken together, the classic histopathological features of GBM were well-represented in this cohort of tumor xenografts.


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)

H&E stains of orthotopic tumors formed in athymic mice, showing hallmark characteristics of GBMs.(A and B) All of the lines showed diffuse invasion of normal parenchymal. The hallmark features of glioblastoma were identified in the brains of mice injected with oncosphere lines, including increased mitotic activity (C, arrows), necrosis (D, designated “N”), and vascular proliferation (E, arrows). The oncosphere lines also demonstrated other histologic features specific to invasive gliomas, including spread through white matter tracts (F), neuronal satellitosis (G), and subventricular tumor formation (H). Original magnification for panel A was 25x; for F and H, 50x; for B and D, 100x; and for C, E, and G, 200x.
© Copyright Policy
Related In: Results  -  Collection

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

pone.0150271.g004: H&E stains of orthotopic tumors formed in athymic mice, showing hallmark characteristics of GBMs.(A and B) All of the lines showed diffuse invasion of normal parenchymal. The hallmark features of glioblastoma were identified in the brains of mice injected with oncosphere lines, including increased mitotic activity (C, arrows), necrosis (D, designated “N”), and vascular proliferation (E, arrows). The oncosphere lines also demonstrated other histologic features specific to invasive gliomas, including spread through white matter tracts (F), neuronal satellitosis (G), and subventricular tumor formation (H). Original magnification for panel A was 25x; for F and H, 50x; for B and D, 100x; and for C, E, and G, 200x.
Mentions: Pathological evaluation showed some heterogeneity between the lines. All the xenografts contained cells with angulated or elongated nuclei and hyperchromasia, characteristic of astrocytomas. All of the xenografts generated showed diffuse invasion into brain parenchyma (Fig 4A and 4B). Furthermore, the standard histologic criteria used to diagnose GBM were identified among the different lines, including increased mitotic activity (Fig 4C), necrosis (Fig 4D), and vascular proliferation (Fig 4E). While significant mitotic activity was seen in all cases, the presence of vascular proliferation and necrosis was variable. The invasive growth was very prominent in many of the lines, including invasion of the corpus callosum and other white matter tracts (Fig 4F). This invasion is characteristic of infiltrating gliomas but often absent in adherent models [8, 9]. Some xenografts demonstrated other features specific to invasive glial neoplasms including neuronal satellitosis (Fig 4G) and subventricular tumor formation (Fig 4H). Taken together, the classic histopathological features of GBM were well-represented in this cohort of tumor xenografts.

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