Novel immunohistochemical methodology for the investigation of stem cell-like subpopulations in glioma spheroids and migratory cells

Viton Cheng, Filomena Estevez, Aruna Chakrabarty, Julia V. Cockle, Susan C Short, Anke Bruning-Richardson

Research output: Contribution to journalMeeting Abstract

Abstract

INTRODUCTION: Targeting tumour invasion may represent a complementary treatment when added to standard care in highly infiltrating tumours such as glioblastomas. Invasiveness is believed to be related to stem cell-like qualities of distinct subpopulations of glioma cells. The characterisation of phenotypic differences between invading and non-invading cells will aid the development of specific anti-migratory drugs. A novel immunohistochemical approach was taken to phenotypically characterise invasive migratory glioma cells after drug treatment in a 3D spheroid invasion assay. METHOD: Spheroids generated from U87 and U251 cells were embedded in a collagen matrix and treated over 72 hours with the anti-migratory drugs LiCl and Bio-Indirubin. A protocol was established to prepare the collagen embedded spheroids and migratory cells for immunohistochemistry (IHC). Markers of proliferation, apoptosis and stemness were optimised for IHC. RESULTS: In both U87 and U251, protein expression profiles in cells maintained in the original core were distinct from profiles in migratory cells. The most striking change of expression was observed for SOX-2 in U251, where drug treatment led to a reduction of SOX-2 levels in core-maintained cells whereas migrating cells exhibited higher levels of SOX-2. We could also quantify drug-induced changes in proliferation and apoptotic events in spheroids generated from both cell lines using this method. CONCLUSION: A novel IHC protocol allowed the investigation of the invasive behaviour of cells in a 3D experimental model. Protein expression profiles of markers of stemness indicate that subpopulations of cells exist within glioma spheroids that may respond differently to anti-migratory drugs.
LanguageEnglish
Pages3
Number of pages1
JournalNeuro-Oncology
Volume17
Issue numberS8
DOIs
Publication statusPublished - Nov 2015
Externally publishedYes
EventBritish Neuro-Oncology Society Annual Meeting: "Neuro-Oncology Across the Ages" - Childhood to Old Age - University of Nottingham, Nottingham, United Kingdom
Duration: 1 Jul 20153 Jul 2015
https://www.bnos.org.uk/bnos-2015-annual-meeting/ (Link to Conference Information)

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Glioma
Stem Cells
Pharmaceutical Preparations
Immunohistochemistry
Collagen
Glioblastoma
Neoplasms
Proteins
Theoretical Models
Therapeutics
Apoptosis
Cell Line

Cite this

Cheng, Viton ; Estevez, Filomena ; Chakrabarty, Aruna ; Cockle, Julia V. ; Short, Susan C ; Bruning-Richardson, Anke. / Novel immunohistochemical methodology for the investigation of stem cell-like subpopulations in glioma spheroids and migratory cells. In: Neuro-Oncology. 2015 ; Vol. 17, No. S8. pp. 3.
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abstract = "INTRODUCTION: Targeting tumour invasion may represent a complementary treatment when added to standard care in highly infiltrating tumours such as glioblastomas. Invasiveness is believed to be related to stem cell-like qualities of distinct subpopulations of glioma cells. The characterisation of phenotypic differences between invading and non-invading cells will aid the development of specific anti-migratory drugs. A novel immunohistochemical approach was taken to phenotypically characterise invasive migratory glioma cells after drug treatment in a 3D spheroid invasion assay. METHOD: Spheroids generated from U87 and U251 cells were embedded in a collagen matrix and treated over 72 hours with the anti-migratory drugs LiCl and Bio-Indirubin. A protocol was established to prepare the collagen embedded spheroids and migratory cells for immunohistochemistry (IHC). Markers of proliferation, apoptosis and stemness were optimised for IHC. RESULTS: In both U87 and U251, protein expression profiles in cells maintained in the original core were distinct from profiles in migratory cells. The most striking change of expression was observed for SOX-2 in U251, where drug treatment led to a reduction of SOX-2 levels in core-maintained cells whereas migrating cells exhibited higher levels of SOX-2. We could also quantify drug-induced changes in proliferation and apoptotic events in spheroids generated from both cell lines using this method. CONCLUSION: A novel IHC protocol allowed the investigation of the invasive behaviour of cells in a 3D experimental model. Protein expression profiles of markers of stemness indicate that subpopulations of cells exist within glioma spheroids that may respond differently to anti-migratory drugs.",
author = "Viton Cheng and Filomena Estevez and Aruna Chakrabarty and Cockle, {Julia V.} and Short, {Susan C} and Anke Bruning-Richardson",
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Novel immunohistochemical methodology for the investigation of stem cell-like subpopulations in glioma spheroids and migratory cells. / Cheng, Viton; Estevez, Filomena; Chakrabarty, Aruna; Cockle, Julia V.; Short, Susan C; Bruning-Richardson, Anke.

In: Neuro-Oncology, Vol. 17, No. S8, 11.2015, p. 3.

Research output: Contribution to journalMeeting Abstract

TY - JOUR

T1 - Novel immunohistochemical methodology for the investigation of stem cell-like subpopulations in glioma spheroids and migratory cells

AU - Cheng, Viton

AU - Estevez, Filomena

AU - Chakrabarty, Aruna

AU - Cockle, Julia V.

AU - Short, Susan C

AU - Bruning-Richardson, Anke

PY - 2015/11

Y1 - 2015/11

N2 - INTRODUCTION: Targeting tumour invasion may represent a complementary treatment when added to standard care in highly infiltrating tumours such as glioblastomas. Invasiveness is believed to be related to stem cell-like qualities of distinct subpopulations of glioma cells. The characterisation of phenotypic differences between invading and non-invading cells will aid the development of specific anti-migratory drugs. A novel immunohistochemical approach was taken to phenotypically characterise invasive migratory glioma cells after drug treatment in a 3D spheroid invasion assay. METHOD: Spheroids generated from U87 and U251 cells were embedded in a collagen matrix and treated over 72 hours with the anti-migratory drugs LiCl and Bio-Indirubin. A protocol was established to prepare the collagen embedded spheroids and migratory cells for immunohistochemistry (IHC). Markers of proliferation, apoptosis and stemness were optimised for IHC. RESULTS: In both U87 and U251, protein expression profiles in cells maintained in the original core were distinct from profiles in migratory cells. The most striking change of expression was observed for SOX-2 in U251, where drug treatment led to a reduction of SOX-2 levels in core-maintained cells whereas migrating cells exhibited higher levels of SOX-2. We could also quantify drug-induced changes in proliferation and apoptotic events in spheroids generated from both cell lines using this method. CONCLUSION: A novel IHC protocol allowed the investigation of the invasive behaviour of cells in a 3D experimental model. Protein expression profiles of markers of stemness indicate that subpopulations of cells exist within glioma spheroids that may respond differently to anti-migratory drugs.

AB - INTRODUCTION: Targeting tumour invasion may represent a complementary treatment when added to standard care in highly infiltrating tumours such as glioblastomas. Invasiveness is believed to be related to stem cell-like qualities of distinct subpopulations of glioma cells. The characterisation of phenotypic differences between invading and non-invading cells will aid the development of specific anti-migratory drugs. A novel immunohistochemical approach was taken to phenotypically characterise invasive migratory glioma cells after drug treatment in a 3D spheroid invasion assay. METHOD: Spheroids generated from U87 and U251 cells were embedded in a collagen matrix and treated over 72 hours with the anti-migratory drugs LiCl and Bio-Indirubin. A protocol was established to prepare the collagen embedded spheroids and migratory cells for immunohistochemistry (IHC). Markers of proliferation, apoptosis and stemness were optimised for IHC. RESULTS: In both U87 and U251, protein expression profiles in cells maintained in the original core were distinct from profiles in migratory cells. The most striking change of expression was observed for SOX-2 in U251, where drug treatment led to a reduction of SOX-2 levels in core-maintained cells whereas migrating cells exhibited higher levels of SOX-2. We could also quantify drug-induced changes in proliferation and apoptotic events in spheroids generated from both cell lines using this method. CONCLUSION: A novel IHC protocol allowed the investigation of the invasive behaviour of cells in a 3D experimental model. Protein expression profiles of markers of stemness indicate that subpopulations of cells exist within glioma spheroids that may respond differently to anti-migratory drugs.

U2 - 10.1093/neuonc/nov284.14

DO - 10.1093/neuonc/nov284.14

M3 - Meeting Abstract

VL - 17

SP - 3

JO - Neuro-Oncology

T2 - Neuro-Oncology

JF - Neuro-Oncology

SN - 1522-8517

IS - S8

ER -