C81
An efficient method for derivation and propagation of glioblastoma cell lines that express markers associated with NG2 lineage
Talal Fael Al-Mahyani1, Siolian Ball1, Koichi Ichimura2, Peter Collins2, Colin Watts1
1University of Cambridge, Department of Clinical Neurosciences, Cambridge, UK, 2University of Cambridge, Department of Pathology, Cambridge, UK
Aims
Glioblastoma (GBM) is highly hetergenous tumour in terms of cellular composition, however, it is still unknown if this cellular heterogeneity implies functional one. Our aim is to develop a protocol to derive and propagate human tumour cell lines from GBM samples under serum-free conditions and to investigate the role of different cellular subpopulations in the neoplastic process.
Method
Fresh GBM specimens were dissociated and cultured in defined serum-free media to form primary spheres. Spheroids were harvested and re-cultured on adhesive substrate to grow as a primary monolayer.
Tumour formation and expansion in vivo was examined. Transcriptional and genetic profiles of GBM cell lines and their parent tumours were compared.
Results
Enforcing primary spheres to grow as primary monolayer allowed subsequent long term propagation of GBM cell lines as secondary spheroid or monolayer cultures. Grafting of GBM cell lines resulted in tumour formation with local invasion and notable migration into host brain.
Array comparative genomic hybridization showed that the molecular cytogenetic profiles of our cell lines were characteristic to their parent tumours.
Notably, markers associated with NG2 lineage (NG2, Olig2, PDGFaR, NKX2.2 and Pax6) were expressed in all cell lines tested and their parent tumours.
Summary
We describe a protocol for the efficient derivation and propagation of GBM cell lines from clinical samples. These cell lines recapitulate GBM in vivo, have characteristic molecular profiles similar to their parent tumours and express markers associated with NG2 lineage which may be of functional importance.
