The molecular heterogeneity of high-grade astrocytomas underlies the difficulties in the development of representative and valuable in vitro experimental models for their studies.The purpose of our study was to estimate the value of astrocytoma-associated antigens (AAAs) – IL13R?2, Fra-1, EphA2 – and the most common molecular aberrations typical for astrocytomas as potential markers to screen the status of tumour-derived cell cultures in vitro.
Methods:
The tumour-derived cell cultures were established from high-grade astrocytomas. The expression analyses of the tested genes were performed via semi-quantitative real-time PCR and subsequently verified by immunohistochemical and immunocytochemical technique.
The analyses of molecular aberrations at DNA level included gene dosage status evaluation based on real-time PCR, sequencing analysis, and loss of heterozygosity (LOH) assay.
Results:
The expression analyses based on semi-quantitative real-time PCR showed that in the final stage of culture the expression level of all tested AAAs was significantly higher or at least comparable to that of primary tumours; however, two expression patterns were observed during cell culture establishment. Analysis at the single cell level via immunocytochemistry also demonstrated an increase of the level of tested proteins and/or selection of tumour cell populations strongly positive for AAAs vs.
other cell types including admixed non-tumoural cells. Confrontation of AAA expression data with the results of molecular analyses at DNA level seems to support the latter, revealing that the expression pattern of astrocytoma-associated antigens in tumour-derived cells in subsequent stages of culture is convergent with changes in the molecular profile of examined cell populations.
Conclusions:
The consistency of the obtained results seems to support the use of the selected AAAs, in particular IL13R?2 and Fra-1, as tools facilitating the establishment of tumour-derived cultures.
However, the intratumoural heterogeneity of high-grade astrocytomas may require further detailed characterisation of the molecular profile of a tumour in order to evaluate the value of the experimental model in relation to the individual context of particular studies.
Author: Monika Witusik-PerkowskaMagdalena ZakrzewskaMalgorzata SzybkaWielislaw PapierzDariusz J JaskolskiPawel P LiberskiBeata Sikorska
Credits/Source: Cancer Cell International 2014, 14:82
Published on: 2014-08-22
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