Development of a spheroid model to study the pathogenesis and regulation of gliomas cell death(Dr. S. Wharton &Prof. Ian Whittle)
2 year project with grant funding of £17,000.00
2000 – 2002
This research project is to study the control and regulation of cell death in gliomas, using a gliomas culture system which allows cells to grow as three dimensional structures (spheroids). As these increase in size they develop diffusion gradients which mimic those seen in life, allowing study of the factors which lead to necrosis and apoptosis. (Necrosis is “accidental” cell death due to an insult which is sufficient to kill the cell. In contrast, apoptosis, or programmed cell death, involves the initiation of a complex and regulated series of cell pathways which finally result in cell death). The development of central necrosis will be studied in detail using electron microscopy. We will examine whether the p53 status influences the susceptibility of the tumour cells to apoptosis and necrosis in this model, and further explore the role of this protein in the cell death response to oxidative stress. Oxidative stress may arise from processes such as ischaemia developing within the tumour, and also as a response to various therapies. It is therefore likely to be an important process in contributing to cell death in these tumours.
Conclusion:
The highly unsaturated fatty acids (HUFA) of the n-6 and n-3 series are involved in cell signalling in normal and transformed cells and have recently been associated with pathways leading to tumour cell death. The antitumour activity of three HUFA (arachidonic acid, gamma linolenic acid and eicosapentaenoic acid) were studied in glioma cells and tissue. Using five gliomas models, including primary cell suspensions and prepared from 46 human gliomas samples and an in vivo rat C6 glioma model, we obtained evidence that, following exposure to HUFA, either administered into the medium surrounding human gliomas cells or in 16 preparations of multicellular spheroids derived from human and rodent gliomas cell lines (C6, MOG, U87, U373) or administered intra-tumourally by infusion using osmotic mini-pumps in 48 rats, gliomas regression and apoptosis were detected. Additionally, synergy between gamma irradiation and HUFA administration was observed in 13 experiments analyzing C6 glioma cell apoptosis in vitro. These pro-apoptotic and antiproliferative activities were observed using both C18 and C20 fatty acids of the n-6 and n-3 series, but no when saturated and monounsaturated C18 and C20 fatty acid preparations were used. In the gliomas infusion model, in addition to the apoptosis detected in gliomas tissue infused with HUFA for 3-7 days, preservation of normal neural tissue and vasculature in adjacent brain was observed. Also, there was little evidence of acute inflammatory infiltration in regressing tumours. Our findings suggest that intraparenchymal infusion of HUFA may be effective in stimulating gliomas regression.