Assignment of 1H nuclear magnetic resonance visible polyunsaturated fatty acids in BT4C gliomas undergoing ganciclovir-thymidine kinase gene therapy-induced programmed cell death.

Polyunsaturated fatty acids (PUFAs), as detected by (1)H nuclear magnetic resonance (NMR) spectroscopy, accumulate into BT4C glioma during ganciclovir-thymidine kinase gene therapy-induced programmed cell death (PCD). In this study, we have quantified the (1)H NMR visible lipids in vivo and characterized their biophysical and biochemical nature in these tumors during PCD both ex vivo and in vitro. Concentrations of (1)H NMR-detectable PUFAs increased 3-fold with pattern recognition identifying CH = CH and CH = CHCH(2)CH = CH as the most significant in monitoring the dynamics of PCD. The increase in PUFAs was equivalent to 70% of that in CH(2)CH(2)CH(2)-saturated lipid peak at 1.3 ppm. Ex vivo tumor samples, obtained from in situ funnel frozen tumors, showed very similar macromolecular peaks, as studied using high-resolution magic angle spinning (1)H NMR at 14.1 T, to those detected in vivo at 4.7 T. Line widths of lipid peaks were not influenced by the spin rate within the range of 1-9 kHz or temperature between 277 and 293 K, showing high degree of (1)H NMR detection of these peaks in vivo. These biophysical results additionally corroborate the idea that cytoplasmic lipid vesicles are the source of (1)H NMR lipid signals. Two-dimensional (1)H NMR ex vivo and tumor lipid extracts in vitro showed that the PUFA signals are in the same chemical compounds and consist of largely 18:1 and 18:2 lipids. Furthermore, it is suggested that the (1)H NMR lipids detected during PCD arise from cell constituent breakdown products forming lipid vesicles into dying cells.