In Vivo, Ex Vivo, and In Vitro One‐ and Two‐Dimensional Nuclear Magnetic Resonance Spectroscopy of an Intracerebral Glioma in Rat Brain: Assignment of Resonances

Abstract

An in vivo study of intracerebral rat glioma using proton-localized NMR spectroscopy showed important modifications of the spectra in the tumor as compared with the contralateral brain. To carry out the assignment of the resonances of the glioma spectra, tumoral and normal rat brain tissues were studied in vivo, ex vivo, and in vitro by one-dimensional and two-dimensional proton spectroscopy. N-Acetylaspartate was found at an extremely low level in the glioma. The change of peak ratio total creatinel 3.2 ppm peak was found to be due to a simultaneous decrease of the total creatine content and an increase of the 3.2 ppm peak. The 3.2 ppm resonance in the glioma spectra has been shown to originate from choline, phosphocholine, glycerophosphocholine, taurine, inositol, and phosphoethanolamine. The increase of the 3.2 ppm peak in the glioma was found to result from the increase of taurine and phosphoethanolamine contents. The peak in the 1.3 ppm region of the glioma spectra was due to both lactate and mobile fatty acids. Moreover, two-dimensional spectroscopy of excised tissues and extracts showed the presence of hypotaurine only in the tumor.

An investigation of human brain tumour lipids by high‐resolution magic angle spinning 1H MRS and histological analysis

“…5). This supports the view that the in vivo 1 H MRS-visible lipid signals in human brain tumours arise from cytoplasmic lipid droplets, as observed in studies of C6 glioma cells and rodent brain tumours (5)(6)(7)(8), and are not just residual lipids from cell membrane breakdown products. A study on endothelial cells exposed to hypoxia showed an increase in lipid body formation to be dependent on O 2 concentration (26).…”

“…1, we give an example of a biopsy specimen from a grade II tumour where there are lipid droplet clusters suggestive of possible islands of hypoxia forming among regions of viable cells, and it is possible that such clusters give rise to the lactate signal often observed in vivo by 1 H MRS (2). This is unlike the development of the implanted C6 rodent tumours, where a core of necrosis develops, with a ring of severely hypoxic cells within the bulk of the tumour (8,10). In contrast, human astrocytomas, by their invasive nature and co-option of existing vasculature, develop in a more heterogeneous manner, with greater intermingling of apoptosis, necrosis, hypoxia, and areas of viable tumour cells.…”

“…Ex vivo studies have further supported this, showing not only a correlation between NMR-visible mobile lipids and percentage of necrosis, but also the presence of mobile lipids in tumours with no necrosis (4). NMR-visible mobile lipids have also been detected by 1 H MRS in cultured brain tumour cells (5)(6)(7), animal brain tumours (8)(9)(10)(11), and human brain tumour biopsy specimens (12). However, the exact metabolic origin and localisation of these lipids has been vigorously debated.…”

“…However, most subsequent studies have shown evidence of the formation of lipid droplets in the cytosol of intact glioma cells (5)(6)(7) or their extracellular presence in the necrotic core of experimental tumours (8)(9)(10). These 1 H MRS lipid resonances are believed to originate from the mobile fatty acyl chains, mainly of TGs, with lesser contributions from free fatty acids and cholesteryl esters (14).…”

“…As previous studies were carried out on C6 glioma cells in vitro, or on tumours grown in rodent brain (5)(6)(7)(8)(9)(10), there is still the need to validate a similar interpretation for the lipid signals of human brain tumours observed by 1 H MRS. We therefore performed high-resolution magic angle spinning (HRMAS) 1 H MRS on whole-biopsy samples obtained from patients with low-and high-grade astrocytomas, to evaluate their NMR-visible lipid content. By using HRMAS, we can obtain high-resolution 1 H spectra from whole-biopsy tissue that can be easily and directly related to in vivo 1 H MRS data (16), but without the confounding factors associated with performing a biochemical extraction of lipids for their assessment.…”