Protocols and Applications of Cellular Metabolomics in Safety Studies Using Precision-Cut Tissue Slices and Carbon 13 NMR
Numerous xenobiotics are toxic to human and animal cells by interacting with their metabolism, but the precise metabolic step affected and the biochemical mechanism behind such a toxicity remain often unknown. In an attempt to reduce the ignorance in this field, we have developed a new approach called cellular metabolomics. This approach, developed in vitro, provides a panoramic view not only of the pathways involved in the metabolism of physiological substrates of any normal or pathological human or animal cell but also of the beneficial and adverse effects of xenobiotics on these metabolic pathways. Unlike many cell lines, precision-cut tissue slices, for which there is a renewed interest, remain metabolically differentiated for at least 24–48 h and allow to study the effect of xenobiotics during short-term and long-term incubations. Cellular metabolomics (or metabolic flux analysis), which combines enzymatic and carbon 13 NMR measurements with mathematical modeling of metabolic pathways, is illustrated in this brief chapter for studying the effect of insulin on glucose metabolism in rat liver precision-cut slices and of valproate on glutamine metabolism in human renal cortical precision-cut slices. The use of very small amounts of test compounds allows to predict their toxic effect and eventually their beneficial effects very early in the research and development processes. Cellular metabolomics is complementary to other omics approaches, but, unlike them, provides functional, mechanistic, and dynamic pieces of information by measuring enzymatic fluxes.
Key wordsToxicology Liver Kidney Cellular metabolomics Carbon 13 NMR
The authors would like to thank Claudie Pinteur, Rémi Nazaret, and Lara Koneckny for technical assistance as well as Claire Morel for secretarial assistance. This work was supported by grants from the European Commission [project numbers: BIO4-CT97-2145 (Euroslice) and STREP 032731 (CellNanoTox)], and from INSERM.
- 1.Warburg O (1923) Versuche an uberiebendem Carcirnomgewebe. Biochem Z 142:317–333Google Scholar
- 5.Bach P, Lock E (1985) The use of renal tissue slices, perfusion and infusion techniques to assess nephrotoxicity related changes. In: Bach PH, Lock EA (eds) Nephrotoxicity assessment and pathogenesis, Monographs of applied toxicology, vol 1. Wiley, New York, pp 505–518Google Scholar
- 7.Bach PH, Vickers AEM, Fisher R et al (1996) The use of tissue slices for pharmacotoxicology studies. ATLA 24:893–923Google Scholar
- 14.Lamprecht W, Trautchold I (1974) Adenosine-5′-triphosphate. Determination with hexokinase and glucose-6-phosphate dehydrogenase. In: Bergmeyer H (ed) Methods of enzymatic analysis, vol 4. Academic Press, New York, pp 2101–2110Google Scholar
- 19.Shaka AJ, Keeler J, Frenkiel T, Freeman R (1983) An improved sequence for broadband decoupling: Waltz 16. J Magn Reson 52:335–338Google Scholar
- 31.Warter JM, Marescaux C, Chabrier G, Rumbach L, Micheletti B, Imler M (1984) Renal glutamine metabolism in man during treatment with sodium valproate. Rev Neurol (Paris) 140:370–371Google Scholar