Metabolic changes in undifferentiated and differentiated human colon adenocarcinoma cells studied by multinuclear magnetic resonance spectroscopy.
Aspects of energetic and intermediary metabolism were studied in a colon adenocarcinoma cell line (HT29) by multinuclear magnetic resonance spectroscopy. Experiments were carried out on the HT29-D4 clone, which was isolated by limit dilution techniques. This clone, usually undifferentiated (D4-UD), can be maintained in a differentiated state (D4-D) in a glucose-free medium. Metabolic data were obtained by NMR analysis of perchloric acid extracts from D4-UD and D4-D cells. Phosphorus-31 and proton NMR spectra showed the presence of a large amount of choline and phosphorylcholine in the differentiated state (400% and 200%, respectively, of the levels found in D4-UD cells). Other differences appeared in the content of phosphocreatine (absent in D4-D cells) and myoinositol (absent in D4-UD cells). Carbon-13 spectra were recorded from perchloric acid extracts of cells incubated with [1-13C]-labeled glucose or [2-13C]-labeled acetate. The data indicated that both types of cells metabolize glucose through the glycolytic pathway to give lactate, but only D4-D cells were able to store glucose as glycogen at a very high level. A mathematical analysis of fluxes through the tricarboxylic acid (TCA) cycle was developed on the basis of models derived from previous 14C tracer studies. The model was based on the steady-state labeling of glutamate carbons by the 13C isotope and gave the fraction of labeled acetyl-Coa entering the TCA cycle, and the activity y of anaplerotic reactions relative to the flux through the citrate synthetase reaction. The data indicated that y greater than 0.3 in all cases. Only 15% and 30% of labeled acetyl CoA entered the TCA cycle in D4-UD and D4-D cells, respectively, under labeled glucose incubation: these values were significantly different upon labeled acetate feeding, reaching 55% for D4-UD cells and 85% for D4-D cells. The main result of this study is that the process of differentiation of HT29 cells is correlated with a large increase in the activity of oxidative metabolism.
Galons
J P
JP
Centre de Résonance Magnétique Biologique et Médicale, URA CNRS, Faculté de Médecine, Marseille, France.
Fantini
J
J
Vion-Dury
J
J
Cozzone
P J
PJ
Canioni
P
P
D000085:Acetates / Q000378:metabolism; D000230:Adenocarcinoma / Q000378:metabolism* / Q000473:pathology; D002247:Carbon Isotopes; D002454:Cell Differentiation; D003110:Colonic Neoplasms / Q000378:metabolism* / Q000473:pathology; D004734:Energy Metabolism; D005947:Glucose / Q000378:metabolism; D006801:Humans; D007294:Inositol / Q000032:analysis; D009682:Magnetic Resonance Spectroscopy / Q000379:methods*; D010759:Phosphorus Isotopes; D010766:Phosphorylation; D011522:Protons; D014407:Tumor Cells, Cultured
D000085:Acetates; D002247:Carbon Isotopes; D010759:Phosphorus Isotopes; D011522:Protons; D007294:Inositol; D005947:Glucose