Effect of glucose and deoxyglucose on the redistribution of calcium in Ehrlich ascites tumour and Zajdela hepatoma cells and its consequences for mitochondrial energetics

Abstract

The distribution of Ca 2+ in intact cells was monitored with fluorescent probes: fura-2 for cytosolic [Ca 2+ ] and rhod-2 for mitochondrial [Ca 2+ ]. It was found that in neoplastic cells, such as Ehrlich ascites tumour and Zajdela hepatoma, but not in non-malignant cells, such as fibroblasts, glucose and deoxyglucose elicited release of Ca 2+ from endoplasmic reticulum stores and an increase in Ca 2+ concentration in the cytosol. Parallel to this, a decrease in the rate of Ca 2+ extrusion from the cell and an enhanced uptake of Ca 2+ by mitochondria were observed. The increase in mitochondrial [Ca 2+ ] was accompanied by an increase in the mitochondrial membrane potential and the reduction state of nicotinamide nucleotides. F 1 F o -ATPase in submitochondrial particles of Zajdela hepatoma was strongly inhibited in the presence of micromolar Ca 2+ concentrations, whereas this activity in submitochondrial particles from rat liver appeared to be less sensitive to Ca 2+ . Indications of glycosylation of Ehrlich ascites tumour cell proteins were also obtained. These data strengthen the proposal [Bogucka, K., Teplova, V.V., Wojtczak, L. and Evtodienko, Y. V. (1995) Biochim. Biophys. Acta 1228, 261±266] that the Crabtree effect is produced by mobilization of cell calcium, which is subsequently taken up by mitochondria and inhibits F 1 F o -ATP synthase.Keywords: Calcium; Crabtree effect; Ehrlich ascites tumour; mitochondrial membrane potential; Zajdela hepatoma.Many kinds of tumour exhibit an unusual reaction towards glucose: addition of millimolar concentrations of glucose to the culture medium results in a decrease in the cell respiration rate. This behaviour, known by the name of its discoverer as the Crabtree effect [1], has also been observed in some non-malignant cells and tissues, such as spermatozoa [2,3], proliferating thymocytes [4], intestinal mucosa [5] and mammalian embryos at their very early developmental stage [6]. Common features of all these tissues, both malignant and normal, are a high proliferation rate and/or a high glycolytic rate. Although the Crabtree effect has been known for seven decades (for an early review see [7]), its molecular mechanism is not fully understood. First attempts to explain it pointed to the competition between respiration and glycolysis for precursors of ATP production, i.e. ADP [8,9] and inorganic phosphate [10±12]. However, this explanation is not satisfactory because deoxyglucose, which is phosphorylated in the cytoplasm but not further metabolized and therefore does not generate ATP, produced even stronger inhibition of respiration than did glucose [13] and phosphate transport into the cell has been shown to be fast enough [13].Other possible explanations of the mechanisms of the Crabtree effect were a shift in intracellular pH [7], a change in permeability properties of the inner mitochondrial membrane [14], a specific isoenzyme pattern of the glycolytic pathway and regulatory behaviour of key enzymes of this pathway [15], and specific topography of enzymes in rap...

The Warburg effect and mitochondrial stability in cancer cells

“…Various mechanisms have been suggested to explain the Crabtree effect in tumor cells: (a) competition between oxidative phosphorylation and glycolysis for ADP and inorganic phosphate (Sussman et al, 1980); (b) a decrease in the cytosolic pH, as a consequence of lactic acid formation, that diminishes the activity of oxidative enzymes (Heinz et al, 1981); (c) damage of the mitochondrial membranes by free radicals, produced as a consequence of glucose catabolism (Yang et al, 1997b); and (d) glucose-induced release of Ca 2+ from the endoplasmic reticulum followed by enhanced uptake of Ca 2+ by the mitochondria, which would inhibit ATP synthase (Wojtczak et al, 1999). The latter explanation would be in agreement with the observed elevation of the Ca 2+ content in tumor mitochondria.…”

Mitochondrial Oxidative Phosphorylation Is Regulated by Fructose 1,6-Bisphosphate

“…In Ehrlich ascites tumors and in Zajdela hepatoma cells, it has been observed that there is a glucose-induced increase in cytoplasmic calcium levels along with an enhanced mitochondrial uptake of this cation. Inside the mitochondria, Ca 2ϩ would inhibit ATP synthase by enhancing the interaction with IF1, its inhibitory subunit (16). However, is not clear whether this Ca 2ϩ accumulation is a common event in all Crabtree-positive cells, since in AS-D30 hepatoma cells, cal-cium levels are not modified after glucose addition (7).…”

Mitochondria Permeabilization by a Novel Polycation Peptide BTM-P1

“…They are a principal energy source in these cells (1) and also are responsible for some mechanisms of apoptosis and necrosis (2-6) as well as for ischemic damage of tissues (4,(7)(8)(9)(10). Mitochondrial energy metabolism is very specific for cancer cells, which are characterized by the Crabtree effect (11)(12)(13)(14), by a very high hexokinase activity of mitochondria (15,16), and by high resistance to permeabilization of mitochondrial membranes by various pro-apoptotic proteins (2,(17)(18)(19)(20). Nevertheless some peptides have been found to be toxic for cancer cells (21)(22)(23)(24)(25)(26)(27)(28).…”