Magnesium and neoplasia: from carcinogenesis to tumor growth and progression or treatment.

Magnesium is involved in a wide range of biochemical reactions that are crucial to cell proliferation, differentiation, angiogenesis, and apoptosis. Changes in magnesium availability have been shown to influence biological responses of immuno-inflammatory cells. Equally plausible seems to be an involvement of magnesium in the multistep and interconnected processes that lead to tumor formation and development; however, the "how" and "when" of such an involvement remain to be defined. Here, we reviewed in vitro and in vivo data that indicated a role for magnesium in many biological and clinical aspects of cancer (from neoplastic transformation to tumor growth and progression or pharmacologic treatment). In adopting this approach we went through a full circle from molecular aspects to observational or epidemiological studies that could reconcile in a unifying picture the otherwise fragmentary or puzzling data currently available on the role of magnesium in cancer.

Vascular endothelial growth factor (VEGF) expression in noise-induced hearing loss.

Noise-induced hearing loss has been associated with alterations in cochlear blood flow. Our study analyzed the expression of Vascular Endothelial Growth Factor (VEGF) and its functional receptors, Flt-1 and Flk-1, in the cochlear structures of noise-exposed and unexposed guinea pigs. VEGF is a prototypical angiogenic agent, with multiple functions on vascular biology, ranging from vascular permeability to endothelial cell migration, proliferation, differentiation, and survival. Acoustic trauma was induced by a continuous pure tone of 6 kHz, at 120 dB SPL for 30 min. Auditory function was evaluated by electrocochleographic recordings at 2-20 kHz for 7 days. Noise-induced cochlear morphological changes were studied by immunohistochemistry and scanning electron microscopy. The expression of VEGF and its receptors was examined by immunohistochemistry and western blotting analysis. The hearing threshold shift reached a level of 60 dB SPL on day 1 after trauma and underwent a partial recovery over time, reaching a value of about 20 dB SPL on day 7. Outer hair cell loss was more prominent in the area located 14-16 mm from the apex. Increased cochlear VEGF expression was observed in noise-exposed animals, in particular at the level of stria vascularis, spiral ligament, and spiral ganglion cells. No changes were observed in the expression of VEGF-receptors. Our data suggest a role for VEGF in the regulation of the vascular network in the inner ear after acoustic trauma and during auditory recovery, with potentially important clinical and therapeutic implications.

Erythrocyte magnesium fluxes in mice with nutritionally and genetically low magnesium status.

Low intracellular magnesium (Mg) contents may be observed in case of severe Mg insufficient intake or because of genetic regulation. This work was conducted to investigate the influence of intracellular Mg content on erythrocyte Mg(2+) influx and efflux in mice with low nutritionally and genetically (MGL and MGH mice) Mg status. C57BL6 mice were fed for 2 wks a diet containing 1000 mg Mg/kg diet Mg (control group), 100 mg Mg/kg diet (Mg-marginal group) or 30 mg Mg/kg diet (Mg deficient group), while mice with low (MGL) and high (MGH) Mg levels were fed a control diet for 2 wks. The quantification of erythrocyte Mg(2+) influx and efflux was performed using a stable isotope of Mg. Our results showed that erythrocyte Mg(2+) influx and efflux were respectively increased and decreased in nutritional Mg deficiency; while in genetically determined Mg status Mg(2+) fluxes were lower in MGL mice compared to MGH mice. Moreover Mg(2+) efflux was significantly correlated to Mg level in erythrocytes in all the mice studied (p < 0.001). In conclusion, erythrocyte Mg(2+) influx and efflux are modulated by low Mg status, namely decreased Mg(2+) efflux compensate for nutritional Mg deficiency, while the genetic regulation of erythrocyte Mg(2+) content depends on modification of Mg(2+) influx.

Erythrocyte magnesium influx and efflux in solid tumor bearing mice.

Mg metabolism is modified in tumors and tumor-bearing organisms. In particular cancer patients often display elevated erythrocyte Mg levels. For a better understanding of the increased erythrocyte Mg content, we attempted to determine Mg fluxes in erythrocytes from tumor-bearing mice by Mg stable isotopes, using a method developed in our laboratory. To characterize the animal Mg status, blood and tissue Mg levels and hematological parameters were assayed. Results showed that in tumor-bearing mice total erythrocyte Mg was about 46% higher than in controls, whereas plasma and tissues Mg levels were not modified; red blood cells and hemoglobin as well as hematocrits were significantly decreased, while mean corpuscular volume and mean corpuscular hemoglobin were slightly but significantly increased in tumor-bearing mice compared to controls (by 3% and 4%, respectively), a picture corresponding to a normochromic, slightly macrocytic anemia. Erythrocyte Mg efflux was about 20% higher (404 + 59 versus 330 + 45 micromol/L, respectively, p < 0.05) in tumor-bearing mice compared to controls, whereas influx was not significantly modified (130 + 11 versus 122 + 19 micromol/L, respectively). Our data therefore exclude that the increased Mg content observed in erythrocytes of tumor-bearing mice is due to decrease of Mg efflux, or to an increase of Mg influx. On the other hand, the increased Mg content observed in erythrocytes of tumor-bearing mice could simply result from an increase of young Mg-enriched erythrocytes produced by the enhanced erythropoiesis which follows tumor-induced anemia.

Magnesium metabolism in mice selected for high and low erythrocyte magnesium levels.

A genetic control of blood magnesium (Mg) levels has been suggested. To investigate the mechanisms and the biologic significance of this genetic regulation, a mouse model, ie, mice selected for low magnesium level (MGL) and high magnesium level (MGH), was developed. The purpose of this study was to explore the Mg status and Mg metabolism in female MGL and MGH mice. We observed that MGL mice had reduced total and ionized plasma Mg, lower erythrocyte Mg, lower tibia, and kidney Mg levels. In contrast, total urinary Mg and (25)Mg levels were significantly higher in MGL mice. MGL mice had smaller total Mg exchangeable pool masses compared with MGH, and fractional transport rates of Mg (exchange constant) were different. In vitro (25)Mg enrichments in erythrocytes from MGL mice were significantly lower. Moreover, Mg efflux from erythrocytes was significantly higher in MGL. In conclusion, this work demonstrates that MGL mice present lower body stores of Mg than MGH mice and lower body Mg retention. This is confirmed at a cellular level by a lower enrichment of (25)Mg in erythrocytes. The lower retention of Mg by MGL erythrocyte in comparison to MGH appears to be partly due to a higher Mg efflux in MGL erythrocyte. It can be hypothesized that a genetic factor that modulates Na(+)/Mg(2+) exchanger activity may be important in the regulation of Mg metabolism. Further investigations on the mechanisms responsible for differences in Mg retention between MGL and MGH mice could contribute to a better understanding of the genetic regulation of cellular Mg.

DNA oxidative damage during differentiation of HL-60 human promyelocytic leukemia cells.

DNA oxidative damage was measured in human promyelocytic leukemia HL-60 cells, in the same cells committed to granulocytic differentiation with dimethyl sulfoxide (DMSO) or all-trans-retinoic acid (RA) and in mature human peripheral granulocytes (HPG). DNA damage was evaluated as single strand breaks and 8-OHdG adducts, measured by single cell electrophoresis or by monoclonal antibodies, respectively. The basal levels of either marker of DNA damage were higher in undifferentiated HL-60 cells than in HPG and DMSO- or RA-differentiated cells. Treatment with H(2)O(2) increased 8-OHdG formation in all cells, but the levels of DNA damage remained higher in undifferentiated cells as compared to the differentiated ones. Three lines of evidence suggested that the higher levels of DNA damage observed in undifferentiated cells were at least in part attributable to a reduced detoxification of reactive oxygen species (ROS). First, undifferentiated cells were shown to accumulate higher levels of dichlorodihydrofluorescein-detectable ROS than HPG and DMSO- or RA-differentiated cells. Second, undifferentiated HL-60 cells were characterized by reduced levels of GSH and lower GSH/GSSG ratios as compared to the differentiated cells. Third, pretreatment of undifferentiated HL-60 cells with antioxidants such as alpha-tocopherol or beta-carotene suppressed the elevation of ROS and the formation of 8-OHdG induced by H(2)O(2). Further evidence for the importance of the oxidant/antioxidant balance was obtained by modulating the iron-catalyzed decomposition of H(2)O(2) to hydroxyl radicals in undifferentiated HL-60 cells. In fact, pretreatment with FeSO(4) increased the formation of 8-OHdG induced by H(2)O(2), whereas pretreatment with the iron chelator deferoxamine produced the opposite effect. These results illustrate correlations between the oxidant/antioxidant balance and DNA damage and suggest that the capability of a cell population to withstand oxidative stress and DNA damage may depend on its degree of differentiation.

Resveratrol, a natural phenolic compound, inhibits cell proliferation and prevents oxidative DNA damage.

Resveratrol (3,4',5-trihydroxystilbene) is a naturally occurring phenolic compound which is present at high levels in wine and has been recently proposed as a potential cancer chemopreventive and chemoterapeutic agent. In this study, we evaluated the antiproliferative activity of resveratrol on a panel of cell lines of various histogenetic origin, including normal rat fibroblasts and mouse mammary epithelial cells compared to human breast, colon and prostate cancer cells. The concentration of resveratrol inhibiting cell growth by 50% (IC(50)) ranged from about 20 to 100 microM. At such concentration, we were unable to detect a significant increase in the apoptotic index in most of the cell lines analyzed. We also studied the effects of resveratrol on cell cycle distribution. The most striking effect was a reduction in the percentage of cells in the G2/M phase which was most frequently associated with an increase of cells in the S phase of the cell cycle. We also found that resveratrol is able to prevent the increase in reactive oxygen species (ROS) following exposure to oxidative agents (i.e. tobacco-smoke condensate (TAR) and H(2)O(2)). Resveratrol also reduced nuclear DNA fragmentation, as assessed by single cell gel electrophoresis (comet test). Taken together our results suggest that resveratrol can act as an antimutagenic/anticarcinogenic agent by preventing oxidative DNA damage which plays a pivotal role in the carcinogenic activity of many genotoxic agents.

Isolation of normal epithelial cells adapted to grow at nonphysiological concentration of magnesium.

Extracellular magnesium (Mg) depletion inhibits the growth of the HC11 normal mammary epithelial cells. In this study we found that an acute increase in extracellular Mg generally exerts a positive effect on the growth of these cells. We also isolated and characterized two derivatives adapted to grow and proliferate at nonphysiological concentration of Mg. The growth properties of the HC-LMg cells at 25 microM Mg were comparable to those of the parental HC11 cells in the regular medium (0.5 mM Mg) despite an increased expression of the CDK inhibitor p27(Kip1). They also showed a reduced dependence from serum to grow. The HC-HMg cells have been adapted to grow and proliferate at an increased (45 mM) Mg concentration. Cell total Mg content was 19.6, 9.7, and 20.1 nmol/mg protein in the HC11, HC-LMg, and HC-HMg cells, respectively. Thus, we have isolated derivatives of normal epithelial cells which are able to maintain Mg content in a physiological range in the face of different extracellular concentration gradients and will be a valuable tool for further studies on the regulation of Mg homeostasis in eukaryotic cells.

Effect of extracellular magnesium on topoisomerase II activity and expression in human leukemia HL-60 cells.

Topoisomerase II (TopoII) is a Mg-dependent enzyme involved in topological modifications of DNA that are crucial to the regulation of cell proliferation and possibly differentiation. To investigate the role of Mg availability in the modulation of TopoII in whole cells, we studied enzyme activity and expression in HL-60 cells grown in the presence of decreasing amounts of extracellular Mg (0.5, 0.03, and 0.01 mM MgSO(4)). In comparison to cells grown in 0.5 mM Mg, cells grown in 0.03 mM Mg exhibited a decrease in TopoII activity, as evidenced by reduced induction of DNA/TopoII cleavable complexes and apoptosis by etoposide and teniposide. Enzyme activity was restored by the readdition of Mg (0.5 and 1.5 mM) in the incubation medium, confirming that this effect was indeed modulated by extracellular Mg. Restriction of Mg to 0.01 mM was associated with a dramatic decrease in TopoII activity resembling that observed in HL-60 cells differentiated by dimethyl sulfoxide treatment. The restriction of Mg, while decreasing enzyme activity, was found to upregulate TopoII protein expression, determined by Western blot analysis. The increase of TopoII protein levels was correlative with the degree of Mg deprivation. Collectively, these results indicate that extracellular levels of Mg may control availability of intracellular Mg, thus affecting the regulation of TopoII activity/expression and downstream processes of cell proliferation and/or differentiation.

Magnesium in cell proliferation and differentiation.

Compelling evidence shows that magnesium (Mg) content is directly correlated to proliferation in normal cells as Mg stimulates DNA and protein synthesis. Some data have demonstrated that upon mitogenic stimuli normal cell are able to increase their intracellular Mg content, likely by activating Mg influx. Mg deprivation, in turn, induces inhibition of DNA and protein synthesis thus promoting growth arrest. From a mechanistic viewpoint, Mg deprivation may influence cell cycle control by upregulating the cyclin inhibitor p27Kip1 thus influencing cyclin E-dependent kinases. In many neoplastic cells, Mg is higher than in normal counterparts and this high Mg is maintained also against concentration gradient. Moderate vs. severe and acute vs chronic effect of Mg deprivation must be distinguished: severe Mg deprivation causes growth arrest also in tumor cells, while chronic Mg deprivation leads to an "adaptation" of tumor cells both to growth rate and Mg content. In tumor cells deranged Mg content and distribution is likely due to an inhibition of Mg efflux via the Na-Mg antiport. When differentiation process is induced by receptor mediated stimuli such as IFN-alpha and ATP, decrease of cell Mg content accompanies with activation of Mg efflux. Transformed cells may thus display high growth rate also because they retain a large amount of Mg. On their whole, these data strongly suggest that regulation of intracellular Mg availability parallels the molecular control of cell proliferation, and maybe also cell differentiation and death.

Magnesium depletion causes growth inhibition, reduced expression of cyclin D1, and increased expression of P27Kip1 in normal but not in transformed mammary epithelial cells.

In this study, we have evaluated the effects of extracellular magnesium restriction on the growth and cell cycle parameters of normal (HC11) and transformed (MCF-7) breast epithelial cell lines. Cells were incubated in medium with different concentrations of Mg2+ (from 0.5 to 0 mM) and the growth rates were determined by [3H]-thymidine incorporation and cell counting. The growth of the HC11 cells was drastically inhibited by Mg2+ depletion whereas the MCF-7 cells were only slightly inhibited (about 50% and 15%, respectively, after incubation in 0.05 mM Mg for 48 h). Cell cycle analyses showed a decrease in the percentage of cells in the S phase when both cell lines were incubated at low Mg2+ concentration. However, while the percentage of cells in both the G0/G1 and G2/M phases was increased in the HC11 cells, only the percentage of cells in the G2/M phase was increased in the MCF-7 cell line. Extracellular magnesium depletion was associated with increased expression of the cyclin-dependent kinase inhibitor p27Kip1 and decreased expression of cyclin D1 in the HC11 but not in the MCF-7 cells. We also demonstrated that Mg2+ depletion does not inhibit kinase activities in the normal HC11 cells and that Mg2+-restricted HC11 cells are still responsive to the epidermal growth factor (EGF)- and insulin-mediated stimulation of cell growth. These data suggest that normal but not transformed mammary epithelial cells are inhibited by extracellular Mg2+ restriction and that this effect might be mediated by changes in the levels of expression of both cyclin D1 and p27Kip1.

Changes in magnesium content and subcellular distribution during retinoic acid-induced differentiation of HL60 cells.

Magnesium (Mg) is required for cellular proliferation; however, the differences in subcellular regulation of Mg between proliferating and differentiated cells has not been determined. We used electron probe microanalysis (EPMA) to investigate the subcellular distribution of Mg in HL60 cells (a promyelocytic leukemia cell line) before and after retinoic acid (RA)-induced differentiation. Most intracellular Mg is bound to ATP and the Mg-ATP complex regulates several metabolic enzymes. We also compared alterations in Mg content following differentiation with the changes in ATP and ADP levels. Using atomic absorption spectrophotometry, we observed a significant decrease (-20%) in cellular Mg content in RA-differentiated HL60 cells. To investigate which intracellular compartments were involved in these changes, we analyzed subcellular elemental composition in freeze-dried cryosections of rapidly frozen undifferentiated and differentiated HL60 cells by EPMA. Following differentiation of HL60 cells, we observed an 18% decrease in Mg content in both the cytoplasm (regions of the cell excluding mitochondria and nuclei) and mitochondria. There was also a significant (40%) decrease in cytoplasmic Ca content after RA-induced differentiation. Nuclear Mg concentration was not significantly different between differentiated and undifferentiated HL60 cells, although differentiation was accompanied by a 30% decrease in the nuclear K/Na ratio. After differentiation, cellular ATP and ADP content decreased by 31 and 40%, respectively. We conclude that during exit from the cell cycle, Mg redistributes within cells and that the decrease in cytoplasmic and mitochondrial Mg is accompanied by a decrease in ATP and ADP content.

Differentiation of HL-60 promyelocytic leukemia cells is accompanied by a modification of magnesium homeostasis.

Magnesium homeostasis in HL-60 promyelocytic leukemia cells was compared to that in neutrophyl-like HL-60 cells obtained by 1.3% DMSO treatment. Magnesium homeostasis was studied by the characterization of magnesium efflux, the identification of intracellular magnesium pools, and the regulation of intracellular ionized Mg2+. In both undifferentiated and neutrophyl-like HL-60 cells, magnesium efflux occurred via the Na-Mg antiporter which was inhibited by imipramine and stimulated by db cAMP and forskolin. Receptor-mediated signals such as ATP, IFN-alpha, or PGE1, which can trigger cAMP-dependent magnesium efflux, were ineffective in undifferentiated HL-60 cells but induced 60-70% increase of magnesium efflux in neutrophyl-like HL-60 cells. Selective membrane permeabilization by the cation ionophore A23187 induced a large magnesium release when cells were treated with rotenone. In both cell populations, the addition of glucose to rotenone-treated cells restored magnesium release to the control level. Permeabilization by 0.005% digitonin provoked the release of 90% cell total magnesium in both cell types. Intracellular [Mg2+]i was 0.15 and 0.26 mM in undifferentiated and neutrophyl-like HL-60 cells, respectively. Stimuli that triggered magnesium efflux, such as db cAMP in undifferentiated and IFN-alpha in neutrophyl-like HL-60 cells, induced a slow but consistent increase of [Mg2+]i which was independent from Ca2+ movements. Overall, these data indicate that magnesium homeostasis is regulated by receptor-mediated magnesium efflux which was modified during differentiation of HL-60 cells. Stimulation of magnesium efflux is paralleled by an increase of [Mg2+]i which reflects a release of magnesium from the bound cation pool.

Magnesium restriction induces granulocytic differentiation and expression of p27Kip1 in human leukemic HL-60 cells.

When cultured in Mg restricted medium, human leukemic HL-60 cells develop morphological and functional granulocytic differentiation. In 0.03 mM Mg, cells display the distinctive features of differentiation, without appreciable inhibition of proliferation. In 0.01 mM Mg, cells show terminal differentiation, accompanied by clear inhibition of proliferation. Such cells accumulate in the G0/G1 phase and subsequently die via apoptosis, similar to HL-60 cells that have been induced to differentiate by DMSO. These phenotypic changes are associated with a marked increase in the expression level of the cyclin dependent kinase inhibitor p27Kip1. Cyclin E expression is also slightly increased in Mg restricted cells, whereas no changes are observed in the expression level of cyclin D1. We also show that during differentiation cell total Mg decreases, whereas [Mg2+]i increases in both Mg-depleted and DMSO-treated cells. These data suggest that the maturation process is paralleled by a redistribution of intracellular Mg, leading to a shift from the bound to the free form. These changes could modulate the kinetics of Mg-dependent enzyme(s) that are involved in the control of the differentiation pathway. We propose that this model may represent an useful tool for the study of the mechanisms of cell differentiation and related events, such as aging and death.

Regulation of magnesium efflux from rat spleen lymphocytes.

Rat spleen lymphocytes (RSL) incubated at 37 degrees C in Mg-free medium (O-trans conditions) exibited Mg2+ efflux with apparent velocity of 0.2 nmol/mg protein/min. After 30 min, this process accounted for the mobilization of about 15% of cell total Mg2+. Half of the Mg2+ efflux depended on extracellular Na+ and was stimulated by cAMP. IFN-alpha significantly enhanced Mg2+ efflux under O-trans conditions as well as in the presence of physiological extracellular Mg2+. Pretreatment of RSL with indomethacin completely abolished IFN-alpha-induced Mg2+ efflux, suggesting a crucial role for cyclooxygenase-dependent arachidonate metabolism. On the other hand, pretreatment of RSL with the PKA inhibitor (Rp)8-Br-cAMPS prevented IFN-alpha stimulation of Mg2+ efflux, indicating the involvement of cAMP. Consistently, both IFN-alpha and exogenous PGE1 increased cAMP from 50 to 125 pmol/mg protein. Altogether these results show that IFN-alpha stimulates Mg2+ efflux by activating arachidonate metabolism and synthesis of prostaglandins. By influencing adenylcyclase activity, PGEs can eventually promote cAMP-dependent Mg2+ efflux, possibly through the activity of a Na-Mg antiport. In RSL, therefore, magnesium movements can be under the control of IFN-alpha and, perhaps, of other cytokines, suggesting the involvement of Mg2+ in cell response to receptor-mediated stimuli.

Regulation of intracellular magnesium in ascites cells: involvement of different regulatory pathways.

Extracellular ATP causes 40% stimulation of Mg2+ efflux from Ehrlich ascites tumor cells (EATC) incubated under 0-trans conditions. ATP also causes a threefold increase of arachidonic acid (AA) metabolite release from [3H]AA-preloaded EATC, indicating that, under these experimental conditions, it induces phospholipase A2 (PLA2) activation. ATP-induced Mg2+ efflux can be prevented by cyclooxygenase inhibition with indomethacin or lysine acetylsalicylate, but not by 5-lipooxygenase inhibition with BWA4C. Mg2+ efflux is also directly stimulated by exogenous AA in a concentration-dependent manner. This phenomenon involves PKA as it is virtually abolished by the specific inhibitor 8-bromoadenosine-3',5'-cyclic monophosphothioate. While stimulating Mg2+ efflux, exogenous AA also increases cAMP content of EATC and this effect can be prevented by cyclooxygenase inhibition. Measurements in mag-fura-2-loaded EATC reveal that stimulation of Mg2+ efflux does not correlate with significant fluctuations of [Mg2+]i. This suggests that Mg2+ efflux is compensated for by mobilization of bound Mg2+, leaving [Mg2+]i unaltered. Altogether these data indicate that Mg2+ efflux can be modulated by extracellular stimuli capable of activating PLA2. This modulation is triggered by cyclooxygenase products which, by activating adenylcyclase, determine an elevation of cAMP. This intracellular messenger upregulates Na-dependent Mg2+ efflux.

cAMP activates magnesium efflux via the Na/Mg antiporter in ascites cells.

In intact ascites cells magnesium efflux is substantially stimulated by cAMP analogues and by low concentrations of prostaglandins (PGE1 and PGE2) which increases the synthesis of cAMP. cAMP stimulates magnesium efflux by directly stimulating the Na-Mg antiporter rather than determining a magnesium release from mitochondria as reported for other cell types. Protein kinase inhibitors abolish the cAMP stimulated magnesium efflux suggesting that the stimulation is mediated by the phosphorylation of the carrier protein, increasing its affinity for intracellular magnesium. These data provide evidence that receptor-mediated stimuli eventually leading to cell activation are directly connected to the regulation of intracellular magnesium.

Characterization of magnesium efflux from Ehrlich ascites tumor cells.

Magnesium efflux from intact Ehrlich ascites tumor cells (EATC) has been characterized under 0-trans conditions. It is shown that a magnesium-extruding mechanism operates in these cells which brings about magnesium efflux up to 20% of the cell total content. EATC magnesium efflux is independent from extracellular Ca2+ and its apparent velocity is not affected by [Mg2+]o up to 160 microM. This extrusion, however, is strictly dependent on extracellular Na+ and it is inhibited by ouabain (1 mM), amiloride (1 mM), imipramine (0.5 mM) and quinidine (1 mM). It is not affected by HMA (5-N,N-hexamethylene) amiloride) (0.5 microM) and vanadate (0.1 mM). Bumetanide (0.5 mM) enhances magnesium extrusion in these cells. EATC magnesium extrusion can be significantly stimulated by db cAMP, forskolin, and IBX (3-isobutyl-1-methyl-xanthine). The intracellular magnesium distribution, studied also by means of the ionophore A23187, is regulated by energy metabolism and cell ATP content. Altogether our data demonstrate that EATC exhibit a magnesium extrusion sustained by Na+ gradient across the plasma membrane and carried out by a Na+/Mg2+ antiport. In these cells magnesium homoeostasis results, regulated efficiently by cell ATP and cAMP content.

The effect of magnesium on glycolysis of permeabilized Ehrlich ascites tumor cells.

We have previously observed that extracellular Mg2+ influences the phosphofructokinase (PFK) activity of intact Ehrlich Ascites tumour cells (EATC). In this study we have investigated the mechanism by which Mg2+ modulates this key glycolytic enzyme in EATC made permeable to the cation by either digitonin or dextran sulphate. Results showed that when Mg2+ is freely permeable to the cytosol, the in vivo PFK activity, calculated as FDP/G6P ratio, is not increased as it is in intact cells. We also observed that in permeabilized cells Mg2+ determines the increase of glucose 6 phosphate (G6P), fructose 1,6 bisphosphate (FDP) and lactate production. We hypothesize that extracellular Mg2+ regulates PFK and glycolysis in these neoplastic cells not by entering the cytosol but by a specific interaction with the plasma membrane.

Magnesium in normal and neoplastic cell proliferation: state of the art on in vitro data.

Information about the involvement of Mg2+ in all biochemical processes that participate in cell proliferation is reviewed in order to define the role of this divalent cation in normal and pathological growth. The lack of conclusive data about cell Mg2+ homeostasis does not suggest any definitive model for its role in the control of cell proliferation. On the other hand, new important information about its absolute requirement in crucial steps of cell activation that can, beside other functions, trigger cell division, strongly support the involvement of Mg2+ in the control of cell proliferation. Studies on the growth of cells in vitro, however, while confirming the indispensible requirement for Mg2+ in extracellular media, do not completely clarify the mechanism(s) or the exact phase/point of the cell cycle where Mg2+ exerts its regulation. Furthermore, the observation that tumour cells grown in culture are influenced by external divalent cations confirms the involvement of Mg2+ in cancer as well as in normal cell proliferation. The proposed explanations (theories, hypotheses) are described and discussed.