Ignacio Núñez de Castro

Antioxidant Enzymes and Human Diseases

OBJECTIVES To describe the importance of the antioxidant enzymes superoxide dismutase, glutathione peroxidase, and catalase working together in human cells against toxic reactive oxygen species, their relationship with several pathophysiologic processes and their possible therapeutic implications. CONCLUSIONS Reactive oxygen species (ROS) are involved in the cell growth, differentiation, progression, and death. Low concentrations of ROS may be beneficial or even indispensable in processes such as intracellular signaling and defense against micro-organisms. Nevertheless, higher amounts of ROS play a role in the aging process as well as in a number of human disease states, including cancer, ischemia, and failures in immunity and endocrine functions. As a safeguard against the accumulation of ROS, several nonenzymatic and enzymatic antioxidant activities exist. Therefore, when oxidative stress arises as a consequence of a pathologic event, a defense system promotes the regulation and expression of these enzymes.

Glutamine and its relationship with intracellular redox status, oxidative stress and cell proliferation/death

Glutamine is a multifaceted amino acid used for hepatic urea synthesis, renal ammoniagenesis, gluconeogenesis in both liver and kidney, and as a major respiratory fuel for many cells. Decreased glutamine concentrations are found during catabolic stress and are related to susceptibility to infections. Besides, glutamine is not only an important energy source in mitochondria, but is also a precursor of the brain neurotransmitter glutamate, which is likewise used for biosynthesis of the cellular antioxidant glutathione. Reactive oxygen species, such as superoxide anions and hydrogen peroxide, function as intracellular second messengers activating, among others, apoptosis, whereas glutamine is an apoptosis suppressor. In fact, it could contribute to block apoptosis induced by exogenous agents or by intracellular stimuli. In conclusion, this article shows evidences for the important role of glutamine in the regulation of the cellular redox balance, including brain oxidative metabolism, apoptosis and tumour cell proliferation.

Antiproliferative effect of dehydrodidemnin B (DDB), a depsipeptide isolated from Mediterranean tunicates

The biological effects of dehydrodidemnin B(DDB), a novel depsipeptide isolated from Aplidium albicans, were studied on Ehrlich carcinoma growing in vivo and in primary cultures, and compared with those reported for Didemnin B (DB). Daily administration of DB or DDB (2.5 micrograms/mouse) almost duplicated the animal life-span and total number of tumour cells decreased by 70-90%. Results suggest a major effect of DDB when administered in the lag phase of growth. DDB behaved as a very potent inhibitor of protein synthesis; consequently, ornithine decarboxylase activity (ODC, EC 4.1.1.17) is drastically reduced by DDB-treatment.

Histamine, polyamines, and cancer

Mammalian ornithine decarboxylase and histidine decarboxylase present common structural and functional features, and their products also share pharmacological and physiological properties. Although accumulated evidence pointed for years to a direct involvement of polyamines and histamine in tumour growth, it has been only in the last few years that new molecular data have contributed to the clarification of this topic. The aim of this commentary is to review the molecular grounds of the role of histamine and polyamines in cancer and to point to possible directions for future research in emerging areas of interest.

Nitrogen Metabolism in Tumor Bearing Mice

In experiments with whole animals infested with a highly malignant strain of Ehrlich ascites tumor cells, serial concentrations of amino acids were determined for host plasma, ascitic fluid, and tumor cells, throughout tumor development. Concentration gradients of glutamine, asparagine, valine, leucine, isoleucine, phenylalanine, tyrosine, histidine, tryptophan, arginine, serine, methionine, and taurine from the host plasma toward the ascitic liquid were established; while on the other hand, concentration gradients from the ascitic liquid toward the plasma were established for glutamate, aspartate, glycine, alanine, proline, and threonine. With the exception of aspartate the concentrations of these amino acids were highest inside the cells. Arginine was the only amino acid not detected in tumor cells. In vitro incubations of tumor cells in the presence of glutamine and/or glucose, as the energy and nitrogen sources, confirmed the amino acid fluxes previously deduced from the observed relative concentrations of amino acids in plasma, ascitic liquid, and tumor cells, suggesting that glutamate, alanine, aspartate, glycine, and serine can be produced by tumors. These findings support that changes in amino acid patterns occurring in the host system are related to tumor development.

Phosphate-activated glutaminase expression during tumor development

Changes in phosphate‐activated glutaminase activities determined in intact cells and isolated mitochondria have been followed during mouse Ehrlich ascites carcinoma development. Glutaminase activities parallel the levels of poly(A)+ RNAs encoding for the mitochondrial phosphate activated glutaminase. During the exponential growth phase, maximum activity was observed and the relative abundance of glutaminase mRNA significantly increased with regard to the stationary growth phase. The presented results show that tumor phosphate‐activated glutaminase is subject to long‐term regulation by differential gene expression.

Dual Role of Plasma Membrane Electron Transport Systems in Defense

Bcause oxidative stress is one of the main sources of severe cellular damage, cells have different defense weapons against reactive oxygen species. Ubiquitous plasma membrane redox systems play a role in defense against oxidative stress damage. On the other hand, a tightly controlled and localized production of reactive oxygen species by a plasma membrane NADPH oxidase can be used as a potent microbicidal weapon. This dual, prooxidant and antioxidant role of plasma membrane electron transport systems in defense is studied and discussed.

Characterization of tomato leaf glutamine synthetase

Abstract Glutamine synthetase was purified to apparent homogeneity from the leaves of light-grown tomato plants. The purification steps involved ammonium sulphate precipitation (40–60%), adsorption by hydroxyapatite, DEAE Sephadex ionic exchange chromatography, and Sephacryl S 300 gel filtration. Only one glutamine synthetase form was found, with Km-values of 5.5 mM, 0.8 mM and 0.4 mM for glutamate, ATP and ammonium, respectively. The in vitro activity depended on Mg2+ or Mn2+ concentrations with positive cooperativity. Enzyme assay pH optima were 7.2 for synthetase, and 6.0 for transferase. The activation energy of the enzymatic reaction was 33.2 kJ/mol. ADP appeared to be a competitive inhibitor with a Ki-value of 0.4 mM. Enzyme inhibition was also produced by aspartate, alanine and phosphoserine.

Transmembrane ferricyanide reductase activity in Ehrlich ascites tumor cells.

A transmembrane ferricyanide reductase activity was assayed in intact Ehrlich ascites tumor cells. Kinetic measurements gave a Km of 0.14 mM and a Vmax of 0.31 mumol/min per 10(6) cells. In short-term batch experiments, this activity was enhanced in the presence of 10 mM lactate, a source of cytosolic NADH. The transmembrane redox activity was accompanied by alkalinization of the cytosol. Both ferricyanide reduction and proton extrusion were diminished in the presence of 0.2 mM amiloride. Several cytotoxic drugs significantly inhibited the ferricyanide reductase activity at concentrations at which they show antineoplastic activity.

Interchange of amino acids between tumor and host

During the growth of a tumor, there are very relevant changes in the metabolism of the host to produce the metabolites rapidly consumed by the tumor. In this context, the exchanges of amino acids between the tumor and its host are especially important; however, they have received little attention. A rigorous study must provide data on the growth curve of the tumor, as well as on amino acid levels in tumor cells, plasma, and metabolically relevant tissues and organs from the host during the whole growth of the tumor. The main conclusions arising from a complete study in a tumor model are discussed.