María L. Tomaro

Cadmium toxicity in plants

Heavy metals are important environmental pollutants and their toxicity is a problem of increasing significance for ecological, evolutionary, nutritional, and environmental reasons. Plants posses homeostatic cellular mechanisms to regulate the concentration of metal ions inside the cell to minimize the potential damage that could result from the exposure to nonessential metal ions. This paper summarizes present knowledge in the field of higher plant responses to cadmium, an important environmental pollutant. Knowledge concerning metal toxicity, including mechanisms of cadmium homeostasis, uptake, transport and accumulation are evaluated. The role of the cell wall, the plasma membrane and the mycorrhizas, as the main barriers against cadmium entrance to the cell, as well as some aspects related to phytochelatin-based sequestration and compartmentalization processes are also reviewed. Cadmium-induced oxidative stress was also considered as one of the most studied topics of cadmium toxicity.

Effect of heavy metal ion excess on sunflower leaves: evidence for involvement of oxidative stress

Abstract The relationship between heavy metal ion toxicity and oxidative stress in plant cells was studied. Leaf segments from 14 day old sunflower seedlings were incubated in solutions containing 0.5 mM Fe(II), Cu(II) or Cd(II) ions for 12 h in the light. Treatment with metal ions studied produced a decrease in chlorophyll and GSH contents as well as increases in lipid peroxidation and lipoxygenase activity. Free radical scavengers, such as sodium benzoate and mannitol, prevented the decrease in chlorophyll and GSH content and the lipid peroxidation and lipoxygenase increases. While Fe(II) and Cd(II) ions caused a decrease in superoxide dismutase activity, Cu(II) ions raised its level. However, all three metal ions caused decreases in other antioxidant enzymes (catalase, ascorbate peroxidase, glutathione reductase and dehydroascorbate reductase). Free radical scavengers protected these enzymes against inactivation. No effect of these scavengers was observed on superoxide dismutase activity. These results indicate that excess Fe(II), Cu(II) or Cd(II) ions produce oxidative damage in plant leaves.

Heme oxygenase and oxidative stress. Evidence of involvement of bilirubin as physiological protector against oxidative damage

Cobalt chloride (CoCl2), a well-known inducer of heme oxygenase, produced a strong increase of in vivo rat liver chemiluminescence (QLV) 6 h after its administration. The activity of antioxidant enzymes, superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) was found to be significantly decreased 9 h after CoCl2 injection. Heme oxygenase activity increased 9 h after treatment, reaching a maximum value around 18 to 24 h after CoCl2 administration. This induction was preceded by a decrease in the intrahepatic GSH pool and an increase in hydrogen peroxide steady state concentration, both effects taking place several hours before induction of the heme-oxygenase. Co-administration of Sn-protoporphyrin IX, a potent inhibitor of heme oxygenase, completely prevented the enzyme induction, increasing the QLV levels. Administration of bilirubin, the end product of heme catabolism in mammals, prevented the heme oxygenase induction as well as the decrease in hepatic GSH and the increase of chemiluminescence when it was administered 2 h before CoCl2 treatment. These results support the proposal that the induction of heme oxygenase by cobalt chloride may be a general response to oxidant stress and, by increasing bilirubin levels, could constitute an important cellular defense mechanism against oxidative damage.

Polyamines as protectors against cadmium or copper-induced oxidative damage in sunflower leaf discs

Abstract Polyamines antioxidant properties were studied in sunflower leaf discs under cadmium and copper induced oxidative stress. Both metals induced oxidative damage in the studied tissue, as evidenced by an increment in lipid peroxidation and a decrease in antioxidant enzymes ascorbate peroxidase (APOX), glutathione reductase (GR) and superoxide dismutase (SOD) activities. Polyamines concentrations were differently affected according to the metal used. Putrescine (Put) was reduced 52% by Cd 2+ and 39% by Cu 2+ , spermidine (Spd) was decreased to 21 and 59% of the control by Cd 2+ and Cu 2+ , respectively and spermine (Spm) content was not modified by any of the metals. Pretreatment with exogenously added polyamines (1 mM) showed that Spm reverted the effect of Cd 2+ and Cu 2+ on lipid peroxidation almost to control values. Neither Spm nor Spd recovered the metal-reduced APOX activity. Besides, both polyamines affected this parameter by themselves. GR activity was completely restored by Spm or Spd and only Spm was effective in reverting copper-reduced SOD activity. This work suggests that polyamines are undoubtedly related to the protection against metal-induced oxidative stress, but more research is necessary to elucidate the precise role they played as antioxidants.

Bilirubin : Its role in cytoprotection against oxidative stress

Bilirubin, the end product of heme catabolism in mammals, is generally regarded as a potentially cytotoxic, lipid-soluble waste product that needs to be excreted. However, in the last 10 years, in vitro and in vivo studies, have demonstrated that bilirubin exhibits potent anti-oxidant properties preventing the oxidative damage triggered by a wide range of oxidant-related stimuli. Therefore, the idea of a beneficial and physiological role for bilirubin in cytoprotection against short and long-lasting oxidant-mediated cell injury is highlighted here.

Polyamine metabolism in sunflower and wheat leaf discs under cadmium or copper stress

Abstract The influence of Cd 2+ or Cu 2+ on polyamine (Pa) metabolism and ethylene evolution was studied in sunflower and wheat leaf-discs. Both metals increased putrescine (Put) content in wheat leaves 280 and 90%, respectively, but reduced this Pa in sunflower leaves by about 50% of the control values. Spermidine content was diminished in cadmium and copper sunflower treated leaves (41 and 79%, respectively) while spermine was reduced by both metals in wheat (around 40%). In sunflower leaves, both metals greatly reduced arginine decarboxylase (ADC) activity, while ornithine decarboxylase (ODC) was only decreased by cadmium. However, in wheat leaves, cadmium increased ADC and ODC activities (210 and 2200% over the control, respectively), and Cu 2+ raised ODC activity fivefold respect to the control, with the resulting increase in Put concentration. Moreover, the decreased diamine oxidase activity observed with both metals in wheat leaves might contribute to the high Put level observed. Cadmium and copper increased ethylene production sixfold in wheat leaves, while in sunflower leaves, only copper produced a great increment in the hormone (300% over the control).

Relationship between antioxidant defence systems and salt tolerance in Solanum tuberosum

A relationship between the antioxidant defence system and salt tolerance in two clones of potato (Solanum tuberosum L.) differing in salt sensitivity was studied. The antioxidant defence system of the sensitive clone responded differently to 100 and 150 mM NaCl. At 100 mM NaCl, growth, dehydroascorbate reductase and catalase activities remained unaltered, but chlorophyll and reduced glutathione content decreased (23% and 35%, respectively), while ascorbate content and superoxide dismutase activity were increased 34% and 63%, with respect to the control (0 mM NaCl). The superoxide dismutase increment was higher under 150 mM NaCl treatment, while a general decrease (except for dehydroascorbate reductase and catalase activities) in all the antioxidant parameters studied was observed in the sensitive clone. Reduced glutathione and ascorbate, the main antioxidant soluble defences, and all antioxidant enzymes (except catalase) were significantly elevated in the tolerant clone compared to the sensitive one when both were grown in the absence of NaCl. Under 100 and 150 mM NaCl treatments, no changes in the antioxidant stress parameters were detected in the tolerant clone. These results suggest a relationship between salt tolerance and the antioxidant defence system in the two clones.

Behaviour of antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells

A relationship between the antioxidant defense system and salt tolerance in two types of sunflower calli differing in salt sensitivity was studied. No reduction in growth occurred in the NaCl-salt-adapted cell line (T) when grown on 175 mM NaCl but growth of the salt-stressed cell line (S) was reduced by 83%. Lipid peroxidation and protein oxidation increased during acute stress of salt stressed cells at 14 and 28 d of the experiment, while salt-adapted calli (T) remained similar to non-shocked (C) values. The antioxidant defense system of callus adapted to growth under NaCl responded differently to 175 mM of salt compared with the corresponding controls under shock treatment. Salt-adapted and salt-stressed calli showed a similar pattern in GSH content at day 14 but at day 28 in S calli, GSH content was increased 100% over the non-shocked calli, while T calli returned to the initial values. In the salt-stressed calli, a general decrease in all the antioxidant enzymes studied (except for glutathione reductase and dehydroascorbate reductase activities) was observed at day 28. Except for catalase, the antioxidant enzymes were elevated constitutively in adapted calli as compared to stressed cells, when both were grown in the absence of NaCl (time 0), and remained unaltered until 28 d after the beginning of the experiment. These results suggest the involvement of an enzymatic antioxidant defense system in the adaptive response to salt stress in Helianthus annuus L. cells.

Effect of cadmium stress on nitrogen metabolism in nodules and roots of soybean plants

The nitrogen metabolism of soybean (Glycine max L.) nodules and roots was studied in plants subjected to two different concentrations (50 and 200 μM) of CdCl2. Nitrogenase activity was decreased in nodules treated with 200 μM Cd2+. In 50 μM Cd2+-treated plants, NH4+ content showed similar values to controls in nodules, but increased by 55% in roots. However, after treatment with 200 μM Cd2+, NH4+ levels increased in both tissues. Glutamate (Glu) and protein contents remained unaltered in nodules treated with 50 μM Cd2+, while at the higher Cd2+ concentration both were decreased. Nevertheless, polyamine content was increased at the two Cd2+ concentrations. In roots, Glu, polyamine and protein levels were significantly diminished at 50 and 200 μM CdCl2. For nitrogen-assimilation enzymes, glutamate dehydrogenase activity was moderately increased in nodules and roots following the lower Cd2+ treatment, though at the higher Cd2+ concentration root enzyme activity returned to control levels. An impressive increase in enzyme activity was found in nodules. In roots, the glutamine synthetase / glutamate synthase pathway was decreased at the two Cd2+ concentrations, though in nodules it was diminished only at 200 μM Cd2+. No changes in protease activity were found in the two tissues treated with 50μMCd2+. However, at 200 μM Cd2+, nodule and root protease activities decreased and increased, respectively. These results suggest that, in general, treatment with Cd2+ affects nitrogen assimilation and metabolism to a greater extent in soybean roots than in nodules.

Polyamines and heavy metal stress: the antioxidant behavior of spermine in cadmium- and copper-treated wheat leaves

Polyamine metabolism, as well as spermine (Spm) antioxidant properties, were studied in wheat leaves under Cd2+ or Cu2+ stress. The oxidative damage produced by both metals was evidenced by an increased of thiobarbituric acid reactive substances (TBARS) and a significant decrease in glutathione under both metal treatments. Ascorbate peroxidase (APOX) and glutathione reductase (GR) activities were reduced by both metals to values ranging from 30% to 64% of the control values. Conversely, copper produced a raise in superoxide dismutase activity. The high putrescine (Put) content detected under Cd2+ stress (282% over the control) was induced by the increased activity of both enzymes involved in Put biosynthesis, arginine decarboxylase (ADC) and ornithine decarboxylase (ODC). However, only ODC activity was increased in wheat leaves subjected to Cu2+ stress, leading to a lower Put rise (89% over the controls). Spermidine (Spd) content was not affected by metal treatments, while Spm was significantly reduced. Pretreatment with Spm completely reverted the metals-induced TBARS increase whereas metals-dependent H2O2 deposition on leaf segments (revealed using diaminobenzidine), was considerably reduced in Spm pretreated leaf segments. This polyamine failed to reverse the depletion in APOX activity and glutathione (GSH) content produced by Cd2+ and Cu2+, although it showed an efficient antioxidant behavior in the restoration of GR activity to control values. These results suggest that Spm could be exerting a certain antioxidant function by protecting the tissues from the metals-induced oxidative damage, though this effect was not enough to completely avoid Cd2+ and Cu2+ effect on certain antioxidant enzymes, though the precise mechanism of protection still needs to be elucidated.