Uptake and toxic effects of heavy metal ions: interactions among cadmium, copper and zinc in cultured cells

Absorption of metal ions by KB, HeLa and L-59 cells has been analyzed by atomic absorption spectrophotometry in the course of culture. Ions of the elements of the fourth period in the periodic chart such as Fe(II), Cu(II), Zn(II), Mn(II) and Ni(II) were not taken up, but those of the higher periods, such as Cd(II), Pb(II), Hg(II) and Ag(I) were were taken up easily. The uptake behavior by the cultured cells was in accordance with the characteristic features of metals, that metals in the fourth period are essential elements, and most of the elements of the fifth and the sixth periods are non-essential or toxic elements. The initial rate of Cd(II) uptake and the Cd(II) concentration has a sigmoidal relationship. Cd(II) was absorbed homotropically through cell membranes. The uptake of Cd(II) was specifically inhibited by Cu(II), but was affected little by Zn(II). The toxicity of Cd(II) to KB cells was greatly enhanced in the presence of Cu(II). On the contrary, the toxicity of Cd(II) was reduced by the addition of Zn(II) at several concentrations of Cd(II). The toxicity of Cd(II) did not depend on the amount of Cd(II) absorbed in the cells, but was determined by cofactors such as Cu(II). The interaction between Cd(II) and Cu(II) may be important for Itai-itai disease.     

Uptake and toxic effects of heavy metal ions: Interactions among cadmium,copper and zinc in cultured cells

Summary Absorption of metal ions by KB, HeLa and L-59 cells has been analyzed by atomic absorption spectrophotometry in the course of culture. Ions of the elements of the fourth period in the periodic chart such as Fe(II), Cu(II), Zn(II), Mn(II) and Ni(II) were not taken up, but those of the higher periods, such as Cd(II), Pb(II), Hg(II) and Ag(I) were were taken up easily. The uptake behavior by the cultured cells was in accordance with the characteristic features of metals, that metals in the fourth period are essential elements, and most of the elements of the fifth and the sixth periods are non-essential or toxic elements.The initial rate of Cd(II) uptake and the Cd(II) concentration has a sigmoidal relationship. Cd(II) was absorbed homotropically through cell membranes. The uptake of Cd(II) was specifically inhibited by Cu(II), but was affected little by Zn(II). The toxicity of Cd(II) to KB cells was greatly enhanced in the presence of Cu(II). On the contrary, the toxicity of Cd(II) was reduced by the addition of Zn(II) at several concentrations of Cd(II). The toxicity of Cd(II) did not depend on the amount of Cd(II) absorbed in the cells, but was determined by cofactors such as Cu(II). The interaction between Cd(II) and Cu(II) may be important for Itai-itai disease.     

Distribution of cadmium,zinc and copper in the musselMytilus edulis. Existence of cadmium-binding proteins similar to metallothioneins

Summary The study of the distribution of Cd, Zn and Cu in homogenates of mussels has shown that Zn and Cu are principally associated with high mol. wt. proteins. The same distribution is observed for Cd in untreated mussels, but in Cd chronically intoxicated animals, the metal is principally bound to low mol. wt. proteins synthesized in response to the uptake of the cation and similar to metallothioneins of vertebrates.This research was carried out in participation in the Belgian National Research and Development Program on the Environment-Water-Sea Project-Office of the Prime Minister-Interministerial Commission for Science Policy and Programmation.I am grateful to Prof.A. Disteche and Dr.Ch. Gerday for theiradvice throughout this work. I thankR. Biondo andN. Gerardin for excellent technical assistance.     

Effects of dosage and cadmium pretreatment on the binding of cadmium in rat bile.

The effects of dosage and of cadmium pretreatment on the binding of cadmium in rat bile were studied. With increasing dose a higher cumulative biliary excretion of Cd was observed and a higher percentage of the Cd was excreted in a low-molecular-weight form. On the other hand, after cadmium pretreatment, a decrease in the cumulative biliary excretion of cadmium was observed but a greater percentage of that excreted into the bile was bound to high molecular weight compounds.     

Effects of dosage and cadmium pretreatment on the binding of cadmium in rat bile

Summary The effects of dosage and of cadmium pretreatment on the binding of cadmium in rat bile were studied. With increasing dose a higher cumulative biliary excretion of Cd was observed and a higher percentage of the Cd was excreted in a low-molecular-weight form. On the other hand, after cadmium pretreatment, a decrease in the cumulative biliary excretion of cadmium was observed but a greater percentage of that excreted into the bile was bound to high molecular weight compounds.     

Subcellular distribution and binding of heavy metals in the untreated liver of the squid; comparison with data from the livers of cadmium and silver-exposed rats

In natural squid liver, about 30% of the total Cd present was found in the cytosolic fraction. A large portion of this Cd was bound to high molecular weight species (mol. wt greater than 70,000). In contrast to Cd, about 60% of the total Ag occurred in the cytosolic fraction; Ag was bound mainly to low molecular weight species (mol. wt less than 20,000).     

Clearance of orally administered115mCd from rat tissues

Summary Peak^115mCd levels in the liver, kidneys, spleen and duodenum of the rat, following a single oral dose, fell exponentially. Half-clearance time for^115mCd is highest in the kidney (30 days) followed by the liver (6.8 days), the spleen (5.5 days) and the duodenum (3.5 days).     

Effects of cadmium on the immune system of mice.

Chronic oral exposure of mice to Cd++ inhibits cell-mediated immunity of delayed type hypersensitivity induced by sheep red blood cells (SRBC). No effect was detected on humoral immune response to SRBC. Spleen cells derived from mice exposed to Cd++ showed in vitro enhanced response to T and B cell mitogens. These results demonstrate that Cd++ exposure alters the immune system of mice.     

The molecular mechanism of zinc and cadmium stress response in plants

When plants are subjected to high metal exposure, different plant species take different strategies in response to metal-induced stress. Largely, plants can be distinguished in four groups: metal-sensitive species, metal-resistant excluder species, metal-tolerant non-hyperaccumulator species, and metal-hypertolerant hyperaccumulator species, each having different molecular mechanisms to accomplish their resistance/tolerance to metal stress or reduce the negative consequences of metal toxicity. Plant responses to heavy metals are molecularly regulated in a process called metal homeostasis, which also includes regulation of the metal-induced reactive oxygen species (ROS) signaling pathway. ROS generation and signaling plays an important duel role in heavy metal detoxification and tolerance. In this review, we will compare the different molecular mechanisms of nutritional (Zn) and non-nutritional (Cd) metal homeostasis between metal-sensitive and metal-adapted species. We will also include the role of metal-induced ROS signal transduction in this comparison, with the aim to provide a comprehensive overview on how plants cope with Zn/Cd stress at the molecular level.     

Metallothionein induction by cadmium and zinc in rat secretory organs

Metallothionein (MT) levels were determined in four secretory organs of the rat following administration of zinc (Zn) and cadmium (Cd). The concentrations of MT in the lacrimal, parotid and adrenal glands of untreated rats were in the range of 2.2-4.9 micrograms/g wet weight tissue while in the pancreas it was shown to be 15.2 micrograms/g. Injection of zinc at total doses of 16, 32 and 80 mg/kg resulted in a 1.8-, 3.2- and 5.9-fold increase in lacrimal MT content, respectively, while a 10.2- and 13.1-fold elevation was observed following treatment with 4 and 8 mg/kg of Cd, respectively. Similar findings were found in the adrenal gland. The parotid MT was elevated 5.9 and 17 times following Zn treatment at doses of 16 and 80 mg/kg respectively, whereas 4 mg/kg of Cd increased MT 14.4 times in this gland. Pancreatic MT was elevated by 39- and 40-fold after injection of Zn at doses of 16 and 32 mg/kg respectively, whereas 4 and 8 mg/kg of Cd caused a 9.8- and 17.9-fold induction, respectively. These results may indicate that secretory organs participate in metabolism of heavy metals in the mammalian body.     

Cadmium as a trace element and cadmium binding components in human cells

Zusammenfassung Fötale menschliche Fibroblasten wurden in einem¹⁰⁹Cd und⁶⁵Zn enthaltenden Kultur medium gezÜchtet.¹⁰⁹Cd wurde in allen Zellen kontinuierlich absorbiert. Mit Gel-Filtration wurde der Zellsaft in 2 Komponenten aufgetrennt, die beide¹⁰⁹Cd enthielten. ähnliche,¹⁰⁹Cd bindende Komponenten wurden auch in Rattenleber-Homogenaten gefunden.     

Effect of platelet activating factor on guinea-pig papillary muscle

Platelet activating factor (PAF) induces a biphasic effect on guinea-pig papillary muscle: 1. a transient positive inotropic effect preceded by an increase in action potential duration (APD); 2. a marked negative effect on inotropism and on APD. Since Ca++ slow action potentials were initially enhanced by PAF and then markedly depressed, it is suggested that PAF specifically interferes with the Ca++ slow channel.     

Dependence of 6β-acetoxy-7α-hydroxyroyleanone block of Kv1.2 channels on C-type inactivation

Voltage-gated K⁺ (Kv) channels exhibit slow or C-type inactivation during continuous depolarization. A selective pharmacological agent targeting C-type inactivation is hitherto lacking. Here, we report that 6β-acetoxy-7α-hydroxyroyleanone (AHR), a diterpenoid compound isolated from Taiwania cryptomerioides, can selectively modify C-type inactivation of Kv1.2 channels. Extracellular, but not intracellular, AHR (50 μM) dramatically accelerated the slow decay of Kv currents and left-shifted the steady-state inactivation curve. AHR blocked Kv currents with an IC₅₀ of 17.7 μM. AHR did not affect the kinetics and voltage-dependence of Kv1.2 channel activation. Channel block by AHR was independent of intracellular K⁺ concentration. In addition, effect of AHR was much attenuated in a Kv1.2 V370G mutant defective in C-type inactivation. Therefore, block of Kv1.2 channels by AHR did not appear to involve direct occlusion of the outer pore but depended on C-type inactivation. AHR could thus be a probe targeting Kv channel C-type inactivation gate.     

Increased erythrocyte permeability to Li and Na in the spontaneously hypertensive rat.

Red blood cells incubated in a physiological medium in which Li replaces Na (LiPSS) gain Li in exchange for Na and K. The rate of Li uptake is modestly but significantly increased in the spontaneously hypertensive rat (SHR) at 37 degrees C and at 22 degrees C. The slow rate of Na gain and K loss during cooling at 2 degrees C was about doubled in unmodified whole blood samples from the SHR.     

Effect of platelet activating factor on guinea-pig papillary muscle

Summary Platelet activating factor (PAF) induces a biphasic effect on guinea-pig papillary muscle: 1. a transient positive inotropic effect preceded by an increase in action potential duration (APD); 2. a marked negative effect on inotropism and on APD. Since Ca⁺⁺ slow action potentials were initially enhanced by PAF and then markedly depressed, it is suggested that PAF specifically interferes with the Ca⁺⁺ slow channel.     

Endogenous oscillatory activity and TTX-sensitive spikes of the heart muscle in early juveniles of the isopod crustaceanLigia exotica

The heart beat of early juveniles of the littoral isopodLigia exotica occurred at a frequency of 250 to 350/min, associated with rhythmic activity of the heart muscle. Each burst was composed of a slow depolarizing potential with superimposed spike potentials. The spike potential was eliminated by perfusion with TTX-containing or Na⁺-free saline. In TTX-saline, the slow potential was unchanged in frequency and amplitude. By current injection into the heart muscle, the rhythm of the slow potential was phase-shifted and its frequency was changed in a membrane potential-dependent manner. These results show that the heart ofLigia early juveniles acts as an endogenous muscle oscillator generating oscillatory slow potentials and Na⁺-dependent spikes.     

Metallothionein induction by cadmium and zinc in rat secretory organs

Summary Metallothionein (MT) levels were determined in four secretory organs of the rat following administration of zinc (Zn) and cadmium (Cd). The concentrations of MT in the lacrimal, parotid and adrenal glands of untreated rats were in the range of 2.2–4.9 g/g wet weight tissue while in the pancreas it was shown to be 15.2 g/g. Injection of zinc at total doses of 16, 32 and 80 mg/kg resulted in a 1.8-, 3.2- and 5.9-fold increase in lacrimal MT content, respectively, while a 10.2- and 13.1-fold elevation was observed following treatment with 4 and 8 mg/kg of Cd, respectively. Similar findings were found in the adrenal gland. The parotid MT was elevated 5.9 and 17 times following Zn treatment at doses of 16 and 80 mg/kg respectively, whereas 4 mg/kg of Cd increased MT 14.4 times in this gland. Pancreatic MT was elevated by 39- and 40-fold after injection of Zn at doses of 16 and 32 mg/kg respectively, whereas 4 and 8 mg/kg of Cd caused a 9.8- and 17.9-fold induction, respectively. These results may indicate that secretory organs participate in metabolism of heavy metals in the mammalian body.     

Recombinant synthesis of mouse Zn3-β and Zn4-α metallothionein 1 domains and characterization of their cadmium(II) binding capacity

Genetic engineering, coupled with spectro scopic analyses, has enabled the metal binding proper ties of the α and β subunits of mouse metallothionein 1 (MT) to be characterized. A heterologous expression system in E.coli has led to high yields of their pure zinc-complexed forms. The cadmium(II) binding properties of recombinant Zn₄-αMT and Zn₃-βMT have been studied by electronic absorption and circular dichroism. The former binds Cd(II) identically to α fragments obtained from mammalian organs, showing that the recombinant polypeptide behaves like the na tive protein. Titration of Zn₃-βMT with CdCl₂ results in the formation of Cd₃-βMT. The addition of excess Cd(II) leads to Cd₄-βMT which, with the extra loading of Cd(II), unravels to give rise isodichroically to Cd₉-βMT. The effect of cadmium-displaced Zn(II) ions and excess Cd(II) above the full metal occupancy of three has been studied using Chelex-100. The Cd₃-βMT species is stable in the presence of this strong metal-chelating agent. Received 20 May 1997; received after revision 7 July 1997; accepted 9 July 1997     

Effect of dimethindene, an antihistaminic drug, on the transmembrane potentials of mammalian myocardium

Dimethindene (DMI) decreased the maximum rate of rise of action potential (AP) without changing the resting potential in cat ventricular myocardium. DMI abolished the histamine-induced slow APs in left atria but not in right ventricular papillary muscles of guinea-pig, suggesting that DMI blocked the histamine H1-receptors.