Correlations between cadmium treatment, oxygen uptake and metallothionein response in liver and kidney from two mice strains.

Oxygen uptake was determined in liver and kidney slices from Balb/C and C3H/HeJ mice at various time points related to a 4-day cadmium (Cd) i.p. injection regime, the latter strain being not only the higher metallothionein (MT) responder, but also more Cd sensitive. In both strains Cd injections caused an increase in O2 uptake in the tissues, being more expressed in the higher MT responsive C3H/HeJ strain. An ability for fast, additional increase of O2 uptake in response to subsequent Cd exposure was also induced during the injection regime. Maximal obtainable MT levels in the two strains were estimated by forced overwhelming pneumococcal infection challenge, giving induction values above those obtained for the Cd injections in the Balb/C strain and equal to those obtained for Cd in the more responsive C3H/HeJ strain. Together with direct defendants against harmful oxygen radicals, MT may act complementary as a radical scavenger and by Cd inactivation, but to a higher degree in the C3H/HeJ compared to the Balb/C strain.

Accumulation of metallothionein and its multiple forms by zinc, cadmium and dexamethasone in human peripheral T and B lymphocytes and monocytes.

Metallothionein (MT) is produced at high rates in isolated monocytes, and T and B lymphocytes during induction in vitro. At optimal concentrations, 125 microM for Zn and 10 microM for Cd and dexamethasone (dex), MT was demonstrated after only 2 h in some cases, and in all cell types substantial levels were measured after 1 day of exposure to all three inductors. With Cd, lower amounts of MT were found, but maximum levels were reached faster than with Zn. The same result was found for dex compared to Zn. Zn and dex in combination showed the same accumulation rate as Zn alone. If the inductors were used in lower concentrations than optimal, reduced accumulation rates occurred, particularly during the first part of the exposure period. No MT was found for concentrations below 5 microM Zn, 1 microM Cd or 0.5 microM dex. The constitutive levels of MT (mean +/- S.E.M.) were 0.11 +/- 0.05, 0.54 +/- 0.3, 0.06 +/- 0.05 and 0.15 +/- 0.08 nmol Cd bound/5 x 10(6) unseparated mononuclear cells (MNC), monocytes, T lymphocytes and B lymphocytes, respectively. Monocytes accumulated 19 times and B lymphocytes 6 times more MT than T lymphocytes after 2 days of exposure to 125 microM Zn. Despite these differences in accumulated amounts of MT, the fold accumulation values were rather similar between the cell types, reflecting corresponding variations in background MT levels. After exposure of unseparated MNC to 125 microM Zn for 2 days, removal of the metal caused constitutive MT levels to be reestablished in 5 days. Five different MT forms, all capable of Cd complexation, were demonstrated in these cells. These forms had kinetically different behaviour during Zn exposure among the cell types, and the response to Cd was different from the Zn response. The results indicate metals to be closely controlled in MNC and emphasize a role for multiple MT forms in the process of regulation.

Cytotoxicity and accumulation of Hg, Ag, Cd, Cu, Pb and Zn in human peripheral T and B lymphocytes and monocytes in vitro.

1. The cytotoxic effects of various heavy metals were assayed by trypan blue exclusion in vitro in human peripheral immune cells separated to high purity. T and B lymphocytes and monocytes were equally sensitive to metals. The individual metals could be ranked in the following decreasing order of cytotoxic potency, Hg approximately Ag > Cd approximately Cu > Pb approximately Zn, based on exposure time and concentration needed to give a particular percentage of dead cells. 2. The cytotoxic effects became irreversible after about 13 hr of metal exposure. 3. Examination by scanning electron microscopy showed that the heavy metals caused serious destruction of the cell membranes. 4. Solubility and uptake of metals into the cells were studied and discussed in relation to the cytotoxic effects. It was concluded that metal binding to cell surfaces or precipitate formation could inhibit ordinary uptake, thereby affecting cytotoxicity. For Pb in monocytes this appeared to lead to uptake of non-toxic complexes, probably by phagocytosis.

Mitogenicity and metallothionein induction: two separate effects of zinc ions on human mononuclear blood cells.

The mitogenic effect of elevated concentrations of zinc ions on human lymphocytes was found to be inhibited by the glucocorticoid hormone dexamethasone. The effect decreased progressively from complete block to partial inhibition when the culture period before the addition of dexamethasone was prolonged. In contrast, dexamethasone did not inhibit an induced formation of metallothionein in mononuclear cells. Furthermore, dexamethasone itself was found to induce small amounts of metallothionein. Apparently, the mitogenic effect and the induction of metallothionein by zinc ions in mononuclear cells occur by mutually independent mechanisms.

Effect of the metal in the reaction between metallothionein and antimetallothionein antibody.

1. An antimetallothionein antibody, raised against Cd-carrying metallothionein, was applied in Western blotting of metallothionein. 2. Treatment of the electroblotted nitrocellulose sheets with metals belonging to the periodic system transition groups Ib and IIb, or with Pb, Ni or Cr, considerably enhanced binding of anti-metallothionein. A similar effect was found when the electroblotted sheets were treated with the strong alkylator N-ethylmaleimide. 3. It seems that the binding of metal to metallothionein modifies the configuration of the antibody binding sites by the formation of metal thiolate complexes. 4. Metal treatment of the nitrocellulose sheets after electroblotting, but before application of the primary antibody, offers a convenient method for use in Western blotting to significantly potentiate the reaction between metallothionein and the antimetallothionein antibody.

Induction of metallothionein production by zinc in human mononuclear cells.

1. Low mol. wt proteins with high Cd-binding capacity were found to be induced by Zn in cultured monocytes and lymphocytes. 2. In T cell cultures one protein was found to be induced by 125 microM Zn for 6 days, while in monocytes and B enriched cells under these conditions two proteins were found, of which the one with higher mol. wt had similar electrophoretic mobility to the T cell protein on polyacrylamide gels. 3. Mol. wt criteria and crossreactivity towards anti-metallothionein (Mt) antibody identified these proteins to be Mts of about 23 and 27 kDa mol. wt. 4. Metal binding studies indicated that monocyte and lymphocyte Mts had a higher affinity to Zn compared to Cd than rat liver Mt and Mts from Cd-resistant substrains of a human epithelial and a murine fibroblast cell line. 5. The presence of the T cell mitogen phytohemagglutinin (PHA) was found not to be necessary for this Mt induction by Zn.