MT Concentrations Research Articles

Absorption and Distribution of Cadmium in Metallothionein-I Transgenic Mice

Metallothionein-I transgenic (MT-TG) mice have higher concentrations of MT in the stomach (10×), small intestine (4×), large intestine (6×), liver (15×), and kidney (5×) than control mice. The purpose of the present study was to use MT-TG mice to determine whether increased concentrations of MT affect cadmium (Cd) absorption and distribution. A single dose of109Cd was given to control and MT-TG mice orally (0.3–300 μmol/kg, 200 μCi/kg) or intravenously (0.03–10 μmol/kg, 20 μCi/kg). Cd concentrations in 15 tissues were quantified 7 days later. Higher MT concentrations in tissues of MT-TG mice had no appreciable effects on the concentration of Cd in tissues compared to controls. An exception to this was the MT-TG mice given the highest dose of Cd (300 μmol Cd/kg, po), which had twice the tissue Cd concentration of controls. Approximately 60% of the Cd administered iv was retained in the tissues; retention of Cd in MT-TG mice was similar to that in controls. In both control and MT-TG mice only 0.1–0.3% of Cd administered po was retained, except for 1–3% at the higher doses (100 and 300 μmol/kg). Cd administered iv distributed mainly to the liver (70%) and kidney (10%) and was independent of dose. In contrast, when administered po, distribution of Cd to the liver increased from 40 to 75% of the dose, whereas distribution to kidney decreased from 30 to 7% as doses were increased from 0.3 to 300 μmol/kg. No difference in pattern of Cd distribution to various organs was observed between control and MT-TG mice. These data indicate that higher concentrations of MT in MT-TG mice do not appear to inhibit the gastrointestinal absorption of Cd nor alter the organ distribution of Cd.

Effect of zinc supplementation on metallothionein, copper, and zinc concentration in various tissues of copper-loaded rats.

The present study was designed to investigate the effects of Zn administration on metallothionein concentrations in the liver, kidney, and intestine of copper-loaded rats. Male CD rats were fed a diet containing 12 mg Cu and 67 mg Zn/kg body wt. They were divided into either acute or chronic experimental protocols. Rats undergoing acute experiments received daily ip injections of either Cu (3 mg/kg body wt) or Zn (10 mg/kg body wt) for 3 d. Chronic experiments were carried out on rats receiving Cu ip injections on d 1, 2, 3, 10, 17, and 24, Cu injections plus a Zn-supplemented diet containing 5 g Zn/kg solid diet, or a Zn-supplemented diet alone. Rats injected Zn or Cu had increased MT concentrations in liver and kidney. Zn produced the most important effects and the liver was the most responsive organ. Rats fed a Zn-supplemented diet had significantly higher MT concentrations in liver and intestine with respect to controls. Increased MT synthesis in the liver may contribute to copper detoxification; the hypothesis of copper entrapment in enterocytes cannot be confirmed.

Transgenic Mice That Overexpress Metallothionein-I Are Protected from Cadmium Lethality and Hepatotoxicity

The purpose of this study was to determine whether metallothionein-I (MT-I) transgenic female mice (MT-TG) are resistant to cadmium (Cd) hepatotoxicity. Female MT-TG mice have 10- to 20-fold higher MT concentrations in liver than control mice and are more resistant to Cd-induced lethality than control mice. CdCl2 (3.7 mg Cd/kg, iv) was lethal to 73% of control mice, but only to 13% of MT-TG mice. Cd administration (3.1 mg/kg, iv) to control mice produced extensive liver injury as evidenced by 20- and 70-fold increases in serum enzyme activities of sorbitol dehydrogenase and alanine aminotransferase, respectively. MT-TG mice are considerably more resistant to Cd-induced hepatotoxicity than control mice, as evidenced by only about one-tenth the elevation in serum enzymes observed in control mice and a lower incidence of hepatocyte necrosis in MT-TG mice. To ascertain the mechanism of this protection, the distribution of Cd to various organs and the subcellular distribution of Cd in liver were determined 2 hr after Cd injection (109CdCl2, 3.5 mg Cd/kg, iv). The hepatic subcellular distribution of Cd was altered markedly in MT-TG mice, with much less Cd distributing to nuclei, mitochondria, and microsomes (25, 42, and 24% of controls, respectively), and more Cd to the cytosol (240% of controls). The increased cytosolic Cd was bound primarily to MT, as determined by G-75 gel chromatography. In addition, primary hepatocyte cultures from MT-TG mice maintained higher levels of MT than hepatocytes from control mice and were more resistant to Cd cytotoxicity than control hepatocytes. In conclusion, studies using MT-I transgenic mice demonstrate that MT protects against Cd lethality and hepatotoxicity, and this hepatoprotective effect of MT is also observed in hepatocyte cultures from MT-TG mice.

Metal accumulation and metallothionein concentrations in tree swallow nestlings near acidified lakes

The authors studied metal accumulation in hepatic and renal tissues of tree swallow (Tachycineta bicolor) nestlings at acidified and nonacid reference lakes in northwestern Ontario. Hepatic concentrations of metallothionein (metal-binding proteins, MT) in tree swallow nestlings were negatively correlated with pH of the nest-site lake. Combined concentrations of Cu and Zn in the liver were correlated with liver MT concentrations, but Cd was not. Although no overt signs of metal toxicity were observed in nestlings near acid lakes, the results clearly provided evidence that metals are transferred from acid lakes to birds and that these metals are correlated with increases in hepatic MT production.

Induction of metallothionein by cadmium-metallothionein in rat liver: A proposed mechanism

Distribution of Cd to various organs following iv administration of CdCl 2 (3.5 mg Cd/kg) resulted in more than 43% of total tissue Cd accumulating in the liver. In contrast, after CdMT administration (0.5 mg Cd/kg), only 1% of the Cd was found in liver. Rats administered CdCl 2 (1.0 mg Cd/kg) had hepatic MT values 30-fold greater than controls and a hepatic Cd concentration of 17 μg/g. In comparison, rats treated with CdMT (0.4 mg Cd/kg) had hepatic MT concentrations 7-fold greater than controls and a hepatic Cd concentration of 0.80 μg/g. However, when hepatic MT levels were normalized to tissue Cd concentrations, induction of MT by CdMT was 5-fold greater than by CdCl 2. Northern and slot-blot analyses of mRNA showed that both CdCl 2 and CdMT coordinately increased MT mRNA. These data suggest that both CdMT and CdCl 2 increase hepatic MT by similar mechanisms. A dose-response increase in MT produced by CdCl 2 indicated a biphasic response, with low doses producing relatively more hepatic MT than higher doses. In addition, the amount of MT produced per unit Cd after CdMT treatment was similar to those observed after low doses of CdCl 2 in the dose-response experiment. These data provide strong evidence to support the conclusion that the apparent potency of CdMT observed here and in previous studies is most likely due to the small amount of Cd distributed to the liver, which is relatively more effective in inducing MT than are higher concentrations.

Influence of thyroparathyroidectomy and thyroxine replacement on Cu and Zn cellular distribution and on the metallothionein level and induction in rats.

Thyroid hormones are involved in copper and zinc distribution in rat tissues. We examined the influence of thyroparathyroidectomy (TPTY) and of a replacement therapy by T4 on Cu and Zn organ distribution. MT levels were also measured both in basal conditions and after induction by cadmium. The results confirm that a lack of T4 modified Cu and Zn in serum and tissues. In serum, TPTY increased Cu (+15%) and ceruloplasmin (+18%), and decreased Zn (-18%). In tissues, Cu was altered in liver (+13%), kidney (-24%), heart (-16%) duodenum (-18%), and Zn in liver (+25%) and kidney (-10%). The soluble fractions (100,000 g supernatant) were mainly affected in liver and kidney, and the subcellular fractions in heart and duodenum. MT levels were modified in basal conditions only in liver (+57%) and kidney (-36%). T4 administration partially prevented the effect of TPTY on both elements and MT concentrations. Therefore, no evidence is provided for a direct role of T4 in the metabolism of MT in a way comparable to the effects of glucocorticoids. However, MT could mediate the consequences of TPTY on metal distribution in certain organs, such as liver and kidney.

Regulation of the isoforms of metallothionein

The objective of this research was to study the regulation of hepatic MT-I and MT-II in rats. Induction of MTs following Cd, Zn, and glucocorticoid treatments was studied in adult rats and the ontogeny of MTs was evaluated in developing rats. The isoforms were quantitated by anion-exchange HPLC and their respective mRNAs were evaluated by Northern blot hybridization with mouse cRNA probes. In untreated adult rat liver, MT-II was more abundant than MT-I. Cd treatment increased MT-I and MT-II to similar concentrations. Low dosages of Zn increased MT-II levels three times higher than MT-I levels, whereas higher dosages increased the isoforms to similar levels. Differences in the degradation of Zn-induced MTs were observed: apparent half-lives for MT-I and MT-II were 12.2 and 21.9 h, respectively. MT-II was the predominant glucocorticoid-inducible isoform. MT-I and MT-II mRNAs increased coordinately after glucocorticoid treatment, but MT-II mRNA was more abundant than MT-I mRNA. In immature rats, MT-I and MT-II were first detected in fetal liver on gestation day 18 and increased to maximum hepatic concentrations during the first week postpartum. Thereafter, MT levels decreased, reaching adult levels by 35 d. In contrast, MT mRNAs reached maximal levels during late gestation that were maintained throughout the first three weeks postpartum. Discoordination between MT mRNA and protein levels is suggestive of translational control of MTs in developing rats. In summary, MT-II is more abundant than MT-I in untreated rats and in rats treated with glucocorticoids or low dosages of zinc. In addition to transcriptional regulations of MTs, translational control of MTs may be important in regulating hepatic MT concentrations, particularly in developing rats, and differences in the degradation of MT-I and MT-II may influence tissue concentrations of the isoforms.

Cadmium-metallothionein nephrotoxicity in the rat: Transient calcuria and proteinuria

After a s.c. injection of 0.4 mg Cd/kg as cadmium-metallothionein (CdMT) in rats, a marked increase in urinary protein concentration appeared at 16–40 h. There was a peak of urinary Cd content during the first 4 h after the treatment. Urinary Ca was increased at 8 h after the CdMT injection and returned to normal level at 32 h. Luminal and basolateral renal membrane vesicles were isolated from both control group and CdMT (0.4 mg Cd/kg) group at 24 h after the injection. Calcium uptake and binding of both fractions were decreased in the group treated with CdMT. Cd, Zn and MT concentrations in the kidney cortex were increased, but Ca concentration was not significantly changed. Since injected CdMT is probably only partly reabsorbed by tubular cells at the dose level of 0.4 mg Cd/kg as CdMT, excessive plasma CdMT is rapidly excreted in urine, explaining the increased Cd excretion during the first few hours observed in the present experiment. Decreased Ca binding in the luminal membranes as observed in vitro could be one of the mechanisms of production of calcuria if occurring in vivo. Another possible explanation of calcuria is that Cd ions released from CdMT into the cytoplasm of the tubular cell, may exert ionic interference with Ca transport across the luminal membranes and produce decreased Ca reabsorption. It is known that a disturbance of Ca metabolism could influence the membrane stability and such a change may contribute to explaining the proteinuria characteristic of CdMT nephrotoxicity. The reversibility of the proteinuria observed after a single dose of CdMT may be related to the induction of metallothionein synthesis in the renal cells.

On the proposed role of metallothionein in the transport of cadmium

In order to test a possible role of MT in the transport of Cd from liver to kidney, we compared the distribution of 109Cd administered either in the form of CdCl 2 or CdMT. Unlike in earlier studies which employed relatively massive amounts of Cd, in the present studies a low total amount of Cd (15–30 μg) was continuously infused over a period of 2–3 days. Results confirmed a preferential renal accumulation of CdMT. High intracellular renal and hepatic MT concentrations, induced by repeated prior injections of Cd, Zn, or Hg, did not alter the tissue distribution of infused CdCl 2 or CdMT. It is interesting to note that in some further experiments, equivalent amounts of extracellular MT depressed renal uptake of CdMT. These findings are consistent with a role of MT in the transport of Cd from liver to kidney.