Renal Metallothionein Levels Research Articles

Effect of Bisphenol A on the Levels of Vitellogenin and Metallothionein in Adult Male Carp, <i>Cyprinus carpio carpio</I. Linnaeus, 1758

Purpose: To investigate the potential role of bisphenol A (BPA) in inducing endocrine disruption in fish.Methods: Adult male koi carps, Cyprinus carpio carpio (n = 60, mean body weight = 125 ± 12.288 g and mean length = 24 ± 1.392 cm) were exposed to three graded concentrations of BPA (10, 100 and 1000 μg/L) for a period of 21 days. A single dose of 17-β estradiol (1 ng/l) was used as positive control.Results: The mean plasma concentration of vitellogenin (VTG) significantly increased in the carps exposed to 100 μg BPA/L reaching a mean value of 14600.0 ± 1400.0 ng/ml in comparison to 1140.00 ± 34.64 ng/ml observed for the control group,. However, BPA at all experimental concentrations had no significant effect on metallothionein levels in the liver, but the levels in the kidney were significantly decreased in all BPA-exposed groups in comparison to the level recorded for control (181.69 ± 0.77 pg/g wet tissue).Conclusion: Exposure to BPA modulates plasma vitellogenin at median concentration and renal metallothionein levels at all concentrations in carp. Thus, these parameters are potential biomarkers in the biomonitoring of the aquatic environment for estrogenic chemicals.Keywords: Carp, Bisphenol A, Vitellogenin, Metallothionein, Endocrine disruptors

Attenuation effects of compatible medicines on arsenical and lead toxicity of Badu Shengji San

Badu Shengji San(BDSJS) is a traditional Chinese medicine (TCM) used for drawing out toxin, eliminating suppuration and promoting granulation. Toxic minerals such as arsenic and lead are the two most important components of BDSJS. Previous hypothesis indicated that according to the compatibility theory of TCM, the toxicity of the entire BDSJS was weaker than that of arsenic and lead, respectively. In the present study, SD rats with injured skin were treated with distilled water and different composition of BDSJS (complete formulations, compatible herbs, mineral medicine containing arsenic and lead, mineral medicine containing arsenic and mineral medicine containing lead) once a day for consecutive 2 weeks. Kidney coefficient and urinary beta-N-acetyl glucosidase (NAG) were used as the indicators of renal toxicity and the content of malondiadehyde (MDA), superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), glutathione (GSH) and metallothionein (MT) in the renal tissue were measured. Our data showed that kidney coefficient, the severity of renal pathological lesion and MT level in the kidney of the entire BDSJS group decreased significantly compared with arsenic and lead group. Additionally, the NAG content of the entire BDSJS group had the decreased trend. The kidney CuZn-SOD level of the entire BDSJS group had the increased trend, but the MDA, GSH-PX, GSH level had no obvious difference. Our results suggested that compatible herbs in BDSJS relieved renal injury induced by arsenic and lead, and the attenuation mechanism may be related to MT and CuZn-SOD, but not to MDA, GSH-PX and GSH directly.

Dietary zinc attenuates renal lead deposition but metallothionein is not directly involved

Chronic lead exposure irreversibly damages the kidneys and may be associated with hypertension and renal insufficiency at sub-clinically toxic levels. Zinc supplementation reduces lead absorption and tissue retention in rodent models but the mechanisms are unknown. Metallothionein (MT) may function in lead detoxification. Our objective was to investigate the effects of marginal zinc (MZ) and supplemental zinc (SZ) intakes on renal lead and zinc accumulation, renal MT immunolocalization and levels. Weanling Sprague Dawley rats were assigned to MZ (8 mg Zn/kg diet), zinc-adequate control (CT; 30 mg Zn/kg), zinc-adequate diet-restricted (DR; 30 mg Zn/kg) or SZ (300 mg Zn/kg) groups, with and without lead acetate-containing drinking water (200 mg Pb/L) for 3 weeks. Kidneys were analyzed for lead and zinc by inductively coupled plasma spectroscopy and MT by immunolocalization and Western blotting. MZ had higher renal lead and lower renal zinc concentrations than CT. SZ was more protective than CT against renal lead accumulation. Renal MT levels reflected dietary intake (SZ > or = DR > or = CT > or = MZ) but lead had no effect on MT staining intensity, distribution, or relative protein amounts. In summary, while SZ lowered renal lead concentration, MT did not appear to function in renal lead accumulation. Future studies should explore alternate mechanisms of renal lead detoxification.

Antioxidants and metallothionein levels in mercury-treated mice

Acute effects of mercury on mouse blood, kidneys, and liver were evaluated. Mice received a single dose of mercuric chloride (HgCl2, 4.6 mg/kg, subcutaneously) for three consecutive days. We investigated the possible beneficial effects of antioxidant therapy (N-acetylcysteine (NAC) and diphenyl diselenide (PhSe)2) compared with the sodium salt of 2,3-dimercapto-1-propanesulfonic acid (DMPS), an effective chelating agent in HgCl2 exposure in mice. We also verified whether metallothionein (MT) induction might be involved in a possible mechanism of protection against HgCl2 poisoning and whether different treatments would modify MT levels and other toxicological parameters. The results demonstrated that HgCl2 exposure significantly inhibited delta-aminolevulinate dehydratase (delta-ALA-D) activity in liver and only DMPS treatment prevented the inhibitory effect. Mercuric chloride caused an increase in renal non-protein thiol groups (NPSH) and none of the treatments modified renal NPSH levels. Urea concentration was increased after HgCl2 exposure. NAC plus (PhSe)2 was partially effective in protecting against the effects of mercury. DMPS and (PhSe)2 were effective in restoring the increment in urea concentration caused by mercury. Thiobarbituric acid-reactive substances (TBARS), aspartate aminotransferase (AST), and alanine aminotransferase (ALT) activities and ascorbic acid levels were not modified after mercury exposure. Mercuric chloride poisoning caused an increase in hepatic and renal MT levels and antioxidant treatments did not modify this parameter. Our data indicated a lack of therapeutic effect of the antioxidants tested.

Increased endogenous antioxidants and reduced basement membrane thickening in Ames dwarf mouse kidney

Experimental evidence suggests that disease- and age-related thickening of glomerular (GBM) and tubular (TBM) basement membranes may be related to increases in reactive oxygen species (ROS). Studies also suggest that antioxidants may function as renoprotectants against such ROS-induced damage. The current study focuses on metallothionein (MT) and glutathione (GSH), both of which exhibit potent antioxidant functions. Ames dwarf mice, which exhibit elevated tissue antioxidant levels (catalase, Cu/Zn superoxide dismutase, GSH) and an average life span significantly greater (50–65%) than wild type controls, were used to test our hypothesis that increased levels of renal MT and GSH are concomitant with decreased rates of renal basement membrane thickening. A competitive ELISA assay was developed to quantitate both endogenously polymerized and “total” (glutaraldehyde-polymerized) renal MT while renal GSH levels were determined by fluorimetry. At 90, 300, 600, and 720 days, dwarf renal MT and GSH levels were significantly higher than controls. Interestingly, TEM morphometry data demonstrated that with increasing age, both the GBMs and TBMs of dwarf animals were thinner (1.1–15.7% and 28.8–45.5%, respectively) than those of age-matched controls. Since MT and GSH are well-documented ROS scavengers, it seems possible that in the Ames dwarf model of delayed aging, increased endogenous antioxidant levels provide renoprotection from oxidative damage and lead to reduced rates of age-related GBM and TBM thickening.

Effects of Iron Overload on Hepatic and Renal Metallothionein Levels in Rats

Metallothionein (MT) is one of the stress proteins induced by various heavy metals and oxidative stress. Although induction by Fe has not been well established, its overload is documented to cause oxidative stress in experimental animals. To investigate alteration in tissue MT levels that were possibly caused by Fe overload, male Wistar rats were fed a diet containing 3.5% Fe(II) fumarate (1.2% as Fe) for up to 21 days. Tissue Fe levels in liver and kidney increased in time-dependent manners up to 7.2 and 2-fold, respectively, by the end of the experiment. Interestingly, plasma Fe levels showed a maximum on day 3, then decreased to the control level. The hepatic MT levels showed a transient decrease on day 3, then turned to increase throughout the experiment period, with the final level higher than that in control animals that fed on a normal diet. MT levels in the kidney decreased to nearly 1/3 of the control on day 3, with the values unchanged thereafter. Although Zn levels in liver, plasma and kidney showed a transient but significant reduction on day 3 of the Fe feeding, they recovered to the control values in the later period. Thus, a time-dependent change in the hepatic MT levels was quite similar to that of the tissue Zn levels. The renal Zn levels could not account for the change in the tissue MT levels, at least in the latter period of the experiment. Hybridization analysis of the MT mRNA in the kidney of Fe-fed rats did not differ from control-diet rats, suggesting Fe feeding would not alter synthetic rates of the renal MT. The turnover rates of the renal MT estimated from a time-dependent alteration after its induction by HgCl2 treatment seemed to be enhanced in Fe-fed rats. Thus, the Fe-induced decrease in the renal MT level could possibly be due to an enhanced turnover rate. Despite the reduced renal MT levels, the Fe-treated rats showed no sign of a renal failure indicated by a stable plasma creatinine levels and sustained increase of body weight, suggesting that the Fe-induced suppression of the renal MT levels would not be one of the toxic effects by this metal. Nevertheless, the HgCl2 injection experiment revealed Fe-fed rats had a slightly higher susceptibility against HgCl2 nephrotoxicity than the control-diet rats.

Effect of lead on ALA-D activity, metallothionein levels, and lipid peroxidation in blood, kidney, and liver of the toadfish Halobatrachus didactylus

The effects of lead (Pb) on ALA-D activity, metallothionein (MT) levels, and lipid peroxidation in liver, kidney, and blood of the toadfish Halobatrachus didactylus were investigated. A time-course experiment was performed with sampling on days 0, 2, 5, and 7 following intraperitoneal Pb injection. This indicated a rank order for lead concentration of kidney>liver>blood in fish exposed to Pb. No significant variation of ALA-D activity was observed in liver and kidney while in blood, a slight decrease of ALA-D activity was found but this was not attributed to acute metal stress. Hepatic and renal MT levels were both affected in different ways by metal uptake. The progressive decrease of MDA concentration in the liver and the lack of a clear induction in kidney suggested the hypothesis that Pb is not a good inductor of lipid peroxidation. The histological and histochemical results demonstrated degenerative effects of lead accumulation on the tissues and the activation of lysosomal responses to induced stress.

Hepatic and renal metallothionein induction by an oral equimolar dose of zinc, cadmium or mercury in mice

The hepatic and the renal subcellular distribution of zinc, cadmium or mercury and induction of tissue metallothionein (MT) at 24, 48 and 72 h following an oral equimolar dose (15 μmol metal/kg) of zinc (II) chloride, cadmium (II) chloride or mercury (II) chloride in male albino mice were investigated. There was a moderate increase in hepatic and renal zinc levels mainly in their nuclear mitochondrial fraction (NMF) 24 h post zinc chloride administration. Subsequently, the hepatic zinc increased and the renal zinc declined with time. The zinc-induced hepatic MT level was maximum at 48 h, which decreased slightly thereafter, while there was no marked increase in renal MT level at any time interval. The cadmium was equally distributed in liver and kidney more in their supernatant cytosol fraction (SCF) than in their NMF at 24 h after a dose of cadmium chloride. The cadmium levels showed a decreasing trend in hepatic fractions and an increasing trend in renal fractions with time. The cadmium-induced hepatic and renal MT were substantial at 24 h post cadmium administration, the former decreased thereafter while the latter enhanced at 48 h before declining. The accumulation of mercury in kidney was 1.5 times that in liver, which was localised more in their SCF than in their NMF at 24 h in response to a dose of mercuric chloride. The mercury levels of hepatic and renal subcellular fractions started declining after 24 h and at 72 h they were significantly lower. The induction of hepatic and renal MT was maximum at 24 h after mercuric chloride administration, which declined thereafter concomitant with the decrease in their mercury levels. However, the MT levels in both the organs remained considerably higher than in normal animals at 72 h post exposure. The results show that the accumulation of metal in liver and kidney follows the order: Hg>Cd>Zn and the induction of MT follows Hg>Cd>Zn in liver and Cd>Hg>Zn in kidney. The alterations in zinc and copper homeostasis were more marked in liver than in kidney and follows the order: Hg>Cd>Zn.

Binding properties of an orally active platinum anti-tumor drug JM216 with metallothionein in vivo.

The abilities of the orally active platinum anti-tumor drug JM216 [trans-bis-acetato-cis-dichloroammine (cyclohexylamine) platinum (IV)] to induce the biosynthesis of metallothionein (MT) were investigated in rabbits given oral administrations or injections s.c. It is revealed that oral administration of JM216 can induce the MT synthesis in the liver but not in the kidney. The hepatic MT contained 7.11 +/- 0.11 Zn and only little Pt or Cu. Injections of JM216 to rabbits can greatly elevate the MT levels in the liver, but increase the renal MT levels only slightly. The MT content as well as Pt concentration in the liver was much higher than that in the kidney. The metal stoichiometry in the purified renal MT was determined to be 4.41 +/- 0.04 Zn, 0.36 +/- 0.11 Pt and 2.50 +/- 0.18 Cu per mole protein. The hepatic MT was still characterized as Zn7MT. Both the treatment with oral administration and injections s.c. cannot lead to the increase of Pt content in the kidney. The oxidation state of platinum in the MT from the kidney was determined to be +2 by X-ray photoelectron spectroscopy. As compared with zinc compounds, JM216 was a very poor stimulator for MT biosynthesis in vivo. Pre-injections with Zn(NO3)2 significantly enhanced the MT levels as well as the Pt concentration compared with that resulting from injections with JM216 alone. Based on the experimental data, the role of MT in relation to its involvement in the metabolism and the mechanism of detoxification of Pt(IV) complexes are discussed.

Protection against chronic cadmium toxicity by caloric restriction

Exposure to cadmium (Cd) can result in nephrotoxicity and osteotoxicity. Because Cd-induced nephrotoxicity involves oxidative stress and caloric restriction decreases oxidative stress, we examined whether reduced caloric intake will protect against Cd-induced nephrotoxicity. In addition, the protection against the osteotoxicity was also examined. Male and female Sprague–Dawley rats were provided drinking water containing 100 mg Cd/l. Since fluid intake relative to the body weight was higher in females as compared to the males, the Cd concentration in their water was reduced to 80 mg/l after 3 months and 65 mg/l after 6.5 months. During the 27 month exposure period the males and females consumed a total of about 5 g Cd/kg body weight. Food was restricted to 20 g/day after the first 3 months. During the unrestricted food intake period Cd exposure reduced the bone density in females by 23%, with a partial recovery and stabilization during the caloric restriction phase. Hepatic and renal Cd accumulation and corresponding metallothionein (MT) levels were very similar in both sexes. The reported critical Cd concentration for nephrotoxicity was reached by 9 months. Renal MT levels were maximum at this time. Despite a 1.5-fold increase in renal Cd concentration over the next 18 months, there was no significant increase in renal MT levels. In spite of high renal Cd levels and lack of availability of sufficient MT, there was no sign of nephrotoxicity, as measured by urinary protein and glucose excretion. It is concluded that caloric restriction prevents Cd-induced nephrotoxicity and also appears to control the osteotoxicity of Cd.

Effects of zinc supplementation during chelating agent administration in cadmium intoxication in rats.

The concomitant administration of zinc during chelation of cadmium (Cd) with meso-2,3-dimercaptosuccinic acid (DMSA) or calcium trisodium diethylenetriamine penta-acetic acid (DTPA) was investigated in male rats subchronically exposed to Cd. The results suggest that the administration of zinc alone after cadmium exposure does not elicit any protective effects. However, when supplemented during treatment with DTPA it produced a significant turnover in the altered biochemical variables and a depletion of Cd concentration in liver and kidneys. Cadmium-induced elevated hepatic metallothionein (MT) contents remained practically unaltered on Zn-DTPA administration, but there was a significant increase in renal MT levels compared to rats administered Cd-DTPA or Cd alone.

Age-dependent changes in metallothionein levels in liver and kidney of the Japanese.

Samples of liver, renal cortex, and medulla were obtained from 55 forensic autopsies (0- to 95-yr-old Japanese). Metallothionein (MT) was determined by the Ag-hem or Cd-hem method. Zinc (Zn), copper (Cu), and cadmium (Cd) were determined by atomic absorption spectrophotometry. The mean levels of MT were 250 micrograms/g in the liver, and 394 micrograms/g (cortex) and 191 micrograms/g (medulla) in the kidney. Age-dependent changes were observed in both the liver and kidney. In the liver, MT level decreased during infancy and increased thereafter with age. Similar age-dependent changes in the levels of Zn and Cu were observed. In the kidney cortex, MT level increased with age, although no correlation was found after middle age. The levels of Cd and Zn also increased with age until middle age; however, they decreased thereafter. These results suggest that age-dependent changes in renal MT levels are associated with accumulation of Cd.

TREATMENT WITH THE ANTIANDROGEN CYPROTERONE ACETATE MODIFIES CADMIUM ACCUMULATION AND METALLOTHIONEIN INDUCTION IN C57 AND C3H MICE

With the continuing concern over both steroid compounds and metals as potential environmental pollutants, the interactions between these compounds could become an important health issue. In our continuing study of the interactions of cadmium with various steroids we recently found that testosterone pretreatment protects against cadmium-induced toxicity in C57 mice but has no effect in C3H mice. The basis of this strain-specific acquired tolerance to cadmium is still unclear. In this study, we examined the effects of the antiandrogen cyproterone acetate (CA) on the toxicokinetics and toxicity of cadmium and on the ability of cadmium to induce metallothionein (MT) in male C57 and C3H mice. To assess the tissue accumulation of cadmium, the mice were given CA (10 mg/kg, sc, at 48, 24, and 0 h) prior to cadmium (10 mumol/kg, 0 h). The tissue distribution of cadmium was markedly affected by CA pretreatment, as accumulation of cadmium in liver of both strains was reduced 24 h after cadmium administration. Renal cadmium levels were not affected in either strain by CA. CA pretreatment reduced testicular accumulation of cadmium in C57 mice but not in C3H mice. Cadmium alone, CA alone, or CA and cadmium in combination were not toxic at the doses studied as reflected by the absence of changes in serum glutamic-oxaloacetic transaminase (GOT) activity, lactate dehydrogenase (LDH) activity, and blood urea nitrogen (BUN). CA alone increased MT synthesis in the liver of both strains of mice. However, the cadmium-induced MT level in the livers of both strains of mice was markedly decreased by CA pretreatment. In the C57 mice cadmium-induced renal MT levels were also substantially reduced by CA pretreatment but not in the C3H mice. In addition, the levels of cadmium-binding protein within testes were not affected by CA with or without additional cadmium treatment. These results indicate that, despite the fact that CA will itself induce MT in the liver, pretreatment with CA reduces cadmium induction of hepatic and, in some cases, renal MT synthesis. This reduction may be secondary to a reduced accumulation of cadmium in the liver, but this does not hold true for the kidney.

Influence of seasonal changes in water temperature on cadmium inducibility of hepatic and renal metallothionein in rainbow trout

Changes in metallothionein (MT) levels in liver and kidney of juvenile rainbow trout ( Oncorhynchus mykiss) were followed for 8 months, as the water temperature was allowed to follow the natural temperature decrease in Tvären, a bay in the Baltic Sea. The temperature dropped from 15 °C to 2 °C during the experimental period. The liver and kidney were analysed for MT, zinc, copper and cadmium. Both the hepatic and renal MT levels were found to increase in the control group during the first 4 months of the experiment, as the water temperature dropped to 4 °C. Once the temperature had stabilized at 2 °C there was a noticeable decrease in both hepatic and renal MT levels. One group of fish was exposed to water-borne cadmium (100 μg Cd/litre) for the duration of the experiment. Cadmium accumulated in both liver and kidney, and reached the highest levels in the kidney. Exposure of the fish to cadmium resulted in marked elevations of kidney MT levels. In the liver the MT levels did not increase above the control levels as the temperature dropped to 2 °C after 6 months. However, once the temperature stabilized at 2 °C (after 8 months) the hepatic MT levels were significantly higher in cadmium-exposed fish than in the control fish.

Induction of renal metallothionein synthesis bycis-diamminedichloroplatinum (II) in glutathione-depleted mice

Effects of glutathione (GSH) depletion on nephrotoxicity of cis-diamminedichloroplatinum(II) (cis-DDP) and induction of renal metallothionein (MT) synthesis elicited by cis-DDP treatment has been studied using mice. Pretreatment with L-buthionine-SR-sulfoximine (BSO) reduced the GSH level in the kidneys to 20% of the control level at the time of cis-DDP administration. Administration of 45 μmol/kg cis-DDP did not change the blood urea nitrogen (BUN) level, determined as an indicator of nephrotoxicity, until 96 h after administration, whereas BSO pretreatment raised the BUN level as early as 48 h after cis-DDP (45 μmol/kg) treatment. The level of renal MT was also increased in the GSH-depleted mice and was changed in parallel with the BUN level after cis-DDP treatment. Level of plasma fibrinogen, one of acute phase proteins, increased by cis-DDP in GSH-depleted mice. The increase was reduced by dexamethasone pretreatment, although the renal MT level increased by BSO and cis-DDP was not changed by dexamethasone pretreatment. These results suggest that the induction of renal MT synthesis by cis-DDP in BSO-pretreated mice was not related with the acute phase responses such as cytokines induction. Although the mechanism of the induction of MT synthesis by cis-DDP in GSH-depleted mice is unclear, it is possible that MT is induced as an antidote against oxidative stress, which may be a major cause for nephrotoxicity of cis-DDP enhanced by GSH depletion. © 1995 Wiley-Liss, Inc.

Cadmium disposition and metallothionein induction in mice: strain-, sex-, age-, and dose-dependent differences

The strain- and sex-related differences in tissue cadmium (Cd) and metallothionein (MT) levels were examined in young adult (4–6 months old) C3H/HeJ, DBA/2J, 129/J, CD-1 and A/J mice, 24 h after a subcutaneous (s.c.) injection of 5–30 μgmmol CdCl 2/kg. In many cases the tissue Cd concentrations were inversely related to the tissue weights. Also, the strain and sex differences were more obvious at 20–30-μmol dose levels. At such doses the hepatosensitive C3H males had as much as 30% higher hepatic Cd concentrations as the resistant DBA males, but similar concentrations to the resistant A/J and CD-1 males. This observation suggests that tissue Cd level is not the only determinant of strain differences in hepatotoxicity in males. Among the females, the A/J had 30–47% higher hepatic Cd concentration than the other strains. Livers of male C3H, CD-1 and 129/J mice had 17–57% higher Cd concentrations than those of the females; the greatest difference was in the C3H strain in which only the males exhibited hepatotoxicity at the 30 μmol dose. In all animals the hepatic MT concentration increased with the increasing Cd concentration. However, the 129/J males and all the females reached a plateau in MT concentration at Cd concentrations of 20–30 μg/g liver. Even at the highest Cd concentration, the C3H males had MT concentrations similar to some of the hepatoresistant males, suggesting that their sensitivity to Cd was not due to compromised MT levels. The renal Cd concentration was similar in males of most strains but not in females. For examples, at the 30-μmol dose 129/J females had a Cd concentration which was 70% higher than in the DBA females. Also, at this dose level the A/J, C3H and 129/J females had 30–50% more Cd than the males. The DBA and C3H males had approximately twice the MT concentration of the A/J and 129/J males at 10 μg Cd/g kidney. At similar Cd concentration, the DBA females also had 1.6–2.0 times the MT concentration of the other females. The renal MT levels in females were 1.4–2.9 times higher than in males. Strains susceptible to the testicular toxicity of Cd had up to three times greater testicular Cd accumulation than the resistant strains. However, there was no increase in testicular MT in any strain. The effect of age on Cd and MT levels was studied in 19-month-old A/J and DBA mice at the 25-μmol dose. The aged mice had 15–42% higher hepatic Cd concentration than the younger animals; however, their MT levels were the same. The renal Cd, but not MT, concentration in the older animals was 25–33% lower than in young animals. Despite the larger body weights the Cd-injected aged males had 22–29% smaller testes, but about the same amount of Cd, as the young males, suggesting that the older DBA males may exhibit testicular toxicity at a lower dose of Cd than the young animals. Thus, it is concluded that not only the dose but also the strain, sex and age of the animals are important factors in determining the disposition of Cd in mice. The differences in the tissue Cd accumulation may relate to variations in hormonal and other intinsic factors affecting the cellular uptake and subcellular distribution of Cd and biosynthesis and degradation of MT.

Pregnancy-associated changes in plasma metallothionein concentration and renal cadmium accumulation in rats.

Pregnancy-associated changes in metallothionein (MT) concentrations in blood plasma were examined using a competitive enzyme-linked immunosorbent assay with a rabbit polyclonal antibody to rat liver MT. Plasma MT of pregnant rats significantly increased after 8 days of gestation and remained high during pregnancy and for 7 days after delivery. Gel filtration showed that both Cu and Zn were associated with the plasma MT of pregnant rats. These results suggest that plasma MT may play a role in the transport of essential metals such as Cu and Zn to fetus during pregnancy, but the source of plasma MT is unknown. Injections of cadmium chloride and cadmium-metallothionein to pregnant rats further increased the plasma MT concentrations. After injection of CdCl2, both MT and Cd concentrations in the liver of the pregnant rats were significantly lower than those of the nonpregnant rats, whereas renal Cd and MT levels were higher in pregnant rats. This increased accumulation of Cd in the kidney of pregnant rats may be related to the increase of plasma MT during pregnancy. About 5% of 6 pg of in vitro added Cd (20 pg/ml) was bound to the MT in the plasma. Therefore, Cd may be transported by the circulating MT to the kidney, leading to an increase in renal accumulation of Cd in pregnant rats.

Effect of Fe-Overload on the Biotransformation of Methylmercury in Rat.

It is well documented that methylmercury (MeHg) in vivo slowly undergoes demethylation reaction to change to inorganic mercury (Hg-i). The cleavage of the C-Hg bond was suggested to occur via a reactive oxygen-mediated process in vitro. To study the possibility of the involvement of hydroxyl radical (·OH) in the demethylation of MeHg in vivo, the combined effect of Fe-overload and carbon tetrachloride (CCl4) treatment on the rate of biotransformation of MeHg was examined in rats. The effects of this treatment on H2O2-scavenging enzyme activities (catalase and glutathione peroxidase) were also studied. Rats were fed 3.5% Fe (II) fumalate-containing diet for 0, 7 or 21 d to load Fe. Feeding of the Fe-containing diet brought about a time-dependent increase of the hepatic Fe levels by nearly 7-fold after 3 weeks. The serum Fe levels showed a maximum on day 7, while the renal levels increased after day 7. TBA-reactive substance levels in the liver significantly increased along with the Fe feeding, and this increase was drastically accelerated by CCl4 treatment, suggesting the effective production of ·OH. The CCl4-induced stimulation of the lipid peroxidation was observed also in serum and kidney, though not as marked as in the liver. The hepatic accumulations of total and inorganic Hg at 72 h after MeHg administration were significantly increased by the Fe-feeding. The proportion of Hg-i to total Hg in this tissue increased markedly by the combined effect of Fe and CCl4 with the concomitant decrease of catalase activity. On the other hand, the renal accumulation of Hg-i drastically decreased by the Fe-load, while the MeHg levels remained unchanged. Since the renal metallothionein levels were found to be lowered after the Fe-feeding, this might account for the reduced retention of Hg-i in the whole kidney. The present results suggest that reactive oxygen species, probably ·OH, induced by Fe and CCl4 treatment may play a critical role in the biotransformation of MeHg in the liver.

Sex differences in hepatic and renal cadmium accumulation and metallothionein induction: Role of estradiol

The role of estradiol in sex differences in hepatic and renal cadmium accumulation and metallothionein (MT) induction was investigated. Male and female rats and castrated males pretreated with estradiol were injected either i.v. or s.c. with 10 μmol CdCl 2/kg. Sex differences in cadmium accumulation and MT induction were apparent after s.c. but not i.v. administration. The female rats accumulated a significantly greater concentration of cadmium in their liver than did the males, as early as 1 hr after the s.c. injection. The elevated levels of cadmium in the females were maintained for at least 10 days. Pretreatment of castrated males with estradiol caused a similarly greater accumulation of cadmium in the liver. Hepatic MT levels peaked in the females at 24 hr and in males 48–72 hr after the cadmium injection and then declined to lower levels. This superinduction of MT occurred only after the s.c. administration of cadmium. MT levels in both sexes plateaued 5 days after the s.c. injection to the levels that were similar to those seen in male and female rats 24 hr after an i.v. injection. In animals injected s.c. with cadmium the renal cadmium levels continued to rise for 5–10 days; however, in animals injected i.v. the levels stabilized within 2hr. The renal MT levels in the females were significantly higher than in the males. Estradiol pretreatment induced renal MT but did not affect renal cadmium accumulation. Thus, the sex differences in hepatic cadmium accumulation and MT induction are affected by the route and time after the administration of cadmium. Furthermore, estradiol causes a more rapid uptake of cadmium by the liver and also an enhanced induction of MT in both the liver and kidney.

The apparent lack of effect of supplementary dietary zinc on zinc metabolism and metallothionein concentrations in the turbot, Scophthalmus maximus (Linnaeus)

Turbot were fed either a basal diet containing approximately 100 mg zinc kg−1 or a similar diet supplemented with 1000 mg kg−1; the fish were sampled over a period of 220 days. At final sampling there was no significant difference between control and test animals in hepatic and renal metallothionein levels, hepatic and renal zinc levels and white muscle, skin and bone zinc concentrations.