metallothionein and Body-Weight

The direct transfer of recombinant DNA to embryos is conceptually a powerful method for the manipulation of the genetic potential of domestic animals, but in practice the technology has yet to fulfil its promise. In this paper, two examples are given of research utilising direct genetic manipulation, both of which are aimed at increasing aspects of productivity in sheep. The first of these involves the modification of the growth hormone status of sheep by the use of the ovine growth hormone gene, the regulation of which has been altered by the use of an ovine metallothionein promoter. While there has been a large amount of research already conducted in this area by many groups, our recent results provide one of the first demonstrations that this approach can increase the growth rates of sheep while maintaining the animals in good health. The second project involves the modification of intermediary metabolism in sheep by the introduction of the cysteine biosynthetic pathway. The results to date demonstrate that it is possible to change intermediary metabolism in animals using our approach but that there are species-specific requirements that must be satisfied in order to make the approach a practical method for improving animal productivity.

Topics: Animals; Animals, Domestic; Animals, Genetically Modified; Body Weight; Cell Nucleus; Cysteine; Female; Gene Transfer Techniques; Growth Hormone; Male; Metallothionein; Nuclear Transfer Techniques; Organ Size; Sheep

Mitochondrial diseases (MD), such as Leigh syndrome (LS), present with severe neurological and muscular phenotypes in patients, but have no known cure and limited treatment options. Based on their neuroprotective effects against other neurodegenerative diseases in vivo and their positive impact as an antioxidant against complex I deficiency in vitro, we investigated the potential protective effect of metallothioneins (MTs) in an Ndufs4 knockout mouse model (with a very similar phenotype to LS) crossed with an Mt1 overexpressing mouse model (TgMt1). Despite subtle reductions in the expression of neuroinflammatory markers GFAP and IBA1 in the vestibular nucleus and hippocampus, we found no improvement in survival, growth, locomotor activity, balance, or motor coordination in the Mt1 overexpressing Ndufs4

Topics: Animals; Ataxia; beta 2-Microglobulin; Biomarkers; Body Weight; Disease Models, Animal; Electron Transport Complex I; Female; Hippocampus; Inflammation; Male; Metabolome; Metallothionein; Mice, Knockout; Mitochondrial Diseases; Motor Activity; Oxidation-Reduction; Oxidative Stress; Reactive Oxygen Species; RNA, Messenger; Survival Analysis

Topics: Animals; Body Weight; Copper; Dietary Supplements; Gene Expression; Iron; Liver; Male; Metallothionein; Minerals; Rats; Rats, Wistar; RNA, Messenger; Sodium Hydroxide; Zinc Oxide

This study was to investigate the effect of zinc pectin oligosaccharides chelate (Zn-POS) on growth performance, serum enzyme activities, tissue zinc accumulation, metallothionein (MT) concentrations, and gene expression of zinc transporters (ZnT) in broilers. Five hundred forty 1-d-old Arbor Acres broiler chicks were randomly assigned to 5 dietary groups with 6 replicates of 18 birds per replicate. The diets were formulated with the same supplemental Zn level (80 mg/kg diet) but different amount of the Zn-POS: 0, 200, 400, 600, and 800 mg Zn-POS/kg diet. ZnSO4 was used to adjust to the desired amount of the Zn (80 mg/kg) in the Zn-POS diets. Broilers were fed with the experimental diets for 42 d including the starter (days 1 to 21) and grower (days 22 to 42) phases. Our results showed that dietary supplementation of Zn-POS linearly and quadratically increased (P < 0.05) the average daily gain and gain-to-feed ratio during 22 to 42 d and 1 to 42 d as well as body weight on day 42, whereas reduced (P < 0.05) the sum of mortality and lag abnormalities in broilers on day 42. Besides, serum alkaline phosphatase and copper-zinc superoxide dismutase activities increased (P < 0.05) linearly and quadratically in response to dietary Zn-POS supplemental level on day 42. Dietary Zn-POS supplementation increased Zn accumulation in serum (linear, P < 0.05), liver (linear, P < 0.05), and pancreas (linear and quadratic, P < 0.05). In addition, Zn-POS supplementation linearly and quadratically increased (P < 0.01, P < 0.05, respectively) MT concentrations in liver and pancreas of broilers. Pancreatic mRNA levels of MT, ZnT-1, and ZnT-2 increased (P < 0.05) linearly and quadratically, and the mRNA expression of metal response element-binding transcription factor-1 increased linearly (P < 0.05), in response to dietary Zn-POS supplementation. In conclusion, supplementation of Zn-POS in the diet increases Zn enrichment in the metabolic organs such as liver and pancreas and promotes productive performance in broilers.

Topics: Animal Feed; Animals; Body Weight; Chickens; Diet; Dietary Supplements; Female; Gene Expression Regulation; Glycine max; Liver; Male; Membrane Transport Proteins; Metallothionein; Minerals; Oligosaccharides; Pancreas; Pectins; Random Allocation; Zinc

To evaluate the role of metallothionein (MT) in sex differences of obesity, we examined the effect of MT on regulation of lipid accumulation in female and male wild type (WT) and MT1/MT2-null (MT-KO) mice.. Male and female WT and MT-KO mice fed standard diet (SD) or high-fat diet (HFD) for 35 weeks. Surgical castration in male mice was also performed to examine the effects of androgen on fat accumulation under HFD condition.. The fat mass and size of adipocytes in white adipose tissue (WAT) was greater in adult MT-KO mice than in WT mice after 35 weeks of SD feeding without gender differences, suggesting a role of MT in limiting WAT development during normal growth in both sexes. In female mice fed HFD, weights of WAT and body were greater in MT-KO mice than in WT mice, indicating that MT had a preventive role against excess fat accumulation. In male mice fed HFD, WAT weight hardly increased in MT-KO mice compared to the increase in WT mice. Surgically castrated WT males fed HFD had lower WAT weight compared with sham-treated mice, although castrated MT-KO males fed HFD had greater increases in WAT weight compared with sham-treated mice and castrated WT males.. These data suggest that MT could enhance the preventive action of estrogen against excess fat accumulation, on the contrary, MT augmented the ability of androgen to increase fat accumulation. MT may act to modify the susceptibility to obesity under sex hormones.

Topics: Adipocytes; Adipogenesis; Adipose Tissue, White; Androgens; Animals; Body Weight; Diet, High-Fat; Dietary Fats; Estrogens; Female; Lipid Accumulation Product; Male; Metallothionein; Mice; Mice, 129 Strain; Mice, Knockout; Obesity; Sex Characteristics; Sex Factors

Metallothioneins are proteins that are involved in intracellular zinc storage and transport. Their expression levels have been reported to be elevated in several settings of skeletal muscle atrophy. We therefore investigated the effect of metallothionein blockade on skeletal muscle anabolism

Topics: Animals; Biomarkers; Body Weight; Cell Size; Gene Silencing; Glucocorticoids; Humans; Hypertrophy; Metallothionein; Mice; Muscle Development; Muscle Fibers, Skeletal; Muscle Strength; Muscle, Skeletal; Muscular Atrophy; Organ Size; Proto-Oncogene Proteins c-akt; Rats; Sarcopenia; Signal Transduction; TOR Serine-Threonine Kinases; Up-Regulation; Zinc

This study explored the potential toxicity of Cd on the Zn bone depletion in prenatal bone formation. Female rats received either tap water, Cd, Zn, or Cd + Zn in their drinking water during gestation, and some markers of bone formation were studied in their fetuses removed at the 20th day of pregnancy (GD20). Cd exposure induced maternal hypozincemia and Zn depletion in the femur of the fetuses. A striking inhibition of bone formation in fetuses, expressed by decreases in femur length, width, and area, by the shortening of diaphysis, and by a decrease in length and area of distal and proximal proliferative zones, was observed in fetuses from Cd-exposed mothers. At the molecular level, Cd caused upregulation of MT-1 and ZIP2 genes and significantly depressed the expression of the ZnT5, colα1, osteocalcin, and ALP genes in the femur. Interestingly, Zn treatment ameliorated the Cd-induced maternal hypozincemia and femoral changes and partially restored the normal histomorphometry of the femur. These results suggest that the observed toxic effects of Cd are, at least in part, mediated by the disruption of maternal Zn metabolism during pregnancy leading to Zn depletion and thus to perturbation of prenatal bone formation.

Topics: Animals; Biomarkers; Body Weight; Bone and Bones; Cadmium; Calcium; Female; Femur; Gene Expression Regulation, Developmental; Liver; Male; Maternal Nutritional Physiological Phenomena; Metallothionein; Osteogenesis; Placenta; Pregnancy; Rats, Wistar; Zinc

Obesity continues to rise as an alarming global epidemic. System level mechanisms, diagnostics, and therapeutics are sorely needed so as to identify at risk individuals and design appropriate population scale interventions. The present study evaluated the protective role of metallothioneins (MTs) against obesity and high-fat diet-induced effects such as insulin resistance in both male and female MT-1+2 knockout and MT-3 knockout mice. As the metabolome is closest to the functional phenotype, changes in metabolite levels were also evaluated, and the direct or indirect involvement of MTs in metabolism examined. MT-1+2-, MT-3 knockout, and wild-type mice were given a high-fat diet for 2 months. Variation in body weight gain, tissue weight, and response to oral glucose tolerance test and insulin tolerance test were determined and compared to mice that received the control diet. Effect of the high-fat diet on the knockout mice were investigated on the metabolome level in specific tissues using metabolomics. Both knockout mice strains were more susceptible to high-fat diet-induced effects, such as weight gain and moderate insulin resistance, with the MT-3 knockout mice most susceptible. Brain tissue of the knockout mice showed most metabolic variation and pointed to possible impairment of mitochondrial function. The protective effect of MTs against high-fat diet and obesity-induced effects such as insulin resistance was evident from our observations. The putative role MTs play in mitochondrial function is possibly the main contributor to the lack of these effects in wild-type mice. Considering the expression profiles of the MT isoforms and similarity in brain metabolic variation in the knockout strains, it appears that they promote mitochondrial function in the hypothalamus, thereby limiting weight gain and insulin resistance. Furthermore, metabolomics research in preclinical models of obesity and in the clinic is warranted in the near future.

Topics: Adipose Tissue; Animals; Blood Glucose; Body Weight; Cerebral Cortex; Diet, High-Fat; Disease Models, Animal; Female; Glucose Tolerance Test; Insulin; Insulin Resistance; Male; Metabolome; Metabolomics; Metallothionein; Metallothionein 3; Mice; Mice, Knockout; Obesity

Parameters such as total number of coelomocytes, riboflavin content in coelomocytes, expression of genes implied in metal homeostasis, and detoxification mechanisms can be used as biomarkers to assess the impact of metals on annelids. Defense biomarkers (detoxification gene expressions and coelomocyte parameters) were investigated in the ecotoxicologically important species Eisenia andrei following in vivo exposure to 5 different metals (zinc, copper, nickel, lead, and cadmium) at known concentrations. Coelomocyte numbers and riboflavin content were not affected by metallic exposure, but metal-specific gene expression variations were evidenced.

Topics: Aminoacyltransferases; Animals; Biomarkers; Body Weight; Cadmium; Gene Expression; Metallothionein; Metals, Heavy; Oligochaeta; Phylogeny; Riboflavin; Sequence Analysis, DNA; Soil Pollutants

The role of zinc (Zn) in the protection of germ cells against testicular toxicants has long been elucidated, but the exact molecular mechanisms have not yet been explored. Cyclophosphamide (CP), one of the most commonly used anticancer drugs survived ages of treatment, but the unwanted toxicity limits its clinical usage. The present investigation was aimed to explore the role of Zn and its associated pathways in CP-induced testicular toxicity in S.D. rat. CP was administered in saline 30 mg/kg 5× weekly for 3 weeks (total dose of 450 mg/kg) by i.p. route, while Zn was supplemented by oral route at the doses of 1, 3, 10mg/kg/day for 3 weeks. CP significantly reduced Zn levels in serum and testes, body and testicular weight, sperm count and motility, spermiogenic cells, plasma testosterone and significantly increased the oxidative stress, sperm head abnormalities, sperm DNA damage with decreased chromatin and acrosome integrity; while Zn supplementation ameliorated the same. The present results demonstrated that Zn supplementation protected against CP-induced testicular damages by modulating metallothionein (MT), tesmin and Nrf2 associated pathways. Thus Zn supplementation during anticancer therapy might be potentially beneficial in reducing the off target effects associated with oxidative stress.

Topics: Animals; Antineoplastic Agents, Alkylating; Body Weight; Cyclophosphamide; DNA Damage; Male; Metallothionein; NF-E2-Related Factor 1; Organ Size; Oxidative Stress; Protective Agents; Rats; Rats, Sprague-Dawley; Sperm Motility; Testis; Testosterone; Zinc

Metal concentrations (Cu, Pb, Zn and Cd) were determined in muscle, gills, soft tissues and eggs in male, non-ovigerous and ovigerous female Callinectes sp. specimens from a reference site in Southeastern Brazil. Metallothionein (MT) and reduced glutathione (GSH) levels were also determined. Results demonstrate that sex has a significant influence on metal, MT and GSH concentrations. Significant maternal transfer of Pb and Zn from ovigerous females to eggs was verified, while female crabs, both ovigerous and non-ovigerous, showed elevated GSH and MT in viscera when compared to males, indicating possible MT role in excreting metals to eggs in ovigerous females of this species. Several strong statistical correlations between metals and MT indicate MTs role in detoxification of both toxic and essential elements in different organs. Pb and Zn were significantly correlated to GSH, indicating oxidative stress caused by the former and a direct link between Zn and GSH in maintaining homeostasis. Regarding human consumption, metal concentrations were lower than the maximum permissible levels established by international and Brazilian regulatory agencies, indicating that this species is safe for human consumption concerning this parameter. The presence of metals in Callinectes sp., however, is still of importance considering that this is a key species within the studied ecosystem and, therefore, plays a major role in the transference of pollutants to higher trophic levels. In addition, the presence of significant metal concentrations found in eggs must be considered in this context, since crab eggs are eaten by several other species, such as shorebirds, seabirds, and fish. Also, to the best of our knowledge, this is the first study regarding both MT and GSH levels in Callinectes sp. eggs and is of interest in the investigation of molecular mechanisms regarding metal exposure in these crustaceans. Data reported in this study support the conclusions from previous reports, provide mechanistic insights regarding metal exposure, metallothionein and oxidative stress induction in this species and also present novel data regarding eggs.

Topics: Animals; Body Weight; Brachyura; Brazil; Female; Gills; Glutathione; Inactivation, Metabolic; Male; Metallothionein; Metals, Heavy; Muscles; Ovum; Oxidative Stress; Sex Characteristics

Copper is required for structural and catalytic properties of a variety of enzymes participating in many vital biological processes for growth and development. Feeds provide most of the copper as an essential micronutrient consumed by animals, but inorganic copper could not be utilized effectively. In the present study, we aimed to develop transgenic mouse models to test if copper utilization will be increased by providing the animals with an exogenous gene for generation of copper chelatin in saliva. Considering that the S. cerevisiae CUP1 gene encodes a Cys-rich protein that can bind copper as specifically as copper chelatin in yeast, we therefore constructed a transgene plasmid containing the CUP1 gene regulated for specific expression in the salivary glands by a promoter of gene coding pig parotid secretory protein. Transgenic CUP1 was highly expressed in the parotid and submandibular salivary glands and secreted in saliva as a 9-kDa copper-chelating protein. Expression of salivary copper-chelating proteins reduced fecal copper contents by 21.61% and increased body-weight by 12.97%, suggesting that chelating proteins improve the utilization and absorbed efficacy of copper. No negative effects on the health of the transgenic mice were found by blood biochemistry and histology analysis. These results demonstrate that the introduction of the salivary CUP1 transgene into animals offers a possible approach to increase the utilization efficiency of copper and decrease the fecal copper contents.

Topics: Animal Feed; Animals; Base Sequence; Blotting, Western; Body Weight; Copper; DNA Primers; Metallothionein; Mice; Mice, Transgenic; Polymerase Chain Reaction; Salivary Glands

Cadmium (Cd) is believed to be one of the most abundant and ubiquitously distributed toxins in the aquatic system. This metal is released to the aquatic environment from both anthropogenic sources, such as industrial, agricultural and urban effluents as well as natural sources, such as rocks and soils. Otherwise, the temperature increase of water bodies, which has been observed due to global climatic changes, has been shown to increase Cd toxicity for several aquatic animal species including fish. In the present study, Nile tilapia, Oreochromis niloticus (L.), (26.0 ± 0.38 g) were reared at 20, 24, 28, or 32 °C and exposed to 0.0 or 0.5mg Cd/L for 8 weeks to investigate effects of water temperature, Cd toxicity and their interaction on fish performance as well as metallothionein (MT) and Cd distribution in different fish organs. It was found that fish reared in Cd-free group at 28 °C showed the optimum growth and feed intake, while Cd-exposed fish showed low growth and feed intake irrespective to water temperature. A synergetic relationship between water temperature and Cd toxicity was observed where Cd toxicity increased as water temperature increased and the worse growth was obtained in Cd-exposed fish reared at 32 °C. Additionally, the highest Cd residues in different fish organs were detected in Cd-exposed fish reared at 32 °C. Similarly, MT concentrations in different fish organs increased as water temperature increased especially in Cd-exposed fish groups. A high positive correlation between MT and Cd concentrations in fish organs was detected. The distribution of MT and Cd levels was in the order of liver>kidney>gills>muscles. The present study revealed that the optimum water temperature suitable for Nile tilapia growth is 28 °C. Additionally, Cd exposure had a deteriorate effect on the growth and health of Nile tilapia. This hazardous effect increased as water temperature increased. Further, liver and kidney were the prime sites of Cd accumulation, while Cd load in the muscles was the lowest as compared to the other investigated organs.

Topics: Animals; Body Weight; Cadmium; Gills; Kidney; Liver; Metallothionein; Muscle, Skeletal; Temperature; Tilapia; Tissue Distribution; Water Pollutants, Chemical

Metallothioneins (MTs) are a family of cysteine-rich low molecular-weight proteins that can act as reactive oxygen species scavengers. Although it is known that the induction of MT expression suppresses various inflammatory disorders, the role of MTs in intestinal inflammation remains unclear. In this study, we investigated the effects of dextran sulfate sodium (DSS) administration in mice with targeted deletions of the MT-I/II genes. Acute colitis was induced by 2% DSS in male MT-I/II double knockout (MT-null) and C57BL/6 (wild-type) mice. The disease activity index (DAI) was determined on a daily basis for each animal, and consisted of a calculated score based on changes in body weight, stool consistency and intestinal bleeding. Histology, colon length, myeloperoxidase (MPO) activity and colonic mRNA expression and the concentration of inflammatory cytokines were evaluated by real-time-PCR and enzyme-linked immunosorbent assay (ELISA). The localization of MTs and macrophages was determined by immunohistological and immunofluorescence staining. To investigate the role of MTs in macrophages, peritoneal macrophages were isolated and their responses to lipopolysaccharide were measured. Following DSS administration, the DAI score increased in a time-dependent manner and was significantly enhanced in the MT-I/II knockout mice. Colonic MPO activity levels and inflammatory cytokines [tumor necrosis factor (TNF)-α, interferon (IFN)-γ and interleukin (IL)-17] production increased following DSS administration, and these increases were significantly enhanced in the MT-I/II knockout mice compared with the wild-type mice. MT-positive cells were detected in the lamina propria and submucosal layer by immunohistochemical and immunofluorescence staining, and were mainly co-localized in F4/80-positive macrophages. The production of inflammatory cytokines (TNF-α, IFN-γ and IL-17) from isolated peritoneal macrophages increased following lipopolysaccharide stimulation, and these increases were significantly enhanced in the macrophages obtained from the MT-I/II knockout mice. These data indicate that MTs play an important role in the prevention of colonic mucosal inflammation in a mouse model of DSS-induced colitis, thus suggesting that endogenous MTs play a protective role against intestinal inflammation.

Topics: Animals; Body Weight; Colitis; Colon; Cytokines; Dextran Sulfate; Enzyme-Linked Immunosorbent Assay; Gene Expression Regulation; Immunohistochemistry; Inflammation; Inflammation Mediators; Intestinal Mucosa; Lipopolysaccharides; Macrophages, Peritoneal; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; RNA, Messenger

Cadmium (Cd) is a pollutant that is harmful to human and animals. The liver is a target for Cd accumulation and it can disrupt Zn homeostasis. Here we examined the interaction of Zn and Cd to determine how these two metals could affect δ-aminolevulinate-dehydratase (δ-ALA-D) and metallothionein (MT), two potential molecular endpoints for Cd hepatotoxicity. Cd exposure (0.25 and 1 mg kg1 body weight, i.p., for 10 days) caused a marked increase in hepatic Zn deposition, which was not modified by treatment with Zn (2 mg kg1 , i.p.). Cd caused a dose-dependent increase in hepatic Cd content that was not modified by Zn. Zn and/or Cd treatment increased hepatic δ-ALA-D activity, although the increase caused by Cd was less marked. Reactivation index of δ-ALA-D by DTT was decreased by Zn and Cd exposure, which indicates that Zn protects enzyme from oxidation. Hepatic MT was increased only after exposure to 1 mg kg(-1) Cd and Zn reduced the stimulation of MT synthesis. The results presented here indicate that Cd can redistribute Zn from non-hepatic tissues to liver and the increase in hepatic Zn deposition can account for the increase in hepatic δ-ALA-D activity after Cd exposure. However, Zn blocked the increase in hepatic MT levels caused by Cd. Thus, the modulation of the two molecular endpoints of Cd toxicity used here was distinct, which indicates that the mechanism(s) involved in Zn and Cd distribution, δ-ALA-D and MT regulation are not coincident.

Topics: Analysis of Variance; Animals; Body Weight; Cadmium; Dose-Response Relationship, Drug; Environmental Pollutants; Liver; Male; Metallothionein; Porphobilinogen Synthase; Rats; Rats, Wistar; Zinc

To evaluate the potential toxicity of copper (Cu) in raptors that may consume Cu bullets, shotgun pellets containing Cu, or Cu fragments as they feed on wildlife carcasses, we studied the effects of metallic Cu exposure in a surrogate, the American kestrel (Falco sparverius). Sixteen kestrels were orally administered 5 mg Cu/g body mass in the form of Cu pellets (1.18-2.00 mm in diameter) nine times during 38 days and 10 controls were sham gavaged on the same schedule. With one exception, all birds retained the pellets for at least 1 h, but most (69%) regurgitated pellets during a 12-h monitoring period. Hepatic Cu concentrations were greater in kestrels administered Cu than in controls, but there was no difference in Cu concentrations in the blood between treated and control birds. Concentration of the metal-binding protein metallothionein was greater in male birds that received Cu than in controls, whereas concentrations in female birds that received Cu were similar to control female birds. Hepatic Cu and metallothionein concentrations in kestrels were significantly correlated. Histopathologic alterations were noted in the pancreas of four treated kestrels and two controls, but these changes were not associated with hepatic or renal Cu concentrations, and no lesions were seen in other tissues. No clinical signs were observed, and there was no treatment effect on body mass; concentrations of Cu, hemoglobin, or methemoglobin in the blood; or Cu concentrations in kidney, plasma biochemistries, or hematocrit. Based on the parameters we measured, ingested Cu pellets pose little threat to American kestrels (and presumably phylogenetically related species), although the retention time of pellets in the stomach was of relatively short duration. Birds expected to regurgitate Cu fragments with a frequency similar to kestrels are not likely to be adversely affected by Cu ingestion, but the results of our study do not completely rule out the potential for toxicity in species that might retain Cu fragments for a longer time.

Topics: Administration, Oral; Animals; Body Weight; Copper; Falconiformes; Female; Firearms; Food Contamination; Kidney; Liver; Male; Metallothionein; Pancreas

Bank voles free living in a contaminated environment have been shown to be more sensitive to cadmium (Cd) toxicity than the rodents exposed to Cd under laboratory conditions. The objective of this study was to find out whether benzo(a)pyrene (BaP), a common environmental co-contaminant, increases Cd toxicity through inhibition of metallothionein (MT) synthesis-a low molecular weight protein that is considered to be primary intracellular component of the protective mechanism. For 6 weeks, the female bank voles were provided with diet containing Cd [less than 0.1 μg/g (control) and 60 μg/g dry wt.] and BaP (0, 5, and 10 μg/g dry wt.) alone or in combination. At the end of exposure period, apoptosis and analyses of MT, Cd, and zinc (Zn) in the liver and kidneys were carried out. Dietary BaP 5 μg/g did not affect but BaP 10 μg/g potentiated rather than inhibited induction of hepatic and renal MT by Cd, and diminished Cd-induced apoptosis in both organs. The hepatic and renal Zn followed a pattern similar to that of MT, attaining the highest level in the Cd + BaP 10-μg/g group. These data indicate that dietary BaP attenuates rather than exacerbates Cd toxicity in bank voles, probably by potentiating MT synthesis and increasing Zn concentration in the liver and kidneys.

Topics: Animals; Apoptosis; Arvicolinae; Benzo(a)pyrene; Body Weight; Cadmium; Dietary Supplements; Drug Synergism; Environmental Pollutants; Female; Immunohistochemistry; In Situ Nick-End Labeling; Kidney; Liver; Metallothionein; Organ Size; Time Factors; Zinc

Microcystins (MC) are usually the predominant cyanotoxins associated with cyanobacterial blooms in natural surface waters. These toxins are well-known hepatotoxic agents that proceed by inhibiting protein phosphatase in aquatic biota; recent studies have also reported oxidative stress and disruption of ion regulation in aquatic organisms. In the present study, young trout (Oncorhynchus mykiss) were exposed to crude extracts of Microsystis aeruginosa for four days at 15 °C. The level of microcystins was calculated to confirm the presence of toxins in these crude extracts: 0, 0.75, 1.8 and 5 μg/L. Protein phosphatase measured in the liver increased by at least 3-fold and is significantly as a result of exposure to these sublethal concentrations of crude extract, his indicates an early defense response against protein phosphatase inhibition from cyanotoxins. This was corroborated by the decreased phosphate content in proteins found in the liver and brain. No increase in glutathione-S transferase (GST) activity was observed and lipid peroxidation was unaffected in both liver and brain tissue exposed to the cyanobacterial extracts. The data revealed that the proportion of the reduced (metal-binding) form of metallothionein (MT) decreased by two-fold relative to the control group (with a concomitant increase in the proportion of the oxidized form). The level of phosphate associated with MT increased by 1.5-fold at the highest concentration of crude extract. Acetylcholinesterase (AChE) activity in brain tissue was decreased after exposure to the highest concentration of crude extract, suggesting a slowdown in neural activity. However, no biotransformation processes or detoxification of GST was triggered. Our findings show early sign of biochemical effects of MC-LR in young trout.

Topics: Acetylcholinesterase; Animals; Body Weight; Brain; Complex Mixtures; Liver; Metallothionein; Microcystis; Oncorhynchus mykiss; Phosphoprotein Phosphatases; Toxicity Tests

Alzheimer's disease (AD) is by far the most commonly diagnosed dementia, and despite multiple efforts, there are still no effective drugs available for its treatment. One strategy that deserves to be pursued is to alter the expression and/or physiological action of endogenous proteins instead of administering exogenous factors. In this study, we intend to characterize the roles of the antioxidant, anti-inflammatory, and heavy-metal binding proteins, metallothionein-1 + 2 (MT1 + 2), in a mouse model of Alzheimer's disease, Tg2576 mice. Contrary to expectations, MT1 + 2-deficiency rescued partially the human amyloid precursor protein-induced changes in mortality and body weight in a gender-dependent manner. On the other hand, amyloid plaque burden was decreased in the cortex and hippocampus in both sexes, while the amyloid cascade, neuroinflammation, and behavior were affected in the absence of MT1 + 2 in a complex manner. These results highlight that the control of the endogenous production and/or action of MT1 + 2 could represent a powerful therapeutic target in AD.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Animals; Behavior, Animal; Body Weight; Disease Models, Animal; Female; Hippocampus; Humans; Male; Metallothionein; Mice; Mice, Knockout; Mice, Transgenic; Peptide Fragments

To examine the effect of increasing dietary zinc (Zn) intake and the lack of metallothionein (MT) expression on activity of small intestinal disaccharidases.. MT-I and II knockout (MT-/-) and wild-type (MT+/+) female mice at 3.5 wk of age were randomly fed with a diet containing 2 (2 Zn), 15 (15 Zn) or 50 (50 Zn) mg Zn/kg (n = 8/group/genotype) for 5 wk. Small intestinal segments (duodenum, jejunum and ileum) were collected and either fixed in 10% formalin for histological analysis or snap frozen in liquid nitrogen for sucrase, lactase and maltase activity analyses.. Plasma Zn was significantly (P < 0.05) lower (33%) in MT-/- compared with MT+/+ mice fed the 2 Zn diet. Villus height and crypt depth were increased by approximately 15% in MT+/+ mice compared with MT-/- mice. Duodenal disaccharidase activities were significantly higher in MT+/+ compared with MT-/- mice particularly in those fed the 2 Zn diet. For the 50 Zn diet, jejunal sucrase and lactase activities were significantly higher in MT-/- (13 313 ± 2314; 4107 ± 364 μmol glucose/well/min/g tissue, respectively) compared with MT+/+ mice (7054 ± 608; 1818 ± 174). Similarly, ileal lactase activities were higher in MT-/- (1480 ± 192) compared with MT+/+ (629 ± 353) mice particularly those fed the 2 Zn diet.. Increasing dietary Zn has little effect on disaccharidases activity in MT wild-type mice. The presence of MT may enhance morphological and functional development of the gut.

Topics: Animals; Body Weight; Diet; Disaccharidases; Female; Intestine, Small; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Organ Size; Random Allocation; Zinc

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.

Topics: Animals; Arsenic; Body Weight; Drugs, Chinese Herbal; Glutathione; Glutathione Peroxidase; Kidney; Lead; Male; Malondialdehyde; Metallothionein; Rats; Rats, Sprague-Dawley; Skin; Superoxide Dismutase

The interactive effects of mixed pollutants in sewage wastewater on biomarker responses were investigated using wild male African sharptooth catfish (Clarias gariepinus) in Morogoro, Tanzania. A total of 58 fish were used, of which 21 were from Mindu dam (reference site) and 22, 9 and 10 from Mafisa, Mazimbu and Mzumbe sewage ponds, respectively. Liver somatic index (LSI) and gonadosomatic index (GSI) were significantly greater (two- to threefold) and (five- to sixfold), respectively, in fish from all sewage ponds. Haemoglobin concentration and gill filament 7-ethoxyresurufin O-deethylase (EROD) activities were significantly higher (1.2-fold and twofold, respectively) in fish from Mzumbe sewage ponds than in fish from Mindu dam, whereas liver EROD activity was significantly higher in fish from Mzumbe and Mafisa sewage ponds (5-fold). A HPLC method for determination of enzymatically formed p-nitrophenyl-glucuronide (PNPG) was developed and applied to measure UDP-glucuronosyl transferase (UGT) activities that was significantly higher in fish from all sewage ponds (2-2.5-fold) than in fish from Mindu dam. Kinetic characteristics and assay dependence of UGT were studied with microsomal preparations. Metallothionein (MT) content was significantly lower (three- to fourfold) in fish from sewage ponds than in fish from Mindu dam, and corresponded with cumulative levels of cadmium, lead and mercury. Condition factor, vitellogenin (Vtg), acetylcholinesterase (AChE) activities in plasma, eyes and brain, haematocrit, plasma protein and cytosolic glutathione S-transferase (GST) activities were comparable in fish from sewage ponds and Mindu dam. Although specific pollutants other than the metals were not identified by chemical analysis, application of a suite of biomarkers in C. gariepinus demonstrated that all sewage ponds were contaminated by pollutants of public health concern.

Topics: Acetylcholinesterase; Animals; Biomarkers; Body Weight; Brain; Catfishes; Chromatography, High Pressure Liquid; Cytochrome P-450 CYP1A1; Environmental Monitoring; Eye; Fish Proteins; Gills; Glucuronosyltransferase; Glutathione Transferase; Gonads; Hemoglobins; Liver; Male; Metallothionein; Metals, Heavy; Microsomes, Liver; Principal Component Analysis; Risk Assessment; Sewage; Tanzania; Vitellogenins; Water Pollutants, Chemical

Traumatic injury to the brain is one of the leading causes of injury-related death or disability, but current therapies are limited. Previously it has been shown that the antioxidant proteins metallothioneins (MTs) are potent neuroprotective factors in animal models of brain injury. The exogenous administration of MTs causes effects consistent with the roles proposed from studies in knock-out mice. We herewith report the results comparing full mouse MT-1 with the independent alpha and beta domains, alone or together, in a cryoinjury model. The lesion of the cortex caused the mice to perform worse in the horizontal ladder beam and the rota-rod tests; all the proteins showed a modest effect in the former test, while only full MT-1 improved the performance of animals in the rota-rod, and the alpha domain showed a rather detrimental effect. Gene expression analysis by RNA protection assay demonstrated that all proteins may alter the expression of host-response genes such as GFAP, Mac1 and ICAM, in some cases being the beta domain more effective than the alpha domain or even the full MT-1. A MT-1-to-MT-3 mutation blunted some but not all the effects caused by the normal MT-1, and in some cases increased its potency. Thus, splitting the two MT-1 domains do not seem to eliminate all MT functions but certainly modifies them, and different motifs seem to be present in the protein underlying such functions.

Topics: Animals; Body Weight; Brain Injuries; Disease Models, Animal; Gene Expression Regulation; Metallothionein; Metallothionein 3; Mice; Mice, Knockout; Motor Activity; Mutation; Nerve Tissue Proteins; Protein Structure, Tertiary; Psychomotor Performance

Cadmium (Cd), one of the most widely distributed heavy metals, is highly toxic to humans and animals. It is well known that zinc (Zn) and selenium (Se) administration reduce the Cd-induced toxicity and that metallothioneins can have a protective effect to mitigate Cd toxicity in biological systems. In this study we report the expression analysis of the two metallothioneines gene classes MT-1 and MT-2 as well as the total metalloprotein content in the liver of rats exposed to Cd (200 ppm), Cd + Zn (200 ppm + 500 ppm), Cd + Se (200 ppm + 0.1 ppm) or Cd + Zn + Se (200 ppm + 500 ppm + 0.1 ppm) in their drinking water for 35 days. Metals accumulation was quantified in rat liver. Cd decreased significantly the hepatic concentrations of Se and increased those of Zn. The treatment of Cd-exposed rats with Se alone or combined with Zn reversed the Cd-induced depletion of Se concentrations in the liver. However, Zn or Zn + Se administration significantly increased the liver Cd uptake and had no effect on the Cd-induced increase in hepatic concentrations of Zn. The molecular assay showed a decreasing trend of MT-1 relative gene expression levels in animals supplemented with Zn (6.87-fold), Se (3.58-fold), and their combination (1.69-fold) when compared to Cd-treated animals (16.22-fold). Upregulation of the MT-2 expression were recorded in all conditions, although fold induction levels were less pronounced than MT-1 expressions. Our data suggest that the well-established protective effect of Zn and Se against Cd-induced toxicity passes through non-MT gene expression mechanisms, being more dependent on the oxidative stress status of the cell.

Topics: Animals; Body Weight; Cadmium; Dietary Supplements; Drug Interactions; Environmental Pollutants; Gene Expression; Liver; Male; Metallothionein; Organ Size; Protective Agents; Rats; Rats, Wistar; Selenium; Zinc

Metals like iron (Fe) and copper (Cu) function as integral components in many biological reactions, and, in excess, these essential metals are toxic, and organisms must control metal acquisition and excretion. We examined the effects of chronic waterborne Cu exposure and the interactive effects of elevated dietary Fe on gene expression and tissue metal accumulation in zebrafish. Softwater acclimated zebrafish exposed to 8 microg/l Cu, with and without supplementation of a diet high in Fe (560 vs. 140 mg Fe/kg food) for 21 days demonstrated a significant reduction in liver and gut Cu load relative to waterborne Cu exposure alone. Gene expression levels for divalent metal transport (DMT)-1, copper transporter (CTR)-1, and the basolateral metal transporter ATP7A in the gills and gut increased when compared with controls, but the various combinations of Cu and high-Fe diet revealed altered levels of expression. Further examination of the basolateral Fe transporter, ferroportin, showed responses to waterborne Cu exposure in the gut and a significant increase with Fe treatment alone in the liver. Additionally, we examined metallothionein 1 and 2 (MT1 and MT2), which indicated that MT2 is more responsive to Cu. To explore the relationship between transcription and protein function, we examined both CTR-1 protein levels and gill apical uptake of radiolabeled Cu64, which demonstrated decreased Cu uptake and protein abundance in the elevated Cu treatments. This study shows that high dietary Fe can significantly alter the genetic expression pattern of Cu transporters at the level of the gill, liver, and gastrointestinal tract.

Topics: Acclimatization; Adenosine Triphosphatases; Animals; Body Weight; Cation Transport Proteins; Copper Radioisotopes; Copper Sulfate; Copper Transporter 1; Copper-Transporting ATPases; Female; Gastrointestinal Tract; Gene Expression Regulation; Gills; Iron, Dietary; Liver; Male; Membrane Transport Proteins; Metallothionein; Transcription, Genetic; Water Pollutants, Chemical; Zebrafish; Zebrafish Proteins

To investigate the effect of repeated stress on DNA damage in seven organs of dyslipidemic mice, and the preventive role of metallothionein (MT).. Female adult 129/Sv wild-type and MT-null mice fed high-fat diet (HFD) were repeatedly subjected to mild stress of fasting or restraint in weeks 2 to 4 of 4-week study period. Serum cholesterol level, DNA damage in the liver, pancreas, spleen, bone marrow, kidney, lung and gastric mucosa, and other parameters were determined.. Body weights were increased in both types of mice fed HFD compared to those fed standard diet (STD), and further increased by 12 h-fasting, while they were markedly decreased by 1-3 h-restraint. Fasting accelerated accumulation of fat in the liver, and increase in serum cholesterol of both types of mice fed HFD. Feeding of HFD increased DNA damage in the pancreas, spleen and bone marrow of both types of mice, compared with those fed STD. In the wild-type mice fed HFD, 24 h-fasting increased DNA damage in the liver and spleen, while restraint increased the damage in the liver, pancreas, spleen and bone marrow. DNA damage in the cells of organs was markedly increased in the MT-null mice. Specifically, damage in the liver, pancreas, spleen and bone marrow was greatly increased with the intensity of stress increased, and the damage was much greater in the restraint mice than in the fasting mice.. MT plays a tissue-dependent preventive role against DNA damage in various murine organs induced by repeated stress.

Topics: Animals; Body Weight; Comet Assay; Dietary Fats; DNA Damage; Dyslipidemias; Fasting; Female; Glutathione; Immobilization; Liver; Metallothionein; Mice; Mice, Knockout; Organ Size; Pancreas; Stress, Physiological

Mycotoxins are secondary metabolites produced by microfungi that are capable of causing disease and death in humans and other animals. This work was aimed at investigation of influence of mouldy wheat contaminated by pathogenic fungi producing mycotoxins on metallothionein levels in hepatic tissue of rats. The rats were administrating feed mixtures with different contents of vitamins or naturally mouldy wheat for 28 days. It was found that the wheat contained deoxynivalenol (80 +/- 5 microg per kg of mouldy wheat), zearalenone (56 +/- 3 microg/kg), T2-toxin (20 +/- 2 microg/kg) and aflatoxins as a sum of B1, B2, G1 and G2 (3.9 +/- 0.2 microg/kg). Rats were fed diets containing 0, 33, 66 and 100% naturally moulded wheat. Control group 0, 33, 66 and 100% contained vitamins according to Nutrient Requirements of Rats (NRC). Other four groups (control group with vitamins, vit33, vit66 and vit100%) were fed on the same levels of mouldy wheat, also vitamins at levels 100% higher than the previous mixtures. We determined weight, feed conversion and performed dissection to observe pathological processes. Changes between control group and experimental groups exposed to influence of mouldy wheat and experimental groups supplemented by higher concentration of vitamins and mouldy wheat were not observed. Livers were sampled and did not demonstrate significant changes in morphology compared to control either. In the following experiments the levels of metallothionein as a marker of oxidative stress was determined. We observed a quite surprising trend in metallothionein levels in animals supplemented with increased concentration of vitamins. Its level enhanced with increasing content of mouldy wheat. It was possible to determine a statistically significant decline (p<0.05) between control group and groups of animals fed with 33, 66 and 100% mouldy wheat. It is likely that some mycotoxins presented in mouldy wheat are able to block the mechanism of metallothionein synthesis.

Topics: Aflatoxins; Animals; Body Weight; Fungi; Liver; Male; Metallothionein; Oxidative Stress; Rats; Rats, Wistar; T-2 Toxin; Trichothecenes; Triticum; Vitamins; Zearalenone

The biochemical mechanisms of Chlorella vulgaris protection against cadmium (Cd)-induced liver toxicity were investigated in male Sprague-Dawley rats (5 weeks of age, weighing 90-110 g). Forty rats were randomly divided into one control and three groups treated with 10 ppm Cd: one Cd without Chlorella (Cd-0C), one Cd with 5% Chlorella (Cd-5C), and one Cd with 10% Chlorella (Cd-10C) groups. The rats had free access to water and diet for 8 weeks. Body weight gain and relative liver weight were significantly lower in the Cd-0C group than in Cd-5C and Cd-10C groups. Rats in the Cd-0C group had significantly higher hepatic concentrations of Cd and metallothioneins (MTs) than in the Cd-5C or Cd-10C group. The hepatic MT I/II mRNA was expressed in all experimental rats. MT II was more expressed in the Cd-5C and Cd-10C groups than in the Cd-0C group. Morphologically, a higher level of congestion and vacuolation was observed in the livers of the Cd-0C group compared to those of the Cd-5C and Cd-10C groups. Therefore, this study suggests that C. vulgaris has a protective effect against Cd-induced liver damage by reducing Cd accumulation and stimulating the expression of MT II in liver. However, the details of the mechanism of C. vulgaris on liver toxicity remains to be clarified by further studies.

Topics: Animals; Body Weight; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Chlorella vulgaris; Dietary Supplements; Hepatocytes; Hyperemia; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Powders; Rats

Free-living bank voles have been shown to be more sensitive to cadmium (Cd) toxicity than the rodents exposed to Cd under laboratory conditions. The present study was designed to find out whether polychlorinated biphenyls (PCBs), common environmental co-contaminants, increase susceptibility to Cd toxicity through inhibition of metallothionein (MT) synthesis-a low molecular weight protein that is considered to be a primary intracellular component of the protective mechanism. For 12 weeks, the male bank voles were provided with diets containing Cd (0.05 microg/g (control) and 10 microg/g dry wt) and PCBs (0, 10 and 50 microg/g dry wt) alone or in combination under long (16 h) and short (8 h) photoperiods. At the end of exposure period, histological examinations and analyses of MT, Cd, Fe and lipid peroxidation in the kidneys and liver were carried out. Dietary PCBs did not affect Cd inducibility of renal MT, but decreased it significantly in the liver; however, no signs of Cd toxicity (measured by histopathology) occurred in both organs. On the contrary, PCBs at the highest dose increased significantly lipid peroxidation in the kidneys and liver (4-fold) only in the bank voles raised under a long photoperiod; the PCB-induced hepatic lipid peroxidation was accompanied by extensive histopathological changes including hepatocyte enlargement, necrosis and steatosis. Co-treatment with dietary Cd significantly suppressed the increase in lipid peroxidation and apparently reduced hepatic damage. These data indicate that (1) dietary PCBs do not enhance Cd toxicity in the kidneys and liver of bank voles and (2) dietary Cd suppresses PCB-induced hepatotoxicity that appears to be photoperiod-dependent.

Topics: Animal Feed; Animals; Arvicolinae; Body Weight; Cadmium; Diet; Kidney; Lipid Peroxidation; Liver; Metallothionein; Organ Size; Polychlorinated Biphenyls

The potential use of metallothioneins (MTs) as biomarkers of trace metal contamination was evaluated for the first time in the Algerian mouse (Mus spretus). Mice were collected seasonally in an abandoned mining area (Aljustrel) and in a reference area, both located in southern Portugal. MT levels were quantified in liver and kidney by differential pulse polarography and hepatic elemental concentrations (Mn, Fe, Cu, Zn, Se) were determined by particle-induced X-ray emission. Hepatic iron and selenium concentrations were elevated in mice from Aljustrel mine when compared to reference animals. MTs levels were averagely higher in mice from Aljustrel than those originated from the reference area. A season-dependent significant effect was found on the hepatic and renal MT concentrations, characterized by higher levels in winter and lower in autumn. In contaminated mice positive relationship between liver elemental contents (Cu in autumn and Fe in winter) and MTs were found. The seasonal variation of MT suggests that probably physiological and environmental factors could influence hepatic and renal MT induction. Results seem to imply that some environmental disturbance occur in the vicinity of the Aljustrel mine. Therefore, for the management purposes MT levels should be followed in liver of M. spretus, especially in winter. Furthermore, other physiological factors that could influence MT expression and turnover in Algerian mouse should also be monitored.

Topics: Animals; Biomarkers; Body Weight; Ecosystem; Environmental Monitoring; Kidney; Liver; Metallothionein; Metals, Heavy; Mice; Mining; Organ Size; Portugal; Seasons; Soil Pollutants

In rats fed a high fat diet (HFD), long-term administration of 3,5-diiodo-L-thyronine (T2), a naturally occurring iodothyronine, was shown to reduce body-weight gain, fat mass, and hepatic lipid accumulation. This work was aimed at investigating the mechanisms of T2 action in the liver of HFD rats. The results show that HFD induces liver lipid peroxidation and stimulates the activity of enzymes involved in hydrogen peroxide (H2O2) metabolism, catalase in particular. Moreover, quantitative RT-PCR revealed HFD-induced upregulation of the transcription factor PPAR alpha, as well as of metallothionein isoforms (MT-1 and MT-2). T2 administration prevented the HDF-induced lipid peroxidation, as well as the increase in H2O2 metabolism, and reduced the upregulation of both PPAR alpha and MT-2. These data demonstrate that in the liver of HFD rats, T2 prevents both lipid accumulation and oxidative stress associated with increased fat metabolism.

Topics: Animal Feed; Animals; Body Weight; Cell Shape; Dietary Fats; Diiodothyronines; Lipid Peroxidation; Lipids; Liver; Male; Metallothionein; Oxidative Stress; PPAR alpha; Rats; Rats, Wistar; Transcription, Genetic

There is clinical evidence linking asthma with the trace element, zinc (Zn). Using a mouse model of allergic inflammation, we have previously shown that labile Zn decreases in inflamed airway epithelium (Truong-Tran AQ, Ruffin RE, Foster PS, Koskinen AM, Coyle P, Philcox JC, Rofe AM, Zalewski PD. Am J Respir Cell Mol Biol 27: 286-296, 2002). Moreover, mild nutritional Zn deficiency worsens lung function. Recently, a number of proteins belonging to the Solute Carrier Family 39 (ZIP) and Solute Carrier Family 30 (ZnT) have been identified that bind Zn and regulate Zn homeostasis. Mice were sensitized, and subsequently aerochallenged, with ovalbumin to induce acute and chronic airway inflammation. Mice received 0, 54, or 100 microg of Zn intraperitoneally. Tissues were analyzed for Zn content and histopathology. Inflammatory cells were counted in bronchoalveolar lavage fluid. Cytokine and Zn transporter mRNA levels were determined by cDNA gene array and/or real-time PCR. Zn supplementation decreased bronchoalveolar lavage fluid eosinophils by 40 and 80%, and lymphocytes by 55 and 66%, in the acute and chronic models, respectively. Alterations in Zn transporter expression were observed during acute inflammation, including increases in ZIP1 and ZIP14 and decreases in ZIP4 and ZnT4. Zn supplementation normalized ZIP1 and ZIP14, but it did not affect mRNA levels of cytokines or their receptors. Our results indicate that inflammation-induced alterations in Zn transporter gene expression are directed toward increasing Zn uptake. Increases in Zn uptake may be needed to counteract the local loss of Zn in the airway and to meet an increased demand for Zn-dependent proteins. The reduction of inflammatory cells by Zn in the airways provides support for Zn supplementation trials in human asthmatic individuals.

Topics: Animals; Anti-Inflammatory Agents; Body Weight; Bronchoalveolar Lavage Fluid; Cation Transport Proteins; Cytokines; Dietary Supplements; Disease Models, Animal; Female; Gene Expression Regulation; Inflammation; Inflammation Mediators; Lung; Metallothionein; Mice; Receptors, Cytokine; Respiratory Hypersensitivity; RNA, Messenger; Zinc

Detection and assessment of the impact of pollution on biological resources imply increasing research on early-warning markers such as metallothioneins in metal exposure. Metallothioneins are cytosolic, low molecular weight proteins, involved principally in essential metal homeostasis and non-essential metal detoxication. Metallothionein synthesis could be influenced by abiotic (season) or biotic (reproduction process) factors directly or indirectly by its effect on metal bioaccumulation (i.e., sex, weight). In a view of using metallothioneins as metal-exposure biomarkers in Gammarus pulex, this study attempts to define the effect of several factors (sex, weight/size and season) on the level of this protein. Metallothionein levels recorded in individuals over a large range of weights indicate a negative correlation between them. Inversely in our conditions, no difference was observed between male and female organisms. During field study, metallothionein level changes were observed with the highest levels in autumn and winter periods. The highest metallothionein levels were observed after the reproduction period, perhaps linked with the metabolic needs of biologically available essential metal such as zinc.

Topics: Amphipoda; Animals; Body Weight; Cadmium; Copper; Environmental Monitoring; Female; Male; Metallothionein; Seasons; Sex Factors

An herbal health care supplement, St John's Wort (SJW, Hypericum perforatum) has become widely used in the treatment of depression, and is known to interact with therapeutic drugs. Here we report a preventive effect of SJW on cisplatin nephrotoxicity in rats. Rats were given SJW (400 mg/kg/day, p.o.) for 10 consecutive days, and were injected with cisplatin (5 mg/kg, i.v.) on the day after the final SJW treatment. Cisplatin treatment increased the serum creatinine level, which is an index of nephrotoxicity, to 1.51+/-0.22 mg/dl (mean+/-SE) from 0.28+/-0.05 mg/dl (control) on day 5 after the cisplatin injection. This increase fell significantly to 0.86+/-0.13 mg/dl by pre-treatment with SJW. Cisplatin-induced histological abnormality of the kidney was blocked by pre-treatment with SJW. When SJW was administered for 10 days, the amounts of renal metallothionein (MT) and hepatic multidrug resistance protein 2 (Mrp2) were increased to 164.8+/-13.0% and 220.8+/-39.3% (mean+/-SE) of controls, respectively. GSH levels in the kidney and liver were not changed. Total and free cisplatin concentration in serum was not influenced by SJW treatment. In conclusion, the results suggest that pre-treatment with SJW may diminish cisplatin nephrotoxicity.

Topics: Animals; Antineoplastic Agents; ATP-Binding Cassette Transporters; Body Weight; Cell Membrane; Cisplatin; Creatinine; Drug Interactions; Glutathione; Hypericum; Kidney; Kidney Diseases; Male; Metallothionein; Rats; Rats, Wistar

The biochemical and histological sequelae resulting from a diet containing 50.20 mg cadmium/kg were studied in Lohmann brown cockerels from hatching until 30 days of age. The additional cadmium chloride (CdCl(2)) to the diet induced the formation of lipid peroxides, which via a chain reaction led to accumulation of malondialdehyde in intestinal mucosa. At the end of the study (after 30 days of cadmium exposure) total protein and metallothionein levels in the intestinal mucosa and the relative ileal and duodenal weight increased. Histological data show that CdCl(2) causes an increase in number of goblet cells and granular lymphocytes in the intestinal mucosa. Down-regulation of the serotonin-positive cells in the cadmium-treated animals was observed. Growth retardation (by 27%) occurred in chicken fed the cadmium-enriched diet for 30 days. Cadmium accumulation in the intestine was markedly higher (154 times) in the cadmium-treated animals compared to the control group. Cadmium induced a decrease in zinc (but not copper) content in intestinal mucosa. We suggest that cadmium uptake triggers an inflammatory and secretory response in chicken small intestine.

Topics: Animals; Body Weight; Cadmium Chloride; Chickens; Intestinal Diseases; Intestinal Mucosa; Lipid Peroxides; Male; Malondialdehyde; Metallothionein; Poultry Diseases

Cadmium (Cd) exposure results in injury to the proximal tubule characterized by polyuria and proteinuria. Kidney injury molecule-1 (Kim-1) is a transmembrane glycoprotein not normally detected in the mature kidney, but is upregulated and shed into the urine following nephrotoxic injury. In this study, we determine if Kim-1 might be a useful early biomarker of Cd nephrotoxicity. Male Sprague-Dawley rats were given daily injections of Cd for up to 12 weeks. Weekly urine samples were analyzed for Kim-1, protein, creatinine, metallothionein, and Clara cell protein CC-16. Significant levels of Kim-1 were detected in the urine by 6 weeks and continued to increase throughout the treatment period. This appearance of Kim-1 occurred 4-5 weeks before the onset of proteinuria, and 1-3 weeks before the appearance of metallothionein and CC-16. Higher doses of Cd gave rise to higher Kim-1 excretion. Reverse transcriptase-polymerase chain reaction (RT-PCR) expression analysis showed that Kim-1 transcript levels were increased after 6 weeks at the low dose of Cd. Immunohistochemical analysis showed that Kim-1 was present in proximal tubule cells of the Cd-treated rats. Our results suggest that Kim-1 may be a useful biomarker of early stages of Cd-induced proximal tubule injury.

Topics: Animals; Biomarkers; Body Weight; Cadmium; Cell Adhesion Molecules; Dose-Response Relationship, Drug; Kidney Tubules, Proximal; Male; Membrane Proteins; Metallothionein; Proteinuria; Rats; Rats, Sprague-Dawley; Uteroglobin

To investigate the inhibitive effect of metallothionein (MT) on DOX-induced cardiac apoptosis and the involved possible mechanisms.. Adult male C57BL mice (6-8 weeks old) were randomly assigned into four groups and given the following treatment: Zinc (ZnSO4, 300 micromol/kg, s.c., once a day for 2 days) or equal volume of physiological saline prior to a single dose of DOX (15 mg/kg, i.p.) or equal volume of saline. Animals were sacrificed on the 4th day after DOX administration and hearts were excised for further studies, including cadmium-hemoglobin affinity assay for MT concentration, ELISA detection of DNA fragmentation and Western blot analysis of Bax and Bcl-2.. DOX administration decreased heart weight by 10% and caused remarkable cardiac apoptosis as demonstrated by DNA fragments, while Zinc pretreatment significantly increased the MT levels and therefore inhibited the cardiac apoptotic effect of DOX. Elevated expression of Bax was obviously observed after DOX treatment, while this elevation was prevented by MT induction by Zinc. On the contrary, Bcl-2 protein level was not altered significantly among each group.. These findings suggest that metallothionein induced by Zinc exhibits protective effects on the cardiac apoptosis of DOX, which might be mediated through the prevention of Bax protein up-regulation by DOX and associated elevation of Bax/Bcl-2 ratio.

Topics: Animals; Apoptosis; bcl-2-Associated X Protein; Body Weight; Doxorubicin; Gene Expression Regulation; Heart; Male; Metallothionein; Mice; Mice, Inbred C57BL; Myocardium; Organ Size; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-bcl-2; Zinc

To understand the interactions between iron and zinc during absorption in iron- and zinc-deficient rats, and their consequences on intestinal oxidant-antioxidant balance.. Twenty-four weanling Wistar-Kyoto rats fed an iron- and zinc-deficient diet (< 6.5 mg Fe and 4.0 mg Zn/kg diet) for 4 wk were randomly divided into three groups (n = 8, each) and orally gavaged with 4 mg iron, 3.3 mg zinc, or 4 mg iron + 3.3 mg zinc for 2 wk. At the last day of repletion, 3 h before the animals were sacrificed, they received either 37 mBq of (55)Fe or (65)Zn, to study their localization in the intestine, using microautoradiography. Hemoglobin, iron and zinc content in plasma and liver were measured as indicators of iron and zinc status. Duodenal sections were used for immunochemical staining of ferritin and metallothionein. Duodenal homogenates (mitochondrial and cytosolic fractions), were used to assess aconitase activity, oxidative stress, functional integrity and the response of antioxidant enzymes.. Concurrent repletion of iron- and zinc-deficient rats showed reduced localization of these minerals compared to rats that were treated with iron or zinc alone; these data provide evidence for antagonistic interactions. This resulted in reduced formation of lipid and protein oxidation products and better functional integrity of the intestinal mucosa. Further, combined repletion lowered iron-associated aconitase activity and ferritin expression, but significantly elevated metallothionein and glutathione levels in the intestinal mucosa. The mechanism of interactions during combined supplementation and its subsequent effects appeared to be due to through modulation of cytosolic aconitase, which in turn influenced the labile iron pool and metallothionein levels, and hence reduced intestinal oxidative damage.. Concurrent administration of iron and zinc corrects iron and zinc deficiency, and also reduces the intestinal oxidative damage associated with iron supplementation.

Topics: Aconitate Hydratase; Animals; Animals, Newborn; Body Weight; Deficiency Diseases; Female; Glutathione Peroxidase; Intestinal Mucosa; Iron; Iron Deficiencies; Metallothionein; Oxidative Stress; Random Allocation; Rats; Rats, Inbred WKY; Superoxide Dismutase; Zinc

Apolipoprotein (apo) A-IV is an anorexigenic gastrointestinal peptide that is also synthesized in the hypothalamus. The goal of these experiments was to determine whether apo A-IV interacts with the central melanocortin (MC) system in the control of feeding. The third ventricular (i3vt) administration of a subthreshold dose of apo A-IV (0.5 microg) potentiated i3vt MC-induced (metallothionein-II, 0.03 nmol) suppression of 30-min feeding in Long-Evans rats. A subthreshold dose of the MC antagonist (SHU9119, 0.1 nmol, i3vt) completely attenuated the anorectic effect of i3vt apo A-IV (1.5 microg). The i3vt apo A-IV significantly elevated the expression of c-Fos in neurons of the paraventricular nucleus of the hypothalamus, but not in the arcuate nucleus or median eminence. In addition, c-Fos expression was not colocalized with proopiomelanocortin-positive neurons. These data support a synergistic interaction between apo A-IV and melanocortins that reduces food intake by acting downstream of the arcuate.

Topics: Animals; Apolipoproteins A; Body Weight; Dose-Response Relationship, Drug; Feeding Behavior; Gene Expression Regulation; Genes, fos; Hypothalamus; Male; Melanocyte-Stimulating Hormones; Metallothionein; Neurons; Pro-Opiomelanocortin; Rats; Rats, Long-Evans

We investigated the effects of genotype (two different clones) and multigenerational Cd-exposure history on Cd toxicity, individual fitness, and biokinetics in populations of a freshwater cladoceran Daphnia magna. The adults of the tolerant (T) clone had longer mean-survival-time than the sensitive (S) clone in both control groups (without Cd-exposure) and continuous Cd-exposure groups, but the two clones showed comparable resistances to acute Cd stress in the recovery groups. The body concentration of metallothionein (MT) played a critical role in handling Cd stress, which mainly accounted for the significant difference between the two clones in terms of survival distribution. High comparability of these two clones in individual fitness parameters and biokinetics suggested that these parameters are unlikely driven by genetic variation. For each specific clone, continuous Cd-exposure inhibited the animal growth, elevated the MT induction, and increased the Cd uptake rate (ingestion rate, assimilation efficiency from dietary phase, and uptake rate from dissolved phase), all of which enhanced the weight-specific Cd accumulation in daphnids' bodies. The strong dependence of biokinetic parameters on environmental factors (e.g., food concentrations, pH, dissolved or dietary metal concentration, and metal exposure histories) rather than on genotypes implied the great potential of using biokinetics in inter-lab comparisons and environmental risk assessments.

Topics: Animals; Body Weight; Cadmium; Daphnia; Environment; Feeding Behavior; Fresh Water; Kinetics; Metallothionein; Reproduction; Subcellular Fractions; Survival Analysis; Water Pollutants, Chemical

People with upper body or visceral obesity have a much higher risk of morbidity and mortality from obesity-related metabolic disorders than those with lower body obesity. In an attempt to develop therapeutic strategies targeting visceral obesity, depot- specific differences in the expression of genes in omental and subcutaneous adipose tissues were investigated by DNA array technology, and their roles in adipocyte differentiation were further examined. We found that levels of metallothionein-II (MT-II) mRNA and protein expression were higher in omental than in subcutaneous adipose tissues. The study demonstrates that MT-II may play an important role in adipocyte differentiation of 3T3L1 preadipocytes, and that N-acetylcysteine (NAC) inhibits the adipocyte differentiation of 3T3L1 cells by repressing MT-II in a time- and dose-dependent manner. Furthermore, the intraperitoneal administration of NAC to rats and mice resulted in a reduction of body weights, and a marked reduction in visceral fat tissues. These results suggest that MT-II plays important roles in adipogenesis, and that NAC may be useful as an anti-obesity drug or supplement.

Topics: 3T3-L1 Cells; Acetylcysteine; Adipocytes; Adipose Tissue; Aged; Animals; Anti-Obesity Agents; Body Weight; Cell Differentiation; Dose-Response Relationship, Drug; Down-Regulation; Female; Humans; Male; Metallothionein; Mice; Mice, Inbred C57BL; Middle Aged; Rats; Rats, Sprague-Dawley; Subcutaneous Fat; Time Factors; Viscera

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.

Topics: Acetylcysteine; Animals; Antioxidants; Benzene Derivatives; Body Weight; Chelating Agents; Drug Therapy, Combination; Free Radical Scavengers; Injections, Subcutaneous; Kidney; Liver; Male; Mercuric Chloride; Mercury Poisoning; Metallothionein; Mice; Organoselenium Compounds; Porphobilinogen Synthase; Sulfhydryl Compounds; Unithiol; Urea

Metallothioneins (MTs) are involved in the cellular metabolism of zinc and in cytoprotection against stress factors. Hippocampus plays a specific role in the body's response to stressors. The present study was conducted to evaluate the effects of zinc on the expression of metallothionein isoforms in the hippocampus of stress rats. The animal model of psychologic stress was developed by restraint for 4 weeks. Wistar rats were randomly assigned to 6 groups: control group, zinc-deficient group, zinc-supplemented group, and the corresponding 3 stress groups. Three separate diets of different zinc contents (1.73 ppm, 17.7 ppm, and 41.4 ppm, respectively) were used in this study. Compared with the control group, the stress groups had higher inductions of MTs and MT-1 and MT-3 mRNA in hippocampus. On the one hand, the expressions of MTs and their mRNAs in hippocampus were downregulated in the zinc-deficient group; however, their expressions were evidently enhanced in the stress zinc-deficient group. MT induction in the zinc-supplemented group was increased. Furthermore, the stress zinc-supplemented group had a more significant yield of MTs and their mRNAs. In addition, the levels of plasma cortisol, interleukin-6 (IL-6), IL-1, and nitric oxide (NO) were increased clearly in the zinc-deficient group and the stress groups. The results suggest that zinc deficiency may decrease and zinc supplementation may increase the expressions of MTs and their mRNAs in hippocampus; moreover, stress can increase their expressions dramatically. The impairment of stress on the body may be involved with the nutrition status of zinc, and zinc deficiency can lower the body's adaptability to stress.

Topics: Animals; Body Weight; Hippocampus; Hydrocortisone; Interleukin-1; Interleukin-6; Metallothionein; Nitric Oxide; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Zinc

In this study, we used an integrated approach to determine whether key biochemical, cellular, and physiological responses were related to growth impairment of cadmium (Cd)-exposed larval topsmelt (Atherinops affinis). Food intake (Artemia franciscana nauplii), oxygen consumption rates, apoptotic DNA fragmentation (TUNEL assay), and metallothionein (MT)-like protein levels, were separately measured in relation to growth of larval topsmelt aqueously exposed to sublethal doses of Cd for 14 days. Cadmium accumulation and concentrations of abundant metals were also evaluated in a subset of fish. Fish in the highest Cd treatments (50 and 100 ppb Cd) were smaller in final mean weight and length, and consumed fewer A. franciscana nauplii than control fish. Food intake was positively correlated with final weight of larval topsmelt in Cd and control treatments; food intake increased as final weight of the fish increased. Oxygen consumption rates were positively correlated with Cd concentration and mean oxygen consumption rates were inversely correlated with final mean weight of topsmelt; the smallest fish were found in the highest Cd treatment and were respiring at higher rates than control fish. Apoptotic DNA fragmentation was concentration-dependent and was associated with diminished growth. Apoptotic DNA fragmentation was elevated in the gill of fish exposed to 50 ppb Cd, and in the gut, gill, and liver of fish exposed to 100 ppb Cd. Metallothionein (MT)-like protein levels in fish from 100 ppb Cd treatments were significantly higher than those in other treatments. Oxygen consumption rates may have increased as a compensatory response to Cd exposure. However, it is likely that the energy produced was allocated to an increased metabolic demand due to apoptosis, MT synthesis, and changes in ion regulation. This diversion of energy expenditures could contribute to growth impairment of Cd-exposed fish.

Topics: Animals; Artemia; Body Weight; Cadmium; Calcium; Eating; In Situ Nick-End Labeling; Larva; Metallothionein; Oxygen Consumption; Smegmamorpha; Water Pollutants, Chemical

Our previous studies have shown that vanadium, a dietary micronutrient, has an inhibitory effect against experimentally induced rat hepatocarcinogenesis. In this study, we evaluated the role of vanadium on some potential protein expression markers of carcinogenesis, such as metallothionein (MT), an intracellular metal-binding protein linked with cell proliferation and apoptosis, Ki-67 nuclear antigen, and p53 tumor suppressor during 2-acetylaminofluorene (2-AAF)-induced (0.05% in basal diet) rat liver preneoplasia. In a short-term regimen, supplementation of vanadium at a dose of 0.5 ppm effectively suppressed the formation of DNA 'comets' (29.55%; P < 0.02), thereby indicating its nongenotoxicity at this particular dose. Vanadium administration throughout the study reduced relative liver weight (RLW), nodular incidence (57.15%), total number, and multiplicity (48.45%) with restoration of hepatic zinc (Zn), magnesium (Mg), selenium (Se), copper (Cu), iron (Fe), and calcium (Ca) contents when compared to the carcinogen control. Moreover, treatment with vanadium significantly abated the expressions of MT and Ki-67, studied at four sequential time points. An increased immunopositivity of p53 protein (1.03 +/- 0.23%; P < 0.02) was found in vanadium-treated rat liver with an elevated apoptotic-labeling index (AI; P < 0.001) as documented by TUNEL assay. Furthermore, a positive correlation between MT expression and Ki-67 labeling along with a strong negative correlation between MT immunoreactivity and AI (r = -0.9000, P = 0.0004 at week 24) at various time intervals suggest that, vanadium-mediated suppression of MT and Ki-67 expressions may be associated with induction of apoptosis. The results thus provide evidence for the first time in support of the potential role of vanadium on induction of p53 and apoptosis with concurrent suppression of MT and Ki-67 in order to have an understanding, in part, of the chemopreventive mechanism of this trace element in limiting neoplastic transformation in a defined model of experimental rat hepatocarcinogenesis.

Topics: 2-Acetylaminofluorene; Animals; Apoptosis; Body Weight; DNA Damage; Gene Expression Regulation; Ki-67 Antigen; Liver; Male; Metallothionein; Metals; Organ Size; Precancerous Conditions; Rats; Rats, Sprague-Dawley; Tumor Suppressor Protein p53; Vanadium

Gammarus sp. individuals were collected at four intertidal sites subjected to direct sources of pollution (marinas, ferry traffic, and harbors) and at one site with no direct source of pollution. Levels of vitellogenin-like proteins (Vtg), metallothioneins (MT), alkali-labile phosphates (ALPs) in proteins, and lipogenic enzyme activities (i.e., glucose-6-dehydrogenase, isocitrate dehydrogenase, and malate enzyme) were measured in whole soft tissues. In exoskeletons, levels of pH-dependent extractable protein and chitin were determined to assess the possible impacts of pollution on exoskeleton integrity and the molting process. Results show that males were consistently heavier than females regardless of site quality but that the whole-body weight of both sexes was significantly lower at polluted sites. Females displayed either induced or decreased Vtg-like proteins at polluted sites, indicating significant changes in gametogenesis activity. MT levels were not sex dependent and tended to be induced at all impacted sites. ALP levels in acetone-fractionated proteins indicate altered phosphate mobilization at some impacted sites, where females tended to display higher ALP levels. Lipogenic enzyme activities did not vary by sex but were readily increased at impacted sites, suggesting a delay in gonad maturation rates. Exoskeleton protein characteristics revealed that the proportion of chitin in exoskeletons was a lower at most impacted site, suggesting disruption of chitin and pH-dependent protein mobilization. Principal component analysis revealed that gammarids collected at affected sites displayed substantial changes in the proportion of chitin, arthropodin, sclerotin, MTs, and intermediary glucose metabolism (glucose-6-phosphate dehydrogenase, and isocitrate dehydrogenase in soft tissues) and thus suffered from disturbed gametogenesis and exoskeleton integrity.

Topics: Animals; Body Weight; Chitin; Crustacea; Discriminant Analysis; Female; Glucosephosphate Dehydrogenase; Isocitrate Dehydrogenase; Malate Dehydrogenase; Male; Metallothionein; Phosphates; Proteins; Quebec; Rivers; Sex Factors; Vitellogenesis; Vitellogenins; Water Pollutants, Chemical

The influence of oral administration of rare earth element cerium (Ce) was studied in relation to metallothionein (MT) and glutathione (GSH) content in the organs of ICR mice, which were administered heavy metal cadmium (Cd) for comparison. Male ICR mice were divided into 9 groups: 1 control group, 4 cerium groups and 4 cadmium groups, each with 4 mice, for a total of 36 mice. Ce groups included a 20 ppm CeCl3 diet (Ce-low) group and a 200 ppm CeCl3 diet (Ce-high) group, as did Cd groups, i.e., a 20 ppm CdCl2 diet (Cd-low) group and a 200 ppm CdCl2 diet (Cd-high) group. Each group was subdivided in 2 groups except a control group: 6-week administration group and 12-week administration group. The level of plasma aspartate aminotransferase(AST) activity, plasma alanine aminotransferase(ALT) activity, plasma cholesterol and plasma triglyceride in the Ce-low, Cd-low, Ce-high, and Cd-high group were higher than that of control group, although there were no significant differences (p > 0.05). By contrast, both Ce and Cd groups had higher levels of MT and GSH in hepatic cells compared to the control group (p < 0.05) and decreased liver tissue level of lipoperoxide (p < 0.05). These groups also had decreased plasma superoxide dismutase (SOD) activity (p < 0.05), and increased plasma level of lipoperoxide (p > 0.05). In conclusion, it is suggested that orally administered Ce increases MT and GSH as an antioxidant in the mouse liver, and these reaction are probably caused by increases in the oxidative stress with Ce.

Topics: Administration, Oral; Animals; Aspartate Aminotransferases; Body Weight; Cadmium; Cerium; Cholesterol; Eating; Glutathione; Lipid Peroxidation; Liver; Male; Metallothionein; Mice; Mice, Inbred ICR; Oxidative Stress; Superoxide Dismutase; Triglycerides

Zinc (Zn) is an essential nutrient that is required in humans and animals for many physiological functions, including immune and antioxidant function, growth, and reproduction. The present study was performed to investigate the effects of three Zn levels, including Zn adequate (35.94 mg/kg, as a control), Zn deficiency (3.15 mg/kg), and Zn overload (347.50 mg/kg) in growing male rats for 6 wk. This allowed for evaluation of the effects that these Zn levels might have on body weight, organ weight, enzymes activities, and tissues concentrations of Zn and Cu. The results showed that Zn deficiency has negative effects on growth, organ weight, and biological parameters such as alkaline phosphatase (ALP) and Cu-Zn superoxide dismutase (Cu-Zn SOD) activities, whereas Zn overload played an effective role in promoting growth, improving the developments of organs and enhancing immune system. Hepatic metallothionein (MT) concentration showed an identical increase tendency in rats fed both Zn-deficient and Zn-overload diets. The actual mechanism of reduction of Cu concentration of jejunum in rats fed a Zn-overload diet might involve the modulation or inhibition of a Cu transporter protein by Zn and not by the induction of MT.

Topics: Alkaline Phosphatase; Animal Nutritional Physiological Phenomena; Animals; Body Weight; Copper; Growth; Liver; Male; Metallothionein; Organ Size; Rats; Rats, Sprague-Dawley; Superoxide Dismutase; Zinc

Newborns are believed to have a higher gastrointestinal uptake of inorganic cadmium (Cd) than adults. However, relevant models for investigations of absorption of dietary Cd in neonates are lacking. In the present study, piglets were exposed to 2 or 20 microg Cd/kg body weight (b wt) per day by repeated oral administrations of CdCl2 dissolved in deionized water or infant follow-up formula from days 0 to 10. Elevated and dose-dependent Cd retention in blood and tissues resulted from this low-dose Cd exposure, as determined at day 11. Follow-up formula reduced Cd uptake in comparison to deionized water. However, Cd distribution to the kidneys was higher when Cd was given in formula than in water. Metallothionein (MT) levels in liver, kidney and duodenum were six, three and two times higher, respectively compared to an un-exposed control. Differences in bioavailability and distribution of low-dose Cd given in various diets to newborns could be detected by the proposed piglet model.

Topics: Animals; Animals, Newborn; Blotting, Western; Body Weight; Cadmium; Cadmium Poisoning; Digestive System; Duodenum; Feces; Ileum; Infant Formula; Iron; Kidney; Liver; Metallothionein; Models, Animal; Swine; Zinc

Growth hormone (GH) plays an incompletely understood role in the development of the central nervous system (CNS). In this study, we use transgenic mice expressing a growth hormone antagonist (GHA) to explore the role of GH in regulating postnatal brain, spinal cord and body growth into adulthood. The GHA transgene encodes a protein that inhibits the binding of GH to its receptor, specifically antagonizing the trophic effects of endogenous GH.. Before 50 days of postnatal age, GHA reduces spinal cord weight more than brain weight, but less than body weight. Thereafter, GHA ceases to inhibit the increase in body weight, which approaches control levels by day 150. In contrast, GHA continues to act on the CNS after day 50, reducing spinal cord growth to a greater extent and for a longer duration than brain growth.. Judging from its inhibition by GHA, GH differentially affects the magnitude, velocity and duration of postnatal growth of the brain, spinal cord and body. GH promotes body enlargement more than CNS growth early in postnatal life. Later, its CNS effects are most obvious in the spinal cord, which continues to exhibit GH dependence well into adulthood. As normal CNS growth slows, so does its inhibition by GHA, suggesting that reduced trophic effects of GH contribute to the postnatal slowing of CNS growth. GHA is a highly useful tool for studying the role of endogenous GH on organ-specific growth during aging.

Topics: Age Factors; Amino Acid Substitution; Animals; Body Weight; Brain; Dietary Supplements; Female; Growth Disorders; Growth Hormone; Humans; Male; Metallothionein; Mice; Mice, Transgenic; Organ Size; Promoter Regions, Genetic; Sex Factors; Spinal Cord; Transgenes; Zinc Sulfate

Concentrations of a metallothionein-like protein (MTLP) in Tubifex tubifex were consistent throughout the various life stages with no significant variation associated with reproductive stage. The mean concentration from cocoon through 10 weeks of age was 2.5 (+/-0.6) nmol MTLP/g. T. tubifex was subjected to a number of potential environmental stressors to determine whether MTLP production could be induced in the absence of metal exposure. There were no significant differences in the concentrations of MTLP between the controls and worms subjected to decreased food supply, increased handling, or reduced dissolved oxygen at a given exposure temperature. However, at 12 degrees C T. tubifex had significantly (P < 0.01) higher MTLP than T. tubifex held at 23 degrees C. There were also significant differences in wet weight and reproduction at the two temperatures, which may have implications for the levels of MTLP.

Topics: Animals; Biomarkers; Body Weight; Larva; Metallothionein; Metals, Heavy; Oligochaeta; Reference Values; Reproduction; Temperature

Elevated growth hormone (GH) concentrations suppress reproductive function in a variety of species, although it is unclear whether GH directly suppresses reproductive performance, or whether GH activates other pathways to achieve these effects. The ovine metallothionein 1a-ovine GH (oMt1a-oGH) transgenic mouse has been used to model the effects of GH on both body composition and reproductive function. A recent report has documented increased leptin levels in obese oMt1a-oGH mice. Given the importance of leptin in modulation of the reproductive endocrine axis, as well as the reports documenting reduced leptin signal transduction in animals with elevated leptin levels, we hypothesized that high leptin concentrations in response to elevated GH would reduce fertility. To determine the effects of high circulating leptin levels on the reproductive endocrine axis, we assessed hypothalamic neuropeptide Y (NPY) and GnRH expression. At weaning, oMt1a-oGH transgenic (TG) and wild-type (WT) female mice were allocated to one of four treatment groups: oMt1a-oGH females chronically expressing the transgene (TG ON); oMt1a-oGH females expressing the transgene from 3 to 8 weeks of age (TG ON/OFF); WT females receiving the transgene stimulus from 3 to 8 weeks of age (WT ON/OFF); and WT females never receiving the transgene stimulus (WT OFF). Eight-week-old females were housed with males for a 2-week period, after which females were isolated from males and allowed to carry pregnancies to term. Body and gonadal fat pad (GFP) weights, along with plasma leptin concentrations, estrous cyclicity, pregnancy rate and litter characteristics, were recorded for each female. Chronic expression of the oMt1a-oGH transgene resulted in larger leaner mice, and inactivation of the transgene produced obese females. Pregnancy rate was reduced in TG ON females when compared with all other groups, and infertility was associated with elevated leptin levels. In addition, high leptin levels were associated with increased NPY expression, suggesting reduced leptin-signaling capacity, which may contribute to suppression of the reproductive axis in oGH animals.

Topics: Animals; Body Composition; Body Weight; Estrous Cycle; Female; Fertility; Gonadotropin-Releasing Hormone; Growth Hormone; Hypothalamus; In Situ Hybridization; Leptin; Litter Size; Metallothionein; Mice; Mice, Transgenic; Neuropeptide Y; Reverse Transcriptase Polymerase Chain Reaction

Dietary isoflavone intake has been linked to cancer prevention and their anti-inflammation activity was examined. Intraperitoneal lipopolysaccharide (LPS) injection in mice led to a decrease in the liver antioxidant glutathione level but this decrease was prevented in mice fed with an isoflavone-containing diet. Similarly, isoflavone diet prevented the inflammation-associated induction of metallothionein (MT) in the intestine; and the induction of manganese superoxide dismutase (Mn-SOD) in the liver. Results from the intestinal cell studies suggest that isoflavones suppress the intestinal response to inflammation by modulating the action of pro-inflammatory cytokine, IL-6. IL-6 secretion and the STAT3 (signal transducer and activator of transcription protein 3) nuclear translocation in response to IL-6 were both decreased by genistein.

Topics: Animals; Antineoplastic Agents; Body Weight; Cell Nucleus; Cells, Cultured; Diet; DNA-Binding Proteins; Genistein; Glutathione; Humans; Interleukin-6; Intestinal Mucosa; Intestines; Isoflavones; Lipopolysaccharides; Liver; Male; Metallothionein; Mice; Mice, Inbred C57BL; Protein Transport; STAT3 Transcription Factor; Superoxide Dismutase; Trans-Activators

Metallothionein (MT) is a metal-binding protein associated with tolerance to metals and oxidative stress. Nickel is a metal carcinogen potentially acting through oxidative attack on critical biomolecules. We investigated the role of MT in nickel carcinogenesis using MT-transgenic mice that constitutively over-express MT-I in all tissues tested. Groups of 25 male MT-transgenic and wild type (C57BL/6; WT) mice received intramuscular injections of nickel subsulfide (Ni3S2) in both thighs at doses of 0 (control), 0.5, or 1.0 mg/site at 12 weeks of age and were observed for 104 weeks. Injection site tumors (ISTs; primarily fibrosarcomas) started occurring 45 weeks after nickel injection and IST incidence was similar in the WT (control - 0%, 0.5 mg/site - 20%, 1.0 mg/site - 40%) and MT-transgenic mice (control - 0%, 0.5mg/site - 28%, 1.0mg/site - 29%.). At the 0.5 mg/site dose the average time to IST in MT-transgenic mice was approximately 13 weeks shorter than in WT mice. Spontaneous lung tumors developed in 25% of control WT mice but none developed in control MT-transgenic mice. A nickel dose-related trend for increased lung tumors occurred in MT-transgenic mice but not in WT mice. Thus, the over-expression of MT did not significantly mitigate the carcinogenic response to nickel.

Topics: Acid Anhydride Hydrolases; Animals; Body Weight; Carcinogens; Gene Expression Regulation, Neoplastic; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Neoplasm Proteins; Neoplasms, Experimental; Nickel; RNA, Messenger

Lead is an environmental nephrotoxicant and probable human carcinogen. Elucidating factors predisposing populations to lead toxicity is an important public health issue. Recently, we found that metallothionein-I/-II double knockout (metallothionein-null) mice that are unable to produce the major forms of metallothionein do not produce lead inclusion bodies, which are thought to mitigate lead toxicity, and were sensitive to the subchronic toxic effects of lead exposure (10 weeks), showing modestly diminished renal function and nephromegaly compared with wild-type (WT) mice. It is unclear how this knockout might impact lead carcinogenesis. Thus, the effects of lead(II) acetate were tested in groups (n = 25) of male metallothionein-null and WT mice receiving drinking water with 0, 1,000, 2,000, or 4,000 parts per million lead for up to 104 weeks. Renal proliferative lesions (adenoma and cystic tubular atypical hyperplasia) were much more common and more severe in lead-exposed metallothionein-null mice than in WT mice. A metastatic renal cell carcinoma also occurred in a lead-treated metallothionein-null mouse, whereas none occurred in WT mice. Lead-induced renal proliferative lesions showed marked overexpression of cyclin D1, a common feature of human renal tumors. Renal lead-containing nuclear inclusion bodies were frequently observed in WT mice but did not form in metallothionein-null mice. Metallothionein was often found associated with the outer portion of these inclusion bodies. Thus, the metallothionein-null mice cannot form renal inclusion bodies, even after protracted lead exposure, and this increases the carcinogenic potential of lead. Poor production of metallothionein may predispose human populations to lead carcinogenicity.

Topics: Animals; Body Weight; Inclusion Bodies; Kidney; Kidney Neoplasms; Lead; Metallothionein; Mice; Mice, Knockout

Cadmium (Cd) and metallothionein (MT) concentrations were determined in the subcellular fractions of the gills of the clam Ruditapes decussatus. Clams were collected monthly during the period of sexual development (from June to August 2001) from two different sites situated in the Gulf of Gabès (Tunisia). These sites differ by their degree of metal contamination. Differences in condition index (CI) were observed in June and August between clams from both sites. In clams from both sites, cadmium was approximately equally distributed between the soluble (S1) and insoluble (C1) fractions. MT levels varied according to month but not sex. Site and body mass of clams seem to be important factors to explain the variation of MT levels compared to cadmium.

Topics: Animals; Bivalvia; Body Weight; Cadmium; Environmental Monitoring; Gills; Metallothionein; Seawater; Tunisia; Water Pollutants, Chemical

Mechanism of testicular toxicity induced by dietary cadmium (Cd) has been less investigated than that following acute Cd injection. In the present study we characterized testicular injury in a small rodent, the bank vole, exposed subchronically to dietary Cd in a quantity of 0.9 micromol/g, and determined the importance of some factors (Cd accumulation, metallothionein (MT), oxidative stress, and zinc (Zn)) in the injury. Dietary Cd induced moderate histopathological changes (hemorrhage in interstitium, necrosis and apoptosis in seminiferous tubule epithelium) in young (1 month old) bank voles fed, for 6 weeks, Fe-adequate (1.1-1.4 micromol/g) and Fe-enriched (4.5-4.8 micromol/g) diets. In contrast, adult (5 months old) bank voles appeared to be resistant to the toxic effects of dietary Cd, despite the fact that testicular Cd contents were higher and MT levels lower than those in the young animals. The Cd-induced histopathological changes and apoptosis were accompanied by increased testicular lipid peroxidation, decreased testicular Zn concentration and elevated levels of hepatic and renal MT and Zn. Supplemental dietary Zn (1.7-1.8 micromol/g) prevented the Cd-induced testicular Zn depletion and injury. The data indicate that dietary Cd produces testicular lesions indirectly, through decreasing testicular Zn, which seems to be due to the sequestration of this element by the Cd-induced hepatic and renal MT.

Topics: Animals; Apoptosis; Arvicolinae; Body Weight; Cadmium; Diet; In Situ Nick-End Labeling; Kidney; Lipid Peroxidation; Liver; Male; Metallothionein; Oxidative Stress; Testis; Time Factors; Tissue Distribution; Zinc

Insulin-like growth factor binding protein-1 (IGFBP-1) is synthesized in the liver and regulates the mitogenic effects of the insulin-like growth factors (IGFs). The evidence that IGFBP-1 plays a role in hepatocarcinogenesis, however, is equivocal. We have, therefore, investigated the development of preneoplastic hepatic lesions in transgenic mice in which the human IGFBP-1 gene is under the control of the mouse metallothionein promoter. The lesions were induced by treating 15-d-old male mice with a single intraperitoneal injection of 5 mg/kg diethylnitrosamine (DENA). Lesions were scored when the mice were 28 wk of age. Quantitative microscopy of liver sections revealed that significantly fewer transgenic mice treated with zinc to activate the transgene had focal lesions compared to either transgenic mice not treated with zinc or wild-type mice treated with zinc (36.4% versus 85.7% and 83.3%, respectively, P < 0.05 in each case). Zinc-treated transgenic mice also had significantly fewer lesions per liver (11.5 +/- 5.0 versus 74.7 +/- 18.4 and 59.4 +/- 15.6, respectively, P < 0.01 in each case) and a smaller percentage of liver volume occupied by lesions (0.2 +/- 0.1 versus 1.4 +/- 0.3 and 1.1 +/- 0.4 respectively, P < 0.05 in each case). Immunohistochemical staining showed that both IGF-I and IGF-II were overexpressed in most of the lesions. These results show that expression of the IGFBP-1 transgene leads to a marked inhibition of hepatic preneoplasia, possibly by decreasing the mitogenic activity of IGF-I and/or IGF-II. This study adds new evidence to the notion that the IGF axis plays an important role in liver cancer development.

Topics: Animals; Animals, Newborn; Body Weight; Carcinogens; Diethylnitrosamine; Gene Expression Regulation; Humans; Insulin-Like Growth Factor Binding Protein 1; Liver; Liver Neoplasms; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Organ Size; Precancerous Conditions; Promoter Regions, Genetic; Zinc

Rats injected with mercuric chloride develop an acute renal tubular necrosis with uremia, which is frequently lethal. Pretreatment for 3 or 7 days with a protein-free diet reduces the mortality, the clinical signs (tremor), and the severity of renal tubular necrosis, and ameliorates the uremic chemical findings in the serum. Similar results followed injection of a nephrotoxic amino acid, D-serine, after pretreatment with a protein-free diet. Indirect evidence suggests that induction of metallothionein may be involved, at least in the experiments with mercury. Acute uremia produced by nephrotoxic chemicals may be useful for further studies of the role of nutrition in uremia, while avoiding the surgical procedures and prolonged observations required for the "remnant kidney" models.

Topics: Acute Disease; Animals; Biomarkers; Body Weight; Diet, Protein-Restricted; Disease Models, Animal; Dose-Response Relationship, Drug; Injections, Subcutaneous; Kidney; Kidney Tubular Necrosis, Acute; Male; Mercuric Chloride; Metallothionein; Phosphates; Rats; Rats, Inbred Lew; Rats, Sprague-Dawley; Sucrose; Time Factors; Treatment Outcome; Urea; Uremia

Because oxidative stress is involved in arterial hypertension, impairment of hepatic antioxidant defences could develop in the course of this disease. Metallothionein (MT), an antioxidant protein, is present in high rates in the liver. The aim of this study was to investigate the effect of a mineralocorticoid-salt treatment on blood pressure, hepatic antioxidant enzyme activities, and cardiac MT levels in transgenic MT null mice compared with control mice to further clarify the role of MT during the experimental development of arterial hypertension. Control and transgenic MT -/- mice were submitted to an 8-week mineralocorticoid-salt treatment. Hepatic glutathione peroxidase, glutathione reductase, superoxide dismutase, and catalase activities and cardiac MT and mineral levels were measured. Mineralocorticoid-salt treatment induced an increase in blood pressure in both transgenic MT -/- and control mice that was associated with an impairment of liver antioxidant status. MT deficiency was associated with modifications of hepatic antioxidant enzyme activities and with a decrease in cardiac iron levels. Adaptive processes of antioxidant systems may explain the absence of an effect of metallothionein deficiency on the development of mineralocorticoid-salt hypertension. The interactions that occur between the in vivo antioxidant systems probably produce a complex regulation of the oxidative balance and consequently prevent antioxidant deficiency.

Topics: Animals; Antioxidants; Blood Pressure; Body Weight; Catalase; Desoxycorticosterone; Drug Administration Schedule; Female; Glutathione Peroxidase; Glutathione Reductase; Heart Rate; Hypertension; Liver; Male; Metallothionein; Mice; Mice, Transgenic; Myocardium; Superoxide Dismutase

Histological and metallothionein (MT) analyses of liver and kidney and measurements of serum electrolytes and hematocrit from lake whitefish (Coregonus clupeaformis), 3.5 years of age, were conducted over a 90-day period that included maximal gonadal maturation in females. Condition factors were higher in females at day 1 and when all data were combined over the 90-day sampling period. Significant differences between females and males of 1.8-fold in hepatic MT concentrations were observed at day 20 when ovaries appeared to be in the most advanced state of maturation, although this parameter was not quantified. Hematocrits were slightly but significantly lower at day 90. No sex differences were observed in the quantitative assessments of epithelium cell height, lumen diameter (LD), tubule diameter (TD), and ratio of LD:TD in the second segment of the proximal tubule (P2); and of hepatocyte area, hepatocyte nuclear diameters, and nucleus area:cytoplasmic area (N:C) ratio. Positive correlations were observed between N:C ratios and fish weight, fork length, and liver weight. No significant correlations between these histological morphometrics and MT concentrations were observed in liver or kidney. The results of this study provide baseline information for investigations designed to determine sublethal effects of metals in lake whitefish.

Topics: Animals; Biometry; Body Weight; Electrolytes; Female; Hematocrit; Kidney; Liver; Male; Metallothionein; Metals, Heavy; Ovary; Reference Values; Salmonidae; Sex Factors; Water Pollutants

The purpose of this study was to investigate the effect of zinc lipoate and zinc sulfate on zinc availability in growing rats. 6 . 6 male albino rats were fed purified diets based on corn starch, egg albumen, sucrose, soy bean oil and cellulose over a 4-week period (diet Ia: 10 mg Zn/kg as zinc sulfate, diet Ib: 10 mg Zn/kg as zinc lipoate, diet IIa: 10 mg Zn/kg as zinc sulfate +0.4% phytic acid, diet IIb: 10 mg Zn/kg as zinc lipoate +0.4% phytic acid, diet IIIa: 20 mg Zn/kg as zinc sulfate + 0.4% phytic acid, diet IIIb: 20 mg Zn/kg as zinc lipoate + 0.4% phytic acid). Zinc lipoate and zinc sulfate both proved to be highly available zinc sources. When 0.4% phytic acid were present in the diets, apparent zinc absorption was generally depressed but was higher from zinc lipoate in tendency than from zinc sulfate. Comparable results were evident for femur zinc, plasma zinc and metallothionein concentrations in liver tissues. This indicates that zinc lipoate could be a valuable zinc source under conditions of low zinc availability. Nevertheless the absence or presence of phytic acid was a more important factor influencing zinc availability than the type of zinc source investigated.

Topics: Alkaline Phosphatase; Animals; Body Weight; Femur; Liver; Male; Metallothionein; Phytic Acid; Rats; Rats, Wistar; Thioctic Acid; Zinc; Zinc Sulfate

The effects of some methyl-containing compounds added to a choline-deficient diet on the metallothionein mRNA level in the rat liver were studied. The addition of choline or carnitine to the choline-deficient diet did not induce a gain in body weight, while the addition of either betaine or methionine to the choline-deficient diet, or of methionine to the choline-deficient diet with choline significantly increased the body weight. The metallothionein mRNA level in the liver of rats fed on the choline-deficient diet was similar to that of rats fed on the choline-deficient diet with choline, betaine or carnitine. However, the addition of methionine to the choline-deficient diet with or without choline caused a marked suppression in the metallothionein mRNA level in the liver. It is thus surmised that the metallothionein mRNA level in the liver might be regulated by the dietary content of methionine.

Topics: Animal Nutritional Physiological Phenomena; Animals; Betaine; Blotting, Northern; Body Weight; Choline; Choline Deficiency; Diet; Liver; Male; Metallothionein; Methionine; Organ Size; Rats; Rats, Wistar; RNA, Messenger

The disposition and toxicity of inhaled elemental mercury (Hg0) vapor for pregnant Long-Evans rats, and potential adverse effects on reproductive outcome were investigated. Rats were exposed to 0, 1, 2, 4, or 8 mg Hg0/m(3) for 2 h/day from gestation day (GD) 6 through GD 15. Maternal toxicity occurred primarily in rats exposed to 4 and 8 mg/m(3) and was manifested as a concentration-related decrease in body weight gain and mild nephrotoxicity. Control rats gained about 13% of their initial body weight during the 10-day exposure. Rats exposed to 4 mg/m(3) Hg0 gained about 7% less than controls, and rats exposed to 8 mg/m(3) Hg0 lost about 17% of their initial body weight during the 10-day exposure period. Maternal kidney weights were significantly increased in the 4 and 8 mg/m(3) concentration groups, and urinalysis revealed increased levels of protein and alkaline phosphatase activity in urine of all Hg0-exposed rats. Dams exposed to 8 mg/m(3) were euthanized in moribund condition on postnatal day (PND) 1. There was no histopathological evidence of toxicity in maternal lung, liver, or kidney of exposed rats at GD 6, GD 15, or PND 1. The incidence of resorptions was significantly increased, litter size and PND 1 neonatal body weights were significantly decreased only in the 8-mg/m(3) group. Total Hg concentrations in maternal tissues increased with increasing number of exposure days and concentration. In general, approximately 70% of Hg was eliminated from maternal tissues during the week following the last exposure (GD 15 to PND 1). Elimination of Hg from maternal brain and kidney was slower than in other tissues, possibly due to higher levels of metallothionein. Total Hg concentrations in fetal tissues increased with increasing number of exposure days and concentration, demonstrating that a significant amount of Hg crossed the placenta. One week after the last exposure, significant amounts of Hg were still present in brain, liver, and kidney of PND 1 neonates. Metallothionein levels in neonatal tissues were not significantly increased by exposure to 4 mg/m(3) Hg0. The total amount of Hg in neonatal brain (ng/brain) continued to increase after termination of inhalation exposure, suggesting a redistribution of Hg from the dam to neonatal brain. These data demonstrate that inhaled Hg0 vapor is distributed to all maternal and fetal tissues in a dose-dependent manner. Adverse effects of Hg on developmental outcome occurred only at a concentration that caused

Topics: Administration, Inhalation; Animals; Body Weight; Dose-Response Relationship, Drug; Embryonic and Fetal Development; Female; Litter Size; Mercury; Metallothionein; Organ Size; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Long-Evans; Tissue Distribution; Urinalysis; Volatilization

Butyl benzyl phthalate (BBP) has been shown to be teratogenic. One mechanism contributing to the teratogenicity of several developmental toxicants, is chemical-induced changes in maternal zinc (Zn) metabolism which result in an increased synthesis of maternal liver metallothionein (Mt), and a subsequent reduction in Zn delivery to the conceptus. We investigated the effects of maternal BBP exposure on maternal-fetal Zn metabolism in Wistar rats. In study I, dams were gavaged with BBP (0,250,1000,1500 or 2000 mg/kg) on gestation days (GD) 11 through 13, and killed on GD 20. Maternal toxicity was evident in the three highest dose groups. Embryo/fetal death and small pup weights and lengths were noted in the 2000 mg BBP/kg group. Fetuses in the 1500 and 2000 mg/kg groups were characterized by poor skeletal ossification, and a high frequency of cleft palate. Rib anomalies were observed in the three highest dose groups. Maternal liver Mt concentrations were only slightly elevated in the 1500 and 2000 mg/kg groups. In study II, dams treated as above, were gavaged with 65Zn and killed 18 h later. While the 2000 mg/kg group had high percentages of 65Zn in some maternal tissues, sequestration of 65Zn in maternal liver was not evident. Thus, BBP is not a strong inducer of Mt, and the teratogenicity of BBP does not appear to be due to alterations in maternal and/or embryonic Zn metabolism.

Topics: Abnormalities, Drug-Induced; Animals; Body Weight; Calcification, Physiologic; Eating; Female; Metallothionein; Phthalic Acids; Plasticizers; Pregnancy; Rats; Rats, Wistar; Reproduction; Teratogens; Tissue Distribution; Trace Elements; Zinc; Zinc Radioisotopes

It is well known that excess dietary histidine induces the metabolic changes in copper and zinc. Therefore, this study was carried out to clarify whether excess dietary histidine alters the gene expressions of metallothionein-1 and metallothionein-2 in the liver and kidney. Male rats were fed the control (ad libitum and pair-fed) or histidine-excess (50 g of L-histidine per kg of diet) diet for 0, 1 and 3 days. The levels of liver metallothionein-1 and metallothionein-2 mRNA were markedly lower in the rats fed the histidine-excess diet as compared to those of the control (ad libitum and pair-fed) diet, when fed for 1 or 3 days. The levels of renal metallothionein-1 and metallothionein-2 mRNA in the rats fed the histidine-excess diet were higher or tended to be higher as compared with the rats fed the control (ad libitum and pair-fed) diet when fed for 1 or 3 days, respectively. At the same time, hepatic copper content was decreased and renal zinc content was increased by dietary histidine. It thus appears, that such a response on the level of liver metallothionein mRNA might be related to the contents of liver copper, but of kidney metallothionein mRNA might be due to the content of zinc.

Topics: Animals; Bacterial Outer Membrane Proteins; Blotting, Northern; Body Weight; Ceruloplasmin; Copper; Diet; Gene Expression Regulation; Histidine; Kidney; Liver; Male; Metallothionein; Organ Size; Rats; Rats, Wistar; Reverse Transcriptase Polymerase Chain Reaction; Weight Gain; Zinc

Exposure to cadmium poses a considerable risk to human health and environmental safety. Earthworms reside in the most contaminated sites on earth, displaying a phenomenal tolerance to toxic heavy metals. They exhibit a distinct metabolic pathway that allows the bio-accumulation of cadmium to yield body burdens in excess of 1/1000th of total dry body weight, a most impressive figure by any standard. However, the precise molecular mechanism underlying this phenomenon remains to be unraveled. This study meets this challenge by fully characterizing the major metal-binding protein in earthworms, namely the two isoforms of metallothionein. Chemical analysis of recombinant protein showed that although both isoforms bind equimolar amounts of cadmium (6 mol), wMT-2 is more stable during proton competition. Furthermore, isoform-specific transcript analysis demonstrated that only wMT-2 is responsive to cadmium in a dose and temporal manner. The specific sequestration of cadmium to wMT-2 protein was confirmed in situ using polyclonal antisera. The latter also provided the means for mapping the cellular and intracellular distribution of metallothionein, thus yielding a holistic insight into its involvement in cadmium transit during absorption, storage, and excretion. The structure-function relationship of wMT-2 and its role in cadmium detoxification through sequestration and compartmentalization is discussed.

Topics: Amino Acid Sequence; Animals; Binding, Competitive; Biological Transport; Body Weight; Cadmium; Cloning, Molecular; Dose-Response Relationship, Drug; Hydrogen-Ion Concentration; Immunohistochemistry; Ions; Metallothionein; Molecular Sequence Data; Oligochaeta; Protein Binding; Protein Isoforms; Recombinant Fusion Proteins; Recombinant Proteins; RNA, Messenger; Sequence Homology, Amino Acid; Time Factors

Metallothionein (MT) functions in zinc (Zn) homeostasis and dietary Zn affects tissue MT concentration. The objective of this study was to investigate the effects of dietary Zn deficiency and 24-h Zn repletion on MT immunolocalization and concentration in the small intestine and liver of growing rats. Three-week-old rats fed Zn-deficient diet (< 1 mg Zn/kg) for 16 d had no MT staining in either small intestine or liver. After 24-h Zn repletion with control diet (30 mg Zn/kg), strong MT staining was observed in intestinal Paneth cells and surface epithelial cells in the proliferative regions of villi. Pair-fed control rats had strong MT staining in liver that was localized around central veins. After 24-h energy repletion, the hepatic MT staining diminished. Furthermore, Zn-deficient rats had significantly reduced intestinal (57%) and hepatic (61%) MT concentrations but unaffected Zn concentrations compared with controls that consumed food ad libitum. Zn repletion for 24 h restored intestinal and hepatic MT concentrations and reduced hepatic Zn concentration. Pair-fed control rats had elevated MT concentration in liver that was normalized by energy repletion. There was a significant positive correlation between tissue Zn and MT concentrations in liver (r = 0.60, P = 0.0001), but not in small intestine. In summary, MT immunolocalization and concentration in rat small intestine and liver were responsive to changes in Zn status, supporting the role of MT in Zn metabolism. Cell-type-specific localization of MT in small intestine after dietary Zn manipulations indicates a function of Zn and MT in gut immunity and intestinal mucosal turnover, and the pattern of hepatic MT distribution with energy restriction may be linked to detoxification processes.

Topics: Animals; Body Weight; Immunohistochemistry; Intestine, Small; Liver; Male; Metallothionein; Rats; Rats, Sprague-Dawley; Tissue Distribution; Zinc

Dogwhelks Nucella lapillus feed mainly on mussels and barnacles, and may experience periods of starvation. We report effects of nutritional state and prey type on the survival, growth, cadmium (Cd) accumulation, metallothionein (MT) induction and glycogen stores in N. lapillus exposed to Cd in water. Adult dogwhelks, with similar shell length (30.0+/-1.5 mm), were either starved or fed to satiation with barnacles Semibalanus balanoides, mussels Mytilus edulis or Cd-dosed M. edulis, and kept in filtered natural seawater (< 0.01 microg Cd 1(-1)) or Cd-contaminated (400 microg Cd 1(-1)) seawater for 80 days. Mortality and individual growth rate were determined. Cd, MT and glycogen were measured in different tissues. Prolonged starvation and exposure to Cd significantly reduced the survivorship of N. lapillus, but feeding could help dogwhelks to combat Cd toxicity and minimise mortality. Extended starvation also caused tissue wastage, leading to higher concentrations of Cd and MT in tissues, whereas fed animals increased in weight and had lower Cd and MT concentrations because of the tissue dilution effect. Prey type significantly affected growth rate of dogwhelks and indirectly influenced Cd accumulation, MT induction and glycogen stores. Eating mussels promoted better growth and higher glycogen reserves than eating barnacles. Individual growth rate decreased with increasing Cd accumulation. Cd-exposed survivors grew faster and consumed more than control animals, implying that these survivors may have better fitness and greater tolerance to Cd toxicity. The use of growth, condition index, MT and glycogen as biomarkers of environmental pollution are discussed. These results indicate a need to incorporate biological data including growth (or at least condition index) and prey type into biomonitoring programmes to allow sound interpretation.

Topics: Animals; Body Weight; Cadmium; Environmental Exposure; Food Chain; Glycogen; Metallothionein; Mollusca; Nutritional Status; Predatory Behavior; Starvation; Tissue Distribution; Water Pollutants, Chemical

Exposing growing oMtla-oGH transgenic mice with the regulatable metallothionein promotor to elevated growth hormone (GH) for three weeks after weaning enhances bone length and adipocyte differentiation. The objective of the present study was to investigate the consequences of highly elevated GH exposure during fetal and early postnatal growth periods on the mature phenotype. Transgene expression, hence elevated GH, was achieved in fetuses and neonates by providing 25 mM ZnSO4 to the drinking water of the dams. Wildtype and oMtla-oGH male and female mice were a) never exposed to the transgene stimulus, b) exposed from birth to 21 d of age, c) exposed through gestation until 21 d of age, d) exposed only through gestation, or e) exposed only during the first 7 d postpartum. At 84 d of age when mature body size was reached, ulna and humerus lengths, and body, liver gonadal fat pad, mesenteric fat pad, and cleaned gastrointestinal (GI) tract weights were recorded. Bone lengths were also determined in a subset of mice at 22 d of age. While early exposure to the elevated GH increased ulna and humerus length at 22 d of age, the early GH levels failed to produce significant changes in adipose content or bone lengths at maturity. However, chronic exposure to slightly elevated GH, as seen in the transgenics never induced to express the transgenic GH, depressed liver and GI weights and increased adipose depot weights and humerus lengths across both sexes. These results suggest that certain tissues in the body, while capable of responding to GH during early developmental periods, are not fully entrained to sustain that growth response once the GH stimulus is withdrawn. Further, the preadipocyte pool appears unable to respond to GH early in development. Finally, the tissues examined exhibited a differential response to the GH suggesting that different tissues possess distinct response thresholds.

Topics: Adipose Tissue; Animals; Animals, Newborn; Body Weight; Bone Development; Digestive System Physiological Phenomena; Female; Gene Expression; Growth Hormone; Humerus; Liver; Male; Metallothionein; Mice; Organ Size; Transgenes; Ulna; Zinc

To clarify toxic effects of long-term oral administration of low dose cadmium (Cd) on the liver and kidney, six groups of female Sprague-Dawley rats were fed a diet containing Cd-polluted rice or CdCl2 at concentrations up to 40 ppm, and killed after 12, 18, and 22 months. With toxicological parameters, including histopathology, there was no evidence of Cd-related hepato-renal toxicity, despite a slight decrease of mean corpuscular volume and mean corpuscular hemoglobin of red blood cells with 40 ppm CdCl2. Dose-dependent accumulation of Cd was observed in the liver and kidneys with peak levels of 130 +/- 42 micrograms/g and 120 +/- 20 micrograms/g, respectively, at 18 months in animals treated with 40 ppm CdCl2. A dose-dependent increase in urinary Cd levels became evident with time. Induction of metallothionein (MT) was also observed in the liver and kidney with a high correlation to the corresponding Cd levels. In the proximal renal tubular epithelia of 40 ppm CdCl2-treated rats at 22 months, prominent accumulation of Cd was observed in secondary lysosomes associated with MT deposits in their exocytotic residual bodies. The results demonstrated that, in contrast to the case with high-dose Cd-administration, renal toxicity is not induced by long-term oral administration of low amounts of Cd, although tissue accumulation does occur. Possible protective mechanisms may be operating.

Topics: Animals; Body Weight; Cadmium Chloride; Dose-Response Relationship, Drug; Environmental Pollutants; Female; Food Contamination; Kidney; Liver; Metallothionein; Organ Size; Organ Specificity; Oryza; Rats; Rats, Sprague-Dawley

Zinc is an essential heavy metal for the normal function of the central nervous system (CNS), but the knowledge of its metabolism and functions is scarce. In this report we have studied the effect of a zinc deficient diet on the regulation of brain metallothioneins (MTs). In situ hybridization analysis revealed that brain MT-I induction by restraint stress was significantly blunted in some but not all brain areas in the mice fed the zinc deficient diet compared to normally fed mice. In contrast, brain MT-I induction by the administration of bacterial lipopolysaccharide (LPS) was not significantly lower in the mice fed the zinc deficient diet. In contrast to MT-I, MT-III mRNA levels were minimally affected by either stress or LPS. Yet, significant decreasing effects of the zinc deficient diet were observed in areas such as the neocortex, CA1-CA3 neuronal layer and dentate gyrus of the hippocampus, and the Purkinje neuronal layer of the cerebellum. These results demonstrate that dietary zinc deficiency impairs the response of brain MTs during both stress and LPS-elicited inflammatory response in a highly specific manner.

Topics: Animals; Body Weight; Brain; Diet; Eating; In Situ Hybridization; Inflammation; Lipopolysaccharides; Male; Metallothionein; Metallothionein 3; Mice; Nerve Tissue Proteins; RNA, Messenger; Stress, Physiological; Zinc

Several studies have recently shown that metallothionein (MT), a protein characterized by a high thiol content and that binds Zn2+ and Cu+, might be involved in the protection against oxidative stress and can act as a free radical scavenger. Oxidative stresses, such as irradiation, increase lipid peroxidation (LP) and subsequent tissue damage through free radical production. The induction of hepatic MT synthesis by gamma-irradiation (20 Gy) at 8, 24, 30 and 48 hrs. post-irradiation in two different age groups of Sprague-Dawley rats (39-40 and 48-49 days old) was studied. LP measured by the thiobarbituric acid reactive substances (TBARS) assay and Cu and Zn levels in liver have also been determined. In the younger group, the gamma-irradiation induced hepatic MT synthesis and increased LP that peaked 24 hrs. after irradiation. During the first 30 hrs. post-irradiation, a positive and statistically significant correlation between hepatic MT content and LP level in liver was found. In the older group, liver MT synthesis was only increased 1.7-fold and LP levels were not altered at 24 hrs. post-irradiation compared with sham-irradiated rats. Therefore it appears that LP is not necessary for induction of MT synthesis by gamma-irradiation.

Topics: Age Factors; Animals; Body Weight; Copper; Gamma Rays; Lipid Peroxidation; Liver; Male; Metallothionein; Rats; Rats, Sprague-Dawley; Time Factors; Zinc

Correlated responses in female reproductive performance were evaluated following short-term selection within full-sib families for increased 8-week body weight in two replicates of four lines of mice: two ovine metallothionein-ovine growth hormone (oMt1a-oGH) transgene-carrier lines, one from a high-growth background (TM) and one from a control background (TC), and two non-transgenic lines, one from each of these genetic backgrounds (NM and NC, respectively). A fifth line (CC), not containing the transgene, served as a randomly selected control. The initial frequency of the oMt1a-oGH transgene construct in the TM and TC lines was 0.5. The frequency of transgenic females sampled at generations 7 and 8 of selection was 84.0% and 6.1% in the TC and TM lines, respectively. No significant female infertility differences were detected between transgene-carrier and non-transgenic lines or between transgenic and non-transgenic mice within carrier lines, whereas high-growth background lines had a higher infertility than control background lines (P < 0.05). Correlated responses in the TC transgene-carrier line were suggestive of reduced reproductive performance as indicated by increased post-implantation mortality (P < 0.05), number of dead fetuses plus implants (P < 0.05), and loss of fetuses from day 16 to parturition (P < 0.001). For the first two traits, the negative correlated responses were accounted for by the reduced performance of transgenic compared with non-transgenic females. Embryos carrying the transgene may also have a lower viability. In contrast, the NC non-transgenic line did not exhibit reduced reproductive performance for these traits. The low frequency of the transgene in the high-growth background TM line was associated with reduced fitness and a lower additive effect for 8-week body weight compared with the control background TC line.

Topics: Animals; Body Weight; Female; Growth Hormone; Infertility, Female; Metallothionein; Mice; Mice, Transgenic; Ovulation; Reproduction; Sheep

The relationship between metallothionein levels and concentrations of several metals and radionuclides was examined in liver tissues of raccoons (Procyon lotor, n = 47) from the Department of Energy's Savannah River Site in South Carolina to determine the applicability of metallothioneins as an initial screening device for exposure assessment in free-living mammals and environmental monitoring. Using a fluorescent marker and a cell sorter to measure metallothionein, a significant positive correlation was found across animals between levels of metallothioneins and concentrations of selenium (Pearson's r = .30), mercury (Pearson's r = .3 1), and copper (Pearson's r = .30) in liver tissue. Arsenic, cobalt, silver, thallium, and tin were below detection limits in most or all liver samples. Other metals, including cadmium, chromium, radiocesium (137-Cs), copper, lead, manganese, strontium, and vanadium, showed only weak and nonsignificant correlations with metallothionein. Concentrations of mercury were correlated with concentrations of selenium (Pearson's r = .73), manganese (Pearson's r = .56), and strontium (Pearson's r = .57). In an a posteriori test, there was a still unexplained positive correlation between mercury (Pearson r = .56), selenium (Pearson r = .54), and radiocesium (Pearson's r = .38) concentrations and background cellular autofluorescence, and a negative correlation of strontium with the latter (Kendall tau = -.38). Background cellular autofluorescence may represent a generalized cellular stress response, or a yet unidentified biomarker. To better understand which metals contribute to the induction of metallothionein, principle component analysis (PCA) was performed. The first three principle components explained 78% of the variance, with highest loadings being from mercury and radiocesium. Metallothionein levels did not correlate well with the principal components from the metals and radiocesium, while autofluorescent background levels tended to correlate better.

Topics: Animals; Arsenic; Body Weight; Cesium Radioisotopes; Copper; Environmental Monitoring; Environmental Pollution; Flow Cytometry; Geography; Liver; Male; Manganese; Mercury; Metallothionein; Metals; Metals, Heavy; Predictive Value of Tests; Raccoons; Selenium; South Carolina; Strontium

Cadmium (Cd) and arsenic (As) are important inorganic toxicants in the environment. Humans certainly have the potential to be exposed to the mixtures of Cd and As, but the toxicological interactions of these inorganic mixtures are poorly defined. Metallothionein (MT) is a cysteine-rich, metal-binding protein that plays an important role in Cd detoxication, but its role in As toxicity is less certain. To examine the role of MT in Cd- and/or As-induced nephrotoxicity, MT-I/II-knockout (MT-null) mice and background-matched wild-type (WT) mice were fed CdCl(2) (100 ppm Cd) in the diet, NaAsO(2) (22.5 ppm As) in the drinking water, or Cd plus As for 4 months. Subsequently, nephrotoxicity was examined by morphological and biochemical techniques. Chronic exposure to Cd produced more renal toxicity than As, and the combination of Cd and As produced even more renal injury than caused by either of the chemicals given alone. In mice receiving Cd plus As, proximal tubule degeneration and atrophy, glomerular swelling and interstitial fibrosis were more severe than those produced by either inorganic. Furthermore, lack of MT rendered MT-null mice more sensitive than WT mice to the nephrotoxicity produced by chronic Cd- and/or As-exposure. MT-null mice were especially susceptible to the toxicity produced by the combination of Cd and As, as evidenced by decreased body weight, enzymuria, glucosuria, proteinuria and nephropathy. In conclusion, this study indicates that As may potentiate Cd nephrotoxicity during the long-term, combined exposure, and that intracellular MT plays a role in decreasing the nephropathy of combined exposure to Cd and As.

Topics: Acetylglucosaminidase; Administration, Oral; Animals; Arsenic; Body Weight; Cadmium; Drug Administration Schedule; Drug Synergism; Glycosuria; Kidney; Kidney Diseases; Liver; Male; Metallothionein; Mice; Mice, Knockout; Organ Size; Proteinuria

The objective of this study was to correlate hepatic and renal cadmium (Cd) accumulation, Cd-binding capacity of metallothionein (MT) and lipid peroxidation with the tissue injury in the male bank voles raised under short (8 h light/16 h dark) and long (16 h light/8 h dark) photoperiods that affect differently Cd accumulation and MT induction in these rodents. The animals were exposed to dietary Cd (0, 40 and 80 microg/g) for 6 weeks. The accumulation of Cd in the liver and kidneys appeared to be dose-dependent in bank voles from the two photoperiod groups; however, the short-photoperiod animals exhibited significantly higher concentrations of Cd in both organs than the long-photoperiod bank voles. Cd-Binding capacity of MT in the liver and kidneys of bank voles from the long photoperiod was sufficiently high to bind and detoxify all Cd ions, while in the animals fed 80 microg Cd/g under the short photoperiod, the concentrations of Cd in both organs exceeded (by about 10 microg/g) the MT capacity. However, similar histopathological changes in the liver (a focal hepatocyte swelling and granuloma) and kidneys (a focal degeneration of proximal tubules) occurred in Cd-80 bank voles from the two photoperiods. Likewise, in either photoperiod group, dietary Cd brought about a similar, dose-dependent decrease in the hepatic and renal lipid peroxidation, which paralleled closely that of the iron (Fe) concentrations. These data indicate that: (1) MT does not protect the liver and kidneys against Cd-induced injury in the bank vole exposed to the higher level of dietary Cd; and (2) lipid peroxidation cannot be responsible for the tissue damage. It is hypothesized that dietary Cd produces histopathological changes indirectly, through depressing the tissue Fe and Fe-dependent oxidative processes.

Topics: Animals; Arvicolinae; Body Weight; Cadmium; Chemical and Drug Induced Liver Injury; Copper; Diet; Iron; Kidney; Kidney Diseases; Light; Lipid Peroxidation; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Photoperiod; Zinc

This study examined the effects of simultaneous exposure to saline and cadmium (Cd) on organ mass and histology of a bird with salt glands, the Pekin duck, Anas platyrhynchos. Three mixed-sex groups, each containing 6 birds, ate duck pellets containing 0, 50, or 300 microg Cd/g, respectively, for 4 1/2 mo and drank 300 mM NaCl. Only females on the high-Cd diet lost body mass. Ingestion of Cd reduced heart mass in females. There was increased mass of Harderian and salt glands in both sexes. Mass of kidneys and liver increased only in males, and the gut mass (also length) increased more in males. Cadmium ingestion also induced (1) inflammation of renal interstitium and degenerative tubular changes, (2) marked degenerative changes in testes, (3) increased heart water content, (4) decreased cytoplasmic volume of liver cells, (5) reduced proportion of basophilic granular cells in chromaffin tissue of the adrenal glands, and (6) in the ileum, increased heterophilia in the lamina propria and, only in females, the apoptosis to mitosis ratio in crypt cells of the epithelium. The ducks' outward appearance gave no indication that ingesting large amounts of cadmium for 4 1/2 mo produced deleterious effects, but the physiological consequences were profound. Both sexes had greatly reduced gonadal mass and the males produced no sperm. The higher dietary level greatly hypertrophied the liver, kidneys, and gut only in males. The cadmium-induced changes in organs, particularly in the gonads, kidneys, and adrenal glands, should greatly impair the health and reproductive capacity of these ducks.

Topics: Animals; Bird Diseases; Body Weight; Cadmium; Cadmium Poisoning; Digestive System; Ducks; Eating; Female; Male; Metallothionein; Organ Size; Salt Gland; Sodium Chloride; Water-Electrolyte Balance

Rainbow trout (Oncorhynchus mykiss) were exposed to sublethal concentrations of copper (Cu, 14 microgram/liter) and zinc (Zn, 57 and 81 microgram/liter) for a 21-day period. The four treatments included a control, a Cu control, a Cu and low-Zn treatment and a Cu and high-Zn treatment. Selected parameters [e.g., hemoglobin (Hb), hematocrit (Hct), plasma glucose, lactate and cortisol, differential leukocyte count, respiratory burst, tissue metal concentrations, hepatic metallothionein (MT), brain acetylcholinesterase (AChE)] were evaluated at 2, 7, 14, and 21 days of exposure. Whole blood and plasma parameters were not altered by exposure to metals. The percentage of lymphocytes was consistently decreased in the three metal treatments, while percentages of neutrophils and monocytes were increased. Respiratory burst activity was elevated in all metal treatments. Gill Zn concentration was highly variable, with no significant alterations occurring. Gill Cu concentration was elevated above control levels in all metal treatments. Gill Cu concentration in the two Cu/Zn treatments was also elevated above levels in the Cu control. Hepatic metal concentrations and MT levels were not altered from control values. Measurements of brain AChE indicated an elevation in this parameter across metal treatments. In general, alterations in physiological parameters appeared to be due to Cu, with Zn having no interactive effect.

Topics: Acetylcholinesterase; Animals; Body Weight; Brain; Copper; Gills; Leukocyte Count; Liver; Metallothionein; Oncorhynchus mykiss; Respiratory Burst; Survival Analysis; Water Pollutants, Chemical; Zinc

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.

Topics: Animals; Body Weight; Bone Density; Bone Diseases; Cadmium; Drinking; Energy Intake; Female; Kidney; Kidney Diseases; Liver; Male; Metallothionein; Rats; Rats, Sprague-Dawley; Sex Factors

Growth hormone (GH) transgenic mice are known to develop hepatocellular adenomas and carcinomas. In order to understand more about hepatocarcinogenesis in the GH-transgenic mouse model we quantitated the rates of hepatocellular proliferation and apoptosis in these mice.. Two lines of GH-transgenic mice and non-transgenic control mice were generated and sacrificed at regular intervals between one and nine months. Hepatocellular replication was measured by in vivo incorporation of bromodeoxyuridine (BrdU) and counting BrdU-positive nuclei in histological liver sections. Serial sections taken from these mouse livers were also assessed for rates of hepatocellular apoptosis using the in situ end-labelling of fragmented DNA (TUNEL) method.. High levels of hepatocellular replication were sustained life-long in this model. Increased rates of hepatocellular proliferation preceded the onset of hepatic inflammation, a prominent feature in the liver pathology of GH-transgenic mice. In tumour tissue, cellular proliferation was up to 17-fold greater than in surrounding non-tumour tissue. Apoptosis rates were also elevated in non-tumour regions of GH-transgenic mouse livers compared to controls. Interestingly, large dysplastic hepatocytes were common in the fraction of cells undergoing apoptosis, especially in older mice with inflamed livers. The increase in the rate of hepatocellular apoptosis in GH-transgenic animals largely balanced the augmented levels of proliferation seen in these mice. In tumour tissue, however, the profound increase in the number of proliferating tumour cells outstripped the increase in apoptosis.. Relatively high and enduring levels of hepatocellular replication and apoptosis precede hepatocarcinogenesis in GH-transgenic mice. Increased cellular proliferation and resistance to apoptosis were evident in tumour growth in older animals.

Topics: Adenoma; Age Factors; Animals; Apoptosis; Body Weight; Carcinoma, Hepatocellular; Cell Division; Female; Growth Hormone; In Situ Nick-End Labeling; Liver; Liver Neoplasms, Experimental; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Mitotic Index; Organ Size

Effects of life-long exposure to high levels of homologous or heterologous growth hormone (GH) and effects of GH resistance on selected parameters of immune function were studied in adult male transgenic mice overexpressing GH releasing hormone (GHRH), bovine (b) GH or an antagonistic bGH analog. In metallothionein I (MT)-bGH transgenic mice with high peripheral levels of bovine GH, there were significant increases in the absolute weight of the thymus and the spleen and in the mitogenic responses of splenocytes to concanavalin A (ConA), lipopolysaccharide (LPS) and phytohemagglutinin (PHA), as compared to age-matched normal animals. There were no significant differences between MT-bGH transgenic and normal mice in splenocyte viability or in delayed-type hypersensitivity measured by the allergic contact dermatitis response to oxazolone. Similar results, including significant stimulation of splenocyte responses to ConA, LPS, and PHA, were obtained in MT-hGHRH transgenic mice in which overexpression of GHRH leads to striking pituitary enlargement and massive elevation of peripheral levels of homologous (mouse) GH. In MT-bGH-antagonist transgenic mice in which overexpression of an antagonistic bGH analog interferes with the actions of endogenous GH, spleen weight was reduced but proliferative responses of splenocytes to ConA, LPS, and PHA were not affected. It is concluded that overexpression of heterologous or homologous GH in transgenic mice can lead to significant stimulation of some parameters of immune function, whereas antagonism of GH action by expression of an antagonistic GH analog does not affect splenocyte responses to mitogens.

Topics: Acromegaly; Animals; Body Weight; Carrier Proteins; Cattle; Cell Division; Cells, Cultured; Gene Expression Regulation; Growth Hormone; Growth Hormone-Releasing Hormone; Hypersensitivity, Delayed; Male; Metallothionein; Mice; Mice, Transgenic; Mitogens; Organ Size; Oxazolone; Promoter Regions, Genetic; Spleen

To investigate the relationship between cadmium (Cd) toxicity, intestinal absorption, and its distribution to various tissues in rats treated orally with minimum amounts of Cd, 14 female rats per dose group per time point were given diets consisting of 28% purified diet and 72% ordinary rice containing Cd-polluted rice (0. 02, 0.04, 0.12, or 1.01 ppm of Cd) or CdCl(2) (5.08, 19.8, or 40.0 ppm of Cd) for up to 8 months. At 1, 4, and 8 months after the commencement of Cd treatment, seven rats per group were euthanized for pathological examinations to determine the Cd concentrations in the liver and kidneys and metallothionein (MT) in the liver, kidneys, intestinal mucosa, serum, and urine. One week before each period of 1, 4, and 8 months, the remaining seven rats in each group were administered a single dosage of (109)Cd, a tracer, to match the amounts of the designated Cd doses (about 1.2 to 2400 microg/kg body wt). They were euthanized 5 days later to determine the distribution of Cd to various tissues. No Cd-related toxic changes were observed. The concentrations of Cd in the liver and kidneys at any time point and MT in the liver, kidney, serum, and urine at 4 and 8 months increased dose-dependently, whereas MT in the intestinal mucosa did not alter markedly at any time point. The distribution rates of Cd to the liver increased dose-dependently (40% at lower doses to 60% at higher doses), whereas those to the kidney decreased dose-dependently (20% at lower doses to 10% at higher doses). The Cd retention rates 5 days after (109)Cd administration (amounts of Cd in various tissues/amounts of Cd administered) ranged from 0.2 to 1. 0% at any time point. These results suggest that the distribution of Cd to the liver and kidneys after the oral administration vary depending on the dosage levels of Cd. The difference of the distribution pattern of Cd to the liver and kidney is probably due to the difference in the form of the absorbed Cd, i.e., free ion or Cd-MT complex, although not closely related to the MT in the intestinal mucosa.

Topics: Administration, Oral; Animals; Body Weight; Cadmium; Cadmium Chloride; Cadmium Radioisotopes; Diet; Female; Food Contamination; Metallothionein; Oryza; Rats; Rats, Sprague-Dawley; Time Factors; Tissue Distribution

To study effects of dietary Cu and Fe levels on the onset of hepatitis in Long-Evans Cinnamon (LEC) rats, female rats (40 days old) were fed a semipurified diet containing 0.1 or 10 mg Cu/kg and 1.5 or 150 mg Fe/kg in a 2 x 2 factorial arrangement for 35 days. At 75 days after birth, LEC rats (+Cu-Fe) fed a Cu-sufficient but Fe-deficient diet (Cu, 10 mg/kg; Fe, 1.5 mg/kg) showed jaundice, with lethargy, anorexia, and malaise. The biochemical variables relating to liver function were significantly increased compared to three other groups, a Cu- and Fe-deficient (-Cu-Fe) group, a Cu-deficient but Fe-sufficient (-Cu+Fe) group, and a Cu and Fe sufficient (+Cu+Fe) group. Furthermore, the +Cu-Fe rat liver showed massive necrosis with huge nuclei. The other three groups presented no biochemical and histological findings of hepatitis. Hepatic Cu and metallothionein concentrations were 289 +/- 87 (mean +/- SD) microg/g liver and 8.7 +/- 1.8 mg/g liver, respectively, in the +Cu-Fe rats. However, in the +Cu+Fe group the values were 196 +/- 28 microg Cu/g liver and 10.8 +/- 1.0 mg/g liver. Hepatic Fe deposition was not influenced significantly by the dietary Cu level. The +Cu-Fe group with jaundice showed the highest free Cu concentration in the liver among the four groups, but the hepatic free Fe concentration was similar to those in the -Cu+Fe and +Cu+Fe groups. Our results indicate that an Fe-deficient diet enhances the deposition of hepatic Cu due to increased absorption of Cu from the gastrointestinal tract. This deposition stimulated the onset of hepatitis.

Topics: Animals; Body Weight; Copper; Female; Ferritins; Hepatitis, Animal; Hepatolenticular Degeneration; Intestinal Absorption; Iron; Iron Deficiencies; Iron, Dietary; Liver; Metallothionein; Rats; Rats, Long-Evans; Specific Pathogen-Free Organisms

Metallothionein (MT) has several putative roles in metal detoxification, in Zn and Cu homeostasis, in scavenging free radicals, and in the acute phase response. Mice of mixed 129/Ola and C57BL/6J background with targeted disruption of MT-I and MT-II genes are more sensitive to toxic metals and oxidative stress. We noted that these animals were larger than most strains of mice, and we systematically studied aspects of their physiology and biochemistry relating to energy metabolism. During the first 2 weeks after weaning, the growth rates of MT-null and C57BL/6J mice were similar, but the transgenic mice became significantly heavier at age 5-6 weeks. At age 14 weeks, the body weight and food intake of MT-null mice was 16 and 30% higher, respectively, compared with C57BL/6J mice. Most 22- to 39-week-old male MT-null mice were obese, as shown by increased fat accretion, elevated obese (ob) gene expression, and high plasma leptin levels, similar to those recorded in Zucker fatty (fa/fa) rats. Seven-week-old MT-null mice also had significantly higher levels of plasma leptin and elevated expression of ob, lipoprotein lipase, and CCAAT enhancer binding protein alpha genes as compared with age-matched C57BL/6J mice. These observations indicate that abnormal accretion of body fat and adipocyte maturation is initiated at 5-7 weeks of age, possibly coincident with sexual maturation. Targeted disruption of MT-I and MT-II genes seems to induce moderate obesity, providing a new obese animal model. A link between MT and the regulation of energy balance is implied.

Topics: Adipose Tissue; Animals; Body Weight; CCAAT-Enhancer-Binding Proteins; DNA-Binding Proteins; Eating; Energy Metabolism; Insulin; Leptin; Liver; Liver Glycogen; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Nuclear Proteins; Obesity; Proteins; Rats; RNA, Messenger; Transcription Factors

Four experiments were conducted to identify several factors that might improve the accuracy and reproducibility of Zn bioavailability assays for chicks. Response of tissue Zn and metallothionein (MT) concentrations to various elevated levels and soluble sources of dietary Zn were measured, as well as the effect of delaying high Zn administration until 7 d posthatching to alleviate the detrimental effect of Zn sulfate on feed intake to 3 wk of age. Bone Zn increased (P < 0.01) in all experiments in response to increasing dietary Zn concentrations. Liver and pancreas MT were affected (P < 0.01) by a source by age interaction and variability that made this criterion unsuitable for bioavailability assays. Lastly, 1-d-old chicks were used to study the effect of delaying feeding of a high-Zn diet up to 7 d of age. The basal diet was fed continuously for 21 d as a control. A diet containing 1,000 ppm Zn was either fed continuously from Day 1, or started on Day 3, 5, or 7. Chicks given high Zn on Day 3, 5, or 7 decreased (P < 0.01) feed intake within 24 h of feeding. Delayed feeding of high dietary Zn might help to alleviate decreased feed intake observed in previous studies. Delaying the onset of high Zn feeding by several days may help alleviate feed intake problems observed with Zn sulfate. Use of either Zn gluconate or Zn acetate as a standard in assays or use of MT synthesis as a bioavailability criterion will probably not be useful to improve accuracy of the estimates.

Topics: Aging; Animals; Biological Availability; Body Weight; Chickens; Diet; Female; Liver; Male; Metallothionein; Pancreas; Zinc; Zinc Acetate; Zinc Oxide; Zinc Sulfate

Kidney is the main target organ of Cd toxicity in humans. Cd-induced nephrotoxicity is thought to be caused by the Cd-metallothionein complex (CdMT) that "leaks" out of the liver and is taken up by the kidney. A single injection of CdMT has therefore been used as a model to study Cd nephropathy for the last 20 years. However, our recent studies reveal discrepancies between renal Cd concentration and nephrotoxic potencies of CdCl2 and CdMT. This study was further designed to critically evaluate whether a single injection of CdMT is an appropriate model to study the mechanism of chronic CdCl2 nephropathy. Age-matched rats were given multiple sc injections of either CdCl2 (0.8 and 1.2 mg Cd/kg) or CdMT (0.05 mg Cd/kg) daily, 6 days/week for 6 weeks, or a single injection of CdMT (0.2-0.6 mg Cd/kg i.p. for 24 h), and the nephrotoxicity was compared. Histologically, chronic CdCl2 or CdMT administration produced damage to the whole kidney, including tubular cell degeneration, apoptosis, and atrophy; interstitial inflammation; glomerular swelling; and sclerosis. In contrast, acute CdMT injection produced severe proximal tubule necrosis as the major feature of its toxicity. Biochemically, chronic exposure to Cd produced polyuria and calciuria, while proteinuria, glucosuria, and enzymuria were mild (2-5x). In contrast, acute CdMT nephrotoxicity was characterized by marked increases in urinary protein (13x), glucose (25x), N-acetyl-beta-d-glucosaminidase (28x), lactate dehydrogenase (100x), and gamma-glutamyltranspeptidase (160x). Serum levels of creatinine and blood urea nitrogen were unchanged following chronic Cd exposure but were markedly elevated (5x) after acute injection of CdMT. Chronic exposure to either CdCl2 or CdMT produced nephrotoxicity at renal Cd concentration of 85 to 110 micrograms/g kidney, while acute CdMT injection produced nephrotoxicity at only 5 to 7 micrograms/g kidney. In conclusion, the present study indicates that the features and mechanisms of renal injury from chronic Cd exposure are quite different from those produced by a single injection of CdMT. Therefore, it is proposed that acute CdMT injection is not an appropriate model for the study of chronic Cd-induced nephrotoxicity.

Topics: Animals; Body Weight; Cadmium Chloride; Female; Glycosuria; Kidney; Kidney Diseases; Metallothionein; Proteinuria; Rats; Rats, Sprague-Dawley

Male and female transgenic mice carrying the regulatable ovine metallothionein-ovine growth hormone (oMt1a-oGH) transgene were exogenously induced to express elevated GH at maturity. Male transgenics responded to continuously elevated GH with smaller white adipose tissue depots (WAT); cessation of transgene stimulation with restoration of basal circulating GH ablated the improvement in WAT. Transgenic males in which the transgene was never exogenously activated expressed low levels of oGH (60.2 ng/ml) and this low, chronic level resulted in 30-50% larger (p < 0.05) gonadal, inguinal, and mesenteric WAT relative to all wildtype controls, transgenics actively expressing the transgene, or transgenics exposed to elevated GH for a limited time. Mature females continued to accrue body mass proportional to circulating GH. Similar to that observed in the males, transgenic females in which the transgene was never exogenously activated exhibited enlarged WAT that were 20-40% larger than the same depots in wildtype control females. However, in contrast to the data for the males, transgenic females that had experienced a four week exposure to elevated GH followed by a return to basal conditions tended to have the greatest WAT depots (p < 0.10). This would indicate that female adipocytes are still capable of responding to elevated GH with proliferation/differentiation at maturity. These data support the hypothesis that the generalized sexual dimorphic pattern of lipid deposition may be attributed to the sexually dimorphic pattern of GH experienced by the two sexes.

Topics: Animals; Body Weight; Female; Growth; Growth Hormone; Lipid Metabolism; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Time Factors

Chronic human exposure to Cd results in kidney injury. It has been proposed that nephrotoxicity produced by chronic Cd exposure is via the Cd-metallothionein complex (CdMT) and not by inorganic forms of Cd. If this hypothesis is correct, then MT-null mice, which cannot form CdMT, should not develop nephrotoxicity. Control and MT-null mice were injected s.c. with a wide range of CdCl2 doses, six times/week for up to 10 weeks, and their renal Cd burden, renal MT concentration, and nephrotoxicity were quantified. In control mice, renal Cd burden increased in a dose- and time-dependent manner, reaching as high as 140 microg Cd/g kidney, along with 150-fold increases in renal MT concentrations, reaching 800 microg MT/g kidney. In MT-null mice, renal Cd concentration (10 microg/g) was much lower, and renal MT was nonexistent. The maximum tolerated dose of Cd in MT-null mice was approximately one-eighth that of controls. MT-null mice were more susceptible than controls to Cd-induced renal injury, as evidenced by increased urinary excretion of protein, glucose, gamma-glutamyltransferase, and N-acetyl-beta-D-glucosaminidase, as well as by increased blood urea nitrogen levels. Kidneys of Cd-treated mice were enlarged and histopathology showed various types of lesions, including proximal tubular degeneration, apoptosis, atrophy, interstitial inflammation, and glomerular swelling. These lesions were more severe in MT-null than in control mice, mirroring the biochemical analyses. These data indicate that Cd-induced renal injury is not necessarily mediated through the CdMT complex and that MT is an important intracellular protein in protecting against chronic Cd nephrotoxicity.

Topics: Acetylglucosaminidase; Animals; Body Weight; Cadmium Chloride; Cadmium Radioisotopes; Female; gamma-Glutamyltransferase; Glycosuria; Kidney; Kidney Diseases; Male; Metallothionein; Mice; Mice, Knockout; Proteinuria

Male Wistar rats at 2, 4, and 8 months were given s.c. injections of CdCl2 at doses of 0.5, 1, and 2 mg Cd/kg, 3 days/week, for 4 weeks. Dose-related adverse effects observed at the end of the injections were as follows: decrease in body weight gain, increases in liver, kidney, and spleen weights, decrease in red blood cell counts, and increase in white blood cell counts accountable by an increased percentage of neutrophils in peripheral white blood cells. Essentially, all of these changes were age-related, i.e., the extents of the effects in each age group were in the order of 8- > 4- > 2-month-old rats when compared at the same dose level of Cd. The sensitive indicators of histological changes were increase in hematopoietic cells in bone marrow and fibrous tissue proliferation in the thymus > fibrous tissue proliferation and hyperplasia of lymph follicle in the spleen > renal tubular degeneration. These histological changes became to be marked at lower doses with aging. Dose-dependent increases in total Cd concentrations in the liver, kidney, and spleen were slightly higher with aging, while metallothionein (MT) contents in these organs were induced exactly in the same pattern as Cd concentrations in each organ in various age groups. This study revealed that the adverse effects after repeated administration of Cd on the kidney, spleen, thymus, and bone marrow worsened with increasing animal age and that this phenomenon could not be explained simply by differences in Cd disposition or MT induction in the organs of the different age groups.

Topics: Age Factors; Animals; Blood Cells; Blood Chemical Analysis; Body Weight; Bone Marrow; Cadmium; Dose-Response Relationship, Drug; Kidney; Lymphoid Tissue; Male; Metallothionein; Rats; Rats, Wistar; Spleen; Thymus Gland; Time Factors

The effect of growth hormone (GH) on cytochrome P450 (CYP) and P450-dependent monooxygenases was studied in 4-, 6-, 8-, and 10-month-old female bovine growth hormone (bGH) transgenic mice that overexpress GH. Nontransgenic female mice (C57/SJL) littermates were used for baseline determinations. The body weights of the bGH mice were approximately 35% greater than those of the controls. The liver weights were 2-fold higher than those of the controls, resulting in a 25-60% increase in liver/body weight ratio during the life span of the bGH mice when compared with the controls. Similar increases in heart and kidney weights were observed. Since the GH transgene was transcriptionally regulated by a metallothionein-I gene promoter, metallothionein concentrations in livers of transgenic and nontransgenic mice were measured. No significant differences were observed. In marked contrast to increases in liver weights, hepatic cytochrome P450 content, benzphetamine N-demethylase, and benzo [a] pyrene hydroxylase activities were decreased by 36, 42 and 75%, respectively. No age-related changes in the decrease of the monooxygenases were observed. Microsomal heme oxygenase (HO) in the liver was induced 44% above the control values. Immunoblot analysis also showed a marked increase in HO-1 in the bGH mice. These results indicate that GH suppresses the carcinogen-metabolizing enzyme benzo [a] pyrene hydroxylase and the drug-metabolizing enzyme benzphetamine N-demethylase. This suppression was accompanied by an induction of HO activity in bGH transgenic mice. The consequences of prolonged exposure to supraphysiological levels of this hormone cannot always be predicted from the known physiological actions of GH.

Topics: Animals; Benzopyrene Hydroxylase; Body Weight; Cattle; Cytochrome P-450 Enzyme System; Female; Growth Hormone; Heart; Kidney; Liver; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microsomes, Liver; Organ Size; Oxidoreductases, N-Demethylating; Promoter Regions, Genetic; Reference Values

Mice transgenic for heterologous and ectopic GH expression serve as models for studying the feedback effects of elevated nonregulated GH on hypothalamic hypophysiotropic neurons as well as on peripheral function. For example, hypothalamic somatostatin expression has been shown to be increased markedly in mice bearing either bovine (b) or human (h) GH transgenes. Human, but not bovine, GH has lactogenic properties in mice, and appears to stimulate PRL-inhibiting tuberoinfundibular dopaminergic (TIDA) neurons. The present study was designed to determine the effect of a lifelong excess of hGH on dopamine (DA) expression in and numbers of TIDA neurons. Male mice of four transgenic lines were examined. The transgenic animals bore constructs of either bGH or hGH fused to either metallothionein (MT) or phosphoenolpyruvate carboxykinase (PEPCK) promoters; brains of transgenic mice were compared morphologically with those of nontransgenic littermates. Formaldehyde-induced catecholamine histofluorescence and tyrosine hydroxylase (TH) immunocytochemistry were examined in alternate brain sections; cell number was quantified for TIDA neurons (area A12) and a nonhypophysiotropic diencephalic DA area, the medial zona incerta (A13). Body weights were higher (P < 0.01) in PEPCK-GH than in MT-GH transgenic mice, as were serum levels of heterologous GH in those lines. In MT-hGH, but not MT-bGH or PEPCK-bGH, transgenic mice, A12 perikaryal fluorescence was enhanced, and ME fluorescence was reduced compared with those in control animals. The reduced ME DA is likely to reflect stimulation of TIDA neurons, because A12 TH-immunoreactive neuron number was increased by 34% in MT-hGH mice compared with that in controls (P < 0.05). In mice bearing the PEPCK-hGH construct, A12 TH neuron number was increased 47% (P < 0.001) compared with that in littermate controls. There were no differences in A13 cell number among animals, and A12 cell numbers in mice expressing bGH did not differ from control values. These results suggest that although extremely high levels of circulating bGH do not stimulate TIDA neurons, lifelong high levels of hGH have a stimulatory and graded effect on developmental differentiation of these cells for TH and DA production, supporting the concept of PRL as a trophic factor for TIDA neurons.

Topics: Animals; Body Weight; Cattle; Dopamine; Fluorescence; Growth Hormone; Human Growth Hormone; Humans; Hypothalamus, Middle; Male; Metallothionein; Mice; Mice, Transgenic; Neurons; Organ Size; Phosphoenolpyruvate Carboxykinase (GTP); Species Specificity; Tyrosine 3-Monooxygenase

Female mice were exposed to a single dose of 0.005 to 5 microg Cd/kg body wt., in order to test the hypothesis that once the Cd-binding capacity of intestinal metallothionein is saturated. Cd becomes more readily available for transfer from the mucosa to the circulatory system, causing an increase in Cd absorption. In this case the binding of Cd to MT would act as a barrier against Cd absorption, thus protecting the organism from accumulation and toxic effects of Cd in target organs such as the kidney. In mice the fractional Cd uptake (% of dose) in the duodenum, which was the main site of Cd uptake in the intestine, was not influenced by the Cd dose 6 h after dosage. However, the percentage of cytosolic Cd associated to MT in the duodenum decreased when the Cd dosage increased from 0.005 or 0.025 microg/kg to 5 microg/kg. Concomitantly, the percentage bound to low-molecular-weight (LMW) ligands increased, indicating saturation of the Cd-binding capacity of MT. Nevertheless, the fractional absorption was not dosage dependent in the dosage interval studied. Moreover, there was no statistically significant correlation between the percentage of cytosolic Cd bound to MT and the percentage of Cd absorbed. Thus, our results do not support the hypothesis that the intestinal Cd absorption is increased when the Cd-binding capacity of intestinal MT is saturated.

Topics: Administration, Oral; Animals; Body Burden; Body Weight; Cadmium; Cytosol; Female; Intestinal Absorption; Intestinal Mucosa; Intestine, Small; Kidney; Liver; Metallothionein; Mice; Mice, Inbred BALB C; Mucous Membrane; Organ Size

Eighteen transgenic mice carrying an ovine metallothionein la-ovine growth hormone (oMTla-oGH) transgene and 18 littermate normal mice were used to investigate the effects of transgene expression and clenbuterol administration on growth performance and skeletal muscle characteristics. The oGH transgene was activated from 21 d of age, and half of the mice were fed 15 ppm clenbuterol from 42 to 70 d of age. All mice were killed at 70 d of age after 4 wk of treatment, and organs and muscles were dissected, weighted, and analyzed. Transgenic mice (TM) gained 2.6 times more than normal mice (NM). However, TM had a significantly lower (-20%, P < .01) proportion of muscle, expressed as percentages of body weights, and a higher percentage of heart (+10%), liver (+26%, P < .01) and spleen (+64%, P < .01) than NM. Clenbuterol improved the weight gain of TM by 20%, compared with 10% for NM. The growth-promoting effect of clenbuterol was almost exclusively confined to skeletal muscle (24% increase) in NM, in contrast to a more generalized growth increase in all tissues including skeletal muscle (11% increase) in TM. The skeletal muscles of TM were longer but smaller in diameter due to 30% smaller muscle fiber cross-sectional area. Clenbuterol increased the muscle fiber size of all fiber types by 60% in NM, compared to 30% in TM. Muscle DNA concentrations and content were higher (P < .05) in TM than in NM, and clenbuterol administration decreased DNA concentrations but not total DNA content for both genotypes. Cathepsin B, C, and H activities were higher (P < .01) in TM muscle, but the significance is not clear at the present time, although it points to a potential for greater protein degradation and(or) turnover rates as suggested by smaller muscle weights.

Topics: Adrenergic beta-Agonists; Aging; Animals; Body Weight; Cathepsins; Clenbuterol; DNA; Female; Genotype; Growth Hormone; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Muscle Development; Muscle Fibers, Skeletal; Muscle, Skeletal; Organ Size; Random Allocation; RNA; Sheep

Mice with a temporally regulatable ovine metallothionein 1a--ovine growth hormone transgene (oMT1a-oGH) were utilized to study the effects of withdrawal of elevated circulating levels of growth hormone (GH) on growth and body composition. The transgene was activated from 21-42 days of age by provision of zinc sulfate in the drinking water. At 42 days, mice were allocated to either activated transgenic (remain on zinc sulfate) or inactivated transgenic (removal of zinc sulfate) groups, and to receive either ad libitum or restricted (80-90% of ad libitum) access to feed. Non-transgenic control mice were treated similarly. Body weights and intakes were recorded weekly. Mice were killed at 70 d and epididymal and subcutaneous fat pads, trimmed hind carcass and various organs were weighed. The main findings of this study are: (1) food-restricted mice possessing an activated oMT1a-oGH transgene fail to demonstrate increased growth, but exhibit significantly reduced levels of fat (P < 0.05) relative to all other genotype x feed level combinations; and (2) inactivation of the oMT1a-oGH transgene, following a period of elevated GH levels, leads to development of obesity as evidenced by two to three fold increases in epididymal and subcutaneous fat pad weights (P < 0.01) relative to both activated transgenic and non-transgenic control mice. These large increases in fat deposition also occurred when intake was restricted to 80-90% of ad libitum levels, indicating that metabolic changes independent of intake occur in these inactivated transgenic mice. It is possible that highly elevated production of GH in activated oMT1a-oGH transgenic mice leads to (1) enhanced promotion of preadipocyte differentiation, leading to increased numbers of adipocytes that, upon cessation of oGH production, are available for lipid deposition resulting in obesity, or (2) alterations in production of or responsiveness to insulin, leading to increased fat deposition upon removal of the chronic anti-lipogenic actions of GH. The oMT1a-oGH transgenic mouse line should provide a new genetic model with which to investigate the mechanisms by which growth hormone affects obesity.

Topics: Adipose Tissue; Animal Feed; Animals; Body Composition; Body Weight; Diet; Energy Intake; Female; Gene Expression Regulation; Growth Hormone; Male; Metallothionein; Mice; Mice, Inbred ICR; Mice, Transgenic; Obesity; Organ Size; Recombinant Proteins; Sulfates; Time Factors; Zinc Compounds; Zinc Sulfate

Cadmium (Cd) chloride was intravenously injected at doses of 0.05 and 0.5 mg/kg/day in ovariectomized rats for 50 weeks, and the chronic Cd exposure-induced nephrotoxicity and anemia were investigated. The rats treated with 0.05 mg/kg Cd showed no apparent hematological, urinary, and histopathological abnormalities. In the 0.5-mg/kg group, renal tubular disorders became marked at 16 weeks, and cortical fibrosis with glomerular dysfunction appeared at 50 weeks. Anemia occurred at 12 weeks in the 0.5-mg/kg group and became increasingly marked with time. The mean corpuscular volume (MCV) and mean corpuscular hemoglobin (MCH) were decreased at 12 and 25 weeks; however, the decreases of MCV and MCH disappeared at 50 weeks. A slight decrease in mean corpuscular hemoglobin concentration was noted at 50 weeks. The blood chemistry from the same group revealed a decrease in plasma iron levels and an increase in total iron binding capacity throughout the administration period. The erythropoietin (EPO) level was increased as the hemoglobin level decreased at 12 weeks, whereas the EPO level was not elevated even when the hemoglobin level was decreased at 50 weeks. These findings showed that renal anemia also occurred in addition to the iron deficiency anemia at 50 weeks.

Topics: Anemia; Animals; Body Weight; Cadmium; Cadmium Poisoning; Drinking; Erythropoietin; Female; Ferritins; Hemoglobins; Injections, Intravenous; Kidney; Kidney Diseases; Liver; Metallothionein; Ovariectomy; Rats; Rats, Sprague-Dawley; Urine

Metallothionein (MT)-bound zinc accumulates when animals are exposed to excess zinc and is depleted under conditions of zinc deficiency, suggesting that MT serves as a means of sequestering excess zinc as well as a zinc reservoir that can be utilized when zinc is deficient. To examine the importance of MT for these processes, mice with null alleles of both MT I and MT II genes were created and the zinc concentration and histological appearance of multiple organs assessed. At birth, the hepatic zinc concentration of these MT-null mice was lower than that of wild-type controls (0.27 +/- 0.02 vs. 0.65 +/- 0.11 micromol zinc/g tissue, P < 0.05). During the next 3 wk of suckling zinc-replete (95 micrograms zinc/g diet) dams, the hepatic zinc concentration of controls fell to 0.42 +/- 0.04 micromol/g but was unchanged in the MT-null mice (0.28 +/- 0.04 micromol/g). The most prominent histological anomaly observed at 3 wk of age was the presence of swollen Bowman's capsules in the kidneys of MT-null mice. When nursing MT-null dams were fed a severely zinc-deficient (1.5 microg/g) diet, kidney development in the MT-null pups was retarded as indicated by the retention of the nephrogenic zone and incomplete tubule development. We suggest that the lack of a hepatic reservoir of zinc jeopardizes the developing kidney in the MT-null mice. In addition to being more sensitive to dietary zinc restriction, MT-null mice are more sensitive to zinc toxicity. When adult mice were challenged with a ramping dose of zinc up to a total of 3700 micromol zinc/kg body weight, MT-null mice had a greater incidence of pancreatic acinar cell degeneration compared with control mice despite accumulating less zinc (2.72 +/- 0.46 vs. 1.23 +/- 0.52 micromol zinc/g pancreas, control and MT-null, respectively, P < 0.05). The results of these experiments suggest that MT I and MT II can protect against both zinc deficiency and zinc toxicity.

Topics: Animals; Animals, Newborn; Body Weight; Bone and Bones; Diet; Genotype; Kidney; Liver; Male; Metallothionein; Mice; Mice, Knockout; Pancreas; Zinc

Forty-eight transgenic mice carrying an ovine metallothionein 1a-ovine growth hormone (oMTla-oGH) transgene and 48 littermate control mice were used to investigate the effect of GH transgene on the growth and biochemical characteristics of skeletal muscle. Transgene expression was initiated in the transgenic mice by the addition of zinc sulfate to the water at 21 d of age; control mice were also supplemented with zinc sulfate. These mice were maintained on zinc sulfate until 84 d of age. Groups of mice (16 controls, 16 transgenics) were killed at 21, 42 and 84 d of age and muscles from the hind leg were dissected, weighed and analyzed. At 84 d, male transgenics were 32% heavier than controls, while female transgenics were 47% heavier. Transgenic mice of both sexes had smaller (p < 0.01) muscles than controls at weaning (21 d). In spite of significantly heavier body weights of transgenic mice at 84 d of age, there were no significant differences in muscle weights. This was due to a significantly lower (p < 0.01) proportion of muscle, expressed as percentages of body weights, in transgenic mice compared with controls. Higher DNA and RNA concentrations at 42 d of age and elevated cathepsins C and H activities at 42 and 84 d of age indicate that muscle protein metabolism is more active in transgenic mice, which are growing at a greater rate than controls from weaning to 84 d of age. The fact that oMTla-oGH transgenic mice inherently have a lower proportion of muscle, compared to controls and that this proportion does not change in spite of transgene activation and 30 fold increase in plasma GH levels, suggests the hypothesis that muscle growth may be controlled by locally produced IGFs, which are essentially independent of circulating GH concentrations.

Topics: Animals; Body Weight; Cathepsins; DNA; Female; Growth Hormone; Hindlimb; Male; Metallothionein; Mice; Mice, Transgenic; Muscle Development; Muscle Proteins; Muscle, Skeletal; RNA

To investigate the extra susceptibility of diabetics to some nephrotoxic agents, adult normal and diabetic Chinese hamsters (6-7 animals in each group) were injected subcutaneously with different doses of cadmium-metallothionein (Cd-MT) equivalent to 0.0, 0.1 or 0.25 mg Cd/kg body weight and the first 24 hr urinary outputs were collected. Several days prior to exposure to the Cd-MT the diabetic hamsters were hyperglycaemic, and plasma insulin levels and body weights were elevated in some of the diabetics. The higher dose of Cd-MT caused significant spillage of N-acetyl-beta-glucosaminidase (U-NAG) activity and protein into the urine of both normal and diabetic animals. The higher dose of Cd-MT was more toxic to the diabetic kidneys because U-NAG levels were higher in the diabetics (2.5-fold higher than normal). U-Cd levels were proportional to the injected Cd-MT dose. U-Zn levels were not consistently affected by the injected Cd-MT although it had contained small amounts of Zn. Therefore, genetic diabetes in the Chinese hamster appears to increase susceptibility to acute cadmium-MT nephrotoxicity. The mechanisms underlying this need to be further investigated.

Topics: Acetylglucosaminidase; Analysis of Variance; Animals; Blood Glucose; Body Weight; Cadmium; Cricetinae; Cricetulus; Diabetes Mellitus; Dose-Response Relationship, Drug; Injections, Subcutaneous; Insulin; Kidney; Metallothionein; Proteinuria

To explore the hypothesis that a second xenobiotic agent is required with excess copper to produce Indian Childhood Cirrhosis, this study investigated the effect of the pyrrolizidine alkaloid retrorsine fed to the mother during the suckling period upon the serial changes in neonatal copper status.. Female Wistar rats with new-born litters were fed either a control or a retrorsine (50 mg/kg) diet. At 0, 4, 8, 11, 15, 18 and 21 days, pups from each litter were weighed, sacrificed and their livers removed for copper, DNA and metallothionein analysis. Serum samples were assayed for caeruloplasmin oxidase activity and albumin.. 1) Higher than adult level of hepatic copper in normal rats which rose post-natally before declining from day 11 after birth, 2) raised hepatic copper concentrations and total copper in the retrorsine group from day 15; levels were higher than adult at birth, 3) reduced serum caeruloplasmin oxidase activity and albumin levels in retrorsine group, but both groups lower than adult, 4) lower hepatic metallothionein levels in retrorsine group, but both groups higher than adult, and 5) reduced liver DNA in the retrorsine group when expressed as total DNA and per gram of tissue. These changes were not secondary to under-nutrition as a small study on under-nourished rat neonates showed that copper handling is not significantly altered when compared to well-nourished rats.. Retrorsine passing to rat neonates via breast milk causes: 1) the accumulation of hepatic copper, 2) impairment of the rise in serum caeruloplasmin, which could indicate a decline in synthesis or failure of copper incorporation into the apo-protein, 3) a decrease in hepatic metallothionein and serum albumin levels, again suggesting diminished protein synthesis, and 4) reduced hepatic DNA indicative of decreased cell number but increased cell size. Accumulation of liver copper but reduction of copper-binding proteins could result in free copper and explain the synergistic hepatotoxicity of copper and retrorsine.

Topics: Animals; Animals, Suckling; Body Weight; Ceruloplasmin; Chemical and Drug Induced Liver Injury; Copper; DNA; Drug Evaluation, Preclinical; Female; Liver; Liver Diseases; Metallothionein; Milk; Nutrition Disorders; Organ Size; Pyrrolizidine Alkaloids; Rats; Rats, Wistar; Serum Albumin

Zinc homeostasis was studied during the induction, growth, and methotrexate (MTX) treatment of Dark Agouti rat mammary adenocarcinomas (DAMA). A progressive fall in plasma Zn concentration (pZn), significant at a tumor burden of less than 1% body weight (bw), was sustained during tumor enlargement to give a 54% reduction in pZn at 16.3% bw (n = 6/group). The hypozincemia was attributed to the increasing Zn demand for tumor growth. Zn content of the 16.3% bw tumors equaled that of muscle (normally 60% of total body Zn). Tumor metallothionein (tMT) was sufficient to bind < 3% of total tumor Zn, and hepatic MT (hMT) remained at basal concentrations during early tumor growth, doubling only in the presence of significant necrosis in large tumors. Methotrexate (MTX, 0.5 mg/Kg im x 2 d) at respective tumor burdens of 5 and 10% bw (n = 9, 10/group) gave 2 therapeutic effects, dependent on tumor size: 1.5% bw tumors in 7 rats remained close to their original size until experiment end when pZn, hMT, and tMT were typical of 5% bw untreated tumors. 2. Tumors in 5 rats given MTX at 10% bw had marked subcapsular necrosis and regression to a size similar to those in group 1; pZn returned toward normal, whereas hMT was 6 times its 5% bw counterpart. Host weight loss was significantly reduced, as were tumor-associated changes in plasma glucose and calcium. In summary, neither tMT nor hMT appears to play a role in the hypozincemia that follows DAMA Zn sequestration and growth. Critically timed MTX can result in tumor regression and return of plasma Zn, Ca, and glucose toward normal. This is associated with an increase in hMT and reduction in host weight loss, suggesting a flow of Zn from the resorbing tumor to the host, enabling the synthesis of hMT and retention of host structural proteins.

Topics: Adenocarcinoma; Animals; Blood Glucose; Body Weight; Calcium; Female; Homeostasis; Liver; Mammary Neoplasms, Experimental; Metallothionein; Methotrexate; Neoplasm Transplantation; Organ Size; Rats; Zinc

The effects of long-term administration of cadmium (Cd) chloride on the bone were studied using ovariectomized rats. The rats were injected iv with the compound at doses of 1.0 and 2.0 mg/kg, 5 days a week, for 13 weeks. The serum concentrations of calcium and inorganic phosphorus were significantly increased from 8 weeks in the 2.0 mg/kg group. The bone Cd content was gradually increased for 13 weeks in a dose-dependent manner. Calcium and phosphorus contents in the bone, and serum levels of parathyroid hormone and osteocalcin, were not significantly different between Cd-treated and control rats. Histopathologically, chronic Cd nephropathy such as tubular atrophy and interstitial fibrosis was observed with clinical polyuria and increased enzymuria. The skeletal changes were detected mainly in the femur and tibia. In the metaphysis of Cd-treated rats, cancellous bone mass increased with time. This change was detected as an increased opacity by a roentgenogram. In the cortical bone of the midshaft haversian canals were dilated with clearly bordered osteoid seams and showed a motheaten pattern in rats in the 2.0 mg/kg group at 13 weeks. In the present study, we report Cd nephropathy and osteomalacic changes in ovariectomized rats with iv injection of CdCl2 for 13 weeks. Although an involvement of the indirect action of Cd through renal failure could not be ruled out in this experiment, our biochemical and pathological data suggested that osteomalacia was induced by a direct action of Cd on the bone through abnormal calcium homeostasis.

Topics: Animals; Body Weight; Cadmium; Calcium; Female; Hematologic Tests; Kidney; Liver; Metallothionein; Osteocalcin; Osteomalacia; Osteopetrosis; Ovariectomy; Parathyroid Hormone; Phosphorus; Rats; Rats, Sprague-Dawley; Urinalysis

An interrelation of Cd-contents with hepato- and nephrotoxicities, and a mechanism for Cd-exclusion from kidneys were investigated in rats that received a single intravenous injection of either CdCl2 or Cd-saturated metallothionein II (Cd-MT-II) with doses of 0.1 and 0.3 mg Cd/kg body weight (b.w.). Between the livers and kidneys, higher uptake of Cd was observed in the liver in terms of CdCl2, and in the kidney in terms of Cd-MT-II. The CdCl2 at the two doses hardly showed any histopathological alterations in the livers and kidneys. The 0.1 mg Cd/kg b.w. of Cd-MT-II showed a slight injury in the kidneys, while hardly in the livers. At Day 1 of the 0.3 mg Cd/kg b.w. of Cd-MT-II, the renal Cd-contents reached the maximal value of 8.22 +/- 0.36 microgram/g wet tissue, and degeneration including necrosis and defluxion of proximal tubular cells were most highly observed. At Day 5 of the 0.3 mg Cd/kg b.w., the renal Cd-contents were lowered to 2.40 +/- 0.24 micrograms/g wet tissue, and fibrosis and regeneration of the proximal tubular cells were remarkably found. These findings strongly suggested that, in the case of administration of Cd-MT-II, the Cd taken into the kidneys was eliminated from there mainly by cellular death of the proximal tubulus and by their resultant defluxion.

Topics: Animals; Body Weight; Cadmium; Injections, Intravenous; Kidney; Liver; Male; Metallothionein; Rats; Rats, Sprague-Dawley

Transgenic mice with a bovine growth hormone gene linked to a mouse metallothionein I promoter (growth hormone transgenics) are a model of chronic growth hormone excess.. Growth of small bowel mucosa in ad libitum-fed growth hormone transgenics and wild type littermates and in growth hormone transgenics pair fed with wild-type littermates were compared.. In both groups, body weight and small bowel weight were greater in growth hormone transgenics. Similarly, mucosal mass was 50%-100% greater in growth hormone transgenics, and the effect was greatest in proximal bowel. Villus height, measured in jejunum, was also greater in growth hormone transgenics. Measurements of mucosal proliferation did not differ between the growth hormone transgenics and wild type. Abundance of insulin-like growth factor-I messenger RNA in bowel was greater in growth hormone transgenics.. Chronic growth hormone excess results in increased growth of small bowel mucosa. This effect appears to be specific because it occurred in ad libitum-fed and diet-restricted growth hormone transgenics, influenced villus height, and was more pronounced in upper than lower small bowel. The effect of chronic growth hormone excess does not appear to be secondary to an increase in the rate of mucosal proliferation, suggesting an effect on lifespan of mucosal cells.

Topics: Animals; beta-Galactosidase; Body Weight; Cell Division; DNA; Energy Intake; Female; Growth Hormone; Insulin-Like Growth Factor I; Intestinal Mucosa; Intestine, Small; Lactase; Male; Metallothionein; Mice; Mice, Inbred Strains; Mice, Transgenic; Organ Specificity; Poly A; Promoter Regions, Genetic; RNA; RNA, Messenger; Sucrase

Pituitary and serum levels of homologous growth hormone (GH) and characteristics of specific GH-releasing factor (GHRF) binding to pituitary homogenates were examined in transgenic mice expressing bovine GH (bGH) gene regulated by different promoters [mouse metallothionein-I (MT) or phosphoenolpyruvate carboxykinase (PEPCK)] and in their normal littermates. Pituitary GH concentration and GHRF binding were reduced by approximately 50% in transgenic MT-bGH mice in which serum bGH levels were about 20 micrograms/l and by approximately 95% in transgenic PEPCK-bGH mice in which serum bGH levels were tenfold higher. Suppression of plasma immunoreactive mouse GH (mGH) levels was detected in MT-bGH but not in PEPCK-bGH animals, presumably due to cross-reaction of the antiserum employed with bGH. Scatchard plots of GHRF binding to washed homogenates of pituitary glands from normal and young adult MT-bGH transgenic mice were curvilinear, indicating the presence of two types of binding sites, with low and high affinities. Both types of binding sites were reduced in number in MT-bGH transgenic mice without changes in their affinity. In 5-7-month-old MT-bGH transgenic mice there were changes in pituitary GH levels, in GHRF binding levels and in characteristics of GHRF binding that closely resembled the alterations described previously in aging rats. We conclude that over-expression of heterologous GH genes in transgenic mice can lead to partial or virtually complete suppression of somatotroph function, depending on the levels of heterologous GH in the circulation, and that transgenic MT-bGH mice exhibit symptoms of remarkably early onset of neuroendocrine aging.

Topics: Age Factors; Animals; Binding Sites; Body Weight; Gene Expression Regulation; Genes, Regulator; Growth Hormone; Growth Hormone-Releasing Hormone; Male; Metallothionein; Mice; Mice, Transgenic; Organ Size; Phosphoenolpyruvate Carboxykinase (GTP); Pituitary Gland; Radioimmunoassay; Recombinant Proteins

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 mumol CdCl2/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-mumol 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 mumol 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 micrograms/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 example, at the 30-mumol 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 micrograms 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-mumol dose.(ABSTRACT TRUNCATED AT 400 WORDS)

Topics: Aging; Animals; Body Weight; Cadmium; Female; Injections, Subcutaneous; Kidney; Liver; Male; Metallothionein; Mice; Mice, Inbred C3H; Mice, Inbred DBA; Organ Size; Sex Factors; Species Specificity; Testis; Tissue Distribution

Previously, we studied the ability of cadmium to initiate or promote tumors in B6C3F1 mice and, contrary to expectation, found that cadmium inhibited development of N-nitrosodiethylamine (NDEA)-initiated and sodium barbital-promoted liver tumors. In this study, the time course of cadmium inhibition of NDEA-initiated tumor formation was studied. A single dose of NDEA (90 mg/kg i.p.) was given at 5 weeks of age (time 0) followed by cadmium (1000 ppm) in drinking water from 2 to 48, 4 to 48, 8 to 48, 16 to 48 and 32 to 48 weeks. The study ended at 48 weeks. NDEA-induced elevations in liver tumor incidence (22 tumor-bearing mice/25 total) over control (5/25) were prevented by cadmium regardless of the period of administration (NDEA + cadmium: 2-48 weeks, 2/25; 4-48 weeks, 1/25; 8-48 weeks, 1/25; 16-48 weeks, 2/25; 32-48 weeks, 6/24). Cadmium alone (2-48 weeks) eliminated (0/25) spontaneously occurring liver tumors (5/25). NDEA-induced lung tumor incidence (25/25) and multiplicity (7.28 tumors/lung) were also reduced by cadmium (maximal decreases 28% and 80%, respectively). Some evidence of a specific deficiency of metallothionein in tumor cells was seen immunohistologically in NDEA-induced hepatic lesions and pulmonary lesions. These results indicate that cadmium prevents or reduces tumor formation in the B6C3F1 mouse liver and lung regardless of the exposure interval and apparently by cell-specific cytotoxicity. Auxiliary studies indicated that in mice bearing multiple liver foci resulting from NDEA treatment there was a marked reduction in basal metallothionein levels and in response to zinc induction.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Anticarcinogenic Agents; Body Weight; Cadmium; Diethylnitrosamine; Liver; Liver Neoplasms, Experimental; Lung; Lung Neoplasms; Male; Metallothionein; Mice; Mice, Inbred C3H; Mice, Inbred Strains; Time Factors

In mice transgenic for growth hormone (GH), the ectopic production of foreign GH causes gigantism and strong inhibition of endogenous pituitary GH. In human (h) GH transgenics, morphologic changes occurred not only in somatotrophs but in lactotrophs, corticotrophs, and gonadotrophs as well. To distinguish between changes attributed to somatotrophic effect of hGH from those caused by its lactogenic effects, we studied the pituitary morphology of bovine (b) GH transgenics since bGH has no lactogenic activity.. Pituitaries from transgenic mice and nontransgenic siblings were studied by immunocytochemistry for adenohypophysial hormones, in situ hybridization for GH, prolactin (PRL), and proopiomelanocortin mRNAs, and electron microscopy.. Transgenic mice had an increased body weight and a significantly decreased pituitary mass. In both sexes, GH immunoreactive cells were markedly reduced in size and moderately decreased in number, and the GH mRNA signal was lower compared with controls; ultrastructurally, in somatotrophs, the cytoplasmic organelles involved in hormone synthesis were inconspicuous. Males were normoprolactinemic, and lactotrophs showed no morphologic changes. In transgenic females, PRL immunoreactive cells were hypertrophic and appeared to be more numerous. In transgenic males, a mild increase in size and number of follicle-stimulating hormone/luteinizing hormone immunoreactive cells was noted. No changes were evident in corticotrophs and thyrotrophs in either sex. In the intermediate and posterior lobes, many corticotrophs and pituicytes were GH immunoreactive, indicating expression of metallothionein-1 (MT)-bGH transgene.. Pituitaries of MT-bGH transgenic mice contained somatotrophs with morphologic features of inhibition. Endogenous GH production was not completely suppressed as indicated by the presence of GH mRNA and GH immunoreactivity. Females had a mild increase in percentage of PRL immunoreactive cells, and previous biochemical data proved that they were hyperprolactinemic. Substantial differences in pituitary morphology between transgenic MT-bGH mice and MT-hGH animals studied previously can be due to the fact that bGH is purely somatotropic, whereas hGH is both somatotropic and lactogenic.

Topics: Adrenocorticotropic Hormone; Animals; Body Weight; Cell Count; DNA; Female; Follicle Stimulating Hormone; Gene Expression; Genes; Growth Hormone; Immunohistochemistry; In Situ Hybridization; Luteinizing Hormone; Male; Metallothionein; Mice; Mice, Transgenic; Microscopy, Electron; Organ Size; Organelles; Pituitary Gland; Pro-Opiomelanocortin; Prolactin; RNA, Messenger; Thyrotropin

Metallothionein (MT) induction in the rat prostate gland was investigated by means of cadmium chloride administration. Ten-week-old male Wistar rats housed with cadmium free food were divided into three groups of six rats each and castrated. After seven days, 1 mg of testosterone propionate per rat was injected subcutaneously once a day until the end of the experiment. After three weeks, rats were injected daily for six days with a physiological saline, 0.3 mg/kg CdCl2, and 0.9 mg/kg CdCl2. MT concentration of the ventral and dorsal lobes was significantly increased in the three groups in proportion to the dose of CdCl2. MT content of the lateral lobe in three groups was also increased, but was not significantly different. Immunohistochemically, MT was induced mainly in the ventral lobe in the basal cells, and in the lateral and dorsal lobes in the epithelial cells. The weights of the prostatic lobes were similar in the three groups, and no histological change was identified. These results suggested that MT in the prostate was induced by cadmium administration, and that it may prevent cellular damage from harmful metals.

Topics: Animals; Body Weight; Cadmium; Cadmium Chloride; Chlorides; Male; Metallothionein; Organ Size; Prostate; Radioimmunoassay; Rats; Rats, Wistar

The comparative influence of dietary zinc status and recombinant human interleukin-1 alpha (rhIL-1 alpha) and recombinant human interleukin-6 (rhIL-6) on metallothionein (MT) gene expression was examined in rat bone marrow and liver. Growing male rats were fed a diet with 5 (restricted), 30 (control), or 180 (supplemented) mg Zn/kg for 14 d. On d 15, rats were injected with 5 micrograms of rhIL-1 alpha or rhIL-6. Marrow metallothionein responded directly to dietary zinc but did not respond to these cytokines. Significantly less zinc accumulated in marrow from the zinc-restricted rats compared with control or supplemented rats. Analysis of metallothionein isoform mRNA expression showed MT-1 is the primary gene expressed in marrow. A significant interaction between dietary zinc and cytokine treatment was observed in the liver. Hepatic metallothionein induction following both rhIL-1 alpha and rhIL-6 injection was directly related to dietary zinc intake. Expression of hepatic metallothionein isoform mRNAs suggested MT-1 responded to zinc and MT-2 responded to cytokines. These results indicate that metallothionein gene expression in both the marrow and the liver responds to dietary zinc status. In contrast, liver metallothionein expression can be altered by these cytokines, which are known to act on many cell types. Furthermore, these results suggest that bone marrow metallothionein could be of importance in the development of marrow cells.

Topics: Administration, Oral; Animals; Body Weight; Bone Marrow; Diet; Dose-Response Relationship, Drug; Gene Expression; Interleukin-1; Interleukin-6; Liver; Male; Metallothionein; Rats; Rats, Sprague-Dawley; RNA, Messenger; Zinc

Cadmium-metallothionein (Cd-MT) may have a role in the pathogenesis and irreversibility of Cd nephrotoxicity. In the present study, rats were injected with 0.3 mg Cd/kg body wt per week as Cd-MT for 5 consecutive weeks and a group of rats (n = 3) was killed 24 hr after each injection. A group of three rats was kept for an additional week after the 5 weeks of Cd-MT injection for recovery. After the first injection, urinary Cd and protein levels and kidney/body wt ratio were increased. The electrophoretic pattern of urinary protein showed increased excretion of low-molecular-weight proteins, especially after the first injection of Cd-MT. Tubular cell necrosis occurred after the first week with renal Cd levels of only 10 micrograms/g and gradually progressed to severe necrosis with inflammation in 3 weeks and then to interstitial fibrosis in 5 weeks. The levels of Cd and MT in kidney increased with repeated injection of Cd-MT, but renal Cd was about 40 micrograms/g after 5 weeks of injection. Urinary Cd and MT levels progressively increased during the Cd exposure period, but returned to pretreatment levels during the sixth week (recovery period). Renal cell necrosis and inflammation were absent at the sixth week, but interstitial fibrosis persisted. This study indicates that nephrotoxicity of Cd in this model is related to urinary excretion of Cd-MT and that renal cell injury may be independent of Cd in the renal cortex. Nephrotoxicity occurs at levels much lower than the proposed critical concentration for Cd (200 micrograms Cd/g) following long-term exposure to CdCl2. However, in the absence of continued Cd exposure from liver or circulation, the Cd-MT-induced renal damage is reversible.

Topics: Analysis of Variance; Animals; Body Weight; Cadmium; Enzyme-Linked Immunosorbent Assay; Kidney; Kidney Tubules, Proximal; Liver; Male; Metallothionein; Necrosis; Proteinuria; Rats; Rats, Sprague-Dawley

The early changes in hepatic metallothionein (MT) and plasma zinc (Zn), copper (Cu), and iron (Fe) were investigated during the induction of adjuvant (AJ) arthritis in rats in conjunction with cyclosporin (CsA) treatment. Plasma Zn decreased after AJ injection (60% of control values at 8 h), and this was associated with a 4.5-fold increase in hepatic MT at 8 h. Plasma Zn was lowest at 16 h (40% of control), whereas hepatic MT concentrations increased to a maximum of 20-fold at 16 h. Changes in plasma Fe paralleled those of Zn, whereas plasma Cu levels were increased. Plasma metal and hepatic MT concentrations returned toward normal from d 1-7. At d 14, when marked paw swelling was apparent, hepatic MT and plasma Cu were again increased and plasma Zn decreased. Administration of CsA decreased MT induction in rats injected with AJ and also caused a marked recovery in plasma Zn and Fe levels. These changes were small but significant even in the early stages (up to 24 h) after AJ injection and were followed by a sustained improvement in all parameters, corresponding to the nonappearance of clinical arthropathy in CsA-treated rats. TNF-alpha and IL-6 production by peritoneal macrophages isolated from AJ-injected rats was significantly decreased by CsA treatment at d 7 and 14. The inhibition of hepatic MT induction during acute and chronic inflammation by cyclosporin emphasizes the role of the immune system in altered metal homeostasis in inflammation.

Topics: Animals; Arthritis, Experimental; Body Weight; Copper; Cyclosporine; Eating; Interleukin-6; Iron; Liver; Macrophages; Male; Metallothionein; Peritoneal Cavity; Rats; Trace Elements; Tumor Necrosis Factor-alpha; Zinc

The influence of 12-O-Tetradecanoylphorbol-13-acetate, an activator of proteinkinase C and A 23187, a calcium ionophore increasing cytosolic free calcium concentration on zinc metabolism was investigated in a study with 24 eight-week old rats. Twenty-four hours before killing, the rats (235 g body weight, 8 per group) were either injected intraperitoneally with TPA (1.6 x 10(-7) mol/kg body weight) or A 23187 (1.6 x 10(-6) mol/kg body weight). Control rats received the solvent dimethylsulfoxide. The application of TPA and A 23187 provoked a marked decline in feed intake accompanied by a reduction in body weight and liver mass. Serum concentrations of zinc were reduced significantly after A 23187 injections. TPA and A 23187 increased liver zinc levels by 20 and 30% respectively, if based on fresh and dry weight. The injections, however, did not alter total liver zinc. Liver metallothionein (MT) concentration was elevated 2.4-fold after TPA administration. The increase in response to A 23187 was only 1.5-fold and not significant. Mucosa MT levels were not altered. Serum activity of alkaline phosphatase was significantly reduced (TPA: -23%, A 23187: -31%). There was no change in serum glucose after injections. However, serum creatinine and urea were increased in response to A 23187. In conclusion, TPA and A 23187 had an effect on zinc metabolism of the rat, most marked in the case of MT induction in the liver. There is evidence that the reduced feed intake caused by TPA and A 23187 resulted in effects indistinguishable from those caused by fasting. Further experiments are needed to clarify whether proteinkinase C and cytosolic free calcium are directly involved in the regulation of zinc metabolism.

Topics: Alkaline Phosphatase; Animals; Blood Glucose; Body Weight; Calcimycin; Creatinine; Eating; Liver; Male; Metallothionein; Organ Size; Rats; Rats, Inbred Strains; Tetradecanoylphorbol Acetate; Urea; Zinc

Transgenic mice were developed by injecting a mouse metallothionein promoter-human growth hormone (Mt-hGH) gene fragment into the pronucleus of C57Bl x DBA/2J-f2 or C57Bl x CBA-f2 one cell embryos. Six founder animals with the C57Bl x DBA genetic background grew 1.3-2.2 times larger than littermate controls and had higher levels of hGH in plasma (4.6-279 mU/l). Three of the four female transgenic founders developed malignant papillar adenocarcinomas of mammary origin at 27-43 weeks of age. One male transgenic founder was successfully mated and two of three female transgenic offsprings developed mammary tumors. To examine if the tumor induction was dependent on the strain of mice used the experiments were repeated using animals with different genetic background. Fourteen female hGH transgenic mice from five founder animals were generated using C57Bl x CBA-f2 mice. Thirteen of the animals had elevated levels of hGH in plasma (7-1960 mU/l) and grew larger than control animals. Nine of the animals developed mammary adenocarcinomas. Four of the hGH expressing animals did not demonstrate macroscopic tumor formation but have not yet been analyzed histologically. The present study suggests that markedly elevated endogenous levels of GH cause mammary carcinoma in hGH transgenic mice. The present animal model might prove useful for studying molecular mechanisms involved in the development of hormonally induced mammary tumors.

Topics: Adenocarcinoma, Papillary; Animals; Body Weight; Female; Growth Hormone; Humans; Male; Mammary Neoplasms, Experimental; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Inbred DBA; Mice, Transgenic; Promoter Regions, Genetic; Recombinant Fusion Proteins

Experiments were conducted with adult male rats to investigate the effects of dietary calcium (Ca) restriction upon intake and tissue distribution of cadmium (Cd), and Cd-metallothionein (Mt) synthesis. Four groups of animals were fed either a low-Ca, semisynthetic diet (0.1% Ca) or the same diet supplemented with 0.8% Ca (normal diet). The caloric intake was similar in all groups. Two groups (low-Ca and normal diet) were used as controls, and two groups (low-Ca and normal diet) received 100 mg/l Cd (as CdCl2) in drinking water. Cd levels in liver, kidney, spleen and red cells were measured in all animals after 8 weeks of treatment. Concomitantly, Mt levels in plasma, liver and kidney were evaluated by radioimmunoassay. Ca deficiency entailed marked and significant increases in accumulation of Cd and synthesis of Mt in all assayed tissues. It is concluded that dietary Ca restriction, independent of caloric intake, enhances Cd intestinal absorption and tissue accumulation, which is followed by increased tissue Mt synthesis.

Topics: Administration, Oral; Animals; Body Weight; Cadmium; Calcium; Calcium, Dietary; Intestinal Absorption; Liver; Male; Metallothionein; Organ Size; Rats; Rats, Inbred Strains; Tissue Distribution

The effect of maternal copper (Cu) deficiency on various proteins was studied to determine if the changes were reversible or persistent with Cu repletion. The functional consequences of these alterations were assessed by exposing the animals to an oxidative stress (endotoxin), and by measuring the formation of benzo[a]pyrene-DNA adducts in vitro. Throughout gestation and lactation, mice were fed a Control diet (10 micrograms Cu/g diet) or a Low Cu diet (1 microgram Cu/g diet). On day 18, the offspring were killed or switched to the Control diet and killed on day 42 following a single injection of saline or endotoxin on day 41. In day-18 offspring, Cu deficiency resulted in decreased hematocrit values, ceruloplasmin activity, liver and tissue Cu levels, and metallothionein concentrations. Cu repletion restored all but metallothionein levels. Early Cu deficiency led to higher brain CuZn superoxide dismutase activity on day 42, and higher levels of brain thiobarbituric acid reactive substances (TBARS) in endotoxin-treated mice. Liver TBARS were lower in day-18 Low Cu offspring and in day-42 Low Cu offspring treated with endotoxin than age-matched Controls. Cytochrome P-450 concentrations were lower in Low Cu, endotoxin-treated males than in Controls. These results show that Cu deficiency-mediated alterations during early development are not immediately reversed with Cu repletion.

Topics: 7-Alkoxycoumarin O-Dealkylase; Animals; Benzo(a)pyrene; Body Weight; Brain; Ceruloplasmin; Copper; Cytochrome P-450 Enzyme System; Embryonic and Fetal Development; Female; Hematocrit; Hemoglobins; Kidney; Liver; Metallothionein; Mice; Mice, Inbred Strains; Organ Size; Pregnancy; Pregnancy Complications; Superoxide Dismutase; Thiobarbiturates; Trace Elements

Pregnant Sprague-Dawley rats were intraperitoneally injected with physiological saline solution (vehicle) or cadmium chloride (CdCl2) at 2.5 mg kg-1 body wt. on days 8, 10, 12 and 14 of gestation. Offspring were examined for renal alkaline phosphatase activity (ALP) on postnatal days (PND) 3 and 12, and for kidney metallothionein (MTh) and for liver, kidney and entire gastrointestinal tract 109Cd content at birth and on PND 3 and 12. No effects were observed on neonatal survival or on body, liver and kidney weights of pups up to PND 12. Newborns born and fed by mothers exposed to CdCl2 during pregnancy exhibited a significant decrease in ALP activity on PND 3. Conversely, no significant changes were observed in newborns lactated by surrogate non-treated mothers. Renal MTh increased with age but was not influenced by maternal treatment. Traces of 109Cd were present in the liver at birth (5-7 ng). Thereafter, 109Cd was mainly found in the gastrointestinal tract of newborns lactated by their biological mothers (610-690 ng on PND 12), with a marginal uptake in the liver (10-12 ng on PND 12). 109Cd was not detectable in the kidneys at any age (less than 4 ng). These results show that prenatal exposure to Cd cannot be the sole aetiological agent in the induction of the subtle and transitory changes in renal biochemistry observed in offspring born and fed by female rats intraperitoneally injected with 2.5 mg CdCl2 kg-1 body wt. on days 8, 10, 12 and 14 of gestation. The results also contradict the role of a direct effect on the kidney.

Topics: Alkaline Phosphatase; Animals; Animals, Newborn; Body Weight; Cadmium Poisoning; Cadmium Radioisotopes; Female; Kidney; Lactation; Liver; Male; Metallothionein; Organ Size; Pregnancy; Rats; Rats, Inbred Strains

In the present study we examined the effects of zinc pretreatment (to induce the renal synthesis of metallothionein) on the renal accumulation and intrarenal distribution of inorganic mercury in uninephrectomized (NPX) and sham-operated (SO) rats 24 h after the animals were given a 0.75, 1.0 or 1.5 mumol/kg intravenous (i.v.) dose of inorganic mercury. We also examined the effects of zinc pretreatment on the nephropathy induced by the three doses of inorganic mercury. Zinc was administered at a dose of 306 mumol/kg (20 mg/kg) subcutaneously (s.c.) in the form of zinc sulfate once daily for 2 consecutive days prior to the administration of inorganic mercury. Following zinc pretreatment, the renal accumulation of injected inorganic mercury increased in both NPX and SO rats treated with the three doses of inorganic mercury, but the increase was significantly greater in the NPX rats. The enhanced accumulation of mercury was associated with an altered pattern in the intrarenal distribution of mercury, particularly in the NPX rats. The increased renal accumulation of mercury in both the NPX and SO rats was due primarily to its increase in the renal cortex. We have recently found that the synthesis of metallothionein in the renal cortex increases in NPX and SO rats given zinc. Therefore, it appears that there is a relationship between the content of preinduced cellular metallothionein in the cortex and the content of mercury that accumulates in the cortex. Zinc pretreatment also prevented the nephropathy induced by the three doses of inorganic mercury from occurring in both the NPX and SO rats. We propose that some of the protection may be related to the altered intrarenal accumulation and distribution of mercury that occurs after pretreatment with zinc. Hepatic accumulation of mercury also increased in both groups of rats, but the increase again was significantly greater in the NPX rats. Our findings show clearly that a significant reduction in renal mass alters the hepatic and renal accumulation of mercury when zinc pretreatment is used to induce the renal and hepatic synthesis of metallothionein. In addition, our findings show that zinc pretreatment protects both normal and remnant kidneys in rats from the nephrotoxic effects of inorganic mercury.

Topics: Animals; Body Weight; Dose-Response Relationship, Drug; Kidney; Liver; Male; Mercuric Chloride; Metallothionein; Nephrectomy; Organ Size; Rats; Rats, Inbred Strains; Zinc

The effects of unilateral nephrectomy and compensatory renal growth on renal metallothionein metabolism were evaluated in the present study. In rats, the renal content of metallothionein increased in proportion to the increase in renal mass after unilateral nephrectomy and compensatory renal growth. However, when zinc was used to induce the synthesis of renal metallothionein, the remnant kidney in uninephrectomized (NPX) rats produced significantly greater amounts of metallothionein on a per gram kidney basis than a normal kidney in sham-operated (SO) rats. In both NPX and SO rats, zinc pretreatment caused metallothionein synthesis to increase primarily in the renal cortex and renal outer stripe of the outer medulla. Zinc pretreatment also changed the pattern for the intrarenal accumulation of inorganic mercury in NPX rats. After pretreatment with zinc, the accumulation of inorganic mercury predominated in the renal cortex rather than in the outer stripe of the outer medulla in the NPX rats. In addition, both NPX and SO rats were afforded complete protection against the nephrotoxic effects of a low, toxic dose of inorganic mercury when they were pretreated with inorganic zinc. The protection is postulated to be related to the alteration in the pattern of renal accumulation of inorganic mercury. In conclusion, the capacity to synthesize metallothionein increases significantly in rats after they have undergone unilateral nephrectomy and compensatory renal growth. The increased capacity of the remnant kidney to synthesize metallothionein may involve adaptive changes both in transcriptional and/or translational controls of metallothionein synthesis.

Topics: Animals; Body Weight; Kidney; Male; Mercuric Chloride; Metallothionein; Nephrectomy; Organ Size; Rats; Rats, Inbred Strains; Zinc

The distribution of cadmium was examined in rats fed diets containing either cadmium-metallothionein (CdMt) or cadmium chloride (CdCl2) for 4 weeks. The test diets contained 3, 10, or 30 mg Cd/kg diet (3, 10, or 30 ppm) as CdMt or 30 mg Cd/kg diet (30 ppm) as CdCl2. A second study was performed to establish the Cd content in liver and kidneys after exposure to low doses of both CdMt and CdCl2 (1.5 and 8 ppm Cd). The feeding of CdMt resulted in a dose- and time-dependent increase of the Cd concentration in liver, kidneys, and intestinal mucosa. Rats fed 30 ppm CdMt consistently showed less Cd accumulation in liver and intestinal mucosa than did rats fed 30 ppm CdCl2. However, renal accumulation in rats fed 30 ppm was similar until Day 28 regardless of Cd form. At lower dietary Cd levels (1.5 and 8 ppm), relatively more Cd is deposited in the kidneys, although even at these doses the kidney/liver ratio of Cd is still higher with CdMt than with CdCl2. Tissue metallothionein (Mt) levels in the intestinal mucosa were relatively constant but always higher after CdCl2 exposure than after CdMt exposure. Mt levels in both liver and kidney increased after CdCl2 or CdMt exposure during the course of study. Although Mt levels in liver were higher after CdCl2 intake (30 ppm) than after CdMt intake (30 ppm), renal Mt concentrations were the same for both groups. In fact on Day 7, CdMt administration resulted in slightly higher Mt levels than CdCl2 administration, suggesting a direct accumulation of exogenous CdMt in the kidneys. In conclusion, after oral exposure to CdMt in the diet there is a relatively higher Cd accumulation in the kidneys. However, the indirect renal accumulation via redistribution of Cd from the liver might be lower than after CdCl2 exposure. Which of these two phenomena is decisive in the eventual level of renal toxicity of Cd after long-term oral intake could determine the toxicological risk of the chronic intake of biologically incorporated Cd.

Topics: Animals; Body Weight; Cadmium; Cadmium Chloride; Diet; Dose-Response Relationship, Drug; Intestinal Mucosa; Kidney; Liver; Male; Metallothionein; Rats; Rats, Inbred Strains

Body weight gain and external body dimensions of MT-hGH transgenic mice were compared with mice (NMRI) selected for high 8-week body weight (N8) and unselected controls derived from the NMRI strain (Pop). The growth curves from day 30 to 120 of transgenic mice exhibited a significantly steeper slope than those of male and female controls and of female N8 mice and did not show sex-related differences. The continuous pattern of GH secretion in transgenic mice is discussed as a possible reason for this phenomenon. Body weight gain of transgenic mice did not significantly exceed that of male N8 mice. None of the groups showed an obvious prolongation of the period of rapid daily weight gain. Maximum body weights of male and female transgenic mice were significantly higher than those of sex-matched controls but not of N8 mice. A drastic loss of body weight of about 25% of the maximum value was observed in the transgenic group prior to death. External body dimensions were largest in MT-hGH transgenic animals, followed by N8 mice and controls. In addition to these absolute measurements, values were related to the cube root of maximum body weight of the same animal. This is the first study that provides a comparative analysis of the effects of GH gene transfer and selection for body weight gain on body growth of mice derived from an outbred strain.

Topics: Animals; Biometry; Body Weight; Female; Growth Hormone; Male; Metallothionein; Mice; Mice, Transgenic; Sex Characteristics; Time Factors; Weight Gain

Growth hormone and mechanical loading are known to be important factors influencing bone growth. We have measured proportions of the skull and the postcranial skeleton of metallothionein-human growth hormone (MT-hGH) transgenic mice expressing high levels of hGH in their serum, of NMRI mice being large as a result of selection for high 8-week body weight (N8), and of unselected controls (Pop) derived from the NMRI strain. Absolute bony dimensions of transgenic mice were as a rule significantly larger than those of controls, the differences ranging between 3% and 32% in males and from 6% to 28% in females. By contrast, the enlargement of skeletal dimensions of N8 mice did not exceed 10% and was restricted to distinct bones. When related to the cube root of maximum body weight of the same animal, bones of controls were as a rule larger than those of N8 and MT-hGH transgenic mice. A detailed analysis of bony dimensions of GH transgenic mice and of mice selected for high body weight was carried out to judge the effects of GH overexpression and mechanical loading due to increased body weight on bone growth. The fact that bones of transgenics were as a rule larger than those of selected mice in spite of both groups reaching similar maximum body weights, suggests that skeletal gigantism in MT-hGH transgenic mice can only in part be a result of increased body weight.

Topics: Animals; Body Weight; Bone Development; Female; Growth Hormone; Male; Metallothionein; Mice; Mice, Transgenic; Sex Characteristics; Time Factors; Weight Gain

Dose-dependent changes in the concentration of metallothionein-1 (MT-1) in rat tissues were determined following subcutaneous administration of paraquat (PQ), a superoxide radical-generating agent. At 24 hr after injection. MT-1 concentrations in the lung increased linearly with PQ dose. Concentrations in the liver increased with dose until a plateau was reached at a dose of 30 mg/kg body wt. In the kidneys, MT-1 concentrations did not increase even at high doses of PQ. Zn was the principal metal bound to MT in the liver. PQ-induced MT-1 synthesis may reflect de novo protein synthesis, since the increase in MT-1 in the liver was reduced by pretreatment of the rats with actinomycin D. Although MT-1 synthesis in the liver was well correlated with lipid peroxidation, as evaluated by measurement of thiobarbituric acid-reactive substances, there was no such correlation in the kidneys. PQ increased the lipid peroxidation in kidneys without any increase in MT-I content. The data suggest that enhanced lipid peroxidation is not necessarily related to the increased MT-1 synthesis caused by radical-generating agents.

Topics: Animals; Body Weight; Dactinomycin; Dose-Response Relationship, Drug; Lipid Peroxidation; Liver; Lung; Male; Metallothionein; Organ Size; Paraquat; Rats; Rats, Inbred Strains; Superoxides; Zinc

To develop transgenic mice, a mouse metallothionein promoter-human growth hormone (Mt-hGH) gene fragment was injected into the pronucleus of C57BI x DBA/2J F2 one-cell embryos. Six founder animals grew larger (1.3-2.2 times) than littermate controls and had higher levels of hGH in plasma (4.6-279 mU/l). At 27-43 weeks of age, 3 of the 4 female transgenic founders developed malignant papillary adenocarcinomas of mammary origin. One of the male transgenic founders was successfully mated and 2 of 3 female transgenic offspring developed mammary tumors. Forty-two female mice with the same genetic background as the transgenic animals and older than 43 weeks served as controls. No palpable tumor was found in this group. Five of the control animals were killed and examined histologically, revealing only normal mammary tissue. Earlier studies have shown that GH is important for growth and development of the mammary gland. Our study suggests that markedly elevated endogenous levels of GH cause mammary carcinoma in a specific strain of transgenic mice. The present animal model might be useful for studying molecular mechanisms that are involved in the development of hormonally induced mammary tumors.

Topics: Adenocarcinoma; Animals; Body Weight; Female; Gene Expression; Genetic Vectors; Growth Hormone; Male; Mammary Neoplasms, Experimental; Metallothionein; Mice; Mice, Transgenic; Promoter Regions, Genetic

Organ weights and the distribution of zinc and copper were compared in HLA/ICR mice that received intraperitoneal injections of 10 micrograms of Serratia marcescens lipopolysaccharide W or of sterile physiologic saline at 2 d of age. Between 5 and 28 d of age, body weight gains were similar in both groups. At 5 and 7 d of age, lipopolysaccharide W-treated mice had significantly lower thymus weights (p less than 0.01). At 7 d of age, liver weight was significantly increased (p less than 0.01) in lipopolysaccharide W-treated mice. Compared with tissue copper concentration in coeval saline-treated mice, lipopolysaccharide W treatment significantly increased copper concentration in thymus at 5 d of age (p less than 0.05) and significantly decreased concentration of this metal in liver at 7 d of age (p less than 0.05) and in spleen at 14 d of age (p less than 0.05). Liver zinc concentration was significantly lower (p less than 0.05) in 28-d-old mice that had received lipopolysaccharide W. When expressed on the basis of total organ burdens of zinc or copper, only the liver burden of zinc in 5-d-old lipopolysaccharide W-treated mice was significantly increased (p less than 0.05). Lipopolysaccharide W treatment consistently decreased copper concentration in liver cytosol and the amounts of zinc and copper bound to metallothionein, a transition metal-binding protein, in liver cytosol. These effects of lipopolysaccharide W on organ size and metal distribution may contribute to the adverse effects that persist after endotoxin exposure in early life.

Topics: Animals; Animals, Newborn; Body Weight; Copper; Endotoxins; Injections, Intraperitoneal; Lipopolysaccharides; Liver; Metallothionein; Mice; Serratia marcescens; Thymus Gland; Zinc

A study has been made of factors which may influence the induction of metallothionein-I (MT-I) synthesis by the superoxide radical generating agent, paraquat (PQ). Hepatic concentrations of zinc (Zn) and MT-I increased in rats injected with PQ (40 mg/kg, s.c.) or fasting, but were greater in the former. Renal concentration of MT-I increased in fasted rats but not in PQ-treated rats. The data suggest that the increase in MT-I concentrations in PQ-treated rats is not caused by reduction in food intake. Administration of PQ increased hepatic concentrations of Zn, MT-I and thiobarbituric acid-reactive substances (TBA-RS), indicating the occurrence of lipid peroxidation. Treatment of rats with vitamin E (400 mg/kg, s.c.) on 4 successive days before injection of PQ prevented only the enhancement of lipid peroxidation. The data indicate that the induction of MT synthesis by PQ is not correlated with enhancement of lipid peroxidation. Similar results were obtained in the liver of rats subjected to the radical-generating conditions, such as fasting and exposure to carbon tetrachloride. Free radicals may induce MT synthesis by direct or indirect mechanisms.

Topics: Animals; Body Weight; Carbon Tetrachloride Poisoning; Fasting; Free Radicals; Kidney; Lipid Peroxides; Liver; Lung; Male; Metallothionein; Organ Size; Oxidation-Reduction; Paraquat; Rats; Rats, Inbred Strains; Stress, Physiological; Vitamin E; Zinc

Zn and Cu concentration, content, and distribution in lung were quantitated in neonatal and adult Lewis rats. Total lung Cu and Zn content rose dramatically during postnatal development, paralleling increases in lung and body weight. Cu concentration was 2-fold higher in adult lung than in neonatal lung whereas Zn concentration was unaffected by developmental age. A Cu-containing protein with an approximate molecular weight of 10,000 Da was identified immunologically as metallothionein (MT). At term, native MT concentration in the lung was 4-fold higher than in adult lungs. By day 4 postpartum, MT concentration was reduced by half and reached a level characteristic of the adult by 7 days postpartum. MT was localized in nuclear and cytoplasmic compartments of positive lung cells by immunofluorescent techniques; initially, nuclear MT staining was more intense than cytoplasmic staining.

Topics: Animals; Animals, Newborn; Body Weight; Copper; Lung; Metallothionein; Molecular Weight; Organ Size; Rats; Rats, Inbred Lew; Zinc

The structural gene coding for human growth hormone (hGH) was fused to the promoter of the mouse gene for metallothionein-1. These fusion genes were introduced into pronuclei of the fertilized eggs of mice by micro-injection. We have injected MT-hGH genes into 121 mouse eggs which were transferred into the oviducts of eleven pseudo-mothers. Seven out of eleven mothers gave birth. Forty-three litters were born. These litters were grown-up and their tails were cut for isolating the DNA. The DNA of each mouse was examined by dot hybridization and Southern blotting. We found that eighteen mice had incorporated the fusion genes and grew larger than control mice. Synthesis of human growth hormone was induced further by zinc, which normally induces metallothionein gene expression. We also isolated the RNA from the livers of transgenic mice. The RNA was hybridized with hGH gene. It showed that on the transcription level the hGH gene was transcribed into mRNA in the transgenic mice. Using the same method to examine the second and third generation of the transgenic mice, we found that the incorporated gene could be inherited for generations. But the body-weight of transgenic mice appeared to decrease in the progeny. The offspring obtained from different combinations of parents (carrying or not carrying hGH gene) were different.

Topics: Animals; Blotting, Northern; Blotting, Southern; Body Weight; Cloning, Molecular; Growth Hormone; Humans; Male; Metallothionein; Mice; Mice, Inbred ICR; Mice, Transgenic

Stress and starvation increased liver metallothionein (MT) and decreased liver glutathione (GSH) levels. Serum cysteine plus cystine levels were increased by stress. The exogenous administration of GSH, while not modifying hepatic GSH content, increased liver MT levels in basal and starved rats but not in stressed rats. Liver and serum cysteine levels were increased by GSH administration, a process partially reverted by the irreversible inhibitor of gamma-glutamyl transpeptidase, alpha-amino-3-chloro-4,5-dihydro-5-isoxazoleacetic acid. Mouse and rat liver MT levels were also increased by buthionine sulfoximine, an inhibitor of GSH synthesis, indicating that GSH is not a necessary precursor of MT. In addition, the hepatic MT content was increased by the administration of cysteine in a dose-response manner. These results suggest that hepatic MT synthesis is elevated by increased cysteine pools, and that MT, GSH and cysteine levels are somehow inter-related. MT, besides GSH, may be contemplated as a putative intracellular reservoir of cysteine in the liver of adult rats.

Topics: Animals; Body Weight; Buthionine Sulfoximine; Cysteine; Glutathione; Isoxazoles; Liver; Male; Metallothionein; Methionine Sulfoximine; Organ Size; Rats; Rats, Inbred Strains; Starvation; Stress, Physiological; Zinc

To examine the combined hepatotoxic and nephrotoxic effects of cadmium and ethanol, rats maintained on an ethanol containing liquid diet (5% w/w) were given cadmium either acutely (3 x 1 mg/kg IP) or subacutely (about 14 mg/kg/day PO for 6 weeks). Parameters tested were cadmium, zinc and copper contents of blood and various organs, metallothionein (MT) contents, polysome profile of liver and kidneys, serum SDH and GPT levels and creatinine clearance. Ethanol reduced the hepatic MT contents without altering the polysome profile and the zinc and copper contents. Cadmium on the other hand raised the MT contents in liver and kidneys. This effect of cadmium predominated in the combined treatment. Morphological examination and functional tests (SDH, GPT, creatinine clearance) indicate that cadmium does not enhance the toxic effects of ethanol, and vice versa.

Topics: Alanine Transaminase; Animals; Body Weight; Brain Chemistry; Cadmium; Chemical and Drug Induced Liver Injury; Copper; Creatinine; Ethanol; Female; Kidney Diseases; L-Iditol 2-Dehydrogenase; Metallothionein; Organ Size; Polyribosomes; Protein Biosynthesis; Rats; Rats, Inbred Strains; Zinc

Pretreatment of rats with Na2MoO4 (1.24 mmol/kg, once a day for 3 days, i.p.) partially protected them against the acute toxicity of CdCl2 (0.075 mmol/kg, once, s.c., 24 h after pretreatment with Na2MoO4). The survival number of rats per total number of rats in the CdCl2-dosed group was 10/10, 8/10, 6/10, 2/10 and 0/10 on 0, 1, 2, 6 and 18 days after treatment with CdCl2 whereas in the group where CdCl2 is given after pretreatment with Na2MoO4 it is 10/10 and 6/10 on 0 and 18 days. The body weight of CdCl2-dosed rats consistently decreased until their death while that of Na2MoO4-CdCl2-dosed rats similarly decreased up to 4 days after exposure to CdCl2 but then increased almost normally. In order to elucidate the mechanism of protective action of Na2MoO4 against the acute toxicity of CdCl2, cellular components such as DNA, inorganic cations and metallothionein were measured in the liver after exposure to CdCl2. The treatment with CdCl2 alone reduced K content and increased Ca content but pretreatment with Na2MoO4 prevented such alterations in the levels of those cations caused by CdCl2. Metallothionein content in the liver was significantly elevated in the CdCl2-treated groups as compared to saline controls although the protein content was higher in the Na2MoO4-CdCl2-dosed group than in the CdCl2-dosed group. There was no difference in the protein content of the liver between saline controls and the Na2MoO4-dosed group. This suggests that Na2MoO4 alleviated the acute toxicity of CdCl2 in the rat and the protective mechanism by the metal is in part related to the enhancement of liver Cd-metallothionein induction.

Topics: Animals; Body Weight; Cadmium; Cadmium Chloride; Electrolytes; Liver; Male; Metallothionein; Molybdenum; Rats; Rats, Inbred Strains

A line of transgenic mice expressing insulin-like growth factor-I (IGF-I) under the control of the mouse metallothionien-1 promoter was crossed to a line of dwarf transgenic mice lacking GH expressing cells that were genetically ablated by diphtheria toxin expression. Mice generated from this cross that carry both transgenes express IGF-I in the absence of GH. These mice grew larger than their GH-deficient transgenic littermates and exhibited weight and linear growth indistinguishable from that of their nontransgenic siblings. These results confirm the suspected role of IGF-I in mediating GH's stimulation of somatic growth, including that of long bones, and illustrates the essential role of GH and IGF-I in the modulation of postnatal growth. Analysis of differences in organ growth among these mice, however, suggests that GH and IGF-I also have growth promoting actions that are independent of one another; GH appears to be necessary for the attainment of normal liver size, while IGF-I can stimulate brain growth.

Topics: Aging; Animals; Body Weight; Bone Development; Brain; Diphtheria Toxin; Female; Gene Expression; Genotype; Growth; Growth Hormone; Insulin-Like Growth Factor I; Liver; Male; Metallothionein; Mice; Mice, Transgenic; Organ Size; Promoter Regions, Genetic; RNA, Messenger; Somatomedins

Ethanol, under certain conditions, alters the metabolism of sulfur amino acids, metallothionein (MT) and zinc. If chronic ethanol administration during pregnancy decreases the availability of sulfur amino acids or Zn, this deficiency could contribute to growth retardation of the fetuses, one of the features of fetal alcohol syndrome. The purpose of this study was to discern whether chronic ethanol administration to pregnant rats alters glutathione (GSH), MT or Zn content of selected tissues of the dams and fetuses. Sprague-Dawley rats were fed from gestational days 5 to 19 either the control diet ad libitum (AF), the ethanol diet ad libitum (EF) or the control diet using the pair-feeding technique (PF). On the 19th day of gestation, total hepatic GSH was significantly lower for the EF and PF dams than for the AF dams. Hepatic MT contents were similar for the AF and EF dams, and hepatic MT content was significantly greater for the PF dams than the AF and EF dams. The three groups did not differ regarding hepatic Zn content of dams or fetuses. In summary, on the 19th day of gestation, chronic ethanol feeding of pregnant rats did not lower the maternal hepatic GSH level below that of PF dams, did not induce hepatic MT in the dams and did not prevent fetuses from achieving body weights and hepatic Zn concentrations equal to those of controls.

Topics: Animals; Birth Weight; Body Weight; Ethanol; Female; Fetal Alcohol Spectrum Disorders; Fetal Growth Retardation; Fetus; Glutathione; Litter Size; Liver; Metallothionein; Organ Size; Pregnancy; Pregnancy, Animal; Rats; Rats, Inbred Strains; Zinc

The effects of iron deficiency and of restriction of food intake on blood and tissue metallothionein-I (MT-I) concentrations in rats were investigated. Compared to ad libitum fed controls, MT-I concentrations in the blood cells of the iron-deficient rats were higher, whereas concentrations in pair-fed control rats were lower. Iron deficiency also increased MT-I concentrations in the bone marrow but concentrations in the liver were unchanged and those in the kidneys were reduced. The MT-I in the blood cells was associated mainly with the lighter cell fractions which were rich in reticulocytes. It is suggested that concentrations of MT-I in blood cells reflect erythropoietic activity.

Topics: Anemia, Hypochromic; Animals; Blood Cells; Body Weight; Bone Marrow; Diet; Eating; Iron; Kidney; Liver; Male; Metallothionein; Rats

N-(n-Propyl)-, N-(n-butyl)-, and N-(n-amyl)-N-dithiocarboxy-D-glucamine were newly synthesized by (a) addition of each n-alkylamine to glucose, (b) high-pressure catalytic reduction of each glucosamine thus formed to the corresponding glucamine, and (c) reaction of the resultant secondary amines with CS2 to form the dithiocarboxy derivatives. Each compound was evaluated as an antagonist of acute cadmium (Cd) toxicity and as a complexing agent for intracellular metallothionein-bound Cd (Cd-MT) in mice. N-Benzyl-N-dithiocarboxy-D-glucamine (BDCG) was used as a positive control compound. Each congener afforded partial or complete protection against the lethal effects of 10.0 mg/kg CdCl2.2.5 H2O, and retarded accumulation of Cd in livers and kidneys when given 2 h after the acutely toxic dose of Cd. Each derivative was also effective in mobilizing Cd from MT-bound sites in livers and kidneys of mice which had received a sub-lethal dose of CdCl2 along with 109CdCl2 2 weeks earlier. Excretion of mobilized Cd was almost exclusively by the fecal route. Potency of the analogs, as well as the octanol/aqueous partition coefficients, increased with the overall length of the N-(n-alkyl) carbon chain. Each compound readily complexed Cd from partially purified Cd-MT in vitro. Serum Cd from mice treated with BDCG was associated principally with proteins of high molecular weight.

Topics: Animals; Body Weight; Cadmium; Chemical Phenomena; Chemistry; Chemistry, Physical; Dose-Response Relationship, Drug; Male; Meglumine; Metallothionein; Mice; Mice, Inbred ICR; Organ Size; Solubility; Sorbitol; Thiocarbamates; Tissue Distribution

Hepatic copper concentration in the guinea-pig increased markedly during the second-half of gestation, attaining a maximum shortly after birth; thereafter, concentration declined rapidly during the neonatal period. Changes in perinatal hepatic copper concentrations paralleled the binding of copper to a cytosolic metallothionein-like component, and the loss of hepatic copper in the neonates coincided with increases in serum copper concentrations. Zinc concentrations of the perinatal liver were low and showed no dramatic developmental changes. The humerus showed striking increases in zinc concentration with gestational age, attaining peak concentration before term and a marked depletion of tissue zinc during the neonatal period.

Topics: Animals; Animals, Newborn; Body Weight; Bone and Bones; Cadmium; Copper; Female; Fetus; Gestational Age; Guinea Pigs; Liver; Metallothionein; Organ Size; Pregnancy; Protein Binding; Zinc

A striking radioresistance has been found in mice which were subjected to various pretreatments to induce metallothionein synthesis in the liver prior to irradiation. The tolerance to lethal damage from an LD50 level of radiation during a 30-day postirradiation period was demonstrated by a highly significant difference (P less than 0.01) in mortality rate between mice given subcutaneously manganese, cadmium, or zinc injection or surgical skin excision of mice and the control mice (no pretreatment). A typical loss in body weight that generally reached a peak 2 weeks after irradiation was observed in the control mice, but mice given a dose of 10 mg manganese per kilogram body weight showed a steady weight increase even a few days after irradiation. The normal level of metallothionein in mouse liver is 20 micrograms/g tissue. This level increased up to 70 micrograms/g tissue following irradiation at 6.3 Gy. Among irradiated mice, metallothionein levels in the liver increased approximately 200-800% after cadmium, manganese, or zinc injection compared to levels of irradiated mice without pretreatment. Mice undergoing 2 X 2-cm2 dermal excision also demonstrated a similar reduction of mortality and high metallothionein contents in liver, i.e., 150-400 micrograms/g. The present results, together with our previous findings (Matsubara et al., 1982, 1983, 1984; Matsubara, 1986), suggest that the body's protective mechanism against radiation strongly correlates with the biosynthesis of metallothionein or metallothionein itself acting as a scavenger of radiation-induced peroxides.

Topics: Acetates; Acetic Acid; Animals; Body Weight; Cadmium; Cadmium Chloride; Chlorides; Cold Temperature; Liver; Male; Manganese; Manganese Compounds; Metallothionein; Mice; Radiation Tolerance; Whole-Body Irradiation; Wounds and Injuries

The effects of starvation and refeeding of 2-wk-old turkey poults on serum and tissue levels of zinc, copper and iron were investigated. Serum concentrations of zinc and copper declined during 4 d of starvation. Refeeding for 24 h following a period of starvation restored serum copper to levels similar to those in the fed controls but failed to elevate zinc levels. Liver concentrations and total quantities of zinc, copper and iron increased throughout starvation. Refeeding the starved poults reduced hepatic metal concentrations but caused a further increase in total metal content. This was apparently related to the large increase in liver mass, and the effect was most pronounced in poults starved 1 d prior to refeeding. Starvation also caused an increased zinc concentration and content and a reduced copper content in the pancreas, duodenal mucosa and kidney. Iron content of the pancreas and kidney increased during starvation, but that of the duodenal mucosa declined. Starvation evoked a progressive increase in the cytosolic zinc concentration from liver, pancreas, duodenal mucosa and kidney. A major part of this increase was accounted for as zinc bound to metallothionein (MT). Refeeding rapidly reduced cytosolic and MT-bound zinc in each of these tissues. It was concluded that starvation and refeeding had major effects on tissue trace metal status. A function is proposed for MT during starvation as a mechanism for the conservation of body zinc stores. Zinc, released as a consequence of tissue catabolism, is repartitioned into a soluble storage site (MT), which can be rapidly mobilized to meet the demands of new tissue synthesis once anabolic metabolism resumes.

Topics: Animal Nutritional Physiological Phenomena; Animals; Body Weight; Copper; Cytosol; Female; Food; Iron; Male; Metallothionein; Organ Size; Poultry Diseases; Starvation; Tissue Distribution; Turkeys; Zinc

To investigate the role of metallothionein (MT) in the sequestration and storage of zinc in newborn rat livers, a cross-fostering experiment was performed in which zinc-deficient (Zn-D) pups were suckled from Zn-sufficient (Zn-S) dams and vice versa. At consecutive days during lactation, groups of pups were killed. The experiment was continued for 22 d. Zinc concentrations in various tissues and MT concentrations in livers and kidneys were analyzed. The retention of injected 65Zn, as well as body weight, was also studied in both groups. Higher whole-body retention of 65Zn in the zinc-depleted (Zn-Dp) rats indicates a decreased zinc turnover. These rats also showed markedly reduced growth. The observation that the zinc-repleted (Zn-Rp) pups, although showing 65Zn retention similar to that of controls, grew less than controls suggests that Zn-Rp may not compensate for gestational zinc deficiency. Zn-Rp pups showed an increased accumulation of zinc into hepatic MT until d 10, whereas Zn-Dp pups showed a more accelerated degradation of MT than controls. These data indicate that hepatic MT levels fluctuate directly in response to dietary zinc status in newborn rats. Various tissues such as spleen, heart, lung and intestine showed no difference in zinc concentration among all groups at d 22 postpartum. Thus the rapid degradation of hepatic MT in zinc deficiency that may occur to maintain the required zinc levels in other tissues supports the role of MT as a zinc storage protein in newborn rats.

Topics: Animals; Animals, Suckling; Body Weight; Copper; Cytosol; Diet; Female; Homeostasis; Kidney; Liver; Metallothionein; Rats; Rats, Inbred Strains; Zinc

Recent reports indicate that the concentrations and total amounts of several essential trace metals in various tissues of genetically obese rodents differ markedly from those in lean controls. In the present studies the absorption, retention and tissue distribution of zinc and constitutive levels of zinc-metallothionein (Zn-MT) in selected tissues were compared in obese (ob/ob) and lean (+/?) C57BL/6J mice. When 5-, 10- and 22-wk-old mice were administered 1.2 mumol 65Zn by stomach tube the apparent absorption of 65Zn by obese mice was 1.5, 2.2 and 3.9 times higher, respectively, than that in age-matched lean mice. Retention of orally administered 65Zn after 96 h was also substantially higher in obese mice than in lean mice. To assess the possible influences of hyperphagia and intestinal hypertrophy on the enhanced apparent absorption of 65Zn by obese mice food intake by an additional group of obese mice was restricted to that of age-matched lean controls. When actual absorption of zinc was determined according to the method of Heth and Hoekstra, groups of ad libitum--fed obese, pair-fed obese and lean mice absorbed 38, 32 and 18% of administered 65Zn, respectively. In contrast, the rate of 65Zn excretion 2-6 d after oral or subcutaneous administration of the metal was similar for obese and lean mice. Unrestricted and pair-fed obese mice had significantly lower percentages of carcass 65Zn present in skin, muscle plus bone, spleen and testes and higher percentages present in liver, small intestine and adipose tissue than lean mice.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Body Weight; Intestinal Absorption; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Obese; Species Specificity; Tissue Distribution; Zinc; Zinc Radioisotopes

A striking resistance to lethal damage from a single dose of 6-8 Gy of X rays has been found in mice which had received various pretreatments to induce metallothionein (MT) synthesis in the liver prior to irradiation. Mice were injected with manganese (10 mg Mn/kg) or cadmium (3 mg Cd/kg) salt subcutaneously, or a patch of dorsal skin (2 X 2 cm2) was excised 1 or 2 days prior to irradiation. The increased tolerance of these mice to radiation was established by a marked decrease of mortality rate, an increase of mean survival time, a reduction of weight loss, and a smaller decrease in the number of leukocytes as compared with the control group. The LD50/30 for control mice was 6.3 Gy, while the corresponding values for the groups pretreated with Mn, Cd, and skin excision were 7.5, 7.7, and 7.9 Gy, respectively. The normal level of MT in mouse liver was approximately 25 micrograms/g tissue. This level increased 2.5- to 3-fold 24 h after 6.3 Gy irradiation. The MT levels of mice pretreated with Cd, Mn, and skin excision were increased 8-, 5-, and 7-fold, respectively, prior to irradiation as compared with the preirradiation control. These results indicate that the induction of MT in mouse liver is a significant factor in the mechanism of protection against radiation.

Topics: Animals; Body Weight; Cadmium; Cadmium Chloride; Chlorides; Dermatologic Surgical Procedures; Lethal Dose 50; Liver; Male; Manganese; Manganese Compounds; Metallothionein; Mice; Mice, Inbred ICR; Radiation Tolerance; Stress, Physiological

Cadmium (Cd) was injected intramuscularly into three species of frogs to examine the number of isoforms in the induced metallothionein (MT). The induced MT was shown to consist of a single isoform in the three species of frogs Rana japonica, R. nigromaculata and Rhacophorus schlegelii. Native MT in the livers of R. japonica and R. nigromaculata was copper (Cu)-MT and the induced MT in the three species was Cd, Cu-MT, the Cd/Cu ratio in the induced MT being different from species to species. The effect of Cd injections on the concentrations of 10 elements in the livers and kidneys of the Rana species was not significant except for Cu.

Topics: Amphibians; Animals; Anura; Body Weight; Cadmium; Cadmium Chloride; Chromatography, High Pressure Liquid; Copper; Kidney; Liver; Metallothionein; Organ Size; Ranidae; Zinc

In a thirty-day inhalation study male Wistar rats were continuously exposed to submicron aerosols of three different cadmium compounds. The cadmium chloride (CdCl2) and cadmium oxide (CdO) aerosol concentrations were 0.1 mg/m3 Cd. Because of its lower solubility the cadmium sulfide (CdS) level was 1 mg/m3 Cd. For CdCl2 and CdO, most of the cadmium was found in the lung cytosolic compartment, but for CdS only 30% of the cadmium was retrieved from the lung cytosols. This was observed both at the end of the inhalation and also after an additional 2-month period in fresh air. The cadmium contents of the lung homogenates, cytosols, and the lung cytosolic metallothionein were found to be twice as much for exposure to CdO than for exposure to CdCl2. For exposure to CdS at cadmium concentrations 10 times higher the same cadmium levels were found as for CdO. These results are confirmed by results from alveolar lavage analysis indicating that in the lung-inhaled CdO is even more available to lung tissue than the very soluble CdCl2, and CdO has an availability 10 times as much as CdS. This study proved that lung compartmental cadmium and metallothionein contents as well as lung lavage analysis can describe the bioavailability of inhaled cadmium.

Topics: Aerosols; Animals; Biological Availability; Body Burden; Body Weight; Bronchi; Cadmium; Metallothionein; Particle Size; Pulmonary Alveoli; Rats; Rats, Inbred Strains; Therapeutic Irrigation; Time Factors

The liver of 7-day-old rats contains high levels of metallothionein (MT) and zinc, which can be reduced markedly following treatment of neonates with dexamethasone (1 mg/kg, s.c.) twice daily on days 3, 4, 5 and 6 postpartum. Prior treatment with trace amounts of 65Zn and 109Cd did neither alter the basal MT and zinc concentrations nor their response to dexamethasone. The turnover of 65Zn or 109Cd in newborn rats was studied by measuring the whole body retention of the radioisotopes. Between days 3 and 7 postpartum, the normal turnover of 65Zn was much faster than that of 109Cd, and dexamethasone treatment resulted in further decrease of 65Zn retention without significantly altering that of 109Cd. The data indicate that dexamethasone enhanced the excretion of 65Zn without any effect on 109Cd excretion. The dexamethasone treatment resulted in marked reduction in both hepatic Zn and MT levels in newborn rats. Gel filtration on Sephadex G-75 columns of hepatic cytosols isolated from 7-day-old rats showed significant amounts of total Zn and 65Zn bound to MT fraction. Moreover, dexamethasone treatment markedly reduced their binding to the MT fraction suggesting that most of the Zn lost from the liver of dexamethasone-treated animals was from MT. About 95% of 109Cd in the liver cytosol was present in the MT fraction of 109Cd-injected rats and dexamethasone had little effect on the binding of 109Cd to MT fraction despite a marked reduction in the binding of total Zn. These data suggest that unlike in adult rats, injection of dexamethasone results in marked decrease in hepatic levels of Zn and MT in neonatal rats.

Topics: Animals; Animals, Newborn; Body Burden; Body Weight; Cadmium; Dexamethasone; Female; Liver; Metallothionein; Organ Size; Rats; Rats, Inbred Strains; Zinc

The effects of maternal Zn, Cu, or Fe deficiencies during late gestation on hepatic levels of metals and metallothionein (MT) and the binding of Zn and Cu to protein fractions were investigated in newborn rats. Timed pregnant rats were fed one of the following diets: Zn deficient (Zn-D), Cu-D, Fe-D, or control from day 12 of gestation until birth. The specific nutritional deficiency status of the dams was confirmed by low plasma levels of the deficient metal. Livers from pups were analyzed for MT, metal content, and metal-protein binding. Maternal Zn-D resulted in a greater than 50% reduction of hepatic MT levels in pups, whereas Cu-D and Fe-D had no significant effects. Pups in each deficient group showed a significant decrease in the hepatic levels of the respective metals. Fractionation of hepatic cytosols from the pups by Sephadex G-75 gel filtration showed that in both Fe-D and Cu-D pups the respective metals were depleted from the high molecular weight protein fraction, whereas in Zn-D pups the Zn was depleted mainly from the MT fraction (Ve/V0 approximately 2). Incorporation of [35S] cysteine into MT fractions was significantly lower in Zn-D pups as compared with control pups. These results indicate that there is a specific effect of the maternal Zn-D on the hepatic storage of Zn as MT in newborn rats.

Topics: Animals; Animals, Newborn; Body Weight; Chromatography, Gel; Copper; Cytosol; Female; Iron; Iron Deficiencies; Liver; Metallothionein; Nutrition Disorders; Pregnancy; Rats; Rats, Inbred Strains; Zinc

Zinc pretreatment is known to ameliorate the acute and chronic effects of the toxic heavy metal, cadmium. However, the ability of zinc to decrease the toxicity of other metals has not been widely investigated. Therefore, this study was designed to determine the effects of zinc pretreatment on the acute toxicity of nickel. Male Fischer rats received either nickel alone (i.p.), zinc alone (s.c.), zinc plus nickel, or saline (i.p. and s.c.; controls). In the lethality studies, the dose of nickel was 115 mumol nickel/kg (as nickel acetate) while for all other studies the dose was 95 mumol nickel/kg. Zinc was given in multiple doses of 300 mumol zinc/kg (as zinc acetate) at -24, 0 and +24 h relative to nickel (total zinc dose 900 mumol/kg) for lethality studies or -24 and 0 h for studies 24 h and under in duration (total dose 600 mumol/kg). Zinc pretreatment significantly increased the 14-day survival of nickel-related rats. Zinc did not, however, prevent the reduction in weight gain over 2 weeks seen with nickel treatment. Histopathologically, at 120 h following nickel exposure, kidneys in the group receiving nickel alone generally showed moderate nephropathy (multifocal proximal tubule degeneration with necrosis) while in the zinc plus nickel group the nephropathy was generally mild. Zinc pretreatment had no apparent effect on the pharmacokinetics of nickel over 24 h as assessed by urinary excretion, blood levels or organ distribution. Zinc pretreatment also did not alter the subcellular distribution of renal nickel 6 h after nickel exposure. Enhanced synthesis of metallothionein did not appear to play a critical role in the reduction of nickel toxicity, since renal concentrations of this metal-binding protein, although elevated compared to control, were not different in rats receiving zinc and nickel or zinc alone. Zinc pretreatment did, however, have marked effect on nickel-induced hyperglycemia, reducing both the duration and severity of elevated blood glucose levels. Results of this study show that zinc can prevent some of the toxic effects of nickel and that the mechanism of this action does not appear to involve either metallothionein or alterations in the pharmacokinetics of nickel.

Topics: Acetates; Acetic Acid; Animals; Blood Glucose; Body Weight; Kidney; Male; Metallothionein; Rats; Rats, Inbred F344; Subcellular Fractions; Tissue Distribution

Rats were injected sc with 0.5 mg Cd/kg, 6 days/week, for up to 26 weeks. Hepatic and renal function and tissue Cd and metallothionein (MT) content were determined in tissues and plasma at various times after Cd injection. Cd in liver and kidney increased linearly for the first 10 weeks of treatment, but thereafter hepatic concentrations of Cd decreased by 33% whereas the content of Cd in kidney remained constant. MT in liver and kidney increased linearly during the first 12 weeks of Cd treatment to 4400 and 2300 micrograms MT/g, respectively, but rose only slightly thereafter. Circulating concentrations of MT progressively increased beginning 2 weeks after Cd treatment and were approximately 10 times control values in rats dosed with Cd for 12 or more weeks. Plasma activities of alanine and aspartate aminotransferase exhibited a time course similar to that observed with MT, and were elevated as early as the sixth week of Cd exposure. Sharp increases in activities of these enzymes also occurred after 10 to 12 weeks of dosing. Hepatic microsomal metabolism of benzo[a]pyrene and ethylmorphine was severely attenuated beginning 4 weeks after Cd. Renal injury occurred after hepatic damage, as evidenced by decreased in vitro p-aminohippuric acid uptake beginning 8 weeks after exposure. Urine outflow increased threefold 11 weeks after Cd exposure began, while urinary protein and Cd excretion increased beginning at Week 9. These data indicate the liver is a major target organ of chronic Cd poisoning, and suggest that Cd-induced hepatic injury, via release of Cd-MT, may play an important role in the nephrotoxicity observed in response to long-term exposure to Cd.

Topics: Animals; Body Weight; Cadmium; Cadmium Poisoning; Chemical and Drug Induced Liver Injury; Kidney; Kidney Diseases; Liver; Liver Diseases; Male; Metallothionein; Organ Size; Rats; Rats, Inbred Strains; Tissue Distribution

Male ICR mice were either given water containing Cd at a level of 192-200 ppm for 45 days (ingestion group), or were injected subcutaneously once a week with Cd (1 mg/kg) as CdCl2 for 7 weeks (injection group). The control group was given Cd-free water. In both Cd groups, the hematocrit and hemoglobin values did not change markedly. In the ingestion group, the Fe concentration decreased greatly in the liver, kidney, spleen, and duodenum. These decreases may be due to depression of Fe absorption from the intestine. In the injection group, Fe increased in the liver, spleen, and duodenum, although it decreased in the kidney. By Sephadex G-200 gel filtration, Fe-proteins in the hepatic supernatants were located in the void volume region of this gel column in both Cd groups. Apparently, Fe was not a component of metallothionein (MT) protein. The hepatic MT induction by Cd resulted in an increase in hepatic supernatant Cu. Serum Cu and ceruloplasmin (Cp) activity were stimulated only in the injection group. The enhancement of Cp activity may possibly be due to the increase in hepatic Cu which was accompanied by an increase in hepatic Fe, rather than a decrease. Our observations suggest that Fe metabolism is influenced differentially by the administration route of Cd.

Topics: Administration, Oral; Animals; Body Weight; Cadmium; Ceruloplasmin; Duodenum; Hematocrit; Hemoglobins; Injections, Subcutaneous; Iron; Liver; Male; Metallothionein; Metals; Mice; Mice, Inbred ICR; Organ Size; Protein Binding; Spleen; Transferrin

Twenty-one male rabbits were divided into three groups: rabbits of two groups were given pelleted food containing cadmium chloride at a dose level of 300 micrograms Cd/g over periods of 44 or 19 weeks. Rabbits of the last group were given ordinary commercial pelleted food and served as controls. Cadmium increased urinary protein and amino acid by week 19 and increased it to a remarkably high level by week 44. After cessation of cadmium exposure, rabbits of the first group (44 weeks exposure group) showed only little recovery from cadmium health effects: proteinuria and aminoaciduria were slightly improved. Depressed hepatic functions were also slightly improved, but did not return to the control level in 24 weeks. Fat and bone metabolism also remained depressed below the control level. Anemia did not also readily recover. On the other hand, rabbits of the second group (19 weeks exposure) recovered from the effects of cadmium: proteinuria and aminoaciduria in most animals disappeared soon after the end of cadmium exposure, plasma GPT fell after 1 week, and hemoglobin and hematocrit returned to normal in 6-11 weeks. The above results show that after cessation of cadmium exposure, mild cadmium-induced health effects were reversible in a short period, while more severe effects were not readily reversible. High performance liquid chromatographic (HPLC) profiles of renal and hepatic cadmium-thionein (Cd-MT) during and after exposure to cadmium showed no correlation to the degree of cadmium health effects, and therefore, did not help to elucidate mechanisms of the recovery from cadmium-induced health effects, probably because cadmium not bound with metallothionein (non-MT-Cd) is responsible for inducing renal effects.

Topics: Amino Acids; Anemia; Animals; Blood; Body Weight; Cadmium; Chromatography, High Pressure Liquid; Copper; Glycosuria; Kidney; Liver; Male; Metallothionein; Molecular Weight; Proteinuria; Rabbits

The relationship of dietary protein to cadmium absorption and tissue deposition was studied in male Sprague-Dawley rats exposed to different levels of cadmium in the drinking water. In animals fed a high-protein or low-protein diet and drinking water containing 25 or 50 ppm cadmium, liver and kidney cadmium and metallothionein were both significantly higher in rats fed the high-protein diet for 2 to 4 months. These differences may possibly be explained by the concentration of cysteine observed between these two diets. When cysteine was added to the low-protein diet to the level observed in the high-protein diet and fed to rats receiving 25 ppm cadmium in the drinking water, significant dietary differences in liver and kidney cadmium and metallothionein were not observed. The importance of dietary protein to cadmium-induced toxicity was also assessed in these studies. The activity of catechol-o-methyltransferase was used as a measure of cadmium-induced toxicity. The activity of this enzyme in the lung, liver and heart was significantly lower in rats fed a low-protein diet than those fed the high-protein diet and 50 ppm cadmium. Metallothionein concentration in the lung and liver from low-protein-fed rats was approximately half the level observed in rats fed the high-protein diet, which suggests a relationship between cadmium-induced toxicity and metallothionein concentrations. These results illustrate the importance of considering dietary protein (and possibly cysteine) when studying cadmium metabolism in experimental animals.

Topics: Animals; Body Weight; Cadmium; Catechol O-Methyltransferase; Cysteine; Dietary Proteins; Kidney; Liver; Male; Metallothionein; Rats; Rats, Inbred Strains

The concentration of metallothionein (MT) in the liver of the perinatal rat is relatively high at term and 7 days after birth and then decreases to barely detectable levels by day 28. The developmental pattern MT-zinc parallels that of MT. When challenged with a single injection of cadmium chloride, the 26-day-old rat responds with a dose-related increase in hepatic MT which sequesters both cadmium and zinc. When the 5-day-old rat is similarly challenged, induction of MT occurs only at the highest dose tested (6 mg Cd/kg); however, due to the pre-existence of MT in these younger rats, cadmium administered at the lower doses still binds to the MT in a dose-related manner. Despite the induction of MT seen in both age groups following the 6.0 mg/kg dose, exposure to that level of the metal produced death in 30% of the younger animals but in only 4% of the older animals. When cadmium was administered to pregnant rats on day 19 of gestation, it was found to produce a dose-related induction of maternal hepatic MT over the following 48 hr. In contrast, maternal exposure to the metal led to a significant depression of fetal hepatic MT over the same time interval.

Topics: Animals; Body Weight; Cadmium Poisoning; Chromatography, Gel; Female; Liver; Male; Maternal-Fetal Exchange; Metalloproteins; Metallothionein; Organ Size; Pregnancy; Rats; Rats, Inbred Strains; Zinc

Topics: Absorption; Aging; Animals; Animals, Newborn; Body Weight; Carrier Proteins; Copper; Digestive System; Female; Intestinal Absorption; Kidney; Liver; Male; Mercury; Metallothionein; Rats; Tissue Distribution

The levels of cadmium and zinc metallothionein in isolated perfused livers of zinc depleted and repleted rats were investigated. Rats (160-200 g or 90-120 g) were fed either a zinc-deficient or zinc-supplemented diet for 62 or 42 days. The 90 to 120 g rats were repleted with 30 ppm Zn2+ in the drinking water for 1, 4 and 10 days after 35 days of depletion, and another group after depletion for 21 days was repleted for 15 days on a 21% casein diet. At appropriate times livers were cannulated, removed and perfused +/- 25 mug Cd2+ as CdCl2, for 2 hours through the portal vein in a perfusion apparatus. Zinc depletion resulted in significant decreases in growth rate (1.4 versus 5.7 g/day) and liver weights. Repletion with Zn for 10 days increased the weight gain to 7.1 g/day. Depletion decreased the incorporation of Zn and Cd into metallothionein by 57 to 60%, while repletion for 1, 4 and 10 days increased the incorporation to nearly 500% of control levels. But, as repletion progressed, zinc in metallothionein fell to normal levels after 15 days. Cadmium in metallothionein did not decrease between 1 and 10 days of repletion. Zinc deficiency also reduced the zinc content of the liver by 44%, which increased to normal levels after 10 days of repletion. These data suport the conclusion that zinc status can influence the incorporation of cadmium and zinc into metallothionein.

Topics: Animals; Body Weight; Cadmium; Liver; Male; Metalloproteins; Metallothionein; Organ Size; Perfusion; Rats; Zinc