metallothionein and Necrosis

Topics: 6-Aminonicotinamide; Animals; Brain; Brain Diseases; Brain Injuries; Central Nervous System; Central Nervous System Diseases; DNA Fragmentation; Humans; Immunohistochemistry; In Situ Nick-End Labeling; Inflammation Mediators; Lymphocyte Activation; Macrophages; Metallothionein; Necrosis; Neovascularization, Pathologic; Neurofibrillary Tangles; Oxidative Stress; Tumor Necrosis Factor-alpha

Reactive oxygen (ROS) and reactive nitrogen species (RNS) produced in vivo at levels that cannot be dealt with adequately by endogenous antioxidant systems can lead to the damage of lipids, proteins, carbohydrates and nucleic acids. Oxidative modification of these molecules by toxic levels of ROS and RNS represents an extreme event that can lead to deleterious consequences such as loss of function. More recently, however, interest has focused on the formation of these species at sub-toxic levels and their potential to act as biological signal molecules. Subtoxic ROS and RNS production can lead to alterations in cellular and extracellular redox state, and it is such alterations that have been shown to signal changes in cell functions. By the use of a variety of cell types it has been shown that numerous cellular processes including gene expression can be regulated by subtle changes in redox balance Examples of this include the activation of certain nuclear transcription factors, and the determination of cellular fate by apoptosis or necrosis. Cellular redox balance is, under normal circumstances, probably under genetic control and maintained by an array of enzymatic systems that ensure that overall reducing conditions prevail. Thiols, by virtue of their ability to be reversibly oxidised, are recognised as key components involved in the maintenance of redox balance. Additionally, increasing evidence suggests that thiol groups located on various molecules act as redox sensitive switches thereby providing a common trigger for a variety of ROS and RNS mediated signalling events. In this review we discuss a number of cellular processes in which ROS and RNS have been implicated in redox signalling mechanisms. Particular attention has been paid to the importance of thiols and thiol-containing molecules in these processes.

Topics: Albumins; Animals; Apoptosis; Cysteine; Disulfides; Glutathione; Glutathione Peroxidase; Humans; Metallothionein; Necrosis; Oxidation-Reduction; Oxidative Stress; Signal Transduction; Sulfhydryl Compounds; Transcription Factors

Lysosomal membrane permeabilization (LMP) induced by oxidative stress has recently emerged as a prominent mechanism behind TNF cytotoxicity. This pathway relies on diffusion of hydrogen peroxide into lysosomes containing redox-active iron, accumulated by breakdown of iron-containing proteins and subcellular organelles. Upon oxidative lysosomal damage, LMP allows relocation to the cytoplasm of low mass iron and acidic hydrolases that contribute to DNA and mitochondrial damage, resulting in death by apoptosis or necrosis. Here we investigate the role of lysosomes and free iron in death of HTC cells, a rat hepatoma line, exposed to TNF following metallothionein (MT) upregulation. Iron-binding MT does not normally occur in HTC cells in significant amounts. Intracellular iron chelation attenuates TNF and cycloheximide (CHX)-induced LMP and cell death, demonstrating the critical role of this transition metal in mediating cytokine lethality. MT upregulation, combined with starvation-activated MT autophagy almost completely suppresses TNF and CHX toxicity, while impairment of both autophagy and MT upregulation by silencing of Atg7, and Mt1a and/or Mt2a, respectively, abrogates protection. Interestingly, MT upregulation by itself has little effect, while stimulated autophagy alone depresses cytokine toxicity to some degree. These results provide evidence that intralysosomal iron-catalyzed redox reactions play a key role in TNF and CHX-induced LMP and toxicity. The finding that chelation of intralysosomal iron achieved by autophagic delivery of MT, and to some degree probably of other iron-binding proteins as well, into the lysosomal compartment is highly protective provides a putative mechanism to explain autophagy-related suppression of death by TNF and CHX.

Topics: Animals; Apoptosis; Autophagy; Carcinoma, Hepatocellular; Cell Line, Tumor; Iron; Liver Neoplasms; Lysosomes; Metallothionein; Necrosis; Oxidative Stress; Rats; Tumor Necrosis Factor-alpha

Inflammatory bowel diseases (IBDs) are recurrent intestinal pathologies characterized by a compromised epithelial barrier and an exaggerated immune activation. Mediators of immune cell infiltration may represent new therapeutic opportunities. Metallothioneins (MTs) are stress-responsive proteins with immune-modulating functions. Metallothioneins have been linked to IBDs, but their role in intestinal inflammation is inconclusive. We investigated MT expression in colonic biopsies from IBDs and acute infectious colitis patients and healthy controls and evaluated MT's role in experimental colitis using MT knockout mice and anti-MT antibodies. Antibody potential to target extracellular MT and its mechanism was tested in vitro. Biopsies of patients with active colitis showed infiltration of MT-positive cells in a pattern that correlated with the grade of inflammation. MT knockout mice displayed less severe acute dextran sulphate sodium (DSS)-induced colitis compared to congenic wild-type mice based on survival, weight loss, colon length, histological inflammation and leukocyte infiltration. Chronic DSS-colitis confirmed that Mt1 and Mt2 gene disruption enhances clinical outcome. Blockade of extracellular MT with antibodies reduced F4/80-positive macrophage infiltration in DSS- and trinitrobenzene sulphonic acid-colitis, with a tendency towards a better outcome. Whole-body single-photon emission computer tomography of mice injected with radioactive anti-MT antibodies showed antibody accumulation in the colon during colitis and clearance during recovery. Necrotic and not apoptotic cell death resulted in western blot MT detection in HT29 cell supernatant. In a Boyden chamber migration assay, leukocyte attraction towards the necrotic cell supernatant could be abolished with anti-MT antibody, indicating the chemotactic potential of endogenous released MT. Our results show that human colitis is associated with infiltration of MT-positive inflammatory cells. Since antibody blockade of extracellular MT can reduce colitis in mice, MT may act as a danger signal and may represent a novel target for reducing leukocyte infiltration and inflammation in IBD patients.

Topics: Acute Disease; Adolescent; Adult; Aged; Aged, 80 and over; Animals; Antibodies; Apoptosis; Biopsy; Case-Control Studies; Chemotaxis, Leukocyte; Chronic Disease; Colitis; Colon; Dextran Sulfate; Disease Models, Animal; Female; HT29 Cells; Humans; Macrophages; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Middle Aged; Necrosis; Severity of Illness Index; Signal Transduction; Time Factors; Trinitrobenzenesulfonic Acid; Young Adult

Zinc is essential for the activities of pancreatic β-cells, especially insulin storage and secretion. Insulin secretion leads to co-release of zinc which contributes to the paracrine communication in the pancreatic islets. Zinc-transporting proteins (zinc-regulated transporter, iron-regulated transporter-like proteins [ZIPs] and zinc transporters [ZnTs]) and metal-buffering proteins (metallothioneins, MTs) tightly regulate intracellular zinc homeostasis. The present study investigated how modulation of cellular zinc availability affects β-cell function using INS-1E cells.. Using INS-1E cells, we found that zinc supplementation and zinc chelation had significant effects on insulin content and insulin secretion. Supplemental zinc within the physiological concentration range induced insulin secretion. Insulin content was reduced by zinc chelation with N,N,N',N-tektrakis(2-pyridylmethyl)-ethylenediamine. The changes in intracellular insulin content following exposure to various concentrations of zinc were reflected by changes in the expression patterns of MT-1A, ZnT-8, ZnT-5, and ZnT-3. Furthermore, high zinc concentrations induced cell necrosis while zinc chelation induced apoptosis. Finally, cell proliferation was sensitive to changes in zinc the concentration.. These results indicate that the β-cell-like function and survival of INS-1E cells are dependent on the surrounding zinc concentrations. Our results suggest that regulation of zinc homeostasis could represent a pharmacological target.

Topics: Animals; Apoptosis; Cation Transport Proteins; Cell Division; Cell Line; Chelating Agents; DNA, Complementary; Dose-Response Relationship, Drug; Ethylenediamines; Gene Expression Profiling; Gene Expression Regulation; In Vitro Techniques; Insulin; Insulin Secretion; Islets of Langerhans; Metallothionein; Necrosis; Rats; Real-Time Polymerase Chain Reaction; RNA, Messenger; Transcription, Genetic; Zinc; Zinc Transporter 8

We studied here the short-term toxicity effects of Cd on the oxidative state and cell death in the gill of freshwater crab Sinopotamon henanense. Crabs were exposed to Cd that resulted in Cd accumulation and a significant increase in the metallothionein (MT) level in the gill, but MT level increased disproportionally compared to the Cd accumulation with an extension of exposure time. Significant changes in the activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx) were observed. An increase in the levels of reactive oxygen species (ROS) and lipid peroxidation (LPO) was detected that will cause oxidative stress. Histological abnormalities of the gills were discovered, including the expansion of gill cavity, a decrease in the numbers of connection of the upper and the lower of the gill lamellae and epithelial cells, and an increase in the number of hemocytes. The results of a TUNEL test and transmission electron microscope (TEM) showed that more gill cells had apoptotic characteristics after 48 h of Cd treatment compared to the control, but epithelial cell necrosis and inflammatory response appeared only after 72 h. It was concluded that (1) Cd induced the ROS production and accumulation through inhibiting antioxidant enzyme activities and exceeding the saturation values of MT binging; (2) Cd led to lipid peroxidation and histopathological alternations; and (3) Cd induced apoptotic response at short time exposure, followed by necrotic features and inflammatory reaction after longer time exposure.

Topics: Animals; Antioxidants; Apoptosis; Cadmium; Decapoda; Gills; Metallothionein; Necrosis; Oxidative Stress; Time Factors; Toxicity Tests, Acute; Water

The widespread use of tributyltin (TBT) as biocides in antifouling paints and agricultural chemicals has led to environmental and marine pollution. Human exposure occurs mainly through TBT contaminated seafood and drinking water. It is a well known endocrine disruptor in mammals, but its molecular mechanism in testicular damage is largely unexplored. This study was therefore, designed to ascertain effects of tributyltin chloride (TBTC) on sertoli-germ cell co-culture in ex-vivo and in the testicular tissue in-vivo conditions. An initial Ca(2+) rise followed by ROS generation and glutathione depletion resulted in oxidative damage and cell death. We observed p38 and JNK phosphorylation, stress proteins (Nrf2, MT and GST) induction and mitochondrial depolarization leading to caspase-3 activation. Prevention of TBTC reduced cell survival and cell death by Ca(2+) inhibitors and free radical scavengers specify definitive role of Ca(2+) and ROS. Sertoli cells were found to be more severely affected which in turn can hamper germ cells functionality. TBTC exposure in-vivo resulted in increased tin content in the testis with enhanced Evans blue leakage into the testicular tissue indicating blood-testis barrier disruption. Tesmin levels were significantly diminished and histopathological studies revealed marked tissue damage. Our data collectively indicates the toxic manifestations of TBTC on the male reproductive system and the mechanisms involved.

Topics: Animals; Apoptosis; Blood-Testis Barrier; Calcium; Calcium Channel Blockers; Cell Death; Coculture Techniques; Coloring Agents; DNA Fragmentation; Enzyme Activation; Free Radical Scavengers; Germ Cells; Glutathione; Male; MAP Kinase Kinase 4; Membrane Potential, Mitochondrial; Metallothionein; Necrosis; Oxidation-Reduction; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Rats; Reactive Oxygen Species; Sertoli Cells; Testicular Diseases; Testis; Trialkyltin Compounds

Manufactured metal (oxide) nanoparticles are entering the aquatic environment with little understanding on their potential health impacts for exposed organisms. Adopting an integrative approach, we investigated effects of particle size and coating on biological responses for two of the most commonly used metal (oxide) nanoscale particles, silver (Ag) and titanium dioxide (TiO₂) in zebrafish embryos. Titanium dioxide nanoparticles (nominally, 4 nm, 10 nm, 30 nm and 134 nm) had little or no toxicity on the endpoints measured. Ag both in nano form (10 nm and 35 nm) and its larger counterpart (600-1600 nm) induced dose-dependent lethality and morphological defects, occurring predominantly during gastrula stage. Of the silver material tested 10 nm nanoparticles appeared to be the most toxic. Coating Ag nanoparticles with citrate or fulvic acid decreased toxicity significantly. In situ hybridisation analysis identified the yolk syncytial layer (YSL) as a target tissue for Ag-nano toxicity where there was a significant induction of the heavy metal stress response gene, metallothionein 2 (Mt2) at sub-lethal exposures. Coherent Anti-stroke Raman Scattering (CARS) microscopy provided no evidence for silver particles crossing the chorionic membrane in exposed embryos. Collectively, our data suggest that silver ions play a major role in the toxicity of Ag nanoparticles.

Topics: Analysis of Variance; Animals; Embryo, Nonmammalian; Metal Nanoparticles; Metallothionein; Necrosis; Particle Size; Silver; Survival Analysis; Titanium; Zebrafish

Inorganic arsenic [As(V)+As(III)] and its metabolites, especially the trivalent forms [monomethylarsonous acid, MMA(III), and dimethylarsinous acid, DMA(III)], are considered the forms of arsenic with the highest degree of toxicity, linked to certain types of cancer and other pathologies. The gastrointestinal mucosa is exposed to these forms of arsenic, but it is not known what toxic effect these species may have on it. The aim of the present work was to evaluate the toxicity and some mechanisms of action of inorganic arsenic and its metabolites [monomethylarsonic acid, MMA(V), dimethylarsinic acid, DMA(V), MMA(III) and DMA(III)] in intestinal epithelial cells, using the Caco-2 human cell line as a model. The results show that the pentavalent forms do not produce toxic effects on the intestinal monolayer, but the trivalent species have a different degree of toxicity. As(III) induces death mainly by necrosis, whereas only apoptotic cells are detected after exposure to MMA(III), and for DMA(III) the percentages of apoptosis and necrosis are similar. The three forms produce reactive oxygen species, accompanied by a reduction in intracellular GSH and lipid peroxidation, the latter being especially notable in the dimethylated form. They also alter the enzyme activity of glutathione peroxidase and catalase and induce expression of stress proteins and metallothioneins. The results indicate that the trivalent forms of arsenic can affect cell viability of intestinal cells by mechanisms related to the induction of oxidative stress. Further studies are needed to evaluate how the effects observed in this study affect the structure and functionality of the intestinal epithelium.

Topics: Apoptosis; Arsenicals; Caco-2 Cells; Cacodylic Acid; Carcinogens, Environmental; Cell Survival; Gene Expression; Glutathione; Heat-Shock Proteins; Herbicides; Humans; Intestinal Mucosa; Lipid Peroxidation; Metallothionein; Mitochondria; Necrosis; Oxidative Stress; Oxidoreductases; Reactive Oxygen Species

Acute hyperadrenergic stressor states are accompanied by cation dyshomeostasis, together with the release of cardiac troponins predictive of necrosis. The signal-transducer-effector pathway accounting for this pathophysiological scenario remains unclear. We hypothesized that a dyshomeostasis of extra- and intracellular Ca2+ and Zn2+ occurs in rats in response to isoproterenol (Isop) including excessive intracellular Ca2+ accumulation (EICA) and mitochondrial [Ca2+]m-induced oxidative stress. Contemporaneously, the selective translocation of Ca2+ and Zn2+ to tissues contributes to their fallen plasma levels. Rats received a single subcutaneous injection of Isop (1 mg/kg body wt). Other groups of rats received pretreatment for 10 days with either carvedilol (C), a β-adrenergic receptor antagonist with mitochondrial Ca2+ uniporter-inhibiting properties, or quercetin (Q), a flavonoid with mitochondrial-targeted antioxidant properties, before Isop. We monitored temporal responses in the following: [Ca2+] and [Zn2+] in plasma, left ventricular (LV) apex, equator and base, skeletal muscle, liver, spleen, and peripheral blood mononuclear cells (PBMC), indices of oxidative stress and antioxidant defenses, mitochondrial permeability transition pore (mPTP) opening, and myocardial fibrosis. We found ionized hypocalcemia and hypozincemia attributable to their tissue translocation and also a heterogeneous distribution of these cations among tissues with a preferential Ca2+ accumulation in the LV apex, muscle, and PBMC, whereas Zn2+ declined except in liver, where it increased corresponding with upregulation of metallothionein, a Zn2+-binding protein. EICA was associated with a simultaneous increase in tissue 8-isoprostane and increased [Ca2+]m accompanied by a rise in H2O2 generation, mPTP opening, and scarring, each of which were prevented by either C or Q. Thus excessive [Ca2+]m, coupled with the induction of oxidative stress and increased mPTP opening, suggests that this signal-transducer-effector pathway is responsible for Isop-induced cardiomyocyte necrosis at the LV apex.

Topics: Adrenergic beta-Agonists; Adrenergic beta-Antagonists; Animals; Antioxidants; Calcium; Capillary Permeability; Carbazoles; Carvedilol; Endothelium; Homeostasis; In Vitro Techniques; Isoproterenol; Male; Metallothionein; Mitochondria, Heart; Myocardium; Myocytes, Cardiac; Necrosis; Oxidative Stress; Propanolamines; Quercetin; Rats; Rats, Sprague-Dawley; Sarcolemma; Stress, Physiological; Superoxide Dismutase; Zinc

Consumption of beverages that contain fructose favors the increasing prevalence of metabolic syndrome alterations in humans, including non-alcoholic fatty liver disease (NAFLD). Although the only effective treatment for NAFLD is caloric restriction and weight loss, existing data show that atorvastatin, a hydroxymethyl-glutaryl-CoA reductase inhibitor, can be used safely in patients with NAFLD and improves hepatic histology. To gain further insight into the molecular mechanisms of atorvastatin's therapeutic effect on NAFLD, we used an experimental model that mimics human consumption of fructose-sweetened beverages. Control, fructose (10% w/v solution) and fructose+atorvastatin (30 mg/kg/day) Sprague-Dawley rats were sacrificed after 14 days. Plasma and liver tissue samples were obtained to determine plasma analytes, liver histology, and the expression of liver proteins that are related to fatty acid synthesis and catabolism, and inflammatory processes. Fructose supplementation induced hypertriglyceridemia and hyperleptinemia, hepatic steatosis and necroinflammation, increased the expression of genes related to fatty acid synthesis and decreased fatty acid β-oxidation activity. Atorvastatin treatment completely abolished histological signs of necroinflammation, reducing the hepatic expression of metallothionein-1 and nuclear factor kappa B binding. Furthermore, atorvastatin reduced plasma (x 0.74) and liver triglyceride (x 0.62) concentrations, decreased the liver expression of carbohydrate response element binding protein transcription factor (x 0.45) and its target genes, and increased the hepatic activity of the fatty acid β-oxidation system (x 1.15). These effects may be related to the fact that atorvastatin decreased the expression of fructokinase (x 0.6) in livers of fructose-supplemented rats, reducing the metabolic burden on the liver that is imposed by continuous fructose ingestion.

Topics: Animals; Atorvastatin; Basic Helix-Loop-Helix Leucine Zipper Transcription Factors; Cyclic AMP-Dependent Protein Kinases; Dietary Carbohydrates; Disease Models, Animal; Down-Regulation; Fatty Acids; Fatty Liver; Fructokinases; Fructose; Gene Expression Regulation; Hepatitis; Heptanoic Acids; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypertriglyceridemia; I-kappa B Proteins; Inflammation Mediators; Lipid Metabolism; Liver; Male; Metallothionein; Necrosis; NF-kappa B; Non-alcoholic Fatty Liver Disease; Oxidation-Reduction; Phosphorylation; Pyrroles; Rats; Rats, Sprague-Dawley; Triglycerides

Mangiferin (MGN), a C-glucosylxanthone was investigated for its ability to protect against methylmercury (MeHg) induced neurotoxicity by employing IMR-32 (human neuroblastoma) cell line. MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] and clonogenic cell survival assays confirmed the efficacy of MGN supplementation in attenuating MeHg-induced cytotoxicity. Pre-treatment with MGN significantly (p<0.01) inhibited MeHg-induced DNA damage (micronuclei, olive tail moment and % tail DNA) thereby demonstrating MGN's antigenotoxic potential. Also, pre-treatment with MGN significantly reduced MeHg-induced oxidative stress, intra-cellular Ca(2+) influx and inhibited depolarization of mitochondrial membrane. MGN pre-treated cells demonstrated a significant (p<0.05) increase in the GSH and GST levels followed by a significant (p<0.05) decrease in malondialdehyde (MDA) formation. In addition, inhibition of MeHg induced apoptotic cell death by MGN was demonstrated by microscopic, Annexin-V FITC and DNA fragmentation assays and further confirmed by western blot analysis. The present findings indicated the protective effect of MGN against MeHg induced toxicity, which may be attributed to its anti-genotoxic, anti-apoptotic and anti-lipid peroxidative potential plausibly because of its free radical scavenging ability, which reduced the oxidative stress and in turn facilitated the down-regulation of mitochondrial apoptotic signalling pathways.

Topics: Annexin A5; Antioxidants; Apoptosis; bcl-2-Associated X Protein; Calcium; Caspase 3; Cell Line, Tumor; Cell Nucleus; Cell Survival; Comet Assay; Cytochromes c; Cytokinesis; Cytoprotection; DNA Damage; DNA Fragmentation; Dose-Response Relationship, Drug; Drug Interactions; Free Radical Scavengers; Humans; Inhibitory Concentration 50; Intracellular Space; Membrane Potential, Mitochondrial; Metallothionein; Methylmercury Compounds; Micronucleus Tests; Necrosis; Neuroblastoma; NF-E2-Related Factor 2; Oxidative Stress; Proto-Oncogene Proteins c-bcl-2; Reactive Oxygen Species; Tubulin; Tubulin Modulators; Tumor Stem Cell Assay; Xanthones

This study investigated the effects of different doses of 17-beta-estradiol (E(2)) in Rhamdia quelen. Groups of males exposed to different doses of E(2) (0.1 mg kg(-)(1), 1 mg kg(-)(1) and 10 mg kg(-)(1)) were compared with non-exposed male and female fish groups. Among the considered biomarkers, no significant differences were observed for micronuclei test, reduced glutathione concentration and lipid peroxidation. All E(2)-treated individuals had decreased glutathione S-transferase activity. Increased catalase and superoxide dismutase activities, increased vitellogenin expression and decreased metallothionein concentration were observed in males treated with the highest dose. Liver of all test groups showed necrotic areas, but cytoplasm vacuolization was again found only in the individuals exposed to highest dose. E(2) causes deleterious hepatic effects to R. quelen, and vitellogenin expression, catalase and superoxide dismutase activity and metallothionein concentration represent appropriate biomarkers for studying E(2) effects. Additionally, the response of some biomarkers was similar in males exposed to E(2) and unexposed females, and therefore exposure to endocrine disruptors may cause consequences for fish populations.

Topics: Animals; Blotting, Western; Catalase; Catfishes; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Estradiol; Estrogens; Female; Fresh Water; Glutathione; Glutathione Peroxidase; Glutathione Transferase; Lipid Peroxidation; Liver; Male; Metallothionein; Micronucleus Tests; Necrosis; Superoxide Dismutase; Vitellogenesis; Vitellogenins; Water Pollutants, Chemical

Zinc is a basic trace element that plays important roles in brain and, consequently, its homeostasis needs to be critically controlled. High zinc concentrations in the interneuron synaptic space may induce neuronal death through mechanisms still partially solved. Undifferentiated pheochromocytoma (PC12) cells have been used to study zinc toxicity. As these cells can be differentiated into neuronal-like cells, the results obtained from differentiated cultures are more useful to understand zinc toxicity in neurons. In this paper, we show by flow cytometry that nerve growth factor (NGF) induces PC12 cells differentiation characterized by cell cycle arrest in the G1/G0 phase, similarly to that observed in serum-deprived cultures. Zinc induces cell death in NGF-differentiated PC12 cultures with an EC(50) value of 143+/-14 microM, which reveals a higher sensitivity with respect to undifferentiated PC12 cultures (EC(50), 308+/-32 microM) and a similar response to that obtained in hippocampal neurons (134+/-12 microM). Thus, the differentiation process appeared responsible for such increase in sensitivity. To further support this tenet, when the NGF differentiation was impaired in presence of 10 microM MK-801, a selective blocker of the N-methyl-d-aspartate (NMDA) receptor that plays a role in the differentiation process, the higher sensitivity to zinc was reverted to an EC(50) value of 241+/-26 microM. Flow cytometry experiments showed that NGF-differentiated PC12 cells in presence of zinc were positive for propidium iodide but not for annexin-V labeling. These results, together with data from fluorescent labeling of nuclear fragmentation, caspase-3 activation, and reactive oxygen species generation, support the view that zinc toxicity in NGF-differentiated PC12 cells takes place mainly through a necrotic process.

Topics: Animals; Caspase 3; Cell Cycle; Cell Death; Cell Differentiation; Cell Survival; Cells, Cultured; Dizocilpine Maleate; Excitatory Amino Acid Antagonists; Hippocampus; Metallothionein; Necrosis; Nerve Growth Factor; Neurons; PC12 Cells; Rats; Reactive Oxygen Species; Trace Elements; Zinc Sulfate

Training under hypoxia has several advantages over normoxic training in terms of enhancing the physical performance. Therefore, we tested the protective effect of hypoxia preconditioning by hypoxia mimetic cobalt chloride against exercise-induced oxidative damage in the skeletal muscles and improvement of physical performance.. Male Sprague-Dawley rats were randomly divided into four groups (n=8), namely control, cobalt-supplemented, training and cobalt with training. The red gastrocnemius muscle was examined for all measurements, viz. free radical generation, lipid peroxidation, muscle damage and antioxidative capacity.. Hypoxic preconditioning with cobalt along with training significantly increased physical performance (33%, P<0.01) in rats compared with training-only rats. Cobalt supplementation activated cellular oxygen sensing system in rat skeletal muscle. It also protected against training-induced oxidative damage as observed by an increase in the GSH/GSSG ratio (36%, P<0.001; 28%, P<0.01 respectively) and reduced lipid peroxidation (15%, P<0.01; 31%, P<0.01 respectively) in both trained and untrained rats compared with their respective controls. Cobalt supplementation along with training enhanced the expression of antioxidant proteins haem oxygenase-1 (HO-1; 1.2-fold, P<0.05) and metallothionein (MT; 4.8-fold, P<0.001) compared with training only. A marked reduction was observed in exercise-induced muscle fibre damage as indicated by decreased necrotic muscle fibre, decreased lipofuscin content of muscle and plasma creatine kinase level (16%, P<0.01) in rats preconditioned with cobalt.. Our study provides strong evidence that hypoxic preconditioning with cobalt chloride enhances physical performance and protects muscle from exercise-induced oxidative damage via GSH, HO-1 and MT-mediated antioxidative capacity.

Topics: Animals; Antioxidants; Cell Hypoxia; Cobalt; Creatine Kinase, MM Form; Cytoprotection; Disease Models, Animal; Dose-Response Relationship, Drug; Glutathione; Heme Oxygenase (Decyclizing); Lactic Acid; Lipid Peroxidation; Lipofuscin; Male; Metallothionein; Mitochondria, Muscle; Mitochondrial Proteins; Muscle, Skeletal; Muscular Diseases; Necrosis; Oxidative Stress; Physical Endurance; Physical Exertion; Rats; Rats, Wistar; Reactive Oxygen Species

Iron is used to cure iron-deficient anaemia but can also be toxic to the intestine. Fe(II) toxicity was investigated using differentiated human intestinal Caco-2 cells treated with 15 and 50 microM of Fe(II)/ascorbate for 2h (acute phase), and followed for 24h after iron removal and replacement of complete culture medium (late phase). During the acute phase damage to tight junctions occurred as demonstrated by an increase in cell monolayer permeability and by partial delocalization of the tight junction protein claudin 4 from the plasma membrane to an intracellular compartment. At the end of the late phase, cells treated with 15 microM Fe(II) showed full restoration of claudin 4 localization to the plasma membrane and their tight junction permeability returned to values close to those of control cells. Conversely, cells treated with 50 microM Fe(II) showed sustained and irreversible damage to the tight junctions, accompanied by apoptosis and necrosis. Activation of NF-kappaB occurred at both Fe(II) concentrations after 30 min of Fe(II) treatment, followed, at the end of the acute phase, by a strong induction of mRNA coding for heat shock protein 70 and metallothionein 2A. Our results indicate that intestinal cells respond to iron toxicity by strongly activating two genes involved in cell response to stress, although the outcome in terms of cell survival is different depending on the dose of treatment, namely almost complete restoration of epithelial permeability and cell survival at 15 microM Fe(II), and progressive and irreversible cytotoxicity leading to apoptosis and necrosis at 50 microM Fe(II).

Topics: Apoptosis; Caco-2 Cells; Gene Expression Regulation; HSP70 Heat-Shock Proteins; Humans; Iron; Metallothionein; Necrosis; NF-kappa B; Oxidative Stress; Permeability; Tight Junctions

A close link between stress protein up-regulation and oxidative damage may provide a novel therapeutic tool to counteract nephrotoxicity induced by toxic metals in the human population, mainly in children, of industrialized countries. Here we analysed the time course of the expression of several heat shock proteins, glucose-regulated proteins and metallothioneins in a rat proximal tubular cell line (NRK-52E) exposed to subcytotoxic doses of inorganic mercury and lead. Concomitantly, we used morphological and biochemical methods to evaluate metal-induced cytotoxicity and oxidative damage. In particular, as biochemical indicators of oxidative stress we detected reactive oxygen species (ROS) and nitrogen species (RNS), total glutathione (GSH) and glutathione-S-transferase (GST) activity. Our results clearly demonstrated that mercury increases ROS and RNS levels and the expressions of Hsp25 and inducible Hsp72. These findings are corroborated by evident mitochondrial damage, apoptosis or necrosis. By contrast, lead is unable to up-regulate Hsp72 but enhances Grp78 and activates nuclear Hsp25 translocation. Furthermore, lead causes endoplasmic reticulum (ER) stress, vacuolation and nucleolar segregation. Lastly, both metals stimulate the over-expression of MTs, but with a different time course. In conclusion, in NRK-52E cell line the stress response is an early and metal-induced event that correlates well with the direct oxidative damage induced by mercury. Indeed, different chaperones are involved in the specific nephrotoxic mechanism of these environmental pollutants and work together for cell survival.

Topics: Animals; Apoptosis; Biomarkers; Cell Line; Cell Proliferation; Cell Survival; Dose-Response Relationship, Drug; Endoplasmic Reticulum; Endoplasmic Reticulum Chaperone BiP; Glutathione; Glutathione Transferase; Heat-Shock Proteins; HSP27 Heat-Shock Proteins; HSP72 Heat-Shock Proteins; Kidney Tubules, Proximal; Lead; Mercuric Chloride; Metallothionein; Mitochondria; Molecular Chaperones; Necrosis; Nitric Oxide; Oxidative Stress; Rats; Reactive Oxygen Species; Time Factors

The present study has been carried out to investigate the protective role of taurine against cadmium (Cd)-induced oxidative impairment in murine liver. Oral administration of cadmium chloride (CdCl(2)) at a dose of 4mg/kg body weight for 6 days increased the accumulation of the Cd in the liver and diminished the liver weight to body weight ratio. The CdCl(2) altered the levels of intracellular trace elements, cofactors of various metalloenzymes and increased the activities of serum marker enzymes related to liver dysfunction. In addition, Cd intoxication also attenuated intracellular antioxidant power, the activities of antioxidant enzymes as well as the levels of cellular metabolites. Moreover, level of hepatic metallothionein, lipid peroxidation, protein carbonylation, DNA fragmentation, concentration of intracellular reactive oxygen species (ROS) and the activities of cytochrome P450s have been increased due to Cd toxicity. In addition to the oxidative impairments, Cd exposure caused hepatic cell death mainly via the necrotic pathway. Oral administration of taurine at a dose of 100mg/kg body weight for 5 days prior to CdCl(2) intoxication prevented the alterations of all the toxic-induced hepatic damages. Histological studies also supported the beneficial role of taurine against Cd-induced hepatic damages. Combining all, results suggest that taurine could protect hepatic tissues against Cd-induced oxidative stress probably through its antioxidant activity.

Topics: Analysis of Variance; Animals; Antioxidants; Cadmium; Cadmium Chloride; Cytochrome P-450 Enzyme System; DNA Fragmentation; Dose-Response Relationship, Drug; Glutathione; Glutathione Transferase; Lipid Peroxidation; Liver; Male; Metallothionein; Mice; Necrosis; Organ Size; Oxidative Stress; Oxidoreductases; Protein Carbonylation; Random Allocation; Reactive Oxygen Species; Sulfhydryl Compounds; Taurine; Trace Elements

Iterations in Ca2+ and Mg2+ balance accompany aldosteronism (inappropriate for dietary Na+ intake). Increased Zn excretion and Zn translocation to injured tissues, including the heart, also occurs. Several causes and consequences of Zn dyshomeostasis in rats receiving aldosterone/salt treatment (ALDOST) were examined. (1) To study the role of urinary acidification in promoting hyperzincuria, acetazolamide (75 mg/kg), a carbonic anhydrase inhibitor, was used as cotreatment to raise urinary HCO3 excretion. (2) To assess Zn levels in the heart, including cardiomyocyte cytosolic free [Zn2+]i and mitochondrial Zn, the expression of metallothionein (MT-I), a Zn binding protein, and biomarkers of oxidative stress were examined. (3) Oxidative stress and cardiac pathology in response to ZnSO4 supplement (40 mg/d) were also studied. Comparison of controls and rats receiving 4 weeks ALDOST revealed the following: (1) an acidification of urine and metabolic alkalosis associated with increased urinary Zn excretion and hypozincemia, each of which were prevented by acetazolamide; (2) a rise in cardiac Zn, including increased [Zn2+]i and mitochondrial Zn, associated with increased tissue MT-I, 8-isoprostane, malondialdehyde, and gp91(phox), coupled with oxidative stress in plasma and urine; (3) ZnSO4 prevented hypozincemia, but not ionized hypocalcemia, and attenuated oxidative stress and microscopic scarring without preventing the vasculitis and perivascular fibrosis of intramural coronary arteries. Thus, the hyperzincuria seen with ALDOST is due to urinary acidification. The oxidative stress that appears in the heart is accompanied by increased tissue Zn serving as an antioxidant. Cotreatment with ZnSO4 attenuated cardiomyocyte necrosis; however, polynutrient supplement may be required to counteract the dyshomeostasis of all 3 cations that accompanies aldosteronism and contributes to cardiac pathology.

Topics: Acetazolamide; Aldosterone; Animals; Calcium; Chronic Disease; Disease Models, Animal; Homeostasis; Hydrogen-Ion Concentration; Hyperaldosteronism; Magnesium; Male; Metallothionein; Myocytes, Cardiac; Necrosis; Oxidative Stress; Rats; Rats, Sprague-Dawley; Urine; Zinc; Zinc Sulfate

In the funnel web spider Agelena labyrinthica (Agelenidae; A. l.), sheet web spider Linyphia triangularis (Linyphiidae; L. t.) and wolf spider Xerolycosa nemoralis (Lycosidae; X. n.) from two differently polluted meadow sites in southern Poland, we studied the relations between antioxidant parameters (glutathione, GSH; glutathione peroxidases, GPOX, GSTPx; catalase, CAT; stress proteins-Hsp70, metallothioneins Mts), the intensity of apoptosis and necrosis, and heavy metal burdens of the midgut gland. Cellular reactions against stress caused by pollutants seemed to be sex-dependent. The concentrations of Zn and Cu in the midgut glands of male A. l. and X. n. were more than double that of the females, from both study sites. In male spiders from the heavily polluted site, both negative correlations (activity of caspase-3-like proteins vs Cu, Zn concentration; number of depolarized mitochondria vs Cu concentration) and positive correlations (number of necrotic cells vs Cu concentrations; activity of CAT vs Zn ) were noted. The defense of males against high metal content and its prooxidative effects is based mainly on GSH and CAT. In females the antioxidative reactions are species-specific and depend mainly on high peroxidase activity and on stress protein level. The increase in the number of apoptotic cells in the midgut gland of female spiders from the heavily polluted site suggests the defensive role of this process in maintaining the proper functioning of this organ.

Topics: Animals; Antioxidants; Apoptosis; Caspase 3; Catalase; Environmental Monitoring; Environmental Pollutants; Female; Gastrointestinal Tract; Glutathione; Glutathione Peroxidase; HSP70 Heat-Shock Proteins; Male; Membrane Potential, Mitochondrial; Metallothionein; Metals, Heavy; Mitochondria; Necrosis; Poland; Species Specificity; Spiders; Stress, Physiological

This study evaluated the efficacy of fish opercular external (skin) and inner (opercular membrane) epithelium as an in vitro model for toxic and other substances studies. The rainbow trout (Oncorhynchus mykiss) operculum was cultured in 12-well dishes containing sterile Leibovitz 15 (L-15) supplemented with glutamine medium during 24h at 9 degrees C, and the effect of copper, a toxic agent, and/or cortisol, an endogenous agent, on the epithelial cells was analyzed using light microscopy techniques. The opercula were submitted to four treatments: (i) control (Cont), L-15 medium only, (ii) 0.28 microM cortisol (Cort), (iii) 100 microM CuSO(4) (Cu), and (iv) 0.28 microM cortisol+100 microM CuSO(4) (Cort-Cu). The tissue condition after 24h incubation was analyzed by staining the mucous cells for neutral and acid mucosubstances. Cellular necrosis was evaluated by measuring the lactate dehydrogenase (LDH) leakage at 12 and 24h incubation. Cellular proliferation, apoptosis, metallothionein (MT) and glucocorticoid receptor (GR) expression were evaluated by immunohistochemistry. The LDH leakage was higher and the proliferating cell nuclear antigen (PCNA) positive-stained cells were lower in Cu treatment in both, epidermis and opercular membrane. Apoptotic cells in the opercular membrane were higher in the Cort and Cort-Cu treatments while, in the epidermis, they were higher in Cu and Cort-Cu treatments. GR-positive stained cells decreased significantly in all treatments in both epithelia and the MT-positive cells increased in the Cu and Cort-Cu treated groups. Copper showed to be a potent toxic stressor killing the cells via necrosis, decreasing the number of PCNA-positive cells and inducing MT synthesis while cortisol did not affect the MT synthesis, although it might stimulate apoptosis. The results are evidence that the opercular epithelia serve as a suitable model for studying in vitro effects of toxic agents, as well as endogenous factors on the cellular responses without interference of the physiological state of fish being useful to predict in vivo toxicity.

Topics: Animals; Apoptosis; Cells, Cultured; Copper; Culture Media; Epithelial Cells; Hydrocortisone; Immunohistochemistry; In Situ Nick-End Labeling; L-Lactate Dehydrogenase; Metallothionein; Mitosis; Necrosis; Oncorhynchus mykiss; Proliferating Cell Nuclear Antigen; Receptors, Glucocorticoid; Skin

A previous study (Oliver, J.R., Mara, T.W., Cherian, M.G. 2005. Impaired hepatic regeneration in metallothionein-I/II knockout mice after partial hepatectomy. Exp. Biol. Med. 230, 61-67) has shown an impairment of liver regeneration following partial hepatectomy (PH) in metallothionein (MT)-I and MT-II gene knockout (MT-null) mice, thus suggesting a requirement for MT in cellular growth. The present study was undertaken to investigate whether MT may play a similar role in hepatic injury and regeneration after acute treatment with thioacetamide (TAA). Hepatotoxicity of TAA is caused by the generation of oxidative stress. TAA was injected ip to both wild-type (WT) and MT-null mice. Mice were killed at 6, 12, 24, 48, 60, and 72 h after injection of TAA (125 mg/kg) or 48 h after injection of saline (vehicle control), and different parameters of hepatic injury were measured. The levels of hepatic lipid peroxidation were increased at 12 h in both types of mice; however, lipid peroxidation was significantly less in WT mice than MT-null mice at 48 h after injection of TAA. Analysis of hepatic glutathione (GSH) levels after TAA injection showed depletion of GSH at 12 h in WT mice and at 6 h in MT-null mice; however, significantly more GSH was depleted early (6-24 h) in MT-null mice than WT mice. An increase in hepatic iron (Fe) levels was observed in both types of mice after injection of TAA, but Fe levels were significantly higher in MT-null mice than WT mice at 6-60 h. The levels of hepatic copper (Cu) and zinc (Zn) were significantly higher in WT mice than MT-null mice at 6-60 h for Cu, and at 24 h and 60 h for Zn, respectively. Histopathological examination showed hemorrhagic necrosis in the liver of both types of mice at 12-72 h, with hepatic injury being more prominent in MT-null mice than WT mice. The hepatic MT levels were increased in WT mice after injection of TAA, and were highest at 24-72 h. Immunohistochemical staining for MT in WT mice indicated the presence of MT in both nucleus and cytoplasm of hepatocytes at 24-72 h after TAA injection. Cell proliferation, as assessed by immunohistochemical staining for proliferating cell nuclear antigen, was detected mainly in the livers of WT mice at 48-72 h after TAA treatment. Hepatic proliferation index in MT-null mice was very low as compared to WT mice during liver regeneration after injection of TAA. These results show that the liver cells of MT-null mice with no functional MT are unable to regenerate afte

Topics: Acute Disease; Animals; Cell Proliferation; Chemical and Drug Induced Liver Injury; Glutathione; Iron; Lipid Peroxidation; Liver; Liver Regeneration; Metallothionein; Mice; Mice, Knockout; Necrosis; Thioacetamide

The introduction of apo-ferritin or the iron chelator DFO (desferrioxamine) conjugated to starch into the lysosomal compartment protects cells against oxidative stress, lysosomal rupture and ensuing apoptosis/necrosis by binding intralysosomal redox-active iron, thus preventing Fenton-type reactions and ensuing peroxidation of lysosomal membranes. Because up-regulation of MTs (metallothioneins) also generates enhanced cellular resistance to oxidative stress, including X-irradiation, and MTs were found to be capable of iron binding in an acidic and reducing lysosomal-like environment, we propose that these proteins might similarly stabilize lysosomes following autophagocytotic delivery to the lysosomal compartment. Here, we report that Zn-mediated MT up-regulation, assayed by Western blotting and immunocytochemistry, results in lysosomal stabilization and decreased apoptosis following oxidative stress, similar to the protection afforded by fluid-phase endocytosis of apo-ferritin or DFO. In contrast, the endocytotic uptake of an iron phosphate complex destabilized lysosomes against oxidative stress, but this was suppressed in cells with up-regulated MT. It is suggested that the resistance against oxidative stress, known to occur in MT-rich cells, may be a consequence of autophagic turnover of MT, resulting in reduced iron-catalysed intralysosomal peroxidative reactions.

Topics: Animals; Apoptosis; Autophagy; Cell Line; Iron; Lysosomes; Macrophages; Metallothionein; Mice; Necrosis; Oxidation-Reduction; Oxidative Stress; Protein Binding; Up-Regulation; Zinc

It has been shown in various systems that zinc is able to antagonize the catalytic properties of the redox-active transition metals iron and copper, although the process is still unclear. Probably, the protective effect of Zn against oxidative stress is mainly due to the induction of a scavenger metal binding protein such as metallothionein (MT), rather than a direct action. To support this hypothesis, in this study, the effects of Zn, Cu, Fe, Zn + Cu and Zn + Fe treatments were investigated in a fibroblast cell line corresponding to an SV40-transformed MT-1/-2 mutant (MT-/-), and in wild type (MT+/+), by valuing metal concentrations and apoptotic and/or necrotic processes. We also investigated the synthesis of MT and the levels of both MT-1 and MT-2 mRNAs. In MT+/+ cells, co-treatment with Zn + Fe caused a decrease in Fe content compared to treatment with Fe alone. After Zn and Zn + Cu exposure the expression of MT-1 and MT-2 isoforms increased with a concomitant increase in MT synthesis. Annexin V-FITC and propidium iodide staining revealed necrotic or apoptotic cells in terminal stages, especially after Fe treatments. Immunofluorescent staining with an anti-ssDNA Mab and annexin detected a lower signal in co-treated cells compared to the single treatments in both cell lines. The intensity and quantity of fluorescence resulting from anti-ssDNA and Annexin V staining of MT null cells was higher compared to wild type cells. These results suggest that Zn alone does not completely exert an anti-oxidant effect against Cu and Fe toxicity, but that induction of MT is necessary.

Topics: Animals; Apoptosis; Cell Line; Cells, Cultured; Copper; Fibroblasts; Gene Expression Regulation; Iron; Metallothionein; Mice; Mice, Knockout; Necrosis; Protein Isoforms; RNA, Messenger; Zinc

This laboratory has shown that both Cd(+2) and As(+3) can malignantly transform human urothelial cells. The present study examined metal resistance and the mechanism of cell death when the parental and malignantly transformed UROtsa cells were exposed to Cd(+2) and As(+3). It was shown that the malignantly transformed UROtsa cells were more resistant to the toxic effects of both metals. The assessment of the mode of cell death demonstrated that the parental UROtsa cells died by both apoptosis and necrosis when exposed to either metal. It was shown that apoptosis was the more prominent mechanism of cell death, accounting for over 50% of cell death. Apoptotic cell death was determined by the observation of fragmented nuclei using 4',6-diamidino-2-phenylindole staining, the formation of a DNA ladder, and the detection of cleaved caspase-3 and caspase-9 products in the cell lysates. Necrotic cell death was determined by measuring the release of lactate dehydrogenase into the growth medium. It was determined that the extent of apoptosis of the malignantly transformed UROtsa cells was decreased and that the extent of necrosis was increased compared to the parental UROtsa cells. These observations are consistent with in vivo studies which suggest that As(+3) can act as a tumor promoter during the regeneration of the bladder urothelium. The present in vitro studies suggest that As(+3)-induced cytotoxicity could set the stage for tissue repair due to its own inherent toxicity to normal urothelium, and then subsequently act as a tumor promoter during the regeneration process through the stimulation of the regrowth of cells that have gained increased resistance to As(+3).

Topics: Apoptosis; Arsenites; Cadmium; Caspases; Cell Line, Transformed; Cell Transformation, Neoplastic; DNA Fragmentation; Dose-Response Relationship, Drug; Drug Resistance, Multiple; Humans; L-Lactate Dehydrogenase; Metallothionein; Necrosis; Urothelium

Photoactivation of hypericin is known to generate reactive oxygen species and induce phototoxic effects. However, modulation of the cellular antioxidant defense would influence the extent and severity of the photodynamic effects. We have previously shown that hypericin-mediated photodynamic therapy (PDT) induced a significant reduction of Glutathione S-transferase activity. In this study, we investigated the phototoxic effects of hypericin-mediated PDT in nasopharyngeal cancer (NPC) in vitro and analyzed the expression of metallothionein (MT), a family of potential free radical scavengers. HK1 NPC cells were subjected to PDT treatment in vitro, and the effects on cell death were analyzed by flow cytometry (using propidium iodide and Annexin V staining) and transmission electron microscopy. The expression profile of MT-1E and MT-2A isoforms (the only functional MT isoforms expressed in HK1 NPC cells) were determined by quantitative real-time RT-PCR. The results showed that hypericin PDT induced necrotic cell death as evidenced by the absence of a subdiploid peak and decreased Annexin-V fluorescence. Ultrastructural examination verified the presence of cell necrosis. There was a significant up-regulation of MT-1E and MT-2A isoforms six hours following PDT, with an approximately 50-fold rise in the expression level of MT-1E and a 15-fold increase of MT-2A. Hence, despite the up-regulation of MT, cells still succumbed to PDT-induced necrosis. It appears that the oxidative stress induced by PDT overwhelmed the antioxidant defense mechanism such as the alteration of MT levels in tumor cells.

Topics: Anthracenes; Flow Cytometry; Humans; In Vitro Techniques; Metallothionein; Nasopharyngeal Neoplasms; Necrosis; Perylene; Photochemotherapy; Photosensitizing Agents; Protein Isoforms; Reverse Transcriptase Polymerase Chain Reaction; Tumor Cells, Cultured; Up-Regulation

To study the possibility of metallothionein (MT) participating in the process of ischemia/reperfusion protection.. The model of pig latissimus dorsi island flap and cultured myocytes were used in this study. The island flap was preconditioned with ischemia and the cultured myocytes were treated with anoxia. MT was detected in the flap and the cultured myocytes instantly and at 12 and 24 hours after the treatment. The protection effect of precondition on flap ischemia/reperfusion damage was observed by recording flap necrosis and the serum LDH level. The protection effect of precondition on myocyte anoxia/reoxygenation damage was observed by recording the cell survival rate and MDA content. The change in the protection effect was also observed after the MT was inhibited by PD098059.. The content of MT in cultured myocytes increased obviously 12 hours after precondition. Similar result was observed in flap tissue 24 hours later. Compared with the non-conditioned flap, the necrosis areas of the preconditioned flaps were smaller, the serum LDH was lower. The survival rate of preconditioned cultured myocytes was higher compared with non-conditioned cells (P < 0.05). The content of MDA and the quantity of discharged LDH were less (P < 0.05). When MT was inhibited by PD098059, the delaying protection of precondition disappeared. The observed quotas at that time were the same with those of simple I/R or A/R groups (P > 0.05).. 24 hours after precondition, the flap and the cultured myocytes are protected from re-damage of I/R or A/R. MT is involved in this process.

Topics: Animals; Cell Hypoxia; Cell Survival; Cells, Cultured; Enzyme Inhibitors; Flavonoids; Ischemic Preconditioning; L-Lactate Dehydrogenase; Malondialdehyde; Metallothionein; Muscles; Necrosis; Reperfusion Injury; Surgical Flaps; Swine; Time Factors

The release of reactive oxygen species (ROS) has been proposed as a cause of streptozotocin (STZ)-induced beta-cell damage. This initiates a destructive cascade, consisting of DNA damage, excess activation of the DNA repair enzyme poly(ADP-ribose) polymerase, and depletion of cellular NAD+. Metallothionein (MT) is an inducible antioxidant protein that has been shown to protect DNA from chemical damage in several cell types. Therefore, we examined whether overexpression of MT could protect beta-cell DNA and thereby prevent STZ-induced diabetes. Two lines of transgenic mice were produced with up to a 30-fold elevation in beta-cell MT. Cultured islets from control mice and MT transgenic mice were exposed to STZ. MT was found to decrease STZ-induced islet disruption, DNA breakage, and depletion of NAD+. To assess in vivo protection, transgenic and control mice were injected with STZ. Transgenic mice had significantly reduced hyperglycemia. Ultrastructural examination of islets from STZ-treated mice showed that MT prevented degranulation and cell death. These results demonstrate that MT can reduce diabetes and confirm the DNA damage mechanism of STZ-induced beta-cell death.

Topics: Animals; Cell Degranulation; Culture Techniques; Diabetes Mellitus, Experimental; DNA Damage; Hyperglycemia; Islets of Langerhans; Metallothionein; Mice; Mice, Inbred Strains; Mice, Transgenic; Necrosis; Streptozocin

Sensitivity to cadmium (Cd)-induced testicular injury varies greatly among mouse strains. For instance, 129/SvJ (129) mice are highly sensitive while C57BL/6J (C57) mice are refractory to Cd-induced testicular injury. Metallothionein (MT), a Cd-binding protein, is thought to be responsible for the strain susceptibility to Cd toxicity. In this study, MT-I/II knockout (MT-null) and wild-type 129 mice were used to determine the role of MT in Cd-induced testicular injury. Two additional strains of mice (C57 and the C57 x 129 F1cross) were also used to help define the role of genetic background in Cd toxicity. Mice were given 5-20 micromol/kg ip CdCl(2) and testicular injury was examined 24 h later by histopathology and testicular hemoglobin concentration. Cd produced dose-dependent testicular injury in all strains of mice, except for C57 mice, in which testicular injury could not be produced. MT-null mice were more sensitive than C57 x 129 mice but were equally sensitive as 129 mice to Cd-induced testicular injury. Fourteen days after 15 micromol/kg ip Cd administration, testicular atrophy was evident in MT-null, 129, and C57 x 129 mice but was absent in C57 mice. The resistance of C57 mice to Cd-induced testicular injury could not be attributed solely to a decreased uptake of (109)Cd nor to a greater amount of testicular MT. Microarray analysis revealed a higher expression of glutathione peroxidase in the testes of C57 mice, as well as genes encoding antioxidant components and DNA damage/repair, but their significance to Cd-induced injury is not immediately clear. Thus, this study demonstrates that it is genetic strain, not MT genotype, that is mechanistically important in determining susceptibility to Cd-induced testicular injury.

Topics: Animals; Cadmium; Crosses, Genetic; DNA Damage; DNA Repair; Genetic Predisposition to Disease; Genotype; Glutathione Peroxidase; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Necrosis; Oligonucleotide Array Sequence Analysis; Organ Size; Phenotype; Species Specificity; Testicular Diseases; Testis

The aim of this study was to investigate the morphologic and biochemical effects of excess dietary copper in young and adult rats of different sex.. Adult Fischer 344 male and female rats were given a diet containing 1500 ppm copper for 18 weeks. Young male and female rats were fed a similar copper-loaded diet from birth until 16 weeks of age. Age- and sex-matched control rats were fed a normal rodent diet (<10 ppm copper). Serum liver enzyme activity was determined in all rats. Livers were sampled for histology, histochemistry (rhodanine), immunohistochemical detection of metallothionein and copper analysis by atomic absorption spectrophotometry. Hepatic metallothionein and zinc concentrations were measured in adult rats.. Excess dietary copper caused substantial liver injury, as evidenced by morphologic changes and increased activity of serum ALT, GGT, and SDH. All copper-loaded rats had significantly (p< 0.05) increased hepatic copper concentrations compared to controls. However, young copper-loaded rats accumulated more hepatic copper, had more severe liver changes, and had higher serum liver enzyme activities than adult rats. Histologic changes in copper-loaded rats consisted of multifocal hepatitis and widespread single-cell necrosis. Cytoplasmic copper was detected histochemically in centroacinar zone 1 (portal) and mid-zone in copper-loaded rats. Immunoreactivity for metallothionein was prominent in necrotic hepatocytes and within inflammatory foci in copper-loaded rats. However, differences in hepatic metallothionein concentrations were not detected between adult copper-loaded and control rats.. Young Fischer 344 rats are more susceptible than adults to copper-induced liver injury.

Topics: Aging; Alanine Transaminase; Animals; Bilirubin; Chemical and Drug Induced Liver Injury; Copper; Diet; Female; gamma-Glutamylcyclotransferase; L-Iditol 2-Dehydrogenase; Liver; Male; Metallothionein; Necrosis; Pregnancy; Rats; Rats, Inbred F344; Sex Characteristics; Zinc

An experimental study was performed on orchiopathy (testis disorder) by using cadmium (Cd) and on the prevention of orchiopathy by the administration of selenium (Se). By a single administration of Cd 1.4 mg/kg (12.4 mumol/kg) or a second administration of Cd 1.4 mg/kg 24 hr after the administration of Cd 1.0 mg/kg (8.9 mumol/kg), the testis of a mouse showed ex-tensive necrosis, and an extreme decrease of glutathione (GSH) concentration accompanied by an increase of thiobarbituric acid reactive substances (TBARS). When Se 1.4 mg/kg (17.7 mumol/kg) was given at the same time as Cd 1.0 mg/kg, the disorder was completely prevented and their serum Cd and Se concentrations were 1165 +/- 268 ng/ml and 534 +/- 128 ng/ml, respectively. However, when Se was given, separately from the Cd administration, either 24 hr or 72 hr before Cd administration, no effect to prevent testis disorder was found. On the other hand, when Se 1.4 mg/kg and Cd 1.0 mg/kg were given simultaneously and then Cd 1.4 mg/kg was administered 24 hr and 72 hr after the simultaneous injection, respectively, there was no sign of disorder caused by the second administration of Cd. When Cd was given after administration of Cd and Se, Cd concentration in the testis (0.88 +/- 0.078 microgram/g and 0.77 +/- 0.03 microgram/g) was about twice as much as the concentration in the case of no administration of Se (0.30 +/- 0.04 microgram/g). The testicular dysfunction could not be explained by the increased Cd concentration in the testis. The groups with high Cd concentration in the testis were accompanied by an increase in metallothionein (92.8 +/- 18.6 micrograms/g and 92.5 +/- 7.3 micrograms/g), but these did not exceed the level of the control group (94.5 +/- 8.4 micrograms/g) which had neither Cd nor Se injections. In the groups with testicular necrosis, concentrations of zinc (Zn) and magnesium (Mg) were decreased while an increase in concentrations of calcium (Ca) and iron (Fe) was observed. These results suggest that Se concentration must be maintained to prevent the testicular disorder caused by Cd.

Topics: Animals; Cadmium; Drug Interactions; Glutathione; Male; Metallothionein; Mice; Mice, Inbred ICR; Necrosis; Selenium; Testicular Diseases; Testis; Thiobarbituric Acid Reactive Substances

Mice express four distinct metallothioneins (MTs) that have similar metal-binding properties. MT-I and MT-II are expressed coordinately in most organs, whereas MT-III is expressed predominantly in a subset of neurons and MT-IV is expressed in certain stratified epithelia. The restricted expression of MT-III suggests that it may severe a specialized function. To test this hypothesis, transgenic mice were generated that express MT-III in the wider expression domain of MT-I. Similar transgenic lines expressing extra MT-I under the same regulation were generated as controls for the effect of over-expression of MT. Transgenic mice that express MT-III ectopically frequently die at 2-3 months of age. The pancreata of moribund mice were abnormally small and histological examination, at various ages, revealed a progressive degeneration of the acinar cells. At early stages multifocal acinar cell eosinophilia and swollen nuclei were seen and this pathology progressed to multifocal acinar cell necrosis and fibrosis. The terminal stages were characterized by a loss of the acinar compartment, leaving the islets embedded in a fibrotic remnant. Other organs of these mice were grossly and histologically normal. All organs examined from mice expressing excess MT-I were unremarkable even though expression of either MT-I or MT-III transgenes resulted in similar accumulations of zinc and copper in the pancreata. This study indicates that pancreatic acinar cells are unusually sensitive to chronic expression of MT-III. The mechanism by which MT-III disrupts pancreatic function is unclear, but the results provide further evidence that MT isoforms exhibit distinct properties and probably serve distinct biological functions.

Topics: Animals; Copper; Diet; DNA Probes; Female; Glucagon; Growth Inhibitors; Insulin; Male; Metallothionein; Metallothionein 3; Mice; Mice, Transgenic; Necrosis; Nerve Tissue Proteins; Pancreas; RNA, Messenger; Zinc

To study the neurophysiological functions of metallothioneins (MTs), localization of MT-I and -II was examined immunohistochemically in a variety of brain lesions in dogs, including infarct, laminar cortical necrosis, hemorrhage, invasive growth of tumour, inflammatory lesions in granulomatous meningoencephalitis and distemper encephalitis. MT-I and -II were demonstrated in both nucleus and cytoplasm of hypertrophic astrocytes in most brain lesions examined regardless of the type, size, localization and duration of the lesions. In addition, MT expression was stronger in a population of hypertrophic astrocytes localizing inside of the surviving brain tissue rather than those localizing at the boundary between the surviving brain tissue and necrotic area, where severe inflammatory changes were developing. These results suggest that MT-I and -II may play roles not only in protection of neurons from metals and free radicals ubiquitous in the inflammatory lesions but also in repair of injured neural tissues.

Topics: Adenocarcinoma; Animals; Astrocytes; Brain Diseases; Brain Neoplasms; Cerebral Hemorrhage; Cerebral Infarction; Distemper; Dog Diseases; Dogs; Encephalitis, Viral; Female; Glial Fibrillary Acidic Protein; Hypertrophy; Inflammation; Male; Meningoencephalitis; Metallothionein; Necrosis; Neoplasm Invasiveness; Pituitary Neoplasms

Zinc exhibits inhibitory effects on apoptosis, and a deficiency in this metal generally causes this type of cell death to occur. In the present study, we found that exposure to zinc results in necrosis of prostate carcinoma cells. When zinc acetate was added to LNCaP or PC-3 cells in monolayer culture, they began to detach from the culture dishes, and viability was lost after 4-8 h. Most of the cell death was found to be due to necrosis as determined by double staining with fluorescein-isothiocyanate-labeled annexin V and ethidium bromide, and by detection of hypodiploid cells. Associated with the induction of necrosis was an increase in low molecular-mass proteins, identified by HPLC analysis to be thymosin beta10, parathymosin and GAGE in LNCaP cells, and thymosin beta4, parathymosin and metallothionein in PC-3. The time course of the increase of thymosin beta10 in LNCaP cells and thymosin beta4 in PC-3 cells was consistent with that of appearance of cell detachment and dead cells. These results indicate that zinc can induce necrosis and suggest that production of proteins including beta-thymosins is involved in induction of processes leading to cell detachment.

Topics: Adenocarcinoma; Annexin A5; Apoptosis; Cell Adhesion; Cell Division; Cell Survival; Copper; DNA, Neoplasm; Humans; Male; Metallothionein; Necrosis; Neoplasm Proteins; Prostatic Neoplasms; Thymosin; Tumor Cells, Cultured; Zinc

Arsenic and cadmium are both priority hazardous substances and human carcinogens. Although there is the potential for simultaneous exposure to both metals, the interactions of cadmium and arsenic are not well defined. We examined the toxicity of these metals when given alone or in alternating sequence to adult male Fischer rats. In the first study, a non-toxic dose of arsenic (22.5 micromol NaAsO2/kg, s.c.) was given 24 h before cadmium (10, 20, or 30 micromol CdCl2/kg, s.c.) and toxicity was assessed 24 h later. Arsenic pretreatment markedly reduced mortality in rats given the high dose of cadmium (9 survivors/10 treated) compared to rats given cadmium alone (2/10). Arsenic pretreatment also reduced cadmium-induced hepatotoxicity, as indicated by serum glutamic oxalacetic transaminase (SGOT) activity, and markedly reduced cadmium-induced testicular hemorrhagic necrosis. Arsenic pretreatment produced an 8-fold increase in hepatic levels of metallothionein (MT), a metal-binding protein often associated with cadmium tolerance. In the second study, a non-toxic dose of cadmium (3 micromol CdCl2/kg, s.c.) was given 24 h before arsenic (68, 79, 84, or 90 micro/mol NaAsO2/kg. s.c.) and toxicity was assessed 24 h later. Cadmium pretreatment did not alter the lethality of the high dose of arsenic and had no effect on arsenic-induced hepatotoxicity. Although cadmium pretreatment had no effect on arsenic toxicity, it produced large increases in hepatic MT (26-fold) before the arsenic challenge and greatly enhanced MT induction after the challenge. Thus, even though both arsenic and cadmium induce MT synthesis, only arsenic pretreatment protects against cadmium intoxication, and cadmium pretreatment does not effect arsenic toxicity. Thus, toxic interactions of arsenic and cadmium appear to depend on the sequence of exposure.

Topics: Animals; Arsenic; Cadmium; Chemical and Drug Induced Liver Injury; Drug Administration Schedule; Drug Interactions; Kidney Diseases; Male; Metallothionein; Necrosis; Rats; Rats, Inbred F344; Testis

Immunocytochemically detectable MT and p53 have been found more commonly in comedo DCIS of the breast with high-grade cytology. The aim of this study is to confirm these findings and to investigate the relationship between MT and p53 in a single large series of cases of DCIS of the breast. To this end, 127 cases of DCIS were classified histologically according to architecture, cytonuclear differentiation (grade), presence and extent of intraduct necrosis, and using the Van Nuys system. Sections were immunostained for p53 and MT (E9) using established techniques, and the extent and intensity of staining were assessed semi-quantitively. The results confirmed that there was generally more MT and p53 positivity in poorly differentiated (grade 3) DCIS with extensive necrosis and that MT expression was greater in grade 2 lesions than p53 expression. However, overall there was no statistically significant correlation between p53 and MT staining. The results indicate that MT and p53 overexpression may arise from independent mechanisms in early breast neoplasia.

Topics: Breast Neoplasms; Carcinoma in Situ; Carcinoma, Ductal, Breast; Cell Transformation, Neoplastic; Female; Gene Expression Regulation, Neoplastic; Humans; Immunohistochemistry; Metallothionein; Necrosis; Tumor Suppressor Protein p53

Exposure of humans to toxic metals and metalloids is a major environmental problem. Many metals, such as cadmium, can be hepatotoxic. However, the mechanisms by which metals cause acute hepatic injury are in many cases unknown. Previous reports suggest a major role for inflammation in acute cadmium induced hepatotoxicity. In initial experiments we found that a non-hepatotoxic dose of cadmium chloride (CdCl2; 2.0 mg/kg, i.v.) markedly increased the clearance rate of colloidal carbon from the blood, which is indicative of enhanced phagocytic activity by Kupffer cells (resident hepatic macrophages). Thus. the objective these studies was to determine the involvement of Kupffer cells in cadmium induced liver injury by inhibiting their function with gadolinium chloride (GdCl3). Male Sprague-Dawley rats were administered GdCl3 (10 mg/kg, i.v.) followed 24 h later by a single dose of CdCl2 (3.0 and 4.0 mg/kg, i.v.). Twenty four hours after CdCl2 administration animals were killed and the degree of liver toxicity was assessed using plasma alanine aminotransferase (ALT), as well as light microscopy. Cadmium chloride administration produced multifocal hepatocellular necrosis and increased plasma ALT activity. Pretreatment with GdCl3 significantly reduced both the morphological changes and hepatic ALT release caused by CdCl2. However, the protection was specific to the liver, and did not alter CdCl2 induced testicular injury, as determined by histopathological damage. In many cases, the inducible cadmium-binding protein, metallothionein (MT) is often an essential aspect of the acquisition of cadmium tolerance in the liver. Although cadmium caused a dramatic induction of hepatic MT (32-fold), GdCl3 caused only a minor increase (2-fold). Combined CdCl2 and GdCl3 treatment did not induce levels to an extent greater than CdCl2 alone. As expected, GdCl3 also caused a slight increase in the amount of cadmium associated with the liver. In cultured hepatocytes isolated from GdCl3 pretreated rats, CdCl2 induced cytotoxicity was not significantly altered compared to control hepatocytes, indicating that the mechanism of tolerance required the presence of other cell types. Thus, GdCl3 attenuation of CdCl2 induced hepatotoxicity does not appear to be caused by increased tissue MT content or a decreased susceptibility of hepatocytes to cadmium. From these data, we concluded that tolerance to cadmium induced hepatotoxicity involves the inhibition of Kupffer cell function which resu

Topics: Alanine Transaminase; Animals; Cadmium Chloride; Carcinogens; Cells, Cultured; Chemical and Drug Induced Liver Injury; Disease Models, Animal; Dose-Response Relationship, Drug; Gadolinium; Gout Suppressants; Injections, Intravenous; Kupffer Cells; Lethal Dose 50; Liver; Liver Diseases; Male; Metallothionein; Necrosis; Phagocytosis; Rats; Rats, Sprague-Dawley; Testis; Tissue Distribution

Previously, we found that oral cadmium (Cd) treatment either prevented or substantially reduced N-nitrosodiethylamine (NDEA)-induced tumor formation in B6C3F1 mouse liver or lung regardless of exposure interval and even when the Cd was given well after tumors were formed. Because Cd salts are powerful emetics, oral exposure would probably be impractical in humans. Thus, we studied suppression of NDEA-initiated tumors in male B6C3F1 mice by a single i.v. dose of Cd. NDEA (776 mumol/kg i.p.) was given at time 0 followed by CdCl2 (16 mumol/kg i.v.) 40 weeks later. This dose of Cd had no effect on body weights through the conclusion of the study at 52 weeks. The NDEA-induced increase in hepatic tumor incidence (19 tumor-bearing mice/22 mice at risk, 86%) over control (5/24, 21%) was remarkably reduced by Cd treatment (13/27, 48%, P < or = .05). Multiplicity and size of liver tumors induced by NDEA (2.18 tumors/liver; 31.6 mm3 mean volume) were also substantially reduced by the Cd exposure (0.96 tumors/liver; 17.1 mm3 mean volume). NDEA-induced lung tumor incidence (22/22, 100%) and multiplicity (5.09 tumors/lung) were modestly, but significantly, reduced by Cd treatment (21/27, 78%; 3.89 tumors/lung). Clear evidence of tumor-specific cytotoxicity was observed as Cd treatment induced a necrotizing effect that was localized only within the hepatic tumors. Metallothionein (MT), an inducible metal-binding protein associated with tolerance to many metal including Cd, was not detected immunohistochemically in mouse liver tumors, even those undergoing Cd-induced necrosis, whereas the surrounding normal liver cells expressed high levels of MT after Cd exposure. Likewise, in human hepatocellular carcinomas MT was only poorly or erratically expressed relative to normal tissue. These results indicate that a single, nontoxic dose of Cd dramatically reduces liver tumor burden through tumor cell-specific necrosis due to a down-regulation of MT expression in hepatic tumors of murine origin and furthermore indicate that a similar down-regulation of MT occurs in human hepatocellular carcinomas.

Topics: Animals; Antineoplastic Agents; Cadmium; Carcinoma, Hepatocellular; Diethylnitrosamine; Down-Regulation; Humans; Liver Neoplasms; Male; Metallothionein; Mice; Necrosis; Tumor Necrosis Factor-alpha

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

Topics: Alanine Transaminase; Animals; Cadmium; Cell Survival; Cells, Cultured; Drug Resistance; Female; L-Iditol 2-Dehydrogenase; Liver; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Necrosis; Survival Rate

The molecular and cellular mechanisms responsible for cytotoxic T lymphocyte (CTL)-induced immunopathology are not well defined. Using a model in which hepatitis B surface antigen (HBsAg)-specific CTL cause an acute necroinflammatory liver disease in HBsAg transgenic mice, we demonstrate that class I-restricted disease pathogenesis is an orderly, multistep process that involves direct as well as indirect consequences of CTL activation. It begins (step 1) almost immediately as a direct antigen-specific CTL-target cell interaction that triggers the HBsAg-positive hepatocyte to undergo programmed cell death (apoptosis). It progresses (step 2) within hours to a focal inflammatory response in which antigen-nonspecific lymphocytes and neutrophils amplify the local cytopathic effect of the CTL. The most destructive pathogenetic function of the CTL, however, is to secrete interferon gamma when they encounter antigen in vivo, thereby activating the intrahepatic macrophage and inducing a delayed-type hypersensitivity response (step 3) that destroys the liver and kills the mouse. We propose that the principles illustrated in this study are generally applicable to other models of class I-restricted, CTL-induced immunopathology, and we suggest that they contribute to the immunopathogenesis of viral hepatitis during hepatitis B virus infection in humans.

Topics: Animals; CD3 Complex; CD4 Antigens; CD8 Antigens; Hepatic Encephalopathy; Hepatitis B Surface Antigens; Hepatitis B virus; Histocompatibility Antigens Class I; Immunoenzyme Techniques; Immunotherapy, Adoptive; Inflammation; Interferon-gamma; Liver; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Transgenic; Necrosis; Promoter Regions, Genetic; T-Lymphocyte Subsets; T-Lymphocytes, Cytotoxic; Tumor Necrosis Factor-alpha

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

Topics: Animals; Cadmium; Cadmium Poisoning; Copper; Kidney; Kidney Tubules, Proximal; Liver; Metalloproteins; Metallothionein; Necrosis; Rats; Zinc

Topics: Animals; Cadmium; Female; Liver; Metalloproteins; Metallothionein; Necrosis; Protein Binding; Rats; Rats, Inbred Strains; Zinc

Repeated s.c. injections of cadmium chloride into rats (3 mg Cd/kg body weight, 4 times a week) caused acute and transitory tubular necrosis after 2 to 3 weeks; recovery was observed after 4 weeks despite continued loading. Kidney copper decreased for the first 3 weeks and was slightly increased after 4 weeks. Distribution profiles of cadmium (Cd), zinc (Zn), copper (Cu) on an SW column revealed that elution profiles of kidney metallothionein changed from typical kidney with high copper content to typical liver metallothioneins with tubular necrosis and restoration.

Topics: Animals; Cadmium; Cadmium Poisoning; Copper; Female; Kidney; Kidney Diseases; Metalloproteins; Metallothionein; Necrosis; Rats; Rats, Inbred Strains; Zinc