Role Of Metallothionein Research Articles

The role of metallothionein and selenium in metal detoxification in the liver of deep-sea fish from the NW Mediterranean Sea

Seven deep-sea fish species were sampled in the Blanes Canyon area (NW Mediterranean) at a depth of 1200m during winter. The concentrations of nine metals were determined in the liver of these species by ICP-MS. Furthermore, the metal detoxification potential was determined for each species by analysing the hepatic metallothionein (MT) content, relations between metals and the molar ratio between MT and/or selected metals. The potential effect of metal content on their physiology was assessed using general stress markers such as the enzyme activities of acetylcholinesterase (AChE) and lactate dehydrogenase (LDH) in muscle.Levels of metals in the seven Mediterranean deep-sea fish species studied were intermediate to equivalent species of fish either from Atlantic waters or hydrothermal vents. The metal detoxification potential varied among species depending on MT, selenium (Se) or zinc (Zn) as reliable mechanisms to handle potential metal toxicity. The role of Se was especially relevant when the liver content of mercury (Hg) was higher. AChE and LDH activities did seem to be affected by metal loads and thus the activities reported would correspond to baseline activities of the selected species.

Metal stoichiometry of isolated and arsenic substituted metallothionein: PIXE and ESI-MS study

The stoichiometric analysis of the metal induced Metallothionein (MT) is pertinent for understanding the metal-MT interactions. Despite innumerable publications on MT, the literature addressing these aspects is limited. To bridge this gap, PIXE and ESI-MS analysis of the commercial rabbit liver MT1 (an isoform of MT), zinc induced isolated rat liver MT1, apo and Arsenic substituted rabbit liver MT1 have been carried out. These techniques in combination provide information about number and the signature of all the metal ions bound to MT. By using ESI-MS in the rabbit MT1, ions of Zn n MT1 (n = 0, 1, 4, 5, 6, 7) whereas, in rat MT1, the Zn1MT1 and Zn5MT1 ions are observed. PIXE analysis shows that some copper along with zinc is also present in the rabbit as well as rat MT1 which could not be assessed with ESI-MS. During As metallation reaction with rabbit MT1, with increase in arsenic concentration, the amount of arsenic bound to MT1 also increases, though not proportionally. The presence of both Zn and Cu in MT1 on Zn supplementation can be related to the role of MT in Zn and Cu homeostasis. Further, the presence of partially metallated MT1 suggests that MT1 may donate fractional amount of metal from it's fully metallated form to other proteins where Zn acts as a cofactor.

Metallothionein I+II Expression as an Early Sign of Chronic Relapsing Experimental Autoimmune Encephalomyelitis in Rats

Metallothioneins (MTs) are small, cysteine-rich proteins which have been implicated in various forms of stress providing cytoprotective action against oxidative injury, DNA damage and apoptosis. Owing to their high affinity for physiological metals, such as zinc and copper MTs are also critical components of regulatory proteins involved in cell growth and multiplication, as well as in the maintenance of immune homeostasis. To elucidate the role of MTs in the pathomechanisms of autoimmune CNS disorders we estimated the expression of MT I+II proteins and the content of free Zn ions in the brain, spinal cord and in the liver early in the course of chronic relapsing experimental autoimmune encephalomyelitis (CR-EAE) pathogenesis, i.e. before the onset of any clinical symptoms. Disease was induced in the genetically susceptible Dark Agouti (DA) rats by subcutaneous injection of bovine brain homogenate in CFA. Control animals were treated with CFA alone. The data, obtained by immuno-histochemistry and in situ fluorescent labeling of free zinc ions, have shown that in the presymptomatic phase of CR-EAE (on the seventh postimmunization day) MTs I+II were markedly upregulated in the cells that form blood-brain and blood-cerebrospinal fluid barriers, as well as in the cerebellar parenchyma and hippocampal dentate gyri. Furthermore, we found that the liver also becomes a site of extensive MTs I+II synthesis shortly after immunization. Simultaneously, tissue content of free zinc ions increased at the sites of MTs induction, reflecting their antioxidative activity. The data, described in this paper point to regulatory and neuroprotective role of MTs in the pathogenesis of CR-EAE.

Expression and functional role of metallothioneins I and II in the spinal cord in inflammatory and neuropathic pain models

In this study, the expression and functional role of metallothioneins I and II (MT-I/II) were evaluated in the spinal cord in rat models of inflammatory and neuropathic pain. Complete Freund's adjuvant (CFA) injection into the hindpaw induced an increase in MT-I/II protein expression in bilateral dorsal and ventral horns throughout the spinal cord, while chronic constriction injury (CCI) of the sciatic nerve induced an increase in MT-I/II expression in the ipsilateral dorsal and ventral horns of the lower lumbar spinal cord. Increased MT-I/II immunoreactivity was predominantly localized to vascular endothelial cells. CFA injection- and CCI-induced MT-I/II expression was inhibited by intrathecal administration of MT-I siRNA. Treatment with MT-I siRNA before CFA injection or at early time points after CCI resulted in a significant attenuation of mechanical allodynia and thermal hyperalgesia, while treatment at later time points had no effect on established pain behaviors. Our results suggest that endogenous MT-I/II might play an important role in the pathogenesis of pain behaviors, participating in the initiation of inflammatory and neuropathic pain rather than in their maintenance.

Sa1846 Metallothionein, an Emerging Danger Signal During Experimental Colitis

BACKGROUND: Danger signals have been postulated as regulators of gut mucosal immunity. During intestinal inflammation, the epithelium is compromised and signals, alerting adjacent cells of tissue damage, are released. Hence, we were interested in metallothioneins (MTs), small proteins which have been identified at inflammation sites.We explored triggers releasing MTs from colon epithelial cells and identified their role as extracellular danger signal during experimental colitis. METHODS: HT29 cells were subjected to the following treatments: 200ng/ml LPS, 200 microM H2O2, hypoxia (~1% oxygen), 100ng/ml TNF-alpha + 300ng/ ml IFN-gamma to induce apoptosis, and repeated freeze/thaw cycles to mimic necrosis. Supernatant was analysed for MT levels using western blot and for lactate dehydrogenase activity (LDH). A Boyden transwell migration assay with blood leukocytes was applied to evaluate the chemotactic potential of extracellular MT and the capacity of monoclonal antiMT antibody (100μg/ml UC1MT) to abolish this. The role of MT as chemo-attractant was further explored using dextran sulphate sodium (DSS)-induced colitis in MT knockout (MT-/-), transgenic (MT+/+) and wild type mice (WT). The therapeutical use of monoclonal therapy was tested in DSSand 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis. Inflammatory cell infiltrate was evaluated in all experiments together with standard inflammation markers. RESULTS: Necrosis and TNF-induced apoptosis resulted in detectable MT levels in supernatant of HT29 cells, which was not the case for LPS, H2O2 nor hypoxia treatment. LDH activity was not increased after stimulation with TNF, ruling out an uncontrolled release of MT from TNF-treated cells. Increased leukocyte migration towards this MT-containing supernatant was detected, whereas the addition of UC1MT was able to overcome this chemo-attraction (p,0.05). Significantly less neutrophil infiltration was observed in MT-/mice compared to MT+/+ and WT mice in DSS colitis (p,0.05). Complementary, i.p. UC1MT treatment reduced the number of F4/80-positive macrophages in DSSand TNBS-induced colitis (p,0.05). Less inflammatory infiltrate was associated with reduced histological inflammation in all three colitis experiments. CONCLUSION: We characterized metallothionein as danger signal released from HT29 cells after necrotic and TNF-induced apoptotic cell death. Inhibiting MT function by monoclonal therapy reduces leukocyte infiltration and represents a novel therapy dampening experimental colitis.

Abstract 4004: Interaction between bone marrow-derived mesenchymal stem cells and tumor cells induces metallothionein gene expression in human colon cancer.

Abstract Bone marrow-derived mesenchymal stem cells (MSCs) are reported to contribute to the formation of tumor stroma. We reported recently that in an orthotopic nude mouse model of human colon cancer, MSCs migrated to tumor stroma, where they differentiated into carcinoma-associated fibroblast (CAF)-like cells and promoted tumor growth and metastasis. Furthermore, the migratory and invasive abilities of KM12SM colon cancer cells were shown by Transwell invasion assay to be enhanced by indirect co-culture with MSCs. However, the molecular mechanisms of this phenomenon have not been elucidated. We performed microarray analysis of gene expression in KM12SM cells and found that expression of metallothionein (MT) mRNA increased markedly by indirect co-culture of KM12SM cells with MSCs. Additionally, in an orthotopic nude mouse model of colon cancer, MT mRNA expression levels were shown to be higher in the tumors produced by KM12SM cells mixed with MSCs than in the tumors produced by KM12SM cells alone. MTs comprise a family of low-molecular-weight, cysteine-rich intracellular proteins that are associated with protection against DNA damage, oxidative stress, and apoptosis. The potential role of MTs in carcinogenesis remains unknown. We examined MT expression in surgical specimens of human colon cancer and colon adenoma by immunohistochemistry. A total of 101 tumors were analyzed. MT immunoreactivity was detected in tumor cells and in atypical stromal cells. Atypical stromal cells with strong MT expression were found in invasive cancers, at the invasive edge, but not in mucosal cancers or adenomas. Levels of MT expression in tumor cells correlated inversely with disease stage. These findings suggest that the interaction between MSCs recruited into tumor stroma and the tumor cells of invasive colon cancers induces and depends on MT expression. Further studies are needed to clarify the specific effects of MTs on tumor cells and stromal cells. Citation Format: Kei Shinagawa, Yasuhiko Kitadai, Ryo Yuge, Mieko Onoyama, Mayu Ohnishi, Shinji Tanaka, Wataru Yasui, Kazuaki Chayama. Interaction between bone marrow-derived mesenchymal stem cells and tumor cells induces metallothionein gene expression in human colon cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4004. doi:10.1158/1538-7445.AM2013-4004

Protective Role of Metallothionein in Chemical and Radiation Carcinogenesis

Metallothionein (MT) is a low molecular weight metal-binding protein induced by endogenous and exogenous stimuli such as cytokines and heavy metals. In 1993 and 1994, two research groups (Choo et al. and Palmiter et al., respectively) produced MT-I/II double-knockout mice (MT-I/II null mice) with null mutations of the MT-I and MT-II genes. Subsequently, MT-I/II null mice have been used to clarify the biological function, physiological role, and pathophysiological relevance of MT by many research groups. Recent studies using MT-I/II null mice to investigate the role of MT in metal toxicity and distribution, oxidative stress, and some disease were reviewed. In addition, several research groups including our laboratory have reported that MT-I/II null mice are highly susceptible to several carcinogenesis caused by 7,12-dimethylbenz[a]anthracene, X-ray, benzo[a]pyrene, N-butyl-N-(4-hydroxybutyl) nitrosamine, lead, and cisplatin. These results suggest that MT is an important protective factor against not only metal toxicity and oxidative stress but also chemical and radiation carcinogenesis. In this review, we present the findings of MT-I/II null mice with regard to the protective role of MT in carcinogenesis and mutagenesis caused by chemical agents and X-ray.

Metallothionein as a Negative Regulator of Pulmonary Inflammation

The integration of knowledge concerning the regulation of metallothionein (MT) with research on its proposed functions is necessary to clarify how MT affects cellular processes. MT expression is induced/enhanced in various tissues by a number of physiological mediators through several response elements in the MT gene promoter. The cellular accumulation of MT depends on the availability of cellular zinc derived from the diet. MT modulates: 1) the binding and exchange/ transport of heavy metals such as zinc, cadmium, or copper under physiological conditions and cytoprotection from their toxicities, and 2) the release of gaseous mediators such as hydroxyl radicals or nitric oxide. In addition, MT reportedly affects a number of cellular processes, such as gene expression, apoptosis, proliferation, and differentiation. Given the genetic approach, the apparently healthy status of MT-deficient mice argues against an essential biological role for MT; however, the molecule may be critical in cells/tissues/organs in times of stress, since MT expression is also evoked/enhanced by various stresses. In particular, because metallothionein (MT) is induced by inflammatory stress, its roles in inflammation are implied. Also, MT expression in the lung can be enhanced by inflammatory stimuli, suggesting that its expression correlates with inflammatory pulmonary diseases. In this paper, we review the role of MT of various inflammatory conditions in the airway.

Obesity and Metallothionein

The development of obesity consists of 3 distinct processes: differentiation of new adipocytes from precursor preadipocytic cells, formation of mature adipocytes, and excess accumulation of lipids in the adipocytes resulting in cell enlargement. This review aims to discuss the possible role of metallothionein (MT) in adipose tissue in the development of obesity. MT is induced in adipose tissue during the development of obesity. In addition, MT plays a preventive role against obesity in female mice. Although the detailed mechanism is presently unknown, one of the factors involved is the mesoderm-specific transcript (MEST/Peg1), adipocyte enlargement factor. MT has a potential to prevent obesity-related diseases, at least in part, through suppression of disease-induced generation of superoxide and endoplasmic reticulum stress and associated damage. Studies on the development of a MT inducer will encourage potential clinical use for obesity- related diseases.

Zinc protects against diabetes-induced pathogenic changes in the aorta: roles of metallothionein and nuclear factor (erythroid-derived 2)-like 2

BackgroundCardiovascular diseases remain a leading cause of the mortality world-wide, which is related to several risks, including the life style change and the increased diabetes prevalence. The present study was to explore the preventive effect of zinc on the pathogenic changes in the aorta.MethodsA genetic type 1 diabetic OVE26 mouse model was used with/without zinc supplementation for 3 months. To determine gender difference either for pathogenic changes in the aorta of diabetic mice or for zinc protective effects on diabetes-induced pathogenic changes, both males and females were investigated in parallel by histopathological and immunohistochemical examinations, in combination of real-time PCR assay.ResultsDiabetes induced significant increases in aortic oxidative damage, inflammation, and remodeling (increased fibrosis and wall thickness) without significant difference between genders. Zinc treatment of these diabetic mice for three months completely prevented the above pathogenic changes in the aorta, and also significantly up-regulated the expression and function of nuclear factor (erythroid-derived 2)-like 2 (Nrf2), a pivotal regulator of anti-oxidative mechanism, and the expression of metallothionein (MT), a potent antioxidant. There was gender difference for the protective effect of zinc against diabetes-induced pathogenic changes and the up-regulated levels of Nrf2 and MT in the aorta.ConclusionsThese results suggest that zinc supplementation provides a significant protection against diabetes-induced pathogenic changes in the aorta without gender difference in the type 1 diabetic mouse model. The aortic protection by zinc against diabetes-induced pathogenic changes is associated with the up-regulation of both MT and Nrf2 expression.

The role of metallothionein in oxidative stress.

Free radicals are chemical particles containing one or more unpaired electrons, which may be part of the molecule. They cause the molecule to become highly reactive. The free radicals are also known to play a dual role in biological systems, as they can be either beneficial or harmful for living systems. It is clear that there are numerous mechanisms participating on the protection of a cell against free radicals. In this review, our attention is paid to metallothioneins (MTs) as small, cysteine-rich and heavy metal-binding proteins, which participate in an array of protective stress responses. The mechanism of the reaction of metallothioneins with oxidants and electrophilic compounds is discussed. Numerous reports indicate that MT protects cells from exposure to oxidants and electrophiles, which react readily with sulfhydryl groups. Moreover, MT plays a key role in regulation of zinc levels and distribution in the intracellular space. The connections between zinc, MT and cancer are highlighted.

Role of metallothionein in murine experimental colitis

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.

Tissue-specificity and phylogenetics of Pl-MT mRNA during Paracentrotus lividus embryogenesis

Metallothioneins (MTs) constitute a family of cysteine-rich, low molecular weight proteins, which generally provide protection against metal toxicity and oxidative stress counteracting the cell damage caused by essential and non-essential heavy metals. Equally important is the physiological role of MTs in the homeostasis of essential metals, which are involved in a wide variety of cellular processes. The aim of this work was to investigate the expression and the territorial localization of Paracentrotus lividus MT (Pl-MT) mRNA during sea urchin development by Quantitative Polymerase Chain Reaction (QPCR) and Whole Mount In Situ Hybridization (WMISH), as well as the phylogenetic comparison with selected MT homologs present in different phyla. We found that Pl-MT mRNA is accumulated in unfertilized eggs and constitutively expressed during development, with very low levels of maternal mRNA at cleavage stages, followed by a significant rise during gastrulation with a peak at the prism stage. Pl-MT mRNA was expressed in the vegetative plate at mesenchyme blastula, later restricted to the endoderm of gastrula embryos and finally to the gut of plutei. Indirect immunofluorescence (IF) using a specific antibody for the endoderm marker Endo1 demonstrated a co-localization with the Pl-MT transcripts in the midgut and hindgut after the intestine differentiation occurs and when larval feeding begins. Our results show for the first time the constitutive temporal and tissue-specific expression of MT in P. lividus embryos, providing new information for studies on the mechanisms controlling basal and induced MT gene expression.The analysis of the phylogenetic relationship of Pl-MT with homologs from different phyla, ranging from yeast to vertebrates, suggests the evolutionary process of these proteins, which could have been selected not only on the basis of their ability to bind metals but also by their tissue-specificity.

P077 Emerging role of the interleukin (IL)-33/ST2 axis in gut mucosal wound healing and epithelial repair

BACKGROUND: Danger signals have been postulated as regulators of gut mucosal immunity. During intestinal inflammation, the epithelium is compromised and signals, alerting adjacent cells of tissue damage, are released. Hence, we were interested in metallothioneins (MTs), small proteins which have been identified at inflammation sites.We explored triggers releasing MTs from colon epithelial cells and identified their role as extracellular danger signal during experimental colitis. METHODS: HT29 cells were subjected to the following treatments: 200ng/ml LPS, 200 microM H2O2, hypoxia (~1% oxygen), 100ng/ml TNF-alpha + 300ng/ ml IFN-gamma to induce apoptosis, and repeated freeze/thaw cycles to mimic necrosis. Supernatant was analysed for MT levels using western blot and for lactate dehydrogenase activity (LDH). A Boyden transwell migration assay with blood leukocytes was applied to evaluate the chemotactic potential of extracellular MT and the capacity of monoclonal antiMT antibody (100μg/ml UC1MT) to abolish this. The role of MT as chemo-attractant was further explored using dextran sulphate sodium (DSS)-induced colitis in MT knockout (MT-/-), transgenic (MT+/+) and wild type mice (WT). The therapeutical use of monoclonal therapy was tested in DSSand 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis. Inflammatory cell infiltrate was evaluated in all experiments together with standard inflammation markers. RESULTS: Necrosis and TNF-induced apoptosis resulted in detectable MT levels in supernatant of HT29 cells, which was not the case for LPS, H2O2 nor hypoxia treatment. LDH activity was not increased after stimulation with TNF, ruling out an uncontrolled release of MT from TNF-treated cells. Increased leukocyte migration towards this MT-containing supernatant was detected, whereas the addition of UC1MT was able to overcome this chemo-attraction (p,0.05). Significantly less neutrophil infiltration was observed in MT-/mice compared to MT+/+ and WT mice in DSS colitis (p,0.05). Complementary, i.p. UC1MT treatment reduced the number of F4/80-positive macrophages in DSSand TNBS-induced colitis (p,0.05). Less inflammatory infiltrate was associated with reduced histological inflammation in all three colitis experiments. CONCLUSION: We characterized metallothionein as danger signal released from HT29 cells after necrotic and TNF-induced apoptotic cell death. Inhibiting MT function by monoclonal therapy reduces leukocyte infiltration and represents a novel therapy dampening experimental colitis.

Oxidation reactivity of zinc–cysteine clusters in metallothionein

Evaluating the reactivity of the metal-thiolate clusters in metallothionein (MT) is a key step in understanding the biological functions of this protein. The effects of the metal clustering and protein environment on the thiolate reactivity with hydrogen peroxide (H(2)O(2)) were investigated by performing quantum theory calculations with chemical accuracy at two levels of complexity. At the first level, the reactivity with H(2)O(2) of a model system ([(Zn)(3)(MeS)(9)](3-), MeS is methanethiolate) of the β domain cluster of MT was evaluated using density functional theory (DFT) with the mPW1PW91 functional. At the second level of complexity, the protein environment was included in the reactant system and the calculations were performed with the hybrid ONIOM method combining the DFT-mPW1PW91 and the semiempirical PM6 levels of theory. In these conditions, the energy barrier for the oxidation of the most reactive terminal thiolate was 21.5kcalmol(-1). This is 3kcalmol(-1) higher than that calculated for the terminal thiolate in the model system [(Zn)(3)(MeS)(9)](3-) and about 7kcalmol(-1) higher than that obtained for the free thiolate. In spite of this rise of the energy barrier induced by the protein environment, the thiolate oxidation by H(2)O(2) is confirmed as a possible way for metal release from MT. On the other hand, the results suggest that the antioxidant role of MT in the living cell cannot be as important as that of glutathione (which bears a free thiol).

Systemic and organ specific metabolic variation in metallothionein knockout mice challenged with swimming exercise

Metallothioneins (MTs) are ubiquitous, multi-functional proteins with key roles in metal homeostasis and redox regulation. Their involvement in cellular energy metabolism is evident from the observation that metallothionein-knockout (MTKO) mice become moderately obese. Transcriptomic studies have also indicated that several genes associated with energy metabolism are differentially expressed in these mice. Although single varying metabolites have been reported, the roles of MTs on a global metabolic level have not been investigated before. In this study, an untargeted, hypothesis-generating metabolomics approach was used to identify and report all metabolites that differ in relative concentration between MT1+2KO, MT3KO and wild-type (WT) mice before and after an exercise (1 h swim) perturbation. PCA and univariate results confirmed that the metabolism of the MTKO mice differs from the WT during unchallenged conditions and hypothetically pointed to increased anabolic activity which could contribute to their previously reported tendency to become obese. Furthermore, the metabolic differences observed in the liver after the 1 h swim indicated that catabolic activity might be impaired in these mice, which could be a consequence of a dysfunction of a common catabolic signal. The MT3KO mice did not show the same metabolic pattern as the MT1+2KO mice as most metabolite concentrations in the brain of these mice were lower after the 1 h swim. The reported metabolic variation contributes to better understand the diverse functionality of these ubiquitous proteins on a global phenotypic level.

Metallothionein blocks oxidative DNA damage in vitro

The role of metallothionein (MT) in mitigation of oxidative DNA damage (ODD) induced by either cadmium (Cd) or the direct oxidant hydrogen peroxide (H(2)O(2)) was systematically examined using MT-I/II double knockout (MT-null) or MT-competent wild-type (WT) cells. Both toxicants were much more lethal to MT-null cells (Cd LC(50) = 6.6 μM; H(2)O(2) LC(50) = 550 μM) than to WT cells (Cd LC(50) = 16.5 μM; H(2)O(2) LC(50) = 930 μM). Cd induced concentration-related MT increases in WT cells, while the basal levels were undetectable and not increased by Cd in MT-null cells. ODD, measured by the immuno-spin trapping method, was minimally induced by sub-toxic Cd levels (1 or 5 μM; 24 h) in WT cells, but markedly increased in MT-null cells (>430 %). Similarly, ODD was induced to higher levels by lower concentrations of H(2)O(2) in MT-null cells than WT cells. Transfection of MT-I into MT-null cells reduced both Cd- and H(2)O(2)-induced cytolethality and ODD. Cd increased the expression of the oxidant defense genes, HO-1, and GSTa2 to a much greater extent in MT-null cells than in WT. Cd or H(2)O(2) exposure increased the expression of key transport genes, Mrp1 and Mrp2, in WT cells but not in MT-null cells. MT protects against Cd- and H(2)O(2)-induced ODD in MT-competent cells possibly by multiple mechanisms, potentially including direct metal ion sequestration and sequestration of oxidant radicals by MT. MT-deficient cells appear to adapt to Cd primarily by turning on oxidant response systems, while MT-competent cells activate MT and transport systems.

Are metallothioneins equally good biomarkers of metal and oxidative stress?

Several researchers investigated the induction of metallothioneins (MTs) in the presence of metals, namely Cadmium (Cd). Fewer studies observed the induction of MTs due to oxidizing agents, and literature comparing the sensitivity of MTs to different stressors is even more scarce or even nonexistent. The role of MTs in metal and oxidative stress and thus their use as a stress biomarker, remains to be clearly elucidated. To better understand the role of MTs as a biomarker in Cerastoderma edule, a bivalve widely used as bioindicator, a laboratory assay was conducted aiming to assess the sensitivity of MTs to metal and oxidative stressors. For this purpose, Cd was used to induce metal stress, whereas hydrogen peroxide (H2O2), being an oxidizing compound, was used to impose oxidative stress. Results showed that induction of MTs occurred at very different levels in metal and oxidative stress. In the presence of the oxidizing agent (H2O2), MTs only increased significantly when the degree of oxidative stress was very high, and mortality rates were higher than 50 percent. On the contrary, C. edule survived to all Cd concentrations used and significant MTs increases, compared to the control, were observed in all Cd exposures. The present work also revealed that the number of ions and the metal bound to MTs varied with the exposure conditions. In the absence of disturbance, MTs bound most (60–70 percent) of the essential metals (Zn and Cu) in solution. In stressful situations, such as the exposure to Cd and H2O2, MTs did not bind to Cu and bound less to Zn. When organisms were exposed to Cd, the total number of ions bound per MT molecule did not change, compared to control. However the sort of ions bound per MT molecule differed; part of the Zn and all Cu ions where displaced by Cd ions. For organisms exposed to H2O2, each MT molecule bound less than half of the ions compared to control and Cd conditions, which indicates a partial oxidation of thiol groups in the cysteine residues through ROS scavenging. The present results suggest that MTs are excellent markers of metal stress, but not of oxidative stress.

Molecular functions of metallothionein and its role in hematological malignancies

Metallothionein (MT) was reported to be a potential negative regulator of apoptosis, and various reports have suggested that it may play roles in carcinogenesis and drug resistance, in at least a portion of cancer cells. The author summarizes the current understanding of the molecular functions of MT for tumor cell growth and drug resistance. These activities are regulated through intracellular metal ion modulation and free radical scavenging. Compared with analyses of solid tumors, few studies have analyzed the roles of MT in hematological malignancies. This review mainly describes the functions of MT in hematopoietic cells. Furthermore, through expression analyses of leukemias and lymphomas, the roles of MT in the biology of these diseases are particularly focused upon.

Correlation among metallothionein expression, intratumoural macrophage infiltration and the risk of metastasis in human cutaneous malignant melanoma

The formation of metastases and the efficacy of systemic therapies in cutaneous malignant melanoma (CMM) depend on the characteristics of the tumour cells and the host immune response. Aberrant expression of metallothionein (MT) has been observed in several types of cancers with poor prognoses. To perform an immunohistochemical study on primary CMM comparing the MT expression of tumours without metastases (n = 23) to that of samples with haematogenous metastases (n = 23) and to examine the correlation between MT staining and immunological markers relevant in CMM progression. The immunohistochemical labelling of different tumour sections was analysed using tissue microarrays for the evaluation of the suitability of this method in future studies. Our results suggest that MT overexpression is significantly more frequent in primary CMM with haematogenous metastases (P = 0.018) and that the overexpression is independent of the Breslow tumour thickness (R = 0.102, P = 0.501). Interestingly, MT overexpression of the tumour cells was correlated with the presence of tumour-infiltrating CD68(+) macrophages (P = 0.003), a known predictive factor for melanoma progression, thereby suggesting a role for MT in the development of a defective host immune response. Furthermore, the presence of CD163(+) macrophages infiltrating the tumours correlated with metastasis formation (P < 0.001), whereas the presence CD1a(+) dendritic cells surrounding the tumours was associated with a lower risk of haematogenous spread (P = 0.003). Our results demonstrate that MT may represent a suitable prognostic factor that can characterize the metastasising ability of CMM and the tumour-promoting host immune response.