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Arsenic trioxide (AsO; ATO, TRISENOX®) is used to treat patients with refractory or relapsed acute promyelocytic leukaemia while its application for treatment of solid cancers like glioblastoma is still under evaluation. In the present study, we investigated the interaction of arsenic trioxide with metallothionein (MT) isoforms as a possible (protective response) resistance of glioblastoma cells to arsenic-induced cytotoxicity. Special attention was focused on MT3, the isoform expressed mainly in the brain. MT3 has low metal inducibility, fast metal binding/releasing properties and outstanding neuronal inhibitory activity. The human astrocytoma (glioblastoma) cell line U87 MG was treated with 0.6, 2 and 6-7 μM arsenic (equivalent to 0.3, 1 and 3-3.5 μM AsO) for 12, 24 or 48 h and gene expression for different MT isoforms, namely MT2A, MT1A, MT1F, MT1X, MT1E and MT3, was measured by real time qPCR using SYBR Green I and Taqman® gene expression assays. TfR, 18S rRNA, GAPDH and AB were tested as reference genes, and the last two evaluated to be appropriate in conditions of low ( GAPDH) and high ( AB) arsenic exposure. The gene expression of MT3 gene was additionally tested and confirmed by restriction enzyme analysis with PvuII. In the given conditions the mRNAs of six MT isoforms were identified in human glioblastoma cell line U87 MG. Depending on arsenic exposure conditions, an increase or decrease of MT gene expression was observed for each isoform, with the highest increase for isoforms MT1X, MT1F and MT2A mRNA (up to 13-fold) and more persistent decreases for MT1A, MT1E and MT3 mRNA. Despite the common assumption of the noninducibility of MT3, the evident MT3 mRNA increase was observed during high As exposure (up to 4-fold). In conclusion, our results clearly demonstrate the influence of As on MT isoform gene expression. The MT1X, MT1F and MT2A increase could represent brain tumour acquired resistance to As cytotoxicity while the MT3 increase is more enigmatic, with its possible involvement in arsenic-related induction of type II cell death. [ABSTRACT FROM AUTHOR]

Abstract: Common silver barb, Puntius gonionotus, exposed to the nominal concentration of 0.06 mg/L Cd for 60 d, were assessed for histopathological alterations (gills, liver and kidney), metal accumulation, and metallothionein (MT) mRNA expression. Fish exhibited pathological symptoms such as hypertrophy and hyperplasia of primary and secondary gill lamellae, vacuolization in hepatocytes, and prominent tubular and glomerular damage in the kidney. In addition, kidney accumulated the highest content of cadmium, more than gills and liver. Expression of MT mRNA was increased in both liver and kidney of treated fish. Hepatic MT levels remained high after fish were removed to Cd-free water. In contrast, MT expression in kidney was peaked after 28 d of treatment and drastically dropped when fish were removed to Cd-free water. The high concentrations of Cd in hepatic tissues indicated an accumulation site or permanent damage on this tissue. [Copyright &y& Elsevier]

The aim of the study was to determine the effect of weaning and the effect of increasing dietary zinc concentrations on the zinc and copper status of weaned piglets (study 1) and to study the effect of high concentrations of dietary zinc and/or copper on zinc and copper status of weaned piglets (study 2). Study 1 included 54 piglets (six litters of nine piglets). One piglet from every litter was killed 1 day before weaning. The remaining 48 piglets were allocated at weaning (28 days) to four dietary zinc treatments (100, 250, 1000 or 2500 ppm) and subsequently killed 1–2, 5–6 or 14–15 days after weaning. Study 2 included 48 piglets (six litters of eight piglets) allocated to four dietary treatments, consisting of low or high dietary zinc (100 or 2500 ppm) in combination with low or high dietary copper (20 or 175 ppm). All piglets in study 2 were killed 5–7 days after weaning. In both studies, the trace mineral status was assessed by zinc and copper concentrations and alkaline phosphatase (AP) activity in plasma and mucosal tissue. In study 2, lymphocyte metallothionein (MT) mRNA and intestinal mucosa MT mRNA concentrations were included as zinc status markers. The results showed that the zinc status, measured as zinc in plasma and mucosa, was not affected by weaning of the piglets. Plasma copper concentrations decreased during the first 2 weeks after weaning. High dietary copper concentrations did not affect the concentration of copper in plasma, but increased the concentration of copper in mucosa and the concentration of zinc in plasma. The dietary zinc treatments increased the zinc concentration in plasma as well as the zinc and MT mRNA concentration in mucosa. Lymphocyte MT mRNA concentrations did not reflect the differences in dietary zinc supplementation. [ABSTRACT FROM AUTHOR]

The relationship between metallothionein mRNA levels and the amounts of copper and zinc in liver, kidney and small intestine by feeding dietary cyclodextrin was examined in growing Wistar rats. α-, β- or γ-cyclodextrin was fed at 50 g/kg of diet for a 7-days period (ad libitum). After feeding, the liver zinc of rats fed β-cyclodextrin was greater than those of rats fed the other three diets. Copper accumulated in kidney of rats fed α- or β-cyclodextrin. Copper content in the small intestine did not show any alterations among rats fed all kinds of diets. The cyclodextrin-supplemented diets were ineffective in zinc content in every organ. There was the greatest level of copper in serum of rats fed β-cyclodextrin, whereas the highest level of serum zinc was observed in rats fed γ-cyclodextrin diet. Northern blot analysis demonstrated that dietary β- and γ-cyclodextrins, but not α-cyclodextrin markedly increased the metallothionein mRNA in the liver, whereas small intestinal metallothionein mRNA levels were markedly decreased. Kidney metallothionien mRNA levels were raised appreciably by all dietary cyclodextrin intakes. Metallothionein gene expressions in liver, kidney and small intestine were not proportional to liver and serum copper or zinc levels in those tissues. These results suggest that regulation of the metallothionein mRNA levels may at least partly involved with the accumulation of metals as copper in liver and kidney of rats fed cyclodextrins. [Copyright &y& Elsevier]

The objective of this study was to assess the effects of Cd and Zn exposure of rainbow trout (Oncorhynchus mykiss) on (a) hepatic glutathione (GSH) levels; and (b) hepatic and branchial metallothionein (MT) mRNA expression. Juvenile rainbow trout were exposed to waterborne Cd (nominal concentrations: 1.5 or 10 μg Cd l−1), Zn (150 or 1000 μg Zn l−1) or Cd/Zn mixtures (1.5 μg Cd l−1 with 200 μg Zn l−1 or 10 μg Cd l−1 with 1000 μg Zn l−1). After 14 and 28 days of treatment, hepatic concentrations of total glutathione, oxidized glutathione (GSSG) and cysteine were determined by means of fluorometric high performance liquid chromatography (HPLC). Branchial and hepatic expression of MT mRNA was measured by means of semi-quantitative RT–PCR. Exposure of trout to Zn did not result in significantly elevated tissue levels of Zn, whereas Cd accumulation factors changed significantly with time and concentration. Despite of the absence of Zn accumulation, hepatic GSH but not MT mRNA levels were significantly altered in Zn-exposed fish. Cd, on the contrary, affected mainly the MT response but not GSH. Also tissue specific differences in the regulation of the two thiol pools were expressed. The thiol response after exposure to metal mixtures could not be explained by simple addition of the effects of the individual metals. The results indicate that cellular thiol pools show different reaction patterns with respect to specific metals and metal mixtures. Under conditions of long-term, low dose metal exposure, the function of GSH appears to go beyond that of a transitory, first line defense. [Copyright &y& Elsevier]

Metallothioneins (MTs), a group of cysteine-rich proteins with a small molecular mass, are known to have metalloregulatory functions. MT gene expression has been demonstrated to be cell type-specific and differentially regulated (possibly related to their germ layer origin and different functional states). In vitro studies suggest that MT-2A, MT-IE, and MT-1F isoforms may be related to breast cancer. In this study, data on MT-2A, MT-1E, MT-1F mRNA analysis via reverse transcription-polymerase chain reaction in invasive ductal breast cancer tissues and their adjacent benign breast tissues from 27 mastectomies are presented. Expression of mRNA in all the three MT isoforms was detected in both cancerous and adjacent benign breast tissues (with MT-2A mRNA expression being the highest). MT-1F expression was significantly higher in benign breast tissues compared with the breast cancers (P=0.017). In situ hybridization confirmed the expression of MT-2A mRNA in the myoepithelial cells of the breast tissues. Immunohistochemical localization of the MT protein revealed that myoepithelial cells consistently expressed the MT protein, while the cancer cells expressed MT with great variation. Based on our immunohistochemical and mRNA analysis, it is likely that the three MT isoforms are specifically expressed in myoepithelial cells of benign breast tissues and cancer cells of the invasive ductal breast cancer tissues. As MT expression occurs in myoepithelial cells and ductal breast cancer cells, our finding supports the proposition that loss of myoepithelial cells in invasive mammary cancers may be compensated in part by changes in the tumor cells, which may subsequently be the basis for studying the role of MT in breast physiology and carcinogenesis. Differential MT-1F expression in breast myoepithelial cells warrants further study. [ABSTRACT FROM AUTHOR]