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Arbuscular mycorrhizal fungi are the most widespread root fungal symbionts, forming associations with the vast majority of plant species. Ectomycorrhizal development alters gene expression in plant symbionts. In this work we examined the effect of arbuscular mycorrhizal fungi spores on the growth and development of Brassica and on the expression of BnMT2 in winter rape. In a pot experiment, rape seedlings growing on different types of sterile and nonsterile soils were inoculated simultaneously with mycorrhizal fungi spores of Acaulospora longula, Glomus geosporum, Glomus mosseae and Scutellospora calospora. As compared with control plants growing in the absence of spores, ten-week-old seedlings of Brassica napus L. in sterile soil inoculated with arbuscular spores had longer shoots and higher fresh biomass of above-ground plant parts. In other types of substrates enriched with mycorrhizal fungi spores, the plants were smaller than non-inoculated plants. The presence of AMF spores stimulated the elongation growth of hypocotyls in both analyzed substrates. BnMT2 expression was highest in plants growing on the sterile substrate. Generally, the presence of mycorrhizal fungi spores appeared to have an adverse effect on the growth of rape plants.
Overexpression of SGTP and/or MT may contribute to various carcinogenic processes and to resistance to anticancer treatment. The importance of these proteins, although clearly established in solid tumours, has not been fully understood in haematopoietic neoplasm. The aim of this study was to determine the expression of MT and SGTP in the bone marrow of patients with MPD. Twenty paraffin-embedded bone marrow core biopsy specimens from newly diagnosed patients with MPD were evaluated — osteomyelofibrosis (OMF), n = 9 and chronic myelocytic leukaemia (CML), n = 11. We demonstrate increased SGTP and MT expression in the bone marrow of MPD patients. In our study levels of MT in OMF patients were higher than in CML. This suggests that MT expression may correlate with bone marrow fibrosis. These data, although based on a relatively small number of patients, raise the possibility that SGTP and MT may play a role in the pathogenesis of MPD. The clinical significance of this phenomenon needs further investigation.
Metallothioneins are low-molecular-weight proteins capable of covalently binding heavy metal ions due to the presence of many cysteine residues in their sequences. We analyzed the predicted amino acid sequences of 19 metallothionein (7 from Arabidopsis thaliana and 12 from Oryza sativa) and their promoter sequences in silico in order to determine the potential regulatory cis-elements present in the promoters of metallothionein genes, from which it is possible to determine the putative functions of these genes. The PlantCARE and PLACE databases provided information about the putative regulatory elements in the metallothionein promoters. Metal response element sequences were found in the promoters of eleven O. sativa and two Arabidopsis metallothionein genes. Copper response elements were identified in both model plants, usually in many copies, particularly in O. sativa. Both the high cysteine content and the presence of metal response motifs in the promoters support the suggestion that metallothioneins play a key role in metal detoxification. The most common putative element in the analyzed promoters was CIRCADIAN, which was present in five A. thaliana and eight O. sativa sequences. The methyl jasmonate response sequence, root-specific expression element and drought response element were found only in O. sativa metallothioneins. Light and low temperature response elements, biotic and abiotic stress elements, an abscisic acid-responsive element and an ethylene-responsive element occur in selected metallothionein promoters of both species. A few promoters have putative organ- and cell-specific regulatory elements. The presence of many different motifs in the promoters of the Arabidopsis and O. sativa genes implies that metallothioneins are general stress response proteins with many important functions in plants, including regulation of their normal development and adaptation to changing environmental conditions.
The aim of this study was to examine the influence of oral intoxication with cadmium and ethanol separately or together on zinc concentrations in liver and kidney of rats. Cadmium was administered at a concentration of 50 mg Cd/dm3 in drinking water for 8 weeks. Ethanol was given in a dose of 5 g/kg of body weight (as a 25% solution) every 12 h for the last 108 h of the experiment. The conducted study has shown that intoxication with cadmium and ethanol alone as well as in combination leads to an increase in zinc concentrations in the liver and kidney. The elevation of zinc levels is highest when the rats are co-exposed to cadmium and ethanol. The deposition of zinc in the liver and kidney of cadmium intoxicated rats is associated with cadmium accumulation and induction of metallothionein synthesis in these organs. Ethanol is also able to induce metallothionein synthesis and an elevation of zinc concentrations in the liver and kidney. As a result, in conditions of co-exposure these effects sum up.
The aim of this study was to present a new analytical method for the quantitative determination of metallothioneins (MT) protein in human body fluids and tissues, in order to determine the level of environmental and industrial exposition to heavy metals. For MTs isolation covalent affinity chromatography with thiol-di- sulphide interchange (CAC-TDI) was applied, which is a modern technique of separation of a high affinity, good repeatability and reproducibility, allowing specific isolation of the thiolproteins and metallothiolproteins. Fundamentals of indirect determination of the contents of metallothioneins protein were worked out throught estimation of the quantities of metals bound with metallothionein protein and adsorbed on covalent affinity chromatography gel as on solid-phase extraction support during a separating process. The (CAC-TDI) gel, specially prepared, was used as a solid phase extraction support (SPE) for preconcent- ration of Hg-Thionein (Hg-Th), Cd-thionein (Cd-Th), Zn-thionein (Zn-Th) and Cu-thionein (Cu-Th) proteins and Hg, Cd, Zn and Cu bonded with MTs from spiked water, urine, human plasma, breast milk and tissues' homogenates.
 Metallothionein (MT) is a low molecular weight cysteine-rich protein with a number of roles in the pro/antioxidant balance and homeostasis of essential metals, such as zinc and copper, and in the detoxification of heavy metals, such as cadmium and mercury. Until now, detection of metallothionein in biological fluids remained difficult because of a lack of a broadly reactive commercial test. Meaningful comparison of the values of metallothionein concentrations reported by different authors using their specific isolation procedures and different conditions of enzyme-linked immunoassay is difficult due to the absence of a reference material for metallothionein. Therefore in the present study, we describe a quantitative assay for metallothionein in biological fluids such as plasma and urine performed by a direct enzyme-linked immunoassay using a commercially; available monoclonal mouse anti-metallothionein clone E9 antibody and commercial standards of metallothionein from rabbit liver and a custom preparation of metallothionein from human liver. The sensitivity of the assay for the standard containing two isoforms MT-I and MT-II from human liver was 140 pg/well. The reactivity of the commercial standards and standards containing two isoforms MT-I and MT-II isolated from human liver in our laboratory with a commercial monoclonal mouse anti-metallothionein clone E9 antibody were similar. This suggests that the described ELISA test can be useful for determination of metallothionein concentration in biological fluids. The concentrations of metallothionein in human plasma, erythrocyte lysate and in urine of smoking and non-smoking healthy volunteers are reported. Tobacco smoking increases the extracellular metallothionein concentration (plasma and urine) but does not affect the intracellular concentration (erythrocyte lysate).
The localizations of metallothionein I and II (MT), a small molecular weight heavy metal binding proteins, and 70-kDa heat shock protein (Hsp70) were investigated by immunohistochemical techniques in brains of lambs that had been injured by congenital copper deficiency. The results were compared with those obtained from control lambs. The morphological findings of the congenital copper deficiency in the central nervous system (CNS) were recorded. The amount of copper in the brain and liver of the lambs and feed of breeding ewes and soil was also assayed by atomic absorption spectrophotometry. The amount of copper in the brain, liver, soil and feed were low. Immunohistochemically, MT and Hsp70 expressions were found to be markedly increased in the CNS of congenital copper deficient lambs compared with control lambs. MT immunoreactivity was prominently found in the astrocytes while strong Hsp70 labelling was in both astrocytes and neurons in the cerebrum, cerebellum, thalamus/hypothalamus and medulla oblongata. Immunohistochemical labelling for both MT and Hsp70 was also seen in the pia mater, ependymal cells and choroid plexi. Present results suggest that the elevated expressions of MT and Hsp70 in astrocytes and neurons are possibly indicating that they are less susceptible to the consequences of cell stress factors and could be exploited to increase selectively their survival in copper deficiency.
The aim of this study was to test a new method for the quantitative determination of metallo- thionein (MT), in order to determine the level of exposure to heavy metals. For MT preconcentration, the solid phase extraction (SPE) method with covalent affinity chromatography with thiol-disulphide interchange (CAC-TDI) gel was applied. In experiment Cd-thionein (Cd-Th) obtained from rats exposed to cadmium salt was used. Protein concentration was determined by indirect method based on metal analysis, bonded with MT protein by atomic absorption spectrometry (AAS). The results showed satisfactory correlation between Cd-Th concentration and the level of exposition for heavy metals.
Tumour growth and expansion are the result of proliferative activity and the capacity to eliminate cells by apoptosis and/or necrosis. The present study was aimed at comparing the apoptosis and proliferation intensity in cells of adenocarcinomas of the large intestine with the expression of metallothionein (MT), the grade of the tumour and the depth to which the tumour infiltrated the intestinal wall. The TUNEL technique and immunocytochemical reactions (expression of caspase-3, Ki-67, MT) were used to detect apoptosis. The results demonstrated augmented levels of all the variables examined, positively correlated with grade of malignancy, G, and with the depth of intestinal wall infiltration by the tumour cells. The testing of apoptosis, proliferation and MT expression may prove useful in the appraisal of the growth and progression of primary adenocarcinomas in the large intestine.
Metalotioneiny (MTs) stanowią rodzinę białek niskocząsteczkowych, które bogate są w reszty cysteinowe i posiadają zdolność do wiązania metali ciężkich. Białka te uczestniczą w utrzymaniu homeostazy metali niezbędnych do życia, a także w detoksyfikacji szkodliwych metali ciężkich. Wykorzystując technikę hybrydyzacji northern przeanalizowano poziom ekspresji metalotioneiny 2 Brassica napus L. (BnMT2) w liściach roślin inokulowanych bakteriami ryzosferowymi. Rośliny rosnące w glebie skażonej metalami ciężkimi (Cd, Cu, Zn i Pb), zainokulowane szczepem Bacteroidetes bacterium (I-116-1) i koinokulowane Pseudomonas fluorescens i Variovorax sp. (LIC1 i ML3-12) zawierały największą ilość transkryptu badanego genu. Równoczesne szczepienie rzepaku bakteriami ryzosferowymi B. bacterium, Ps. fluorescens i Variovorax sp. (I-116-1, LIC1 i ML3-12) oraz B. bacterium i Ps. fluorescens (I-116-1 i LIC1) wpływało na nieznaczne zwiększenie poziomu ekspresji BnMT2 w porównaniu do roślin nieinokulowanych. Najniższy poziom ekspresji BnMT2 stwierdzono u roślin rosnących w obecności Ps. fluorescens (LIC1). Poziom ekspresji metalotioneiny 2 w liściach rzepaku zależał od rodzaju bakterii ryzosferowych obecnych w glebie.
The involvement of some low-molecular thiol compounds in the mechanisms of peroxidative action of cadmium (Cd) and ethanol (EtOH) was studied. Concentrations of reduced glutathione (GSH), metallothionein (Mt) and thiol (-SH) groups in protein and non-protein fractions were assessed in the homogenates of the liver and kidney of rats exposed to Cd (50 Cd/dm³ of drinking water) and EtOH (5 g EtOH/kg body weight/24 h, intragastrically), singly or in combination, for 12 weeks. Exposure to Cd caused a reduction in the concentration of GSH and non-protein SH groups in the liver and kidneys with a simultaneous increase in Mt level in these organs. The concentration of total SH groups increased only in kidneys. Administration of EtOH had no effect on Mt concentration in both organs, but caused a reduction in the concentration of GSH and non-protein SH groups. A reduction in the level of total SH groups following exposure to EtOH was also noted in the liver. In the group of rats with a simultaneous exposure to Cd and EtOH, GSH concentration was decreased in the liver compared to the control and Cd-exposed animals, and in the kidney in comparison to the control and EtOH-receiving rats. Following the combined exposure to Cd and EtOH, the concentration of non-protein SH groups decreased in the liver and kidneys in comparison to the control and Cd-exposed rats, and in the liver also in comparison to the EtOH group. Mt concentration increased in the liver and kidneys of animals exposed to a combination of Cd and EtOH, compared to the control and EtOH group, but was reduced compared to the Cd group. Combined administration of Cd and EtOH caused an increase in the concentration of total SH groups in the kidneys compared to the control, Cd and EtOH groups. A negative correlation was found between GSH concentration and malondialdehyde (MDA) levels and positive correlation between Mt and MDA. The intensity of lipid peroxidation as well as GSH and Mt concentrations influencing this process in the state of combined exposure to Cd and EtOH results both from independent actions of these substances and interactions between them. The study outcome seems to indicate that the Cd- and EtOH-induced reduction in GSH and non-protein SH groups in the liver and kidneys may be one of the mechanisms that leads to lipid peroxidation in these organs.
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