Metallothionein 2 activation by pravastatin reinforces epithelial integrity and ameliorates radiation-induced enteropathy.

BACKGROUND: Radiotherapy or accidental exposure to ionizing radiation causes severe damage of healthy intestinal tissues. Intestinal barrier function is highly sensitive to ionizing radiation, and loss of epithelial integrity results in mucosal inflammation, bacterial translocation, and endotoxemia. Few studies have of epithelial integrity as a therapeutic target to treat radiation toxicity. Here, we examined the effects of pravastatin (PS) and the molecular mechanisms underlying epithelial integrity on radiation-induced enteropathy. METHODS: The radio-mitigative effects of PS were evaluated in a minipig model by quantifying clinical symptoms, and performing histological and serological analyses and mRNA sequencing in intestinal tissues. To evaluate the role of intercellular junctions on radiation damage, we used tight junction regulator and metallothionein 2 (MT2) as treatments in a mouse model of radiation-induced enteropathy. Caco-2 monolayers were used to examine functional epithelial integrityand intercellular junction expression. FINDING: Using a minipig model of pharmaceutical oral bioavailability, we found that PS mitigated acute radiation-induced enteropathy. PS-treated irradiated minipigs had mild clinical symptoms, lower intestinal inflammation and endotoxin levels, and improved gastrointestinal integrity, compared with control group animals. The results of mRNA sequencing analysis indicated that PS treatment markedly influenced intercellular junctions by inhibiting p38 MAPK signaling in the irradiated intestinal epithelium. The PS-regulated gene MT2 improved the epithelial barrier via enhancement of intercellular junctions in radiation-induced enteropathy. INTERPRETATION: PS regulated epithelial integrity by modulating MT2 in radiation-damaged epithelial cells. These findings suggested that maintenance of epithelial integrity is a novel therapeutic target for treatment of radiation-induced gastrointestinal damage. FUNDING: As stated in the Acknowledgments.

Efeito da exposição a produtos fumígenos sobre a expressão de metalotioneínas durante a artrite experimental e sobre linfócitos T / Effect of exposure to tobacco-related products on metallothionein expression in the experimental arthritis and on T lymphocytes functions

Produtos liberados pela queima do cigarro convencional (CC) estão relacionados com a progressão clínica da artrite reumatoide (AR). Produtos fumígenos não combustíveis surgiram com a premissa de apresentarem menor toxicidade que o CC, dentre os quais está o tabaco aquecido (heat-not-burn tobacco; HNBT). Neste projeto investigamos os efeitos do HNBT sobre eventos envolvidos na AR, focando na sintomatologia, expressão de metalotioneínas (MTs), e na biologia de linfócitos T CD4+ primários e da linhagem Jurkat. Exposições in vivo ao ar, CC ou HNBT foram realizadas 2 vezes ao dia, 1 hora cada (12 CC ou 24 HNBT/hora), nos dias 14-21 da indução da artrite induzida por antígeno (AIA) em camundongos C57Bl/6. Foram realizadas análises dos parâmetros clínico da doenças, histopatologia e imunohistoquímica; quantificação de nicotina e cotinina séricas por cromatografia líquida acoplada a espectrometria de massas (MS). Os efeitos das exposições in vitro sobre linfócitos T foram mensurados por citometria de fluxo e ELISA. A concentração de metais emitidas pelo CC ou HNBT durante as exposições foram mensurados por MS com plasma acoplado. Camundongos expostos ao CC apresentaram intensa inflamação pulmonar, expressões acentuadas de MTs hepáticas e pulmonares e exacerbação dos parâmetros de AIA quando comparados ao grupo expostos ao HNBT. Animais expostos ao CC ou ao HNBT apresentaram redução na celularidade de órgãos linfoides. Somente a exposição in vitro ao CC causou estresse oxidativo e secreção de citocinas inflamatórias, ativação do receptor de hidrocarbonetos arila (AhR) e polarização de células Th17. Diferentemente, exposição ao CC ou ao HNBT provocaram redução da secreção de IL-2 e proliferação de células Jurkat. A exposição de células Jurkat à nicotina mimetizou os efeitos inibitórios da exposição ao HNBT sobre a secreção de IL-2 e proliferação de linfócitos T. O CC liberou maiores concentrações de metais nas câmaras de exposição. Associados, nossos resultados mostram que embora exposições ao HNBT não exacerbem parâmetros inflamatórios de AIA e nem em funções linfócitos T, ambos produtos prejudicam a celularidade de órgãos linfoides e a proliferação e secreção de IL-2 por linfócitos T

Products released by burning conventional cigarettes (CC) are related to the worsening of rheumatoid arthritis (RA). Non-combustible smoking products appeared with the premise of presenting less toxicity than the CC, among which is the heated tobacco (heat-not-burn tobacco; HNBT). Here, we investigate the effects of HNBT on events involved in RA, focusing on symptoms, expression of metallothioneins (MTs), and on the biology of primary CD4+ T lymphocytes and the Jurkat T cell lineage. In vivo exposures to air, CC or HNBT were performed twice a day, 1 hour each (12 CC or 24 HNBT / hour), on days 14-21 of the induction of antigen-induced arthritis (AIA) in C57Bl / 6 mice. Analyzes of the clinical parameters of the AIA, histopathology, and immunohistochemistry were performed; quantification of nicotine and cotinine by liquid chromatography coupled to mass spectrometry (MS). The in vitro effects of exposures on T lymphocytes were measured by flow cytometry and ELISA. The concentration of metals released by the CC or HNBT during the exposures was measured by MS with coupled plasma. Mice exposed to CC showed intense pulmonary inflammation, marked expressions of hepatic and pulmonary MTs, and exacerbation of AIA parameters when compared to the group exposed to HNBT. Animals exposed to CC or HNBT showed a reduction in the cellularity of lymphoid organs. Only in vitro exposure to CC caused oxidative stress and secretion of inflammatory cytokines, activation of the aryl hydrocarbon receptor (AhR), and polarization of Th17 cells. However, exposure to CC or HNBT led to reduced secretion of IL-2 and proliferation of Jurkat cells. The exposure of Jurkat T cells to nicotine mimicked the inhibitory effects of exposure to HNBT on IL-2 secretion and T lymphocyte proliferation. The CC released higher concentrations of metals in the exposure chambers. In association, our results show that although exposures to HNBT do not exacerbate inflammatory parameters of AIA or T lymphocyte functions, both products impair lymphoid organ cell function and the proliferation and secretion of IL-2 by T lymphocytes

The physiological role and toxicological significance of the non-metal-selective cadmium/copper-metallothionein isoform differ between embryonic and adult helicid snails.

Metal regulation is essential for terrestrial gastropods to survive. In helicid snails, two metal-selective metallothionein (MT) isoforms with different functions are expressed. A cadmium-selective isoform (CdMT) plays a major role in Cd2+ detoxification and stress response, whereas a copper-selective MT (CuMT) is involved in Cu homeostasis and hemocyanin synthesis. A third, non-metal-selective isoform, called Cd/CuMT, was first characterized in Cantareus aspersus. The aim of this study was to quantify the transcriptional activity of all three MT genes in unexposed and metal-exposed (Cd, Cu) embryonic Roman snails. In addition, the complete Cd/CuMT mRNA of the Roman snail (Helix pomatia) was characterized, and its expression quantified in unexposed and Cd-treated adult individuals. In embryos of Helix pomatia, the Cd/CuMT gene was induced upon Cu exposure. Its transcription levels were many times higher than that of the other two MT genes, and also exceeded by far the Cd/CuMT mRNA concentrations of adult snails. In the hepatopancreas of adult Roman snails, no Cd/CuMT could be detected at the protein level, irrespective of whether the snails had been exposed to Cd or not. This contrasts with the situation in the near relative, Cantareus aspersus. It appeared that the 3'-UTR of the Cd/CuMT mRNA differed largely between Cantareus aspersus and Helix pomatia, being larger in the latter species, with a number of putative binding sites for proteins and miRNAs known to inhibit mRNA translation. We suggest this as a possible mechanism responsible for the lack of Cd/CuMT protein expression in adult Roman snails.

Protective effects of Zn2+ against cobalt nanoparticles and cobalt chloride-induced cytotoxicity of RAW 264.7cells via ROS pathway.

Recent concerns have emerged surrounding the toxicity that cobalt may represent when used in MOM implants. Owing to corrosion and wear of MOM implants, the subsequent released cobalt nanoparticles (CoNPs) or Co ions (Co2+) can cause adverse reactions, such as the generation of pseudotumors, extensive necrosis, early osteolysis, and implants failure. The present study confirmed that CoNPs and Co2+ can induce dose- and time-dependent cytotoxicity with increasing reactive oxygen species (ROS) levels. Additionally, using metallothionein (MT), a heavy metal-binding protein, the present study assessed the protective effects of Zn2+ against CoNPs and Co2+-induced cytotoxicity of RAW 264.7 cells through ROS pathway. Further studies are needed to explore the underlying protective mechanisms in vitro. However, the current findings indicate that the ROS pathway may be a potential target for therapeutic interventions.

Phenotypic and biochemical alterations in relation to MT2 gene expression in Plantago ovata Forsk under zinc stress.

Plantago ovata Forsk is an annual herb with immense medicinal importance, the seed and husk of which is used in the treatment of chronic constipation, irritable bowel syndrome, diarrhea since ancient times. Zinc, an essential metal, is required by plants as they form important components of zinc finger proteins and also aid in synthesis of photosynthetic pigments such as chlorophyll. However, in excess amount Zn causes chlorosis of leaf and shoot tissues and generate reactive oxygen species. The present study is aimed at investigating the changes in expression levels of MT2 gene in Plantago ovata under zinc stress. Data show up to 1.66 fold increase in expression of PoMT2 in 1000 µM ZnSO4·7H2O treated sample. Our study also describes alteration of MT2 gene expressions in Plantago ovata as observed through Real time PCR (qPCR) done by [Formula: see text] method. In this study we have observed an upregulation (or induction) in the PoMT2 gene expression level in 500 and 800 µM ZnSO4·7H2O treated samples but found saturation on further increasing the dose to 1000 µM of ZnSO4·7H2O. Determination of the phenotypic and biochemical changes in Plantago ovata due to exposure to zinc stress of concentrations 500, 800 and 1000 µM revealed oxidative stress. The enhanced expression of MT2 gene in Plantago ovata has a correlation with the increased total antioxidant activity and increased DPPH radical scavenging activity.

Profiling the physiological and molecular response to sulfonamidic drug in Procambarus clarkii.

Sulfamethoxazole (SMZ) is one of the most widely employed sulfonamides. Because of the widespread use of SMZ, a considerable amount is indeed expected to be introduced into the environment. The cytotoxicity of SMZ relies mainly on arylhydroxylamine metabolites (S-NOH) of SMZ and it is associated with the production of reactive oxygen species (ROS). There is limited information about the toxic potential of SMZ at the cellular and molecular levels, especially in aquatic and/or non-target organisms. In the present study, the red swamp crayfish (Procambarus clarkii), being tolerant to extreme environmental conditions and resistant to disease, was used as a model organism to profile the molecular and physiological response to SMZ. Haemolymphatic-immunological parameters such as glucose serum levels and total haemocyte counts were altered; moreover, a significant increase in Hsp70 plasma levels was detected for the first time. Variations at the transcriptional level of proinflammatory genes (cyclooxygenase-1, COX 1, and cyclooxygenase-2, COX 2), antioxidant enzymes (glutathione-S-transferase, GST and manganese superoxide dismutase MnSOD), stress response and Fenton reaction inhibitor genes (heat-shock protein 70 HSP70, metallothionein, MT and ferritin, FT) were evaluated, and alterations in the canonical gene expression patterns emerged. Considering these results, specific mechanisms involved in maintaining physiological homeostasis and adaptation in response to perturbations are suggested.

[Functions of metallothioneins and a system of antioxidant defense under the effect of Co- and Zn-containing nanocomposites on crucian carp (Carassius auratus gibelio)].

The effect of metal-nanocomposites (Me-NC) of cobalt and zinc (Co- and Zn-NC, correspondingly) synthecized on the basis of vinylpyrrolidone (PS) on the metal-accumulative proteins with antioxidant potential metallothioneins (MT) in crucian carp (Carassius auratus gibelio) was studied. Fish was subjected to the effect of Co-NC, Zn-NC, Co2+, Zn2+ or polymer carrier (PC) in the concentrations correspondent to 50 microg x Co/l or 100 microg x Zn/l during 14 days. It was shown that the MTs response is highly specific for the nature of metal, both in ion and Me-NC form: the effect of Co and Co-NC provoked the elevation of total MT concentration (MT-SH) and activation of antioxidant defence, whereas Zn and Zn-NC induced the decrease of the concentration of MT-SH and the inhibition of antioxidant defense. All the exposures provoked the decrease of the concentration of immunoreactive chelating MT form (MTi) and reduced glutathione, activation of anaerobiosys and Mn-superoxide dismutase, and also decrease of the concentration of proteins and lipids oxidative injury products. It was accompanied by the increase of the content of erythrocytes with nuclear abnormalities but did not cause the decrease of choline esterase activity. According to the rate of MT-SH and MTi concentrations, antioxidant potential of MTs is determined by its apoform. Our data indicate that partial biodegradation of Me-NC occurs in the organism of crucian carp.

Zinc supplementation attenuates metallothionein and oxidative stress changes in kidney of streptozotocin-induced diabetic rats.

Zinc is an element that under physiological conditions preferentially binds to and is a potent inducer of metallothionein under physiological conditions. The present study was conducted to explore whether zinc supplementation morphologically and biochemically protects against diabetic nephropathy through modulation of kidney metallothionein induction and oxidative stress in streptozotocin-induced diabetic rats. Thirty-two Wistar albino male rats were equally divided into four groups. The first group was used as untreated controls and the second group was supplemented with 30 mg/kg/day zinc as zinc sulfate. The third group was treated with streptozotocin to induce diabetes and the fourth group was treated with streptozotocin and supplemented with zinc as described for group 2. The blood glucose and micro-albuminuria levels, body and kidney weights were measured during the 42-day experimental period. At the end of the experiment, the kidneys were removed from all animals from the four groups. Diabetes resulted in degenerative kidney morphological changes. The metallothionein immunoreactivity level was lower and the kidney lipid peroxidation levels were higher in the diabetes group than in the controls. The metallothionein immunoreactivity levels were higher in the tubules of the zinc-supplemented diabetic rats as compared to the non-supplemented diabetic group. The zinc and metallothionein concentrations in kidney tissue were higher in the supplemented diabetic group compared to the non-supplemented diabetes group. The activity of glutathione peroxidase did not change in any of the four groups. In conclusion, the present study shows that zinc has a protective effect against diabetic damage of kidney tissue through stimulation of metallothionein synthesis and regulation of the oxidative stress.

A metallothionein mimetic peptide protects neurons against kainic acid-induced excitotoxicity.

Metallothioneins I and II (MTI/II) are metal-binding proteins overexpressed in response to brain injury. Recently, we have designed a peptide, termed EmtinB, which is modeled after the beta-domain of MT-II and mimics the biological effects of MTI/II in vitro. Here, we demonstrate the neuroprotective effect of EmtinB in the in vitro and in vivo models of kainic acid (KA)-induced neurotoxicity. We show that EmtinB passes the blood-brain barrier and is detectable in plasma for up to 24 hr. Treatment with EmtinB significantly attenuates seizures in C57BL/6J mice exposed to moderate (20 mg/kg) and high (30 mg/kg) KA doses and tends to decrease mortality induced by the high KA dose. Histopathological evaluation of hippocampal (CA3 and CA1) and cortical areas of mice treated with 20 mg/kg KA shows that EmtinB treatment reduces KA-induced neurodegeneration in the CA1 region. These findings establish EmtinB as a promising target for therapeutic development.

Predictors of blood mercury levels in older urban residents.

OBJECTIVE: The objective of this study was to describe the distribution and predictors of blood mercury levels in an adult population. METHODS: This was a cross-sectional analysis of first-visit data (2001-2002) on a random sample of 474 subjects from the Baltimore Memory Study. RESULTS: After adjustment for race/ethnicity, education, assets, and diabetes, persons in the highest quartile of fish consumption had median mercury levels 1.82 times above the levels in the lowest quartile, while those in the highest education category had median mercury levels 1.57 times higher than levels in the lowest category. Nine percent of subjects were above the reference dose recommended by the Environmental Protection Agency, which is high compared with that found by the Centers for Disease Control and Prevention in women of childbearing age. CONCLUSIONS: These findings offer guidance for targeted education and possible new insights regarding the kinetics of mercury.