Metallothionein mRNA Expression Research Articles

Effect of initial periodontal therapy on metallothionein levels in smokers and non-smokers with periodontitis.

The aim of this study was to determine the effect of non-surgical periodontal therapy (NSPT) on mRNA expression of metallothionein (MT) and its levels in serum, saliva and gingival crevicular fluid (GCF) of smokers (S) and non-smokers (NS) with periodontitis (P).A total of 100 participants were included: 48 periodontally healthy (PH) subjects (24 S [PH + S] and 24 NS [PH + NS]) and 52 patients with P (27 S [P + S] and 25 NS [P + NS]). Clinical parameters were recorded, and biofluids (serum, saliva and GCF) and gingival tissue samples were obtained at baseline in all groups and 3months after NSPT in P groups. MT levels in biofluids were determined by ELISA. In gingival tissues, MT-mRNA expression was quantified using real-time PCR. mRNA expression of MT and its levels in biofluids were significantly higher in P + S compared to other groups, and the differences between P + NS and PH + S were non-significant. A significant decrease was observed for MT levels in biofluids, and MT-mRNA expression in periodontitis patients after NSPT. In conclusion, smoking and periodontitis are associated with higher MT expression which decreases after NSPT. MT as an oxidative stress biomarker and its therapeutic role in periodontitis should be investigated in future studies.Clinical trial registration: The study was prospectively registered at Clinical Trials Registry-India (ctri.nic.in) as CTRI/2018/08/015427 on August 23, 2018.

Developmental regulation of zinc homeostasis in differentiating oligodendrocytes

Oligodendrocytes develop through sequential stages and understanding pathways regulating their differentiation remains an important area of investigation. Zinc is required for the function of enzymes, proteins and transcription factors, including those important in myelination and mitosis. Our previous studies using the ratiometric zinc sensor chromis-1 demonstrated a reduction in intracellular free zinc concentrations in mature MBP+ oligodendrocytes compared with earlier stages (Bourassa et al., 2018). We performed a more detailed developmental study to better understand the temporal course of zinc homeostasis across the oligodendrocyte lineage. Using chromis-1, we found a transient increase in free zinc after O4+,O1− pre-oligodendrocytes were switched from proliferation medium into terminal differentiation medium. To gather other evidence for dynamic regulation of free zinc during oligodendrocyte development, qPCR was used to evaluate mRNA expression of major zinc storage proteins metallothioneins (MTs) and metal regulatory transcription factor 1 (MTF1), which controls expression of MTs. MT1, MT2 and MTF1 mRNAs were increased several fold in mature oligodendrocytes compared to oligodendrocytes in proliferation medium. To assess the depth of the zinc buffer, we assayed zinc release from intracellular stores using the oxidizing thiol reagent 2,2′-dithiodipyridine (DTDP). Exposure to DTDP resulted in ∼ 100% increase in free zinc in pre-oligodendrocytes but, paradoxically more modest ∼ 60% increase in mature oligodendrocytes despite increased expression of MTs. These results suggest that zinc homeostasis is regulated during oligodendrocyte development, that oligodendrocytes are a useful model for studying zinc homeostasis in the central nervous system, and that regulation of zinc homeostasis may be important in oligodendrocyte differentiation.

Effect of salinity on the toxicokinetics, oxidative stress, and metallothionein gene expression in Meretrix meretrix exposed to cadmium

To understand the effect of salinity on the toxicokinetics, oxidative stress, and detoxification of cadmium-exposed Meretrix meretrix, M. meretrix were acclimatized to different salinities (8, 14, 20, 26, and 32 ppt) for 14 d, exposed to 10 μg/L Cd for 7 d, followed by a 28-day depuration period. The internal Cd concentration was determined, and the activities of antioxidant enzymes (superoxide dismutase (SOD), catalase (CAT), and glutathione-S-transferase (GST)), and the malondialdehyde (MDA) content were measured. The mRNA expression levels of antioxidant enzyme (Cu/Zn SOD, CAT) and detoxification-related genes metallothionein (MT) were analyzed. The mean concentrations of Cd in M. meretrix tissues were in the order gill > digestive gland > mantle > axe foot. The Cd uptake rate in the four tissues decreased with increasing salinity (range: 14–26 ppt). The Cd elimination half-lives were the highest at 8 ppt and 14 ppt salinity. Cadmium activated the four oxidative stress-related related enzymes in the gills. At the end of accumulation period, Cd exposure at 20 ppt salinity significantly increased the expression of Cu/Zn SOD. CAT expression was significantly inhibited at 20 ppt salinity, but was induced at 32 ppt. MT mRNA expression was only induced under Cd at 20 ppt salinity. At the end of depuration period, Cu/Zn SOD expression was inhibited at salinities of 8, 14, and 26 ppt. The results indicated that SOD, CAT, GST, MDA, Cu/Zn SOD, CAT, and MT were sensitive to cadmium in a water environment, and can be used as indicators of marine heavy metal pollution.

Vascular protection afforded by zinc supplementation in human coronary artery smooth muscle cells mediated by NRF2 signaling under hypoxia/reoxygenation

Zinc (Zn) has antioxidant, anti-inflammatory and anti-proliferative actions, with Zn dysregulation associated with coronary ischemia/reperfusion injury and smooth muscle cell dysfunction. As the majority of studies concerning Zn have been conducted under non-physiological hyperoxic conditions, we compare the effects of Zn chelation or supplementation on total intracellular Zn content, antioxidant NRF2 targeted gene transcription and hypoxia/reoxygenation-induced reactive oxygen species generation in human coronary artery smooth muscle cells (HCASMC) pre-adapted to hyperoxia (18 kPa O2) or normoxia (5 kPa O2). Expression of the smooth muscle marker SM22-α was unaffected by lowering pericellular O2, whereas calponin-1 was significantly upregulated in cells under 5 kPa O2, indicating a more physiological contractile phenotype under 5 kPa O2. Inductively coupled plasma mass spectrometry established that Zn supplementation (10 μM ZnCl2 + 0.5 μM pyrithione) significantly increased total Zn content in HCASMC under 18 but not 5 kPa O2. Zn supplementation increased metallothionein mRNA expression and NRF2 nuclear accumulation in cells under 18 or 5 kPa O2. Notably, NRF2 regulated HO-1 and NQO1 mRNA expression in response to Zn supplementation was only upregulated in cells under 18 but not 5 kPa. Furthermore, whilst hypoxia increased intracellular glutathione (GSH) in cells pre-adapted to 18 but not 5 kPa O2, reoxygenation had negligible effects on GSH or total Zn content. Reoxygenation-induced superoxide generation in cells under 18 kPa O2 was abrogated by PEG-superoxide dismutase but not by PEG-catalase, and Zn supplementation, but not Zn chelation, attenuated reoxygenation-induced superoxide generation in cells under 18 but not 5kPaO2, consistent with a lower redox stress under physiological normoxia. Our findings highlight that culture of HCASMC under physiological normoxia recapitulates an in vivo contractile phenotype and that effects of Zn on NRF2 signaling are altered by oxygen tension.

Proximal tubular transport of Metallothionein-Mercury complexes and protection against nephrotoxicity

Mercury (Hg) is an important environmental toxicant to which humans are exposed on a regular basis. Mercuric ions within biological systems do not exist as free ions. Rather, they are bound to free sulfhydryl groups (thiols) on biological molecules. Metallothionein (MT) is a cysteine-rich, metal-binding protein that has been shown to bind to heavy metals and reduce their toxic effects in target cells and organs. Little is known about the effect of MT on the handing and disposition of Hg. Therefore, the current study was designed to test the hypothesis that overexpression of MT alters the corporal disposition of Hg and reduces its nephrotoxicity. Furthermore, the current study examined the transport of Hg-MT complexes in isolated proximal tubules. Rats were treated with saline or Zn followed by injection with a non-nephrotoxic (0.5 µmol kg−1), moderately nephrotoxic (1.5 µmol kg−1), or significantly nephrotoxic (2.25 µmol kg−1) dose of HgCl2 (containing radioactive Hg). Pretreatment with Zn increased mRNA expression of MT and enhanced accumulation of Hg in the renal cortex of male and female rats. In addition, injection with Zn also protected animals from Hg-induced nephrotoxicity. Studies using isolated proximal tubules from rabbit kidney demonstrated that Hg-MT is taken up rapidly at the apical and basolateral membranes. The current findings suggest that at least part of this uptake occurs through an endocytic process. This study is the first to examine the uptake of Hg-MT complexes in isolated proximal tubules. Overall, the findings of this study suggest that supplementation with Zn may be a viable strategy for reducing the risk of Hg intoxication in at-risk populations.

Parental cadmium exposure during the spawning period reduces cadmium sensitivity through the antioxidant system in rare minnow (Gobiocypris rarus) larvae

Cadmium (Cd) is a noxious heavy metal widely dispersed in aquatic systems. Parental Cd exposure of fish species at environmental concentrations has been shown to cause deformities and stunted growth in their offspring. However, the long-term effects and the mechanisms underlying parental Cd exposure in fish species on Cd sensitivity in their offspring remain unclear. To explore the impacts of parental Cd exposures on Cd sensitivity, rare minnow (Gobiocypris rarus) larvae whose parents were exposed to Cd at 0, 5 or 10 μg/L for 28 days were established. Results showed that parental Cd exposure in rare minnow increased the Cd content of its larvae. In terms of malformation rate, mortality rate and total length at 7 days of rare minnow larvae, parental Cd exposure at 5 or 10 μg/L reduced Cd sensitivity. Further mechanistic investigation demonstrated that parental Cd exposure significantly upregulated the expression of antioxidant gene regulated by nuclear factor erythroid 2-related factor 2 (Nrf2) and nuclear factor-kappa B (NF-кB) in rare minnow larvae. In addition, parental Cd exposure significantly elevated the level of reactive oxygen species (ROS) and malondialdehyde (MDA), but markedly decreased catalase (CAT), superoxide dismutase (SOD) and oxidized glutathione (GST) activity. The impact of parental Cd exposure to metallothionein (MT) content and the expression of MT mRNA, a detoxifying metallothionein, showed that parental Cd exposure of rare minnow induced oxidative stress in the larvae. Meanwhile, these results indicated that parental Cd exposure in rare minnow reduced the Cd sensitivity of the larvae via activating the Nrf2-mediated antioxidant system. This project helps us to further understand the toxicological mechanism of Cd in fish species and properly assess its potential ecological risk.

Toxicity response to benzo[α]pyrene exposure: Modulation of immune parameters of the bay scallop, Argopectenirradians

Bay scallops were exposed to four BaP concentrations (0.5, 1.0, 10 and 50 μg/L) for 72 h to elucidate their immune response. Immune parameters were evaluated by measuring nitric oxide (NO) levels in hemolymph. Additionally, we measured peptidoglycan recognition proteins (PGRP), fibrinogen-domain-containing protein (FReDC1), metallothionein (MT), and heat shock protein (HSP) 70 mRNA expression in digestive diverticula. NO as well as FReDC1 and MT expression in each BaP group increased significantly over time except for the BaP 0.5 group. The PGRP and HSP70 mRNA expression in the BaP 50 group increased in the range 6–24 h and then decreased. In situ hybridization also confirmed that there was higher MT mRNA expression in the BaP 50 group than in the control group at 72 h. Our results suggest that higher levels of BaP dampened scallop immune responses, while simultaneously reducing their ability to cope with oxidative stress and DNA damage. BaP exposure can be considered a potential immune inducer in bay scallop.

Gestational Cd Exposure in the CD-1 Mouse Sex-Specifically Disrupts Essential Metal Ion Homeostasis.

In CD-1 mice, gestational-only exposure to cadmium (Cd) causes female-specific hepatic insulin resistance, metabolic disruption, and obesity. To evaluate whether sex differences in uptake and changes in essential metal concentrations contribute to metabolic outcomes, placental and liver Cd and essential metal concentrations were quantified in male and female offspring perinatally exposed to 500 ppb CdCl2. Exposure resulted in increased maternal liver Cd+2 concentrations (364 µg/kg) similar to concentrations found in non-occupationally exposed human liver. At gestational day (GD) 18, placental Cd and manganese concentrations were significantly increased in exposed males and females, and zinc was significantly decreased in females. Placental efficiency was significantly decreased in GD18-exposed males. Increases in hepatic Cd concentrations and a transient prenatal increase in zinc were observed in exposed female liver. Fetal and adult liver iron concentrations were decreased in both sexes, and decreases in hepatic zinc, iron, and manganese were observed in exposed females. Analysis of GD18 placental and liver metallothionein mRNA expression revealed significant Cd-induced upregulation of placental metallothionein in both sexes, and a significant decrease in fetal hepatic metallothionein in exposed females. In placenta, expression of metal ion transporters responsible for metal ion uptake was increased in exposed females. In liver of exposed adult female offspring, expression of the divalent cation importer (Slc39a14/Zip14) decreased, whereas expression of the primary exporter (Slc30a10/ZnT10) increased. These findings demonstrate that Cd can preferentially cross the female placenta, accumulate in the liver, and cause lifelong dysregulation of metal ion concentrations associated with metabolic disruption.

Relative Bioavailability of Broiler Chickens Fed with Zinc Hydroxychloride and Sulfate Sources for Corn-Soybean Meal.

This study was conducted to determine relative bioavailability (RBV) of basic zinc chloride (BZC) compared to zinc sulfate monohydrate (ZSM) for broilers. A randomized design involving a 2 × 3 factorial arrangement of the different treatment regimens plus one negative control was set up for this study. A total of 630 newly hatched male AA broiler chicks were randomly allocated to 42 different pens (15 chickens/pen) and assigned to 7 dietary treatments in a completely randomized design. The diet was supplemented with 0, 20, 40, or 80mg of Zn mg/kg of feed in the form of ZSM or BZC. The results showed that zinc supplementation altered average daily gain (ADG) and feed conversion ratio (FCR) (P < 0.05) for both zinc sources. It was observed that the weight gain increased linearly (P < 0.01) and FCR decreased linearly as dietary BZC and ZSM concentration increased. Moreover, compared with chickens fed with ZSM, chickens fed with BZC had higher ADG and lower FCR from days 0 to 14 (P < 0.05), and higher activity of plasma alkaline phosphatase (ALP) (P < 0.05), total superoxide dismutase (T-SOD), and CuZn superoxide dismutase (CuZn-SOD) (P < 0.01) in the plasma of chickens fed with BZC at zinc level 80mg/kg at day 14. The pancreas divalent metal-ion transporter-1 (DMT1) mRNA expression of chickens fed with BZC was found to be significantly enhanced at day 28, and the pancreas metallothionein (MT) mRNA expression for BZC fed group was also markedly increased at Zn levels of 20 and 40mg/kg respectively. The relative bioavailability (RBV) of BZC (Zn sulfate 100%) based on ADG in the starter phase was 110.82%, whereas the tibia zinc content, as well as the activities of plasma ALP and CuZn-SOD, and the pancreas MT mRNA level were in the range between 108 and 119%. It was thus concluded that BZC was more efficacious than Zn sulfate and could serve as a potentially novel zinc source in the broilers.

Effects of Zinc Glycinate on Growth Performance, Serum Biochemical Indexes, and Intestinal Morphology of Yellow Feather Broilers

The purpose of this study was to investigate the effects of zinc glycinate (Gly-Zn) on growth performance, serum biochemical index, intestinal morphology, and hepatic metallothionein (MT) mRNA expression in the liver of yellow feather broilers. A total of 540 18-day-old yellow feather broilers were randomly divided into three groups: control group (basal diet), ZnSO4 group (basal diet plus 60 mg Zn/kg from ZnSO4), and Gly-Zn group (basal diet plus 60 mg Zn/kg from zinc glycinate). Each treatment group had 6 replicates with 30 birds in each replicate. The experiment lasted for 42 days (18 to 59 days of age). The results showed that Gly-Zn supplementation significantly improved the average daily gain (ADG) and average daily feed intake (ADFI) of broilers during 18 to 39 days of age compared with that in the control group (P < 0.05) but not different from the ZnSO4 group. The Gly-Zn group had higher glutathione peroxidase (GSH-Px) (P < 0.05) and lower malondialdehyde (MDA) concentrations than the broilers in the control and ZnSO4 group. It was also observed that zinc content in the tibia of Gly-Zn group broilers was higher than the control and ZnSO4 group (P < 0.05). The results of intestinal morphology parameters showed that the Gly-Zn group significantly increased the villus height in duodenum and jejunum (P < 0.05) and decreased crypt depth in duodenum and ileum compared to the control group. However, there were no significant differences between the Gly-Zn group and ZnSO4 group in duodenum and ileum regarding intestinal morphology parameters. The Gly-Zn group significantly increased mRNA expression of MT in the liver than both control and ZnSO4 groups (P < 0.05). Collectively, the results indicated that supplementing 60 mg Zn/kg through zinc glycinate improved growth performance and serum indexes as well as intestinal morphology of yellow feather broilers. It also regulates MT gene expression more effectively than the ZnSO4 group at the transcriptional level.

Effect of medical herbs in Tian-Dong-Tang-Gan powder on the oxidative stress induced by ammonia and nitrite in Litopenaeus vannamei

In order to evaluate the effect of different doses of Tian-Dong-Tang-Gan powder (TDTGP) on the immune function of Litopenaeus vannamei, and the molecular mechanism of anti-ammonia stress and anti-nitrite stress, and screening for the best dose, a 35-day feeding experiment and a 72-h ammonia stress or nitrite stress experiment were carried out. The results showed that when compared with the blank control group, the different doses of TDTGP could increase the body weight (BW), percent weight gain (PWG), body length (BL) and percent body length growth (PBLG), and the activities of acid phosphatase (ACP), phenol oxidase (PO), inducible nitric oxide synthase (i-NOS) and the total antioxidant capacity (T-AOC) in the hemolymph of L. vannamei during the feeding experiment, and the effect of TDTGP-4 treatment group was the best. When exposed ammonia or nitrite stress for 72 h, the PO activity was decreased in the hemolymphs, and the mRNA expression of PPO, MT, GSH-Px, Mn-SOD, HSP70 and HIF-1α in the hepatopancreas of L. vannamei. However, different TDTGP doses were fed to the shrimp followed by the application of ammonia and nitrite stresses. The activities of ACP, PO, i-NOS and T-AOC in the hemolymphs remained stable, and the mRNA expression of PPO, ACP, metallothionein (MT), glutathione peroxidase (GSH-Px), superoxide dismutase (Mn-SOD), heat shock protein 70 (HSP70), Hypoxia inducible factor 1α (HIF-1α) and other factors in the hepatopancreas could be maintained or improved to varying degrees in comparison with the blank control group and positive group. These results indicated that the TDTGP can improve growth performance and non-specific immune response of L. vannamei, and by maintaining the mRNA expression of the antioxidative stress factor, such as MT, GSH-PX, Mn-SOD, HSP70, HIF-1α and PPO and ACP in the hepatopancreas, acting against the damage of hepatopancreas caused by ammonia and nitrite stress in L. vannamei. The changes in BW, WG, activities of PO and ACP, and mRNA expression levels of PO, Mn-SOD and ACP indicated that the optimum supplement of dietary TDTGP should be 4 g per·kg.

Effects of dietary zinc levels on growth performance, digestive enzyme activities, plasma physiological response, hepatic antioxidant responses and metallothionein gene expression in juvenile spotted sea bass ( Lateolabrax maculatus )

The current study was conducted to investigate the effects of dietary zinc on growth performance, body composition, feed utilization, digestive enzyme activities, plasma physiological response, antioxidant responses and metallothionein (MT) gene expression in juvenile spotted sea bass, Lateolabrax maculatus. Six experimental diets (D1-6) were formulated with increasing levels of Zn (0, 20, 40, 80, 160 and 320 mg/kg, respectively) supplied as ZnSO4·H2O. The concentrations of Zn in experimental diets were 36, 55, 74, 110, 175 and 327 mg/kg diet determined by analysis. Each experimental diet was assigned to triplicate groups of 25 L. maculatus for a period of 56 days. The weight gain (WG) and specific growth rate (SGR) in the D4, D5 and D6 groups were significantly increased than that of the control group (p < 0.05), while the feed conversion ratio (FCR) significantly decreased (p < 0.05), respectively. The zinc contents of plasma and vertebra were significantly affected by dietary zinc levels, with the highest zinc content of plasma and vertebra occurring in D6 group (p < 0.05). The levels of plasma alkaline phosphatase (AKP), glucose and catalase (CAT) in the D5 and D6 groups were significantly increased compared with those in the control group (p < 0.05). The content of aspartate aminotransferase (AST) and cortisol (COR) in the D3 and D4 groups was significantly decreased compared with that in the control group (p < 0.05). The intestinal Na+, K+-ATP and CK activities in D3 group were significantly higher than those in the control group (p < 0.05). Compared with the control, the D3 group significantly increased the activities of SOD and T-AOC (p < 0.05), and the D5 group significantly decreased the content of MDA (p < 0.05). The expression of intestinal metallothionein (MT) mRNA in D5 and D6 groups was significantly higher than that of fish in control, D2 and D3 groups (p < 0.05), respectively. Based on the broken-line regression analysis of SGR and AKP, the optimal dietary zinc levels for maximal growth of juvenile L. maculatus were estimated to be 92 and 107 mg/kg diet, respectively.

Lead and cadmium content in Nile tilapia (Oreochromis niloticus) from Egypt: A study for their molecular biomarkers

This study was conducted to examine the residual levels of lead (Pb) and cadmium (Cd) in Nile tilapia (Oreochromis niloticus) collected from three locations (Abbassa, Damietta, and El-Husseiniya) in Egypt. Moreover, the effects of these heavy metals on the antioxidant status in fish were also studied. A trial was conducted to establish some molecular biomarkers reflecting the adverse biological responses towards heavy metals exposure in Nile tilapia by following up changes in the mRNA expression of metallothionein (MT) and heat shock protein (HSP70a). The obtained results indicated that both Pb and Cd residual levels in the fish muscles exceeded maximum permissible limits. Both gills and livers had higher levels of Pb and Cd compared with the muscular tissue. Contamination with heavy metals had a significant alteration on the antioxidant status in Nile tilapia. MT and HSP70a mRNA expressions in the livers of Nile tilapia collected from locations with high Pb and Cd contamination were significantly upregulated. The major novelty of the current study is studying the effects of Pb, and Cd on the antioxidant status in fish, and the development of two molecular biomarkers (MT and HSP70) for the Nile tilapia exposure to such pollutants that can be used as tools to understand the fish response and adaptation to the exposure to high levels of Pb, and Cd.

Zinc alleviates the heat stress of primary cultured hepatocytes of broiler embryos via enhancing the antioxidant ability and attenuating the heat shock responses.

Zinc (Zn) has been shown to attenuate the adverse effects of heat stress on broilers, but the mechanisms involving this process remain unclear. We aimed to investigate possible protective mechanisms of Zn on primary cultured hepatocytes of broiler embryos subjected to heat stress. Three experiments were conducted. In Exp. 1, hepatocytes were treated with 0, 50, 100, 200, or 400 μmol/L added Zn as inorganic Zn sulfate (iZn) for 12, 24 or 48 h. In Exp. 2, cells were exposed to 40 °C (a normal temperature [NT]) and 44 °C (a high temperature [HT]) for 1, 2, 4, 6, or 8 h. In Exp. 3, cells were preincubated with 0 or 50 μmol/L Zn as iZn or organic Zn lysine chelate (oZn) for 8 h under NT, and then incubated with the same Zn treatments under NT or HT for 4 or 6 h. The biomarkers of antioxidative status and heat stress in cells were measured. The results in Exp. 1 indicated that 50 μmol/L Zn and 12 h incubation were the optimal conditions for increasing antioxidant ability of hepatocytes. In Exp. 2, the 4 or 6 h incubation under HT was effective in inducing heat shock responses of hepatocytes. In Exp. 3, HT elevated (P < 0.01) malondialdehyde content and expressions of heat shock protein 70 (HSP70) mRNA and protein, as well as HSP90 mRNA. However, Zn supplementation increased (P < 0.05) copper zinc superoxide dismutase (CuZnSOD) activity and metallothionein mRNA expression, and effectively decreased (P < 0.05) the expressions of HSP70 mRNA and protein, as well as HSP90 mRNA. Furthermore, oZn was more effective (P < 0.05) than iZn in enhancing CuZnSOD activity of hepatocytes under HT. It was concluded that Zn (especially oZn) could alleviate heat stress of broiler hepatocytes via enhancing their antioxidant ability and attenuating heat shock responses.

Assessment spermatogenic cell apoptosis and the transcript levels of metallothionein and p53 in Meretrix meretrix induced by cadmium

Cadmium (Cd) has been widely used in industry and can accumulate in the water, soil, and food. Meretrix meretrix is one of the marine shellfishes cultivated for economic purpose in China. The increasing Cd levels in coastal marine water could adversely affect the economic benefits of shellfish cultivation. In the present study, M. meretrix were exposed to different Cd2+ concentrations (0, 1.5, 3, 6, and 12 mg L-1) for 5 d to evaluate the effects of Cd on spermatogenic cell. The Cd accumulation, survival rate and the indices of oxidative stress and apoptosis were determined in the spermatogenic cells of M. meretrix. The expression levels of p53 and metallothionein (MT) mRNA were also measured in the spermatogenic cells. Cd accumulation and the mortality rate of spermatogenic cells were found to increase in a dose-response manner with Cd2+ concentrations. Histopathology changes, especially the damage of membranous structure, were more severe as the Cd2+ levels in the testis became higher. The indexes of oxidative stress, including reactive oxygen species, malondialdehyde, protein carbonyl derivates and DNA-protein crosslinks all increased after exposure to Cd2+. However, the total antioxidant capacity gradually decreased with the increasing Cd2+ concentration. In addition, exposure to Cd2+ increased the apoptotic rate and caspase-3 and 9 activities but decreased the level of mitochondrial membrane potential and cytochrome C oxidase in the spermatogenic cells. MT mRNA expression increased in lower Cd2+ concentration treated groups whereas decreased in higher groups, while the p53 mRNA expression increased in a dose-response manner with Cd2+ and was positively correlated with the oxidative damage indices. These results indicated that Cd2+ caused oxidative stress and p53 induced apoptosis in the spermatogenic cells, and thus decreased the survival rate of sperm cells. This finding highlights that Cd can reduce the reproductive capacity of M. meretrix, thus threatening to wild shellfish populations and reducing the efficiency of shellfish farming.

Dietary exposure to methyl mercury chloride induces alterations in hematology, biochemical parameters, and mRNA expression of antioxidant enzymes and metallothionein in Nile tilapia.

Methyl mercury chloride "MMC" (CH3ClHg) is an ubiquitous environmental toxicant that causes a variety of adverse effects. In the present study, we investigated the effects of sub-chronic toxicity of MMC on Nile tilapia (Oreochromis niloticus) through the evaluation of growth performance and hematological, biochemical, and oxidative stress biomarkers. From 150 healthy fish, five equally sized treatment groups were created: a control (CT) group fed with a basal diet and four MMC treatment groups exposed to 0.5, 1, 1.5, and 2 mg of MMC per kg of basal diet for 60 days. MMC exposure significantly reduced the growth performance and survival of O. niloticus and decreased red blood cell count and hemoglobin concentration. Treated fish exhibited normocytic normochromic anemia in addition to leucopenia, lymphopenia, granulocytopenia, and monocytopenia. Moreover, MMC exposure significantly affected liver function, including a reduction in the total protein levels while increasing cholesterol and triglyceride levels. It also markedly increased the production of stress biomarkers such as glucose and cortisol levels. Furthermore, MMC significantly elevated the levels of hepatic enzymes, induced tissue damage, and caused inflammation, as indicated by the upregulation of mRNA expression of hepatic metallothionein. Finally, MMC exposure induced oxidative stress by altering the antioxidant status of the liver and downregulating the mRNA expression of superoxide dismutase, glutathione peroxidase, and glutathione S-reductase. In conclusion, MMC toxicity induced hematological and biochemical alterations, leading to an enhanced state of oxidative stress in O. niloticus.

Cadmium Accumulation and mRNA Expression Associated with Detoxification in Various Organs of Black Seabream (Acanthopagrus schlegelii) Exposed to Cadmium-contaminated Diet

Cadmium toxicity is proportional to its ionic concentration, and with long-term exposure, it interferes with ion regulation to maintain homeostasis and affects growth, reproduction, development, behavior, and immune and endocrine systems in organisms. Particularly, cadmium has a long residence time in the body, and if continuously absorbed, it accumulates in tissues, causing bioconcentration. Here, black seabream (Acanthopagrus schlegelii) was exposed to a cadmium-treated diet to investigate cadmium accumulation and mRNA expression of metallothionein (MTII) and heat shock protein (HSP90). The tissue-specific cadmium concentration showed a significant correlation with cadmium concentration and time in all treated tissues except the muscle. Cadmium accumulation in each organ decreased as follows: intestine > liver > gill > muscle. The MT II mRNA level initially increased in the intestine, before decreasing with time, different from the pattern in the other organs. In contrast, the liver showed an increasing trend of MT II mRNA level after the initial increase in the intestine. An increase in the MT II mRNA level in the gill was observed only under high cadmium concentration. Besides, the HSP90 mRNA level initially increased in the intestine and liver, but decreased with time, similar to that in the control group on day 28. mRNA expression of both MT II and HSP90 was observed in the gill and no significant difference was observed in the muscle. The results suggest that the cadmium accumulation pattern with dietary exposure to cadmium was different in each organ in relation to the protective genes metallothionein (MT II) and heat shock protein (HSP90).

Effects of organic zinc on tibia quality, mineral deposit, and metallothionein expression level of aged hens

ABSTRACTThe study aimed to determine the effects of methionine hydroxy analog chelate zinc on the tibia quality, mineral deposit, apparent retention of nutrients, and liver metallothionein (MT) expression level of aged laying hens. A total of 960 layers (Hy-Line Grey, 57 wk old) were randomly assigned into 4 groups, and each group had 8 replicates of 30 hens. During the first 2 wk, groups were fed a basal diet without extra zinc (Zn: 35.08 mg/kg). During the ensuing 14 wk, 4 levels of Zn (inorganic Zn: 80 mg/kg; organic Zn: 20, 40, 80 mg/kg) were added to the diet. The results indicated that both the Zn source and level did influence tibia strength and calcium (Ca) and Zn concentrations of tibia (P < 0.05), whereas there were no differences in the copper (Cu) and phosphorus (P) concentrations of the tibia and the tibia length (P > 0.05). Moreover, dietary supplementation with 40 or 80 mg/kg of organic Zn showed higher Zn and Ca concentrations in the tibia and higher tibia strength. The Cu concentration in the liver showed no difference among the 4 treatments, whereas the Zn concentration in the liver increased with the increasing Zn level. The apparent retention of P, iron (Fe), and manganese (Mn) was not affected by the Zn level or source (P > 0.05). However, the organic Zn group increased the apparent retention of Cu, Zn, Ca, crude protein (CP), and energy, and the group supplemented with 40 or 80 mg/kg of organic Zn obtained significant effects (P < 0.05). Moreover, dietary supplementation with 40 or 80 mg/kg organic Zn increased the MT mRNA expression of the liver at week 72, whereas 20 mg/kg of organic Zn decreased it (P < 0.05). In conclusion, this study suggested that an optimum dietary (40 mg/kg) organic Zn level plays a key role in promoting the apparent retention of minerals and nutrients, trace element deposit, and MT mRNA expression.

Expression of Metallothionein after Administration of Aspirin, Vitamin C or Zinc Supplement in the DMH Induced Colon Carcinoma in Rat

Background:Chemoprevention refers to the use of specificnatural or synthetic chemical agents to suppress the development and progression to carcinoma. The purpose of this study was to assess the effect of aspirin, vitamin C or zinc on the metallothionein (MT) mRNA gene expression as well as MT protein content byimmunohistochemistry andradioimmunoassay (RIA) in 1, 2-dimethyl hydrazine (DMH) induced cancerous colonic tissuein rats.Methods:Rats were randomly divided into three groups, group 1 (aspirin), group 2 (vitamin C) group 3 (zinc), each of which was further sub divided into two groups and given subcutaneous injections of DMH (30 mg/kg body weight) twice a week for 3 months and sacrificed at either 4 months (A-precancer model) or at 6 months (B-cancer model). The control groups were administered 0.5 ml saline subcutaneously. All the 3 groups were simultaneouslyadministered aspirin, vitamin Cor zinc supplement respectively from the beginning till the end of the study.Results:It was observed that rats co-treated with aspirin, vitamin C or zinc resulted in a significant increase in the colonic MT mRNA expression in the precancer and cancer model as compared to the saline only controls. MT protein expression showed a 60%, 64% and 78% immunopositivity in the co-treated groups respectively. The mean MT content in the precancer and the cancer model was restored to near normal levels in all the three co-treated groups.Conclusion:These results suggest that co-administration of aspirin, vitamin C or zinc resulted in a significant increase in MT mRNA gene expression, MT protein expression and MT protein content which could possibly be one of the reasons for a chemo protective effect against progression to colonic cancer in a chemically induced DMH model in rat. Zinc supplement had a greater effect on metallothionein expression than aspirin or vitamin C.

Effect of Supplementation of Nano Zinc Oxide on Nutrient Retention, Organ and Serum Minerals Profile, and Hepatic Metallothionein Gene Expression in Wister Albino Rats

A study was conducted to validate the effects of nano form of zinc (NZn) on nutrient digestibility, zinc retention, organ and serum zinc profile, and hepatic metallothionein gene expression in Wistar albino rats (WAR). Nano zinc (NZn) was synthesized through chemical method, by using 0.45M zinc nitrate [Zn(NO3)2.6H2O] and 0.9M sodium hydroxide (NaOH). The NZn particle in its oxide form was characterized by TEM-EDAX and XRD, and found to be in nano range (below 100 nm. Zinc was supplemented to the Wistar albino rats (WAR) through synthetic semi-purified diet either without Zn, or as inorganic zinc (IZn; 25mg/kg), or as synthesized NZn (25, 12.5, 6.25, 3.125 or 50mg/kg DM) for 60days. The zinc content was observed to be significantly (P < 0.05) higher in liver, bone, kidney, and serum due to NZn supplementation where NZn-50 had highest zinc content and control had the least, without affecting Fe, Mn, and Cu. NZn at 12.5mg/kg group rats were either comparable or better than IZn at 25mg/kg in terms of zinc retention, CP digestibility, zinc level in serum, liver, bone, and kidney suggesting its better bioavailability simultaneously also reduced fecal excretion of zinc to the environment. Metallothionein mRNA expression was upregulated in NZn at 25mg/kg and NZn at 50mg/kg than IZn at 50mg/kg. Thus, in WAR, NZn at half of the ICAR recommendation (25mg/kg DM) is as effective as inorganic zinc at 100% of recommended dose.