Variation of metal and metallothionein concentrations in a natural population of Ruditapes decussatus

The spatial and seasonal variation of total and subcellular distribution of Cd, Cu, and Zn was followed in different tissues (gills, digestive gland, and remaining tissues) of the clam Ruditapes decussatus collected along a metal contamination gradient in the Ria Formosa lagoon (southern Portugal) and compared with metallothionein (MT) concentrations.Total metal concentrations decreased according to the sequence digestive gland > gills > remaining tissues for Cd, digestive gland approximately gills > remaining tissues for Cu and gills > digestive gland > remaining tissues for Zn. MT concentrations in these tissues decreased according to the same sequence observed for Cd. In all the tissues, the highest subcellular concentration was in the cytosol for Cd and Cu and in the pellet for Zn. Among the three metals, Cd concentrations showed the most evident spatial variation. In all tissues, total and subcellular Cd concentrations decreased from the inner parts of the lagoon toward the ocean. However, no significant spatial or seasonal variation occurred in clam tissues for the other two metals, though marginal elevated Cu concentrations were observed in the inner parts of the lagoon. Therefore, Cu subcellular distribution in clam tissues was not significantly altered by Cu changes in the lagoon and are the baseline levels for normal metabolism of this clam population. The fact that total Zn concentrations remained unchanged both spatial and seasonal suggested that these clams regulate Zn in their tissues.In the three tissues, MT bind most significantly to Cd and Cu, while Zn, although binding to MT, is preferably bound to other ligands. MT concentrations showed the same spatial and seasonal variation of Cd and were significantly related with total and heat-treated cytosolic Cd in all tissues. For Cu a significant relationship between MT and total or cytosolic Cu was only observed in the remaining tissues. No relationship was observed between MT and total or cytosolic Zn concentrations. Metals and MT concentrations increased with the increase in the condition index for the gills and the digestive gland and decreased from the remaining tissues.Cd concentrations in the gills increased only in the heat-treated cytosolic fraction while Zn in this fraction decreased. Thus Cd concentrations in this tissue displaced Zn from the MT-fraction, leading to a modification of the soluble/insoluble Zn ratio once total Zn concentrations remained unchanged. This modification reflects a perturbation in the normal metabolism in this tissue due to the excess of Cd present. With the exception of the gills, Zn subcellular distribution in the other two tissues was similar among sites and season.The model that describes the relationship between MT, metals, and weight in the gills, digestive gland and remaining tissues also indicates that Cd was the only metal that influence MT synthesis significantly in all the tissues. The induced and/or existent MT was sufficient to bind free Cd ions present in the cells, preventing any damage to cellular metabolism in this clam population. Therefore, MT in the gills and digestive gland of R. decussatus can be used as an early warning signal for Cd exposure and are a useful biomarker to assess the toxicological status of this population in the Ria Formosa lagoon.     

Kinetic model of cadmium accumulation and elimination and metallothionein response in Ruditapes decussatus

The aim of the present study was to determine the response of metallothionein (MT) during Cd accumulation and elimination in different tissues of the estuarine bivalve Ruditapes decussatus exposed to two nominal Cd concentrations (4 and 40 microg/L) for 40 d, followed by a depuration period of 50 d. Cadmium was accumulated in all tissues of R. decussatus at both exposure concentrations, and the accumulation was tissue dependent. Use of the kinetic model showed that in the gills and remaining tissues, Cd was assimilated faster at the beginning of the exposure and decreased with time, possibly limited by the diffusion rate of this metal within the cell. In the digestive gland, however, the Cd was continuously accumulated. This could reflect that the Cd uptake rate is considerably higher than the loss rate and, therefore, that this tissue has a higher capacity to accumulate Cd compared to the other two tissues. Moreover, the application of this kinetic model in the different subcellular fractions showed that the bioconcentration factor was significantly higher in the low-molecular-weight fraction (where MT is found), suggesting that this fraction binds Cd faster, with a high uptake rate (K(u) = 32/d), and eliminates this metal more slowly (K(1) = 0.005/d). During the depuration phase, MT decreased simultaneously with Cd elimination in all tissues, although with a shorter half-life. In conclusion, the MT response prevented Cd in the tissues of R. decussatus from interfering in the normal clam metabolism; therefore, MT acts as a detoxification mechanism of Cd.     

Differential induction of glutathione S-transferases in the clam Ruditapes decussatus exposed to organic compounds

Studies of glutathione S-transferase (GST) induction were performed in the Mediterranean clam Ruditapes decussatus after controlled exposure to organics in holding tanks. Clams were treated with phenobarbital (PB), benzo[a]pyrene (BaP), and 2,2-bis-(p-chlorophenyl)-1,1-dichlorethylene (p,p'-DDE). Three different substrates, i.e., 1-chloro-2,4-dinitrobenzene (CDNB), ethacrynic acid (ETHA), and paranitrobenzene chloride (PNBC), were used to determine GST activities in order to distinguish the isoenzymes induced by contamination. The isoforms conjugating ETHA were significantly induced by treatment with PB and BaP, whereas exposure to p,p'-DDE induced isoforms conjugating CDNB and ETHA. An antibody against affinity-purified GSTs from R. decussatus was prepared by injection into rabbit. The serum containing the antibody gave a positive reaction with both the purified GSTs from R. decussatus and the low molecular weight GSTs from rat. Subcellular fractions from both control and treated animals were analyzed by Western blot. Cytosolic extracts from clams contaminated with PB and p,p'-DDE showed a 24-kDa band in addition to the 26-kDa band recognized by the antibody. Results of these studies suggest that, in R. decussatus, organics may induce GSTs belonging to the pi class.     

In situ potential use of metallothionein as a biomarker of cadmium contamination in Ruditapes decussatus

Cadmium (Cd) and metallothionein (MT) concentrations were analysed in the gills and digestive gland of clams Ruditapes decussatus collected from two sites of the Gulf of Gabès (Tunisia) which differ by their degree of pollution. The effect of biotic (maturity stages, protein, sex and body mass) and abiotic factors (temperature, salinity, site and season) on Cd and MT concentrations showed that in the reference site “Bordj d’Ungha” physiological changes caused by gamete development and maturity contributed more to changes in MT concentrations than the bioavailable Cd concentrations; whereas, in the contaminated site “El Hofra”, MT induction responded to raised Cd bioavailabilities rather than to physiological changes. With a view to using MTs as metal exposure biomarker in monitoring program, our results show that the analysis in the digestive gland seem to be more relevant than in gills, and that males appear as the most promising candidates compared to females in the determination of this biomarker.     

Pollution-Tolerant Allele in Fingernail Clams (Musculium transversum)

For nearly 50 years, the fingernail clam (Musculium transversum) was believed to be virtually eliminated from the Illinois River. In 1991, workers began finding substantial populations of M. transversum in the Illinois River including several beds in and around the highly polluted Chicago Sanitary District. In order to determine if populations of M. transversum from polluted sites exhibited any genetic response to the high levels of toxins and to examine the genetic structure of several populations of M. transversum for any changes due to the population crash, starch-gel electrophoresis was performed on M. transversum from three Illinois River localities and four Mississippi River basin locations. The sampled populations produced an inbreeding coefficient (F_IS) of 0.929, indicating that the populations were highly inbred. The results of a suspected founder effect due to a bottleneck was suggested by an F_ST= 0.442. The isozyme Glucose-6-phosphate isomerase-2 (Gpi-2) produced allelic frequency patterns that were consistent with expected patterns of a pollution-tolerant allele. Polluted sites exhibited elevated frequencies of Gpi-2 ¹⁰⁰ whereas nonpolluted sites exhibited elevated frequencies of Gpi-2 ⁷⁴ . This frequency pattern suggested that natural selection was occurring in populations under severe toxic pressures, leading to an increase in the frequency of the allele Gpi-2 ¹⁰⁰ . Therefore, Gpi-2 ¹⁰⁰ is a possible pollution-tolerant mutation in M. transversum. Received: 29 June 1996/Accepted: 8 February 1998     

Polycyclic aromatic hydrocarbons concentrations and biomarker responses in the clam Ruditapes decussatus transplanted in the Ria Formosa lagoon

Clams Ruditapes decussatus were transplanted in the Ria Formosa lagoon and the variation of PAH concentrations in the whole soft tissues measured, along with a suite of biomarkers, including the following: (a) phase I and phase II metabolism of xenobiotics enzymes: benzo[a]pyrene hydroxylase (BPH) and glutathione S-transferase (GST); (b) antioxidant enzymes: superoxide dismutase, catalase and glutathione peroxidases and (c) lipid peroxidation (LPO) levels. Individual PAHs were differently accumulated and eliminated by R. decussatus. During the metabolisation of PAHs by R. decussatus BPH was clearly induced in the digestive gland. Moreover, ROS lead to the induction of protective antioxidant enzymes still causing oxidative damage to membranes. Therefore, BPH seems to be a relevant indicator of PAHs in R. decussatus.     

DNA Alterations Triggered by Environmentally Relevant Polymetallic Concentrations in Marine Clams Ruditapes philippinarum and Polychaete Worms Hediste diversicolor

We exposed marine clams (Ruditapes philippinarum) and aquatic worms (Hediste diversicolor) to environmentally relevant concentrations of two metal mixtures each containing three divalent metals [(C₁ in µg/L) cadmium (Cd) 1, mercury (Hg) 0.1, and lead (Pb) 4] and [C₂ in µg/L) Cd 17, Hg 1.1, and Pb 55]. Animals collected in the Arcachon Bay were exposed for 8 days in microcosms made up of a mixed biotope consisting of a water column and natural marine sediment both taken up from the Arcachon Bay. Bioaccumulation analysis showed a significant increase of Cd, Hg, and Pb in clams, particularly at C₂ concentration in the water column reaching, in soft body, 2.3 ± 0.3 µg Cd/g, 0.7 ± 0.2 µg Hg/g, and 45 µg Pb/g dry weight (dw). DNA alterations and upregulation of the cox1 mitochondrial gene were also observed in clam gill after exposure to the metal blend. For worms exposed to the C₂ metal blend, DNA alterations and significant increase of Cd and Hg concentrations were observed reaching 0.5 ± 0.1 µg Cd/g and 2 ± 0.6 µg Hg/g dw.     

氯菊酯对大鼠脑组织一氧化氮的影响

为了研究氯菊酯对大鼠脑组织一氧化氮(NO)的作用,采用腹腔注射,按200mg·kg^-1·d^-1分别给予大鼠氯菊酯1天、3天和5天后,测定氯菊酯对大脑皮层、海马和小脑组织中一氧化氮合酶(NOS)活性、亚硝酸根(NO₂^-)含量和环鸟苷酸(cGMP)含量的影响.结果显示:1天组所测脑区NOS活性和cGMP含量无明显变化;3天组小脑NOS活性降低,大脑皮层和小脑cGMP减少,而海马cGMP明显增加;5天组大脑皮层和小脑NOS活性明显降低,小脑cGMP含量明显减少;3个时间组所测脑区NO₂^-含量与对照组相比无明显变化.结果表明:氯菊酯染毒持续时间不同对大鼠脑组织NO的作用不同,可能在其毒作用的发生发展中具有重要意义.     

Effects of Metal Exposures on Juvenile Clams,Mercenaria mercenaria

Bulletin of Environmental Contamination and Toxicology - Elevated metal concentrations occur throughout the coastal zone due to urbanization and various anthropogenic activities. The lethal...     

Variation of metallothionein and metal concentrations in the digestive gland of the clam Ruditapes decussatus: sex and seasonal effects

Metallothionein (MT) and metal (Cd, Cu, and Zn) concentrations were determined in the digestive gland of male and female Ruditapes decussatus. Clams were collected monthly during the period of sexual development, from June to September, at two different sites of a coastal lagoon, the Ria Formosa Lagoon, on the south coast of Portugal. The MT concentrations were determined in the heat-treated cytosolic fraction of the digestive gland of both male and female clams. Total metal (Cd, Cu, and Zn) concentrations in the heat-treated cytosol of the digestive gland were also determined in both sexes. The MT and metal concentrations in the digestive gland were not sex dependent in this species. Therefore, these results suggest that random samples can be used to determine MT and metal concentrations in the digestive gland of R. decussatus. Seasonal and site-specific dependency, however, were detected for MT, Cd, and Cu concentrations. The maximum concentration levels of MT, Cd, and Cu were detected in July and August for both sexes in the same area. Zinc concentrations, however, did not show marked seasonal variations, indicating that this species is able to regulate zinc concentrations. The highest MT concentrations were significantly related to the highest metal concentrations.     

Arsenic Bioaccumulation and Biotransformation in Clams (Asaphis violascens) Exposed to Inorganic Arsenic: Effects of Species and Concentrations

Bulletin of Environmental Contamination and Toxicology - High arsenic (As) concentrations are found in marine clams, usually as less-toxic arsenobetaine (AsB). However, when clams were exposed to...     

氯丁二烯所致接触工人不良健康效应的生物标记研究

本研究对６８例氯丁二烯（ＣＢＤ）接触工人和４９例健康人进行了调查。结果显示，ＣＢＤ接触组血清中甘胆酸（ＣＧ）含量明显升高，而血清中丙氨酸转氨酶、硫酸锌浊度试验和麝香草酚浊度试验无明显改变；全血谷胱甘肽过氧化物酶（ＧＳＨ—Ｐｘ）活性和血清铜蓝蛋白（ＣＰ）含量明显降低，这与动物实验结果基本一致，即ＣＢＤ在体内生成的活性中间物可导致氧化性损害。血清ＣＧ含量、全血ＧＳＨ—ｐｘ活性及血清ＣＰ可能是接触ＣＢＤ所致不良健康效应的生物标记。     

Effects of Calcium on the Uptake and Elimination of Cadmium and Zinc in Asiatic Clams

We examined the effects of Ca, one of the major ions contributing to water hardness, on the uptake and elimination of Cd and Zn in the Asiatic clam Corbicula fluminea, a common bivalve species in the rivers and streams of Southern China. Over the wide range of dissolved Cd (4.5 to 446 nM) and Zn (15.3 to 1529 nM) concentrations, uptake of both metals increased with increasing dissolved metal concentration, showing Michaelis-Menten–type saturation kinetics. At each tested metal concentration, increased Ca concentration led to a significantly lower metal uptake. When the Ca, Cd, Zn concentrations were expressed as free-ion activities, Cd and Zn uptake data fitted the Michaelis-Menten inhibition model well. We also quantified the effects of the two Ca blockers on metal uptake by the clams. Verapamil significantly inhibited the uptake Cd and Zn, but the inhibitive effect of lanthanum on metal uptake was more evident for Cd than for Zn. Ca did not significantly affect the assimilation efficiency of either metal from ingested phytoplankton, nor did it affect the elimination of the two metals during a 1-month depuration period. Our study has shown that Ca inhibited the uptake of metals from water; such an effect could be predicted based on the free-metal ion activities. Ca did not seem to have a direct effect on metal assimilation from food or metal elimination from the bivalve.     

The Effects of a Mediterranean Diet Intervention on Targeted Plasma Metabolic Biomarkers among US Firefighters: A Pilot Cluster-Randomized Trial

Metabolomics is improving the understanding of the mechanisms of the health effects of diet. Previous research has identified several metabolites associated with the Mediterranean Diet (MedDiet), but knowledge about longitudinal changes in metabolic biomarkers after a MedDiet intervention is scarce. A subsample of 48 firefighters from a cluster-randomized trial at Indianapolis fire stations was randomly selected for the metabolomics study at 12 months of follow up (time point 1), where Group 1 (n = 24) continued for another 6 months in a self-sustained MedDiet intervention, and Group 2 (n = 24), the control group at that time, started with an active MedDiet intervention for 6 months (time point 2). A total of 225 metabolites were assessed at the two time points by using a targeted NMR platform. The MedDiet score improved slightly but changes were non-significant (intervention: 24.2 vs. 26.0 points and control group: 26.1 vs. 26.5 points). The MedDiet intervention led to favorable changes in biomarkers related to lipid metabolism, including lower LDL-C, ApoB/ApoA1 ratio, remnant cholesterol, M-VLDL-CE; and higher HDL-C, and better lipoprotein composition. This MedDiet intervention induces only modest changes in adherence to the MedDiet and consequently in metabolic biomarkers. Further research should confirm these results based on larger study samples in workplace interventions with powerful study designs.     

Effects of Paraquat on the Freshwater Fish Channa punctata (Bloch): Non-Enzymatic Antioxidants as Biomarkers of Exposure

Paraquat is a quaternary herbicide widely used for broadleaf weed control, which has been known to be a highly toxic compound for humans and animals. Therefore, analysis and development of biomarkers of exposure are undoubtedly valuable in evaluating the toxicity of paraquat contaminated water bodies. The effect of a single exposure of paraquat (1 ppm) for 24 h on various non-enzymatic antioxidants was studied in freshwater fish Channa punctata (Bloch). The levels of the reduced glutathione were significantly reduced in the liver and gill of exposed fish. The total and protein thiol levels were increased in all the tissues of the exposed fish. The non-protein thiol levels were reduced in liver and gill. The levels of ascorbic acid increased in liver. The uric acid level increased significantly in kidney and decreased significantly in gill of the exposed fish. The findings of the present investigation demonstrate the oxidative-stress-inducing potential of the herbicide paraquat in fish. This work indicated the possibility of using non-enzymatic antioxidants as biomarkers of exposure to environmental contamination and subsequent validation as a sensitive system for biomonitoring and ecotoxicological risk assessment.     

Biomarkers for virus-induced hepatocellular carcinoma (HCC)

Hepatocellular carcinoma (HCC) is one of the most common cancers worldwide, and is advanced by severe viral hepatitis B or C (HBV or HCV) as well as alcoholic liver disease. Many patients with early disease are asymptomatic therefore HCC is frequently diagnosed late requiring costly surgical resection or transplantation. The available non-invasive detections systems are based on the clinical utility of alpha fetoprotein (AFP) measurement, together with ultrasound and other more sensitive imaging techniques. The hallmark of liver disease and its propensity to develop into fully blown HCC is depended on several factors including the host genetic make-up and immune responses. While common symptoms involve diarrhea, bone pain, dyspnea, intraperitoneal bleeding, obstructive jaundice, and paraneoplastic syndrome, the evolution of cell and immune markers is important to understand viral induced liver cancers in humans. The circulating miRNA, cell and immune based HCC biomarkers are imperative candidates to successfully develop strategies to restrain liver injury. The current molecular genetics and proteomic analysis have lead to the identification of number of key biomarkers for HCC for earlier diagnosis and more effective treatment of HCC patients. In this review article, we provide latest updates on the biomarkers of HBV or HCV-associated HCC and their co-evolutionary relationship with liver cancer.