Individual and combined cytotoxic effects of cadmium, copper, and nickel on brown cells of Mercenaria mercenaria

An assay based on the lysosomal incorporation of neutral red dye by brown cells of Mercenaria mercenaria (Bivalvia) was used to measure the cytotoxicity of Cd2+, Cu2+, and Ni2+ singly and in combination. Cytotoxicity was a linear function of Cd2+ concentration 0.1-1.5 mM and Cu2+ concentration between 10 and 100 microM. Nickel was not cytotoxic at concentrations as high as 10 mM. The presence of Cu2+ lessened the cytotoxic effect of Cd2+. Ni2+ did not affect cytotoxicity in combination with either Cu2+ or Cd2+.Ni2+ inflated estimates of cell survival by the neutral red assay in this study. Cells exposed to Ni2+ yielded measured quantities of neutral red dye in excess of those measured in cells from control treatments.     

Exposure of human proximal tubule cells to cd2+, zn2+, and Cu2+ induces metallothionein protein accumulation but not metallothionein isoform 2 mRNA.

The organization of the human metallothionein (MT) gene family is more complex than the commonly used mouse and rat models. The human MTs are encoded by a family of genes consisting of 10 functional and 7 nonfunctional MT isoforms. One objective of this study was to determine if the accumulation of MT protein in cultures of human proximal tubule (HPT) cells exposed to metals is similar to that expected from the knowledge base obtained from rodent models. To accomplish this objective, HPT cells were exposed to both lethal and sublethal concentrations of Cd2+, Zn2+, Cu2+, Ag2+, Hg2+, and Pb2+ and MT protein levels were determined. The results were in general agreement with animal model studies, although there were some exceptions, mainly in areas where the animal model database was limited. In clear agreement with animal models, Cd2+, Zn2+, and Cu2+ were demonstrated to be potent inducers of MT protein accumulation. In contrast to the similarity in MT protein expression, we obtained evidence that the human renal MT-2 gene has a unique pattern of regulation compared to both animal models and human-derived cell cultures. In the present study, we determined that MT-2A mRNA was not induced by exposure of HPT cells to Cd2+ or the other metals, a finding in contrast to studies in both animal models and other human cell culture systems in which a high level of MT-2 mRNA induction occurs upon exposure to Cd2+ or Zn2+. While MT protein expression may be similar between humans and animal models, this finding provides initial evidence that regulation of the genes underlying MT protein expression may be divergent between species.     

Rabbit lung after inhalation of manganese chloride: a comparison with the effects of chlorides of nickel, cadmium, cobalt, and copper

Rabbits were exposed to aerosols of MnCl2 (mass median aerodynamic diameter 1 micron) in metal concentrations of 1.1 and 3.9 mg/m3 for 4-6 weeks, 5 days/week, 6 h/day. The effects of alveolar type II cells, phospholipids, alveolar macrophages, and lung structure in general were compared with earlier reported effects of Ni2+, Cd2+, Cu2+, and Co2+. Except for a significant increase in the diameter of the alveolar macrophages after exposure to the higher Mn2+ concentration, no abnormalities were seen. The results of this and earlier studies indicate that these five metal ions have different, specific effects on the alveolar part of the lung.     

Protective effects of Ca2+, Mg2+, Cu2+, and Ni2+ on mercury and methylmercury toxicity to a cyanobacterium.

Toxicological investigations of the impact of inorganic mercury (Hg2+) and methylmercury (CH3Hg+) in terms of growth, NH4+ uptake, in vivo glutamine synthetase (transferase) activity, and regulation of toxicity by Ca2+, Mg2+, Cu2+ and Ni2+ in the diazotrophic cyanobacterium Nostoc calcicola Bréb. have been completed. Photoautotrophic growth of the cyanobacterium was extremely sensitive to both mercury compounds, CH3Hg+ being 2.5 times more toxic than Hg2+. Although NH4+ uptake was 6 times more sensitive than in vivo GS activity against the two mercurials, both processes had a greater susceptibility toward CH3Hg+. On the basis of Km and Vmax, it is suggested that both mercury species inhibit such metabolic events noncompetitively. Ca2+, Mg2+, Cu2+, and Ni2+ did not change the nature of inhibition and effectively antagonized the Hg2+ and CH3Hg+ toxicities in the sequence Ca2+ greater than Mg2+ much greater than Cu2+ greater than Ni2+.     

Effects of the heavy metals Cu2+, Ni2+, Pb2+, and Zn2+ on some physiological parameters of the lichen Usnea amblyoclada

The effect of Cu2+, Ni2+, Pb2+, and Zn2+ on some physiological parameters of the lichen Usnea amblyoclada and the selective uptake of Cu2+ and Pb2+ was assessed. Fresh thalli were soaked in single or mixed metallic solutions. The concentration of chlorophylls and malondialdehyde; the dry weight/fresh weight ratio as well as the water content and the concentration of Cu, Ni, Pb, and Zn were measured in the treated and control thalli. The exposure to Cu, Ni, and Pb solutions caused several changes on the parameters measured; no differences were found with Zn. A stronger ability for binding Pb2+ was also observed. The results suggest that Cu2+ was the most harmful cation followed by Pb and Ni. Consequently, the damage observed in U. amblyoclada thalli when it is used as a biomonitor in polluted areas is possibly due to the presence of these heavy metals, masking the effect of other gaseous pollutants.     

Effect of cadmium on transmembrane Na+ and K+ transport systems in human erythrocytes.

The effects of cadmium (Cd2+) on Na+,K(+)-ATPase in disrupted human erythrocyte membranes and on various transmembrane Na+ and K+ transport systems in intact erythrocyte suspensions were studied. Cadmium2+ inhibited the erythrocyte Na+,K(+)-ATPase enzyme with a 50% inhibition at a Cd2+ concentration of 6.25 microM. The Cd2+ inhibition in the human erythrocyte was non-competitive with respect to Na+,K+, and ATP. Cadmium2+ exerted no acute effect, however, on the Na+,K(+)-ATPase pump activity as measured by the ouabain sensitive 86Rb uptake or Na+ efflux in intact red blood cells. Cadmium2+ also inhibited the Ca2+ dependent K+ channels in human red blood cells, whereas it had no effect on Na+,K+ cotransport, Na+,Li+ countertransport, anion carrier, and the number of active Na+ pump units. The data indicate that in human erythrocytes under acute conditions Cd2+ exerts an inhibitory effect on Na+,K(+)-ATPase enzyme in disrupted erythrocytes and the Ca2+ stimulated K+ efflux in intact red blood cells without affecting the Na+ pump, Na+,K+ cotransport, and Na+,Li+ countertransport activity.     

Effect of cadmium on transmembrane Na+ and K+ transport systems in human erythrocytes.

The effects of cadmium (Cd2+) on Na+,K(+)-ATPase in disrupted human erythrocyte membranes and on various transmembrane Na+ and K+ transport systems in intact erythrocyte suspensions were studied. Cadmium2+ inhibited the erythrocyte Na+,K(+)-ATPase enzyme with a 50% inhibition at a Cd2+ concentration of 6.25 microM. The Cd2+ inhibition in the human erythrocyte was non-competitive with respect to Na+,K+, and ATP. Cadmium2+ exerted no acute effect, however, on the Na+,K(+)-ATPase pump activity as measured by the ouabain sensitive 86Rb uptake or Na+ efflux in intact red blood cells. Cadmium2+ also inhibited the Ca2+ dependent K+ channels in human red blood cells, whereas it had no effect on Na+,K+ cotransport, Na+,Li+ countertransport, anion carrier, and the number of active Na+ pump units. The data indicate that in human erythrocytes under acute conditions Cd2+ exerts an inhibitory effect on Na+,K(+)-ATPase enzyme in disrupted erythrocytes and the Ca2+ stimulated K+ efflux in intact red blood cells without affecting the Na+ pump, Na+,K+ cotransport, and Na+,Li+ countertransport activity.     

Divalent metal inhibition of non-haem iron uptake across the rat duodenal brush border membrane

Duodenal Fe2+ uptake is essential to body Fe2+ homeostasis, but the interaction of metals with the uptake process remains unclear. The present study compared the effects of four essential trace metals (Mn2+, Zn2+, Co2+ and Ni2+) with two toxic metals (Pb2+ and Cd2+) on Fe2+ uptake across the brush border membrane of villus-attached duodenal enterocytes. Everted rat duodenum was exposed to buffer containing 0.2 mm-59Fe2+-ascorbate with or without the competing metal (2 mm) and the tissue was then processed for autoradiography allowing Fe2+ uptake to be determined at specific crypt-villus regions. The quantification method ensured that uptake by cells, rather than Fe2+ binding to the tissue surface, was measured. Fe2+ uptake was significantly inhibited by Cd2+ in upper villus enterocytes only and Pb2+ was without effect on Fe2+ uptake. The inhibition by Cd2+ was not due to general cell damage as judged by the release of lactate dehydrogenase from tissue into incubation fluid. Essential divalent trace metals reduced uptake significantly along the whole length of the crypt-villus axis. Cd2+ uptake, measured separately, took place at all regions of the villus-crypt axis, highest uptake being into crypt enterocytes. The very different uptake profiles for Cd2+ and Fe2+ suggests that the divalent metal transporter 1 is not the principal transporter of Cd2+. The addition of Fe2+ to incubation buffer inhibited Cd2+ uptake by both crypt and villus enterocytes. The possibility that the inhibitory actions of Fe2+ and Cd2+ on the uptakes of Cd2+ and Fe2+ respectively can be explained by a non-competitive action or the involvement of an additional metal transporter is discussed.     

Toxicity of heavy metals using sperm cell and embryo toxicity bioassays with Paracentrotus lividus (Echinodermata: Echinoidea): comparisons with exposure concentrations in the Lagoon of Venice,Italy

Sperm cell and embryo toxicity tests using the Mediterranean sea urchin Paracentrotus lividus Lmk were performed to assess the toxicity of As3+, Cd2+, Cr3+, Ni2+, Pb2+, Cu2+, Zn2+, and Hg2+. The aim of this study was to improve information about the comparative sensitivity of sea urchin bioassays to the heavy metals, which are an important cause of contamination in the ecosystem of the Lagoon of Venice. Considering the data in mM/L, the order of toxicity is Hg2+ > Cu2+ > Zn2+ > As3+ > or = Cr3+ > or = Cd2+ > or = Pb2+ > or = Ni2+ for the sperm cell test and Hg2+ > or = Pb2+ > Cu2+ > Zn2+ > Cd2+ > Ni2+ > As3+ > or = Cr3+ for the embryo test. New toxicity data for metals expressed as median effective concentration (EC50) and no-observed-effect concentration (NOEC) are reported for the Mediterranean species. Accurate observations of embryotoxic effects at increasing metal concentrations were done, detecting some different behaviors in metal toxicity. Toxicity data compared with water column and pore-water concentrations recorded in the Lagoon of Venice (Italy) demonstrate the potential ability of bioassays using sea urchin to detect important contaminants in this ecosystem.     

镉、铅、乐果和对硫磷雌激素样联合作用初步研究

目的探讨镉、铅、乐果和对硫磷对未成年雌性小鼠雌激素样联合作用。方法选择断乳后17天龄SPF级雌性昆明小鼠192只,体重(25±2)g,随机分为16个组:空白对照组,氯化镉、醋酸铅、乐果和对硫磷组单独作用12个组,联合作用3个组。给药浓度为氯化镉、醋酸铅、乐果和对硫磷LD50(分别为9.3、140、45和3mg/kg)的1/20、1/50及1/100,给药体积为10ml/kg BW,每天腹腔注射,连续3天。实验第4天,处死小鼠,测量子宫脏器系数、子宫上皮细胞高度及子宫雌激素受体(ER)表达。结果氯化镉1/50LD50、醋酸铅1/50LD50及联合作用1/100LD50剂量组能显著改变子宫脏器系数;氯化镉各剂量组、醋酸铅1/20LD50剂量组和联合作用的1/20LD50、1/50LD50剂量组可引起未成年雌性小鼠子宫上皮细胞高度的显著增加,即引起子宫内膜的明显增生;除氯化镉和乐果的1/20LD50、1/50LD50剂量组其他所有实验组与空白对照比较均能显著增强雌激素受体(ER)的表达。结论镉、铅、乐果和对硫磷对未成年雌性小鼠雌激素样联合作用与单因素作用存在显著差异,且在不同剂量不同指标中表现不尽相同。     

The effects of some divalent metals on cardiac and branchial Ca2+-ATPase in a freshwater fish Saccobranchus fossilis

The heart and gill of a freshwater fish Saccobranchus fossilis have been shown to contain a Ca2+-activated ATPase involved in Ca2+ transport. Enzyme showed optimal activity at 3 mM Ca2+ and 3 mM ATP for gills and at 3 mM Ca2+ and 1 mM ATP for heart. Mg2+ was equally effective in stimulating enzyme activity but was not essential for hydrolysis. Maximum activity was found in heart ventricular muscles as compared to gills. Among all the metals tested Hg2+ was the most toxic (IC50, 0.75 and 0.85 microM for heart and gill, respectively) followed by Pb2+, Mn2+, and Cd2+. The inhibition was concentration dependent and reached almost 100% with each metal at the highest concentration. Stimulation of enzyme activity was observed at lower concentrations of Mn2+ and Cd2+ but not with Pb2+ and Hg2+. Stimulation was more pronounced with Mn2+ than with Cd2+ in both heart and gills. The results indicated that the inhibitory effect of these metals might be through the Ca2+-ATPase which is a manifestation of the calcium pump in various tissues.     

Heavy Metal Resistant of E. coli Isolated from Wastewater Sites in Assiut City, Egypt

Twelve isolates of E. coli were isolated from wastewater of El-Malah canal located in Assiut, Egypt and were checked for their heavy metal tolerance. One isolate has tested for its multiple metal resistances and found to be plasmid mediated with molecular weights 27 and 65 kb for hexa- and trivalent chromium. It was identified as E. coli ASU 7. Its minimal inhibitory concentration (MIC) for Cu(2+), Co(2+), Ni(2+), Zn(2+), Cr(6+), Cr(3+), Cd(2+) and Pb(2+) were 1.57, 2.55, 1.7, 9.17, 0.48, 7.69, 4.4 and 3.1 mM, respectively. Growth kinetics was determined under Cr(6+) and Cr(3+) stress. SDS-PAGE of protein profile shows that 10 ppm (0.19 mM) of Cr(6+) induces new protein with molecular weight 23 kDa.     

Effects of temperature on the sensitivity of sludge worm Tubifex tubifex Müller to selected heavy metals

The present study evaluates the effect of temperature on the sensitivity of the freshwater tubificid sludge worm Tubifex tubifex Müller to 10 heavy metal ions. Metals used in this study were cadmium, chromium, cobalt, copper, iron, lead, manganese, mercury, nickel, and zinc. The acute toxicity of these heavy metals was studied at 15, 20, 25, and 30 degrees C. The percentage mortality, relative toxicity, and EC50 values and their 95% confidence limits from 24 to 96 h were determined at varying temperatures. The EC50 values (mg/liter) of metal ions at 15 degrees C were Hg2+, 0.034; Cu2+, 0.340; Cr6+, 1.846; Zn2+, 10.99; Ni2+, 25.10; Cd2+, 56; Fe3+, 86.09; Co2+, 239.39; Pb2+, 456.76; and Mn2+, 164.55. At 30 degrees C the values were Hg2+, 0.014; Cu2+, 0.031; Cr6+, 0.872; Zn2+, 3.37; Ni2+, 18; Cd2+, 28.55; Fe3+, 71.26; Co2+, 95.35; Pb2+, 165.22; and Mn2+, 239.39. The results indicate that the acute toxicity of cadmium, chromium, cobalt, copper, lead, mercury, nickel, and zinc increases with temperature increase. The toxicity of manganese was not influenced by temperature, and temperature had little effect on iron toxicity. The rank order of toxicity of metal ions at 15, 20, 25, and 30 degrees C is presented and discussed. It is concluded that temperature is an important factor in short-term acute toxicity tests. The study indicates that seasonal temperature changes are an important variable in determining the amount of heavy metals that may be safely released from metal industries and other similar sources into the aquatic environment. Influence of temperature on the short- and long-term toxicity of chemicals should be considered for establishing appropriate water-quality criteria and standards to protect aquatic flora and fauna and human health.     

Interactions between cadmium and radiation in the killing of Escherichia coli

CdCl2 (10 microM) added to the growth medium caused an extension to the lag phase in the growth of Escherichia coli. Radiation sensitivity in the extended lag phase was increased, particularly at low radiation doses. However, exposure to Cd2+ did not affect log-phase cultures which are normally more sensitive than lag-phase cultures. The increase in radiation resistance normally seen in stationary phase was delayed and reduced. When the cells had adapted to growth in the Cd2+-containing medium, further Cd2+ exposure was ineffective unless a Cd2+-free medium was provided for several hours. Prior growth in 25 microM Zn2+ partially protected against the effects of Cd2+ on growth and radiation sensitivity.     

胃癌组织中CD4+、CD25+调节性T淋巴细胞的表达及意义

目的 探讨CD4+、CD25+调节性T淋巴细胞在胃癌患者胃组织中的表达及意义.方法 利用流式细胞仪对76例胃癌及癌旁组织中的CD4+、CD25+调节性T淋巴细胞与CD8+T淋巴细胞进行定量及量化关系分析.结果 CD4+、CD25+调节性T淋巴细胞的阳性率在胃癌、癌旁组织、正常时照组分别为(11.2±0.9)%、(6.4±1.1)%、(4.1±0.8)%.癌组织与癌旁组织和对照组比较,差异有统计学意义(t=2.03、2.05,P<0.05).在肿瘤组织,随着CD4+、CD25+调节性T淋巴细胞的增加,CD8+T淋巴细胞出现了快速减少趋势.而在癌旁组织中却没有此现象出现.结论 CD4+、CD25+调节性T淋巴细胞通过对CD8+T淋巴细胞的抑制参与胃癌细胞抗肿瘤免疫的作用.     

Nickel speciation in the presence of different sources and fractions of dissolved organic matter

This study evaluated nickel (Ni) speciation in the presence of different fractions (humic acid (HA), fulvic acid (FA)) and sources (natural sediment, Suwannee River, peat moss) of dissolved organic matter (DOM) at Ni concentrations toxicologically relevant to the freshwater amphipod, Hyalella azteca. The free Ni ion, Ni(2+), was measured in reconstituted water (with or without DOM) using a miniaturized ion-exchange technique (IET). Ni speciation from these experiments was compared to calculated results obtained from equilibrium modelling (WHAM, Model VI). While it is known that Ni will complex with DOM, it was found that under acutely toxic Ni exposure concentrations ([Ni(Total)]=5mg/L, or 85.1 microM) representative surface-water DOC concentrations ( approximately 10mg/L) played little or no role in Ni speciation. Conversely, at sublethal Ni exposure concentrations ([Ni(Total)]=0.2 and 0.5 microg/L, or 3.4 and 8.51 microM, respectively) DOM significantly affected Ni speciation with [Ni(2+)] decreasing with increasing concentration of DOM. It was found that for similar concentrations of DOC (same fraction, different sources), the measured Ni(2+) concentrations were reduced (relative to the control), but similar to one another. Conversely, at similar DOC concentrations, the HA fraction reduced Ni(2+) levels to a greater extent than the associated FA fraction. Overall, this study provides proof of principle that Suwannee River and peat humic substances are suitable analogues for natural sediment pore-water DOM when evaluating Ni bioavailability in freshwater.     

Heat shock protein 27 expression in human proximal tubule cells exposed to lethal and sublethal concentrations of CdCl2.

The expression of hsp 27 mRNA and protein was determined in cultured human proximal tubule (HPT) cells exposed to lethal and sublethal concentrations of Cd2+ under both acute and extended conditions. Initial procedures demonstrated that HPT cells display the classic stress response following physical and chemical stress. Heat stress (42.5 degrees C for 1 hr) caused an increase in both hsp 27 mRNA and protein as well as a shift in the protein to a more phosphorylated state. Results were similar when the cells were subjected to chemical stress (exposure to 100 microM sodium arsenite for 4 hr). Acute exposure to 53 microM CdCl2 for 4 hr also resulted in an increase in hsp 27 mRNA and protein and a shift to the more phosphorylated protein isoform. Extended Cd2+ exposure involved continuous treatment with Cd2+ at both lethal and sublethal levels over a 16-day time course. The results of this treatment showed that chronic exposure to Cd2+ failed to increase either hsp 27 mRNA or protein expression in HPT cells, even at lethal Cd2+ concentrations. In fact, hsp 27 protein levels decreased as compared to controls at both lethal and sub-lethal exposure to Cd2+. These findings imply that hsp 27 expression in human proximal tubule cells may have two distinct modes depending on the nature (acute vs. chronic) of the stress.     

Chemopreventive properties of trans-resveratrol against the cytotoxicity of chloroacetanilide herbicides in vitro

The beneficial effect of trans-resveratrol (RESV) on health is well documented. Our aim was to study the putative preventive effect of RESV on the cytotoxicity of frequently used herbicides (alachlor, acetochlor). Estrogen receptor positive (ER+) MCF-7 human mammary carcinoma, HepG2 (ER+) human hepatocellular carcinoma and VERO estrogen receptor negative (ER-) non-transformed monkey fibroblast cell lines were treated with alachlor and acetochlor (2-500 microg/ml) as toxic agents, and RESV (10 microM) as preventive agent. The MTT dye reduction assay was performed to test cytotoxicity, and flow cytometry to test cell proliferation and apoptosis. RESV is not cytotoxic in the concentration range of 1-100 microM on neither cell lines examined after 24 h, but cytotoxic on Vero and MCF-7 cells at 100 microM after 48h, and on all three cell lines after 72 h. On both ER+ cell lines a stimulation of viability occurs in the low concentration range (0.5-12.5 microM) as detected by the MTT assay. Cell cycle analysis of the culture shows a significant increase of S-phase cells at low concentrations of RESV (10-50 microM) and a decrease in the 100-200 microM concentration range. The ratio of apoptotic cells significantly increases after the administration of 50 microM RESV, depending on the incubation time. The cytotoxicity of 20-65 microg/ml alachlor and 10-65 microg/ml acetochlor was significantly decreased by the addition of 10 microM RESV in Vero ER- cells whereas no significant change was detected on ER+ cell lines MCF-7 and HepG2. These results show that RESV protects non-transformed ER- cells, but has no such effect on ER+ tumor cells.     

Evaluation of metallothionein as a biomarker of single and combined Cd/Cu exposure in Dreissena polymorpha

The effects of metal mixture (Cd+Cu) versus single-metal exposure on total MT response and bioaccumulation were investigated in the freshwater bivalve Dreissena polymorpha. A two-month exposure period, including two levels of contamination, was chosen for each of the two metals: 5, 10 microg/L for Cu, and 2, 20 microg/L for Cd, with mixtures of, respectively, 5 microg/L Cu+2 microg/L Cd, 5 microg/L Cu+20 microg/L Cd, 10 microg/L Cu+2 microg/L Cd, and 10 microg/L Cu+20 microg/L Cd. Total MT contents were assessed by an Ag-saturation method, and metals contents were determined by atomic absorption spectrometry.Results at the whole-organism level showed a significant and early increase of total MT biosynthesis after exposure to Cd. This increase was significantly correlated with Cd bioaccumulation. By contrast, Cu did not modify total MT response, and mussels limited Cu bioaccumulation. The mixture either did not influence or only weakly influenced metal accumulation and MT response to Cu and Cd after long-term exposure. Our results suggest that the form of MT existing in D. polymorpha was not Cu-inducible. This could limit the use of MT in D. polymorpha as a biomarker of heavy metal pollution in freshwater ecosystems.