Short communication: Staphylococcus aureus infection modulates expression of drug transporters and inflammatory biomarkers in mouse mammary gland.

Mastitis is the most common disease in dairy herds worldwide and is often caused by Staphylococcus aureus. Little is known about the effect of mastitis on transporters in the mammary gland and the effect on transporter-mediated secretion of drugs into milk. We studied gene expressions of ATP-binding cassette and solute carrier transporters in S. aureus-infected mammary glands of mice. On d 7 of lactation, NMRI mice were inoculated with 1,000 cfu of S. aureus in 2 mammary glands and with a saline vehicle in 2 control glands. Gene expression of the transporters, Bcrp, Mdr1, Mrp1, Oatp1a5, Octn1, and Oct1, and of Csn2, the gene encoding ß-casein, were determined in mammary glands at 72 h after treatment. As biomarkers of the inflammatory response gene, expressions of the cytokines Il6, Tnfα, and the chemokine Cxcl2 were measured. Despite a high individual variation between the 6 animals, some characteristic patterns were evident. The 3 inflammatory biomarkers were upregulated in all animals; Csn2 was downregulated compared with controls in all animals, although not statistically significantly. Both Mrp1 and Oatp1a5 were statistically significantly upregulated and Bcrp was downregulated. Gene expression of Bcrp followed the expression of Csn2 in each of the animals, indicating a possible co-regulation. The findings demonstrate that S. aureus infection has an effect on expression of drug transporters in the mammary gland, which may affect secretion of drugs into milk and efficacy of drug therapy.

Meeting report: international workshop on endocrine disruptors: exposure and potential impact on consumers health.

The French Agency for Food, Environmental and Occupational Health and Safety (Anses) hosted a two-day workshop on Endocrine Disruptors: Exposure and Potential Impact on Consumers Health, bringing together participants from international organizations, academia, research institutes and from German, Swedish, Danish and French governmental agencies. The main objective of the workshop was to share knowledge and experiences on endocrine disruptors (ED) exposure and potential impact on consumers' health, to identify current risk assessment practices and knowledge gaps and issue recommendations on research needs and future collaboration. The following topics were reviewed: (1) Definition of ED, (2) endpoints to be considered for Risk assessment (RA) of ED, (3) non-monotonic dose response curves, (4) studies to be considered for RA (regulatory versus academic studies), (5) point of departure and uncertainty factors, (6) exposure assessment, (7) regulatory issues related to ED. The opinions expressed during this workshop reflect day-to-day experiences from scientists, regulators, researchers, and others from many different countries in the fields of risk assessment, and were regarded by the attendees as an important basis for further discussions. Accordingly, the participants underlined the need for more exchange in the future to share experiences and improve the methodology related to risk assessment for endocrine disrupters.

Fate of nickel subsulfide during carcinogenesis studied by autoradiography and X-ray powder diffraction.

Sarcomas in mice were induced by i.m. and s.c. administration of 63Ni- and 35S-labeled nickel subsulfide (Ni3S2), and the fate of the Ni3S2 was studied in tumors and normal tissues during carcinogenesis. Whole-body autoradiography showed a gradual loss of solubilized 63Ni and 35S radioactivity from the site of injection. There was also a loss of nonsolubilized dust particles which appeared to be phagocytized by reticuloendothelial cells in the liver, spleen, and regional lymph nodes. Microautoradiography showed that the totally dominating radioactivity within both the 63Ni3S2- and the Ni3(35)S2-induced tumors was associated with dust particles. There was no specific or excessive localization of solubilized radioactivity in the tumors or in metastases (when present). Two patterns of localization of dust particles within the tumors were observed: one with particles concentrated in a central part of the tumor and one with the particles present in the periphery of the tumor. X-ray powder diffraction of the insoluble crystalline material in the tumors indicated that a conversion of the alpha Ni3S2 to alpha Ni7S6 and beta NiS had occurred.

Fungicide prochloraz induces oxidative stress and DNA damage in vitro.

Prochloraz is widely used in horticulture and agriculture, e.g. as a post-harvest anti-mold treatment. Prochloraz is a known endocrine disruptor causing developmental toxicity with multiple mechanisms of action. However, data are scarce concerning other toxic effects. Since oxidative stress response, with formation of reactive oxygen species (ROS), is a common mechanism for different toxic endpoints, e.g. genotoxicity, carcinogenicity and teratogenicity, the aim of this study was to investigate if prochloraz can induce oxidative stress and/or DNA damage in human cells. A cell culture based in vitro model was used to study oxidative stress response by prochloraz, as measured by the activity of the nuclear factor erythroid 2-related factor 2 (Nrf2), a key molecule in oxidative defense mechanisms. It was observed that prochloraz induced oxidative stress in cultured human adrenocortical H295R and hepatoma HepG2 cells at non-toxic concentrations. Further, we used Comet assay to investigate the DNA damaging potential of prochloraz, and found that non-toxic concentrations of prochloraz induced DNA damage in HepG2 cells. These are novel findings, contradicting previous studies in the field of prochloraz and genotoxicity. This study reports a new mechanism by which prochloraz may exert toxicity. Our findings suggest that prochloraz might have genotoxic properties.

In vitro screening of inhibition of PPAR-γ activity as a first step in identification of potential breast carcinogens.

Alcohol consumption and increased estrogen levels are major risk factors for breast cancer, and peroxisome proliferator-activated receptor γ (PPAR-γ) plays an important role in alcohol-induced breast cancer. PPAR-γ activity is inhibited by ethanol, leading to increased aromatase activity and estrogen biosynthesis ultimately leading to breast cancer. If other organic solvents inhibit PPAR-γ activity, they should also lead to increased oestrogen biosynthesis and thus be potential breast carcinogens. Ten commonly used hydrophilic organic solvents were first tested in a cell-based screening assay for inhibitory effects on PPAR-γ transactivation. The chemicals shown to inhibit PPAR-γ were tested with vectors encoding PPAR-γ with deleted AB domains and only the ligand-binding domain to rule out unspecific toxicity. Next, the effects on biosynthesis of estradiol, testosterone and oestrone sulphate were measured in the H295R steroidogenesis assay after incubation with the chemicals. Ethylene glycol, ethyl acetate, and dimethyl sulphoxide inhibited PPAR-γ transactivation in a dose-dependent manner. The inhibitory effect on PPAR-γ was specific for PPAR-γ since the AB domain of PPAR-γ was required for the inhibitory effect. In the second step, ethylene glycol significantly increased production of oestradiol by 19% (p < 0.05) and ethyl acetate inhibited production of testosterone (p < 0.05). We here show that screening of 10 commonly used organic solvents for the ability to inhibit PPAR-γ transactivation followed by a well-established steroidogenesis assay for production of sex hormones in exposed H295 R cells may provide a screening tool for potential breast carcinogens. This initial screening thus identified ethylene glycol and possibly ethyl acetate as potential breast carcinogens.

Observation of direct photons in central 158A GeV (208)P(208)b+Pb collisions

A measurement of direct photon production in 208Pb+208Pb collisions at 158A GeV has been carried out in the CERN WA98 experiment. The invariant yield of direct photons in central collisions is extracted as a function of transverse momentum in the interval 0.51.5 GeV/c. The result constitutes the first observation of direct photons in ultrarelativistic heavy-ion collisions. It could be significant for diagnosis of quark-gluon-plasma formation.

Alterations in renal heme biosynthesis during metal nephrotoxicity.

The regulation of the heme biosynthetic pathway in the kidney by various metals has been reviewed. In addition, a study on the effects of lead on renal heme biosynthesis after acute treatment of rats has been reported. Chronic low-level lead exposure in rats results in relatively small effects on renal heme biosynthetic pathway enzymes. After acute treatment of rats with lead, no effects on ALAD or UROS and mild, transitory effects on ALAS and ferrochelatase are observed. The intracellular binding of lead within intranuclear inclusion bodies in the proximal tubule cells and to high-affinity cytosolic lead-binding proteins probably protects sensitive subcellular systems, such as the heme pathway, from lead toxicity. Chronic exposure to methyl mercury results in increased urinary excretion of uro- and coproporphyrins in rats, mediated via inhibition of ferrochelatase and UROS and stimulation of ALAS. A tissue-specific inhibition of ALAD occurs in the kidney after treatment of rats with indium. Acute treatment of rats with nickel, platinum, tin, antimony, bismuth, and cobalt results in induction of heme oxygenase, followed by decreased microsomal heme content and ALAS stimulation in the kidney.

Methylmercury induced alterations in the nerve growth factor level in the developing brain.

Pre- and early postnatal stages in the development of the central nervous system (CNS) are very sensitive to the toxic effects of methylmercury. The influence of methylmercury on the level of nerve growth factor (NGF) during the development of CNS was studied. Sprague-Dawley rats were exposed indirectly throughout the fetal and suckling periods until weaning on postnatal day 25 (P 25) via their dams given methylmercury in the diet (3.9 mg/kg diet). In addition, after weaning offsprings were exposed directly to methylmercury via the diet until postnatal day 50 (P 50). The level of NGF was analyzed in cortical areas and in the septum with a sensitive enzyme immunoassay. The pups exposed to MeHg exhibited a 50% elevation in the level of NGF in the hippocampus on P 25 and P 50 compared to control animals. Concomitantly, the level of NGF decreased by 30% in the septum on P 25 and P 50, suggesting that the retrograde transport of NGF from hippocampus to septum could be affected by the exposure of methylmercury. The exact mechanism by which the low level of mercury is affecting the NGF concentration in the developing brain is yet unknown. The increase of NGF in the hippocampus and the decrease of NGF measured in the septum could reflect altered conditions for neurotrophic support in these areas of the brain as a result of the exposure to heavy metal. Thus, this finding might indicate a connection between exposure of heavy metals and neurodegeneration, such as that found in the basal forebrain in Alzheimer's disease.

Lead-induced inclusion bodies in rat kidney after perinatal treatment with lead and disulfiram.

The presence of inclusion bodies in renal proximal tubules was studied in rats exposed to lead and/or disulfiram (tetraethylthiuram disulfide). Pregnant rats were treated with only lead acetate (0.25% Pb in the drinking water), only disulfiram (0.1 mmol/kg p.o. twice a week) or with both lead acetate and disulfiram from day 1 of pregnancy and until the offspring were 4 weeks of age. After parturition the disulfiram was given s.c. directly to the offspring instead of to the dams. Treatment was discontinued at weaning and tissue samples from renal cortex were studied by electron microscopy. In lead-treated dams inclusions were present in nuclei of renal proximal tubule cells in the 3 segments with the highest incidence in the middle segment. Inclusions were also present in the cytoplasm. In the offspring, indirectly exposed to lead via the dams, inclusions were present in all 3 segments. No inclusions were present in control rats or in disulfiram-treated rats. Combined treatment with lead and disulfiram resulted in a marked decrease in the incidence of inclusion bodies both in the dams and in the offspring compared to in rats treated with only lead. Diethyldithiocarbamate, a major metabolite of disulfiram, forms a lipophilic complex with lead, and is known to cause pronounced effects on the tissue distribution of lead. The present investigation shows that lead inclusion bodies are formed in the offspring indirectly exposed to lead via the dams during gestation and lactation. Concurrent exposure to disulfiram reduces the incidence of inclusion bodies in renal proximal tubules, probably due to formation of a lead-dithiocarbamate complex.

Response of rat hepatocyte cultures to cadmium chloride and cadmium-diethyldithiocarbamate.

Cellular effects of cadmium (Cd) were studied in primary cultures of rat hepatocytes incubated with cadmium chloride (CdCl2) or cadmium-diethyldithiocarbamate (Cd(DTC)2), labelled with 109Cd. The lipid-soluble complex Cd(DTC)2 was rapidly taken up into the cells and a maximal concentration was reached after 4 h incubation. On the other hand, incubation with CdCl2 resulted in a slow, continuous accumulation of Cd for up to 20 h. Cd was found to be associated with proteins to a higher extent when added to the incubation medium as CdCl2 than when added as Cd(DTC)2, which in addition to differences in lipophilicity of the Cd compounds partly explains the differences in Cd uptake. Subcellular distribution studies showed that a significantly higher proportion of Cd was associated with the total particulates fraction in cells after incubation with Cd(DTC)2 compared to CdCl2 (32 and 19%, respectively). The activities of glutathione reductase and succinic dehydrogenase were inhibited to a similar extent by the 2 Cd compounds. Alcohol dehydrogenase was more strongly affected by CdCl2 than by Cd(DTC)2, although the uptake of Cd was 3-4 times higher in cells incubated with Cd(DTC)2 than in those incubated with CdCl2. The results from the present study show that DTC can increase the transport of Cd into the cell by complex formation with Cd. Compared to CdCl2 the Cd(DTC)2 complex was less toxic as indicated by the biochemical parameters used.

Immunomodulating effects after perinatal exposure to methylmercury in mice.

The influence of methylmercury on the developing immune system was studied in offspring from Balb/c mice exposed to 0, 0.5 or 5 mg Hg/kg as methylmercury in the diet. Dams were exposed for 10 weeks prior to mating, during gestation and lactation. Pups were exposed to mercury until day 15 of lactation, thereafter the pups were given control milk and control diet. Samples for mercury analysis were collected from the pups on days 22 and 50, and for immunological studies on days 10, 22 and 50. The exposure resulted in significantly increased total Hg concentrations in whole blood on day 22 and 50 in offspring from the 5 mg Hg/kg group, and in offspring from the 0.5 mg Hg/kg group on day 22. On day 50, blood mercury levels had decreased to background levels in the 0.5 mg Hg/kg group. Increased numbers of splenocytes and thymocytes were found in offspring from the 0.5 mg Hg/kg group. Flow cytometry analysis of thymocytes revealed increased numbers and altered proportions of lymphocyte subpopulations within the thymus in offspring from both of the exposed groups. The proliferative response of splenocytes to the B-cell mitogen LPS was increased in offspring from dams exposed to 5 mg Hg/kg, and the primary antibody response to a viral antigen was stimulated in pups from dams exposed to 0.5 mg Hg/kg. The present results indicate that placental and lactational transfer of mercury affects thymocyte development and stimulates certain mitogen- or antigen-induced lymphocyte activities in mice.

Cellular response after mobilization of metals by diethyldithiocarbamate in rat hepatocyte cultures.

Cellular effects and mobilization of metals by diethyldithiocarbamate (DTC) were studied in primary cultures of rat hepatocytes incubated with lead acetate (PbAc), lead-diethyldithiocarbamate (Pb(DTC)2), cadmium chloride (CdCl2), cadmium-diethyldithiocarbamate (Cd(DTC)2), mercuric acetate (HgAc) and methylmercuric chloride (MeHgCl). In cells pretreated with inorganic Pb, Cd and Hg, the cellular levels of Cd and Pb were somewhat decreased or unchanged after incubation with DTC, while the cellular concentration of Hg was increased. Cells preincubated with MeHgCl showed a marked decrease in cellular Hg concentration upon DTC treatment. In Pb(DTC)2-treated cells the Pb concentration was increased when incubated with DTC, while DTC caused a decrease in Cd concentrations of cells preincubated with Cd(DTC)2. The activity of alcohol dehydrogenase (ADH) in cells incubated with CdCl2, Cd(DTC)2 and HgAc was significantly decreased. In Hg-treated cells the ADH activity was further decreased after incubation with DTC, whereas the activity in Cd-treated cells recovered gradually after incubation with increasing concentrations of DTC. The activity of the enzyme delta-aminolevulinic acid dehydratase (ALAD) was significantly inhibited in cells treated with PbAc and Pb(DTC)2, but could be restored to 80% and to almost 100%, respectively, of control activity after incubation with DTC. It is suggested that, in the absence of excess DTC, a decomposition of Pb(DTC)2 takes place after penetration into the cell, resulting in inhibition of ALAD by the released Pb. In the presence of excessive amounts of DTC, the equilibrium is shifted towards Pb(DTC)2 and Pb in the complex form is not available for ALAD. These studies suggest that DTC decreases cellular effects of Pb and Cd despite unchanged or even increased cellular concentrations of the metals, while the antidotal efficacy of DTC on inorganic Hg toxicity seems to be of low value.

Protein binding of mercury in milk and plasma from mice and man--a comparison between methylmercury and inorganic mercury.

Inorganic mercury has previously been shown to be excreted to milk from plasma to a higher extent than methylmercury. Protein binding of mercury as methylmercury and inorganic mercury in whey and plasma from mouse and man was studied in order to get a better understanding of the transport of mercury into milk. Mice were administered a single i.v. dose of 0.25 mg Hg/kg body weight labelled with (CH3)203HgCl or 203HgCl2, resulting in 11 ng Hg/g milk and 38 ng Hg/g milk after 1 h, respectively. Milk and plasma from mice and man were also incubated with the respective radiolabelled compound (150 ng Hg/g milk or plasma). Casein, fat and whey fractions in milk from methylmercury treated mice were found to contain 11, 39 and 34%, respectively, and from inorganic mercury treated mice 31, 15 and 41%, respectively, of the total amount of mercury in milk. Serum albumin was a major mercury binding protein in whey and plasma from mice for both methylmercury and inorganic mercury, as demonstrated by FPLC gel filtration and anion-exchange chromatography and further characterised by SDS-PAGE for whey. In addition, anion-exchange chromatography indicated that inorganic mercury, but not methylmercury, in whey from mouse milk formed a dimer of serum albumin. The unbound fraction of mercury in whey and plasma from mice was very small (<0.7%), and somewhat higher in plasma and whey from man. It is concluded, that the unbound fraction in plasma cannot be a determining factor for the observed differences in milk excretion between the two mercury compounds. Instead, it is suggested that methylmercury and to some extent inorganic mercury are transferred from plasma into milk using albumin as a passive carrier.

Combined lead acetate and disulfiram treatment-induced alterations of glial fibrillary acidic protein (GFA) immunoreactive astrocytes in brain smears.

Dithiocarbamates are known to form lipid-soluble complexes with lead and greatly increase brain lead levels. The present study was undertaken to investigate whether lead acetate, when administered together with disulfiram (Antabuse, metabolite of dithiocarbamate) during development, would induce morphological changes in brain astrocytes. Female Sprague-Dawley rats were given 0.25% lead acetate in the drinking water from day one of pregnancy and this treatment was continued after birth until the litters were 4 weeks old. In addition, some dams received disulfiram in a dose of 0.1 mmol/kg p.o. twice weekly and after parturition this dose was given s.c. directly to the offspring twice a week. Lead acetate and disulfiram treatments were discontinued at weaning and animals sacrificed 3 weeks later. Samples of parietal cortex, hippocampal formation and cerebellar cortex were dissected out and smeared onto glass-slides and astrocytes were visualized in toto using immunohistochemistry with antibodies against glial fibrillary acid protein (GFA), enabling morphometric analysis with a computerized image analyser. Animals treated with lead acetate showed a minor increase in the size of the GFA-immunoreactive astrocytes in parietal cortex smears, while animals treated with disulfiram showed no difference in size or form compared to controls. However, in combined lead acetate and disulfiram-treated animals a profound increase in astrocyte size and an increase in the number of processes of the individual GFA-immunoreactive astrocytes could be demonstrated in parietal cortex. No significant changes were noted in GFA-immunoreactive astrocytes of hippocampal smears following the different treatments, while GFA-immunoreactive astrocytes in cerebellar cortex smears were significantly smaller and had reduced number or processes following the combined lead acetate and disulfiram treatment compared to lead acetate treatment or controls. It is concluded that combined exposure to lead acetate and disulfiram during development induces regionally specific changes in GFA-immunoreactive astrocyte morphology. Furthermore, the present study demonstrates the usefulness of smear preparations combined with computerized image analysis to study the morphology of GFA-immunoreactive astrocytes as an index of toxic effects in CNS.

Dithiocarbamate-induced redistribution and increased brain uptake of lead in rats.

The effects of dithiocarbamates on brain uptake and tissue distribution of 203Pb were studied in rats injected intravenously with 100 microCi of 203Pb (28.6 nmol/kg b.wt.) as lead acetate. The dithiocarbamates--sodium diethyldithiocarbamate (DEDTC), tetraethylthiuram disulphide (disulfiram), sodium dimethyldithiocarbamate (DMDTC) and tetramethylthiuram disulphide (thiram)--were administered perorally in doses of 2 mmol/kg. Half of the dose was given 2h before and half immediately after the injection of 203Pb. The concentration of 203Pb in tissues, plasma and erythrocytes, and in urine and feces was determined by gamma counting. Four hours after injection of 203Pb, the dithiocarbamate-treated rats had a significantly higher concentration of lead in brain, liver and lung and a significantly lower concentration of lead in kidney, femur and erythrocytes compared to controls. At 72 h the brain concentration of lead in rats administered thiram, DMDTC, disulfiram or DEDTC was 100, 15, 10, and 4 times higher respectively, than in controls. At 72 h these rats also had higher levels of lead in liver, lung and kidney than did controls. The kidneys of dithiocarbamate-treated rats had a higher concentration of lead at 72 h than at 4 h, and during this time the lead concentration in femur had increased in the treated groups as well as in the controls. Excretion was mainly via feces. In controls about 15% of the dose was excreted in feces at 72 h. Fecal excretion at 72 h in DEDTC and DMDTC-treated rats was about 25% and in thiram and disulfiram-treated rats about 7% of the dose. The total urinary excretion of lead at 72 h was about 9% of the dose in controls, 5% in the DEDTC and DMDTC-treated groups and about 1% in the thiram and disulfiram-treated groups. The results indicate that a lipid-soluble complex between lead and the dithiocarbamates is formed in vivo and that these complexes have a high capacity to penetrate the blood-brain barrier and be retained in brain due to binding to lipid rich brain constituents. The toxicity of the complexes is not known, but it is possible that there is intracellular release of inorganic lead after metabolism/decomposition of the complex.

Synthesis, Structure, and Reactivity of Arylchlorobis(dialkyl sulfide)platinum(II) Complexes.

Complexes trans-[PtRCl(SR'(2))(2)], where R = Ph, mesityl, and p-anisyl and R' = Me or Et, have been synthesized and their crystal and molecular structures determined. Crystals of trans-[PtPhCl(SEt(2))(2)] (2) are triclinic (P&onemacr;) with a = 10.112(6) Å, b = 13.158(2) Å, c = 14.714(5) Å, alpha = 102.48(2) degrees, beta = 94.394(4) degrees, gamma = 90.22(3) degrees, and Z = 4. Crystals of trans-[Pt(mesityl)Cl(SMe(2))(2)] (4) are monoclinic (P2(1)/c) with a = 13.158(2) Å, b = 9.170(1) Å, c = 16.013(3) Å, beta = 120.93(2) degrees, and Z = 4, and crystals of [Pt(p-anisyl)Cl(SMe(2))(2)] (5) are monoclinic (P2(1)/n) with a = 9.879(4) Å, b = 8.128(2) Å, c = 19.460(5) Å, beta = 96.56(3) degrees, and Z = 4. All complexes are square-planar, featuring Pt-Cl distances between 2.40 and 2.42 Å, indicating a large ground-state trans influence of the aryl group. The coordination geometry is maintained in methanol and chloroform solution as shown by (1)H-NMR spectra. The kinetics of substitution of the labile chloride trans to aryl by various nucleophiles has been studied in methanol by variable-temperature and -pressure stopped-flow spectrophotometry. A two-term rate law with a well-developed solvolytic pathway is followed. Negative entropies and volumes of activation indicate an associative mode of activation in all cases, independent of steric blocking of the axial sites and a large Pt-Cl ground-state bond-weakening. Comparison of the reaction rates of the present series of complexes with their bis(phosphine) analogues and with related cyclometalated compounds shows that the triethylphosphine complexes are 2-3 orders of magnitude less reactive than the thioether complexes, which in turn are a factor 10-20 less reactive than the cyclometalated ones. This reactivity increase can be rationalized mainly in terms of a decrease in steric hindrance in the series. There seems to be no inherent differences with regard to trans labilizing ability of the aryl ligands in the various types of complexes, including the cyclometalated ones.

Increased lead uptake and inhibition of ALAD-activity in isolated rat hepatocytes incubated with lead-diethyldithiocarbamate complex.

Dithiocarbamates can form lipid soluble complexes with lead and are known to markedly increase tissue uptake of lead and potentiate toxic effects of lead in rats. Cellular effects of the interactions between lead and diethyldithiocarbamate were studied in primary cultures of rat hepatocytes. The cells were incubated with lead acetate (PbAc) or lead-diethyldithiocarbamate complex (Pb(DTC)2), labelled with 203Pb. The lipid soluble Pb(DTC)2 was rapidly taken up in the cells and after 30 min incubation the cellular levels of lead were approximately 40 times higher in cells incubated with Pb(DTC)2 than in cells incubated with a similar concentration of PbAc. The maximal cellular uptake of lead was reached after 4 h incubation with Pb(DTC)2, while incubation with PbAc caused a slow continuously increasing uptake of lead during the 20 h incubation. The enzyme delta-aminolevulinic acid dehydratase (ALAD) was inhibited to a much higher extent by Pb(DTC)2 compared to PbAc after incubations with similar concentrations of lead. Maximal inhibition of ALAD activity was reached at a cellular concentration of 0.5-1 nmol Pb/mg protein, irrespective of which form of lead was used in the incubation. Pb(DTC)2 was shown to inhibit ALAD activity also in vitro when incubated with purified ALAD enzyme. The rapid and high intracellular uptake and cellular response of Pb(DTC)2, shown in the present study, may explain the drastic effects of dithiocarbamates on lead distribution and toxicity previously shown in vivo.

Effect of disulfiram on milk transfer and tissue distribution of lead in the neonatal rat.

The effect of disulfiram (tetraethylthiuram disulphide) on uptake and tissue distribution of 203Pb was studied in neonatal rats exposed to lead via dams' milk. In the dams, treatment with disulfiram greatly increased the 203Pb concentration in brain and liver and decreased 203Pb concentration in plasma and erythrocytes, as compared to controls given only 203Pb. However, in the pups of disulfiram-treated dams total uptake of 203Pb was reduced by 50% and the concentration of 203Pb in brain and liver was significantly reduced. Consistent with these findings, the level of 203Pb in the maternal milk was lower in the disulfiram-treated dams compared to controls. It is suggested, that the lower lactational transfer and uptake of lead in the neonatal rat after treatment of the dams with disulfiram is due to retention and strong binding of lead to tissue components in the dams after formation of a lead-dithiocarbamate complex.

Increased availability of mercury in rat hepatocytes by complex formation with diethyldithiocarbamate.

The availability of different chemical forms of mercury (Hg) was studied in primary cultures of rat hepatocytes incubated with mercuric acetate (HgAc), mercuric diethyldithiocarbamate (Hg(DTC)2) or methyl mercuric chloride (MeHgCl), labelled with 203Hg. The uptake of Hg was linearly related to the concentration in the medium and increased in the order Hg(DTC)2 greater than MeHgCl greater than HgAc when similar concentrations of Hg were used. A maximum concentration of Hg was reached after 4 h incubation with Hg(DTC)2 while incubation with HgAc and MeHgCl resulted in a slow continuous accumulation of Hg for up to 24 h. Hg added as HgAc was bound to proteins in the incubation medium to a greater extent than Hg added as Hg(DTC)2 or MeHgCl. Differences in affinity to the medium, as well as in lipophilicity, may partly explain the observed differential uptake of these Hg compounds. The enzyme alcohol dehydrogenase was inhibited by HgAc and Hg(DTC)2 to a similar extent at comparable cellular concentrations of Hg. On the other hand, glutathione reductase was inhibited to a higher degree by HgAc than by Hg(DTC)2, indicating that Hg(DTC)2 remains at least temporarily in the complexed form and that the enzyme is less susceptible to Hg in this form. Both enzymes were much less susceptible to MeHgCl than to HgAc or Hg(DTC)2. The results from the present study indicate that diethyldithiocarbamate can increase the transport of Hg across the cellular membrane by complex formation with Hg and thereby increase the toxicity of Hg.

Methyl mercury exposure via placenta and milk impairs natural killer (NK) cell function in newborn rats.

The effect of methyl mercury (MeHg) exposure (3.9 micrograms/g diet) on the development of immune function was studied in the newborn Sprague-Dawley rat after MeHg exposure via placenta and/or milk. No consistent alterations were observed between control and treated offspring (at the age of 15 days) on the following parameters: body weights, lymphoid organ weights or cell number, and the lymphoproliferative response to B-cell mitogen. The lymphoproliferative response to T-cell mitogen was increased in thymocytes (by 30-48%), but decreased in splenocytes (by 30-32%). This decreased activity was only observed in the groups exposed during lactation. White blood cell counts (WBC) were increased in all groups. Natural killer (NK) cell activity was reduced (by 42%, P less than 0.01) in the group that was exposed both via placenta and milk. These results indicate that placental and lactational transfer of MeHg does adversely affect the developing immune system of the rat.