Vanillin-coumarin hybrid molecule as an efficient fluorescent probe for trace level determination of Hg(II) and its application in cell imaging.

An efficient Hg(2+) selective fluorescent probe (vanillin azo coumarin, VAC) was synthesized by blending vanillin with coumarin. VAC and its Hg(2+) complex were well characterized by different spectroscopic techniques like (1)H NMR, QTOF-MS ES(+), FTIR and elemental analysis as well. VAC could detect up to 1.25 µM Hg(2+) in aqueous methanol solution through fluorescence enhancement. The method was linear up to 16 µM of Hg(2+). Negative interferences from Cu(2+), Ni(2+), Fe(3+), and Zn(2+) were eliminated using EDTA as a masking agent. VAC showed a strong binding to Hg(2+) ion as evident from its binding constant value (2.2×10(5)), estimated using Benesi-Hildebrand equation. Mercuration assisted restricted rotation of the vanillin moiety and inhibited photoinduced electron transfer from the O, N-donor sites to the coumarin unit are responsible for the enhancement of fluorescence upon mercuration of VAC. VAC was used for imaging the accumulation of Hg(2+) ions in Candida albicans cells.

L-carnitine protects against nickel-induced neurotoxicity by maintaining mitochondrial function in Neuro-2a cells.

Mitochondrial dysfunction is thought to be a part of the mechanism underlying nickel-induced neurotoxicity. L-carnitine (LC), a quaternary ammonium compound biosynthesized from the amino acids lysine and methionine in all mammalian species, manifests its neuroprotective effects by improving mitochondrial energetics and function. The purpose of this study was to investigate whether LC could efficiently protect against nickel-induced neurotoxicity. Here, we exposed a mouse neuroblastoma cell line (Neuro-2a) to different concentrations of nickel chloride (NiCl2) (0.25, 0.5, 1, and 2 mM) for 24 h, or to 0.5 mM and 1 mM NiCl2 for various periods (0, 3, 6, 12, or 24 h). We found that nickel significantly increased the cell viability loss and lactate dehydrogenase (LDH) release in Neuro-2a cells. In addition, nickel exposure significantly elevated reactive oxygen species (ROS) and malondialdehyde (MDA) levels, disrupted the mitochondrial membrane potential (ΔΨ(m)), reduced adenosine-5'-triphosphate (ATP) concentrations and decreased mitochondrial DNA (mtDNA) copy numbers and mtRNA transcript levels. However, all of the cytotoxicities and mitochondrial dysfunctions that were triggered by nickel were efficiently attenuated by pretreatment with LC. These protective effects of LC may be attributable to its role in maintaining mitochondrial function in nickel-treated cells. Our results suggest that LC may have great pharmacological potential in protecting against the adverse effects of nickel in the nervous system.

Signal transducer and activator of transcription 1 (STAT1) is essential for chromium silencing of gene induction in human airway epithelial cells.

Hexavalent chromium (Cr(VI)) promotes lung injury and pulmonary diseases through poorly defined mechanisms that may involve the silencing of inducible protective genes. The current study investigated the hypothesis that Cr(VI) actively signals through a signal transducer and activator of transcription 1 (STAT1)-dependent pathway to silence nickel (Ni)-induced expression of vascular endothelial cell growth factor A (VEGFA), an important mediator of lung injury and repair. In human bronchial airway epithelial (BEAS-2B) cells, Ni-induced VEGFA transcription by stimulating an extracellular regulated kinase (ERK) signaling cascade that involved Src kinase-activated Sp1 transactivation, as well as increased hypoxia-inducible factor-1 alpha (HIF-1 alpha) stabilization and DNA binding. Ni-stimulated ERK, Src, and HIF-1 alpha activities, as well as Ni-induced VEGFA transcript levels were inhibited in Cr(VI)-exposed cells. We previously demonstrated that Cr(VI) stimulates STAT1 to suppress VEGFA expression. In BEAS-2B cells stably expressing STAT1 short hairpin RNA, Cr(VI) increased VEGFA transcript levels and Sp1 transactivation. Moreover, in the absence of STAT1, Cr(VI), and Ni coexposures positively interacted to further increase VEGFA transcripts. This study demonstrates that metal-stimulated signaling cascades interact to regulate transcription and induction of adaptive or repair responses in airway cells. In addition, the data implicate STAT1 as a rate limiting mediator of Cr(VI)-stimulated gene regulation and suggest that cells lacking STAT1, such as many tumor cell lines, have opposite responses to Cr(VI) relative to normal cells.

Protective role of vitamin E on nickel and/or chromium induced oxidative stress in the mouse ovary.

In the present study, we report the invivo effects of nickel chloride (NiCl2; 8 and 16 mg/kg body weight) and/or potassium dichromate (K2Cr2O7; 5 and 10mg/kg body weight) in the ovary of adult mice. The protective role of vitamin E (2mg/kg body weight) along with their combination was also studied. Nickel and/or chromium to mice enhanced the levels of lipid peroxides in the ovary, which was accompanied by a significant decline in the levels of protein, glutathione, total ascorbic acid and activities of superoxide dismutase and catalase. Supplementation of vitamin E along with NiCl2 + K2Cr2O7 significantly lowered the levels of lipid peroxidation and enhanced the antioxidant status. Findings of the present study suggest that vitamin E exerts its protective effect against nickel and/or chromium induced toxicity by preventing lipid peroxidation and protecting antioxidant system in the mouse ovary.

Protective effect of cactus (Opuntia ficus indica) cladode extract upon nickel-induced toxicity in rats.

The purpose of this study carried out on male Wistar rats, was to evaluate the protective effects of regular ingestion of juice from the prickly pear cactus (Opuntia ficus indica) cladodes against nickel chloride toxicity. Rats were given either normal tap water or water containing 25% of cactus juice for one month. Then, rats of each group were injected daily, for 10 days, with either NiCl(2) solution (4mg (30micromol)/kg body weight) or with the same volume of saline solution (300mM NaCl). Significant increases of lactate dehydrogenase, aspartate aminotransferase, alanine aminotransferase activities and of cholesterol, triglycerides and glucose levels were observed in blood of nickel-treated rats. In the liver, nickel chloride was found to induce an oxidative stress evidenced by an increase in lipid peroxidation and changes in antioxidant enzymes activities. Superoxide-dismutase (SOD) activity was found to be increased whereas glutathione peroxidase and catalase activities were decreased. These changes did not occur in animals previously given cactus juice, demonstrating a protective effect of this vegetal extract.

Critical appraisal of respirometric methods for metal inhibition on activated sludge.

This paper evaluates the merit of oxygen uptake rate measurements for the assessment of metal inhibition on activated sludge. For this purpose, experiments are conducted to calculate EC50 levels of nickel and hexavalent chromium using the ISO 8192 procedure, yielding results that are highly variable and difficult to correlate, depending on the type of substrate and the initial food to microorganism ratio. Similar experiments based on continuous respirometric measurements to give the entire oxygen uptake rate profile provide a much better insight on the impact of inhibition on different biochemical processes taking place in the reactor. The results indicate that percent reduction of the amount of dissolved oxygen utilized after an appropriate reaction time is a much better index for the assessment of the inhibitory effects.

Acorus calamus extracts and nickel chloride: prevention of oxidative damage and hyperproliferation response in rat kidney.

Nickel, a major environmental pollutant, is known for its clastogenic, toxic, and carcinogenic potential. In this article, we report the effect of Acorus calamus on nickel chloride (NiCl2)-induced renal oxidative stress, toxicity, and cell proliferation response in male Wistar rats. NiCl2 (250 micromol/kg body weight/mL) enhanced reduced renal glutathione content (GSH), glutathione- S-transferase (GST), glutathione reductase (GR), lipid peroxidation (LPO), H2O2 generation, blood urea nitrogen (BUN), and serum creatinine with a concomitant decrease in the activity of glutathione peroxidase (GPx) (p < 0.001). NiCl2 administration also dose-dependently induced the renal ornithine decarboxylase (ODC) activity several-fold as compared to salinetreated control rats. Similarly, renal DNA synthesis, which is measured in terms of [3H] thymidine incorporation in DNA, was elevated following NiCl2 treatment. Prophylactic treatment of rats with A. calamus (100 and 200 mg/kg body weight po) daily for 1 wk resulted in the diminution of NiCl2- mediated damage, as evident from the downregulation of glutathione content, GST, GR, LPO, H2O2 generation, BUN, serum creatinine, DNA synthesis (p < 0.001), and ODC activity (p < 0.01) with concomitant restoration of GPx activity. These results clearly demonstrate the role of oxidative stress and its relation to renal disfunctioning and suggest a protective effect of A. calamus on NiCl2-induced nephrotoxicity in a rat experimental model.

Evaluation of PDTC for removing Ni(II) from human serum transferrin in vitro.

Pyrrolidine dithiocarbamate (PDTC)can be an oral chelator with pKa=3.300+/-0.002. It behaves as a bidentate ligand at serum pH. The effect of pH on Ni2+-Tf indicated that the maximum adsorption was at pH=7.4. The effective Ni-PDTC binding constant was determined (logk=11.1+/-0.1) for the 1:2 Ni(PDTC)2 complex using UV-vis spectra. The isosbestic point at 298 indicated that the complexation reaction was done directly (without side reaction). Removal of Ni from transferrin (Tf) was investigated by reverse titration of PDTC at 25 masculineC and pH=7.4 using UV-vis spectra. PDTC is able to remove 25% of Ni from human serum transferrin.

Zinc protects rat liver histo-architecture from detrimental effects of nickel.

This study was designed to examine the protective potential of zinc on the histoarchitecture distortion induced by nickel in rats. Male Sprauge Dawley (S.D) rats received either nickel alone in the form NiSO(4) x 6H(2)O at a dose of 800 mg/l in drinking water, zinc alone in the form of ZnSO(4) x 7H(2)O at a dose of 227 mg/l in drinking water, or nickel plus zinc or drinking water alone for a total duration of eight weeks. The effects of different treatments were studied on rat liver histoarchitecture by using both light and transmission electron microscopes. Normal control and zinc treated animals revealed normal histology of liver, however, nickel treated animals resulted in drastic alterations of normal hepatic histoarchitecture, after 8 weeks of treatment. Administration of zinc to nickel treated rats resulted in marked improvement in the structure of hepatocytes, thus emphasizing the protective potential of zinc in restoring the altered hepatic histoarchitecture close to the histoarchitecture of normal animals.

Reduction of nickel-induced contact hypersensitivity reactions by topical tea tree oil in humans.

OBJECTIVE AND DESIGN: Whilst the anti-microbial properties of tea tree oil (TTO) are established, the anti-inflammatory effects of TTO in human skin remain largely anecdotal and require evaluation. This study examined the effect of topically applied TTO on nickel-induced contact hypersensitivity reactions in human dorsal skin. TREATMENT: TTO (100%), a 5% TTO lotion, a placebo lotion (no TTO), or 100% macadamia oil were applied at days 3 and 5 after nickel exposure. METHODS: The flare area and erythema index were measured on days 3, 5 and 7. The regulatory effects of TTO were also investigated on the proliferative response to nickel or polyclonal mitogens by peripheral blood mononuclear cells from nickel-sensitive and control subjects. RESULTS: TTO (100%) significantly reduced the flare area and erythema index when compared to the nickel-only sites. With respect to the erythema index, the anti-inflammatory effects were predominantly, but not exclusively, seen in a subgroup of nickel-sensitive subjects with a prolonged development phase of nickel-induced contact hypersensitivity response. The 5% TTO lotion, the placebo lotion and the 100% macadamia oil were all without significant effect. TTO significantly inhibited proliferation to nickel but not to non-specific polyclonal mitogens by peripheral blood mononuclear cells from nickel-sensitive subjects. CONCLUSIONS: Topical application of 100% TTO may have therapeutic benefit in nickel-induced contact hypersensitivity in human skin. The mode of action of TTO requires further investigation, but may be an effect on the antigen presenting cells or the antigen presenting process in nickel-induced contact hypersensitivity, as well as vascular changes associated with this response.

Responses to nickel in the proteome of the hyperaccumulator plant Alyssum lesbiacum.

A proteomic analysis of the Ni hyperaccumulator plant Alyssum lesbiacum was carried out to identify proteins that may play a role in the exceptional degree of Ni tolerance and accumulation characteristic of this metallophyte. Of the 816 polypeptides detected in root tissue by 2D SDS-PAGE, eleven increased and one decreased in abundance relative to total protein after 6-week-old plants were transferred from a standard nutrient solution containing trace concentrations of Ni to a moderately high Ni treatment (0.3 mM NiSO4) for 48 h. These polypeptides were identified by tandem mass spectrometry and the majority were found to be involved in sulphur metabolism (consistent with a re-allocation of sulphur towards cysteine and glutathione), protection against reactive oxygen species, or heat-shock response. In contrast, very few polypeptides were found to change in abundance in root or shoot tissue after plants were exposed for 28 days to 0.03 mM NiSO4, a concentration representing the optimum for growth of this species but sufficient to lead to hyperaccumulation of Ni in the shoot. Under these conditions, constitutively expressed genes in this highly Ni-tolerant species may be sufficient to allow for effective chelation and sequestration of Ni without the need for additional protein synthesis.

Identification of membrane calcium channels essential for cytoplasmic and nuclear calcium elevations induced by vascular endothelial growth factor in human endothelial cells.

Vascular endothelial growth factor (VEGF) is mitogenic for endothelial cells and has been shown to induce angiogenesis and endothelial cell migration through stimulation of endothelial tyrosine-kinase receptors. Here, using confocal microscopy and the patch-clamp technique on endothelial cells, membrane permeability to calcium as well as cytoplasmic and nuclear free calcium levels have been investigated in the first stages of tyrosine-kinase receptor activation by VEGF. VEGF (0.5nM) as well as inositol trisphosphate (IP3) induced an activation of membrane calcium-permeable channels exhibiting a similar low conductance in the range of 10 pS. The VEGF-triggered activation of these calcium channels, mediated by IP3 and involving the intracellular calcium stores, results in an increase in both cytoplasmic and nuclear calcium levels in endothelial cells, potentially modulating gene expression. Finally, the effect of Ni2+, a calcium channel blocker, on endothelial cell proliferation has been studied. The results show that inhibition of extracellular calcium influx significantly inhibits VEGF-induced cell proliferation. In the process of cell stimulation by VEGF, and possibly by other growth factors, activation of calcium channels could then be a key step in calcium-regulated gene expression and cell activation. These results suggest that the use of calcium channel blockers could be a novel way of prevention or reversion of VEGF-induced tumoral angiogenesis.

AP-1-dependent induction of plasminogen activator inhibitor-1 by nickel does not require reactive oxygen.

Inhalation of nickel dust has been associated with an increased incidence of pulmonary fibrosis. Nickel may promote fibrosis by transcriptionally activating plasminogen activator inhibitor (PAI)-1 and inhibiting fibrinolysis. The current studies examined whether nickel stimulated the PAI-1 promoter though an oxidant-sensitive activator protein (AP)-1 signaling pathway. Addition of nickel to BEAS-2B human airway epithelial cells stimulated intracellular oxidation, induced c-Jun and c-Fos mRNA levels, increased phospho- and total c-Jun protein levels, and elevated PAI-1 mRNA levels over a 24-h time course. Pretreatment of the cells with antioxidants did not affect increased c-Jun protein or PAI-1 mRNA levels. Expression of the dominant negative inhibitor of AP-1, TAM67, prevented nickel-stimulated AP-1 DNA binding, AP-1-luciferase reporter construct activity, and PAI-1 mRNA levels. Overexpression of c-Jun, however, failed to induce the AP-1 luciferase reporter construct or PAI-1 mRNA levels. These data indicated that nickel activated AP-1 through an oxidant-independent pathway and that basal AP-1 is necessary for nickel-induced expression of PAI-1.

Two-dimensional crystallization of a membrane protein on a detergent-resistant lipid monolayer.

Two-dimensional crystals of a membrane protein, the proton ATPase from plant plasma membranes, have been obtained by a new strategy based on the use of functionalized, fluorinated lipids spread at the air-water interface. Monolayers of the fluorinated lipids are stable even in the presence of high concentrations of various detergents as was established by ellipsometry measurements. A nickel functionalized fluorinated lipid was spread into a monolayer at the air-water interface. The overexpressed His-tagged ATPase solubilized by detergents was added to the subphase. 2D crystals of the membrane protein, embedded in a lipid bilayer, formed as the detergent was removed by adsorption. Electron microscopy indicated that the 2D crystals were single layers with dimensions of 10 microm or more. Image processing yielded a projection map at 9 A resolution, showing three well-separated domains of the membrane-embedded proton ATPase.

Specific amino acids moderate the effects on Ni2+ on the testosterone production of mouse leydig cells in vitro.

The purpose of this investigation was to study the effectiveness of two nickel-binding amino acids, histidine (His) and cysteine (Cys), to prevent the inhibitory action of Ni2+ on testosterone (T) production by mouse primary Leydig cell culture. The maximal human chorionic gonadotropin (hCG)-stimulated T response was measured by radioimmunoassay (RIA) in the culture media. Three types of experiments were performed. In a concentration-response study, Ni2+ (62.5 to 1,000 microM) was added to the cells simultaneously with equimolar or twice the equimolar concentrations of His or Cys and the cultures were maintained for 48 h. Nickel-induced reduction in T production was completely prevented by equimolar concentrations of His at Ni2+ concentrations of 125, 250, and 500 microM; equimolar or twice the equimolar concentrations of His were only partially effective at 1,000 microM Ni2+. Protective action of Cys was complete only at the lowest concentration of Ni2+ (125 microM). In a second series, the cells were incubated for various times (0.5 to 48 h) with 1,000 microM Ni2+ in the presence of 2,000 microM His or Cys. Increasing the time of incubation, the protective effect of both amino acids against Ni2+ was reduced. In a third series, attempts were made to reverse the action of 1,000 microM Ni2+ after incubation with cells for various times (0.5 to 24 h), followed by exposure to 2,000 microM His or Cys. Cell cultures were maintained for 48 h. A partial recovery of hCG-stimulated T production could be observed only if the amino acid was added not later than 4 h after the metal. This time was also required to elicit the T depression produced by Ni2+. Administration of either His or Cys at later times had no effect. Our results show that both His and Cys are able to moderate the effects of Ni2+ on Leydig cell T production, depending on the concentration of this metal ion, as well as on amino acid. However, at higher Ni2+ concentrations the complete protection by His or Cys is only temporary. Administration of these amino acids after the Ni2+-produced decrease in T production was not able to reverse the process.

Perfil de risco entre os pacientes com história de atopia quanto às dermatites alérgicas de contato / Risk profile among patients with a history of atopy for allergic contact dermatitis

Verificar a freqüência da associaçäo entre a presença de histórico pessoal e/ou familiar de atopia e a positividade dos testes epicutâneos para avaliaçäo de dermatite alérgica de contato; Verificar as especifidades quanto aos tipos de substâncias implicadas, à faixa etária e ao grau de intensidade das reaçöes aos testes. Foram avaliados 136 pacientes que consultaram no Setor de Alergia Cutânea do Serviço e realizaram os testes epicutâneos entre 1998 e 2000. Utilizou-se uma bateria padräo com 30 substâncias, com retirada dos testes em 48 horas e leitura final em 96 horas. Realizou-se aferiçäo dos resultados de acordo com as normas internacionais.Näo houve diferença estatisticamente significativa entre os grupos com e sem história de atopia quanto à freqüência de positividade aos testes epicutâneos. O grau de intensidade das reaçöes foi maior no grupo com história de atopia (OR: 1,36). O grupo com história de atopia teve uma média de idade de 39,8 anos e o grupo sem a história uma média de 45,9 anos. Entre os atópicos, verificou-se o sulfato de níquel com significância estatística como a principal substância (p<0,05). Näo há diferenças da freqüência em geral quanto à reatividade aos testes epicutâneos. Os pacientes com história de atopia pessoal e/ou familiar provavelmente constituem um grupo de risco para o desenvolvimento de dermatites alérgicas de contato mais intensas e säo, particularmente, reativos ao sulfato de níquel.

Ellagic acid ameliorates nickel induced biochemical alterations: diminution of oxidative stress.

Nickel, a major environmental pollutant is known for its clastogenic, toxic and carcinogenic potentials. The present investigation shows that ellagic acid proves to be exceptional in the amelioration of the nickel-induced biochemical alterations in serum, liver and kidney. Administration of nickel (250 micromol Ni/kg body wt) to female Wistar rats, resulted in increase in the reduced glutathione (GSH) content [kidney (*P<0.05) and liver (**P<0.001)] and Glutathione-S-transferase (GST) and glutathione reductase (GR) activities [kidney and liver, (**P<0.001)]. Ellagic acid treatment to the intoxicated rats leads to the formation of soluble ellagic acid-metal complex which facilitates excretion of nickel from the cell or tissue, thus ameliorating nickel-induced toxicity, as evident from the down regulation of GSH content, GST and GR activities with concomitant restoration of glutathione peroxidase (GPx) activity in liver and kidney. Our data shows that ellagic acid maintains cell membrane integrity through sequestration of metal ions from the extracellular fluid, as evident from the alleviated levels of serum glutamate oxaloacetate transaminase, (SGOT), serum glutamate pyruvate transaminase (SGPT) and lactate dehydrogenase (LDH) when compared to nickel treated group. Similarly, the enhanced blood urea nitrogen (BUN) and serum creatinine levels that are indicative of renal injury showed a reduction of about 45 and 40%, respectively. The data also show that treatment of ellagic acid after 30 min of nickel administration exhibits maximum inhibition in a dose-dependent manner. In summary, our data suggests that ellagic acid act as an effective chelating agent in suppressing nickel-induced renal and hepatic biochemical alterations.

Vicinal-thiol-containing molecules enhance but mono-thiol-containing molecules reduce nickel-induced DNA strand breaks.

Several thiol-containing molecules (TCM) are currently used as antidotes for nickel, and vicinal TCM seem to be more effective in mobilizing tissue nickel than are mono TCM. Using single cell alkaline electrophoresis, we have shown that the vicinal TCM, meso-2, 3-dimercaptosuccinic acid (DMSA), 2,3-dimercaptopropane-1-sulfonate, and 2,3-dimercaptopropanol markedly enhanced, whereas the mono TCM, D-penicillamide, glutathione, beta-mercaptoethanol, and diethyl dithiocarbomate, reduced nickel chloride (Ni)-induced DNA breaks in a human leukemia cell line, NB4 cells. Ni or TCM alone did not induce plasmid DNA breaks in test tubes and neither did Ni plus mono TCM; however, Ni plus vicinal TCM did. Vicinal TCM did, but mono TCM did not generate H(2)O(2) in solution. H(2)O(2) alone did not, but H(2)O(2) plus Ni induced plasmid DNA breaks. Although Ni plus glutathione did not break DNA, Ni plus glutathione plus H(2)O(2) did. The Ni-DMSA-induced DNA breaks in NB4 cells, as well as in plasmids, were completely prevented by d-mannitol or partially prevented by several antioxidants. Therefore, the DNA breaks induced by Ni plus vicinal TCM seem to be due to the complex of Ni with TCM in concert with the H(2)O(2) produced by the vicinal TCM. The results that DMSA at a concentration as low as 5 microM enhanced the Ni-induced DNA breaks suggest a further evaluation of the TCM as nickel chelators is needed.

Effects of nickel chloride on reproduction of the rat and possible antagonistic role of selenium.

Nickel (10-100 ppm added as NiCl2) was studied to determine its effects on reproduction of Wistar rats. In nine experimental groups, females, males or both were exposed to nickel in drinking water. In one female group and one male group, the drinking water was also supplemented with 0.3 ppm selenium (added as Na2SeO3). Breeding success and the growth and viability of pups were recorded. Nickel, copper and zinc concentrations in kidneys, liver and skin (with fur) of the females, males and pups were determined with an atomic absorption spectrophotometer. In addition, histology of the male testes (from control and nickel-exposed groups) was studied. The female exposures started 14, 28 or 100 days before copulation and continued during pregnancy and lactation. When the males were exposed (for 28 or 42 days before copulation), NiCl2 reduced both the number of pregnancies and the number of pups born. In the testes, NiCl2 induced shrinkage of the seminiferous tubules, which seemed to close some of the tubules. In the tubules, NiCl2 decreased the number of basal spermatogonia. When the females or both parents were exposed to NiCl2, pup mortality during lactation was high. However, when the females were drinking NiCl2 supplemented with selenium, all the pups survived and development of the total mass of the litters was even better than in the control group. In the same way, in males, selenium supplementation of the drinking water protected those pups that were born; but fertility was lower than with the control treatment. In the tissues studied, nickel accumulated most in the kidneys and then in the liver and skin. In each type of organ, there was a clear dose response relationship. In the pups, in particular, selenium (given to the females) increased the amount of nickel in tissues compared with corresponding administration of nickel without selenium. In summary, selenium seemed to counteract the deleterious effects of NiCl2 on the reproduction of rats.

Nickel subsulfide is similar to potassium dichromate in protecting normal human fibroblasts from the mutagenic effects of benzo[a]pyrene diolepoxide.

The cellular response to multiple carcinogen treatment has not been extensively studied, even though the effect of individual carcinogens is, in many cases, well known. We have previously shown that potassium dichromate can protect normal human fibroblasts from the mutagenic effects of benzo[a]pyrene diolepoxide (BPDE), and that this effect may be via an oxidative stress mechanism [Tesfai et al. (1998) Mutat Res 416:159-168]. Here, we extend our previous work by showing that nickel subsulfide can produce the some effect. Normal human fibroblasts, preincubated with nickel subsulfide for 46 hr followed by a coincubation of nickel subsulfide and BPDE for 2 hr, showed a dramatic reduction in the mutant frequency of the hypoxanthine (guanine)phosphoribosyl-transferase (HPRT) gene when compared to cells treated only with BPDE. The preincubation period with nickel subsulfide was necessary to see the antagonistic effect, since it was not observed if the cells were simply incubated with both carcinogens for 2 hr. The extent of the antagonistic effect was nickel subsulfide dose-dependent and also appeared to be species-specific, since the effect was not observed when Chinese hamster fibroblasts were tested. Finally, the antagonistic effect of the nickel subsulfide was eliminated by vitamin E, suggesting that production of reactive oxygen species by the nickel may be required. This data, along with our previous work, suggest that the antagonistic effect we observe is not chromium-specific, and that it could be species-specific.