Mitigative role of garlic and vitamin E against cytotoxic, genotoxic, and apoptotic effects of lead acetate and mercury chloride on WI-38 cells.

BACKGROUND: Lead acetate (Led) and mercury chloride (Mer) represent important ecological and public health concerns due to their hazardous toxicities. Naturally found products play a vital role in chemopreventive agent innovation. The current study aimed to assess the modifying effect of garlic (Gar) and/or vitamin E (Vit E) against the half-maximal inhibitory concentration (IC50) Led and/or Mer-induced cytotoxic, genotoxic and apoptotic effects. METHODS: Human lung cells (WI-38) were pretreated with Gar and/or Vit E for 24h and then treated with Led and/or Mer either alone or with their combination for 24h. Cytotoxicity of Led and Mer and the viability of Gar and Vit E were assessed using MTT assay. The alkaline comet assay was used to assess DNA damage, whereas QRT-PCR was performed to evaluate p53, Bax, and Bcl2 mRNA-expression. RESULTS: The results of this study showed that IC50 of Led was (732.72µg/mL) and for Mer was (885.83µg/mL), while cell viability effective dose for Gar was (300µg/mL) and for Vit E was (26,800µg/mL). Treating cells with the IC50-concentration of Led or Mer or their combination using half IC50 of both of them induced severe DNA-damage. Bax-expression was increased, while p53 and Bcl2-expressions were decreased. Pretreatment of cells with Gar and/or Vit E ameliorated the previous alternations. CONCLUSIONS: Led and Mer can induce oxidative stress and change the expressions of apoptosis-related proteins in WI-38 cells. Gar and Vit E may be promising protective candidate agent against the toxic effect of heavy metals.

Arginine decarboxylase: A novel biological target of mercury compounds identified in PC12 cells.

Mercury compounds are well-known toxic environmental pollutants and potently induce severe neurotoxicological effects in human and experimental animals. Previous studies showed that one of the mechanisms of mercury compounds neurotoxicity arose from the over-activation of the N-methyl d-aspartate (NMDA)-type glutamate receptor induced by increased glutamate release. In this work, we aimed to investigate the molecular mechanisms of Hg compounds neurotoxicities by identifying their biological targets in cells. Firstly, the inhibitory effects of four Hg compounds, including three organic (methyl-, ethyl- and phenyl-mercury) and one inorganic (Hg2+) Hg compounds, on the activity of arginine decarboxylase (ADC), a key enzyme in the central agmatinergic system, were evaluated. They were found to inhibit the ADC activity significantly with methylmercury (MeHg) being the strongest (IC50=7.96nM). Furthermore, they showed remarkable inhibitory effects on ADC activity in PC12 cells (MeHg>EtHg>PhHg>HgCl2), and led to a marked loss in the level of agmatine, an endogenous neuromodulatory and neuroprotective agent that selectively blocks the activation of NMDA receptors. MeHg was detected in the immunoprecipitated ADC from the cells, providing unequivocal evidence for the direct binding of MeHg with ADC in the cell. Molecular dynamics simulation revealed that Hg compounds could form the coordination bond not only with cofactor PLP of ADC, but also with substrate arginine. Our finding indicated that MeHg could attenuate the neuroprotective effects of agmatine by the inhibition of ADC, a new cellular target of MeHg, which might be implicated in molecular mechanism of MeHg neurotoxicity.

Thymol, a monoterpene phenolic derivative of cymene, abrogates mercury-induced oxidative stress resultant cytotoxicity and genotoxicity in hepatocarcinoma cells.

Thymol (TOH) was investigated for its ability to protect against mercuric chloride (HgCl2 )-induced cytotoxicity and genotoxicity using human hepatocarcinoma (HepG2) cell line. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay confirmed the efficacy of TOH pretreatment in attenuating HgCl2 -induced cytotoxicity. Pretreatment with TOH inhibited HgCl2 -induced genotoxicity, depolarization of mitochondrial membrane, oxidative stress, and mitochondrial superoxide levels. Interestingly, TOH (100 µM) alone elevated the intracellular basal glutathione S-transferase (GST) levels and TOH pretreatment abrogated the decrease in glutathione, GST, superoxide dismutase, and catalase levels even after HgCl2 intoxication. Furthermore, TOH was also capable of inhibiting HgCl2 -induced apoptotic as well as necrotic cell death analyzed by flowcytometric analysis of cells dual stained with Annexin-FITC/propidium iodide. The present findings clearly indicate the cytoprotective potential of TOH against HgCl2 -induced toxicity, which may be attributed to its free radical scavenging ability which facilitated in reducing oxidative stress and mitochondrial damage thereby inhibiting cell death.

Full recovery from a potentially lethal dose of mercuric chloride.

INTRODUCTION: Mercuric chloride poisoning is rare yet potentially life-threatening. We report a case of poisoning with a potentially significant amount of mercuric chloride which responded to aggressive management. CASE REPORT: A 19-year-old female presented to the Emergency Department with nausea, abdominal discomfort, vomiting of blood-stained fluid, and diarrhea following suicidal ingestion of 2-4 g of mercuric chloride powder. An abdominal radiograph showed radio-opaque material within the gastric antrum and the patient's initial blood mercury concentration was 17.9 µmol/L (or 3.58 mg/L) at 3 h post-ingestion. Given the potential toxicity of inorganic mercury, the patient was admitted to the intensive care unit and chelation with dimercaprol was undertaken. Further clinical effects included mild hemodynamic instability, acidosis, hypokalemia, leukocytosis, and fever. The patient's symptoms began to improve 48 h after admission and resolved fully within a week. DISCUSSION: Mercuric chloride has an estimated human fatal dose of between 1 and 4 g. Despite a reported ingestion of a potentially lethal dose and a high blood concentration, this patient experienced mild to moderate poisoning only and she responded to early and appropriate intervention. Mercuric chloride can produce a range of toxic effects including corrosive injury, severe gastrointestinal disturbances, acute renal failure, circulatory collapse, and eventual death. Treatment includes close observation and aggressive supportive care along with chelation, preferably with 2,3-dimercapto-1-propane sulfonate or 2,3-meso-dimercaptosuccinic acid.

Effects of selenite and chelating agents on mammalian thioredoxin reductase inhibited by mercury: implications for treatment of mercury poisoning.

Mercury toxicity is a highly interesting topic in biomedicine due to the severe endpoints and treatment limitations. Selenite serves as an antagonist of mercury toxicity, but the molecular mechanism of detoxification is not clear. Inhibition of the selenoenzyme thioredoxin reductase (TrxR) is a suggested mechanism of toxicity. Here, we demonstrated enhanced inhibition of activity by inorganic and organic mercury compounds in NADPH-reduced TrxR, consistent with binding of mercury also to the active site selenolthiol. On treatment with 5 µM selenite and NADPH, TrxR inactivated by HgCl(2) displayed almost full recovery of activity. Structural analysis indicated that mercury was complexed with TrxR, but enzyme-generated selenide removed mercury as mercury selenide, regenerating the active site selenocysteine and cysteine residues required for activity. The antagonistic effects on TrxR inhibition were extended to endogenous antioxidants, such as GSH, and clinically used exogenous chelating agents BAL, DMPS, DMSA, and α-lipoic acid. Consistent with the in vitro results, recovery of TrxR activity and cell viability by selenite was observed in HgCl(2)-treated HEK 293 cells. These results stress the role of TrxR as a target of mercurials and provide the mechanism of selenite as a detoxification agent for mercury poisoning.

Concomitant administration of sodium 2,3-dimercapto-1-propanesulphonate (DMPS) and diphenyl diselenide reduces effectiveness of DMPS in restoring damage induced by mercuric chloride in mice.

The effect of combined therapy with diphenyl diselenide (PhSe)(2) and sodium 2,3-dimercapto-propane-1-sulphonate (DMPS) against alterations induced by mercury (Hg(2+)) was evaluated. Mice were exposed to mercuric chloride (HgCl(2)) (1mg/kg, subcutaneously) for two weeks. After that, mice received (PhSe)(2) (15.6 mg/kg), or DMPS (12.6 mg/kg), or a combination of both for one week. Thiobarbituric acid-reactive substances (TBARS), ascorbic acid and Hg(2+) levels and glutathione S-transferase (GST) and catalase (CAT) activities were carried out in kidney. Hematological parameters, plasmatic bilirubin, uric acid, urea and creatinine levels as well as lactate dehydrogenase (LDH) activity were determined. (PhSe)(2) or DMPS restored the increase in LDH activity and TBARS, bilirubin, uric acid, urea and creatinine levels caused by HgCl(2). The levels of erythrocytes, hemoglobin and hematocrit reduced by HgCl(2) exposure were restored by (PhSe)(2) or DMPS administration in mice. Leukocyte and platelet counts modified by HgCl(2) exposure were restored by (PhSe)(2) or DMPS therapy. DMPS restored the increase in Hg(2+) levels induced by exposure to HgCl(2). Concomitant administration of (PhSe)(2) and DMPS reduced the effectiveness of DMPS in restoring damage induced by HgCl(2). Combined therapy with (PhSe)(2) and DMPS was less effective than isolated therapies in restoring the damage induced by HgCl(2) in mice.

Effects of chelators on mercury, iron, and lead neurotoxicity in cortical culture.

Chelation therapy for the treatment of acute, high dose exposure to heavy metals is accepted medical practice. However, a much wider use of metal chelators is by alternative health practitioners for so called "chelation therapy". Given this widespread and largely unregulated use of metal chelators it is important to understand the actions of these compounds. We tested the effects of four commonly used metal chelators, calcium disodium ethylenediaminetetraacetate (CaNa2EDTA), D-penicillamine (DPA), 2,3 dimercaptopropane-1-sulfonate (DMPS), and dimercaptosuccinic acid (DMSA) for their effects on heavy metal neurotoxicity in primary cortical cultures. We studied the toxicity of three forms of mercury, inorganic mercury (HgCl2), methyl mercury (MeHg), and ethyl mercury (thimerosal), as well as lead (PbCl2) and iron (Fe-citrate). DPA had the worst profile of effects, providing no protection while potentiating HgCl2, thimerosal, and Fe-citrate toxicity. DMPS and DMSA both attenuated HgCl2 toxicity and potentiated thimerosal and Fe toxicity, while DMPS also potentiated PbCl2 toxicity. CaNa2EDTA attenuated HgCl2 toxicity, but caused a severe potentiation of Fe-citrate toxicity. The ability of these chelators to attenuate the toxicity of various metals is quite restricted, and potentiation of toxicity is a serious concern. Specifically, protection is provided only against inorganic mercury, while it is lacking against the common form of mercury found in food, MeHg, and the form found in vaccines, thimerosal. The potentiation of Fe-citrate toxicity is of concern because of iron's role in oxidative stress in the body. Potentiation of iron toxicity could have serious health consequences when using chelation therapy.

Effect of astaxanthin on kidney function impairment and oxidative stress induced by mercuric chloride in rats.

Reactive oxygen species are implicated as mediators of tissue damage in the acute renal failure induced by inorganic mercury. Astaxanthin (ASX), a carotenoid with potent antioxidant properties, exists naturally in various plants, algae, and seafoods. This paper evaluated the ability of ASX to prevent HgCl(2) nephrotoxicity. Rats were injected with HgCl(2) (0 or 5 mg/kg b.w., sc) 6h after ASX had been administered (0, 10, 25, or 50mg/kg, by gavage) and were killed 12h after HgCl(2) exposure. Although ASX prevented the increase of lipid and protein oxidation and attenuated histopathological changes caused by HgCl(2) in kidney, it did not prevent creatinine increase in plasma and delta-aminolevulinic acid dehydratase inhibition induced by HgCl(2). Glutathione peroxidase and catalase activities were enhanced, while superoxide dismutase activity was depressed in HgCl(2)-treated rats when compared to control and these effects were prevented by ASX. Our results indicate that ASX could have a beneficial role against HgCl(2) toxicity by preventing lipid and protein oxidation, changes in the activity of antioxidant enzymes and histopathological changes.

Effects of BSO, GSH, Vit-C and DMPS on the nephrotoxicity of mercury.

OBJECTIVE: To study the effects of BSO, GSH, Vit-C and DMPS on the nephrotoxicity of mercury. METHODS: The rats in groups 1, 2 and 3 were sc injected with 0.75, 1.5 and 2.5 mg/kg HgCl2, respectively. Fourth group rats were ip injected with 0.5 mmol/kg BSO and 4h later sc administrated with 0.75 mg/kg HgCl2. The rats in groups 5, 6 and 7 were ip injected with 3 mmol/kg GSH, 4 mmol/kg Vit-C, 200 micromol/kg DMPS, respectively, and 2 h later sc administrated with 2.5 mg/kg HgCl2. Eighth group rats were sc injected with saline as a control. Mercury concentrations in the liver, renal cortex and urine, urinary NAG, ALP, LDH activities, protein and BUN contents were determined. RESULTS: Urinary NAG, ALP activities, protein and BUN contents in the rats of BSO pretreatment group were significantly higher than that of 0.75 mg/kg HgCl2 alone group and control group. As compared with 2.5 mg/kg HgCl2 alone group, urinary NAG, ALP, LDH activities, urinary protein and BUN contents decreased significantly. CONCLUSION: BSO pretreatment could enhance the renal toxicity of mercury and GSH, Vit-C and DMPS pretreatment had antagonistic effects on nephrotoxicity of mercury.

[Experimental study on the effects of BSO, GSH, vitamin C and DMPS on the nephrotoxicity induced by mercury].

OBJECTIVE: To study the renal toxicity caused by mercury administrated once and to observe the effects of buthionine sulfoximine (BSO), gluthionein (GSH), vitamin C (VC), and sodium 2,3-dimercato-1-propanesulfonate (DMPS) pretreatment on the nephrotoxicity of mercury. METHODS: Sixty-four Wistar rats were divided randomly into eight groups, i. e., control group, low, middle and high dose mercury groups and BSO, GSH, VC, DMPS pretreatment groups. The low, middle, and high dose mercury group rats were subcutaneously (sc) injected with 0.75, 1.5, and 2.5 mg/kg HgCl2, respectively. The BSO pretreatment group rats were intraperitoneally (ip) injected with 0.5 mmol/kg BSO and four hours later sc administrated with 0.75mg/kg HgCl2. The GSH, VC and DMPS pretreatment group rats were ip injected with 3 mmol/kg GSH, 4mmol/kg VC, 200 micromol/kg DMPS, respectively, and two hours later sc administrated with 2.5 mg/kg HgCl2. The control group rats were sc injected with saline at corresponding time. The volume of injection was 5 ml/kg body weight. The 12 h urine samples were collected after 12 hours. After 48 hours, the blood samples were collected and then centrifuged to get the serum. The liver and renal cortex were also removed. Mercury contents in the liver, renal cortex, and urine samples were measured. Urinary NAG, ALP, LDH activities, urinary protein and BUN contents were also determined. RESULTS: Mercury concentrations in the liver, renal cortex, and urine samples increased with mercury dose increasing. Mercury contents in the renal cortex presented evident dose-effect relationship. Mercury concentrations in the liver of high-dose mercury group were higher significantly than that of low, middle-dose mercury group, and control group. The concentrations of urinary mercury in the middle and high dose mercury groups were higher significantly than that of control group. Compared with 0.75mg/kg HgCl2 alone group, BSO pretreatment increased mercury concentrations in the liver, but decreased the concentrations in the renal cortex and urine. Mercury concentrations in the liver of GSH, VC and DMPS pretreatment groups were lower than that of 2.5 mg/kg HgCl2 alone group. Urinary NAG, ALP, LDH activities, urinary protein and BUN contents increased with mercury dose increasing, and the values in the animals of 2.5 mg/kg HgCl2 mercury group were higher significantly than that of control, 0.75 and 1.5 mg/kg HgCl2 groups. Urinary NAG, ALP activities, urinary protein and BUN contents in the rats of BSO pretreatment were higher than that of 0.75 mg/kg HgCl2 alone group and control group. Compared with 2.5 mg/kg HgCl2 alone group, urinary NAG, ALP, LDH activities, urinary protein and BUN contents decreased significantly. CONCLUSION: Mercury concentrations in the liver, renal cortex, and urine of the rats increased with mercury dose increasing. BSO pretreatment could enhance the renal toxicity induced by mercury, however, GSH, VC, and DMPS pretreatment had antagonistic effects on nephrotoxicity of the mercury.

2,3-Dimercapto-1-propanol does not alter the porphobilinogen synthase inhibition but decreases the mercury content in liver and kidney of suckling rats exposed to HgCl2.

Heavy metals have received great attention as environmental pollutants mainly because once introduced in the biological cycle they are incorporated in the food chain. Especially the mercury toxicity due to a diversity of effects caused by different chemical species should be emphasized. Heavy metal intoxication has been treated with chelating agents such as 2,3-dimercapto-1-propanol (BAL). However, the efficacy of this treatment is questionable due to the lack of specific effect on the toxic metal. The present study examined the effects of HgCl2 exposure (five doses of 5.0 mg/kg between ages 8 to 12 days) on physiological parameters, on porphobilinogen synthase activity, and on mercury content in liver, kidneys and brain from suckling rats. The effect of BAL (one dose of 12.5-75 mg/kg) applied 24 hr after mercury intoxication on these parameters was also investigated. The results demonstrate that HgCl2 intoxication induced a decrease of corporal weight gain as well as brain weight and an increase in renal weight. The inhibition of porphobilinogen synthase from liver and kidney, is still significant and was not modified by subsequent BAL treatment. However, BAL altered two effects induced by mercury: increase in death percentage and decrease in mercury contents in liver and kidney. The increase of mortality induced by mercury was not promoted by metal redistribution to brain nor by the increase of porphobilinogen synthase inhibition induced by metal. More investigations are necessary to determine if the different effects of BAL on intoxication by metals are possibly related to other tissues and/or if the probable metal-chelating complex formed is more toxic than the metal itself.

Melatonin protects against mercury(II)-induced oxidative tissue damage in rats.

Mercury exerts a variety of toxic effects in the body. Lipid peroxidation, DNA damage and depletion of reduced glutathione by Hg(II) suggest an oxidative stress-like mechanism for Hg(II) toxicity. Melatonin, the main secretory product of the pineal gland, was recently found to be a potent free radical scavenger and antioxidant. N-Acetylcysteine, a precursor of reduced glutathione and an antioxidant, is used in the therapy of acute heavy metal poisoning. In this study the protective effects of melatonin in comparison to that of N-acetylcysteine against Hg-induced oxidative damage in the kidney, liver, lung and brain tissues were investigated. Wistar albino rats of either sex (200-250 g) were divided into six groups, each consisting of 8 animals. Rats were intraperitoneally injected with 1) 0.9% NaCl, control (C) group; 2) a single dose of 5 mg/kg mercuric chloride (HgCl2), Hg group; 3) melatonin in a dose of 10 mg/kg, 1 hr after HgCl2 injection, Hg-melatonin group; 4) melatonin in a dose of 10 mg/kg one day before and 1 hr after HgCl2 injection, melatonin-Hg-melatonin group; 5) N-acetylcysteine in a dose of 150 mg/kg, 1 hr after HgCl2 injection, Hg-N-acetylcysteine group, and 6) N-acetylcysteine in a dose of 150 mg/kg one day before and 1 hr after HgCl2 injection, N-acetylcysteine-Hg-N-acetylcysteine group. Animals were killed by decapitation 24 hr after the injection of HgCl2. Tissue samples were taken for determination of malondialdehyde, an end-product of lipid peroxidation; glutathione (GSH), a key antioxidant, and myeloperoxidase activity, an index of neutrophil infiltration. The results revealed that HgCl2 induced oxidative tissue damage, as evidenced by increases in malondialdehyde levels. Myeloperoxidase activity was also increased, and GSH levels were decreased in the liver, kidney and the lungs. All of these effects were reversed by melatonin or N-acetylcysteine treatment. Since melatonin or N-acetylcysteine administration reversed these responses, it seems likely that melatonin or N-acetylcysteine can protect all these tissues against HgCl2-induced oxidative damage.

Selenium protection against mercury-induced apoptosis and growth inhibition in cultured K-562 cells.

Selenium and mercuric chloride (MC) interactions regarding effects on cell growth and cell death have been studied. Human K-562 cells were pretreated or simultaneously treated with either selenite (5 or 50 microM) or selenomethionine (10 or 50 microM) and with MC (35 or 50 microM). The 35-microM MC treatments resulted in a clear inhibition of cell growth with no obvious difference between mercury-treated and mercury-selenium-treated cells. Furthermore, the apoptotic frequency was similar at all observations for all selenium treatments with 35 microM MC. In the simultaneously treated selenite and 50- microM MC combinations, a selenite-dependent protection was shown both by increased cell growth and by lower apoptotic frequency at 48 and 96 h of exposure. Both treatments with selenomethionine showed protection observed as an increased cell growth at 48 and 96 h and as decreased apoptotic frequency at 96 h of exposure.

Profile of nonprotein thiols, lipid peroxidation and delta-aminolevulinate dehydratase activity in mouse kidney and liver in response to acute exposure to mercuric chloride and sodium selenite.

The effects of mercury (Hg(2+)) and selenite (Se(4+)) on delta-aminolevulinic acid dehydratase (delta-ALA-D) activity, 2-thiobarbituric acid reactive substances (TBARS) and nonprotein sulfhydryl content (NPSH) in mouse kidney and liver were investigated. Male mice were given a single i.p. injection of Hg(2+) and/or Se(4+) (25 micromol/kg) and were killed at 6, 12, 24 and 48 h after treatment. Hg(2+) inhibited renal delta-ALA-D at 6 and 12 h after treatment. Se(4+) abolished the inhibitory effect of mercury on renal delta-ALA-D at 12 h after treatment. Renal and hepatic NPSH content decreased after Hg(2+) exposure and selenite inhibited, at least in part, the Hg-induced oxidation of renal and hepatic NPSH. Se(4+) and Hg(2+), when injected alone, did not alter hepatic or renal TBARS levels; however, simultaneous exposure to these compounds increased hepatic and renal TBARS levels at 12 and 48 h after treatment, respectively. Present results suggest that selenium abolishes the interaction of Hg(2+) with sulfhydryl groups of protein and nonprotein sources.

Antioxidants inhibit mercuric chloride-induced early vasculitis.

In the Brown Norway (BN) rat, mercuric chloride (HgCl(2)) induces a T(h)2-dominated autoimmune syndrome which includes an early phase of mast cell-dependent vasculitis. We have shown in vitro that oxidative stress up-regulates IL-4 in mast cells and predisposes to degranulation. The aim of this study was to determine whether administration of antioxidants inhibits HgCl(2)-induced early vasculitis in vivo, and, if so, to examine whether modulation of the oxidative/antioxidative balance influences IgE and IL-4 expression by mast cells in situ. Groups of rats were given HgCl(2) + saline, HgCl(2) + N-acetyl-L-cysteine (NAC), saline + saline or saline + NAC respectively and blood was taken and animals killed 48 h later. NAC significantly reduced both HgCl2-induced early vasculitis and HgCl(2)-enhanced IgE expression on mast cells with a trend to a decrease in HgCl(2)-enhanced IL-4 expression in these cells. In addition, there was an increased rat mast cell protease (RMCP) II concentration in the serum after HgCl(2) injection and the elevated levels of RMCP II stimulated by HgCl(2) were totally abolished by the administration NAC in the HgCl(2) + NAC group. However, there was no significant change in serum total IgE concentrations between the HgCl(2) + saline group and the HgCl(2) + NAC group. The non-sulphydryl-containing antioxidants desferrioxamine and pyruvate demonstrated a similar effect in inhibiting HgCl(2)-induced early vasculitis. Our data show that administration of an antioxidant to BN rats reduces HgCl(2)-induced early vasculitis, suggesting that oxidative stress plays a role in the pathogenesis of HgCl(2)-induced early vasculitis. This finding may have implications for the understanding of the initiation in this experimental model of T(h)2 cell-driven autoimmunity and possibly of analogous human diseases.

Protective effect of vitamin E against mercuric chloride reproductive toxicity in male mice.

Mercury intoxication has been associated with male reproductive toxicity in experimental animals and mercury may have the potential to produce adverse effects on fertility in men. Vitamin E may protect against toxic effects of mercury in the liver and other tissues. To investigate the protective role of vitamin E against mercuric chloride toxicity for the testis, epididymis, and vas deferens of adult male mice, animals were treated with either mercuric chloride 1.25 mg/kg/day, vitamin E 2 mg/kg/kg, or a combination of the two treatments. Control animals were treated with water. Treatments were administered by daily gavage for 45 days. An additional group of animals treated with mercuric chloride were permitted to recover for 45 days after mercuric chloride treatments. Parameters studied included serum testosterone, epididymal sperm count, motility, and morphology, epididymal and vas deferens adenosine triphosphatase (ATPase), phosphorylase, sialic acid, glycogen and protein, testicular succinate dehydrogenase (SDH), phosphatases, cholesterol, ascorbic acid, and glutathione. Fertility was evaluated by sperm positive vaginal smears after overnight cohabitation with a female. Mercuric chloride produced a reduction in epididymal sperm count, sperm motility, and sperm viability, and there were no sperm-positive smears in this group. Biochemical tests from the male reproductive organs were also altered by mercuric chloride treatment. Coadministration of vitamin E with mercuric chloride prevented the changes in sperm and biochemical parameters and was associated with control rates of sperm positive smears after cohabitation. Animals given vitamin E with mercuric chloride also had lower concentrations of mercury in the testis, epididimyis, and vas deferens. Permitting animals to recover for 45 days after mercuric chloride treatment resulted in partial recovery of sperm and biochemical parameters. Vitamin E cotreatment has a protective role against mercury-induced male reproductive toxicity.

Heterotrimeric G-protein and signal transduction in the nematode-trapping fungus Arthrobotrys dactyloides.

The fungus Arthrobotrys dactyloides produces specialized constricting rings to trap and then consume nematodes. The signal transduction pathway involved in the nematode-trapping process was examined. Mastoparan, an activator of G-protein, had a stimulatory effect on the inflation of ring cells, whereas a G-protein inhibitor, pertussis toxin, prevented ring-cell expansion. The 40-kDa G alpha of heterotrimeric G-proteins was specifically ADP-ribosylated by pertussis toxin. Using an antibody specific to the 35-kDa subunit G beta, we showed that immunogold-labeled G beta was more concentrated in ring cells than in the hyphae. In the absence of nematodes, the rings could be inflated by either pressurizing the culture in a syringe, raising intracellular Ca2+ concentrations, or adding warm water. We used these methods to reveal differences in responses to antagonists. The results support a model in which the pressure exerted by a nematode on the ring activates G-proteins in the ring cells. The activation leads to an increase in cytoplasmic Ca2+, activation of calmodulin, and finally the opening of water channels. The ring cells expand to constrict the ring and thus immobilize the nematode.

Lead reduces the nephrotoxicity of mercuric chloride.

The nephrotoxic effect of a single intraperitoneal dose of mercuric chloride (HgCl(2); 6 mg/kg) on adult CD-1 female mice was reduced at 24 and 48 h after injection, by a 48-h pretreatment nontoxic dose of lead acetate (Pb; 5 mg/kg) delivered by intravenous tail-vein injection (intravenous). While protection is temporally associated with lead-induced mitosis, occurring about 39 h after intracardiac lead injection (D. D Choie and G. W. Richter, 1974, Lab. Invest. 30, 447-451), the mechanism of the observed protection remains to be established.

Structural basis of the antagonism between inorganic mercury and selenium in mammals.

Mercuric chloride toxicity in mammals can be overcome by co-administration of sodium selenite. We report a study of the mutual detoxification product in rabbit plasma, and of a Hg-Se-S-containing species synthesized by addition of equimolar mercuric chloride and sodium selenite to aqueous, buffered glutathione. Chromatographic purification of this Hg-Se-S species and subsequent structural analysis by Se and Hg extended X-ray absorption fine structure (EXAFS) spectroscopy revealed the presence of four-coordinate Se and Hg entities separated by 2.61 A. Hg and Se near-edge X-ray absorption spectroscopy of erythrocytes, plasma, and bile of rabbits that had been injected with solutions of sodium selenite and mercuric chloride showed that Hg and Se in plasma samples exhibited X-ray absorption spectra that were essentially identical to those of the synthetic Hg-Se-S species. Thus, the molecular detoxification product of sodium selenite and mercuric chloride in rabbits exhibits similarities to the synthetic Hg-Se-S species. The underlying molecular mechanism for the formation of the Hg-Se-S species is discussed.

Levamisole induces interleukin-18 and shifts type 1/type 2 cytokine balance.

Immune responses can be classified, according to the predominant cytokines involved, into type 1 (featuring interferon-gamma, IFN-gamma) and type 2 (featuring interleukin-4, IL-4); imbalance between type 1 and type 2 cytokine compartments has been implicated in many human diseases. Levamisole is a drug with an unknown mode of action that has been used to boost immunity in infectious diseases including leprosy, and in some cancers. To test the hypothesis that levamisole acts by inducing a shift to a type 1 immune response, we used Brown Norway (BN) rats, which are markedly biased to type 2 responses. BN rats treated with levamisole showed a dose-dependent rise in serum IFN-gamma and fall in serum immunoglobulin E (IgE) level. Detailed analysis of cytokine gene expression showed upregulation of IFN-gamma and downregulation of IL-4 messenger RNA. This coincided with marked upregulation of IL-18, a recently characterized cytokine with potent activity in stimulating IFN-gamma production. IL-12 was not induced. Further, the type 2 response induced in BN rats by mercuric chloride was markedly attenuated when rats were pretreated with levamisole: there was a 2-log reduction in maximum serum IgE level and marked attenuation of IL-4 gene upregulation. These data indicate that levamisole acts by resetting the immune balance towards a type 1 response via induction of IL-18. Our findings provide a direction for development of more specific immunomodulating therapy.