The antioxidant defence mechanisms of parasite and host after chronic Hymenolepis diminuta infestation of the rat.

The aim of the present study was to determine antioxidant defence mechanisms in the rat and Hymenolepis diminuta after long-term infestation. We determined levels of oxidative stress markers, and activity of antioxidant enzymes in the rat small intestine and in particular parts of H. diminuta. Observed changes in antioxidant enzymes activity in H. diminuta and the rat intestine indicate the defence against parasitic infestation and probably allowed parasite to adapt and live in oxidative stress.

The effects of sodium diethyldithiocarbamate in fibroblasts V79 cells in relation to cytotoxicity, antioxidative enzymes, glutathione, and apoptosis.

Sodium diethyldithiocarbamate (DETC) is the main metabolite of disulfiram. Recently, we reported that mechanism of disulfiram cytotoxicity in V79 cells might be partially connected with thiol redox-state imbalance. Here, we examined the effect of DETC on the level of intracellular glutathione (GSH), protein oxidation (measured as PC-protein carbonyl content), lipid peroxidation (measured as TBARS-thiobarbituric acid reactive substances), antioxidant enzymatic defense, as well as on apoptosis. We used V79 Chinese hamster fibroblasts cells with and without modulated glutathione (GSH) level by N-acetyl-L-cysteine (NAC). We showed that treatment with DETC at concentrations that cause a moderate increase in thiol-state imbalance but not cell death stimulates oxidative stress measured as increased level of PC and TBARS, adaptive response of GSH-related enzymes and apoptosis. Our results show that cellular effects of DETC are partially attributable to the initial redox cellular state, since the increase of GSH level by NAC pre-treatment prevented the observed changes.

Evaluation of mutagenic activity of the organo-selenium compound Selol by use of the Salmonella typhimurium mutagenicity assay.

We examined the mutagenic activity of the anti-oxidant Selol, an organo-selenium compound, by use of the Salmonella typhimurium mutagenicity assay (Ames test) with strains TA97a, TA98, TA100, TA 1535 and TA102 in the absence and in the presence of metabolic activation with an S9 fraction from Aroclor-induced rat liver. Doses were 330, 500, 1000 and 5000 microg per plate. Selol contains the element selenium (valency, +4) in its structure and it may have chemopreventive and anticancer activity. Selol was found to be non-toxic and non-mutagenic for test doses up to 5% per plate (which designates the declared content of Selenium (+4) as 5000 microg per plate) in all the S. typhimurium strains.

Enzymatic antioxidant defense in isolated rat hepatocytes exposed to cadmium.

The aim of the study was the evaluation of cadmium effects on the activity of antioxidant enzymes in rat hepatocytes. The studies were conducted with isolated rat hepatocytes incubated for 1 or 2 hours in a modified (deprived of carbonates with phosphates) Williams' E medium (MWE) in the presence of cadmium chloride (25, 50 and 200 microM). Hepatocytes incubated in the MWE medium without cadmium chloride were used as a control. The application of the modified Williams' E medium allowed for the appearance of cadmium compounds in a soluble form that is indispensable for suitable estimation of its toxic action. There were evaluated markers of the oxidative stress such as: concentration of thiobarbiturate reactive substances (TBARS)--proportional to the level of lipid peroxidation, concentration of reduced glutathione (GSH), and the activity of antioxidant enzymes, including superoxide dismutase (SOD1 and SOD2), catalase (CAT), total glutathione peroxidase (GSHPx), selenium--dependent glutathione peroxidase (SeGSHPx), glutathione transferase (GST) and glutathione reductase (GSHR). Alterations of antioxidant enzymes activity, the level of TBARS and GSH in isolated rat hepatocytes caused by cadmium in vitro, were shown to depend on the concentration and time of exposure of cells to this metal. The increased level of TBARS and GSH was observed as well as changes in the activity of antioxidant enzymes. The activity of SOD isoenzymes and CAT was increased, whereas GSHPx and GST were decreased. These results indicate that cadmium induces oxidative stress followed by alterations in the cellular antioxidant enzyme system in isolated rat hepatocytes.

Changes in antioxidant defense systems induced by thiram in V79 Chinese hamster fibroblasts.

The role of antioxidant defence systems in protection against oxidative damage of lipids and proteins induced by fungicide thiram during in vitro exposure was investigated in cultured Chinese hamster V79 cells with normal, depleted, and elevated glutathione (GSH) levels. We analyzed the catalytic activities of superoxide dismutases (SOD1 and SOD2), Se-dependent and Se-independent glutathione peroxidases (GSH-Px), glutathione reductase (GR), and catalase (CAT), as well as total glutathione/glutathione disulfide ratio (GSH(total)/GSSG). Thiram treatment resulted in an increase in activities of SOD1, Se-dependent GSH-Px, and GR at the highest tested dose (150 microM). On the contrary, inhibition of CAT and Se-independent GSH-Px activities, and no significant changes in the level of SOD2 activity was observed at any tested doses (100-150 microM). GSH(total)/GSSG ratio in the 100 microM thiram treated cells was not significantly changed comparing to the control, despite significant decrease of GSH total (50%). In 150 microM thiram treated cells the ratio falls to 43% of control value. Pretreatment with l-buthionine sulfoximine (L-BSO), an inhibitor of GSH synthesis, significantly enhanced decrease in CAT and Se-independent GSH-Px activities, as well as GSH(total)/GSSG ratio, and reduced Se-dependent GSH-Px activity, following exposure to thiram. Simultaneously, L-BSO pretreatment enhanced increase in SOD1 activity, and had no effect on SOD2, following thiram exposure. Pretreatment with N-acetyl cysteine (NAC), a GSH precursor, prevented enzymatic changes in CAT, Se-dependent GSH-Px, GR, SOD1 activities, and significantly decreased SOD2 activity following exposure to thiram. GSH(total)/GSSG ratio was restored to the control value. This study suggests that following the changes in antioxidant defense systems thiram can act through the production of free radicals.

Induction of rat liver cytochrome P450 isoenzymes CYP 1A and CYP 2B by different fungicides, nitrofurans, and quercetin.

The genotoxic activity of environmental xenobiotics is manifested either in their direct interaction with cellular genetic material or in provoking secondary events, among which reactive oxygen species (ROS) production is a common phenomenon. Both pathways can be mediated by the activity of the cytochrome P450 monooxygenase system. We studied induction of the CYP 1A or CYP 2B monooxygenases in rat liver by the fungicides: thiram, captan, captafol, dodine and the drugs: nitrofurazone, furazolidone and the plant flavonoid: quercetin. A cytochrome P450 induction assay (CYPIA test) was used. S9 prepared from livers of rats treated with the test compounds were used to activate ethidium bromide (EtBr) (CYP 1A isoenzyme) or cyclophosphamide (CPA) (CYP 2B isoenzyme) in the Ames test. It was found that among the tested compounds, the most potent inducer of CYP 1A was furazolidone (3 x 80 mg/kg). Less potent was thiram (1 x 100mg/kg), as well as quercetin (3 x 80 mg/kg), and captafol (1 x 30 mg/kg). On the other hand, thiram (1 x 100 mg/kg), captafol (1 x 30 mg/kg), and quercetin (3 x 80 mg/kg) were most potent in the CYP 2B isoenzyme induction, while furazolidone (3 x 80 mg/kg), and nitrofurazone (3 x 80 mg/kg) appeared to be less potent in this respect. Captan and dodine (3 x 80 mg/kg) did not affect the activity of any of the cytochrome P450 isoenzymes.

Bacterial mutagenicity of dithiocarbamate fungicide thiram.

In the present work, within a project of re-evaluation of authorized pesticides coordinated by PZH (National Institute of Hygiene) we aimed at looking for a mechanism of induction of chromosomal aberrations by thiram. We checked its ability to damage bacterial DNA.

[Selected short-term bacterial and eukaryotic tests used for detection of genotoxicity of environmental chemical contaminants]. / Wybrane krótkoterminowe testy bakteryjne i na organizmach eukariotycznych, stosowane do oceny genotoksycznosci srodowiskowych zanieczyszczen chemicznych.

During the last years human exposure to environmental chemical contaminants (mutagenic/carcinogenic agents) has greatly increased. Evaluation of the biological effect of human exposure to mutagenic/carcinogenic agents in short-term tests is very important. In present paper the bacterial and eukaryotic tests for detection of genotoxic effect of environmental chemical contaminants have been described.

[The effect of nitrofurazone and furazolidone on induction of cytochrome P-450 in the CYPIA test]. / Okreslenie wplywu nitrofurazonu i furazolidonu na indukcje cytochromu P-450 w tescie-CYPIA.

The double CYPIA-TEST (cytochrome P-450) induction assay) was used to discriminate between the forms of cytochrome P-450 (the 3-methylcholantrene type (3-MC) or phenobarbital type (PB)) induced by nitrofurans; nitrofurazone and furazolidone. The test consisted of the studies by the technique of Ames of the ability of the S9 preparation obtained from livers of rats induced by nitrofurans to cause the metabolic activation of ethidium bromide (EtBr)--for induction of 3-MC type or cyclophosphamide (CPA) for PB type of cytochrome P-450. The mutagenicity of activated EtBr and CPA was checked with TA98 and TA100 of S. typhimurium respectively. It was found that only furazolidone at cumulative dose 3 x 80 mg/kg of body weight induce the 3-MC-type of cytochrome P-450.

Studies of the ability of rhamnetin and isorhamnetin to alkylate bacterial DNA.

The alkylation of bacterial DNA by two flavonoids: rhamnetin and isorhamnetin was examined using of strains Escherichia coli K-12: AB1157 (ada+) and (ada3). Rhamnetin and isorhamnetin and their putative metabolites formed in the presence of the S9 mix did not alkylate DNA at 0-6 of guanine.

Isolation and studies of the mutagenic activity of saponins in the Ames test.

Mutagenic activity of medicagenic acid, medicagenic acid 3-0-glucopyranoside and soyasaponin I was tested by the Ames method with S. typhimurium strains TA97, TA98, TA100, TA102 in the absence and the presence of metabolic activation (S9 mix). These saponins have been isolated and identified from alfalfa roots and clover Trifolium incarnatum seeds. All of them were found to be non-toxic and non-mutagenic for testing doses.

A study of the genotoxic potential of flavonoids using short-term bacterial assays.

Genotoxic activities of flavonoids (quercetin, rhamnetin, isorhamnetin, apigenin, luteolin) were investigated using two short-term bacterial assays. In the "repair test" in Salmonella typhimurium (strains TA1538 uvrB- and TA1978 uvrB+) the flavonoids studied did not introduce any damage into the DNA recognized by UvrABC nuclease (correndonuclease II). The results of the SOS-Chromotest in Escherichia coli K-12 strains PQ37 (tag+, alk+) and PQ243 (tagA, alkA) indicated that flavonoids only weakly induced the SOS system. The addition of a liver activation system (S9 mix) did not increase the mutagenic effect of the flavonoids tested. Two compounds: rhamnetin, isorhamnetin and their putative metabolites formed in the presence of the S9 mix did not alkylate DNA at N-3 of adenine.

Nonidentity of subunits of human kidney arginase A1 and human liver arginase A5.

The main forms of arginase A1 from human kidney and A5 from human liver were purified to homogeneity. Molecular weight of both forms of enzyme approximates 120,000. In the presence of EDTA these arginases dissociate into single type distinct subunits. M(r) of both kinds of subunits is 30,000. Similarly as native arginase forms, they differ in electric charge and display complete immunological incompatibility.