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  • Author: Maria Minta x
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Comparison of albendazole cytotoxicity in terms of metabolite formation in four model systems

Abstract

Introduction: Albendazole is used to treat endoparasitic diseases in animals and humans. After oral administration, it is quickly oxidised into its pharmacologically active metabolite albendazole sulfoxide and then to sulfone. However, it is not clear which compound is responsible for toxic effects towards mammalian cells. Material and Methods: The model systems comprised cultures of isolated rat hepatocytes, two hepatoma cell lines (FaO, HepG2), and non-hepatic Balb/c 3T3 line. Cells were exposed for 24, 48, and 72 h to eight concentrations of albendazole ranging from 0.05 to 100 μg/mL. At all three time points cytotoxic effects were assessed by MTT assay and metabolites in the culture media were determined by LC-MS/MS analysis. Results: The effective concentrations EC50-72h showed that Balb/c 3T3 cells were the most sensitive to albendazole (0.2 ±0.1 μg/mL) followed by FaO (1.0 ±0.4 μg/mL), and HepG2 (6.4 ±0.1 μg/mL). In the case of isolated hepatocytes this value could not be attained up to the highest concentration used. Chemical analysis revealed that the concentrations of albendazole in hepatocytes and HepG2 and FaO culture media gradually decreased with incubation time, while the concentrations of its metabolites increased. The metabolism in isolated hepatocytes was dozens of times greater than in HepG2 and FaO cells. Two metabolites (albendazole sulfoxide, albendazole sulfone) were detected in isolated hepatocytes and HepG2 culture medium, one (albendazole sulfoxide) in FaO culture medium and none in Balb/c 3T3. Conclusion: The obtained data indicate that metabolism of albendazole leads to its detoxification. The lower cytotoxic potential of metabolites was confirmed in the independent experiments in this study.

Open access
Cytotoxicity of Some Nitroimidazole Derivatives - Comparative Studies on Human and Rat Hepatoma Cell Lines

Abstract

The cytototoxic potential of metronidazole, tinidazole, ronidazole, and ornidazole, using human and rat hepatoma cell lines (HepG2 and FaO) in culture was assessed. The cells were treated with drugs for 24, 48 and 72 h at 37 °C in 5% CO2 at concentrations of 0.1 to 200 μg/mL. Following the treatment period, the cells were assayed by four independent assays: MTT reduction, neutral red uptake (NRU), total protein content (TPC), and LDH leakage. The results suggest that nitroimidazoles are of low cytotoxic potential (EC50 >200μg/mL). The exception was ronidazole, which demonstrated a distinct endpoint sensitivity related to the species. EC50 (μg/mL) in human cells were: in MTT assay - 196±5.5 and 122±9.3 at 24 and 48 h, respectively, and in NRU assay - 150±1.25 at 72 h. Based on minimal toxic concentrations (EC20) for ronidazole, determined by all methods used in HepG2 cells, it could be concluded that their sensitivity was as follows: MTT>NRU>LDH>TPC.

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Influence of dietary soy isoflavones on immature hamster uterotrophic and Hershberger assays

Abstract

To select appropriate diet for hamsters used in the uterotrophic and Hershberger assays two rodent diets were compared: Murigran (Agropol, Poland) and Altromin 7010 (Altromin Spezialfutter GmbH&Co., Germany). The contents of bioactive compounds in feeds were evaluated by liquid chromatography, and their oestrogenic activity by yeast enhanced green fluorescent protein assay. In opposition to Altromin, Murigran contained high amounts (μg/kg) of genistein (765 600) and daidzein (132 000), and the oestrogenic activity of these compounds, expressed as 17β-oestradiol equivalent concentration (EEQ), was found to be 9.54 μg EEQ/kg. In in vivo study, Murigran induced a high degree of oestrogenisation in immature hamsters, and females failed to exhibit a normal uterine response to recommended dose of a model oestrogen agonist 17α-ethinyloestradiol. There was no influence of the diet on the weight of five accessory sex organs (ASO): ventral prostate, seminal vesicles with coagulating glands, levator ani bulbocavernosus muscles, Cowper`s glands, and glans penis of control males. However, the impact on ASO response to model androgen agonist, testosterone propionate was observed. The obtained results provide the evidence that phytooestrogen-rich feed modulates the oestrogenic and androgenic response to chemicals.

Open access
Influence of fluoroquinolones on viability of Balb/c 3T3 and HepG2 cells

Abstract

The cytotoxic potential of fluoroquinolones (enrofloxacin, ciprofloxacin, difloxacin, sarafloxacin, danofloxacin, norfloxacin and marbofloxacin) was investigated using mouse fibroblasts Balb/c 3T3 and human hepatoma HepG2 cell lines. The cells were exposed for 24, 48, and 72 h to drugs at eight concentrations ranged from 0.78 to 100 μg/mL. Four independent cytotoxicity assays were applied, in which various endpoints were assessed: mitochondrial activity - MTT reduction, lysosomal activity - neutral red uptake, total protein content, and cellular membrane integrity - lactate dehydrogenase release. Mean effective cytotoxic concentrations (EC50) calculated at different time points from concentration-response curves ranged from 10 to 100 μg/mL. The most affected endpoint in both cell lines was mitochondrial activity. The EC50-MTT-72 h <10 μg/mL was found for difloxacin, marbofloxacin (fibroblasts), sarafloxacin, and norfloxacin (HepG2). The data shows that cytotoxicity of the fluoroquinolones appears after longer exposure of both cell cultures to these compounds.

Open access
Differential toxicities of albendazole and its two main metabolites to Balb/c 3T3, HepG2, and FaO lines and rat hepatocytes

Abstract

Introduction: The cytotoxicity of anthelmintic agent, albendazole (ABZ) and its two major metabolites, sulfoxide (ABZSO) and sulfone (ABZ-SO2), on non-hepatic Balb/c 3T3 line, two hepatoma cell lines (FaO, HepG2), and isolated rat hepatocytes was investigated. Material and Methods: Cell cultures were exposed for 24, 48, and 72 h to eight concentrations of the compounds ranging from 0.05 to 100 μg/mL (ABZ) and from 0.78 to 100 μg/mL (ABZ-SO and ABZ-SO2). Three different assays were applied in which various biochemical endpoints were assessed: lysosomal activity - neutral red uptake (NRU) assay, proliferation - total protein contents (TPC) assay and lactate dehydrogenase (LDH) leakage assay. Results: The most toxic was albendazole whose EC50 values calculated from the concentration effect curves ranged from 0.2 to 0.5 μg/mL (Balb/c 3T3 ) and from 0.4 to 73.3 μg/mL (HepG2). Rat hepatoma line and isolated rat hepatocytes were less sensitive to the impact of ABZ. Toxic action expressed as EC50 was recorded after 72 h exposure only in LDH release assay at 0.8 μg/mL and 9.7 μg/mL respectively. The toxicity of metabolites was much lower. The most sensitive to ABZ-SO were fibroblasts and EC50-72h values were similar in all three assays used, i.e. NRU (14.1 μg/mL), TPC (15.8 μg/mL), and LDH (20.9 μg/mL). In the case of ABZ-SO2 the mean effective concentrations were the highest, and could be reached only in one LDH assay. These values (μg/mL) were as follows: 65.3 (FaO), 65.4 (HepG2), 75.8 (hepatocytes), and 77.4 (Balb/c 3T3). Conclusion: The differences in in vitro toxicity of albendazole depend on metabolic ability of the cellular models. Primary cultured rat hepatocytes represent a valuable tool to study the impact of biotransformation on the cytotoxicity of drugs.

Open access
Usefulness of immature golden hamster (Mesocricetus auratus) as a model for uterotrophic assay

Abstract

The present study assessed the sensitivity of immature hamster uterotrophic assay to reference oestrogen agonists/antagonists in order to develop a sensitive model for evaluation of endocrine-active compounds in diets. After performing a baseline for control animals, the sensitivity of immature females (postnatal day 18) to reference compounds was evaluated in a three-day uterotrophic assay. The absolute and adjusted dry uterine weights, fold induction over control for absolute wet uterine weight, and wet uterine weight/body weight ratio (%) were used as endpoints. The significantly active doses for reference oestrogens were as follows: 0.6 μg/kg for 17α-ethinyloestradiol (s.c.); 1 μg/kg/day (s.c.) and 40 μg/kg (p.o.) for diethylstilboestrol; 40 mg/kg (s.c.) and 160 mg/kg (p.o.) for bisphenol A. Co-treatment with tamoxifen at a dose of 1 mg/kg significantly antagonised the uterotrophic effect induced by 1 μg/kg 17α-ethinyloestradiol, and showed the attenuated proliferative effect in histopathological examination. We found immature hamster uterotrophic assay as a sensitive model that could be a good alternative to the rat assay.

Open access
Cytotoxic effects of the synthetic oestrogens and androgens on Balb/c 3T3 and HepG2 cells

Abstract

The aim of the study was to test and compare the cytotoxic potential of two synthetic oestrogens: diethylstilboestrol (DES) and ethinyloestradiol (EE2) and two androgens: testosterone propionate (TP) and trenbolone (TREN) on two cell lines. The fibroblast cell line Balb/c 3T3 and the hepatoma cell line HepG2 were selected. To get more insight into the mode of toxic action, four methods were used, which evaluated different biochemical endpoints: mitochondrial activity (3-(4,5-dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide reduction assay), lysosomal activity (neutral red uptake assay), total protein content, and lactate dehydrogenase release. Cytotoxicity was assessed after 24, 48, and 72 h exposure to eight concentrations ranging from 0.78 to 100 μg/mL. Concentration- and time- dependent effects were observed. Depending on the line and assay used, half maximal effective concentration after 72 h (EC50-72h) values ranged as follows: DES 1-13.7 μg/mL (Balb/c 3T3) and 3.7-5.2 μg/mL (HepG2); EE2 2.1-14.3 μg/mL (Balb/c 3T3) and 1.8-7.8 μg/mL (HepG2); TP-14.9-17.5 μg/mL (Balb/c 3T3), and 63.9- 100 μg/mL (HepG2); and TREN 11.3-31.4 μg/mL (Balb/c 3T3) and 12.5-59.4 μg/mL (HepG2). The results revealed that oestrogens were more toxic than androgens and the most affected endpoint was mitochondrial activity. In contrast to oestrogens, for which EC50-72h values were similar in both lines and by all assays used, Balb/c 3T3 cells were more sensitive than HepG2 cells to TP.

Open access