Embryonic midbrain micromass cultures were exposed for five days to ochratoxin A (OTA) at seven concentrations (ranging from 0.16 to 10 µg/mL). Cell viability was assessed in neutral red uptake test (NRU), and differentiation -by immunoenzymatic determination of structural proteins (β III -tubulin, MAP2, GFAP) expression level as well as by computer image analysis. Dose dependent decrease in cell number and differentiation was observed. Concentration-response curves were analysed and the mean inhibition concentrations (µg/mL) for cytotoxicity (IC 50 ) and differentiation (ID 50 ) were calculated. There were no significant differences in the sensitivity of neurons in early and late stage of differentiation and astrocytes to the toxic activity of this compound. For all endpoints ID 50 value was very low (< 10 µg/mL) so OTA was classified as a strong teratogen. IC 50 / ID 50 ratios <2 pointed out that with harmful action of OTA the basic cytotoxicity should be connected.
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.
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% CO 2 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 (EC 50 >200µg/mL). The exception was ronidazole, which demonstrated a distinct endpoint sensitivity related to the species. EC 50 (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 (EC 20 ) 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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