engleski

Cytotoxicity of environmental estrogens in fish cell line - impact of serum

Fish cell lines are becoming increasingly important in the aquatic ecotoxicology. The discrepancy between whole fish (acute toxicity) and fish cell lines (cytotoxicity) is mostly attributed to the lower bioavailability of tested substances in vitro. During the in vitro testing, the cells are cultivated in the media with serum which can modify the outcome of cytotoxicity studies. This is mainly due to binding of test substance to serum proteins, which affects the concentration-effect ratio. In this study, we tested the impact of serum on the diethylstilbesterol (DES) and 17α-ethynylestradiol (EE2) cytotoxicity in CCO (Channel Catfish Ovary) cell line. The cells were treated with DES and EE2 (0.1 to 10 μg/mL) in DMEM medium with 2.5 %, 5 % or 10 % (v/v) foetal bovine serum (FBS) or serum stripped with charcoal (Cs-FBS). After 72 hours of treatment, the viability of CCO cells was determined by WST-1 method. DES (0.1 μg/mL) induced stimulatory effect in range of 15 % to 40 % compared to control, while higher concentrations (5 μg/mL and 10 μg/ml) had inhibitory effects of 15 % to 60 % and 40 to 70 %, respectively. The same was observed for EE2, the stimulation of proliferation (15 % to 30 %) with 0.1 μg/mL and inhibition of 5 μg/mL and 10 μg/mL (20 % to 35 % and 30 % to 50 %). Calculated EC50 values showed that the influence of DES and EE2 on CCO cells depend on the type and quantity of serum. Further research is necessary to elucidate which culture conditions better correlate with in vivo toxicity and thus improve the prediction of human acute toxicity, which is the ultimate goal of alternative in vitro testing.

CCO cells; diethylstilbestrol; EC50; 17alpha-ethynylestradiol; foetal bovine serum

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