Combinatory estrogenic effects of bisphenol A in mixtures with alternariol and zearalenone in human endometrial cells

Humans are typically exposed to mixtures of substances, whereby their bioactivity can be significantly altered by co-occurring compounds. Thus, over the last years, research on combinatory effects has gained increasing attention. In particular, several xenoestrogens have been recently reported to interact synergistically, among them alternariol (AOH) and zearalenone (ZEN), two toxins produced by molds which contaminate crops or food commodities. Bisphenol A (BPA) is a potential food contaminant arising from its use in plastics and represents a well-known xenoestrogen, acting as an endocrine disruptor. However, little research was yet conducted on its impact on the bioactivity of other xenoestrogens, and vice versa. Thus, in this study, we focused on combinatory estrogenic effects of BPA with AOH and ZEN in Ishikawa cells, which represent a well-established, estrogen-sensitive human cell model. Estrogenic stimuli of the single compounds and binary combinations in constant concentration ratios were measured by assessing the activity of alkaline phosphatase, a natural reporter gene for estrogen receptor activation. In parallel, cytotoxicity was monitored by neutral red assay. For statistical analysis of combinatory effects the "combination index" model was applied. In combination with ZEN, BPA was found to cause additive estrogenic effects. Mixtures of BPA with AOH expressed moderately antagonistic to nearly additive combinatory effects, depending on the concentration ratio. Although no synergistic effects were measured in the applied chemical mixtures, additive estrogenic stimuli were observed, underlining the importance to consider the cumulative impact of endocrine active factors out of different sources and structural classes.