Abstract
We have reported an approach for evaluating developmental toxicity through an FGF-SRF signal reporter assay utilizing human iPS cells (S. Kanno et al., iScience, 2022). Signal interactions are vital for the regulation of fetal development, and certain developmental toxicants are known to disrupt signaling pathways. Considering these facts, we hypothesized that developmental toxicity eventually relates to signal disruption, and thus established a signal reporter assay. In this assay, chemiluminescence associated with FGF signaling activity was continuously monitored, demonstrating that signal disruption occurred at different time points depending on developmental toxicants. By integrating signal disruption over time, it was possible to accurately distinguish known developmental toxicants. In this study, we further improved the approach for more comprehensive analysis of signal disruption. In our previous approach, chemiluminescence was manually measured at 2, 4, 6, 8, 10, and 24 hours of culture. Because continuous measurements over several days were difficult to perform, disruption between the time points might be missed and longer-term disruption tests were not possible. To address these issues, we conducted the assay using a real-time luminescence measurement system. The continuous monitoring of chemiluminescence reveals that there were two peaks in for 72 hours, unlike the single peak in 24 hours. Furthermore, we found that some developmental toxicants, such as valproic acid, enhanced signal disruption after more than 24 hours and these were readily detected by continuous and long-term luminescence monitoring. Our approach holds great potential as a screening tool for developmental toxicants.
