Developmental Toxicity Assessment Using Human iPSCs by Automated Measurement of FGF Signaling Disruption

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. we hypothesized that developmental toxicity eventually relates to signal disruption, and thus established a signal reporter assay (DynaLux/c). 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. However, chemiluminescence was　measured manually in this method. Thereby, it was difficult to capture detailed temporal changes and to measure during the night. Therefore, we automated luminescence measurements, establishing a method enabling detailed and prolonged luminescence assessments. The continuous monitoring of chemiluminescence reveals that there were two peaks in 72 hours, unlike the single peak in 24 hours. Moreover, substances with developmental toxicity causing expanded disruption after 24 hours, such as valproic acid, were more accurately detected. The automation of luminescence measurements demonstrated the potential for more precise tracking of signal disruption. Future efforts will focus on optimizing measurement conditions and increasing the number of test substances to further elucidate the utility of this method.