304 THE DEVELOPMENTAL TOXICITY EVALUATION OF 5-FLUOROURACIL AND INDOMETHACIN IN HUMAN EMBRYONIC STEM CELLS

Abstract

Embryonic stem cells have pluripotency and differentiate into and constitute the cells and tissues of our body. In this study, using human embryonic stem cells (hESC), we evaluated novel methods for screening toxicological chemicals during developmental process. We elucidated developmental toxicity of two well-known chemicals, 5-fluorouracil (5-FU) and indomethacin (Indo) in hESC. The undifferentiated hESC were treated with the chemicals (10^–4 to 10⁴ µM of 5-FU and Indo) in a dose-dependent manner during 1 to 3 days. Surface markers (SSEA-4, TRA-1-60, and TRA-1-81) expressed only in undifferentiated hESC were monitored by immunocytochemistry to ensure the characterisation of undifferentiated hESC. Moreover, expression of embryonic stem cell-specific genes was assessed with real-time PCR after treatment of 5-FU and Indo (10^–2, 10⁰, and10² µM of 5-FU and Indo). The expression of surface markers was not significantly affected by treatment of 5-FU and Indo. The expression of transcription factors (Oct-4, Sox-2, Nanog, and hTERT) was significantly decreased by high concentrations of 5-FU and Indo (10² µM). However, no difference was observed in treatment of low concentration of 5-FU and Indo (10^–2 µM). Taken together, these results suggest that 5-FU and Indo have cytotoxic effects, and modulate the expression of transcription factors that have pivotal roles in undifferentiated hESC. Therefore, we suggest that hESC may have potential to test toxicity of chemicals during embryonic developmental stage.