STUDY QUESTION: How does activation of aryl hydrocarbon receptor (AHR) signaling affect human trophoblast cell development and differentiation? SUMMARY ANSWER: AHR activation alters gene expression without impairing the ability of trophoblast cells to maintain a stem cell state or differentiate into essential cell types, such as extravillous trophoblast (EVT) cells or syncytiotrophoblast (ST), while promoting the production of 2-methoxy estradiol (2ME), which may impact placental development. WHAT IS KNOWN ALREADY: The placenta serves both as a nutrient delivery system and a protective barrier against environmental toxins. AHR signaling is known to mediate cellular responses to environmental pollutants, potentially affecting trophoblast cell function, but the specific impacts of AHR activation on these cells were not fully understood. STUDY DESIGN, SIZE, DURATION: This study utilized an in vitro model of human trophoblast stem (TS) cells to investigate the downstream effects of AHR activation. The study focused on both undifferentiated TS cells and cells undergoing differentiation. PARTICIPANTS/MATERIALS, SETTING, METHODS: Human TS cells were used as a model system. Researchers examined the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exposure in TS cells maintained in their stem state and in TS cells induced to differentiate into EVT cells or ST. The study assessed changes in gene expression, particularly focusing on CYP1A1 and CYP1B1, as well as the production of 2ME. MAIN RESULTS AND THE ROLE OF CHANCE: AHR activation stimulated the expression of CYP1A1 and CYP1B1, key genes associated with AHR signaling, in both undifferentiated and differentiating trophoblast cells. While AHR activation did not impact the ability of the cell to remain in a stem state or differentiate, it increased the production of 2ME, which may influence placentation. These effects were dependent on AHR signaling. LARGE SCALE DATA: n/a. LIMITATIONS, REASONS FOR CAUTION: This study was conducted in vitro, which may not fully replicate in vivo conditions. Further research is needed to confirm whether these findings apply to placental development in humans. WIDER IMPLICATIONS OF THE FINDINGS: The results suggest that AHR signaling activated by environmental pollutants could have a significant impact on placental development through mechanisms involving AHR activation. These findings may have broader implications for understanding how environmental factors affect fetal development. STUDY FUNDING/COMPETING INTEREST(S): This work was funded by the National Institutes of Health: ES028957, HD020676, ES029280, HD105734, HD112559, and the Sosland Foundation. The authors declare no conflicts of interest.