Tension-induced enhancement of SIX1 expression during preplacodal ectoderm differentiation from human induced pluripotent stem cells

Based on observations of <i>in vivo</i> morphogenesis, differentiation is expected to be regulated by mechanical cues. However, the detail mechanisms remain largely unknown. A previous study using human pluripotent stem cells (hPSCs) demonstrated that neural plate border (NPB) specification was enhanced by mechanical force. However, it is unknown whether mechanical force is also involved in the specification of the preplacodal ectoderm (PPE), which is derived from the NPB. Here, we verified the validity of the PPE induction method in stretch chambers, and conducted the stretching stimuli experiments. When repetitive stretching stimuli were applied from Day 2 to 10 or Day 2 to 7, expression of the PPE marker <i>SIX1</i> was increased. However, this increase was not observed when the stimuli were applied from Day 5 to 10, suggesting there is a critical period of sensitivity to mechanical forces. Immunofluorescent staining revealed lower active β-catenin signals in the cell sheet in the stretched samples compared to those in the controls, suggesting a negative correlation between stretching stimuli and Wnt signaling. Our finding suggests that mechanical force is important in PPE differentiation.