Activation of Wnt/ß-catenin signaling in ESC promotes rostral forebrain differentiation in vitro

Wnt/ß-catenin signaling is crucial for maintenance of pluripotent state of embryonic stem cell (ESC). However, it is unclear how Wnt/ß-catenin signaling affects the differentiation ability of ESC, especially with regard to rostral forebrain cells. Here, using Rax, rostral forebrain marker, and Wnt/ß-catenin reporter lines, we report ratio of Rax⁺ and Wnt responding tissue (Wnt⁺) patterns, which were affected by seeding number of ESC in three-dimensional culture system. Surprisingly, we found ß-catenin level and localization are heterogeneous in ESC colony by immunostaining and time-laps imaging of ß-catenin-mEGFP signals. Moreover, activation of Wnt signaling in ESC promoted expression level and nuclear localization of ß-catenin, and mRNA levels of Wnt antagonists, axin2 and dkk1, leading to upregulating Wnt/ß-catenin reporter in ESC state and Rax expression at differentiation culture day 7. Together, our results suggest that activation of Wnt signaling in ESC promotes the differentiation efficacy of rostral forebrain cells. Wnt-priming culture method may provide a useful tool for applications in the areas of basic science and molecular therapeutics for regenerative medicine.