Activation of Wnt/β-Catenin Signaling Pathway Enhances the Derivation of Buffalo (Bubalus bubalis) Embryonic Stem Cell-Like Cells.

Wnt/β-Catenin signaling pathway plays an important role in maintaining self-renewal and pluripotency of human and mouse embryonic stem cells (ESCs). Activation of Wnt/β-Catenin signaling pathway by glycogen synthase kinase-3 (GSK3) inhibitor, the Wnt signaling agonist, could maintain the pluripotency of human and mouse ESCs in the presence of serum. However, the role of signaling pathway in the derivation of buffalo ESCs remains unclear. In this study, we used GSK3 inhibitors (6-bromoindirubin-3'-oxime [BIO] and CHIR99021) and investigated the effect of Wnt/β-Catenin activation on colony formation, proliferation, self-renewal, and pluripotency of Chinese swamp buffalo (buffalo) embryonic stem cell-like cells (ES-like cells), which were isolated from blastocysts. The results showed that buffalo ES-like cells displayed typical morphological characteristics of pluripotent stem cells: positive for alkaline phosphatase staining, expression of pluripotent markers, including OCT4, SOX2, SSEA-1, SSEA-4, LIN28, CH1, NANOG, and the proliferative markers, PCNA and C-MYC. Furthermore, activation of Wnt/β-Catenin signaling pathway by GSK3 inhibitors could promote colony formation and proliferation of buffalo ES-like cells and maintain their undifferentiated state, and upregulate the expression levels of pluripotent-related genes and proliferation-related genes. These results indicated that Wnt/β-Catenin signaling pathway plays an important role in the derivation and pluripotency of buffalo ES-like cells.