Although there are many studies about pluripotent stem cells, little is known about pluripotent pathways and the difficulties of maintaining the pluripotency of bovine cells in vitro. Here, we investigated differently expressed genes (DEG) in bovine embryo-derived stem-like cells (eSLCs) from various origins to validate their distinct characteristics of pluripotency and differentiation. We identified core pluripotency markers and additional markers which were not determined as pluripotency markers yet in bovine eSLCs. Using the KEGG database, TGFβ, WNT, and LIF signaling were related to the maintenance of pluripotency. In contrast, some DEGs related to the LIF pathway were down-regulated, suggesting that reactivation of the pathway may be required for the establishment of true bovine embryonic stem cells (ESCs). Interestingly, oncogenes were co-down-regulated, while tumor suppressor genes were co-up-regulated in eSLCs, implying that this pattern may induce abnormal teratomas. These data analyses of signaling pathways provide essential information on authentic ESCs in addition to providing evidence for pluripotency in bovine eSLCs.