Generating stable livestock pluripotent stem cells (PSCs) can be used for complex genome editing, cellular agriculture, gamete generation, regenerative medicine and in vitro breeding schemes. Over the past decade, significant progress has been made in characterizing pluripotency markers for livestock species. In this study, we investigated embryo development and gene expression of the core pluripotency triad (OCT4, NANOG, SOX2) and cell lineage commitment markers (REX1, CDX2, GATA4) in the presence of three small molecules and their combination [PD0325901 (FGF inhibitor), SB431542 (TGFβ inhibitor), and CHIR99021 (GSK3B inhibitor)] from day 2–7 post-insemination in goat.

Significant reduction in rate of blastocyst formation was observed when SB was used along with PD or CHIR and their three combinations had more sever effect. SB and CHIR decreased the expression of SOX2 while increasing the GATA4 expression. PD decrease the relative expression of NANOG, OCT4 and GATA4, while increased the expression of REX1. Among the combination of two molecules, only SB + CHIR combination significantly decreased the expression of GATA4, while the combination of the three molecules significantly decreases the expression of NANOG, SOX2 and CDX2.

According to these results, the inhibition of the FGF signaling pathway, by PD may lead to blocking the hypoblast formation as observed by reduction of GATA4. OCT4 and NANOG expressions did not show signs of maintenance pluripotency. GATA4, NANOG and OCT4 in the PD group were downregulated and REX1 as EPI-marker was upregulated thus REX1 may be considered as a marker of EPI/ICM in goat.

生成稳定的家畜多能干细胞（PSC）可用于复杂的基因组编辑、细胞农业、配子生成、再生医学和体外育种计划。在过去的十年中，在表征牲畜物种的多能性标记方面取得了重大进展。在这项研究中，我们研究了在三种小分子及其组合存在的情况下胚胎发育和核心多能性三联体（OCT4、NANOG、SOX2）和细胞谱系定型标记（REX1、CDX2、GATA4）的基因表达[PD0325901（FGF抑制剂） ）、SB431542（TGFβ 抑制剂）和 CHIR99021（GSK3B 抑制剂）] 山羊授精后第 2-7 天。

当SB与PD或CHIR一起使用时，观察到囊胚形成率显着降低，并且它们的三种组合具有更严重的效果。SB和CHIR降低了SOX2的表达，同时增加了GATA4的表达。PD使NANOG、OCT4和GATA4的相对表达减少，而REX1的表达增加。两种分子的组合中，只有SB+CHIR组合显着降低了GATA4的表达，而三种分子的组合显着降低了NANOG、SOX2和CDX2的表达。

根据这些结果，PD 对 FGF 信号通路的抑制可能导致阻碍下胚层形成，如GATA4减少所观察到的。OCT4和NANOG表达没有显示出维持多能性的迹象。PD组中GATA4、NANOG和OCT4表达下调，而作为EPI标志物的REX1表达上调，因此REX1可能被认为是山羊EPI/ICM的标志物。