The Arbas cashmere goat is a unique biological resource that plays a vital role in livestock husbandry in China. LCDM is a medium with special small molecules (consisting of human LIF, CHIR99021, (S)-(+)-dimethindene maleate, and minocycline hydrochloride) for generation pluripotent stem cells (PSCs) with bidirectional developmental potential in mice, humans, pigs, and bovines. However, there is no report on whether LCDM can support for generation of PSCs with the same ability in Arbas cashmere goats. In this study, we applied LCDM to generate goat induced PSCs (giPSCs) from goat fetal fibroblasts (GFFs) by reprogramming. The derived giPSCs exhibited stem cell morphology, expressing pluripotent markers, and could differentiate into three germ layers. Moreover, the giPSCs differentiated into the trophectoderm lineage by spontaneous and directed differentiation in vitro. The giPSCs contributed to embryonic and extraembryonic tissue in preimplantation blastocysts and postimplantation chimeric embryos. RNA-sequencing analysis showed that the giPSCs were very close to goat embryos at the blastocyst stage and giPSCs have similar properties to typical extended PSCs (EPSCs). The establishment of giPSCs with LCDM provides a new way to generate PSCs from domestic animals and lays the foundation for basic and applied research in biology and agriculture.

The Arbas cashmere goat is a unique biological resource that plays a vital role in livestock husbandry in China. LCDM is a medium with special small molecules (consisting of human LIF, CHIR99021, (S)-(+)-dimethindene maleate, and minocycline hydrochloride) for generation pluripotent stem cells (PSCs) with bidirectional developmental potential in mice, humans, pigs, and bovines. However, there is no report on whether LCDM can support for generation of PSCs with the same ability in Arbas cashmere goats. In this study, we applied LCDM to generation goat induced PSCs (giPSCs) from goat fetal fibroblasts (GFFs) by reprogramming. The derived giPSCs exhibited stem cell morphology, expressing pluripotent markers, and could differentiate into three germ layers. Moreover, the giPSCs differentiated into the trophectoderm lineage by spontaneous and directed differentiation in vitro. The giPSCs contributed to embryonic and extraembryonic tissue in preimplantation blastocysts and postimplantation chimeric embryos. RNA-sequencing analysis showed that the giPSCs were very close to goat embryos at the blastocyst stage and giPSCs have similar properties to typical extended PSCs (EPSCs). The establishment of giPSCs with LCDM provides a new way to generate high quality of PSCs from domestic animals and lays the foundation for basic and applied research in biology and agriculture.

Methyl-CpG binding domain protein 3 (MBD3) is a member of the nucleosome remodeling and deacetylase (NuRD) corepressor protein complex, which plays a pivotal role in embryonic development, pluripotent stem cell (PSC) differentiation, and induced pluripotent stem cell (iPSC) reprogramming. However, whether MBD3 is also involved in iPSC differentiation into primordial germ cell-like cells (PGCLCs) is unclear. In this study, bovine iPSCs generated by MBD3 knockdown (MBD3 KD) during reprogramming were subjected to PGCLC differentiation using a monolayer cell culture method. Our results revealed that MBD3 KD enhanced the ability of bovine iPSCs to differentiate into PGCLCs. MBD3-KD bovine iPSC-derived PGCLCs exhibited the characteristics of in vivo primordial germ cells (PGCs) in a migratory state. MBD3 KD increased NODAL signal transduction during the induction of PGCLCs and was associated with increased expression of PRDM1 and SOX15. Moreover, MBD3 KD increased the bivalent modification of transforming growth factor beta (TGF-β)-related genes concomitant with the upregulation of DPPA4 and DNMT3L expression in bovine iPSCs, and this epigenetic change was associated with enhanced NODAL signaling. Our results indicate that MBD3 KD during reprogramming favors the differentiation of bovine iPSCs into PGCLCs, further highlighting the impact of MBD3 on histone modifications during germ cell development.