Student Seoul National University, Seoul-t'ukpyolsi, Republic of Korea
Abstract: The derivation of trophoblast stem cell (TS) lines from porcine embryos produced in vitro provides a powerful platform for studying placental development and embryo-maternal interactions in ungulates. Porcine TS offer a valuable model for studying placental development and implantation in ungulates due to their dynamic nature and ability to form non-invasive epitheliochorial placentas. In this study, we aimed to optimize culture conditions for inducing and maintaining porcine TS from in vitro-produced embryos. Initially, we replicated culture conditions based on previous studies, which led to an increase in the expression of epithelial-mesenchymal transition (EMT)-related genes. As the cells were sub-cultured, noticeable changes in cell morphology were observed, prompting us to identify specific chemical combinations that could either maintain the original cell shape or revert the cells to their initial morphology. Taken together, we confirmed the expression of TS-specific markers in both the pig embryos and the outgrown cells, validating their identity and lineage. To establish stable TS lines, we systematically evaluated various media compositions, growth factors, and signaling modulators to determine their effects on cell adhesion, colony formation, and stemness maintenance. During the derivation process, we identified distinct subpopulations of TE cells and classified the subtype that exhibited the highest capacity for maintaining stemness as trophoblast stem cells (TS). Combinations of specific signaling molecules were identified as critical for preserving TS-specific marker expression, while promoting robust attachment and colony formation. This comprehensive analysis provides new insights into the molecular mechanisms governing TS behavior, offering a foundation for further studies on trophoblast differentiation and placental development in pigs. Our results contribute to a broader understanding of TS biology, with potential applications in improving reproductive efficiency and addressing placental dysfunctions in livestock.
Funding Source: This work was supported by the BK21 Four program, the Korea Evaluation Institute of Industrial Technology (KEIT) [20012411]; and the National Research Foundation of Korea (NRF) grant [2023R1A2C1005026].
