Animal Bioscience

  • 제36권8호
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  • Pages.1180-1189
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  • 2023
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  • 2765-0189(pISSN)
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  • 2765-0235(eISSN)
아세아태평양축산학회 (Asian Australasian Association of Animal Production Societies)
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The number of primitive endoderm cells in the inner cell mass is regulated by platelet-derived growth factor signaling in porcine preimplantation embryos

  • Jong-Nam Oh (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Mingyun Lee (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Gyung Cheol Choe (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Dong-Kyung Lee (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Kwang-Hwan Choi (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Seung-Hun Kim (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Jinsol Jeong (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University) ;
  • Chang-Kyu Lee (Department of Agricultural Biotechnology, Animal Biotechnology Major, and Research Institute of Agriculture and Life Sciences, Seoul National University)
  • 투고 : 2022.12.21
  • 심사 : 2023.02.15
  • 발행 : 2023.08.01
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초록

Objective: Discovering the mechanism of cell specification is important to manipulate cellular lineages. To obtain lineage-specific cell lines, the target lineage needs to be promoted, and counterpart lineages should be suppressed. Embryos in the early blastocyst stage possess two different cell populations, the inner cell mass (ICM) and trophectoderm. Then, cells in the ICM segregate into epiblasts (Epi) and primitive endoderm (PrE). PrE cells in embryos show specific expression of platelet-derived growth factor (PDGF) and its receptor, PDGF receptor A (PDGFRA). In this study, we suppressed PDGF signaling using two methods (CRISPR/Cas9 injection and inhibitor treatment) to provide insight into the segregation of embryonic lineages. Methods: CRISPR/Cas9 RNAs were injected into parthenogenetically activated and in vitro fertilized embryos. The PDGF receptor inhibitor AG1296 was treated at 0, 5, 10, and 20 µM concentration. The developmental competence of the embryos and the number of cells expressing marker proteins (SOX2 for ICM and SOX17 for PrE) were measured after the treatments. The expression levels of the marker genes with the inhibitor were examined during embryo development. Results: Microinjection targeting the PDGF receptor (PDGFR) A reduced the number of SOX17-positive cell populations in a subset of day 7 blastocysts (n = 9/12). However, microinjection accompanied diminution of Epi cells in the blastocyst. The PDGF receptor inhibitor AG1296 (5 µM) suppressed SOX17-positive cells without reducing SOX2-positive cells in both parthenogenetic activated and in vitro fertilized embryos. Within the transcriptional target of PDGF signaling, the inhibitor significantly upregulated the Txnip gene in embryos. Conclusion: We identified that PDGF signaling is important to sustain the PrE population in porcine blastocysts. Additionally, treatment with inhibitors was a better method to suppress PrE cells than CRISPR/Cas9 microinjection of anti-PDGF receptor α gene, because microinjection suppressed number of Epi cells. The PDGF receptor might control the number of PrE cells by repressing the proapoptotic gene Txnip. Our results can help to isolate Epi-specific cell lines from blastocysts.

키워드

과제정보

This work was supported by the BK21 Four program, the Korea Evaluation Institute of Industrial Technology (KEIT) through the Alchemist project funded by the Ministry of Trade, Industry and Energy (MOTIE; 20012411), and the National Research Foundation of Korea (NRF) grant funded by the government of Republic of Korea (NRF-2021R1A2C4001837).

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