Department of Obstetrics and Gynecology, Key Laboratory for Major Obstetric Diseases of Guangdong Province, Key Laboratory of Reproduction and Genetics of Guangdong Higher Education Institutes, The Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, Guangdong, China.
Yunnan Key Laboratory of Primate Biomedical Research, Institute of Primate Translational Medicine, Kunming University of Science and Technology, Kunming, China.
Cell Prolif. 2022 Nov;55(11):e13317. doi: 10.1111/cpr.13317. Epub 2022 Jul 26.
This study aims to optimize the human extended pluripotent stem cell (EPSC) to trophectoderm (TE)-like cell induction with addition of EGF and improve the quality of the reconstructing blastoids.
TE-like cells were differentiated from human EPSCs. RNA-seq data analysis was performed to compare with TE-like cells from multiple human pluripotent stem cells (hPSCs) and embryos. A small-scale compound selection was performed for optimizing the TE-like cell induction and the efficiency was characterized using TE-lineage markers expression by immunofluorescence stanning. Blastoids were generated by using the optimized TE-like cells and the undifferentiated human EPSCs through three-dimensional culture system. Single-cell RNA sequencing was performed to investigate the lineage segregation of the optimized blastoids to human blastocysts.
TE-like cells derived from human EPSCs exhibited similar transcriptome with TE cells from embryos. Additionally, TE-like cells from multiple naive hPSCs exhibited heterogeneous gene expression patterns and signalling pathways because of the incomplete silencing of naive-specific genes and loss of imprinting. Furthermore, with the addition of EGF, TE-like cells derived from human EPSCs enhanced the TE lineage-related signalling pathways and exhibited more similar transcriptome to human embryos. Through resembling with undifferentiated human EPSCs, we elevated the quality and efficiency of reconstructing blastoids and separated more lineage cells with precise temporal and spatial expression, especially the PE lineage.
Addition of EGF enhanced TE lineage differentiation and human blastoids reconstruction. The optimized blastoids could be used as a blastocyst model for simulating early embryonic development.
本研究旨在通过添加 EGF 优化人扩展多能干细胞（EPSC）向滋养外胚层（TE）样细胞的诱导，并提高重构胚泡的质量。
从人 EPSC 中分化出 TE 样细胞。进行 RNA-seq 数据分析，以比较来自多种人类多能干细胞（hPSC）和胚胎的 TE 样细胞。进行小规模化合物筛选以优化 TE 样细胞诱导，并用免疫荧光染色检测 TE 谱系标志物表达来表征效率。通过三维培养系统，使用优化的 TE 样细胞和未分化的人 EPSC 来生成胚泡。进行单细胞 RNA 测序，以研究优化的胚泡向人类囊胚的谱系分离。
人 EPSC 来源的 TE 样细胞表现出与胚胎 TE 细胞相似的转录组。此外，由于幼稚特异性基因不完全沉默和印迹丢失，多种原始 hPSC 来源的 TE 样细胞表现出异质的基因表达模式和信号通路。此外，添加 EGF 可增强人 EPSC 来源的 TE 样细胞中与 TE 谱系相关的信号通路，并表现出与人类胚胎更相似的转录组。通过与未分化的人 EPSC 相似，我们提高了重构胚泡的质量和效率，并分离出更多具有精确时空表达的谱系细胞，特别是 PE 谱系。
添加 EGF 可增强 TE 谱系分化和人类胚泡重建。优化的胚泡可用作模拟早期胚胎发育的囊胚模型。
