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Primary specification of blastocyst trophectoderm by scRNA-seq: New insights into embryo implantation.

ABSTRACT: Mechanisms of implantation such as determination of the attachment pole, fetal-maternal communication, and underlying causes of implantation failure are largely unexplored. Here, we performed single-cell RNA sequencing on peri-implantation embryos from both humans and mice to explore trophectoderm (TE) development and embryo-endometrium cross-talk. We found that the transcriptomes of polar and mural TE diverged after embryos hatched from the zona pellucida in both species, with polar TE being more mature than mural TE. The implantation poles show similarities in cell cycle activities, as well as in expression of genes critical for implantation and placentation. Embryos that either fail to attach in vitro or fail to implant in vivo show abnormalities in pathways related to energy production, protein metabolism, and 18S ribosomal RNA m⁶A methylation. These findings uncover the gene expression characteristics of humans and mice TE differentiation during the peri-implantation period and provide new insights into embryo implantation.

SUBMITTER: Liu D

PROVIDER: S-EPMC9365277 | biostudies-literature | 2022 Aug

REPOSITORIES: biostudies-literature

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Primary specification of blastocyst trophectoderm by scRNA-seq: New insights into embryo implantation.

Liu Dandan D Chen Yidong Y Ren Yixin Y Yuan Peng P Wang Nan N Liu Qiang Q Yang Cen C Yan Zhiqiang Z Yang Ming M Wang Jing J Lian Ying Y Yan Jie J Zhai Fan F Nie Yanli Y Zhu Xiaohui X Chen Yuan Y Li Rong R Chang Hsun-Ming HM Leung Peter C K PCK Qiao Jie J Yan Liying L

Science advances 20220810 32

Mechanisms of implantation such as determination of the attachment pole, fetal-maternal communication, and underlying causes of implantation failure are largely unexplored. Here, we performed single-cell RNA sequencing on peri-implantation embryos from both humans and mice to explore trophectoderm (TE) development and embryo-endometrium cross-talk. We found that the transcriptomes of polar and mural TE diverged after embryos hatched from the zona pellucida in both species, with polar TE being mo ...[more]

PMID: 35947672

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Project description:BackgroundA key step in embryo implantation is the adhesion to and invasion of the endometrium by the blastocyst trophectoderm. The envelope proteins of HERV-W and -FRD (human endogenous retrovirus-W and -FRD), syncytin-1 and syncytin-2, are mainly distributed in the placenta, and play important roles in the development of the placenta. The placenta originates from the trophectoderm of the blastocyst. It is unclear whether the envelope proteins of HERV-W and -FRD have an effect on the development of the trophectoderm and whether they have any association with the implantation of the blastocyst.MethodsThe whole-genome amplification products of the human blastocyst trophectoderm were used to measure the copy number of syncytin-1 and syncytin-2 using real time qPCR. In addition, clinical data associated with the outcome of pregnancies was collected, and included age, body mass index (BMI), basic follicle stimulating hormone(bFSH), rate of primary infertility and oligo-astheno-teratospermia, the thickness of the endometrium on the day of endometrial transformation, the levels of estrogen and progestin on the transfer day, the days and the morphological scores of the blastocysts. The expression of mRNA and the copy numbers of syncytin-1 and syncytin-2 in H1 stem cells, and in differentiated H1 cells, induced by BMP4, were measured using real time qPCR.ResultsThe relative copy number of syncytin-1 in the pregnant group (median: 424%, quartile: 232%-463%, p < 0.05) was significantly higher than in the non-pregnant group (median: 100%, quartile: 81%-163%). There was a correlation (r s = 0.681, p < 0.001) between the copy number of syncytin-1 and blastocyst implantation after embryo transfer. As the stem cells differentiated, the expression of NANOG mRNA decreased, and the expression of caudal type homeobox 2(CDX2) and β-human chorionic gonadotropin (β-hCG) mRNAs increased. Compared to the undifferentiated cells, the relative expression of the syncytin-1 mRNA was 1.63 (quartile: 0.59-6.37, p > 0.05), 3.36 (quartile: 0.85-14.80, p > 0.05), 10.85 (quartile: 3.39-24.46, p < 0.05) and 67.81 (quartile: 54.07-85.48, p < 0.05) on day 1, 3, 5 and 7, respectively, after the differentiation. The relative expression of syncytin-2 was 5.34 (quartile: 4.50-10.30), 7.90 (quartile: 2.46-14.01), 57.44 (quartile: 38.35-103.87) and 344.76 (quartile: 267.72-440.10) on day 1, 3, 5 and 7, respectively, after the differentiation (p < 0.05). The copy number of syncytin-1 increased significantly during differentiation.ConclusionPreceding the transfer of frozen embryos, the increased copy number of syncytin-1 in the blastocyst trophectoderm was associated with good outcomes of pregnancies.

Project description:In mammalian embryonic development, blastocyst hatching is essential for normal implantation and development of the fetus. We reported previously that blastocysts hatching out of the zona pellucida (ZP) exhibited site preferences that were associated with pregnancy outcomes. To characterize these site differences, we analyzed the transcriptomes in the following developing mouse blastocysts within 16 h of hatching: expanding (E), hatching from the A-site (A), B-site (B), and C-site (C), hatched (H), and non-hatching (N). By principal component analysis and hierarchical cluster analysis, we determined that the gene expression profiles of A and B blastocysts, which resulted in good fertility, clustered closely. C and N blastocysts, which resulted in poor fertility, clustered closely, but distantly from A and B. Embryos hatched at B- vs. C-sites, with good vs. poor pregnancy, showed 178 differentially expressed genes (DEGs), mainly involved in immunity, which correlated positively with birth rate. These DEGs were primarily regulated by transcription factors TCF24 and DLX3. During blastocyst hatching, immune-related genes were regulated, such as Ptgs1, Lyz2, Il-α, Cfb (upregulated) and Cd36 (downregulated). By immunofluorescence staining, we found C3 and IL-1β on the extra-luminal surface of the trophectoderm of the hatched blastocyst, suggesting that they play a role in maternal-fetal interactions. As the blastocysts developed from the expanding to the fully hatched state, 307 DEGs were either upregulated by transcription factor ATOH8 or downregulated by SPIC to switch on immune pathways. Based on the hatching outcome, we identified three transcription patterns in developing blastocysts, with complex changes in the transcriptional regulation network of failed hatched blastocysts vs. successfully hatched blastocysts. We developed a LASSO regression-based model using DEGs Lyz2, Cd36, Cfb, and Cyp17a1 to predict implantation success. This study revealed the diverse, multidimensional developmental fates of blastocysts during short-term hatching and indicated that the immune properties of the embryo had a major effect on blastocyst hatching outcomes. We suggest that transcriptional changes and their regulation during the development of the preimplantation blastocyst affect implantation. This study contributes to our understanding of the role of transcriptional changes in mammalian embryonic development during hatching and their effect on maternal-fetal interactions.

Dataset Information

Primary specification of blastocyst trophectoderm by scRNA-seq: New insights into embryo implantation.

Publications

Primary specification of blastocyst trophectoderm by scRNA-seq: New insights into embryo implantation.

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