Single-cell insights into development of the bovine placenta

Abstract

A central determinant of pregnancy success is proper development of the conceptus (embryo/fetus and associated extraembryonic membranes including the placenta). Although the gross morphology and histology of the bovine placenta has been well studied, the cellular and molecular mechanisms regulating placenta development and trophoblast differentiation and function remain essentially undefined. Here single cell transcriptome (scRNA-seq) analysis was performed on the day 17 bovine conceptus and chorion of day 24, 30 and 50 conceptuses (n = 3–4 samples per day) using the 10X Genomics platform. Bioinformatic analyses identified cell types and their ontogeny including trophoblast, mesenchyme, and immune cells. Loss of IFNT expressing trophoblast uninucleate cells (UNC) occurred between days 17 and 30, whereas binucleate cells (BNC), identified based on expression of placental lactogen (CSH2) and specific pregnancy-associated glycoproteins (PAGs) genes, first appeared on day 24. Several different types of UNC were present in day 24, 30 and 50 samples, but only one (day 24) or two types of BNC (days 30 and 50). Cell trajectory analyses provided a conceptual framework for UNC development and BNC differentiation, and bioinformatic analyses identified candidate transcription factors governing differentiation and function of the trophoblasts. The digital atlas of cell types in the developing bovine conceptus reported here serves as a resource to discover key genes and biological pathways regulating its development during the critical periods of implantation and placentation.

Single-cell transcriptional landscapes of bovine peri-implantation development

“…By integrating peri-implantation trophoblast cell lineages with the recently two published single-cell transcriptomes of trophoblasts from day 17 through day 195 of pregnancy in cattle, 14 , 39 we constructed a comprehensive transcriptomic road map of trophoblast lineage development and differentiation. Clustering analysis classified trophoblast lineages into three major groups based on the development stage (cluster 1: day 12–14; cluster 2: day 16–18; cluster 3: day 24–195 ( Figure 4 A).…”

“…In our dataset, we identified six different sub-lineages of trophoblast cells, classified into two categories as spheroid TB and elongated TB. By integrated analysis early trophoblast lineages with the published more advanced trophoblasts after implantation and placentation, 14 , 39 we further constructed a comprehensive transcriptomic road map of trophoblast lineage development and differentiation. While BNC cells are not present in the peri-implantation embryos, these newly identified elongated TB cells have up-regulated machinery for the BNC cells and represent an important stage of trophoblast cell fate that is responsible for pregnancy maintenance in bovine prior to the emergence of binucleate cells.…”

“…G1916 Fertagyl (Gonadorelin) Merck Animal Health NDC 57926-477 Folltropin (Follicle-Stimulating Hormone) Vetoquinol NDC 17030-102-70 Lutalyse (Dinoprost Tromethamine Injection) Zoetis NDC 54771-1327 Critical commercial assays Chromium Next GEM Single Cel 3′ Reagent Kit v3.1 Dual Index 10x Genomics Cat# 1000268 ViewRNA-ISH cell assay kit Thermo Fisher Cat# QVC0001 ViewRNA-ISH cell 740 Module Thermo Fisher Cat#QVC0200 Deposited data Bovine scRNA-seq data from this paper This paper GSE234335 Bovine scRNA-seq data from day 17–50 Davenport et al. 14 , 39 GSE234524 Bovine sc RNA-seq data from day 195 Davenport et al. 14 , 39 GSE214407 Mouse scRNA-seq data from 4.5 dpf embryos Nowotschin et al.…”

“… 14 , 39 GSE234524 Bovine sc RNA-seq data from day 195 Davenport et al. 14 , 39 GSE214407 Mouse scRNA-seq data from 4.5 dpf embryos Nowotschin et al. 40 GSE123046 Sheep scRNA-seq data from 12, 16, 19 dpf embryos Jia et al.…”

Single-Cell Sequencing Technology in Ruminant Livestock: Challenges and Opportunities

“…Furthermore, the application of single-cell genomics in livestock research has the potential to accelerate the pace of genetic improvement by facilitating the identification of cell-type-specific gene expression, along with gene regulatory elements and networks associated with various traits important to livestock production . Adipose Nuclei RNA GEO GSE211707 [7] Embryo (8-and 16-cell stage) Cell RNA GEO GSE99210 [8] Embryo (peri-implantation) Cell RNA GEO GSE234335 [9] Embryo (SCNT) Cell RNA SRA PRJNA727165 [10] Embryo (trophectoderm) Cell RNA GEO GSE200216 [11] Fetal Gonads Cell RNA GEO GSE162952 [12] Milk Somatic Cells Cell RNA ENA PRJEB73560 [13] Muscle Cell RNA GEO GSE205347 [14] Muscle Cell RNA, ATAC GSA CRA006626 [15] PBMC * Cell RNA, ATAC GEO GSE225962 [16] PBMC * Cell RNA, ATAC GEO GSE166473 [17] Placenta (developing) Cell RNA GEO GSE234524 [18] Placenta (mature) Nuclei RNA GEO GSE214407 [19] Primary Mammary Epithelial Cells Cell RNA FAANG PRJEB41576 [20] Ruminal epithelial cells Cell RNA GEO GSE166473 [21] Satellite cells Cell RNA GEO GSE184128 [22] Satellite cells Cell RNA GEO GSE211428 [23] Despite the potential benefits, the application of sc/snRNA-seq and sc/snATAC-seq in ruminant livestock species presents challenges. Standardized computational analysis pipelines that are species-agnostic are essential, and the discovery and annotation of cell populations can be difficult in non-model organisms and for tissues such as the rumen, which are unique to these species [44][45][46].…”