Trophoblast Cell Subtypes and Dysfunction in the Placenta of Individuals with Preeclampsia Revealed by Single-Cell RNA Sequencing

Abstract

Trophoblasts, important functional cells in the placenta, play a critical role in maintaining placental function. The heterogeneity of trophoblasts has been reported, but little is known about the trophoblast subtypes and distinctive functions during preeclampsia (PE). In this study, we aimed to gain insight into the cell type-specific transcriptomic changes by performing unbiased single-cell RNA sequencing (scRNA-seq) of placental tissue samples, including those of patients diagnosed with PE and matched healthy controls. A total of 29,006 cells were identified in 11 cell types, including trophoblasts and immune cells, and the functions of the trophoblast subtypes in the PE group and the control group were also analyzed. As an important trophoblast subtype, extravillous trophoblasts (EVTs) were further divided into 4 subgroups, and their functions were preliminarily analyzed. We found that some biological processes related to pregnancy, hormone secretion and immunity changed in the PE group. We also identified and analyzed the regulatory network of transcription factors (TFs) identified in the EVTs, among which 3 modules were decreased in the PE group. Then, through in vitro cell experiments, we found that in one of the modules, CEBPB and GTF2B may be involved in EVT dysfunction in PE. In conclusion, our study showed the different transcriptional profiles and regulatory modules in trophoblasts between placentas in the control and PE groups at the single-cell level; these changes may be involved in the pathological process of PE, providing a new molecular theoretical basis for preeclamptic trophoblast dysfunction.

Identification of Key Genes and Pathways Associated with Preeclampsia by a WGCNA and an Evolutionary Approach

“…For example, EIF4E in the C6 cluster is involved in protein synthesis and a key checkpoint in the control of the rate of mRNA translation [ 55 ], and EIF4E has a critical role in the pathology of various types of cancer [ 56 , 57 ]. Consistent with these findings, it has been recently shown that the genes with upregulated expression in extravillous trophoblast (EVT) were involved in translation by using single-cell RNA sequencing [ 51 ]. In contrast, we observed that PS genes were mainly enriched in the cyan-associated C3 and C5 clusters.…”

“…It was also reported that hypoxia, ischemia, and the production of proinflammatory cytokines associated with PE lead to protein misfolding and initiate endoplasmic reticulum (ER) stress [ 48 , 49 , 50 ]. Furthermore, Zhou et al recently showed that the genes with upregulated expression in syncytiotrophoblast (SCT) were involved in protein folding by using single-cell RNA sequencing [ 51 ]. These observations suggest that cyan and magenta module genes are related to some extent to the functions that regulate protein processing and folding, and thus these genes may contribute to the PE risk during placenta development and maintenance.…”

Single cell RNA sequencing research in maternal fetal interface

“…The 10X genomics Chromium platform builds on the Gem Code technology, which has been frequently used for Single cell transcriptome research in human placenta and decidua ( Vento-Tormo et al, 2018 ; Wang et al, 2021a ; Du et al, 2021 ; Huang et al, 2021 ; Yang et al, 2021 ; Zhou et al, 2022a ). The core of the technology is a Gel bead in Emulsion (GEM) ( Zheng et al, 2017 ).…”

“… Zhang et al (2021) revealed the difference between the preeclampsia and healthy pregnancy groups by analyzing the transcripts of 11,518 cells at the single cell level and found that GO terms enriched in EVT from patients with preeclampsia were mainly associated with immune responses, suggesting altered placental immune function in preeclampsia. Zhou et al (2022b) classified EVT into four subtypes and found that pro-inflammatory, immune, and oxidative stress-related pathways in preeclampsia EVT subtypes were activated. Further analysis revealed that the transcription factors CEBPB, ATF3 , and GTF2B were significantly reduced in expression in preeclamptic EVT, and CEBPB and GTF2B were selected for the next functional study, which revealed that knockdown of these two molecules significantly reduced cell viability and invasiveness, and were hypothesized to be involved in the pathogenesis of Pre-eclampsia ( Zhou et al, 2022b ).…”

“… Zhou et al (2022b) classified EVT into four subtypes and found that pro-inflammatory, immune, and oxidative stress-related pathways in preeclampsia EVT subtypes were activated. Further analysis revealed that the transcription factors CEBPB, ATF3 , and GTF2B were significantly reduced in expression in preeclamptic EVT, and CEBPB and GTF2B were selected for the next functional study, which revealed that knockdown of these two molecules significantly reduced cell viability and invasiveness, and were hypothesized to be involved in the pathogenesis of Pre-eclampsia ( Zhou et al, 2022b ). Overall, scRNA-seq provides a fresh molecular theoretical basis for preeclampsia trophoblast dysfunction, which may help in the diagnosis and treatment of this disease.…”