ANP promotes HTR-8/SVneo cell invasion by upregulating protein kinase N 3 via autophagy inhibition.

Preeclampsia is a gestational disease characterized by two major pathological changes-shallow trophoblast invasion and impaired spiral artery remodeling. Atrial natriuretic peptide (ANP) is a kind of peptide hormone that regulates blood pressure, while the lack of active ANP participates in preeclampsia pathogenesis. However, the underlying mechanism of how ANP modulates trophoblasts function remains unclarified. Here, we performed isobaric tags for relative and absolute quantification (iTRAQ) in ANP-treated HTR-8/SVneo cells and identified Protein Kinase 3 (PKN3) as the downstream factor of ANP, which was downregulated in preeclamptic placenta. Chromatin immunoprecipitation analysis and luciferase assays showed that NFYA was one of the transcription factors for the PKN3 promoter, which was also regulated by ANP treatment in HTR-8/SVneo cells. Transmission electron microscopy and Western Blotting in HTR-8/SVneo cells indicated that ANP inhibited autophagy via AMPK-mTORC1 signaling, while excess autophagy was observed in preeclamptic placenta. The increased expression of PKN3 and enhanced cell invasion ability in HTR-8/SVneo cells induced by ANP could be abolished by autophagy activation or transfection with PKN3 shRNA or NFYA shRNA or NPR-A shRNA via regulating the invasion-related genes and the epithelial mesenchymal transition molecules. Our results demonstrated that ANP could enhance trophoblast invasion by upregulating PKN3 via NFYA promotion through autophagy inhibition in an AMPK/mTORC1 signaling-dependent manner.

CD74 deficiency reduces trophoblast invasion and proliferation mediated by SIRT1 in preeclampsia.

In brief: Preeclampsia (PE) is a severe complication that leads to major maternal and fetal mortality and morbidity, and one of its causes is extravillous trophoblast (EVT) dysfunction. This study revealed the role of CD74 in the invasion and proliferation of EVTs. Abstract: PE is a severe hypertensive disorder during pregnancy, and one of its causes is the dysfunction of EVTs. In this study, we analyzed single-cell RNA sequencing (scRNA-seq) data of placentas from PE patients and the sirtuin 1 (SIRT1) heterozygous knockout mouse model, which exhibited typical PE-like symptoms. We identified 134 differentially expressed genes (DEGs) with similar trends in EVTs of PE patients and in parietal trophoblast giant cells (P-TGCs) of Sirt1-/- (HO) placentas from Sirt1+/- (HE) pregnant mice. Interestingly, Kyoto Encyclopedia of Genes and Genomes analysis showed that 134 overlapping genes were related to the MAPK signaling pathway. We validated several DEGs using immunofluorescence at the protein level. Finally, we selected CD74 for further experiments, which showed a decrease in EVTs of PE patients and in P-TGCs of Sirt1-/- placentas from Sirt1+/- pregnant mice. Additionally, cell proliferation assays and transwell assays showed that the proliferation and invasion abilities were decreased in CD74 knockdown HTR8/SVneo cells using lentivirus transfection, which can be improved by adding the SIRT1 agonist SRT1720 or metformin, an agonist of the MAPK signaling pathway. Importantly, the expression of CD74 can be positively regulated by SIRT1. These data suggest that CD74 plays an important protective role in the pathogenesis of preeclampsia by regulating the MAPK signaling pathway, which can be regulated by SIRT1.

Crosstalk between Placental Trophoblast and Decidual Immune Cells in Recurrent Miscarriage.

Recurrent miscarriage (RM) is a pregnancy complication associated with dysregulation of the maternal-fetal interface. We aimed to identify dysfunctional interactions between trophoblast cells and decidual immune cells in RM. We downloaded single-cell RNA sequencing (scRNA-seq) datasets (GSE214607) from the Gene Expression Omnibus (GEO) datasets for further analysis using the R software. The data comprised of paired placental and decidual tissues, including those from patients diagnosed with RM and matched healthy controls. A total of 22976 cells were identified in 11 cell types, including trophoblasts, immune cells, and other cells. We divided trophoblast cells into three types and analyzed their interactions with decidual immune cells. Additionally, we re-clustered NK&T cells and macrophages, identified differentially expressed genes (DEGs), enriched their functions, and compared the cell interactions with trophoblast cells in each cell type. Our single-cell atlas of the maternal-fetal interface revealed alterations in the cellular organization of the decidua and placenta, cell type-specific transcriptome, and cell communication between immune and non-immune cells in RM, which are critical for illuminating the pathophysiology of RM.

APOA1 Is a Novel Marker for Preeclampsia.

Preeclampsia (PE) is one of the pregnancy complications, leading to major maternal and fetal morbidity and mortality; however, the underlying mechanisms of PE still remain unclear. We aimed to explore the role of apolipoprotein A1 (APOA1) in the pathophysiology of PE. The expression of APOA1 was elevated in both plasma and placental tissues, as detected by Western blotting, immunohistochemistry, and a qRT-PCR assay. Importantly, we detected the concentration of APOA1 using the ELISA assay in normal control women (n = 30) and women with preeclampsia (n = 29) from a prospective cohort study. The concentration of APOA1 was not significantly altered in plasma during early and mid-term gestation of the PE patients compared to the NP patients; however, it was elevated during late gestation. Additionally, the concentration of APOA1 was positively associated with systolic blood pressure during late gestation. The proliferation and invasion of trophoblast were all increased in HTR8/SVneo cells transfected with APOA1 siRNA and decreased in HTR8/SVneo cells treated with the recombinant human APOA1 protein (rhAPOA1). Additionally, we used public datasets to investigate the downstream genes of APOA1 and qRT-PCR for validation. Furthermore, we explored the transcriptional activity of peroxisome proliferator-activated receptor gamma (PPARγ) in APOA1 by using a luciferase assay, which showed that the APOA1 promoter was activated by PPARγ. Additionally, the inhibitory effect of rhAPOA1 on the ability of trophoblast invasion and proliferation can be rescued by the PPARγ inhibitor. Our findings suggest the crucial role of APOA1 in PE, which might provide a new strategy for the prevention and treatment of PE.

Oxytocin use in trial of labor after cesarean and its relationship with risk of uterine rupture in women with one previous cesarean section: a meta-analysis of observational studies.

BACKGROUND: Trial of labor after a previous cesarean delivery (TOLAC) has reduced the rate of cesarean sections (CS). Nevertheless, the widespread use of TOLAC has been limited by an increase in adverse outcomes, the most serious one being the risk of symptomatic uterine rupture, which is possibly associated with oxytocin. In this meta-analysis, we explored the risk association between oxytocin use and uterine rupture in TOLAC. METHODS: Multiple electronic databases (PubMed, Embase, Web of Science, and Google Scholar) were searched for cross-sectional studies reporting on TOLAC, oxytocin and uterine rupture, which were published between January 1986 and October 2019. The bias-corrected Hedge's g was calculated as the effect size using the random-effects model. A two-sample Z test was used to compare the differences in synthetic rates between groups. The Newcastle-Ottawa Scale (NOS) was used to evaluate the risk of bias. Quality of the evidence was assessed with the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) certainty ratings system. RESULTS: A total of 14 studies, which included 48,457 women who underwent TOLAC, met the inclusion criteria. The pooled rate of vaginal birth after a cesarean section (VBAC) and the rate of uterine rupture in spontaneous labor were 74.3 and 0.7%, respectively. In addition, the pooled rate of VBAC and the rate of uterine rupture in the induction labor group was 60.7 and 2.2%, respectively. The women who had spontaneous labor had a significantly higher rate of VBAC (p = 0.001) and a lower rate of uterine rupture (p = 0.0003) compared to induced labor. The pooled rates of uterine rupture in women using oxytocin and women not using oxytocin in TOLAC were 1.4% and 0.5%, respectively, and the difference was significant (p = 0.0002). Also, the synthetic rate of uterine rupture in oxytocin augmentation among women with spontaneous labor and women who had a successful induction of labor were 1.7% and 2.2%, respectively, without significant difference (p = 0.443). CONCLUSIONS: Women with induced labor had a higher risk of uterine rupture compared to women with spontaneous labor following TOLAC. Oxytocin use may increase this risk, which could be influenced by the process of induction or individual cervix condition. Consequently, simplified and standardized intrapartum management, precise protocol, and cautious monitoring of oxytocin use in TOLAC are necessary.

Decreased expression of JHDMID in placenta is associated with preeclampsia through HLA-G.

The exact mechanism of preeclampsia (PE) remains unclear, accumulating researches have indicated multiple epigenetic factors relate to PE and histone methylation plays a crucial role in modifying the gene expression. So we aimed to confirm that abnormal expression of histone demethylase JHDM1D contributes to PE and lower expression of HLA-G in PE. We tested the expression of JHDM1D, H3K9me2, and H3K27me2 in the placentas of PE and normal control (NC)women who had a healthy pregnancy with Immunohistochemistry and we found that JHDM1D, H3K9me2, and H3K27me2 were all mainly expressed in the nuclei of the extra-villous trophoblasts (EVTs). JHDM1D was lower expressed in PE than in NC placentas, corresponding with the mRNA level and protein level with qTR-PCR and western blot, while H3K9me2 and H3K27me2 were higher expressed in PE. We further investigated the biological functions of JHDM1D in HTR-8/SVneo cells. We found that siJHDM1D inhibited cell growth after 24 h of the transfection and reduced the invasion, while increasing the apoptosis of HTR-8/SVneo. We then constructed the siJHDM1D stable cell line and confirmed with CHIP-qPCR that siJHDM1D inhibited the expression of HLA-G through increased the enrichment of H3K9me2 and H3K27me2 in the JHDM1D bounding region of HLA-G. Taken together, our study confirms that decreased expression of JHDM1D is associated with PE through down-regulating HLA-G and casts new light to the diagnosis and therapy of PE.

Identification of key microRNAs and genes in preeclampsia by bioinformatics analysis.

Preeclampsia is a leading cause of perinatal maternal-foetal mortality and morbidity. The aim of this study is to identify the key microRNAs and genes in preeclampsia and uncover their potential functions. We downloaded the miRNA expression profile of GSE84260 and the gene expression profile of GSE73374 from the Gene Expression Omnibus database. Differentially expressed miRNAs and genes were identified and compared to miRNA-target information from MiRWalk 2.0, and a total of 65 differentially expressed miRNAs (DEMIs), including 32 up-regulated miRNAs and 33 down-regulated miRNAs, and 91 differentially expressed genes (DEGs), including 83 up-regulated genes and 8 down-regulated genes, were identified. The pathway enrichment analyses of the DEMIs showed that the up-regulated DEMIs were enriched in the Hippo signalling pathway and MAPK signalling pathway, and the down-regulated DEMIs were enriched in HTLV-I infection and miRNAs in cancers. The gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) enrichment analyses of the DEGs were performed using Multifaceted Analysis Tool for Human Transcriptome. The up-regulated DEGs were enriched in biological processes (BPs), including the response to cAMP, response to hydrogen peroxide and cell-cell adhesion mediated by integrin; no enrichment of down-regulated DEGs was identified. KEGG analysis showed that the up-regulated DEGs were enriched in the Hippo signalling pathway and pathways in cancer. A PPI network of the DEGs was constructed by using Cytoscape software, and FOS, STAT1, MMP14, ITGB1, VCAN, DUSP1, LDHA, MCL1, MET, and ZFP36 were identified as the hub genes. The current study illustrates a characteristic microRNA profile and gene profile in preeclampsia, which may contribute to the interpretation of the progression of preeclampsia and provide novel biomarkers and therapeutic targets for preeclampsia.