Pathogenesis Of Preeclampsia Research Articles

A hidden proteome encoded by circRNAs in human placentas: Implications for uncovering preeclampsia pathogenesis.

CircRNA-encoded proteins (CEPs) are emerging as new players in health and disease, and function as baits for the common partners of their cognate linear-spliced RNA encoded proteins (LEPs). However, their prevalence across human tissues and biological roles remain largely unexplored. The placenta is an ideal model for identifying CEPs due to its considerable protein diversity that is required to sustain fetal development during pregnancy. The aim of this study was to evaluate circRNA translation in the human placenta, and the potential roles of the CEPs in placental development and dysfunction. Multiomics approaches, including RNA sequencing, ribosome profiling, and LC-MS/MS analysis, were utilised to identify novel translational events of circRNAs in human placentas. Bioinformatics methods and the protein bait hypothesis were employed to evaluate the roles of these newly discovered CEPs in placentation and associated disorders. The pathogenic role of a recently identified CEP circPRKCB119aa in preeclampsia was investigated through qRT-PCR, Western blotting, immunofluorescence imaging and phenotypic analyses. We found that 528 placental circRNAs bound to ribosomes with active translational elongation, and 139 were translated to proteins. The CEPs showed considerable structural homology with their cognate LEPs, but are more stable, hydrophobic and have a lower molecular-weight than the latter, all of which are conducive to their function as baits. On this basis, CEPs are deduced to be closely involved in placental function. Furthermore, we focused on a novel CEP circPRKCB119aa, and illuminated its pathogenic role in preeclampsia; it enhanced trophoblast autophagy by acting as a bait to inhibit phosphorylation of the cognate linear isoform PKCβ. We discovered a hidden circRNA-encoded proteome in the human placenta, which offers new insights into the mechanisms underlying placental development, as well as placental disorders such as preeclampsia. Key points A hidden circRNA-encoded proteome in the human placenta was extensively identified and systematically characterised. The circRNA-encoded proteins (CEPs) are potentially related to placental development and associated disorders. A novel conserved CEP circPRKCB119aa enhanced trophoblast autophagy by inhibiting phosphorylation of its cognate linear-spliced isoform protein kinase C (PKC) β in preeclampsia.

Comparative Immune Profiles: Platelet-to-Lymphocyte and Neutrophil-to-Lymphocyte Ratios in Preeclampsia versus Healthy Women

Background & Objective: Pre-eclampsia is a hypertensive disorder linked with immune dysregulation and systemic inflammation in pregnancy. An elevated neutrophil count alongside a relatively stable lymphocyte count leads to an increased Neutrophil-to-Lymphocytic Ratio (NLR), serving as a parameter of widespread systemic inflammation. Platelet to lymphocyte ratio (PLR) is a also used as a hematological marker that reflects the balance between platelets and lymphocytes. A higher NLR and PLR may reflect increased endothelial activation and damage, contributing to the pathogenesis of preeclampsia. The objective of the present study was to compare the Neutrophil to lymphocytic ratio (NLR as well as platelet-to-lymphocytic ratio (PLR) between women affected by preeclampsia (PE) and those with uncomplicated pregnancies. Methods: It was a cross-sectional comparative study. Fifty-five registered preeclamptic patients were enrolled as cases along with fifty-five healthy pregnant women. Total leukocyte counts, including neutrophils and lymphocytes as well as platelet count, were calculated using an automated hematology analyzer, and the Neutrophil-to-Lymphocyte Ratio (NLR) as well as Platelet to Lymphocyte ratio were subsequently calculated. Results: Higher value of TLC and neutrophils were recorded in pre-eclampsia. The normotensive women had higher platelet as well as lymphocyte count. NLR was significantly high in the preeclampsia contrary to PLR ratio that showed a higher value in normotensive pregnancies. Conclusions: In conclusion, our study highlights significant alterations in the immune-inflammatory profile of patients with preeclampsia compared to controls. The observed high neutrophil-to-lymphocyte ratio (NLR) and low platelet-to-lymphocyte ratio (PLR) suggest a distinct systemic inflammatory response associated with preeclampsia.

Detection and assessment of immune and stromal related risk genes to predict preeclampsia: A bioinformatics analysis with dataset.

This study aimed to investigate immune score and stromal score-related signatures associated with preeclampsia (PE) and identify key genes for diagnosing PE using bioinformatics analysis. Four microarray datasets, GSE75010, GSE25906, GSE44711, and GSE10588 were obtained from the Gene Expression Omnibus database. GSE75010 was utilized for differential expressed gene (DEGs) analysis. Subsequently, bioinformatic tools such as gene ontology, Kyoto Encyclopedia of Genes and Genomes, weighted gene correlation network analysis, and gene set enrichment analysis were employed to functionally characterize candidate target genes involved in the pathogenesis of PE. The least absolute shrinkage and selection operator regression approach was employed to identify crucial genes and develop a predictive model. This method also facilitated the creation of receiver operating characteristic (ROC) curves, enabling the evaluation of the model's precision. Furthermore, the model underwent external validation through the other three datasets. A total of 3286 DEGs were identified between normal and PE tissues. Gene ontology and Kyoto Encyclopedia of Genes and Genomes analyses revealed enrichments in functions related to cell chemotaxis, cytokine binding, and cytokine-cytokine receptor interaction. weighted gene correlation network analysis identified 2 color modules strongly correlated with immune and stromal scores. After intersecting DEGs with immune and stromal-related genes, 13 genes were selected and added to the least absolute shrinkage and selection operator regression. Ultimately, 7 genes were screened out to establish the risk model for discriminating preeclampsia from controls, with each gene having an area under the ROC curve >0.70. The constructed risk model demonstrated that the area under the ROC curves in internal and the other three external datasets were all greater than 0.80. A 7-gene risk signature was identified to build a potential diagnostic model and performed well in the external validation group for PE patients. These findings illustrated that immune and stromal cells played essential roles in PE during its progression.

The downregulation of NSUN5 may contribute to preeclampsia†.

Preeclampsia (PE) is a complication of pregnancy characterized by the new onset of hypertension after 20weeks of gestation. The incidence of PE is steadily rising, posing a significant threat to the lives of both the pregnant woman and the fetus. Most studies on PE pathogenesis currently focus on the placenta, but maternal decidualization forms the foundation for placental growth and development. Recent studies have shown that impaired decidualization is also a cause of PE. Decidualization is a process where endometrial stromal cells gradually transform into secretory decidual cells during early pregnancy. While NSUN5 encodes a member of a conserved family of proteins, its role in pregnancy remains unknown. In this study, we conducted experiments and observed a significant downregulation of NSUN5 expression in severe PE decidual tissues compared to those of normal pregnant women. When inducing decidualization in vitro, we found an increase in NSUN5 expression. However, when we used siRNA to knockdown NSUN5 expression, the process of decidualization was prevented. Moreover, we observed a decrease in ATP content during both cell decidualization and after knockdown of NSUN5. Finally, through immunoprecipitation combined with mass spectrometry, we discovered that the protein ATP5B interacts with NSUN5. Furthermore, after knocking down ATP5B using siRNA, we observed impaired decidualization. Moreover, transfection with siRNA to suppress NSUN5 resulted in a decrease in ATP5B expression. These significant findings provide strong evidence that NSUN5 plays a crucial role in decidualization and is closely associated with the development of PE through its interaction with ATP5B.

Decreased Extracellular Vesicle Vasorin in Severe Preeclampsia Plasma Mediates Endothelial Dysfunction.

Preeclampsia (PE) is a serious pregnancy complication affecting 5-8% of pregnancies globally. It is a leading cause of maternal and neonatal morbidity and mortality. Despite its prevalence, the underlying mechanisms of PE remain unclear. This study aimed to determine the potential role of vasorin (VASN) in PE pathogenesis by investigating its levels in extracellular vesicles (EV) and its effects on vascular function. We conducted unbiased proteomics on urine-derived EV from severe PE (sPE) and normotensive pregnant women (NTP), identifying differential protein abundances. Out of one hundred and twenty proteins with ≥ ±1.5-fold regulation at P<0.05 between sPE and NTP, we focused on Vasorin (VASN), which is downregulated in sPE in urinary EV, in plasma EV and in the placenta and is a known regulator of vascular function. We generated EV with high VASN content from both human and murine placenta explants (Plex EV), which recapitulated disease-state-dependent effects on vascular function observed when treating murine aorta rings (MAR) or human aortic endothelial cells (HAEC) with murine or human plasma-derived EV. In normal murine pregnancy, VASN increases with gestational age (GA), and VASN is decreased in plasma EV, in placenta tissue and in Plex EV after intravenous administration of adenovirus encoding short FMS-like tyrosine kinase 1 (sFLT-1), a murine model of PE (murine-PE). VASN is decreased in plasma EV, in placenta tissue and in EV isolated from conditioned media collected from placenta explants (Plex EV) in patients with sPE as compared to NTP. Human sPE and murine-PE plasma EV and Plex EV impair migration, tube formation, and induces apoptosis in human aortic endothelial cells (HAEC) and inhibit acetylcholine-induced vasorelaxation in murine vascular rings (MAR). VASN over-expression counteracts the effects of sPE EV treatment in HAEC and MAR. RNA sequencing revealed that over-expression or knock down of VASN in HAEC results in contrasting effects on transcript levels of hundreds of genes associated with vasculogenesis, endothelial cell proliferation, migration and apoptosis. The data suggest that VASN, delivered to the endothelium via EV, regulates vascular function and that the loss of EV VASN may be one of the mechanistic drivers of PE. What is NewVASN in circulating plasma EV in sPE is reduced compared with VASN content in plasma EV of gestational age-matched pregnant women.VASN is encapsulated and transported in EV and plays a pro-angiogenic role during pregnancy.VASN should be explored both for its pro-angiogenic mechanistic role and as a novel biomarker and potential predictive diagnostic marker for the onset and severity of PE.What Are the Clinical Implications?VASN plays a role in maintaining vascular health and the normal adaptive cardiovascular response in pregnancy. A decrease of VASN is observed in sPE patients contributing to cardiovascular maladaptation.Strategies to boost diminished VASN levels and/or to pharmacologically manipulate mechanisms downstream of VASN may be explored for potential therapeutic benefit in PE.The decrease in EV-associated VASN could potentially be used as a (predictive) biomarker for PE.

Placental neutrophil reverse trans-migration and maternal serum neutrophil extracellular trap expression in HIV infection co-morbid pre-eclampsia in women of African ancestry

Neutrophil extracellular traps (NETs) and placental neutrophil reverse transmigration (r-TM) are implicated in the pathogenesis of pre-eclampsia (PE). However, the role of the comorbidity of PE and human immunodeficiency virus (HIV) infection in placental neutrophil r-TM and serum NETs remains unknown. Human placental tissue (n = 160) and serum (n = 80) samples were obtained post-ethical approval and divided by pregnancy type and HIV status and across the study population. Immunohistochemistry and morphometry were performed to localize and quantify junctional adhesion molecule-C (JAM-C) expression as an inverse marker of neutrophil r-TM within placental villi. An enzyme-linked immunosorbent assay (ELISA) was performed to quantify the concentration of citrullinated histone H3 (cit-H3) as a marker of NETs. GraphPad Prism (version 8.0.2) was used to compare the results, and a p value of p < 0.05 was considered statistically significant. The localization of JAM-C was observed on the syncytiotrophoblasts (STBs) and endothelial cells of placental villi. The immunoexpression of JAM-C was elevated in PE vs. normotensive (N) placentae. In the exchange villi, JAM-C immunoexpression was higher in the N+ve vs. N-ve group. However, in PE comorbid HIV infection, JAM-C expression was lower in the PE+ve vs. PE-ve group. Citrullinated histone-H3 concentration was lower in the N+ve vs. N-ve group but elevated in early-onset PE (EOPE)+ve vs. late-onset PE (LOPE)+ve group. These results indicate that PE and HIV-infected placentae individually express elevated JAM-C, manifesting in less neutrophil r-TM. However, in exchange villi of PE comorbid with HIV infection reduced JAM-C enhances neutrophil r-TM, thus supporting the synergistic effect of PE comorbid with HIV.

CircHIPK3/miR-124 affects angiogenesis in early-onset preeclampsia via CPT1A-mediated fatty acid oxidation.

Multiple theories have been proposed to explain the pathogenesis of early-onset preeclampsia (EOPE), and angiogenic dysfunction is an important part of this pathogenesis. Carnitine palmitoyltransferase (CPT1A) is a key rate-limiting enzyme in the metabolic process of fatty acid oxidation (FAO). FAO regulates endothelial cell (EC) proliferation during vascular germination and is also essential for ab initio deoxyribonucleotide synthesis, but its role in EOPE needs to be further elucidated. In the present study, we investigated its functional role in EOPE by targeting the circHIPK3/miR-124-3p/CPT1A axis. In our study, reduced expression of circHIPK3 and CPT1A and increased expression of miR-124-3p in placental tissues from patients with EOPE were associated with EC dysfunction. Here, we confirmed that CPT1A regulates fatty acid oxidative activity, cell proliferation, and tube formation in ECs by regulating FAO. Functionally, knockdown of circHIPK3 suppressed EC angiogenesis by inhibiting CPT1A-mediated fatty acid oxidative activity, which was ameliorated by CPT1A overexpression. In addition, circHIPK3 regulates CPT1A expression by sponging miR-124-3p. Hence, circHIPK3 knockdown reduced fatty acid oxidation in ECs by sponging miR-124-3p in a CPT1A-dependent manner and inhibited EC proliferation and tube formation, which may have led to aberrant angiogenesis in EOPE. Thus, strategies targeting CPT1A-driven FAO may be promising approaches for the treatment of EOPE. KEY MESSAGES: Decreased Carnitine palmitoyltransferase (CPT1A) expression in preeclampsia(PE). CPT1A overexpression promotes FAO activity and tube formation in ECs. CircHIPK3 can affect CPT1A expression and impaire angiogenesis of EOPE. CircHIPK3 regulates CPT1A expression by acting as a ceRNA of miR-124-3p in HUVECs. Confirming the effect of circHIPK3/miR-124-3p/CPT1A axis on EOPE.

Circ_0007611 modulates the miR-34c-5p/LPAR2 cascade to suppress proliferation and enhance apoptosis of HTR-8/SVneo cells

IntroductionThe aberrant biological behaviors of trophoblast cells actively take part in the pathogenesis of preeclampsia (PE). Herein, we defined the action of the circular RNA (circRNA) circ_0007611 on trophoblast cell apoptosis and growth to understand its role in PE. MethodsExpression of circ_0007611, miR-34c-5p and lysophosphatidic acid receptor 2 (LPAR2) mRNA was analyzed by qPCR. LPAR2 protein was determined by western blotting. Cell proliferation was analyzed by EdU assay. We assessed apoptosis through flow cytometry and analysis of caspase3 activity and apoptosis-related marker proteins. The binding of miR-34c-5p and circ_0007611 or LPAR2 was verified by dual-luciferase and pull-down assays. ResultsCirc_0007611 and LPAR2 levels were augmented, while miR-34c-5p was diminished in blood samples of PE. Circ_0007611 deficiency repressed cell apoptosis and enhanced the growth of HTR-8/SVneo cells. Circ_0007611 interacted with miR-34c-5p, and miR-34c-5p depletion reversed circ_0007611 deficiency-induced HTR-8/SVneo cell apoptotic inhibition and growth enhancement. MiR-34c-5p targeted LPAR2, and circ_0007611 affected LPAR2 expression via miR-34c-5p competition. Circ_0007611 deficiency-induced HHTR-8/SVneo cell apoptotic inhibition and growth enhancement were also counteracted by LPAR2 overexpression. DiscussionCirc_0007611 modulates the miR-34c-5p/LPAR2 cascade to enhance apoptosis and inhibit proliferation in HTR-8/SVneo cells, thereby contributing to the progression of PE.

Lipidomic signatures in patients with early-onset and late-onset Preeclampsia

BackgroundPreeclampsia is a pregnancy-specific clinical syndrome and can be subdivided into early-onset preeclampsia (EOPE) and late-onset preeclampsia (LOPE) according to the gestational age of delivery. Patients with preeclampsia have aberrant lipid metabolism. This study aims to compare serum lipid profiles of normal pregnant women with EOPE or LOPE and screening potential biomarkers to diagnose EOPE or LOPE.MethodsTwenty normal pregnant controls (NC), 19 EOPE, and 19 LOPE were recruited in this study. Untargeted lipidomics based on ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to compare their serum lipid profiles.ResultsThe lipid metabolism profiles significantly differ among the NC, EOPE, and LOPE. Compared to the NC, there were 256 and 275 distinct lipids in the EOPE and LOPE, respectively. Furthermore, there were 42 different lipids between the LOPE and EOPE, of which eight were significantly associated with fetal birth weight and maternal urine protein. The five lipids that both differed in the EOPE and LOPE were DGTS (16:3/16:3), LPC (20:3), LPC (22:6), LPE (22:6), PC (18:5e/4:0), and a combination of them were a potential biomarker for predicting EOPE or LOPE. The receiver operating characteristic analysis revealed that the diagnostic power of the combination for distinguishing the EOPE from the NC and for distinguishing the LOPE from the NC can reach 1.000 and 0.992, respectively. The association between the lipid modules and clinical characteristics of EOPE and LOPE was investigated by the weighted gene co-expression network analysis (WGCNA). The results demonstrated that the main different metabolism pathway between the EOPE and LOPE was enriched in glycerophospholipid metabolism.ConclusionsLipid metabolism disorders may be a potential mechanism of the pathogenesis of preeclampsia. Lipid metabolites have the potential to serve as biomarkers in patients with EOPE or LOPE. Furthermore, lipid metabolites correlate with clinical severity indicators for patients with EOPE and LOPE, including fetal birth weight and maternal urine protein levels.

Association of catechol-o-methyltransferase gene polymorphism with preeclampsia and biomarkers of oxidative stress: Study protocol for a prospective case-control study in Pakistan.

Preeclampsia is one of the three leading causes of worldwide maternal mortality. Oxidative stress-mediated endothelial damage is expected to be an ultimate common mechanism in the pathophysiology of preeclampsia. The role of bioamines is also well-established in the induction of preeclampsia. This project is aimed to understand the factors which may affect the induction, progression, and aggravation of preeclampsia and oxidative stress during pregnancy. This study will explore the methylation pattern of the Catechol-O-methyltransferase gene to determine its role in the pathogenesis of preeclampsia, association of Val158Met polymorphism with a wide range of oxidative stress biomarkers, major antioxidants vitamins, and blood pressure regulating amines in preeclamptic Pakistani women. In this prospective case-control study, 85 preeclamptic and 85 normotensive pregnant women will be recruited in their third trimesters. DNA will be extracted from peripheral blood and Val158Met polymorphism in the Catechol-O-methyltransferase gene will be examined on PCR amplified product digested with Hin1II (NlaIII) restriction enzyme, further validated by Sanger sequencing. Methylation-sensitive PCR will also be performed. Oxidative stress biomarkers, antioxidant vitamins, bioamines, and catechol-O-methyltransferase levels will be measured by ELISA. The data will be used to correlate maternal and fetal outcomes in both groups. This study will help to identify and understand the multifactorial path and cause-effect relationship of gene polymorphism, oxidative stress biomarkers, major antioxidants vitamins, and blood pressure regulating amines in the pathogenesis and aggravation of preeclampsia in the Pakistani population. The outcome of this project will be particularly helpful in reducing the incidence of preeclampsia and further improving its management.

CircPAPPA2 plays a role in preeclampsia pathogenesis via regulation of the miR-942/miR-5006-3p

CircRNAs are a class of endogenous non-coding RNAs implicated in the pathogenesis of many pregnancy related diseases, one of which is pre-eclampsia (PE). This study aims to investigate the role of CircPAPPA2 (circbase ID: hsa_circ_0015382) in regulating the migration and invasion of trophoblast cells. RNA sequencing was used to identify the differentially expressed circRNAs in placenta of PE and normal pregnant women. Quantitative polymerase chain reaction (qRT-PCR) was used to verify the expression of circPAPPA2 and two miRNAs (miR-942-5p, 5006-3p) in placenta of PE and normal pregnant women. CCK8 and transwell experiments were performed to assess the function of circPAPPA2 in PE development.The interaction between circPAPPA2 and miR-942-5p/miR-5006-3p was verified by dual-luciferase reporter assay. Finally, bioinformatics analyzed with gene ontology, Kyoto Encyclopedia of the target genes. The results showed that the expression of circPAPPA2 was increased in placenta of PE pregnant women. Also, circPAPPA2 impedes trophoblasts cell proliferation and invasion. Moreover, the expression of circPAPPA2 was positively correlated with systolic blood pressure and urine protein. In addition, circPAPPA2 serves as a sponge of miR-942-5p and miR-5006-3p. In conclusion, CircPAPPA2 regulates trophoblasts cell proliferation and invasion by mediating the miR-942/miR-5006-3p.

КЛИНИКО-ПАТОФИЗИОЛОГИЧЕСКИЕ АСПЕКТЫ МИКРОВЕЗИКУЛЯРНОГО СОСТАВА ПЕРИФЕРИЧЕСКОЙ КРОВИ У БЕРЕМЕННЫХ С ПРЕЭКЛАМПСИЕЙ

Abstract. Introduction. Preeclampsia is a pregnancy complication characterized by the development of arterial hypertension after 20 weeks of gestation, proteinuria, and/or oedemas. The pathogenesis of preeclampsia remains unclear to this day. Some studies have shown the importance of microvesicles in the pathogenesis of various pathological conditions. Involvement of microvesicles in the preeclampsia pathogenesis is a relevant issue. Taking into account all of the above, we have made an attempt to consider the pathogenesis of preeclampsia from the perspective of clinical and laboratory data. Aim. To establish the importance of microvesicles circulating in peripheral blood, their quantity and phenotype in the pathogenesis of preeclampsia. Materials and Methods. A one-stage cohort study was conducted, including 105 pregnant women: 30 pregnant women with severe preeclampsia (group 1), 35 pregnant women with mild preeclampsia (group 2), and 40 pregnant women without hypertensive syndrome (group 3), at 29–40 weeks of gestation. All patients were delivered surgically (Joel-Cohen laparotomy, cesarean section in the lower segment). All pregnant women underwent blood sampling to define the number and phenotypes of microvesicles (using flow cytometry) and laser scanning microscopy of the clot to confirm the presence of microvesicles of placental origin containing tissue factor, as well as the lipopolysaccharide of gram-negative microorganisms. Results and Discussion. In the group of patients with severe preeclampsia, a statistically significant increase was found in the number of placental, erythrocyte microvesicles, microvesicles with the tissue factor and lipopolysaccharide, compared to the group of patients with mild preeclampsia and pregnant women without hypertensive syndrome. In the group of patients with mild preeclampsia, a statistically significant increase was found in microvesicles with tissue factor and lipopolysaccharide, compared to pregnant women without hypertensive syndrome. The data obtained from the study confirm the fact that severe preeclampsia is a separate nosological entity, most likely pregnancy-associated thrombotic microangiopathy. Conclusions. The data obtained from the study indicate the need for further research to clarify the importance of microvesicles in the preeclampsia pathogenesis.

Alleviation of preeclampsia-like symptoms through PlGF and eNOS regulation by hypoxia- and NF-κB-responsive miR-214-3p deletion

Preeclampsia is caused by placental hypoxia and systemic inflammation and is associated with reduced placental growth factor (PlGF) and endothelial nitric oxide synthase (eNOS) levels. The molecular signaling axes involved in this process may play a role in the pathogenesis of preeclampsia. Here, we found that hypoxic exposure increased hypoxia-inducible factor-1α (HIF-1α)/Twist1-mediated miR-214-3p biogenesis in trophoblasts, suppressing PlGF production and trophoblast invasion. TNF-α stimulation increased NF-κB-dependent miR-214-3p expression in endothelial cells, impairing eNOS expression and causing endothelial dysfunction. Synthetic miR-214-3p administration to pregnant mice decreased PlGF and eNOS expression, resulting in preeclampsia-like symptoms, including hypertension, proteinuria, and fetal growth restriction. Conversely, miR-214-3p deletion maintained the PlGF and eNOS levels in hypoxic pregnant mice, alleviating preeclampsia-like symptoms and signs. These findings provide new insights into the role of HIF-1/Twist1- and NF-κB-responsive miR-214-3p-dependent PlGF and eNOS downregulation in the pathogenesis of preeclampsia and establish miR-214-3p as a therapeutic or preventive target for preeclampsia and its complications.

Small RNAs in the pathogenesis of preeclampsia

Preeclampsia is a major contributor to maternal and fetal morbidity and mortality. The disorder can be classified into early- and late-onset subtypes, both of which evolve in two stages. The first stage comprises the development of pre-clinical, utero-placental malperfusion. Early and late utero-placental malperfusion have different causes and time courses. Early-onset preeclampsia (20 % of cases) is driven by dysfunctional placentation in the first half of pregnancy. In late-onset preeclampsia (80 % of cases), malperfusion is a consequence of placental compression within the confines of a limited uterine cavity. In both subtypes, the malperfused placenta releases stress signals into the maternal circulation. These stress signals trigger onset of the clinical syndrome (the second stage). Small RNA molecules, which are implicated in cellular stress responses in general, may be involved at different stages. Micro RNAs contribute to abnormal trophoblast invasion, immune dysregulation, angiogenic imbalance, and syncytiotrophoblast-derived extracellular vesicle signalling in preeclampsia. Transfer RNA fragments are placental signals known to be specifically involved in cell stress responses. Disorder-specific differences in small nucleolar RNAs and piwi-interacting RNAs have also been reported. Here, we summarise key small RNA advances in preeclampsia pathogenesis. We propose that existing small RNA classifications are unhelpful and that non-biased assessment of RNA expression, incorporation of non-annotated molecules and consideration of chemical modifications to RNAs may be important in elucidating preeclampsia pathogenesis.

Combined analysis of the effects of hypoxia and oxidative stress on DNA methylation and the transcriptome in HTR-8/SVneo trophoblast cells.

The alterations in DNA methylation and transcriptome in trophoblast cells under conditions of low oxygen and oxidative stress have major implications for pregnancy-related disorders. However, the exact mechanism is still not fully understood. In this study, we established models of hypoxia (H group) and oxidative stress (HR group) using HTR-8/SVneo trophoblast cells and performed combined analysis of genome-wide DNA methylation changes using reduced representation bisulphite sequencing and transcriptome expression changes using RNA sequencing. Our findings revealed that the H group exhibited a higher number of differentially methylated genes and differentially expressed genes than the HR group. In the H group, only 0.90% of all differentially expressed genes displayed simultaneous changes in DNA methylation and transcriptome expression. After the threshold was expanded, this number increased to 6.29% in the HR group. Notably, both the H group and HR group exhibited concurrent alterations in DNA methylation and transcriptome expression within Axon guidance and MAPK signalling pathway. Among the top 25 differentially methylated KEGG pathways in the promoter region, 11 pathways were commonly enriched in H group and HR group, accounting for 44.00%. Among the top 25 KEGG pathways in transcriptome with significant differences between the H group and HR group, 10 pathways were consistent, accounting for 40.00%. By integrating our previous data on DNA methylation from preeclamptic placental tissues, we identified that the ANKRD37 and PFKFB3 genes may contribute to the pathogenesis of preeclampsia through DNA methylation-mediated transcriptome expression under hypoxic conditions.

Revealing ANXA6 as a Novel Autophagy-related Target for Pre-eclampsia Based on the Machine Learning

Background: Preeclampsia (PE) is a severe pregnancy complication associated with autophagy. Objective: This research sought to uncover autophagy-related genes in pre-eclampsia through bioinformatics and machine learning. Methods: GSE75010 from the GEO series was subjected to WGCNA to identify key modular genes in PE. Autophagy genes retrieved from the THANATOS overlapped with the modular genes to yield PErelated autophagy genes. Furthermore, the crucial step involved the utilization of two machine learning algorithms (LASSO and SVM-RFE) for dimensionality reduction. The candidate gene was further verified by quantitative reverse transcription polymerase chain reaction, western blot, and immunohistochemistry. Preliminary experiments were conducted on HTR-8/SVneo cell lines to explore the role of candidate genes in autophagy regulation. Results: WGCNA identified 291 genes from 5 hubs, and after overlapping with 1087 autophagy-related genes obtained from THANATOS, 42 PE-related ARGs were identified. ANXA6 was recognized as a potential target through SVM-RFE and LASSO analyses. The mRNA and protein expression of ANXA6 were verified in placenta samples. In HTR8/SVneo cells, modulating ANXA6 expression altered autophagy levels. Knocking down ANXA6 resulted in an anti-autophagy effect, which was reversed by treatment with CAL101, an inhibitor of PI3K, Akt, and mTOR. Conclusion: We observed that ANXA6 may serve as a possible PE action target and that autophagy may be crucial to the pathogenesis of PE.

ROLE OF CYTOKINES IN PRE-ECLAMPSIA: A REVIEW

Preeclampsia is a serious and life-threatening pregnancy complication. Reduced uteroplacental perfusion and oxygen tension impaired trophoblastic differentiation and invasion altered placental production of immunomodulators and growth factors are all considered important aspects of the aetiology of the condition. The placenta expresses a variety of pro and anti-inflammatory cytokines, adipokines and cytokine-like angiogenic growth factors, production of which is altered in preeclampsia, driven (at least in part) by hypoxia. Altered levels of cytokines have been measured in the circulation of women with preeclampsia. While the placenta undoubtedly makes an important contribution to plasma cytokine levels, production by maternal peripheral blood mononuclear cells (PBMCs) and other tissues is also likely to be significant, although to what extent remains undetermined. Increased placental expression of soluble receptors occurs with preeclampsia, resulting in elevated circulating concentrations which confer impaired angiogenesis, deficient placental vascularisation, increased placental apoptosis and endothelial dysfunction. The extent to which these changes reflect a response to the disorder, as opposed to being a causative factor in the development of maternal disease is a matter of some debate. Nevertheless, convincing evidence is now accruing that autocrine/paracrine interactions between placental cytokines/growth factors and the maternal endothelium plays a central role in the pathogenesis of preeclampsia.

The effects of leptin on human cytotrophoblast invasion are gestational age and dose-dependent.

Leptin and its receptors are expressed by the human placenta throughout gestation, yet the role of leptin in early human placental development is not well characterized. Leptin is overexpressed in the placentas from preeclamptic (PE) pregnancies. PE can result from the impaired invasion of fetal placental cells, cytotrophoblasts (CTBs), into the maternal decidua. We hypothesized that elevated leptin levels would impair human CTB invasion. The effects of leptin on the invasion of human CTBs were evaluated in three cell models, HTR-8/SVneo cells, primary CTBs, and placental villous explants using invasion assays. Further, leptin receptor expression was characterized in all three cell models using RT-PCR. Further phosphokinase assays were performed in HTR-8/SVneo cells to determine signaling pathways involved in CTB invasion in response to differential leptin doses. We found that, prior to 8 weeks gestation, leptin promoted CTB invasion in the explant model. After 11 weeks gestation in explants, primary CTBs and in HTR-8/SVneo cells, leptin promoted invasion at moderate but not at high concentrations. Further, leptin receptor characterization revealed that leptin receptor expression did not vary over gestation, however, STAT, PI3K and MAPK pathways showed different signaling in response to varied leptin doses. These data suggest that the excess placental leptin observed in PE may cause impaired CTB invasion as a second-trimester defect. Leptin's differential effect on trophoblast invasion may explain the role of hyperleptinemia in preeclampsia pathogenesis.

#1715 Role of T cell profile (Th1/Th2/Th17/Treg) and podocyturia in the peripheral blood of patients with preeclampsia versus normal pregnancy

Abstract Background and Aims The concept of the balance between Th17/Treg cell immunity plays important role in determining pregnancy outcomes. Does Treg/TH17 play a role in pathogenesis of preeclampsia associated proteinuria and its implication on irreversible renal damage was the research question. Method This hospital based prospective study conducted in the Department of obstetrics and gynecology at a tertiary care medical college between October 2018 till October 2020 and assessment of T cell profiling and urinary podocyturia among all pregnancies with clinical evidence of preeclampsia and compared it with normal pregnancy monitoring for maternal and foetal outcomes. Repeat assessment of Th profiling and podocyturia was done after 3 months post partum to look for any residual effect. Results A total of 40 patients of preeclampsia and 40 control were enrolled in the study with median age group (24.58 ± 1.22 vs 25.16 ± 1.01). The percentage of Th1 and Th2 were higher for the preeclampsia group as compared to the control group with no change in the percentage of Th17 subsets while Th17 1 were found higher and percentage of Treg (8.1 ± 3.19 vs 2.3 ± 1,05, p = 0.001) lower in the peripheral blood of the preeclampsia group at time of delivery however the ratio of Th1/Treg, Th2/Treg, Th17/Treg and Th17.1/Treg were found significantly lowered after 6 weeks of delivery among those with preeclampsia nearly comparable to the normal pregnancy group The quantitative assessment of podocyte in urine using antinephrin showed increase numbers among preeclamptic pregnancies compared to normal yet it settled within 6 weeks of delivery. Conclusion Women with preeclampsia per se does not increase the risk to exacerbation of kidney disease however greater research mandates to assess the women at greatest risk of kidney disease after pre-eclampsia to have good maternal and foetal outcomes.

TJP1 suppresses trophoblast cell invasion by expressing E2F8 in the human placenta

Adequate extravillous trophoblast (EVT) invasion into the maternal decidua is important for human placental development. We identified that E2F transcription factor 8 (E2F8) suppresses EVT invasion, and that tight junction protein-1 (TJP1) is a potential downstream target gene of E2F8. We investigated the role of TJP1 in the human placenta and regulation of TJP1 expression by E2F8. TJP1 expression decreased in E2F8 knockdown HTR-8/SVneo cells. TJP1 and E2F8 were co-expressed in villi in the first-trimester placenta and in EVTs and villi in the third-trimester placenta. TJP1 was significantly increased in the pre-eclamptic compared with control placenta. TJP1 knockdown increased the invasion of HTR-8/SVneo cells, while TJP1 overexpression inhibited cell invasion. Halo-E2F8 overexpression significantly increased TJP1 expression and TJP1 transcription compared with control placenta. Our findings suggest that E2F8 promotes TJP1 transcription, and that TJP1 expression by E2F8 inhibits EVT invasion. TJP1 and E2F8 may be related to pre-eclampsia pathogenesis.