Identification of HTRA4 as a Transcriptional Target of p63 in Trophoblast.

The placenta plays a crucial role in pregnancy success. ΔNp63α (p63), a transcription factor from the TP53 family, is highly expressed in villous cytotrophoblasts (CTBs), the epithelial stem cells of the human placenta, and is involved in CTB maintenance and differentiation. We examined the mechanisms of action of p63 by identifying its downstream targets. Gene expression changes were evaluated following overexpression and knockdown of p63 in the JEG3 choriocarcinoma cell line, using microarray-based RNA profiling. High-temperature requirement A4 (HTRA4), a placenta-specific serine protease involved in trophoblast differentiation and altered in preeclampsia, was identified as a gene reciprocally regulated by p63, and its expression was characterized in primary human placental tissues by RNA-sequencing and in situ hybridization. Potential p63 DNA-binding motifs were identified in the HTRA4 promoter, and p63 occupancy at some of these sites was confirmed using chromatin immunoprecipitation, followed by quantitative PCR in both JEG3 and trophoblast stem cells. These data begin to identify members of the transcriptional network downstream of p63, thus laying the groundwork for probing mechanisms by which this important transcription factor regulates trophoblast stemness and differentiation.

Evaluating the role of DNA methyltransferases in trophoblast fusion and preeclampsia development: Insights from methylation-regulated genes.

The etiology of preeclampsia (PE), a complex and multifactorial condition, remains incompletely understood. DNA methylation, which is primarily regulated by three DNA methyltransferases (DNMTs), DNMT1, DNMT3A, and DNMT3B, plays a vital role in early embryonic development and trophectoderm differentiation. Yet, how DNMTs modulate trophoblast fusion and PE development remains unclear. In this study, we found that the DNMTs expression was downregulated during trophoblast cells fusion. Downregulation of DNMTs was observed during the reconstruction of the denuded syncytiotrophoblast (STB) layer of placental explants. Additionally, overexpression of DNMTs inhibited trophoblast fusion. Conversely, treatment with the DNA methylation inhibitor 5-aza-CdR decreased the expression of DNMTs and promoted trophoblast fusion. A combined analysis of DNA methylation data and gene transcriptome data obtained from the primary cytotrophoblasts (CTBs) fusion process identified 104 potential methylation-regulated differentially expressed genes (MeDEGs) with upregulated expression due to DNA demethylation, including CD59, TNFAIP3, SDC1, and CDK6. The transcription regulation region (TRR) of TNFAIP3 showed a hypomethylation with induction of 5-aza-CdR, which facilitated CREB recruitment and thereby participated in regulating trophoblast fusion. More importantly, clinical correlation analysis of PE showed that the abnormal increase in DNMTs may be involved in the development of PE. This study identified placental DNA methylation-regulated genes that may contribute to PE, offering a novel perspective on the role of epigenetics in trophoblast fusion and its implication in PE development.

DNA methylation-mediated 11ßHSD2 downregulation drives the increases in angiotensin-converting enzyme and angiotensin II within preeclamptic placentas.

Preeclampsia (PE) is a complex human-specific complication frequently associated with placental pathology. The local renin-angiotensin system (RAS) in the human placenta, which plays a crucial role in regulating placental function, has been extensively documented. Glucocorticoids (GCs) are a class of steroid hormones. PE cases often have abnormalities in GCs levels and placental GCs barrier. Despite extensive speculation, there is currently no robust evidence indicating that GCs regulate placental RAS. This study aims to investigate these potential relationships. Plasma and placental samples were collected from both normal and PE pregnancies. The levels of angiotensin-converting enzyme (ACE), angiotensin II (Ang II), cortisol, and 11ß-hydroxysteroid dehydrogenases (11ßHSD) were analyzed. In PE placentas, cortisol, ACE, and Ang II levels were elevated, while 11ßHSD2 expression was reduced. Interestingly, a positive correlation was observed between ACE and cortisol levels in the placenta. A significant inverse correlation was found between the methylation statuses within the 11ßHSD2 gene promoter and its expression, meanwhile, 11ßHSD2 expression was negatively correlated with cortisol and ACE levels. In vitro experiments using placental trophoblast cells confirmed that active GCs can stimulate ACE transcription and expression through the GR pathway. Furthermore, 11ßHSD2 knockdown could enhance this activating effect. An in vivo study using a rat model of intrauterine GCs overexposure during mid-to-late gestation suggested that excess GCs in utero lead to increased ACE and Ang II levels in the placenta. Collectively, this study provides the first evidence of the relationships between 11ßHSD2 expression, GCs barrier, ACE, and Ang II levels in the placenta. It not only contributes to understanding the pathological features of the placental GCs barrier and RAS under PE conditions, also provides important information for revealing the pathological mechanism of PE.

The GPI-anchor biosynthesis pathway is critical for syncytiotrophoblast differentiation and placental development.

The glycosylphosphatidylinositol (GPI) biosynthetic pathway in the endoplasmic reticulum (ER) is crucial for generating GPI-anchored proteins (GPI-APs), which are translocated to the cell surface and play a vital role in cell signaling and adhesion. This study focuses on two integral components of the GPI pathway, the PIGL and PIGF proteins, and their significance in trophoblast biology. We show that GPI pathway mutations impact on placental development impairing the differentiation of the syncytiotrophoblast (SynT), and especially the SynT-II layer, which is essential for the establishment of the definitive nutrient exchange area within the placental labyrinth. CRISPR/Cas9 knockout of Pigl and Pigf in mouse trophoblast stem cells (mTSCs) confirms the role of these GPI enzymes in syncytiotrophoblast differentiation. Mechanistically, impaired GPI-AP generation induces an excessive unfolded protein response (UPR) in the ER in mTSCs growing in stem cell conditions, akin to what is observed in human preeclampsia. Upon differentiation, the impairment of the GPI pathway hinders the induction of WNT signaling for early SynT-II development. Remarkably, the transcriptomic profile of Pigl- and Pigf-deficient cells separates human patient placental samples into preeclampsia and control groups, suggesting an involvement of Pigl and Pigf in establishing a preeclamptic gene signature. Our study unveils the pivotal role of GPI biosynthesis in early placentation and uncovers a new preeclampsia gene expression profile associated with mutations in the GPI biosynthesis pathway, providing novel molecular insights into placental development with implications for enhanced patient stratification and timely interventions.

Overexpression of long noncoding RNA DUXAP8 inhibits ER-phagy through activating AKT/mTOR signaling and contributes to preeclampsia.

Preeclampsia (PE) is a life-threatening pregnancy-specific complication with controversial mechanisms and no effective treatment except delivery is available. Currently, increasing researchers suggested that PE shares pathophysiologic features with protein misfolding/aggregation disorders, such as Alzheimer disease (AD). Evidences have proposed defective autophagy as a potential source of protein aggregation in PE. Endoplasmic reticulum-selective autophagy (ER-phagy) plays a critical role in clearing misfolded proteins and maintaining ER homeostasis. However, its roles in the molecular pathology of PE remain unclear. We found that lncRNA DUXAP8 was upregulated in preeclamptic placentae and significantly correlated with clinical indicators. DUXAP8 specifically binds to PCBP2 and inhibits its ubiquitination-mediated degradation, and decreased levels of PCBP2 reversed the activation effect of DUXAP8 overexpression on AKT/mTOR signaling pathway. Function experiments showed that DUXAP8 overexpression inhibited trophoblastic proliferation, migration, and invasion of HTR-8/SVneo and JAR cells. Moreover, pathological accumulation of swollen and lytic ER (endoplasmic reticulum) was observed in DUXAP8-overexpressed HTR8/SVneo cells and PE placental villus trophoblast cells, which suggesting that ER clearance ability is impaired. Further studies found that DUXAP8 overexpression impaired ER-phagy and caused protein aggregation medicated by reduced FAM134B and LC3II expression (key proteins involved in ER-phagy) via activating AKT/mTOR signaling pathway. The increased level of FAM134B significantly reversed the inhibitory effect of DUXAP8 overexpression on the proliferation, migration, and invasion of trophoblasts. In vivo, DUXAP8 overexpression through tail vein injection of adenovirus induced PE-like phenotypes in pregnant rats accompanied with activated AKT/mTOR signaling, decreased expression of FAM134B and LC3-II proteins and increased protein aggregation in placental tissues. Our study reveals the important role of lncRNA DUXAP8 in regulating trophoblast biological behaviors through FAM134B-mediated ER-phagy, providing a new theoretical basis for understanding the pathogenesis of PE.

GPR124 promotes trophoblast proliferation, migration, and invasion and inhibits trophoblast cell apoptosis and inflammation via JNK and P38 MAPK pathways.

Early-onset preeclampsia, which occurrs before 34 weeks of gestation, is the most dangerous classification of preeclampsia, which is a pregnancy-specific disease that causes 1% of maternal deaths. G protein-coupled receptor 124 (GPR124) is significantly expressed at various stages of the human reproductive process, particularly during embryogenesis and angiogenesis. Our prior investigation demonstrated a notable decrease in GPR124 expression in the placentas of patients with early-onset preeclampsia compared to that in normal pregnancy placentas. However, there is a lack of extensive investigation into the molecular processes that contribute to the role of GPR124 in placenta development. This study aimed to examine the mechanisms by which GPR124 affects the occurrence of early-onset preeclampsia and its function in trophoblast. Proliferative, invasive, migratory, apoptotic, and inflammatory processes were identified in GPR124 knockdown, GPR124 overexpression, and normal HTR8/SVneo cells. The mechanism of GPR124-mediated cell function in GPR124 knockdown HTR8/SVneo cells was examined using inhibitors of the JNK or P38 MAPK pathway. Downregulation of GPR124 was found to significantly inhibit proliferation, invasion and migration, and promote apoptosis of HTR8/SVneo cells when compared to the control and GPR124 overexpression groups. This observation is consistent with the pathological characteristics of preeclampsia. In addition, GPR124 overexpression inhibits the secretion of pro-inflammatory cytokines interleukin (IL)-8 and interferon-γ (IFN-γ) while enhancing the secretion of the anti-inflammatory cytokine interleukin (IL)-4. Furthermore, GPR124 suppresses the activation of P-JNK and P-P38 within the JNK/P38 MAPK pathway. The invasion, apoptosis, and inflammation mediated by GPR124 were partially restored by suppressing the JNK and P38 MAPK pathways in HTR8/SVneo cells. GPR124 plays a crucial role in regulating trophoblast proliferation, invasion, migration, apoptosis, and inflammation via the JNK and P38 MAPK pathways. Furthermore, the effect of GPR124 on trophoblast suggests its involvement in the pathogenesis of early-onset preeclampsia.

The interaction of ER stress and autophagy in trophoblasts: navigating pregnancy outcome†.

The endoplasmic reticulum is a complex and dynamic organelle that initiates unfolded protein response and endoplasmic reticulum stress in response to the accumulation of unfolded or misfolded proteins within its lumen. Autophagy is a paramount intracellular degradation system that facilitates the transportation of proteins, cytoplasmic components, and organelles to lysosomes for degradation and recycling. Preeclampsia and intrauterine growth retardation are two common complications of pregnancy associated with abnormal trophoblast differentiation and placental dysfunctions and have a major impact on fetal development and maternal health. The intricate interplay between endoplasmic reticulum stress, and autophagy and their impact on pregnancy outcomes, through mediating trophoblast differentiation and placental development, has been highlighted in various reports. Autophagy controls trophoblast regulation through a variety of gene expressions and signaling pathways while excessive endoplasmic reticulum stress triggers downstream apoptotic signaling, culminating in trophoblast apoptosis. This comprehensive review delves into the intricacies of placental development and explores the underlying mechanisms of preeclampsia and intrauterine growth retardation. In addition, this review will elucidate the molecular mechanisms of endoplasmic reticulum stress and autophagy, both individually and in their interplay, in mediating placental development and trophoblast differentiation, particularly highlighting their roles in preeclampsia and intrauterine growth retardation development. This research seeks to the interplay between endoplasmic reticulum stress and impaired autophagy in the placental trophoderm, offering novel insights into their contribution to pregnancy complications.

Upregulation of Siglec-6 induces mitochondrial dysfunction by promoting GPR20 expression in early-onset preeclampsia.

BACKGROUND: Preeclampsia, especially early-onset preeclampsia (EO-PE), is a pregnancy complication that has serious consequences for the health of both the mother and the fetus. Although abnormal placentation due to mitochondrial dysfunction is speculated to contribute to the development of EO-PE, the underlying mechanisms have yet to be fully elucidated. METHODS: The expression and localization of Siglec-6 in the placenta from normal pregnancies, preterm birth and EO-PE patients were examined by RT-qPCR, Western blot and IHC. Transwell assays were performed to evaluate the effect of Siglec-6 on trophoblast cell migration and invasion. Seahorse experiments were conducted to assess the impact of disrupting Siglec-6 expression on mitochondrial function. Co-IP assay was used to examine the interaction of Siglec-6 with SHP1/SHP2. RNA-seq was employed to investigate the mechanism by which Siglec-6 inhibits mitochondrial function in trophoblast cells. RESULTS: The expression of Siglec-6 in extravillous trophoblasts is increased in placental tissues from EO-PE patients. Siglec-6 inhibits trophoblast cell migration and invasion and impairs mitochondrial function. Mechanismly, Siglec-6 inhibits the activation of NF-κB by recruiting SHP1/SHP2, leading to increased expression of GPR20. Notably, the importance of GPR20 function downstream of Siglec-6 in trophoblasts is supported by the observation that GPR20 downregulation rescues defects caused by Siglec-6 overexpression. Finally, overexpression of Siglec-6 in the placenta induces a preeclampsia-like phenotype in a pregnant mouse model. CONCLUSIONS: This study indicates that the regulatory pathway Siglec-6/GPR20 has a crucial role in regulating trophoblast mitochondrial function, and we suggest that Siglec-6 and GPR20 could serve as potential markers and targets for the clinical diagnosis and therapy of EO-PE.

RNA-binding protein SORBS2 increases human trophoblast cell migration via stabilizing HK2 mRNA in preeclampsia.

In brief: SORBS2, an RNA-binding protein, is identified as a regulator of aerobic glycolysis, which is essential for trophoblast migration and placental development. Reduced SORBS2 expression in preeclampsia may impair trophoblast migration by affecting mRNA stability and glycolysis, suggesting its role in the disease's pathogenesis. Abstract: Insufficient trophoblast migration and impaired uterine spiral artery remodeling are implicated in the pathogenesis of preeclampsia, contributing to inadequate placentation. However, the molecular mechanism underlying this process remains unclear. Aerobic glycolysis, which produces substantial lactate, is crucial for establishing a favorable microenvironment for early uterine preparation and supporting embryo implantation and trophoblast migration. In the present study, we have demonstrated that SORBS2, an RNA-binding protein, regulated aerobic glycolysis and significantly improved trophoblast migration in vitro. Our results showed that SORBS2 expression was significantly reduced in human PE placentas and trophoblasts during hypoxia. Overexpression of SORBS2 enhanced cell proliferation and migration, whereas knockdown of SORBS2 decreased these functions in HTR-8/SVneo cells. Mechanistic studies have demonstrated that SORBS2 directly interacts with the 3' untranslated regions (UTRs) of key glycolysis-related genes, specifically HK2. This interaction results in enhanced stability of HK2 and activation of glycolysis. Moreover, silencing HK2 abrogated the enhancement of proliferation and migration of HTR-8/SVneo cells induced by SORBS2. In conclusion, our findings suggest that the downregulation of SORBS2 may contribute to the pathogenesis of preeclampsia by regulating mRNA stability and inhibiting trophoblast migration during placentation.

Phenylbutyric acid inhibits hypoxia-induced trophoblast apoptosis and autophagy in preeclampsia via the PERK/ATF-4/CHOP pathway.

Preeclampsia (PE) is a common pregnancy complication with a high mortality rate. Abnormally activated endoplasmic reticulum stress (ERS) is believed to be responsible for the destruction of key placental cells-trophoblasts. Phenylbutyric acid (4-PBA), an ERS inhibitor, is involved in regulating the development of ERS-related diseases. At present, how 4-PBA affects trophoblasts and its mechanisms is still unclear. In this study, PE cell models were established by stimulating HTR-8/SVneo cells with hypoxia. To verify the underlying mechanisms of 4-PBA on PE, CCT020312, an activator of PERK, was also used. The results showed that 4-PBA restored hypoxia-induced trophoblast viability, inhibited HIF-1α protein expression, inflammation, and PERK/ATF-4/CHOP pathway. Hoechst 33342 staining and flow cytometry results confirmed that 4-PBA decreased hypoxia-induced apoptosis in trophoblasts. The results of the JC-1 analysis and apoptosis initiation enzyme activity assay also demonstrated that 4-PBA inhibited apoptosis related to the mitochondrial pathway. Furthermore, by detecting autophagy in trophoblasts, an increased number of autophagic vesicles, damaged mitochondria, enhanced dansylcadaverine fluorescence, enhanced levels of autophagy proteins Beclin-1, LC3II, and decreased p62 were seen in hypoxia-stimulated cells. These changes were reversed by 4-PBA. Furthermore, it was observed that CCT020312 reversed the effects of 4-PBA on the viability, apoptosis, and autophagosome number of hypoxia-induced trophoblasts. In summary, 4-PBA reduces autophagy and apoptosis via the PERK/ATF-4/CHOP pathway and mitochondrial pathway, thereby restoring the viability of hypoxic trophoblasts. These findings provide a solid evidence base for the use of 4-PBA in PE treatment and guide a new direction for improving the outcomes of patients with PE.

LINC01410 accelerates the invasion of trophoblast cells by modulating METTL3/Fas.

BACKGROUND: Insufficient trophoblast invasion, culminating in suboptimal uterine spiral artery remodeling, is pinpointed as a pivotal contributor to preeclampsia (PE) development. LINC01410 has been documented to be increased in various neoplasms, and is significantly associated with the invasive capabilities of tumor cells. Nonetheless, its function and the mechanisms in the pathogenesis of PE require further investigation. METHODS AND RESULTS: LINC01410 and methyltransferase-like 3 (METTL3) were ectopically expressed in HTR-8/Svneo cells via lentiviral transduction. Subsequently, the cells' invasive capabilities and apoptosis rates were evaluated employing Transwell assays and flow cytometry, respectively. The interplay between LINC01410 and METTL3, alongside the m6A methylation of FAS, was probed through RNA immunoprecipitation (RIP). Additionally, the association between FAS and METTL3 was elucidated via Coimmunoprecipitation (Co-IP) assays. The protein level of NF-κB, BAX, and BCL-2 in LINC01410-overexpressing cells was detected by Western blot. Our findings revealed that LINC01410 elevation increased the invasive ability of HTR-8/Svneo cells, directly impacting METTL3 then leading to its reduced expression. Conversely, heightened METTL3 expression mitigated invasiveness while enhancing apoptosis in these cells. Moreover, METTL3's interaction with FAS led to increased FAS expression, subject to m6A methylation. A surge in LINC01410 markedly decreased both mRNA and protein levels of FAS. Furthermore, LINC01410 overexpression significantly reduced NF-κB and BAX protein levels while augmenting BCL-2. CONCLUSIONS: Upregulation of LINC01410 expression promotes trophoblast cell invasion by inhibiting FAS levels through modified m6A alteration and suppressing the NF-κB pathway. These findings underscore the pivotal role of LINC01410 in regulating trophoblast cell invasion and propose it as a promising therapeutic strategy for preventing or alleviating PE. This offers valuable insights for the clinical treatment of PE, for which definitive targeted therapy methods are currently lacking.

Relationship Between Anatomic Features of the Placenta, the Type of Abnormal Placental Cord Insertion and Adverse Pregnancy Outcomes in Singleton Pregnancies: A Prospective Observational Study.

INTRODUCTION: This study aimed to evaluate the potential value of placental anatomic features and various types of normal and abnormal cord insertion types in predicting adverse maternal-fetal outcomes in singleton pregnancies. We also tried to assess the association between these outcomes and various types of placental cord insertion. METHOD: This prospective observational study was performed on singleton pregnancies. For each patient placental features including diameter, thickness, type of cord insertion, and the shortest distance between the cord insertion point and placental edge were recorded. The relationship between these factors and the development of multiple adverse pregnancy outcomes including preterm labor, intrauterine fetal death (IUFD), and the rate of neonatal intensive care unit (NICU) admissions were evaluated and reported. RESULTS: Overall 308 patients were enrolled in the study. Smoker mothers had significantly smaller placentas (P-value = .008), and those with lower diameter placentas were more likely to suffer from IUFD (P-value = .0001). Shorter placental cord insertion distances led to more episodes of preterm labor (P-value = .057). Eccentric-type placental cord insertion was significantly associated with the development of preeclampsia (P-value = .006). DISCUSSION: Abnormalities in placental diameter and cord insertion can lead to significant maternal-fetal complications including preterm labor, IUFD, and preeclampsia.

Pathologic maternal and neonatal outcomes associated with programmed embryo transfer: potential etiologies and strategies for prevention.

PURPOSE: In the first of two companion papers, we comprehensively reviewed the recent evidence in the primary literature, which addressed the increased prevalence of hypertensive disorders of pregnancy, late-onset or term preeclampsia, fetal overgrowth, postterm birth, and placenta accreta in women conceiving by in vitro fertilization. The preponderance of evidence implicated frozen embryo transfer cycles and, specifically, those employing programmed endometrial preparations, in the higher risk for these adverse maternal and neonatal pregnancy outcomes. Based upon this critical appraisal of the primary literature, we formulate potential etiologies and suggest strategies for prevention in the second article. METHODS: Comprehensive review of primary literature. RESULTS: Presupposing significant overlap of these apparently diverse pathological pregnancy outcomes within subjects who conceive by programmed autologous FET cycles, shared etiologies may be at play. One plausible but clearly provocative explanation is that aberrant decidualization arising from suboptimal endometrial preparation causes greater than normal trophoblast invasion and myometrial spiral artery remodeling. Thus, overly robust placentation produces larger placentas and fetuses that, in turn, lead to overcrowding of villi within the confines of the uterine cavity which encroach upon intervillous spaces precipitating placental ischemia, oxidative and syncytiotrophoblast stress, and, ultimately, late-onset or term preeclampsia. The absence of circulating corpus luteal factors like relaxin in most programmed cycles might further compromise decidualization and exacerbate the maternal endothelial response to deleterious circulating placental products like soluble fms-like tyrosine kinase-1 that mediate disease manifestations. An alternative, but not mutually exclusive, determinant might be a thinner endometrium frequently associated with programmed endometrial preparations, which could conspire with dysregulated decidualization to elicit greater than normal trophoblast invasion and myometrial spiral artery remodeling. In extreme cases, placenta accreta could conceivably arise. Though lower uterine artery resistance and pulsatility indices observed during early pregnancy in programmed embryo transfer cycles are consistent with this initiating event, quantitative analyses of trophoblast invasion and myometrial spiral artery remodeling required to validate the hypothesis have not yet been conducted. CONCLUSIONS: Endometrial preparation that is not optimal, absent circulating corpus luteal factors, or a combination thereof are attractive etiologies; however, the requisite investigations to prove them have yet to be undertaken. Presuming that in ongoing RCTs, some or all adverse pregnancy outcomes associated with programmed autologous FET are circumvented or mitigated by employing natural or stimulated cycles instead, then for women who can conceive using these regimens, they would be preferable. For the 15% or so of women who require programmed FET, additional research as suggested in this review is needed to elucidate the responsible mechanisms and develop preventative strategies.

The relationship between Toll-like receptor-4 genes and preeclampsia outcomes.

PURPOSE: The study aimed to analyse the relationship of the rs4986790 locus of the TLR4 gene with the overall risk of preeclampsia, including both its early and late forms. METHODS: The study used standard genetic analysis methods such as DNA extraction, PCR amplification, and genotyping of the rs4986790 locus of the TLR4 gene to assess the association with the development of preeclampsia and peripartal stroke in 207 pregnant women from the southern regions of Kazakhstan from 2016 to 2022, of whom 103 had peripartal stroke on the background of preeclampsia (the main group) and 104 preeclampsia (comparative group). RESULTS: The results of the study demonstrate that the AG and AG + GG genotypes at the rs4986790 locus of the TLR4 gene are significantly associated with an increased risk of developing an early form of preeclampsia. This opens up a new perspective in the identification of genetic markers that can serve as indicators of a tendency to develop preeclampsia in earlier periods of pregnancy. CONCLUSION: It was noted that the rs4986790 locus did not show a statistically significant association with the risk of late preeclampsia. An important aspect of the study revealed the relationship of genotypes with the development of peripartal stroke on the background of preeclampsia. This study offers practical insights for creating targeted genetic screening and personalised treatments for preeclampsia, aiming to improve patient outcomes. To fully understand the molecular mechanisms underlying the identified association, additional research is required to identify deeper molecular pathways and relationships, and to develop new strategies for the prevention and treatment of preeclampsia.

Pathologic maternal and neonatal outcomes associated with programmed embryo transfer.

PURPOSE: In this first of two companion papers, we critically review the evidence recently published in the primary literature, which addresses adverse maternal and neonatal pregnancy outcomes associated with programmed embryo transfer cycles. We next consider whether these pathological pregnancy outcomes might be attributable to traditional risk factors, unknown parental factors, embryo culture, culture duration, or cryopreservation. Finally, in the second companion article, we explore potential etiologies and suggest strategies for prevention. METHODS: Comprehensive review of primary literature. RESULTS: The preponderance of retrospective and prospective observational studies suggests that increased risk for hypertensive disorders of pregnancy (HDP) and preeclampsia in assisted reproduction involving autologous embryo transfer is associated with programmed cycles. For autologous frozen embryo transfer (FET) and singleton live births, the risk of developing HDP and preeclampsia, respectively, was less for true or modified natural and stimulated cycles relative to programmed cycles: OR 0.63 [95% CI (0.57-0.070)] and 0.44 [95% CI (0.40-0.50)]. Though data are limited, the classification of preeclampsia associated with programmed autologous FET was predominantly late-onset or term disease. Other adverse pregnancy outcomes associated with autologous FET, especially programmed cycles, included increased prevalence of large for gestational age infants and macrosomia, as well as higher birth weights. In one large registry study, FET was associated with fetal overgrowth of a symmetrical nature. Postterm birth and placenta accreta not associated with prior cesarean section, uterine surgery, or concurrent placenta previa were also associated with autologous FET, particularly programmed cycles. The heightened risk of these pathologic pregnancy outcomes in programmed autologous FET does not appear to be attributable to traditional risk factors, unknown parental factors, embryo culture, culture duration, or cryopreservation, although the latter may contribute a modest degree of increased risk for fetal overgrowth and perhaps HDP and preeclampsia in FET irrespective of the endometrial preparation. CONCLUSIONS: Programmed autologous FET is associated with an increased risk of several, seemingly diverse, pathologic pregnancy outcomes including HDP, preeclampsia, fetal overgrowth, postterm birth, and placenta accreta. Though the greater risk for preeclampsia specifically associated with programmed autologous FET appears to be well established, further research is needed to substantiate the limited data currently available suggesting that the classification of preeclampsia involved is predominately late-onset or term. If substantiated, then this knowledge could provide insight into placental pathogenesis, which has been proposed to differ between early- and late-onset or term preeclampsia (see companion paper for a discussion of potential mechanisms). If a higher prevalence of preeclampsia with severe features as suggested by some studies is corroborated in future investigations, then the danger to maternal and fetal/neonatal health is considerably greater with severe disease, thus increasing the urgency to find preventative measures. Presupposing significant overlap of these diverse pathologic pregnancy outcomes within subjects who conceive by programmed embryo transfer, there may be common etiologies.

Exosomal encapsulation of miR-3198 promotes proliferation and migration of trophoblasts in preeclampsia.

PURPOSE: Preeclampsia (PE) is a vascular remodeling disorder cloesly linked to trophoblast dysfunction, involving defects in their proliferation, migration, and apoptosis. Maternal exosomal microRNAs (miRNAs) have been reported to play pivotal roles in the development of PE. However, the mechanism underlying the role of maternal exosomes in trophoblast dysfunction regarding the development of PE is poorly understood. METHODS: Plasma exosomes from maternal peripheral blood were collected from pregnant women with PE and from those with normal pregnancy. Bioinformatics analysis was used to identify significantly differentially expressed miRNAs under these two conditions. The expression of the miR-3198 gene in plasma exosomes was detected using quantitative real-time polymerase chain reaction. Dual luciferase reporter assay was used to confirm binding of miR-3198 and 3'UTR region of WNT3. Cell proliferation was examined using the Cell Count Kit-8 and EdU assays, and flow cytometry was performed to detect apoptosis and cell cycle. Changes in cell migration were examined using transwell and scratch assays. RESULTS: Patients with PE showed decreased expression of plasma-derived exosomal miR-3198. The proliferation and migration abilities of HTR-8/SVneo and primary human trophoblast cells were both improved when cocultured with miR-3198-rich exosomes. Exposure to miR-3198-enriched exosomes facilitated cell cycle progression but reduced apoptosis in HTR-8/SVneo cells. Notably, overexpression of miR-3198 partially prevented the inhibitory effects of WNT3 on proliferation and migration in HTR-8/SVneo cells. CONCLUSION: Exosomal miR-3198 in the maternal peripheral blood may regulate the biological functions of trophoblasts by targeting WNT3 and influence the development of diseases of placental origin.

The effect of bariatric surgery on polycystic ovary syndrome patients' obstetric and neonatal outcomes: a population-based study.

PURPOSE: To examine the effect of bariatric surgery (BS) on obstetric and neonatal outcomes in patients with polycystic ovary syndrome (PCOS). METHODS: A retrospective population-based cohort study utilizing the Healthcare Cost and Utilization Project Nationwide Inpatient Sample database, including women who delivered in the third trimester or had a maternal death in the USA (2004-2014). We compared obstetric and neonatal outcomes between groups in three analyses: (1) Primary analysis-women with an ICD-9 PCOS diagnosis who underwent BS compared to pregnant PCOS patients without BS. (2) Sub-group analysis-PCOS women with BS compared to obese PCOS women (body mass index (BMI) ≥ 30 kg/m2) without BS. (3) Women with and without PCOS who underwent BS. RESULT: In the primary analysis, pregnant PCOS women who underwent BS (N = 141), compared to pregnant PCOS women without BS (N = 14,741), were less likely to develop pregnancy-induced hypertension (PIH) (9.2% vs. 16.2%, respectively, aOR 0.39, 95% CI 0.21-0.72) and gestational diabetes mellitus (GDM) (9.9% vs. 18.8, aOR 0.40, 95% CI 0.23-0.70). In the sub-group analysis, PCOS women with BS, compared to obese PCOS women without BS (N = 3231), were less likely to develop gestational hypertension, preeclampsia, and preeclampsia or eclampsia superimposed on hypertension (P < 0.05). Lastly, PCOS patients with BS had a higher cesarean section rate when compared to non-PCOS patients with BS (N = 9197) (61.7% vs. 49.2%, aOR 1.48, 95% CI 1.05-2.09), with otherwise comparable obstetric and neonatal outcomes. CONCLUSIONS: BS in PCOS patients was associated with reduced risks for GDM and PIH when compared to PCOS controls without BS and reduced risk for gestational hypertension, preeclampsia, and preeclampsia or eclampsia superimposed on hypertension when compared to obese PCOS controls without BS. Moreover, BS was associated with reduced inherent pregnancy risks of PCOS, almost equating them to those of non-PCOS counterparts.

Alpha 1,3 N-Acetylgalactosaminyl Transferase (GTA) Impairs Invasion Potential of Trophoblast Cells in Preeclampsia.

Preeclampsia (PE) is a pregnancy-specific disorder associated with shallow invasion of the trophoblast cells and insufficient remodeling of the uterine spiral artery. Protein glycosylation plays an important role in trophoblast cell invasion. However, the glycobiological mechanism of PE has not been fully elucidated. In the current study, employing the Lectin array, we found that soybean agglutinin (SBA), which recognizes the terminal N-acetylgalactosamine α1,3-galactose (GalNAc α1,3 Gal) glycotype, was significantly increased in placental trophoblast cells from PE patients compared with third-trimester pregnant controls. Upregulating the expression of the key enzyme α1,3 N-acetylgalactosaminyl transferase (GTA) promoted the biosynthesis of terminal GalNAc α1,3 Gal and inhibited the migration/invasion of HTR8/SVneo trophoblast cells. Moreover, the methylation status of GTA promoter in placental tissues from PE patients was lower than that in the third trimester by methylation-specific PCR (MSP) and bisulfite sequencing PCR (BSP) analysis. Elevated GTA expression in combination with the DNA methylation inhibitor 5-azacytidine (5-AzaC) treatment increased the glycotype biosynthesis and impaired the invasion potential of trophoblast cells, leading to preeclampsia. This study suggests that elevated terminal GalNAc α1,3 Gal biosynthesis and GTA expression may be applied as the new markers for evaluating placental function and the auxiliary diagnosis of preeclampsia.

Behind the Curtain of Abnormal Placentation in Pre-Eclampsia: From Molecular Mechanisms to Histological Hallmarks.

Successful human pregnancy needs several highly controlled steps to guarantee an oocyte's fertilization, the embryo's pre-implantation development, and its subsequent implantation into the uterine wall. The subsequent placenta development ensures adequate fetal nutrition and oxygenation, with the trophoblast being the first cell lineage to differentiate during this process. The placenta sustains the growth of the fetus by providing it with oxygen and nutrients and removing waste products. It is not surprising that issues with the early development of the placenta can lead to common pregnancy disorders, such as recurrent miscarriage, fetal growth restriction, pre-eclampsia, and stillbirth. Understanding the normal development of the human placenta is essential for recognizing and contextualizing any pathological aberrations that may occur. The effects of these issues may not become apparent until later in pregnancy, during the mid or advanced stages. This review discusses the process of the embryo implantation phase, the molecular mechanisms involved, and the abnormalities in those mechanisms that are thought to contribute to the development of pre-eclampsia. The review also covers the histological hallmarks of pre-eclampsia as found during the examination of placental tissue from pre-eclampsia patients.

Altered 5-methylcytosine modification of mRNA is involved in the pathogenesis of pre-eclampsia.

5-Methylcytosine (m5 C) is a prevalent RNA modification in messenger RNAs (mRNAs). Despite its abundance, its role in the decidua of pre-eclampsia (PE) remains elusive. In this study, we utilized methylated RNA immunoprecipitation sequencing (MeRIP-seq) and RNA-sequencing (RNA-seq) to map m5 C peaks and mRNA expression profile in the decidua of human early-onset PE (EPE), late-onset PE (LPE), and normal pregnancy (NP). Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway analyses elucidated potential roles of the differentially methylated mRNAs (DMGs) and differentially expressed mRNAs in decidualization pathways. Integrative analysis of MeRIP-seq and RNA-seq data pinpointed 50 candidate genes linked to PE, marked by both differentially methylated m5 C peaks and congruent expression changes. To validate these observations, we selected nine genes for verification via quantitative PCR. Our results underscore the precision and reproducibility of our bioinformatics approach. Importantly, we propose that changes in m5 C modification and expression of relevant mRNA might influence the pathogenesis of PE by hampering decidualization. This work shines light on the distinct mRNA m5 C modification patterns and expression profiles in the decidua of PE, implicating pivotal signaling disruptions and decidualization impediments in the onset of PE.