Embryo Quality Conditions the Secretory Profile of Tolerogenic Dendritic Cell DC-10 During the Peri-Implantation Period.

PROBLEM: The decidualization process conditions monocytes to the immunosuppressive and tolerogenic dendritic cell (DC)-10 profile, a DC subset with high IL-10 production. Since the implantation process implies an embryo-endometrium-immune crosstalk, here we focused on the ability of embryonic soluble factors to modify decidual DC conditioning accordingly with its quality. METHOD OF STUDY: Human endometrial stromal cell line (HESC) decidualized with medroxyprogesterone and dibutyryl-cAMP (Dec) was stimulated with human embryo-conditioned media (ECM), classified as normal (ND) or impaired developed (ID) for 48 h (n = 18/group). Monocytes isolated from six healthy women were differentiated to DCs with rhGM-CSF+rhIL-4 in the presence/absence of conditioned media (CM) from decidualized cells stimulated with ECM or nontreated. RESULTS: We found that decidualized cells stimulated with ECM sustain a myeloid regulatory cell profile on monocyte-derived culture with increased frequency of CD1a-CD14+ and CD83+CD86low cells. ND-Dec sustained the higher expression of the DC-10 markers, HLA-G and IL-10 whereas ID-Dec diminished IL-10 production (ID-Dec: 135 ± 37.4 vs. Dec: 223.3 ± 49.9 pg/mL, p < 0.05). The treatment with ECM-Dec sustained a higher IL-10 production and prevented the increase of CD83/CD86 after LPS challenge regardless of embryo quality. Notably, TNF-α production increased in ID-Dec cultures (ID-Dec: 475.1 ± 134.7 vs. Dec: 347.5 ± 98 pg/mL, p < 0.05). CONCLUSIONS: Although remaining in a tolerogenic profile compatible with DC-10, DCs can differentially respond to decidual secreted factors based on embryo quality, changing their secretome. These results suggest that in the presence of arrested embryo, DCs could differentially shape the immunological microenvironment, contributing to arrested embryo clearance during the menstrual phase.

[Notch1/Akt/Foxo1 Pathway Regulated by Kisspeptin Is Involved in Endometrial Decidualization in Patients With Recurrent Spontaneous Abortion]. / kisspeptin调控Notch1/Akt/Foxo1é€šè·¯å‚ä¸Žå¤å‘æ€§æµäº§æ‚£è€ å­å®«å† è†œèœ•è†œåŒ–.

Objective: Kisspeptin, a protein encoded by the KISS1 gene, functions as an essential factor in suppressing tumor growth. The intricate orchestration of cellular processes such as proliferation and differentiation is governed by the Notch1/Akt/Foxo1 signaling pathway, which assumes a central role in maintaining cellular homeostasis. In the specific context of this investigation, the focal point lies in a meticulous exploration of the intricate mechanisms underlying the regulatory effect of kisspeptin on the process of endometrial decidualization. This investigation delves into the interplay between kisspeptin and the Notch1/Akt/Foxo1 signaling pathway, aiming to elucidate its significance in the pathophysiology of recurrent spontaneous abortion (RSA). Methods: We enrolled a cohort comprising 45 individuals diagnosed with RSA, who were admitted to the outpatient clinic of the Reproductive Center at the Second Affiliated Hospital of Soochow University between June 2020 and December 2020. On the other hand, an additional group of 50 women undergoing elective abortion at the outpatient clinic of the Family Planning Department during the same timeframe was also included. To comprehensively assess the molecular landscape, Western blot and RT-qPCR were performed to analyze the expression levels of kisspeptin (and its gene KISS1), IGFBP1 (an established marker of decidualization), Notch1, Akt, and Foxo1 within the decidua. Human endometrial stromal cells (hESC) were given targeted interventions, including treatment with siRNA to disrupt KISS1 or exposure to kisspeptin10 (the bioactive fragment of kisspeptin), and were subsequently designated as the siKP group or the KP10 group, respectively. A control group comprised hESC was transfected with blank siRNA, and cell proliferation was meticulously evaluated with CCK8 assay. Following in vitro induction for decidualization across the three experimental groups, immunofluorescence assay was performed to identify differences in Notch1 expression and decidualization morphology between the siKP and the KP10 groups. Furthermore, RT-qPCR and Western blot were performed to gauge the expression levels of IGFBP1, Notch1, Akt, and Foxo1 across the three cell groups. Subsequently, decidualization was induced in hESC by adding inhibitors targeting Notch1, Akt, and Foxo1. The expression profiles of the aforementioned proteins and genes in the four groups were then examined, with hESC induced for decidualization without adding inhibitors serving as the normal control group. To establish murine models of normal pregnancy (NP) and RSA, CBA/J×BALB/c and CBA/J×DBA/2 mice were used. The mice were respectively labeled as the NP model and RSA model. The experimental groups received intraperitoneal injections of kisspeptin10 and kisspeptin234 (acting as a blocker) and were designated as RSA-KP10 and NP-KP234 groups. On the other hand, the control groups received intraperitoneal injections of normal saline (NS) and were referred to as RSA-NS and NP-NS groups. Each group comprised 6 mice, and uterine tissues from embryos at 9.5 days of gestation were meticulously collected for observation of embryo absorption and examination of the expression of the aforementioned proteins and genes. Results: The analysis revealed that the expression levels of kisspeptin, IGFBP1, Notch1, Akt, and Foxo1 were significantly lower in patients diagnosed with RSA compared to those in women with NP (P<0.01 for kisspeptin and P<0.05 for IGFBP1, Notch1, Akt, and Foxo1). After the introduction of kisspeptin10 to hESC, there was an observed enhancement in decidualization capability. Subsequently, the expression levels of Notch1, Akt, and Foxo1 showed an increase, but they decreased after interference with KISS1. Through immunofluorescence analysis, it was observed that proliferative hESC displayed a slender morphology, but they transitioned to a rounder and larger morphology post-decidualization. Concurrently, the expression of Notch1 increased, suggesting enhanced decidualization upon the administration of kisspeptin10, but the expression decreased after interference with KISS1. Further experimentation involved treating hESC with inhibitors specific to Notch1, Akt, and Foxo1 separately, revealing a regulatory sequence of Notch1/Akt/Foxo1 (P<0.05). In comparison to the NS group, NP mice administered with kisspeptin234 exhibited increased fetal absorption rates (P<0.001) and decreased expression of IGFBP1, Notch1, Akt, and Foxo1 (P<0.05). Conversely, RSA mice administered with kisspeptin10 demonstrated decreased fetal absorption rates (P<0.001) and increased expression levels of the aforementioned molecules (P<0.05). Conclusion: It is suggested that kisspeptin might exert its regulatory influence on the process of decidualization through the modulation of the Notch1/Akt/Foxo1 signaling cascade. A down-regulation of the expression levels of kisspeptin could result in suboptimal decidualization, which in turn might contribute to the development or progression of RSA.

Progesterone inactivation in decidual stromal cells: A mechanism for inflammation-induced parturition.

The process of human parturition involves inflammation at the interface where fetal chorion trophoblast cells interact with maternal decidual stromal (DS) cells and maternal immune cells in the decidua (endometrium of pregnancy). This study tested the hypothesis that inflammation at the chorion-decidua interface (CDI) induces labor by negating the capacity for progesterone (P4) to block labor and that this is mediated by inactivation of P4 in DS cells by aldo-keto reductase family 1 member C1 (AKR1C1). In human, Rhesus macaque, and mouse CDI, AKR1C1 expression increased in association with term and preterm labor. In a human DS cell line and in explant cultures of term human fetal membranes containing the CDI, the prolabor inflammatory cytokine, interleukin-1ß (IL-1ß), and media conditioned by LPS-stimulated macrophages increased AKR1C1 expression and coordinately reduced nuclear P4 levels and P4 responsiveness. Loss of P4 responsiveness was overcome by inhibition of AKR1C1 activity, inhibition of AKR1C1 expression, and bypassing AKR1C1 activity with a P4 analog that is not metabolized by AKR1C1. Increased P4 activity in response to AKR1C1 inhibition was prevented by the P4 receptor antagonist RU486. Pharmacologic inhibition of AKR1C1 activity prevented parturition in a mouse model of inflammation-induced preterm parturition. The data suggest that inflammatory stimuli at the CDI drive labor by inducing AKR1C1-mediated P4 inactivation in DS cells and that inhibiting and/or bypassing of AKR1C1-mediated P4 inactivation is a plausible therapeutic strategy to mitigate the risk of inflammation-associated preterm birth.

Trophoblastic signals facilitate endometrial interferon response and lipid metabolism, ensuring normal decidualization.

The decidua plays a crucial role in providing structural and trophic support to the developing conceptus before placentation. Following embryo attachment, embryonic components intimately interact with the decidual tissue. While evidence indicates the participation of embryo-derived factors in crosstalk with the uterus, the extent of their impact on post-implantation decidual development requires further investigation. Here, we utilize transgenic mouse models to selectively eliminate primary trophoblast giant cells (pTGCs), the embryonic cells that interface with maternal tissue at the forefront. pTGC ablation impairs decidualization and compromises decidual interferon response and lipid metabolism. Mechanistically, pTGCs release factors such as interferon kappa (IFNK) to strengthen the decidual interferon response and lipoprotein lipase (LPL) to enhance lipid accumulation within the decidua, thereby promoting decidualization. This study presents genetic and metabolomic evidence reinforcing the proactive role of pTGC-derived factors in mobilizing maternal resources to strengthen decidualization, facilitating the normal progression of early pregnancy.

Lysosome dynamics during human endometrial stromal cells decidualization: effect of para-nonylphenol.

During the process of decidualization, the stromal cells of the endometrium change dynamically to create a favorable environment for embryo implantation. Lysosome activity has often been associated with physiological changes in the endometrium during the preimplantation period and early pregnancy. In this study, the effect of para-nonylphenol (p-NP), an endocrine disruptor, on human immortalized endometrial stromal cells (tHESCs) was investigated. After exposure to p-NP (1 nM and 1 pM), the cells were examined for the decidualization markers connexin-43, insulin like growth factor binding protein 1 (IGFBP1), and prolactin. In addition, the effect of p-NP on lysosome biogenesis and exocytosis was investigated by examining the expression and localization of the transcription factor EB (TFEB) and that of the lysosomal-associated membrane protein 1 (LAMP-1). Finally, we evaluated the effect of p-NP on extracellular matrix (ECM) remodeling using a fibronectin assay. Our results showed that p-NP reduced the expression of prolactin protein, increased the nuclear localization of TFEB, and induced the increase and translocation of the lysosomal protein LAMP-1 to the membrane of tHESCs. The data indicate an impairment of decidualization and suggest an increase in lysosomal biogenesis and exocytosis, which is supported by the higher release of active cathepsin D by tHESCs. Given the importance of cathepsins in the processing and degradation of the ECM during trophoblast invasiveness and migration into the decidua, our results appear to be clear evidence of the negative effects of p-NP on endometrial processes that are fundamental to reproductive success and the establishment of pregnancy.NEW & NOTEWORTHY Endocrine disruptors, such as para-nonylphenol, affect the decidualization of human endometrial stromal cells with an impact on decidualization itself, lysosome biogenesis and exocytosis, and extracellular matrix remodeling. All these alterations may negatively impact embryo implantation with the success of reproduction and the establishment of pregnancy.

The Long Non-Coding RNA Gene AC027288.3 Plays a Role in Human Endometrial Stromal Fibroblast Decidualization.

During the secretory phase of the menstrual cycle, endometrial fibroblast cells begin to change into large epithelial-like cells called decidual cells in a process called decidualization. This differentiation continues more broadly in the endometrium and forms the decidual tissue during early pregnancy. The cells undergoing decidualization as well as the resulting decidual cells, support successful implantation and placentation during early pregnancy. This study was carried out to identify new potentially important long non-coding RNA (lncRNA) genes that may play a role in human endometrial stromal fibroblast cells (hESF) undergoing decidualization in vitro, and several were found. The expression of nine was further characterized. One of these, AC027288.3, showed a dramatic increase in the expression of hESF cells undergoing decidualization. When AC027288.3 expression was targeted, the ability of the cells to undergo decidualization as determined by the expression of decidualization marker protein-coding genes was significantly altered. The most affected markers of decidualization whose expression was significantly reduced were FOXO1, FZD4, and INHBA. Therefore, AC027288.3 may be a major upstream regulator of the WNT-FOXO1 pathway and activin-SMAD3 pathways previously shown as critical for hESF decidualization. Finally, we explored possible regulators of AC027288.3 expression during human ESF decidualization. Expression was regulated by cAMP and progesterone. Our results suggest that AC027288.3 plays a role in hESF decidualization and identifies several other lncRNA genes that may also play a role.

LIF-STAT signaling in decidual cells: a possible role in embryo implantation and early pregnancy.

In this study, we investigate the effects of miRNA-138-5p and probable G-protein coupled receptor 124 (GPR124)-regulated inflammasome and downstream leukemia inhibitory factor (LIF)-STAT and adhesion molecule signaling in human decidual stromal cells. After informed consent was obtained from women aged 25-38 years undergoing surgical termination of the normal pregnancy and spontaneous miscarriage after 6-9 weeks of gestation, human decidual stromal cells were extracted from the decidual tissue. Extracellular vesicles (EVs) with microRNA (miRNA) between cells have been regarded as critical factors for embryo-maternal interactions on embryo implantation and programming of human pregnancy. MicroRNA-138-5p acts as the transcriptional regulator of GPR124 and the mediator of downstream inflammasome. LIF-regulated STAT activation and expression of integrins might influence embryo implantation. Hence, a better understanding of LIF-STAT and adhesion molecule signaling would elucidate the mechanism of microRNA-138-5p- and GPR124-regulated inflammasome activation on embryo implantation and pregnancy. Our results show that microRNA-138-5p, purified from the EVs of decidual stromal cells, inhibits the expression of GPR124 and the inflammasome, and activates the expression of LIF-STAT and adhesion molecules in human decidual stromal cells. Additionally, the knockdown of GPR124 and NLRP3 through siRNA increases the expression of LIF-STAT and adhesion molecules. The findings of this study help us gain a better understanding the role of EVs, microRNA-138-5p, GPR124, inflammasomes, LIF-STAT, and adhesion molecules in embryo implantation and programming of human pregnancy.

CD4+ T cell heterogeneity in gestational age and preeclampsia using single-cell RNA sequencing.

A balance between pro-inflammatory decidual CD4+ T cells and FOXP3+ regulatory T cells (FOXP3+ Tregs) is important for maintaining fetomaternal tolerance. Using single-cell RNA-sequencing and T cell receptor repertoire analysis, we determined that diversity and clonality of decidual CD4+ T cell subsets depend on gestational age. Th1/Th2 intermediate and Th1 subsets of CD4+ T cells were clonally expanded in both early and late gestation, whereas FOXP3+ Tregs were clonally expanded in late gestation. Th1/Th2 intermediate and FOXP3+ Treg subsets showed altered gene expression in preeclampsia (PE) compared to healthy late gestation. The Th1/Th2 intermediate subset exhibited elevated levels of cytotoxicity-related gene expression in PE. Moreover, increased Treg exhaustion was observed in the PE group, and FOXP3+ Treg subcluster analysis revealed that the effector Treg like subset drove the Treg exhaustion signatures in PE. The Th1/Th2 intermediate and effector Treg like subsets are possible inflammation-driving subsets in PE.

Decidual natural killer cells dysfunction is caused by IDO downregulation in dMDSCs with Toxoplasma gondii infection.

Myeloid-derived suppressor cells (MDSCs) play a crucial role in maintaining maternal-fetal tolerance by expressing some immune-suppressive molecules, such as indoleamine 2,3-dioxygenase (IDO). Toxoplasma gondii (T. gondii) infection can break the immune microenvironment of maternal-fetal interface, resulting in adverse pregnancy outcomes. However, whether T. gondii affects IDO expression in dMDSCs and the molecular mechanism of its effect are still unclear. Here we show, the mRNA level of IDO is increased but the protein level decreased in infected dMDSCs. Mechanistically, the upregulation of transcriptional levels of IDO in dMDSCs is regulated through STAT3/p52-RelB pathway and the decrease of IDO expression is due to its degradation caused by increased SOCS3 after T. gondii infection. In vivo, the adverse pregnancy outcomes of IDO-/- infected mice are more severe than those of wide-type infected mice and obviously improved after exogenous kynurenine treatment. Also, the reduction of IDO in dMDSCs induced by T. gondii infection results in the downregulation of TGF-ß and IL-10 expression in dNK cells regulated through Kyn/AhR/SP1 signal pathway, eventually leading to the dysfunction of dNK cells and contributing the occurrence of adverse pregnancy outcomes. This study reveals a novel molecular mechanism in adverse pregnancy outcome induced by T. gondii infection.

Interleukin-1 beta signals through the ERK signalling pathway to modulate human placental trophoblast migration and invasion in the first trimester of pregnancy.

INTRODUCTION: Interleukin-1 beta (IL-1ß) can promote cell migration, invasion and metastasis in various cancer cells. The mechanism of its role in human trophoblast has not been fully investigated. Therefore, we aimed to investigate the expression level of IL-1ß in first trimester decidua and placenta and its potential role in regulation of extravillous trophoblast cell (EVT) invasion and migration. METHODS: First trimester placenta and decidua were collected to study the expression levels of IL-1ß and its receptors by immunohistochemical staining. Primary isolates of first trimester EVT or the HTR-8/SVneo trophoblast like cell line were used to assess migration and invasion. Matrix metalloproteinase levels were assessed by gelatin zymography and ELISA. The phosphorylation profile of signaling pathway proteins was detected with the Proteome Profiler Human Phospho-Kinase Array Kit. Differentially expressed proteins in cells was detected and verified by Western Blot. RESULTS: IL-1ß, its receptors and antagonist are expressed in first trimester placenta and decidua, exogenous IL-1ß stimulates trophoblast cell outgrowth, migration and invasion through the ERK signaling pathway. IL-1ß was significantly increased in the placenta at 6-7 weeks gestation compared with 8-9 weeks gestation (P < 0.0001). Transwell and RTCA assays indicated that IL-1ß stimulates the invasion and migration of EVT. In addition, IL-1ß promoted the phosphorylation of ERK 1/2. It also promoted the expression of MMP2 and MMP9 in EVT as demonstrated by gelatin zymography assay and enzyme linked immunosorbent assay. DISCUSSION: This study demonstrated IL-1ß expression in placenta and decidua, and that it regulates EVT invasion and migration.

Role of maternal-fetal immune tolerance in the establishment and maintenance of pregnancy.

ABSTRACT: Normal pregnancy is a contradictory and complicated physiological process. Although the fetus carries the human leukocyte antigen (HLA) inherited from the paternal line, it does not cause maternal immune rejection. As the only exception to immunological principles, maternal-fetal immune tolerance has been a reproductive immunology focus. In early pregnancy, fetal extravillous trophoblast cells (EVTs) invade decidual tissues and come into direct contact with maternal decidual immune cells (DICs) and decidual stromal cells (DSCs) to establish a sophisticated maternal-fetal crosstalk. This study reviews previous research results and focuses on the establishment and maintenance mechanism of maternal-fetal tolerance based on maternal-fetal crosstalk. Insights into maternal-fetal tolerance will not only improve understanding of normal pregnancy but will also contribute to novel therapeutic strategies for recurrent spontaneous abortion, pre-eclampsia, and premature birth.

Alterations in the pro-resolving lipid mediator machinery within first trimester maternal tissue: Implications in decidualization and miscarriage risk.

A pivotal event in uterine receptivity and human reproduction is the differentiation of endometrial stromal cells into decidual cells, known as decidualization. Decidualization is interlinked with its inflammatory environment. Our study aimed to investigate the presence and role of pro-resolving lipid mediators in first trimester maternal tissue. We assessed the levels of LXA4 and RvD1, along with their metabolic LOX enzymes, in elective (control) and sporadic miscarriage samples. We investigated the effects of LXA4 and RvD1 on decidualization using primary endometrial stromal cells and the immortalized endometrial stromal St-T1b cell line. The upregulation of 12- and 15-LOX expression was observed in pregnancy tissue after sporadic miscarriage, suggesting an inflammatory imbalance. Furthermore, incubation with these lipid mediators led to a decrease in decidualization biomarkers PRL and IGFBP-1, accompanied by morphological changes indicative of aberrant differentiation. The expression of LOX enzymes in decidual natural killer cells suggests their involvement in regulating the inflammatory surroundings and the extent of decidualization.

Tryptophan in the mouse diet is essential for embryo implantation and decidualization.

Introduction: Nutritional deficiency occurs frequently during pregnancy and breastfeeding. Tryptophan (Trp), an essential amino acid which is critical for protein synthesis, serves as the precursor for serotonin, melatonin, and kynurenine (Kyn). The imbalance between serotonin and kynurenine pathways in Trp metabolism is closely related to inflammation and depression. This study assessed the effects of Trp deficiency on mouse early pregnancy. Methods: Embryo implantation and decidualization were analyzed after female mice had been fed diets containing 0.2% Trp (for the control group), 0.062% Trp (for the low Trp group) and 0% Trp (for the Trp-free group) for two months. The uteri of the mice were collected on days 4, 5, and 8 of pregnancy for further analysis. Results: On day 8 of pregnancy, the number of implantation sites were found to be similar between the control and the low Trp groups. However, no implantation sites were detected in the Trp-free group. On day 5 of pregnancy, plane polarity- and decidualization-related molecules showed abnormal expression pattern in the Trp-free group. On day 4 of pregnancy, there was no significant difference in uterine receptivity molecules between the low-Trp group and the control group, but uterine receptivity was abnormal in the Trp-free group. At implantation sites of the Trp-free group, IDO and AHR levels were markedly elevated. This potentially increased levels of Kyn, 2-hydroxy estradiol, and 4-hydroxy estradiol to affect decidualization. Conclusions: Trp-free diet may impair decidualization via the IDO-KYN-AHR pathway.

In vitro mRNA-S maternal vaccination induced altered immune regulation at the maternal-fetal interface.

BACKGROUND: Maternal-fetal immunology is intricate, and the effects of mRNA-S maternal vaccination on immune regulation at the maternal-fetal interface require further investigation. Our study endeavors to elucidate these immunological changes, enhancing our comprehension of maternal and fetal health outcomes. By analyzing immune profiles and cytokine responses, we aim to provide valuable insights into the impact of mRNA-S vaccination on the delicate balance of immune regulation during pregnancy, addressing critical questions in the field of reproductive pharmacology. OBJECTIVES: This investigation sought to examine the prospective influence of mRNA-S-based vaccines and extracellular vesicles (EVs) containing the Spike (S) protein at the maternal-fetal interface. Our primary emphasis was on evaluating their effects on maternal decidua cells and fetal chorion trophoblast cells (hFM-CTCs). METHODS: We validated the generation of EVs containing the S protein from small human airway epithelial cell lines (HSAECs) following mRNA-S vaccine exposure. We assessed the expression of angiotensin-converting enzyme 2 (ACE2) gene and protein in fetal membranes and the placenta, with specific attention to decidual cells and fetal membrane chorion cells. To assess cellular functionality, these cells were exposed to both recombinant S protein and EVs loaded with S proteins (eSPs). RESULTS: Our findings revealed that cells and EVs subjected to mRNA-S-based vaccination exhibited altered protein expression levels of S proteins. At the feto-maternal interface, both placental and fetal membrane tissues demonstrated similar ACE-2 expression levels. Among individual cellular layers, syncytiotrophoblast cells in the placenta and chorion cells in the fetal membrane exhibited elevated ACE-2 expression. Notably, EVs derived from HSAECs activated the MAPK pathway in decidual cells. Additionally, decidual cells displayed a substantial increase in gene expression of chemokines like CXCL-10 and CXCL-11, as well as proinflammatory cytokines such as IL-6 in response to eSPs. However, the levels of Ccl-2 and IL-1ß remained unchanged in decidual cells under the same conditions. Conversely, hFM-CTCs demonstrated significant alterations in the proinflammatory cytokines and chemokines with respect to eSPs. CONCLUSION: In conclusion, our study indicates that mRNA-S-based maternal vaccination during pregnancy may influence the maternal-fetal interface's COVID-19 interaction and immune regulation. Further investigation is warranted to assess safety and implications.

Long noncoding RNA BMPR1B-AS1 stability regulated by IGF2BP2 affects the decidualization in endometriosis patients through the SMAD1/5/9 pathway.

Endometriosis (EMs)-related infertility commonly has decreased endometrial receptivity and normal decidualization is the basis for establishing and maintaining endometrial receptivity. However, the potential molecular regulatory mechanisms of impaired endometrial decidualization in patients with EMs have not been fully clarified. We confirmed the existence of reduced endometrial receptivity in patients with EMs by scanning electron microscopy and quantitative real-time PCR. Here we identified an lncRNA, named BMPR1B-AS1, which is significantly downregulated in eutopic endometrium in EMs patients and plays an essential role in decidual formation. Furthermore, RNA pull-down, mass spectrometry, RNA immunoprecipitation, and rescue analyses revealed that BMPR1B-AS1 positively regulates decidual formation through interaction with the RNA-binding protein insulin-like growth factor 2 mRNA-binding protein 2 (IGF2BP2). Downregulation of IGF2BP2 led to a decreased stability of BMPR1B-AS1 and inhibition of activation of the SMAD1/5/9 pathway, an inhibitory effect which diminished decidualization in human endometrial stromal cells (hESCs) decidualization. In conclusion, our identified a novel regulatory mechanism in which the IGF2BP2-BMPR1B-AS1-SMAD1/5/9 axis plays a key role in the regulation of decidualization, providing insights into the potential link between abnormal decidualization and infertility in patients with EMs, which will be of clinical significance for the management and treatment of infertility in patients with EMs.

Long-chain acyl-CoA synthetase-4 regulates endometrial decidualization through a fatty acid ß-oxidation pathway rather than lipid droplet accumulation.

OBJECTIVE: Lipid metabolism plays an important role in early pregnancy, but its effects on decidualization are poorly understood. Fatty acids (FAs) must be esterified by fatty acyl-CoA synthetases to form biologically active acyl-CoA in order to enter the anabolic and/or catabolic pathway. Long-chain acyl-CoA synthetase 4 (ACSL4) is associated with female reproduction. However, whether it is involved in decidualization is unknown. METHODS: The expression of ACSL4 in human and mouse endometrium was detected by immunohistochemistry. ACSL4 levels were regulated by the overexpression of ACSL4 plasmid or ACSL4 siRNA, and the effects of ACSL4 on decidualization markers and morphology of endometrial stromal cells (ESCs) were clarified. A pregnant mouse model was established to determine the effect of ACSL4 on the implantation efficiency of mouse embryos. Modulation of ACSL4 detects lipid anabolism and catabolism. RESULTS: Through examining the expression level of ACSL4 in human endometrial tissues during proliferative and secretory phases, we found that ACSL4 was highly expressed during the secretory phase. Knockdown of ACSL4 suppressed decidualization and inhibited the mesenchymal-to-epithelial transition induced by MPA and db-cAMP in ESCs. Further, the knockdown of ACSL4 reduced the efficiency of embryo implantation in pregnant mice. Downregulation of ACSL4 inhibited FA ß-oxidation and lipid droplet accumulation during decidualization. Interestingly, pharmacological and genetic inhibition of lipid droplet synthesis did not affect FA ß-oxidation and decidualization, while the pharmacological and genetic inhibition of FA ß-oxidation increased lipid droplet accumulation and inhibited decidualization. In addition, inhibition of ß-oxidation was found to attenuate the promotion of decidualization by the upregulation of ACSL4. The decidualization damage caused by ACSL4 knockdown could be reversed by activating ß-oxidation. CONCLUSIONS: Our findings suggest that ACSL4 promotes endometrial decidualization by activating the ß-oxidation pathway. This study provides interesting insights into our understanding of the mechanisms regulating lipid metabolism during decidualization.

Single-cell transcriptomic analysis of decidual immune cell landscape in the occurrence of adverse pregnancy outcomes induced by Toxoplasma gondii infection.

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that can lead to adverse pregnancy outcomes, particularly in early pregnancy. Previous studies have illustrated the landscape of decidual immune cells. However, the landscape of decidual immune cells in the maternal-fetal microenvironment during T. gondii infection remains unknown. METHODS: In this study, we employed single-cell RNA sequencing to analyze the changes in human decidual immune cells following T. gondii infection. The results of scRNA-seq were further validated with flow cytometry, reverse transcription-polymerase chain reaction, western blot, and immunofluorescence staining. RESULTS: Our results showed that the proportion of 17 decidual immune cell clusters and the expression levels of 21 genes were changed after T. gondii infection. Differential gene analysis demonstrated that T. gondii infection induced the differential expression of 279, 312, and 380 genes in decidual NK cells (dNK), decidual macrophages (dMφ), and decidual T cells (dT), respectively. Our results revealed for the first time that several previously unknown molecules in decidual immune cells changed following infection. This result revealed that the function of maternal-fetal immune tolerance declined, whereas the killing ability of decidual immune cells enhanced, eventually contributing to the occurrence of adverse pregnancy outcomes. CONCLUSIONS: This study provides valuable resource for uncovering several novel molecules that play an important role in the occurrence of abnormal pregnancy outcomes induced by T. gondii infection.

Deficiency of UCHL1 results in insufficient decidualization accompanied by impaired dNK modulation and eventually miscarriage.

BACKGROUND: Miscarriage is a frustrating complication of pregnancy that is common among women of reproductive age. Insufficient decidualization which not only impairs embryo implantation but disturbs fetomaternal immune-tolerance, has been widely regarded as a major cause of miscarriage; however, the underlying mechanisms resulting in decidual impairment are largely unknown. METHODS: With informed consent, decidual tissue from patients with spontaneous abortion or normal pregnant women was collected to detect the expression profile of UCHL1. Human endometrial stromal cells (HESCs) were used to explore the roles of UCHL1 in decidualization and dNK modulation, as well as the mechanisms involved. C57/BL6 female mice (7-10 weeks old) were used to construct pregnancy model or artificially induced decidualization model to evaluate the effect of UCHL1 on mice decidualization and pregnancy outcome. RESULTS: The Ubiquitin C-terminal hydrolase L1 (UCHL1), as a deubiquitinating enzyme, was significantly downregulated in decidua from patients with miscarriage, along with impaired decidualization and decreased dNKs. Blockage of UCHL1 led to insufficient decidualization and resultant decreased expression of cytokines CXCL12, IL-15, TGF-ß which were critical for generation of decidual NK cells (dNKs), whereas UCHL1 overexpression enhanced decidualization accompanied by increase in dNKs. Mechanistically, the promotion of UCHL1 on decidualization was dependent on its deubiquitinating activity, and intervention of UCHL1 inhibited the activation of JAK2/STAT3 signaling pathway, resulting in aberrant decidualization and decreased production of cytokines associated with dNKs modulation. Furthermore, we found that inhibition of UCHL1 also disrupted the decidualization in mice and eventually caused adverse pregnancy outcome. CONCLUSIONS: UCHL1 plays significant roles in decidualization and dNKs modulation during pregnancy in both humans and mice. Its deficiency indicates a poor pregnancy outcome due to defective decidualization, making UCHL1 a potential target for the diagnosis and treatment of miscarriage.

Reproductive Immunology and Pregnancy 2.0.

This Special Issue comprises original articles in the field of clinical studies whose major topics concern the genetic and immunological aspects of miscarriage and pre-eclampsia, the isolation of decidua macrophages and Hofbauer cells in the placenta for diagnostic purposes, and epigenetic mechanisms that trigger labor [...].

NEK2 promotes the development of ovarian endometriosis and impairs decidualization by phosphorylating FOXO1.

Ovarian endometriosis is a common gynecological disease, and one of its most significant symptoms is infertility. In patients with endometriosis, defects in endometrial decidualization lead to impaired endometrial receptivity and embryo implantation, thus affecting early pregnancy and women's desire to have children. However, the mechanisms underlying the development of endometriosis and its associated defective decidualization are unclear. We find that NEK2 expression is increased in the ectopic and eutopic endometrium of patients with endometriosis. Meanwhile, NEK2 interacts with FOXO1 and phosphorylates FOXO1 at Ser184, inhibiting the stability of the FOXO1 protein. Importantly, NEK2-mediated phosphorylation of FOXO1 at Ser184 promotes cell proliferation, migration, invasion and impairs decidualization. Furthermore, INH1, an inhibitor of NEK2, inhibits the growth of ectopic lesions in mouse models of endometriosis and promotes endometrial decidualization in mouse models of artificially induced decidualization. Taken together, these findings indicate that NEK2 regulates the development of endometriosis and associated disorders of decidualization through the phosphorylation of FOXO1, providing a new therapeutic target for its treatment.