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Problem: Recurrent pregnancy loss (RPL) is defined as the failure of two or more pregnancies and affects approximately 5% of couples, often without a clear cause. The etiologies of RPL include factors such as maternal age, endocrine dysfunction, uterine abnormalities, chromosomal abnormalities, thrombophilias, infections, and autoimmune disorders. However, these conditions account for only 50%–60% of RPL cases. Research has explored whether an altered immune system, compared to the physiological state, may be linked to RPL. This review aims to determine whether specific immunophenotypes are associated with unexplained Recurrent Pregnancy Loss (uRPL) and whether targeted therapies addressing specific immunophenotypic alterations can improve pregnancy outcomes. Methods: A literature review was conducted using Pubmed/Medline, Scopus, and Embase databases, analyzing data from 95 articles published between 2001 and 2023. The roles of various cells of the immune system (B lymphocytes, T lymphocytes, natural killer cells, macrophages) in different tissues (peripheral blood, menstrual blood) were specifically investigated in women with uRPL. Discussion and Conclusion: Specific immunophenotypes have been demonstrated to be associated with this condition. However, there is a need to standardize immunophenotyping assays and conduct more trials to stratify RPL risk and improve potential therapeutic strategies. [ABSTRACT FROM AUTHOR]

Yanyu Zhong,1,&ast; Nan Wang,1,&ast; Sihui Lu,2 Yaqian Lu,2 Xin Pan,3 Ying Zhou1 1Reproductive Medicine Centre, The First Affiliated Hospital of Soochow University, Suzhou, People’s Republic of China; 2Department of Obstetrics and Gynecology, The Fourth Affiliated Hospital of Soochow University, Suzhou, People’s Republic of China; 3Department of Obstetrics and Gynecology, The First Affiliated Hospital of Soochow University, Suzhou, People’s Republic of China&ast;These authors contributed equally to this workCorrespondence: Ying Zhou, Reproductive Medicine Centre, The First Affiliated Hospital of Soochow University, No. 899 Pinghai Road, Gusu District, Suzhou, Jiangsu Province, 215008, People’s Republic of China, Tel +86-0512-67973182, Email zhouyingzzzyyy@163.com Xin Pan, Department of Obstetrics and Gynecology, The First Affiliated Hospital of Soochow University, No. 899 Pinghai Road, Gusu District, Suzhou, Jiangsu Province, 215008, People’s Republic of China, Tel +86-0512-67973182, Email panxinxin9@outlook.comObjective: This study aimed to analyze uterine artery and spiral artery hemodynamics in patients with unexplained recurrent pregnancy loss (URPL) with varying pregnancy outcomes.Methods: 174 pregnant women with URPL and 144 pregnant women without adverse pregnancy histories were enrolled in this retrospective study. Based on pregnancy outcomes, these patients were divided into two groups: normal pregnancy outcomes (URPL-N, n=138) and adverse pregnancy outcomes (URPL-A, n=36). Control group participants were categorized into normal pregnancy outcomes (CON-N, n=129) and adverse pregnancy outcomes (CON-A, n=15). We compared uterine artery and spiral artery hemodynamics during different stages of gestation and the predictive value of these parameters for pregnancy outcomes.Results: URPL-N group had fewer pregnancy losses and lower BMI compared to URPL-A group (P< 0.05). Spiral artery hemodynamics in URPL-N and CON-N groups were lower than those in URPL-A and CON-A groups during the mid-luteal phase, 11– 13 weeks, 15– 17 weeks, and 19– 21 weeks of gestation, respectively. Uterine artery hemodynamics ((Pulsatility index (mPI), resistive index (mRI), and systolic-to-diastolic ratio (mS/D)) in the mid-luteal period were lower in URPL-N group than URPL-A group. Similarly, in CON-N group were lower than CON-A group. The URPL-A and CON-A groups had higher uterine artery and spiral artery hemodynamics when compared to the URPL-N and CON-N groups. Spiral artery hemodynamics exhibited larger areas under the ROC curve compared to uterine artery parameters.Conclusion: Abnormal hemodynamics in these arteries may contribute to URPL and adverse pregnancy outcomes. Spiral artery hemodynamics are more reliable predictors of pregnancy outcomes than uterine artery parameters.Keywords: unexplained recurrent pregnancy loss, spiral artery, uterine artery, blood flow, pregnancy outcome

With unknown etiology and limited treatment options, unexplained recurrent pregnancy loss (URPL) remains a thorny problem. Ferroptosis, a newly identified type of cell death, has been shown to be crucial in the development in reproductive disorders. This study aims to explore the specific mechanism of ferroptosis in URPL and to uncover whether alpha-lipoic acid (ALA) can inhibit ferroptosis, and then exert a protective effect in URPL.

Background Methods Findings Interpretation Understanding the pathogenesis of unexplained recurrent pregnancy loss is paramount for advancing effective treatments. Various biological processes, including spermatogenesis and embryo development, are tightly regulated by N6‐methyladenosine modifications. However, few studies have focused on the impact of sperm N6‐methyladenosine modifications on embryonic development. Therefore, we aimed to study altered N6‐methyladenosine‐mediated messenger RNA methylation modifications in the spermatozoa of male partners from couples experiencing unexplained recurrent pregnancy loss, to identify potential diagnostic markers and explore their potential molecular mechanisms in pregnancy loss and embryogenesis.Methylated RNA immunoprecipitation (MeRIP) sequencing and RNA sequencing were conducted on the spermatozoa of men from couples in the ‘unexplained recurrent pregnancy loss’ group (n = 6), and the fertility control group (n = 6). To identify the role of the detected key genes, zebrafish model embryos were studied, and multi‐omics (transcriptomics, proteomics, and metabolomics) analyses helped to explore the molecular mechanism of abnormal embryogenesis.Comparing unexplained recurrent pregnancy loss with the fertility control group, 217 N6‐methyladenosine peaks were significantly upregulated, and 40 were downregulated in the spermatozoa. The combined analyses of spermatozoa‐methylated RNA immunoprecipitation sequencing and RNA sequencing indicated that N6‐methyladenosine methylation and the expression of SEMA5A, MT‐ATP6, ZNF662, and KDM4C were significantly different. In zebrafish embryos, the altered expression of the four genes increased embryonic mortality and malformations by disturbing several key signaling pathways and zygotic genome activation.This study highlights the paternal epigenome, which could be one of the reasons for faulty embryogenesis leading to pregnancy loss. The N6‐methyladenosine modification, the most prevalent RNA modification, contributes to the exploration and understanding of the paternal epigenome in the maintenance of pregnancy and fetal growth and development. The four genes identified in this study may serve as potential diagnostic markers and elucidate novel molecular mechanisms of embryogenesis. [ABSTRACT FROM AUTHOR]

Introduction: Unexplained recurrent pregnancy loss (URPL), affecting approximately 1%–5% of women, exhibits a strong association with various maternal factors, particularly immune disorders. However, accurately predicting pregnancy outcomes based on the complex interactions and synergistic effects of various immune parameters without an automated algorithm remains challenging. Material and methods: In this historical cohort study, we analyzed the medical records of URPL patients treated at Xiangya Hospital, Changsha, China, between January 2020 and October 2022. The primary outcomes included clinical pregnancy and miscarriage. Predictors included complement, autoantibodies, peripheral lymphocytes, immunoglobulins, thromboelastography findings, and serum lipids. Least absolute shrinkage and selection operator (LASSO) analysis and logistic regression analysis was performed for model development. The model's performance, discriminatory, and clinical applicability were assessed using area under the curve (AUC), calibration curve, and decision curve analysis, respectively. Additionally, models were visualized by constructing dynamic and static nomograms. Results: In total, 502 patients with URPL were enrolled, of whom 291 (58%) achieved clinical pregnancy and 211 (42%) experienced miscarriage. Notable differences in complement, peripheral lymphocytes, and serum lipids were observed between the two outcome groups. Moreover, URPL patients with elevated peripheral NK cells (absolute counts and proportion), decreased complement levels, and dyslipidemia demonstrated a significantly increased risk of miscarriage. Four models were developed in this study, of which Model 2 demonstrated superior performance with only seven predictors, achieving an AUC of 0.96 (95% CI: 0.93–0.99) and an accuracy of 0.92. A web‐based platform was established to visually present model 2 and to facilitate its utilization by clinicians in outpatient settings (available from: https://yingrongli.shinyapps.io/liyingrong/). Conclusions: Our findings suggest that the implementation of such prediction models could serve as valuable tools for providing comprehensive information and facilitating clinicians in their decision‐making processes. [ABSTRACT FROM AUTHOR]

The majority of unexplained recurrent pregnancy loss (URPL) cases have been attributed to immune abnormalities. Inappropriate changes in microbiota could lead to immune disorders. However, the specific role of uterine cavity microbiota in URPL remains unclear, and only a limited number of related studies are available for reference. We utilized double-lumen embryo transfer tubes to collect uterine cavity fluid samples from pregnant women in their first trimester. Subsequently, we conducted 16S rRNA sequencing to analyze the composition and abundance of the microbiota in these samples. For this study, we enlisted 10 cases of URPL and 28 cases of induced miscarriages during early pregnancy. Microbial communities were detected in all samples of the URPL group (100 %, n = 10), whereas none were found in the control group (0 %, n = 28). Among the identified microbes, Lactobacillus and Curvibacter were the two most dominant species. The abundance of Curvibacter is correlated with the number of NK cells in peripheral blood (r = −0.759, P = 0.018). This study revealed that during early pregnancy, Lactobacillus and Curvibacter were the predominant colonizers in the uterine cavity of URPL patients and were associated with URPL. Consequently, alterations in the dominant microbiota may lead to adverse pregnancy outcomes. • Most unexplained URPL cases have been attributed to immune abnormalities. • Inappropriate changes in microbiota could lead to immune disorders. • Lactobacillus and Curvibacter were predominant in URPL patients' uterine cavity. • Alterations in the dominant microbiota may lead to adverse pregnancy outcomes. [ABSTRACT FROM AUTHOR]

Abstract Objective Recurrent pregnancy loss (RPL) patients have higher absolute numbers of decidual natural killer (dNK) cells with elevated intracellular IFN-γ levels leading to a pro-inflammatory cytokine milieu, which contributes to RPL pathogenesis. The main objective of this study was twofold: first to explore the regulatory effects and mechanisms of villus-derived exosomes (vEXOs) from induced abortion patients or RPL patients at the level of intracellular IFN-γ in dNK cells; second to determine the validity of application of vEXOs in the treatment of unexplained RPL (uRPL) through in vitro experiments and mouse models. Methods Exosomes were isolated from villus explants by ultracentrifugation, co-cultured with dNK cells, and purified by enzymatic digestion and magnetically activated cell sorting. Flow cytometry, enzyme-linked immunosorbent assays, and RT-qPCR were used to determine IFN-γ levels. Comparative miRNA analysis of vEXOs from induced abortion (IA) and uRPL patients was used to screen potential candidates involved in dNK regulation, which was further confirmed by luciferase reporter assays. IA-vEXOs were electroporated with therapeutic miRNAs and encapsulated in a China Food and Drug Administration (CFDA)-approved hyaluronate gel (HA-Gel), which has been used as a clinical biomaterial in cell therapy for > 30 years. In vivo tracking was performed using 1,1-dioctadecyl-3,3,3,3-tetramethylindotricarbocyaine iodide (DiR) labelling. Tail-vein and uterine horn injections were used to evaluate therapeutic effects of the engineered exosomes in an abortion-prone mouse model (CBA/J × DBA/2 J). Placental growth was evaluated based on placental weight. IFN-γ mRNA levels in mouse placentas were measured by RT-qPCR. Results IFN-γ levels were significantly higher in dNK cells of uRPL patients than in IA patients. Both uRPL-vEXOs and IA-vEXOs could be efficiently internalized by dNK cells, whereas uRPL-vEXOs could not reduce the expression of IFN-γ by dNK cells as much as IA-vEXOs. Mechanistically, miR-29a-3p was delivered by vEXOs to inhibit IFN-γ production by binding to the 3′ UTR of IFN-γ mRNA in dNK cells. For in vivo treatment, application of the HA-Gel effectively prolonged the residence time of vEXOs in the uterine cavity via sustained release. Engineered vEXOs loaded with miR-29a-3p reduced the embryo resorption rate in RPL mice with no signs of systemic toxicity. Conclusion Our study provides the first evidence that villi can regulate dNK cell production of IFN-γ via exosome-mediated transfer of miR-29a-3p, which deepens our understanding of maternal–fetal immune tolerance for pregnancy maintenance. Based on this, we developed a new strategy to mix engineered vEXOs with HA-Gel, which exhibited good therapeutic effects in mice with uRPL and could be used for potential clinical applications in uRPL treatment. Graphical Abstract

Purpose Previous studies have reported the involvement of long noncoding RNAs (lncRNAs) in reproductive diseases via the regulation of target genes. This study aimed to determine whether lnc-prostate androgen-regulated transcript 1 (lnc-PART1)could be used as a biomarker of unexplained recurrent pregnancy loss (URPL) and a possible predictor of poor pregnancy outcomes in women with URPL.Materials and methods Sixty patients with URPL and 15 healthy women were included in this study. PART1 expression was detected in plasma and endometrial tissues using a quantitative reverse transcription polymerase chain reaction. Logistic regression and receiver operating characteristic curve analyses were performed to analyze the association between PART1 expression and pregnancy outcomes in women with URPL.Results The expression of PART1transcript variant 2 was significantly up-regulated in the endometrial specimens from patients with URPL compared to control tissues. High tissue expression levels of PART1transcript variant 2 were associated with poor pregnancy outcomes in women with URPL, indicating that it could serve as a potential risk factor. Additionally, PART1 could serve as a potential risk factor for adverse pregnancy outcomes in patients with URPL (OR = 4.374; 95% CI = 1.052–18.189; p = .042).Conclusion lncRNA PART1 transcript variant 2 was highly expressed in patients with URPL. Therefore, it is important to conduct in-depth studies on the relationship between PART1 expression and URPL.

Abstract Throughout pregnancy, the decidua is predominantly populated by NK lymphocytes expressing Killer immunoglobulin-like receptors (KIR) that recognize human leukocyte antigen-C (HLA-C) ligands from trophoblast cells. This study aims to investigate the association of KIR-HLA-C phenotypes in couples facing infertility, particularly recurrent pregnancy loss (RPL) and recurrent implantation failure (RIF), in comparison to a reference population and fertile controls. This observational, non-interventional retrospective case–control study included patients consecutively referred to our Reproductive Immunology Unit from 2015 to 2019. We analyzed the frequencies of KIR and HLA-C genes. As control groups, we analyzed a reference Spanish population for KIR analysis and 29 fertile controls and their male partners for KIR and HLA-C combinations. We studied 397 consecutively referred women with infertility and their male partners. Among women with unexplained RPL (133 women) and RIF (176 women), the centromeric (cen)AA KIR genotype was significantly more prevalent compared to the reference Spanish population (p = 0.001 and 0.02, respectively). Furthermore, cenAA was associated with a 1.51-fold risk of RPL and a 1.2-fold risk of RIF. Conversely, the presence of BB KIR showed a lower risk of reproductive failure compared to non-BB KIR (OR: 0.12, p

Throughout pregnancy, the decidua is predominantly populated by NK lymphocytes expressing Killer immunoglobulin-like receptors (KIR) that recognize human leukocyte antigen-C (HLA-C) ligands from trophoblast cells. This study aims to investigate the association of KIR-HLA-C phenotypes in couples facing infertility, particularly recurrent pregnancy loss (RPL) and recurrent implantation failure (RIF), in comparison to a reference population and fertile controls. This observational, non-interventional retrospective case–control study included patients consecutively referred to our Reproductive Immunology Unit from 2015 to 2019. We analyzed the frequencies of KIR and HLA-C genes. As control groups, we analyzed a reference Spanish population for KIR analysis and 29 fertile controls and their male partners for KIR and HLA-C combinations. We studied 397 consecutively referred women with infertility and their male partners. Among women with unexplained RPL (133 women) and RIF (176 women), the centromeric (cen)AA KIR genotype was significantly more prevalent compared to the reference Spanish population (p = 0.001 and 0.02, respectively). Furthermore, cenAA was associated with a 1.51-fold risk of RPL and a 1.2-fold risk of RIF. Conversely, the presence of BB KIR showed a lower risk of reproductive failure compared to non-BB KIR (OR: 0.12, p < 0.001). Women and their partners with HLA-C1C1/C1C1 were significantly less common in the RPL-Group (p < 0.001) and RIF-Group (p = 0.002) compared to the control group. Moreover, the combination of cenAA/C1C1 in women with C1C1 partners was significantly higher in the control group than in the RPL (p = 0.009) and RIF (p = 0.04) groups, associated with a 5-fold increase in successful pregnancy outcomes. In our cohort, the cenAA KIR haplotype proved to be a more accurate biomarker than the classic AA KIR haplotype for assessing the risk of RPL and RIF, and might be particularly useful to identify women at increased risk among the heterogeneous KIR AB or Bx population. The classification of centromeric KIR haplotypes outperforms classical KIR haplotypes, making it a better indicator of potential maternal–fetal KIR-HLA-C mismatch in patients. [ABSTRACT FROM AUTHOR]

Purpose: The goal of this study is to determine whether any balanced translocation (BT) had been missed by previous karyotyping in patients with unexplained recurrent pregnancy loss (uRPL). Methods: This case series included 48 uRPL-affected couples with normal karyotypes. The embryos from these couples have all undergone preimplantation testing for aneuploidies (PGT-A). Based on the PGT-A's results, 48 couples could be categorized into two groups: 17 couples whose multiple embryos were detected with similar structural variations (SVs, segmental/complete) and 31 couples without such findings but who did not develop any euploid embryo despite at least three high-quality blastocysts being tested. The peripheral blood sample of each partner was then collected for mate-pair sequencing (MPseq) to determine whether any of them were BT carriers. Results: MPseq analyses identified 13 BTs in the 17 couples whose multiple embryos had similar SVs detected (13/17, 76.47%) and three BTs in the 31 couples without euploid embryo obtained (3/31, 9.7%). Among the 16 MPseq-identified BTs, six were missed due to the limited resolution of G-banding karyotyping analysis, and the rest were mostly owing to the similar banding patterns and/or comparable sizes shared by the two segments exchanged. Conclusion: A normal karyotype does not eliminate the possibility of carrying BT for couples with uRPL. The use of PGT-A allows us to perceive the "carrier couples" missed by karyotyping analysis, providing an increased risk of finding cryptic BTs if similar SVs are always detected on two chromosomes among multiple embryos. Nonetheless, certain carriers with translocated segments of sub-resolution may still go unnoticed. [ABSTRACT FROM AUTHOR]

Problem: Recurrent pregnancy loss (RPL) affects up to 4% of couples attempting to conceive. RPL is unexplained in over 50% of cases and no effective treatments exist. Due to the immune system's pivotal role during implantation and pregnancy, immune‐mediated RPL may be suspected and immunomodulatory treatments like intravenous immunoglobulin (IVIg) have been administered but remain controversial. The goal of our study was to evaluate our center's 6 year‐outcomes and to develop a framework for IVIg use in RPL. Method of the study: Retrospective, single‐center cohort study. All patients having received IVIg for unexplained RPL at the McGill Reproductive Immunology Clinic (MRIC) from January 2014 to December 2020 were included if maternal age was <42 years, body mass index (BMI) < 35 kg/m2, non‐smoker and having had ≥3 consecutive RPL despite previous treatment with aspirin and progesterone. IVIg 0.6–0.8 g/kg was given prior to conception and monthly during pregnancy until 16–20 weeks' gestation. We compared IVIg treated patient's outcomes to a separate "natural history cohort". This cohort was composed of patients consulting at the McGill recurrent pregnancy loss clinic and the MRIC over a 2‐year period (January 2020 to December 2021) with similar inclusion criteria as the treatment cohort but did not receive IVIg or other immunomodulatory treatments. The association of IVIg with outcomes (compared to no IVIg) was evaluated among the groups of patients with primary RPL and secondary RPL. The primary outcome was live birth rate (LBR), secondary outcomes included IVIg safety, obstetrical, and neonatal complications. Results: Among 169 patients with unexplained RPL that were included in the study, 111 had primary RPL (38 exposed to IVIg and 83 controls) and 58 had secondary RPL (nine exposed to IVIG and 49 controls). Among patients with primary RPL (n = 111), the LBR was 64.3% (18/28) among patient exposed to IVIg compared to 43.4% (36/83) in controls (p = 0.079); regression analysis adjusting for BMI and number of previous miscarriages showed benefit favoring the use of IVIg (OR = 3.27, CI 95% (1.15–10.2), p = 0.03) when evaluating for live birth. In the subgroup of patients with ≥5 previous RPL and primary RPL (n = 31), IVIg was associated with higher LBR compared to control (10/15 (66.7%) vs. 3/16 (18.8%); p = 0.0113) but not the in the sub‐group of patients with <5 miscarriages and primary RPL (8/13 (61.5%) vs. 33/67 (49.3%); p = 0.548). IVIG treatment did not improve LBR in patients with secondary RPL in our study (3/9 (33.3%) vs. 23/49 (47%); p = 0.495). There were no serious adverse events in the IVIg treatment group, obstetrical/neonatal complications were similar between groups. Conclusion: IVIg may be an effective treatment for patients with RPL if appropriately used in specific groups of patients. IVIg is a blood product and subject to shortages especially with unrestricted off‐label use. We propose considering IVIg in well‐selected patients with high order RPL who have failed standard medical therapy. Further mechanistic studies are needed to understand immune‐mediated RPL and IVIg's mode of action. This will enable further refinement of treatment criteria and the development of standardized protocol for its use in RPL. [ABSTRACT FROM AUTHOR]

Objective: Changes of platelet distribution width (PDW) have been observed in women with unexplained recurrent pregnancy loss (URPL). However, relevant previous studies showed inconsistent results. We performed a meta-analysis to comprehensively evaluate the association between PDW and URPL. Methods: Observational studies reporting the difference of PDW between women with and without URPL were retrieved by searching PubMed, Embase, Web of Science, Wanfang, and CNKI. A random-effect model was used to pool the results by incorporating potential heterogeneity. Results: Eleven case-control studies with 1847 women with URPL and 2475 healthy women were included. Age was matched for all studies between cases and controls. Pooled results showed women with URPL had significantly increased level of PDW (mean difference [MD]: 1.54%, 95% CI: 1.04 to 2.03, p < 0.05; I2 =77%). Subgroup analysis showed consistent result for URPL defined as ≥ 2 (MD: 1.45%, p = 0.003) and ≥ 3 (MD: 1.61%, p < 0.001) failed clinical pregnancies, and in studies compared to women with normal pregnancy (MD: 2.02%, p < 0.001) and non-pregnant healthy women (MD: 1.34%, p < 0.001). Results of the meta-analysis also showed that increment of PDW was associated with higher odds of URPL (per 1 unit, odds ratio: 1.26, 95% CI: 1.17 to 1.35, p < 0.001; I2 = 0%). Conclusions: Women with URPL had significantly increased levels of PDW compared to healthy women without URPL, which suggest that higher PDW may predict the risk of URPL. [ABSTRACT FROM AUTHOR]

Nhat Nguyen Ngoc,1 My Tran Ngoc Thao,2 Sang Trieu Tien,3 Son Vu Tung,4 Hoang Le,5 Hung Ho Sy,6 Tung Nguyen Thanh,1 Son Trinh The1 1Military Institute of Clinical Embryology and Histology, Vietnam Military Medical University, Hanoi, 12108, Vietnam; 2Département de formation Biologie moléculaire et cellulaire, Sorbonne University, Paris, 75006, France; 3Department of Biology and Genetics, Vietnam Military Medical University, Hanoi, 12108, Vietnam; 4Department of Epidemiology, Vietnam Military Medical University, Hanoi, 12108, Vietnam; 5IVFTA, Tam Anh General Hospital, Hanoi, 100000, Vietnam; 6Department of Obstetrics and Gynecology, Hanoi Medical University, Hanoi, 100000, VietnamCorrespondence: Son Trinh The; Tung Nguyen Thanh, Military Institute of Clinical Embryology and Histology, Vietnam Military Medical University, Hanoi, 12108, Vietnam, Email trinhtheson@vmmu.edu.vn; nguyenthanhtung@vmmu.edu.vnObjective: To investigate the relationship between unexplained recurrent pregnancy loss (URPL) and polymorphisms of homocysteine metabolism-related genes in women.Materials and Methods: A case–control study included 90 women with two or more consecutive unexplained pregnancy losses and 92 controlled women without miscarriage history; the female participants were in the age category of 18– 35 years. The high-resolution melting technique was used to detect the single-nucleotide variants related to homocysteine metabolism disorder, namely MTHFR C677T, MTHFR A1298C, MTR A2756G, and MTRR A66G polymorphism.Results: The MTHFR C677T polymorphism had significantly correlation with URPL. Indeed, the frequency of the677T allele and genotypes (677CT, 677TT) in the URPL group was significantly higher than that in the control group (p < 0.05). However, the allele, as well as genotype distribution of MTHFR A1298C, MTR A2756G, and MTRR A66G polymorphisms showed no significant difference (p > 0.05). MTHFR 677CT-1298AC genotype combination led to a 9.0-fold increased risk of URPL (OR 9.0; 95% CI, 2.25– 35.99; p = 0.001), while the risk increased 10.0-fold (OR 10.0; 95% CI, 1.8– 55.53; p = 0.008) when participants had more than the 3 variant loci.Conclusion: The MTHFR C677T polymorphism was a risk factor for URPL, and determining the MTHFR C677T polymorphism had a potential prediction of URPL risk. Moreover, the MTHFR C677T and MTHFR A1298C joint mutants might have a synergistic effect on URPL. Conversely, there is a lack of evidence suggesting the URPL risk of MTHFR A1298C, MTR A2756G, and MTRR A66G polymorphisms.Keywords: unexplained recurrent pregnancy loss, homocysteine, MTHFR C677T, MTHFR A1298C, MTR A2756G, MTRR A66G

Recurrent pregnancy loss (RPL) puzzles 1–3% of women of childbearing age worldwide. Immunological factors account for more than 60% of cases of unexplained RPL (URPL); however, the underlying mechanism remains unclear. Here, using single-cell sequencing data and functional experiments with clinical samples, we identified a distinct population of CCR1+ decidual macrophages (dMφ) that were preferentially enriched in the decidua from normal early pregnancies but were substantially decreased in patients with URPL. Specific gene signatures endowed CCR1+ dMφ with immunosuppressive and migration-regulatory properties, which were attenuated in URPL. Additionally, CCR1+ dMφ promoted epithelial-to-mesenchymal transition (EMT) to promote trophoblast migration and invasion by activating the ERK1/2 signaling pathway. Decidual stromal cell (DSC)-derived CCL8 was the key regulator of CCR1+ dMφ as CCL8 recruited peripheral CCR1+ monocytes, induced a CCR1+ dMφ-like phenotype, and reinforced the CCR1+ dMφ-exerted modulation of trophoblasts. In patients with URPL, CCL8 expression in DSCs was decreased and trophoblast EMT was defective. Our findings revealed that CCR1+ dMφ play an important role in immune tolerance and trophoblast functions at the maternal–fetal interface. Additionally, decreased quantity and dysregulated function of CCR1+ dMφ result in URPL. In conclusion, we provide insights into the crosstalk between CCR1+ dMφ, trophoblasts, and DSCs at the maternal–fetal interface and macrophage-targeted interventions of URPL. [ABSTRACT FROM AUTHOR]

Objective: To explore the roles of advanced paternal age (APA) and abnormal paternal weight on embryo quality and pregnancy outcomes for unexplained recurrent pregnancy loss (uRPL) couples who underwent preimplantation genetic testing for aneuploidies (PGT-A). Methods: This study included 779 uRPL couples who underwent their first PGT-A cycles between 2014 and 2018. Male patients’ aging and nutritional status were quantified by paternal age and body mass index (BMI). Routine semen parameters and sperm DNA fragmentation index (DFI) were used to reflect the seminal quality. Blastocyst formation rate and aneuploidy rate were used to reflect the embryo quality. Cycle cancellation rate, implantation rate, pregnancy loss rate, and live birth rate were measured to evaluate the treatment efficiency from IVF. To remove the interference of maternal age, only the women younger than 38 years old were included. After univariate screening, interaction tests were performed in a generalized linear model (GLM) to further examine the effects of paternal age and BMI on each outcome indicator. Results: In the total population (779 cycles), there were no statistical differences in aneuploidy rate, cycle cancellation rate, implantation rate, pregnancy loss rate, and live birth rate, whether stratified by paternal age or paternal BMI. Similar results occurred in the younger men (

Recurrent pregnancy loss (RPL) puzzles 1–3% of women of childbearing age worldwide. Immunological factors account for more than 60% of cases of unexplained RPL (URPL); however, the underlying mechanism remains unclear. Here, using single-cell sequencing data and functional experiments with clinical samples, we identified a distinct population of CCR1+ decidual macrophages (dMφ) that were preferentially enriched in the decidua from normal early pregnancies but were substantially decreased in patients with URPL. Specific gene signatures endowed CCR1+ dMφ with immunosuppressive and migration-regulatory properties, which were attenuated in URPL. Additionally, CCR1+ dMφ promoted epithelial-to-mesenchymal transition (EMT) to promote trophoblast migration and invasion by activating the ERK1/2 signaling pathway. Decidual stromal cell (DSC)-derived CCL8 was the key regulator of CCR1+ dMφ as CCL8 recruited peripheral CCR1+ monocytes, induced a CCR1+ dMφ-like phenotype, and reinforced the CCR1+ dMφ-exerted modulation of trophoblasts. In patients with URPL, CCL8 expression in DSCs was decreased and trophoblast EMT was defective. Our findings revealed that CCR1+ dMφ play an important role in immune tolerance and trophoblast functions at the maternal–fetal interface. Additionally, decreased quantity and dysregulated function of CCR1+ dMφ result in URPL. In conclusion, we provide insights into the crosstalk between CCR1+ dMφ, trophoblasts, and DSCs at the maternal–fetal interface and macrophage-targeted interventions of URPL.

This study aims to investigate the expression levels of TNF-α, IFN-γ, IL-4, and IL-10 in dNK cells and determine whether or not the MAPK signal pathway is involved in the regulation of cytokine secretion by dNK cells at the maternal–fetal interface. In this study, we collected decidua specimens from patients with apparently normal pregnant and unexplained recurrent pregnancy loss (URPL) and extracted dNK cells by enzymatic digestion. Then the expression of cytokines were analyzed by flow cytometry and Real-Time PCR respectively. The secretions of both IFN-γ and TNF-α in dNK cells in URPL were significantly higher than those in normal pregnancy. Furthermore, p38/MAPK inhibitors can inhibit the secretion of four cytokines in normal pregnancy, while in URPL cases, p38/MAPK inhibitors only significantly inhibit the secretion of IL-4 and IFN-γ. ERK inhibitors had no effect on the expression of all four cytokines and JNK/MAPK inhibitors varied on different cytokines. URPL is associated with a NK1 cytokine profile. MAPK signaling pathway is involved in the regulation of cytokine secretion by decidual NK cells at maternal–fetal interface. [ABSTRACT FROM AUTHOR]

Recurrent pregnancy loss is a common obstetric complication affecting approximately 1–2% of reproductive population worldwide, but the precise causes for approximately a half of such patients remain unexplained. In this study, we compared the expression profiles of messenger RNA (mRNA), long non-coding RNA (lncRNA), microRNA (miRNA), and circular RNA (circRNA) in villi tissues from patients with unexplained recurrent pregnancy loss (URPL) and elective termination of pregnancy (ETP) using whole-transcriptome sequencing. A number of differentially expressed RNAs were confirmed by real-time PCR analysis. As a result, we identified a total of 1,703 mRNAs, 798 lncRNAs, 199 miRNAs, and 163 circRNAs that were significantly differentially expressed between villi tissues from URPL and ETP. The data of real-time PCR were consistent with those of the sequencing results. Gene ontology and Kyoto Encyclopedia of Genes and Genomes pathway analysis revealed that the majority of differentially expressed mRNAs and target genes of ncRNAs were associated with focal adhesion, extracellular matrix-receptor interaction, and the PI3K-Akt signaling pathway. Additionally, two co-expression networks (lncRNA–miRNA–mRNA and lncRNA-circRNA–miRNA–mRNA) were constructed based on the correlation analysis between the differentially expressed RNAs. Taken together, this study provides a large number of valuable candidates for elucidating regulatory mechanisms of ncRNAs, which may ultimately assist in understanding the pathogenesis of URPL. [ABSTRACT FROM AUTHOR]

Previous studies have suggested that trophoblast cells inhibit the proliferation of peripheral natural killer cells and that the level of peripheral natural killer cells decrease in the middle and late pregnancy stage among healthy women. The change in peripheral natural killer cell level during early pregnancy and the relationship between the change in peripheral natural killer cell level and pregnancy outcomes among women with unexplained recurrent pregnancy loss have not been sufficiently explored. This study aimed to characterize the level of prepregnancy peripheral natural killer cells in comparison with those in early pregnancy among women with unexplained recurrent pregnancy loss and to determine if the change in the level of peripheral natural killer cells from prepregnancy to early pregnancy can predict pregnancy outcomes. In this prospective cohort study, 1758 women with recurrent pregnancy loss were recruited between January 2017 and December 2021 among whom 252 women with unexplained recurrent pregnancy loss had prepregnancy and early pregnancy (4–6 weeks gestation) peripheral natural killer cell measurements. These 252 women were divided into 2 groups, namely those with a lower gestational peripheral natural killer cell level (group 1) when compared with prepregnancy levels and those who did not (group 2). The respective outcomes of these groups in terms of live birth and pregnancy loss were comparatively analyzed using chi-square and Student's t tests. Candidate factors that could influence live birth were selected using the Akaike information criterion. The participates were then randomly divided into training and testing groups. A multivariable logistic regression analysis was performed and a nomogram was created to assess the possibility of live birth. The predictive accuracy was determined by the area under the receiver operating characteristic curve and validated by plotting the predicted probabilities and the observed probabilities. A Hosmer-Lemeshow test was used to assess the goodness of fit. When early gestational peripheral natural killer cell levels were compared with prepregnancy peripheral natural killer cell levels, 61.5% (154) of women had a comparatively lower early-gestational peripheral natural killer cell level and 38.9% (98) of women had an increase or no change in the peripheral natural killer cell level. The live birth rate in group 1 was 89.0% (137/154), which was significantly higher than the rate of 49.0% (48/98) in group 2 (P <.001). A decrease in the peripheral natural killer cell level (odds ratio, 1.36; 95% confidence interval, 1.22–1.55; P <.001) and the anti-Muellerian hormone level (odds ratio, 1.41; 95% confidence interval, 1.14–1.81; P =.003) were important predicting factors for a higher live birth rate. Female body mass index (odds ratio, 0.97; 95% confidence interval, 0.82–1.15; P =.763) and parity (odds ratio, 1.61; 95% confidence interval, 0.71–4.12; P =.287) also were predicting factors. Furthermore, the area under the receiver operating characteristic curve of the model to diagnose of live birth was 0.853 with a sensitivity of 81.6% and a specificity of 78.0% using the training data set. And the Hosmer-Lemeshow test showed that the model was a good fit (p =6.068). We report a comparative decrease in the peripheral natural killer cell levels in early gestation when compared with prepregnancy cell levels in more than 60% of women with unexplained recurrent pregnancy loss at 4 to 6 weeks of gestation. When compared with prepregnancy peripheral natural killer cell levels, a decrease in the peripheral natural killer cell level during early pregnancy might be a useful predictor of the live birth rate among women with unexplained recurrent pregnancy loss. [Display omitted] [ABSTRACT FROM AUTHOR]

Decidual polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) are essential to immune tolerance during pregnancy. A reduction in the number of these cells is associated with unexplained recurrent pregnancy loss (URPL). In our previous study, we reported that PMN-MDSCs are a group of mature neutrophils that are activated by the decidua microenvironment. In the present study, we show that the decidua microenvironment induces substantial lipid accumulation in neutrophils during their differentiation to PMN-MDSCs. Lower levels of lipid accumulation are detected in PMN-MDSCs from URPL patients, and the amount of lipid in the PMN-MDSCs is positively correlated with the proportion of PMN-MDSCs. Next, we demonstrate that decidua-derived IL6 with the presence of arachidonic acid upregulates fatty acid-binding protein 5 (FABP5) via the phosphorylation of signal transducer and activator of transcription 3 (STAT3). Fy -60ABP5 then continuously stimulates intracellular lipid accumulation. Increased intracellular lipid accumulation mediates arachidonic acid metabolism, a pathway that is significantly activated by the induction of the decidua microenvironment, to stimulate the synthesis of prostaglandin E2 (PGE2) and finally induce the differentiation of PMN-MDSCs. To summarize, decidua-derived IL6 facilitates the differentiation of PMN-MDSCs from neutrophils via the pSTAT3/FABP5/PGE2 pathway. Defects in the process may result in impaired differentiation and dysfunction of PMN-MDSCs in URPL. These findings enhance our understanding of the physiological mechanisms of immune tolerance in pregnancy and provide therapeutic options for URPL. [ABSTRACT FROM AUTHOR]

Decidual polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) are essential to immune tolerance during pregnancy. A reduction in the number of these cells is associated with unexplained recurrent pregnancy loss (URPL). In our previous study, we reported that PMN-MDSCs are a group of mature neutrophils that are activated by the decidua microenvironment. In the present study, we show that the decidua microenvironment induces substantial lipid accumulation in neutrophils during their differentiation to PMN-MDSCs. Lower levels of lipid accumulation are detected in PMN-MDSCs from URPL patients, and the amount of lipid in the PMN-MDSCs is positively correlated with the proportion of PMN-MDSCs. Next, we demonstrate that decidua-derived IL6 with the presence of arachidonic acid upregulates fatty acid-binding protein 5 (FABP5) via the phosphorylation of signal transducer and activator of transcription 3 (STAT3). Fy -60ABP5 then continuously stimulates intracellular lipid accumulation. Increased intracellular lipid accumulation mediates arachidonic acid metabolism, a pathway that is significantly activated by the induction of the decidua microenvironment, to stimulate the synthesis of prostaglandin E2 (PGE2) and finally induce the differentiation of PMN-MDSCs. To summarize, decidua-derived IL6 facilitates the differentiation of PMN-MDSCs from neutrophils via the pSTAT3/FABP5/PGE2 pathway. Defects in the process may result in impaired differentiation and dysfunction of PMN-MDSCs in URPL. These findings enhance our understanding of the physiological mechanisms of immune tolerance in pregnancy and provide therapeutic options for URPL.

Purpose: Previous studies have reported the involvement of long noncoding RNAs (lncRNAs) in reproductive diseases via the regulation of target genes. This study aimed to determine whether lnc-prostate androgen-regulated transcript 1 (lnc-PART1)could be used as a biomarker of unexplained recurrent pregnancy loss (URPL) and a possible predictor of poor pregnancy outcomes in women with URPL., Materials and Methods: Sixty patients with URPL and 15 healthy women were included in this study. PART1 expression was detected in plasma and endometrial tissues using a quantitative reverse transcription polymerase chain reaction. Logistic regression and receiver operating characteristic curve analyses were performed to analyze the association between PART1 expression and pregnancy outcomes in women with URPL., Results: The expression of PART1transcript variant 2 was significantly up-regulated in the endometrial specimens from patients with URPL compared to control tissues. High tissue expression levels of PART1transcript variant 2 were associated with poor pregnancy outcomes in women with URPL, indicating that it could serve as a potential risk factor. Additionally, PART1 could serve as a potential risk factor for adverse pregnancy outcomes in patients with URPL (OR = 4.374; 95% CI = 1.052-18.189; p  = .042)., Conclusion: lncRNA PART1 transcript variant 2 was highly expressed in patients with URPL. Therefore, it is important to conduct in-depth studies on the relationship between PART1 expression and URPL.

This real-world study aimed to assess the clinical efficacy and safety of intravenous immunoglobulin (IVIG) in treating unexplained recurrent pregnancy loss (uRPL) patients. A total of 238 patients with uRPL were enrolled in this retrospective study. According to the treatment, patients were assigned into treatment group (n=184, receiving IVIG+low molecular weight heparin [LMWH]) and control group (n=54, only LMWH). The primary outcome was the live birth rates (LBR) after 24 weeks of gestation. In addition, we performed subgroup analyses to assess the LBR in uRPL patients with age (≥30 years or <30 years), body mass index (BMI, 18-24 kg/m 2 or ≥24 kg/m 2 ), number of previous pregnancy losses (2 or ≥3), gestational week of first medication, and IVIG medication regimen during pregnancy. The LBR showed a significant increase after IVIG treatment compared to the control group (77.7 % vs. 53.7 %, P=0.001). Multivariable logistic regression indicated that IVIG was associated with increased LBR (aOR=4.383, 95 %CI 2.024-9.489). Besides, we observed a significantly lower incidence of decreased platelet count (1.6 % vs. 7.4 %, P=0.049) in the treatment group. Subgroup analyses showed that IVIG treatment was associated with improved LBR in uRPL patients with age of <30 years (aOR=4.012, 95 %CI 1.428-11.266), previous pregnancy losses ≥3 times (aOR=17.327, 95 %CI 2.525-118.921), and BMI of 18-24 kg/m 2 (aOR=8.914, 95 %CI 2.985-26.618). IVIG treatment significantly improves the LBR in uRPL patients. These findings from real-world data provide evidence for the safety and efficacy of IVIG as a viable treatment for uRPL patients in clinical practice., Competing Interests: Declaration of Competing Interest The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Recurrent pregnancy loss (RPL) is a common fertility problem that affects 1%-2% of couples all over the world. Despite exciting discoveries regarding the important roles of the decidual natural killer cell (dNK) and regulatory T cell in pregnancy, the immune heterogeneity in patients with unexplained recurrent pregnancy loss (URPL) remains elusive. Here, we profiled the transcriptomes of 13,953 CD45+ cells from three normal and three URPL deciduas. Based on our data, the cellular composition revealed three major populations of immune cells including dNK cell, T cell, and macrophage, and four minor populations including monocytes, dendritic cell (DC), mast cell, and B cell. Especially, we identified a subpopulation of CSF1+ CD59+ KIRs-expressing dNK cells in normal deciduas, while the proportion of this subpopulation was decreased in URPL deciduas. We also identified a small subpopulation of activated dDCs that were accumulated mainly in URPL deciduas. Furthermore, our data revealed that in decidua at early pregnancy, CD8+ T cells exhibited cytotoxic properties. The decidual macrophages expressed high levels of both M1 and M2 feature genes, which made them unique to the conventional M1/M2 classification. Our single-cell data revealed the immune heterogeneity in decidua and the potentially pathogenic immune variations in URPL.

Recurrent pregnancy loss (RPL) is a common fertility problem that affects 1%-2% of couples all over the world. Despite exciting discoveries regarding the important roles of the decidual natural killer cell (dNK) and regulatory T cell in pregnancy, the immune heterogeneity in patients with unexplained recurrent pregnancy loss (URPL) remains elusive. Here, we profiled the transcriptomes of 13,953 CD45+ cells from three normal and three URPL deciduas. Based on our data, the cellular composition revealed three major populations of immune cells including dNK cell, T cell, and macrophage, and four minor populations including monocytes, dendritic cell (DC), mast cell, and B cell. Especially, we identified a subpopulation of CSF1+ CD59+ KIRs-expressing dNK cells in normal deciduas, while the proportion of this subpopulation was decreased in URPL deciduas. We also identified a small subpopulation of activated dDCs that were accumulated mainly in URPL deciduas. Furthermore, our data revealed that in decidua at early pregnancy, CD8+ T cells exhibited cytotoxic properties. The decidual macrophages expressed high levels of both M1 and M2 feature genes, which made them unique to the conventional M1/M2 classification. Our single-cell data revealed the immune heterogeneity in decidua and the potentially pathogenic immune variations in URPL. [ABSTRACT FROM AUTHOR]

Objective. To investigate the difference of prolactin (PRL) expression in peripheral blood and decidual stromal cells (DSCs) between unexplained recurrent pregnancy loss (URPL) and normal early pregnancy. Methods. Eighty cases of URPL patients in our hospital from March 2018 to March 2019 were recruited. Seventy women with induced abortion and 190 normal pregnancy women were selected as controls. Fasting peripheral blood and decidua tissues were collected. The level of PRL in peripheral blood was detected by enzyme-linked immunosorbent assay. Quantitative real time PCR (RT-qPCR), Western bloting (WB) and immunohistochemistry (IHC) were used to detect the PRL expression of DSCs. Independent sample t test was used to compare the data of the two different groups, and one-way analysis of variance test was used to compare the data of the groups. Results. In the URPL group, 21 cases were excluded for chromosomal abnormality, and 59 cases were finally included. There were 70 cases in the induced abortion group. One hundred and seventy-seven cases in the normal pregnancy group completed follow-up, of which 157 cases were classified as final live birth in the normal pregnancy group, and the rest 20 cases subsequently had pregnancy loss, of which 19 miscarriages occurred in the first 10 weeks of gestation, classified as pregnancy loss in the normal pregnancy group. The PRL levels of peripheral blood in the final live birth in the normal pregnancy group and induced abortion group were similar, with no significant difference between the two groups in the same gestation, and they all increased significantly with advancing gestation (normal pregnancy with final live birth group: 5-5+6 week vs 6-7+6 week, P = 0.002; 6-7+6 week vs 8-9+6 week,P = 0.012;Induced abortion group: 5-5+6 week vs 6-7+6 week, P = 0.015; 6-7+6 week vs 8-9+6 week, P = 0.023). The levels of PRL in the peripheral blood of patients with URPL had no significant change with advancing gestation, and they were significantly lower than those of women with final live birth in the normal pregnancy group and induced abortion women in the same gestation from the 6th week (6-7+6week: P = 0.012, 0.024; 8-9+6 week: P = 0.015, 0.003). In addition, the levels of PRL in the peripheral blood of the women in the normal pregnancy group who later suffered miscarriage were similar to those in the URPL group of the same gestation. The PRL expression of DSCs in the induced abortion group increased significantly in both RNA and protein level with advancing gestation (5-5+6 week vs 6-7+6 week: RT-qPCR, P = 0.020; WB, P = 0.010; IHC,P = 0.030. 6-7+6 week vs 8-9+6 week: RT-qPCR, P = 0.011; WB, P = 0.034; IHC, P = 0.012). However, the increase of PRL expression of DSCs in the URPL group was delayed: the PRL expression of DSCs in women with 6-7+6week and 8-9+6 week of gestation was significantly lower than that in women accepting induced abortion of the same gestation (6-7+6 week: RT-qPCR, P = 0.012; WB, P = 0.014; IHC,P = 0.028. 8-9+6 week: RT-qPCR, P = 0.011; WB, P = 0.030;IHC, P = 0.026). Conclusion. PRL expression of the decidua is defective. The PRL expression of DSCs in URPL patients is deficient, and the lack of decidualization of DSCs may be related to the occurrence of URPL. The low level of PRL in peripheral blood after 6 weeks of gestation may indicate embryonic dysplasia. [ABSTRACT FROM AUTHOR]

Myeloid-derived suppressor cells (MDSC), especially polymorphonuclear MDSC (PMN-MDSC), accumulate in maternal-fetal interface during pregnancy and are involved in the maintenance of immune tolerance. Decreased PMN-MDSC is associated with pregnancy complications such as unexplained recurrent pregnancy loss (URPL). In the present study we showed decreased PMN-MDSC in the URPL group compared with the normal pregnancy (NP) group, and PMN-MDSC was the major subset of MDSC in human decidua with potent immune suppression activity. We then performed gene expression profile and found that human decidual PMN-MDSC in the NP and URPL groups showed different gene and pathway signature, including apoptosis. Apoptosis of decidual PMN-MDSC was mediated by TNF-related apoptosis–induced ligand (TRAIL) in a Caspase 3 dependent manner. TRAIL was expressed in decidua and upregulated in decidua of the URPL group. Notably, of all the membrane TRAIL receptors, only DcR2 was down-regulated in PMN-MDSC in the URPL group. In vitro experiment demonstrated that DcR2 blockade sensitized PMN-MDSC to TRAIL-mediated apoptosis. Together, these data indicate that increased TRAIL and reduced DcR2 on PMN-MDSC sensitize PMN-MDSC response to TRAIL-induced apoptosis in the URPL group, which is responsible for decreased accumulation of PMN-MDSC in URPL.

Résumé Contexte Les fausses couches à répétition (FCR) sont définies lorsqu’au moins trois fausses couches ont eu lieu au cours du premier trimestre. Bien que la cause la plus fréquente soit l’aneuploïdie embryonnaire, et malgré un bilan chez la femme et un caryotype du couple, dans environ 50% des cas, les FCR restent inexpliquées. L’implication masculine a été peu étudiée et les résultats restent discordants. Ainsi, nous avons réalisé une étude cas-témoins prospective et multicentrique afin d’investiguer l’implication du gamète mâle dans les FCR inexpliquées. Méthodes Un total de 33 cas et de 27 témoins ont été inclus recrutés au sein de trois hôpitaux universitaires. Nous avons étudié les facteurs environnementaux et familiaux à partir d’un questionnaire détaillé ainsi que les données de l’examen andrologique, les caractéristiques du sperme, la fragmentation de l’ADN et la chromatine du spermatozoïde en utilisant le sperm chromatine structure assay (SCSA) et le test du TUNEL, ainsi que l’aneuploïdie des spermatozoïdes grâce à la méthode d’hybridation in situ de sonde chromosomique (FISH). Le test de Mann-Whitney et les tests exacts de Wilcoxon ou de Fisher ont été utilisés. Une corrélation de Spearman non-paramétrique a été réalisée afin d’analyser la relation entre les divers paramètres de sperme et les résultats de fragmentation d’ADN du sperme et les résultats de la FISH. Résultats Nous avons trouvé des différences significatives entre les cas et les témoins pour le délai de conception, l’indice de masse corporelle (IMC), les antécédents familiaux d’infertilité et le milieu de vie. Chez les cas, la mobilité totale des spermatozoïdes et le pourcentage de spermatozoïdes normaux étaient significativement diminués. Aucune différence n’a été trouvée entre les cas et les témoins concernant la fragmentation de l’ADN des spermatozoïdes ou l’intégrité de la chromatine. Chez les cas, la fréquence des spermatozoïdes présentant une aneuploïdie, une hyperhaploïdie et une disomie du chromosome 18 étaient significativement augmentée. Conclusions Cette étude cas-témoins prospective est. l’une des plus importantes ayant investigué à la fois les facteurs environnementaux, les caractéristiques des spermatozoïdes, la fragmentation et la chromatine de l’ADN des spermatozoïdes et les anomalies chromosomiques des spermatozoïdes en rapport avec les fausses couches à répétition inexpliquée. L’originalité de notre étude réside dans l’examen andrologique complet et la recherche des facteurs de risque et des antécédents reproductifs. D’autres études sont nécessaires pour confirmer les liens entre les FCR inexpliquées et les antécédents familiaux masculins d’infertilité ou de fausses couches. L’augmentation de l’aneuploïdie des spermatozoïdes observée chez les cas présentant des FCR inexpliquées plaide en faveur d’une étiologie masculine. Ces données ouvrent la voie à d’autres études pour démontrer l’utilité d’un dépistage génétique préimplantatoire chez les hommes présentant une augmentation de l’aneuploïdie des spermatozoïdes dont les partenaires présentent des FCR inexpliquées.

Background and Objectives: Unexplained recurrent pregnancy loss (RPL) has been a challenging issue in diagnosis. Methylenetetrahydrofolate reductase (MTHFR) gene polymorphisms (677C > T, 1298A > C) and plasminogen activator inhibitor type 1 (PAI-1) 4G/5G gene polymorphism have been proposed as risk factors of unexplained RPL. This study aimed to identify relationships between MTHFR 677C > T, 1298A > C gene polymorphisms and PAI-1 4G/5G gene polymorphism on unexplained RPL. Materials and Methods: A case-control study was performed on two groups: pregnant women with unexplained RPL (i.e., two or more pregnancy losses) as the case group, and those with at least one or more live births and no history of RPL as the control group. All pregnant women were assessed for the presence of MTHFR gene polymorphisms and PAI-1 4G/5G polymorphism by using the amplification refractory mutation system polymerase chain reaction (ARMS-PCR) technique. Associations between MTHFR 677C > T, 1298A > C gene polymorphisms, and PAI-1 4G/5G gene polymorphism with unexplained RPL were evaluated. Results: There were 73 pregnant women (43 cases and 30 controls) recruited. The rates of MTHFR 677C > T among cases were: TT allele 32.6%, CT allele 34.8%, and CC allele 32.6%, while the frequencies of these alleles were: TT alleles 10%, CT allele 36.7%, and CC allele 53.3% (p = 0.049). MTHFR 677C > T gene polymorphism was associated with an increased risk of unexplained RPL (for TT allele vs. CT + CC allele: OR = 3.25, 95%CI: 1.02-10.35, p = 0.028; for T vs. C: OR = 2.53, 95%CI: 1.25-5.11, p = 0.09). MTHFR 1298A > C gene polymorphism and PAI-1 4G/5G gene polymorphism were not related to unexplained RPL. Conclusion: MTHFR 677C > T gene polymorphism was related to unexplained RPL in Vietnamese pregnant women. Further longitudinal studies should be performed to examine this relationship. [ABSTRACT FROM AUTHOR]

Introduction: There has a close relationship between the balance of T helper 17 (Th17) cells and Foxp3+ regulatory T cells (Treg) and unexplained recurrent pregnancy loss (URPL). The present study is to investigate the regulatory effect of daphnetin, which is extracted from Daphne odora Var, on the balance of Th17 and Treg cells in the peripheral blood mononuclear cells (PBMCs) from patients with URPL. Material and methods: PBMCs were isolated from 35 pregnant women with URPL and 35 women with normal early pregnancies, respectively and treated with daphnetin for three days. Flow cytometry was performed to measure the proportions of Th17 and Treg cells. The level of expression of IL-6, TGF-β1 and IL-2 were detected using enzyme-linked immunosorbent assay (ELISA) and the level of expression of FoxP3, RORγt, signal transducers and activators of transcriotion 3 (STAT3) and STAT5 were detected by RT-PCR. Results: The concentrations of Th17-type cytokines IL-2 were significantly decreased in the URPL group after treatment (p < 0.01). Treg-type cytokines such as TGF-β1 and IL-6 were significantly increased after treatment (p < 0.01). At the same time, daphnetin may induce a decrease in the ratio of RORγt to Foxp3 and a Treg cell bias, which would be beneficial for pregnancy maintenance. Futhermore the expression level of STAT3 were higher in the URPL patients whereas STAT5 were lower than those in the control subjects. Conclusions: In conclusion, daphnetin may have regulatory effect on the balance of Th17 and Treg cells in the peripheral blood mononuclear cells from patients with URPL.. [ABSTRACT FROM AUTHOR]

Myeloid-derived suppressor cells (MDSC), especially polymorphonuclear MDSC (PMN-MDSC), accumulate in maternal-fetal interface during pregnancy and are involved in the maintenance of immune tolerance. Decreased PMN-MDSC is associated with pregnancy complications such as unexplained recurrent pregnancy loss (URPL). In the present study we showed decreased PMN-MDSC in the URPL group compared with the normal pregnancy (NP) group, and PMN-MDSC was the major subset of MDSC in human decidua with potent immune suppression activity. We then performed gene expression profile and found that human decidual PMN-MDSC in the NP and URPL groups showed different gene and pathway signature, including apoptosis. Apoptosis of decidual PMN-MDSC was mediated by TNF-related apoptosis–induced ligand (TRAIL) in a Caspase 3 dependent manner. TRAIL was expressed in decidua and upregulated in decidua of the URPL group. Notably, of all the membrane TRAIL receptors, only DcR2 was down-regulated in PMN-MDSC in the URPL group. In vitro experiment demonstrated that DcR2 blockade sensitized PMN-MDSC to TRAIL-mediated apoptosis. Together, these data indicate that increased TRAIL and reduced DcR2 on PMN-MDSC sensitize PMN-MDSC response to TRAIL-induced apoptosis in the URPL group, which is responsible for decreased accumulation of PMN-MDSC in URPL. [ABSTRACT FROM AUTHOR]

Problem: To determine whether patients with unexplained recurrent pregnancy loss (URPL) can benefit from pre‐conception immunotherapy or on the early phase of the first trimester. Method of study: The prospective follow‐up study which involved pre‐conception patients diagnosed with URPL following rigorous etiology screening in the medical center of recurrent pregnancy loss. In this study, pre‐conception immunotherapy included lymphocyte immunotherapy (pre‐LIT). Post‐conception immunotherapy (post‐IM) included LIT or intravenous immunoglobulin (IVIG). Patients were recommended to undergo post‐IM immediately from human chorionic gonadotrophin (hCG) elevation. Autoimmune antibodies (AIA) and anti‐paternal lymphocytotoxic antibodies (APLA) were tested before and after pre‐LIT. Favorable outcome was defined as pregnancy over 14 weeks. Unfavorable outcomes included biochemical pregnancy loss (BPL) and pregnancy loss with clear implantation location (PLCIL). Results: In this study, URPL accounted for 12.9% of recurrent pregnancy loss (217/1682). Frequency of BPL was significantly lower in patients with post‐IM than that without post‐IM [2.8% vs 28.2%; adjusted relative risk (aRR), 0.06; 95% confidence interval (CI), 0.01‐0.24]. There was a significant positive conversion in the AIA induced by pre‐LIT (0.0% vs 31.0%). Frequency of PLCIL in patients with positive iatrogenic AIA conversion induced by pre‐LIT was higher than that in patients without AIA conversion [30.4% vs 5.8%; aRR, 7.53; 95% CI, 1.31‐43.34]. Conclusion: Pre‐LIT of patients with URPL contributed to a positive iatrogenic AIA conversion, which was associated with an increased risk of PLCIL. Post‐IM immediately initiated from the time of hCG elevation can reduce the incidence of BPL. [ABSTRACT FROM AUTHOR]

This case-control study was conducted to assess whether neopterin (NP) and anti-Mullerian hormone (AMH) can be used as markers in the condition of unexplained recurrent pregnancy loss (RPL). To the best of our knowledge, this is the first work which has studied the association between AMH, NP and RPL. A total of 110 patients were included; 53 women who had at least two consecutive unexplained miscarriages (<12 weeks) made up the study group. The control group was established from 57 women who never had a miscarriage and had at least one healthy birth history. Peripheral blood samples were collected from each patient to analyse the AMH and NP concentrations. The results suggested that mean level of AMH (1.38 ± 0.683 ng/ml) in the patient's group was significantly lower than the control group (1.84 ± 0.718 ng/ml). The mean levels of NP were significantly higher in the patient group (1.69 ± 0.486 vs. 1.38 ± 0.431 ng/ml). IMPACT STATEMENT What is already known about this subject? To the best of our knowledge, no previous studies about the association among AMH, NP and RPL were found in the literature. What do the results of this study add? This pioneer study demonstrates the significant relationship between the unexplained RPL with the increased levels of NP and decreased AMH. What are the implications of these findings for clinical practice and/or further research? NP and AMH may play an effective role in illuminating the condition of unexplained RPL. High levels of NP and low values of AMH in patients with RPL can be used as predictive markers for this clinical situation. If the causes of high levels of NP and low levels of AMH can be better illuminated, new treatments towards these causes can be developed to help such patients become childbearing. [ABSTRACT FROM AUTHOR]

We assessed the outcome of pregnancy in women with a history of recurrent pregnancy loss (RPL) following treatment with low-dose human menopausal gonadotropin (HMG)+progesterone or progesterone alone. This single-center retrospective cohort study included data from women diagnosed with RPL and treated between February 2005 and December 2012 with one cycle of HMG + progesterone or progesterone alone. Primary endpoint was the rate of ongoing pregnancies and losses by treatment, age (<38 vs. ≥38 years) and in the subgroup with unexplained RPL. Of 169 RPL patients, 35.5% (n = 60) received HMG + progesterone and 64.5% (n = 109) progesterone alone. Compared to progesterone alone, HMG + progesterone led to a lower, although not significant, frequency of losses (3.3% vs. 11.9%, p = .09) and a twofold higher rate of ongoing pregnancies (41.7% vs. 19.3%, p = .002). Similar results were obtained in the subgroup of patients with unexplained RPL (ongoing pregnancies: 48.1% upon HMG + progesterone vs. 21.3% upon progesterone, p = .03; losses: 0% vs. 8.5%, respectively, p = .29) and in those <38 years (ongoing pregnancies: 47.4% vs. 18.8%, respectively, p = .003; losses: 5.3% vs. 10.9% respectively, p = .47). These findings suggest that HMG in women with RPL may reduce the rate of miscarriages and increase that of live births regardless of RPL cause and in women aged <38 years. [ABSTRACT FROM AUTHOR]

Objective: To establish a statistical model to predict thrombophilia in patients with unexplained recurrent pregnancy loss (URPL).Methods: A retrospective case-control study was conducted at Ren Ji Hospital, Shanghai, China, from March 2014 to October 2016. The levels of D-dimer (DD), fibrinogen degradation products (FDP), activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT), fibrinogen (Fg), and platelet aggregation in response to arachidonic acid (AA) and adenosine diphosphate (ADP) were collected. Receiver operating characteristic curve analysis was used to analyze data from 158 UPRL patients (≥3 previous first trimester pregnancy losses with unexplained etiology) and 131 non-RPL patients (no history of recurrent pregnancy loss). A logistic regression model (LRM) was built and the model was externally validated in another group of patients.Results: The LRM included AA, DD, FDP, TT, APTT, and PT. The overall accuracy of the LRM was 80.9%, with sensitivity and specificity of 78.5% and 78.3%, respectively. The diagnostic threshold of the possibility of the LRM was 0.6492, with a sensitivity of 78.5% and a specificity of 78.3%. Subsequently, the LRM was validated with an overall accuracy of 83.6%.Conclusion: The LRM is a valuable model for prediction of thrombophilia in URPL patients. [ABSTRACT FROM AUTHOR]

In this study, recent research focusing on recurrent pregnancy loss (RPL) are reviewed. Recurrent pregnancy loss is a devastating reproductive health burden that affects about 5% of couples trying to conceive globally. Currently, there are few evidence-based diagnostic and treatment strategies for RPL. More so, the number of unexplained etiology cases in patients with RPL arrives at 50%. Here, we discuss the progress in diagnosis and treatment of unexplained RPL, as well as recommended treatment strategies and controversial etiologies. • Genetic abnormalities, mostly single nucleotide polymorphisms, are related to URPL. • Single-cell RNA sequencing is utilized in discovery of new cell subgroups at the maternal-fetal interface. • Lymphocyte immunotherapy may boost therapeutic effect in URPL patients. • IVIG can improve the live birth rate of PRL patients with dysimmunity. [ABSTRACT FROM AUTHOR]

Bisphenol analogues are widely used in consumer products such as disposable dinnerware, canned food, personal care products, bottled beverages, and more, and dietary exposure is the main pathway. Bisphenol A is used to manufacture synthetic resins and commercial plastics in large quantities. According to epidemiological and animal studies, bisphenols disrupt the reproductive, immunological, and metabolic systems. These analogues are estrogenic like Bisphenol A, although human studies are limited. We did a thorough search of the literature on the toxicity of bisphenol on reproductive and endocrine systems in pregnancy, focusing particularly on human studies. Hence, we present a comprehensive literature review on this topic.​​​​​​​ During our literature search, three epidemiological studies and one human observational study demonstrated a substantial link between bisphenol toxicity and recurrent miscarriages.​​​​​​​ The aforementioned research shows that bisphenol may harm pregnancy and cause miscarriages. We believe this is the first literature review on the topic., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2023, Agarwal et al.)

Low levels of folate combined with high levels of homocysteine may cause unexplained recurrent pregnancy loss (URPL). However, the relationships between polymorphisms in genes of the folate metabolic pathway and URPL remain controversial. We conducted a case-control study to explore polymorphisms of the major folate pathway genes, including methylenetetrahydrofolate reductase (MTHFR) 677C>T, MTHFR 1298A>C, methionine synthase (MTR) 2756A>G, methionine synthase reductase (MTRR) 66A>G and reduced folate carrier 1 (RFC-1) 80A>G, and their associations with URPL. Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to determine the distributions of MTHFR, MTR and RFC-1 polymorphisms, and the results were validated using direct sequencing. The polymorphisms in MTRR were determined using direct sequencing. Haplotypes were analyzed using SHEsis, an online tool for biological analysis. We found that the MTHFR 677T allele and the 677T/1298A/2756A/66A/80G haplotype were risk factors for URPL, while the MTR 2756G allele and the 677C/1298A/2756A/66A/80A haplotype exhibited protective effects on susceptibility to URPL in a Chinese Han population from the Hangzhou area. [ABSTRACT FROM AUTHOR]

KIAA0319L, PXK and JAZF1 gene polymorphisms were investigated to determine whether they conferred susceptibility to unexplained recurrent pregnancy loss (URPL) in a group of Chinese Han patients. Genotyping and sequencing of the single nucleotide polymorphisms (SNP) rs2275247(A/G) in KIAA0319L , rs2176082(C/T) and rs6445975(G/T) in PXK and the rs1635852(C/T) in J AZF1 were carried out in 84 couples with URPL and 102 healthy couples with at least one live birth. Frequencies of the SNP rs2176082(C/T) in PXK gene were significantly different between women with URPL and control women: P < 0.05; OR 95% CI 0.530 (0.287 to 0.979); OR 95% CI 0.482 (0.254 to 0.911) but were not significantly different after Bonferroni correction. The frequencies of the SNP rs2176082(C/T) in PXK gene showed no difference between the husband of a woman with URPL and a control husband: OR 95% CI 1.494 (0.821 to 2.721); OR 95% CI 1.567 (0.841 to 2.921). No statistically significant differences were observed in the distribution of any genotype or allele frequency or any genetic model of the other three SNPs between couples with URPL and control couples. Therefore, the rs2176082(C/T) polymorphism of PXK might play a possible role in the development of URPL in Chinese Han women. [ABSTRACT FROM AUTHOR]

Recurrent pregnancy loss (RPL) puzzles 1-3% of women of childbearing age worldwide. Immunological factors account for more than 60% of cases of unexplained RPL (URPL); however, the underlying mechanism remains unclear. Here, using single-cell sequencing data and functional experiments with clinical samples, we identified a distinct population of CCR1 + decidual macrophages (dMφ) that were preferentially enriched in the decidua from normal early pregnancies but were substantially decreased in patients with URPL. Specific gene signatures endowed CCR1 + dMφ with immunosuppressive and migration-regulatory properties, which were attenuated in URPL. Additionally, CCR1 + dMφ promoted epithelial-to-mesenchymal transition (EMT) to promote trophoblast migration and invasion by activating the ERK1/2 signaling pathway. Decidual stromal cell (DSC)-derived CCL8 was the key regulator of CCR1 + dMφ as CCL8 recruited peripheral CCR1 + monocytes, induced a CCR1 + dMφ-like phenotype, and reinforced the CCR1 + dMφ-exerted modulation of trophoblasts. In patients with URPL, CCL8 expression in DSCs was decreased and trophoblast EMT was defective. Our findings revealed that CCR1 + dMφ play an important role in immune tolerance and trophoblast functions at the maternal-fetal interface. Additionally, decreased quantity and dysregulated function of CCR1 + dMφ result in URPL. In conclusion, we provide insights into the crosstalk between CCR1 + dMφ, trophoblasts, and DSCs at the maternal-fetal interface and macrophage-targeted interventions of URPL., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sang, Li, Xu, Chen, Li and Du.)

Recurrent pregnancy loss (RPL) is a multifactorial disorder, both genetic and environmental factors contribute to the development of RPL. Recently, the association between endothelial nitric oxide synthase (eNOS) gene polymorphisms and unexplained recurrent pregnancy loss (URPL) has been reported, and the results were inconsistent. Hence, we performed the meta-analysis to drive a more precise estimation of association between eNOS polymorphisms and URPL. Odds ratio (OR) and its 95 % confidence interval were calculated under co-dominant (AA vs. BB, TT vs. GG) and additive (A vs. B, T vs. G) genetic models. Studies of eNOS intron 4 VNTR and Glu298Asp were separated by ethnicities. 13 studies included 1,769 URPL cases and 1,376 healthy controls on eNOS intron 4 VNTR polymorphism, and 11 studies were involved in Glu298Asp polymorphism with 1,498 URPL cases and 1,123 healthy controls. The integrated results showed that eNOS Glu298Asp polymorphism was associated with URPL [ORs were 1.91 (1.42-2.56), P < 0.001; 1.67 (1.36-2.04), P < 0.001, respectively]. When analyses were separated by ethnic subgroups, the association between eNOS Glu298Asp polymorphism and URPL was only observed in East Asians [OR = 1.88 (1.52-2.33), P < 0.001 under additive model], and there was no association between eNOS intron 4 VNTR polymorphism and URPL in Caucasians and East Asians. The results indicated a significant association between eNOS Glu298Asp polymorphism and URPL in East Asians. No association was observed between eNOS intron 4 VNTR polymorphism and URPL in Caucasians and East Asians. [ABSTRACT FROM AUTHOR]