Cause Of Recurrent Miscarriage Research Articles

Identification and RT-qPCR Validation of Biomarkers Based on Butyrate Metabolism-Related Genes to Predict Recurrent Miscarriage.

To date, the cause of recurrent miscarriage (RM) in at least 50% of patients remains unknown. However, no study has explored the correlation between butyrate metabolism-related genes (BMRGs) and RM. RM-related datasets (GSE165004, GSE111974, GSE73025, and GSE179996) were obtained from the Gene Expression Omnibus (GEO) database. First, 595 differentially expressed genes (DEGs) were identified between the RM and control samples in GSE165004. Subsequently, 213 differentially expressed BMRGs (DE-BMRGs) were identified by considering the intersection of DEGs with BMRGs. The protein-protein interaction (PPI)network of DE-BMRGs contained 156 nodes and 250 edges, and a key module was obtained. In total, four biomarkers (ACTR2, ANXA2, PFN1, and OAS1) were acquired through least absolute shrinkage and selection operator (LASSO), support vector machine-recursive feature elimination (SVM-RFE), and random forest (RF). Immune analysis revealed two immune cells and three immune-related gene sets that were significantly different between the RM and control groups, namely, T helper cells, regulatory T cells (Treg), MHC class I, parainflammation, and type I IFN response. In addition, a TF-mRNA network based on the top 100 nodes ranked in the order of connectivity was created, including 100 nodes and 253 edges, such as MTERF2-ACTR2, NKX23-PFN1, STAT1-OAS1, and SP100-ANXA2. Finally, 3 drugs (withaferin A, N-ethylmaleimide, and etoposide) were predicted to interact with 2 biomarkers (ANXA2 and ACTR2). Eventually, ANXA2 and OAS1 were significantly downregulated, and PFN1 was markedly overexpressed in the RM group, as determined by reverse transcription quantitative polymerase chain reaction (RT-qPCR). Our findings authenticated four butyrate metabolism-related biomarkers for the diagnosis of RM, providing a scientific reference for further studies on RM treatment.

Study on pharmacokinetic and tissue distribution of hyperin, astragalin, kaempferol-3-O-β-D-glucuronide from rats with multiple administrations of Semen Cuscutae processed with salt solution with effect of treating recurrent spontaneous abortion.

Semen Cuscutae is a traditional Chinese medicine (TCM) that tonifies the kidneys and prevents miscarriage. According to Chinese medicine theory, kidney deficiency is one of the main causes of recurrent spontaneous abortion (RSA). The previous studies showed that raw product of Semen Cuscutae (SP) and Semen Cuscutae processed with salt solution (YP) have ameliorative effects on RSA, and that YP is superior to SP. However, the active components of YP to ameliorate RSA remain unclear and require further studies. The objective of this study is to investigate the active components of YP in ameliorating RSA. First, a rat model of RSA was established using hydroxyurea in combination with mifepristone. Aqueous decoction of YP was given by gavage to rats. Second, pregnant rats were sampled on days 5, 7, 9, 10 and 12 during the modelling period. The content of Hyperin (HY), astragalin (AS) and kaempferol-3-O-β-D-glucuronide (KA) in blood and liver, heart, spleen, lung and kidney tissues were detected by liquid chromatography-mass spectrometry (LC-MS). The pharmacodynamic indicators including progesterone (P), chorionic gonadotropin β (β-HCG), estradiol (E2), tumor necrosis factor-α (TFN-α), interleukin 4 (IL-4), and tryptophan (TRP) were measured by enzyme-linked immunosorbent assay (ELISA) Pearson's correlation analysis and grey relational analysis were used to establish the relationship between the pharmacodynamic indexes and chemical constituents. The pharmacokinetic results showed that the area under curve (AUC) value of KA was the largest. The tissue distribution results showed that astragalin was widely distributed in liver, heart, spleen, lung and kidney in the RSA model rats, while HY was detected only in the uterus, and KA was detected only in the kidney. The pearson correlationl analysis showed that KA was significantly and positively correlated with the contents of E2, P, β-HCG and TRP. Both AS and HY were significantly negatively correlated with the content of TNF-α, respectively. This study reveals the pharmacokinetics and tissue distribution of KA, AS and HY in rats with RSA. It was elucidated that all three were involved in the regulation of progesterone levels and immune function. It initially revealed the mechanism of action of YP in enhancing the improvement of RSA, and it provided a theoretical basis for the quality assessment of YP.

Glutamine metabolism promotes human trophoblast cell invasion via COL1A1 mediated by PI3K-AKT pathway

Abnormal trophoblast invasion function is an important cause of recurrent spontaneous abortion (RSA). Recent research has revealed a connection between glutamine metabolism and RSA. However, the interplay between these three factors and their related mechanisms remains unclear. To address this issue, we collected villus tissues from 10 healthy women with induced abortion and from 10 women with RSA to detect glutamine metabolism. Then, the trophoblast cell line HTR-8/SVneo was used in vitro to explore the effect of glutamine metabolism on trophoblast cells invasion, which was tested by transwell assay. We found that the concentration of glutamine in the villi of the normal pregnancy group was significantly higher than that in the RSA group. Correspondingly, the expression levels of key enzymes involved in glutamine synthesis and catabolism, including glutamine synthetase and glutaminase, were significantly higher in the villi of the normal pregnancy group. Regarding trophoblast cells, glutamine markedly enhanced the proliferative and invasive abilities of HTR-8/SVneo cells. Additionally, collagen type I alpha 1 (COL1A1) was confirmed to be a downstream target of glutamine, and glutamine also activated the PI3K-AKT pathway in HTR-8/SVneo cells. These findings indicate that glutamine metabolism facilitates the invasion of trophoblasts by up-regulating COL1A1 expression through the activation of the PI3K-AKT pathway, but the specific mechanism of COL1A1 requires further study.

Genetic landscape of thrombophilia in recurrent miscarriages.

The etiology of recurrent miscarriage (RM) is extremely heterogeneous, encompassing genetic, immunological, anatomical, endocrine, thrombophilic, infectious, and uterine abnormalities. Thrombophilia is a major contributor to pregnancy complications, potentially harming the fetus and jeopardizing the continuation of pregnancy. Therefore, successful pregnancy outcomes depend on maintaining a delicate balance between coagulation and fibrinolytic factors, crucial for ensuring the adjustment of the basal plate to facilitate adequate placental perfusion. Despite numerous studies shedding light on the role of thrombophilic factors and genetic variations in RM, the exact pathogenesis remains unclear. It is imperative to systematically rule out thrombophilia and other related factors responsible for pregnancy disorders and RMs to guide appropriate and active management strategies. Addressing thrombophilia continues to present challenges in terms of effective treatment. The current review aims to address the heterogeneity of RM as a therapeutic challenge, emphasizing the need for standardized diagnostic tests and well-designed multicenter research trials to gather robust, evidence-based data on thrombophilic causes of RM and provide effective treatment. The goal is to enhance the understanding of thrombophilic factors and genetic landscapes associated with RM through various approaches, including candidate gene studies, genome-wide association studies, and high-throughput sequencing. Meta-analyses have underscored the significance of genetic aberrations in RM, highlighting the necessity for identifying critical mutations implicated in the etiopathogenesis of miscarriages to pave the way for implementation of targeted clinical therapies.

CBLL1 promotes endometrial stromal cell senescence via inhibiting PTEN in recurrent spontaneous abortion.

Recurrent spontaneous abortion (RSA) is a common pregnancy-related disorder. Cbl proto-oncogene like 1 (CBLL1) is an E3 ubiquitin ligase, which has been reported to vary with the menstrual cycle in the endometrium. However, whether CBLL1 is involved in the occurrence and development of RSA remains unclear. This study aimed to investigate the effects of CBLL1 on RSA. We analyzed the expression of CBLL1 in the decidua of RSA patients, as well as its functional effects on cellular senescence, oxidative stress, and proliferation of human endometrial stromal cells (HESCs). RNA sequencing was employed to identify a key downstream target gene regulated by CBLL1. We found that CBLL1 was upregulated in the decidua of RSA patients. Additionally, overexpression of CBLL1 promoted HESC senescence, increased oxidative stress levels, and inhibited proliferation. Phosphatase and tensin homolog located on chromosome 10 (PTEN) was identified as one of the important downstream target genes of CBLL1. Invivo experiments demonstrated that CBLL1 overexpression in the endometrium caused higher embryo absorption rate in mice. Consequently, elevated CBLL1 expression is a potential cause of RSA, representing a novel therapeutic target for RSA.

BMP4 in Human Endometrial Stromal Cells Can Affect Decidualization by Regulating FOXO1 Expression.

Recurrent spontaneous abortion (RSA) is defined as the loss of 2 or more consecutive intrauterine pregnancies with the same sexual partner in the first trimester. Despite its significance, the etiology and underlying mechanisms of RSA remain elusive. Defective decidualization is proposed as one of the potential causes of RSA, with abnormal decidualization leading to disturbances in trophoblast invasion function. To assess the role of bone morphogenetic protein 4 (BMP4) in decidualization and RSA. Decidual samples were collected from both RSA patients and healthy controls to assess BMP4 expression. In vitro cell experiments utilized the hESC cell line to investigate the impact of BMP4 on decidualization and associated aging, as well as its role in the maternal-fetal interface communication. Subsequently, a spontaneous abortion mouse model was established to evaluate embryo resorption rates and BMP4 expression levels. Our study identified a significant downregulation of BMP4 expression in the decidua of RSA patients compared to the normal control group. In vitro, BMP4 knockdown resulted in inadequate decidualization and inhibited associated aging processes. Mechanistically, BMP4 was implicated in the regulation of FOXO1 expression, thereby influencing decidualization and aging. Furthermore, loss of BMP4 hindered trophoblast migration and invasion via FOXO1 modulation. Additionally, BMP4 downregulation was observed in RSA mice. Our findings highlighted the downregulation of BMP4 in both RSA patients and mice. BMP4 in human endometrial stromal cells was shown to modulate decidualization by regulating FOXO1 expression. Loss of BMP4 may contribute to the pathogenesis of RSA, suggesting potential avenues for abortion prevention strategies.

“White spots” in the diagnostic search for the causes of recurrent miscarriage

Recurrent pregnancy loss is not only a medical, but also a social problem of a multifactorial nature. This problem remains unsolvable despite the constant search and application of new diagnostic and therapeutic strategies. The presented review examines the literature data on evidence-based methods for diagnosing the causes of miscarriage, as well as methods whose use remains controversial. Special attention is paid to the immunological foundations of a normal pregnancy formation, the violation of which creates the prerequisites for a recurrent miscarriage. The paper also analyzes current data on the role of endometrial receptivity and the contribution of the male factor to the problem of miscarriage, and presents prospects for further research on the problem of recurrent pregnancy loss.

Dysregulation of CREB5 Impairs Decidualization and Maternal-Fetal Interactions by Inhibiting Autophagy in Recurrent Spontaneous Abortion.

Clinically, recurrent spontaneous abortion (RSA) is a pregnancy illness that is difficult to treat. Impaired decidualization is a documented cause of RSA, but the etiology and mechanism are still unknown. cAMP-responsive element binding protein 5 (CREB5) is a member of the ATF/CREB family. CREB5 has been reported to be related to pathological pregnancy, but there are few related studies on this topic in patients with RSA, and the underlying mechanism is unclear. We collected decidual tissues from RSA patients and healthy pregnant women to measure the expression level of CREB5, PRL, IGFBP1, ATG5, LC3B, and SQSTM/p62. Then, the changes in CREB5 expression and autophagy levels were measured in human endometrial stromal cells (hESCs) during decidualization. The expression levels of PRL and IGFBP1 were tested in sh-CREB5/ov-CREB5 hESCs after decidualization induction, and the autophagy level in sh-CREB5/ov-CREB5 hESCs was measured without decidualization induction. The decidualization ability of sh-CREB5 and ov-CREB5 hESCs treated with an autophagy inducer or inhibitor was measured. To investigate the effect of CREB5 in hESCs on the invasion and migration of HTR8/SVneo cells, we performed a coculture experiment. Finally, we examined the expression of CREB5 and autophagy key proteins in mouse decidual tissues by constructing an abortion mouse model. In our study, we found that the expression of CREB5 was unusually elevated in the uterine decidua of RSA patients, but the expression of PRL, IGFBP1, and autophagy were decreased. During the decidualization of hESCs, the expression of CREB5 gradually decreases in a time-dependent manner with increasing autophagy. Moreover, by knocking down or overexpressing CREB5 in hESCs, it was found that CREB5 can impair decidualization and reduce autophagy in hESCs. Furthermore, the damage caused by CREB5 in terms of decidualization can be reversed by the addition of an autophagy inducer (rapamycin). In addition, CREB5 can increase the secretion of proteins (IL-1β and TGF-β1) in hESCs to inhibit trophoblast invasion and migration. Our data support the supposition that CREB5 disturbs the decidualization of endometrial stromal cells and interactions at the maternal-fetal interface by inhibiting autophagy and that its abnormal upregulation and dysfunction may lead to RSA. It may function as a diagnostic and therapeutic target for RSA. Similarly, we found that in the spontaneous abortion mouse model, the expression of CREB5 in the decidual tissue of the abortion group was significantly increased, and autophagy was decreased.

Molecular cytogenetic analysis of multi-miscarriage products of conception in clinical cases from Al-Anbar Governorate, west of Iraq

Most clinical miscarriages often occur throughout the first trimester of pregnancy, with fetal chromosomal abnormalities being identified as the primary reason for such occurrences. The objective is to analyze the fetal chromosomal aberrations in the product of conception among Iraqi patients suffering from recurrent miscarriages. The cross-sectional study was performed on 60 cases of products of conception in women suffering from multiple miscarriages, obtained from Department of Obstetrics and Gynecology is located in Ramadi Teaching Hospital for Child and Maternity, as well as other Private Clinics in the Ramadi City. Long-term culture of conventional cytogenetic analysis using the G-banding technique was employed to determine the chromosomal disorder of fetal tissue part or villus samples. Fetal chromosomal abnormalities were detected in 86.7 %. Numerical chromosomal abnormalities were revealed in 98.1 %, while structural abnormalities were detected in 1.9 %. Additionally, the commonest gestation loss occurs in parents under 35 years in the first trimester (92.3 %). Trisomy 21 was the most frequent (46.2 %) in gestational loss. Fetal chromosomal abnormalities have been linked with gestational loss in Iraqi couples. Therefore, it is recommended that cytogenetic analysis should be performed to identify the genetic cause of recurrent miscarriage. This is important for providing appropriate genetic counseling and educating couples about the risk of future pregnancies.

Preconceptual administration of doxycycline in women with recurrent miscarriage and chronic endometritis: protocol for the Chronic Endometritis and Recurrent Miscarriage (CERM) trial, a multicentre, double-blind, placebo-controlled, adaptive randomised trial with an embedded translational substudy

IntroductionRecurrent miscarriage is a common condition with a substantial associated morbidity. A hypothesised cause of recurrent miscarriage is chronic endometritis (CE). The aetiology of CE remains uncertain. An association between...

#216 : High Live Birth Rates After Low Dose Lymphocyte Immunization Therapy (LD-LIT) with Partner Lymphocytes in Recurrent Miscarriage Patients

Background and Aims: Recurrent Miscarriage (RM) affects 1–2% of couples trying to conceive, with 50% remaining unexplained. Meta-analyses suggest LIT improves pregnancy outcomes for RM couples. However, results are conflicting due to differing criteria and protocols. This study evaluates low dose LIT effectiveness in patients with unexplained RM and Th1/Th2/Treg paradigm disorders. Method: Self-controlled, prospective study of 23 RM patients receiving partner lymphocyte immunization from August 2021 to October 2022, with [Formula: see text]3 miscarriages less than 12 weeks when endometrium at ET was [Formula: see text]8mm and PGT embryos were used. Patients with abnormal Th1/Th2/Treg cell proportions and biomarkers were treated with paternal lymphocytes twice before embryo transfer (ET) and once post-pregnancy confirmation. Results: Significant decrease in Th1 and increase in Th2 cytokines like TNF-a and IFN-g and significantly increase in Treg cells and TGF-b noted and blocking antibodies detected in all patients after LIT therapy. 20/23 (87%) post LIT conceived and 4 delivered healthy full-term babies (17%) and 3 patients have yet to undergo ET. 48% (11/23) uneventful ongoing pregnancies crossed [Formula: see text]14 weeks. 3 patients had miscarriages (13%). 1 patient (4%) did not conceive. 1(4%) conceived spontaneously ([Formula: see text]25 wks) and 1(4%) opted to try to conceive naturally. Cumulative uneventful pregnancy outcome 65%. Conclusion: LIT was associated with high live birth rates, especially in women with RM. It not only ameliorates patient’s cellular immune function, but also further increases patient’s pregnancy success rate with high safety, which is worthy of clinical application and promotion. Being small size self-controlled study, this result needs to be validated with large we’ll planned RCTs. LIT may be considered a safe and effective treatment for RM/RIF in IVF patients in individual cases based on immunoprofiling (Th1/Th2/Treg paradigm disorders), after ruling out other potential causes of RM or RIF.

Prospective study on biochemical markers causing recurrent miscarriages in Iraqi women of reproductive age

Objective: To compare the levels of elemental calcium, vitamin D3, malondialdehyde and super oxide dismutaseenzymes in serum of women having recurrent miscarriages with normal healthy pregnant women.Method: The prospective study was conducted at Al-Karkh Maternity Hospital, Baghdad, Iraq, and Al-Alawia MaternityHospital, Baghdad, Iraq, from February to August 2022, and comprised adult pregnant women having recurrentmiscarriages without being hypertensive or diabetic in group A and normal healthy pregnant women in control groupB. Blood samples were obtained from all the subjects. Besides, placental tissues were obtained from some membersfrom both the groups. Levels of elemental calcium, vitamin D3, malondialdehyde and superoxide dismutase enzymesin serum were evaluated and compared between the groups. Malondialdehyde and super oxide dismutase enzymesin the placental tissues were also compared. Data was analysed using SPSS 20.Results: Of the 140 women, 70(50%) were in each of the 2 groups. The overall age range of the sample was 18-43years. Placental tissues were obtained from 40(57.14%) in group A and 30(42.85%) in group B. In group A, miscarriagewas reported in the first trimester in 45(62.3%) cases, 21(30%) in the second trimester and 4(5.7%) in the third trimester.Mean level of serum elemental calcium, vitamin D3, malondialdehyde and super oxide dismutase were significantlydifferent between the groups (p<0.05), and the same was the case with the placental tissues (p<0.05).Conclusion: The causes of recurrent miscarriage in women could be attributed to changes in the levels of elementalcalcium, vitamin D3, super oxide dismutase and malondialdehyde in serum and placental tissues.Keywords: Pregnancy, Abortion, Spontaneous, Malondialdehyde, Cholecalciferol, Placenta, Vitamins, Oxides.

Heparin for Women With Recurrent Miscarriage and Inherited Thrombophilia (ALIFE2): An International Open-Label, Randomized Controlled Trial

ABSTRACT Thrombophilia has been implicated in the cause of recurrent miscarriage, which affects approximately 3% of couples trying to conceive. International professional guidelines recommend heparin anticoagulation for antiphospholipid syndrome, an acquired thrombophilia responsible for approximately 15% of recurrent miscarriage, but not for other inherited thrombophilias due to an absence of evidence. Many clinicians prescribe heparin to women with recurrent miscarriage and inherited thrombophilia despite the professional recommendations. This international, open-label, randomized controlled trial aimed to compare the effect of low-molecular weight heparin (LMWH) and standard pregnancy surveillance on livebirth rates in women with recurrent miscarriage and inherited thrombophilia. Women aged 18–42 years with recurrent miscarriages who were attempting to conceive or less than 7 weeks pregnant and had an inherited thrombophilia were recruited across 40 hospitals in 5 countries. Women were randomly assigned to LMWH or no LMWH in a 1:1 ratio. Women randomized to LMWH self-administered it once a day subcutaneously, beginning as soon as possible after a positive pregnancy test and before 7 weeks of gestation and continuing throughout pregnancy. The primary study outcome was livebirth after 24 weeks of gestation. Livebirth was compared across randomized treatment groups using an χ2 test with continuity correction, then a sensitivity analysis with logistic regression to adjust for stratification factors. A total of 326 women were randomized between August 2012 and January 2021. Of these, 164 were randomized to LMWH plus standard care and 162 to standard care alone. In the standard care group, 30 patients ultimately received LMWH for thromboprophylaxis per professional treatment guidelines. The mean age of participants was 33 years, and the median number of miscarriages before randomization was 3 (interquartile range, 2–4), with two thirds of patients having a history of 3 or more miscarriages. The most common thrombophilia diagnoses were heterozygosity for factor V Leiden, prothrombin G20210A mutation, and protein S deficiency. The livebirth rate was 72% (116/162) in the LMWH group, and 71% (112/158) in the standard care group, and no statistical significant was detected between the groups even after adjustment (odds ratio, 1.08; 95% confidence interval, 0.65–1.78; P = 0.77). No differences in adverse pregnancy outcomes or complications were observed between the groups. Easy bruising was reported by 45% (73) women in the LMWH group and 10% (16) in the standard care group. This randomized controlled trial demonstrates that although LMWH is safe in women with recurrent pregnancy loss and inherited thrombophilia, it does not result in an increased live birth rate compared with standard pregnancy surveillance.

Analysis of chromosomal structural variations in patients with recurrent spontaneous abortion using optical genome mapping.

Background and aims: Certain chromosomal structural variations (SVs) in biological parents can lead to recurrent spontaneous abortions (RSAs). Unequal crossing over during meiosis can result in the unbalanced rearrangement of gamete chromosomes such as duplication or deletion. Unfortunately, routine techniques such as karyotyping, fluorescence in situ hybridization (FISH), chromosomal microarray analysis (CMA), and copy number variation sequencing (CNV-seq) cannot detect all types of SVs. In this study, we show that optical genome mapping (OGM) quickly and accurately detects SVs for RSA patients with a high resolution and provides more information about the breakpoint regions at gene level. Methods: Seven couples who had suffered RSA with unbalanced chromosomal rearrangements of aborted embryos were recruited, and ultra-high molecular weight (UHMW) DNA was isolated from their peripheral blood. The consensus genome map was created by de novo assembly on the Bionano Solve data analysis software. SVs and breakpoints were identified via alignments of the reference genome GRCh38/hg38. The exact breakpoint sequences were verified using either Oxford Nanopore sequencing or Sanger sequencing. Results: Various SVs in the recruited couples were successfully detected by OGM. Also, additional complex chromosomal rearrangement (CCRs) and four cryptic balanced reciprocal translocations (BRTs) were revealed, further refining the underlying genetic causes of RSA. Two of the disrupted genes identified in this study, FOXK2 [46,XY,t(7; 17)(q31.3; q25)] and PLXDC2 [46,XX,t(10; 16)(p12.31; q23.1)], had been previously shown to be associated with male fertility and embryo transit. Conclusion: OGM accurately detects chromosomal SVs, especially cryptic BRTs and CCRs. It is a useful complement to routine human genetic diagnostics, such as karyotyping, and detects cryptic BRTs and CCRs more accurately than routine genetic diagnostics.

Imbalance of immunoregulatory molecular cellular mechanisms as a cause of recurrent miscarriage

Introduction. Recurrent pregnancy loss (RPL) is an early spontaneous termination of pregnancy before 20 weeks, which is defined as two or more miscarriages. Most of the known causes associated with the pathophysiology of RPL include endocrine disorders, antiphospholipid syndrome, intrauterine infection, anatomical defects of the uterus, etc. However, in approximately 50% of cases, the cause of the pathogenesis of RPL remains unclear and may be associated with a violation of immune mechanisms, such as maternal tolerance to fetal alloantigens and controlled inflammation, which play a decisive role in successful pregnancy. Purpose of the review. To summarize current knowledge about the molecular cellular immune mechanisms that ensure the induction and maintenance of maternal-fetal tolerance and highlight the association between impaired immunoregulation and the development of RPL. Methods. The materials were the results of research on the topic over the past 23 years. Publications included in the Pubmed and eLibrary.ru databases were analyzed. Results. This review provides information about the immune regulation of pregnancy, which is carried out through the interaction of molecular mediators and effector cells of the innate and adaptive immunity. Current evidence points to a key role of the immune system in the pathophysiology of RPL. Successful pregnancy requires a finely regulated and tightly controlled balance between immune activation and tolerance to fetal antigens. Conclusion. The main events occur in the uteroplacental zone, where trophoblast cells and maternal lymphocytes come into close contact. The search for biomarkers for pregnancy complications is the focus of scientists.

P-529 High Live Birth Rates After Low Dose Immunization Therapy (LIT) Pre and Post Pregnancy With Partner Lymphocytes In Patients With Recurrent Miscarriage (RM)

Abstract Study question Could Lymphocyte Immunization Therapy effectively improve pregnancy and live birth rate in Unexplained Recurrent Miscarriage patients? Summary answer The use of paternal lymphocytes pre and post pregnancy significantly increases the pregnancy success and Live Birth Rate In Patients With Recurrent Miscarriage (RM) What is known already RM affects 1–2% of couples who attempt to conceive and has been defined as three consecutive pregnancy losses within 20 weeks of pregnancy confirmation. Although many potential causes have been established for RM, about 50% of these remain idiopathic and unexplained owing to immunology factors. Meta-analyses have observed better effectiveness and safety of LIT in treating couples with RM shown improvement in pregnancy outcomes. Still, results are conflicting due to different screening criteria and therapeutic protocols. Objective of present study is to evaluate effectiveness of low dose LIT in patients with uRM and Th1/Th2/Treg paradigm disorders Study design, size, duration In self-controlled, prospective study, after IEC clearance and obtaining Informed consent, 19 RM patients underwent immunization with partner lymphocytes from August 2021 to October 2022. Inclusion criteria were at least three clinical pregnancies that culminated in abortion before 20 weeks of gestation or at least three cycles of IVF/ICSI with at least two PGT+ embryos and >8mm endometrium during ET and no pregnancy, normal coagulation tests, autoimmune antibodies negative, normal couple karyotyping. Participants/materials, setting, methods Maternal BMI of 30 and age of 45 was the upper limit with peripheral blood Th1/Th2/Treg cells proportions and concentrations of TNF-a, IFN-g, TGF-β1, IL-2, IL-4, IL-6, IL-10, and blocking antibodies were detected using flowcytometry/ELISA as per Seragen’s Immuno profiling platform. 30-50ml paternal blood was used to purify lymphocytes which were intradermally administered to the wife before Embryo Transfer and post-pregnancy confirmation. Main results and the role of chance Proportion of Th1 cells was significantly decreased while the proportions of Th2 cells and Treg cells were significantly increased in immunotherapy patients after treatment. In addition, the concentration of TGF-β1 in serum was significantly higher after immunotherapy than before. The concentration of TNF-a, IFN-g was significantly high before therapy were significantly decreased after therapy. LIT effectively induced the production of blocking antibodies in all the patients. 16/19 (84%) uRM patients underwent ET post LIT became pregnant and 4 of the pregnant patient had delivered healthy babies (21%). 42% (8/19) of patients have reported uneventful ongoing pregnancies crossed >14weeks. 4 patients who were pregnant for more than 12-14 weeks had miscarriages (21%). 1 patient (5%) did not conceive. 1 patient (5%) conceived spontaneously and crossed 20 weeks of uneventful gestation. 1 patient (5%) opted to try to conceive naturally hence ET is not planned. The cumulative positive uneventful pregnancy outcome was 68%, approximately the same as the figures reported earlier. LIT is associated with high live birth rates, especially in women with RM. It not only ameliorates patient’s cellular immune function but also further increases patient’s pregnancy success rate with high safety, which is worthy of clinical application and promotion. Limitations, reasons for caution This prospective self-control study with small sample size lacks randomized control group. Beneficial effects observed in this study are in sync with published data. Larger study with patients recruited based on inclusion criteria of unexplained recurrent miscarriage due to immunological factors is proposed for recommending LIT as routine therapy. Wider implications of the findings Our study findings supports LIT as a beneficial treatment for RM/RIF in IVF patients. LIT may be considered safe and effective therapy in individual cases and based on the immunoprofiling (Th1/Th2/Treg paradigm disorders), after all other potential causes of RM or RIF have been ruled out. Trial registration number Not applicable

P-504 Pregnancy outcome of intravenous immunoglobulin in women with recurrent pregnancy loss

Abstract Study question Does intravenous immunoglobulin(IVIG) increase live births in recurrent miscarriage(RM) women with cellular immune abnormality such as high NK cell level and cytotoxicity and TNF-α/IL-10 ratio? Summary answer IVIG treatment increased the live birth rate of RM women with abnormal cellular immunity. What is known already Various causes of RM have been identified but even with a detailed evaluation, almost half of the cases have unidentified etiologies. Immune imbalance is one of the proposed potential etiologies of these idiopathic RM. To regulate abnormal cellular immunity, treatment such as IVIG is proposed to improve pregnancy outcomes. However, the use of IVIG in RM is still controversial. In addition, one of the widely used indicators of immune imbalance in pregnancy is the NK cell level, but there is a disagreement in setting the cut-off value. The percentage varies from 12 to 18%. Study design, size, duration RM was defined as women with two or more spontaneous abortions and in total, 987 RM women visited Department of Obstetrics and Gynaecology, Konyang University Hospital(KYUH) from January 2007 to December 2020. Only those with a full evaluation and known treatment outcome were included. A total 204 idiopathic RM and 209 RM women with known aetiology were enrolled. Participants/materials, setting, methods We investigated the pregnancy outcome by sorting the patients into seven subgroups depending on abnormal cellular immunity including NK cell level, NK cell cytotoxicity and TNF-α/IL-10 ratio. Then, to verify the cut-off value(16.1%) of NK cell level which we set in our previous study, patients were classified into three groups according to their NK cell level: 1) <12%(low), 2) 12-16%(moderate), 3) >16%(high). Main results and the role of chance Among all RM women with at least one abnormal cellular immunity were treated with IVIG and the overall live birth rate (LBR) was 80.2%. The group which did not have IVIG treatment showed an overall LBR of 78.0%. Within the 7 groups with abnormal cellular immunity, the group with both high NK cell toxicity and TNF-α/IL-10 ratio showed the highest LBR, 100% of LBR and the group with both high NK cell level and TNF-α/IL-10 ratio showed the lowest treatment outcome, 71.4%. To verify the adequacy of our cut-off value, after excluding those with high NK cell cytotoxicity and TNF-α/IL-10 ratio, 152 patients were classified into low, moderate, and high level of NK cells. LBR were 81.0%, 72.5%, and 76.9%. Limitations, reasons for caution The study was designed retrospectively, resulting in numerous follow-up losses. It is a single-centered study that may not be enough to generalize the diagnosis method and treatments from these findings. Moreover, IVIG is an expensive drug that may lead to certain patient preferences. Wider implications of the findings The study may provide evidence in selecting RM patients with immune abnormalities to be treated with IVIG and raise their pregnancy outcomes. We can also apply these results to the reproductive failure group. A detailed evaluation and an evidence-based treatment are important. Trial registration number not applicable

Recurrent Pregnancy Loss Etiology, Risk Factors, Diagnosis, and Management. Fresh Look into a Full Box.

Recurrent pregnancy loss is a complex health challenge with no universally accepted definition. Inconsistency in definitions involves not only the number of spontaneous abortions (two or three) that are accepted for recurrent pregnancy loss but the types of pregnancy and gestational age at miscarriage. Due to the heterogeneity of definitions and criteria applied by international guidelines for recurrent pregnancy loss, the true incidence of recurrent miscarriage, which is reported to range from 1% to 5%, is difficult to estimate. Moreover, the exact etiology of recurrent pregnancy loss remains questionable; thus, it is considered a polyetiological and multifactorial condition with many modifiable and non-modifiable factors involved. Even after thoroughly evaluating recurrent pregnancy loss etiology and risk factors, up to 75% of cases remain unexplained. This review aimed to summarize and critically analyze accumulated knowledge on the etiology, risk factors, relevant diagnostic options, and management approach to recurrent pregnancy loss. The relevance of various factors and their proposed roles in recurrent pregnancy loss pathogenesis remains a matter of discussion. The diagnostic approach and the management largely depend on the etiology and risk factors taken into consideration by a healthcare professional as a cause of recurrent miscarriage for a particular woman or couple. Underestimation of social and health consequences of recurrent pregnancy loss leads to compromised reproductive health and psychological well-being of women after miscarriage. Studies on etiology and risk factors for recurrent pregnancy loss, especially idiopathic, should be continued. The existing international guidelines require updates to assist clinical practice.

Expression of Apoptotic and Ubiquitin-Proteasome Genes Are Altered in Blastocoel Fluid From Preimplantation Euploid IVF Embryos That Result in Miscarriage [ID: 1365769

INTRODUCTION: Infertility is one of the leading causes of recurrent miscarriage, affecting approximately 14% of reproductive-aged patients. Recurrent miscarriages are presumed to be the result of embryonic chromosomal abnormalities such as aneuploidy. Literature has indicated that apoptotic mechanisms may play a role in selective elimination of aneuploid cells during preimplantation embryo development. This study aims to identify the expression patterns of an apoptotic gene and a gene encoding a ubiquitin ligase within blastocoel fluid-conditioned media from preimplantation euploid in vitro fertilization (IVF) embryos that resulted in miscarriage to identify specific genes associated with poor implantation outcomes. METHODS: Candidate genes were identified through whole-transcriptome (RNA-seq) analysis of blastocoel fluid from euploid IVF embryos that resulted in successful or unsuccessful implantation. Blastocoel fluid from euploid IVF embryos resulting in miscarriage or successful implantation was selected for specific gene expression analysis using RT-qPCR and Taqman-specific primers. BCL2L12 and SHARPIN gene expression was assessed, compared to GAPDH control. RESULTS: Expression of BCL2L12, an antiapoptotic gene, and SHARPIN, a gene that encodes an ubiquitin ligase, was altered in blastocoel fluid samples associated with miscarriage. Initial studies indicate that SHARPIN has a higher expression in blastocoel fluid from samples associated with negative implantation outcomes. CONCLUSION: Identification of gene expression patterns during preimplantation development that are associated with miscarriage may allow for an additional embryo quality metric to improve the success of each IVF embryo transfer potentially reducing the number of miscarriages. Additionally, these metrics could be expanded to allow for the identification of potential high-risk pregnancies soon after implantation, without any harm to the fetus.

Cytogenetic Analysis of Patients with Recurrent Miscarriages

Objective: To evaluate the cytogenetic analysis of patients with recurrent miscarriages. Study design and setting: Cross-sectional study, Department of Hematology, Armed Forces Institute of Pathology, Rawalpindi from February 2022 to August 2022. Methodology: 196 patients (98 couples) of recurrent miscarriages within the reproductive age group were included. Cases with known anatomical or endocrinal causes of recurrent miscarriages were excluded. Couples with abnormal reproductive tract anatomy or abnormal endocrine functions were excluded. A standardized system for human cytogenetic nomenclature was used for identifying all chromosomal aberrations. Axioscope microscopes (MetaSystems, Germany) were used for visualizing the metaphases, and MetaSystems software (MetaSystems, Germany) was used to determine the karyotype of each metaphase. Data were analyzed using the student t-test and Chi-square test. A p-value =0.05 was considered significant. Results: Of 98 couples, most of the couples experienced 3 miscarriages. The difference in ages between males and females was significant (p-value <0.001). Chromosomal abnormalities were found in 7 (7.2%) of females and 5 (5.2%) of males. Positive family history of RPL was noted in 27 (13.8%) of the participants. A total of 12/196 (6.1%) males and females experiencing RPL had chromosomal anomalies. Out of these 1 individual (0.5%) had structural aberration, 1(0.5%) numerical abnormality, and 10 (5.1%) were found to have Chromosome Polymorphism. Conclusions: Translocations, numerical aberrations, and chromosomal polymorphism are common cytogenetic abnormalities noted in cases with RPL. Clinicians should refer such couples for karyotyping to rule out the possible genetic causes of recurrent miscarriages