Role In Recurrent Pregnancy Loss Research Articles

Regulatory role of miR-125a expression with respect to its target genes LIFR, ERBB2 and STAT3 in the pathogenesis of recurrent pregnancy losses.

Studies have investigated miR-125a for its predictable role in recurrent pregnancy loss (RPL) cases to regulate many biological events required for the maintenance of pregnancy by regulating its confirmed target genes LIFR, ERBB2 and STAT3. The present study included 40 cases of women with at least two RPLs in ≤20 weeks of gestation against 40 healthy multiparous women without a previous history of abortion. Expression analysis of ERBB2, LIFR, STAT3 and miR-125a was conducted by quantitative real-time PCR (qPCR). The expression of miR-125a was significantly lower in the plasma of RPL cases (P = 0.0001) and showed a significantly increased mean expression level in product of conception (2.56-fold, P < 0.0001). Among the target gene of miR-125a, ERBB2 and STAT3 gene expression level was significantly increased (2.58-fold, P = 0.04; 1.87-fold, P = 0.025), respectively in RPL cases while the LIFR gene revealed comparable expression (P = 0.64). Furthermore, expression analysis of ERBB2 gene with respect to its regulatory miR-125a cases depicted a significant association (P = 0.0005). Kaplan-Meier survival analysis revealed cases with low miR-125a expression had significantly shorter time to miscarriages, (log-rank P = 0.02). Also, decreased expression of miR-125a significantly conferred >2-fold increased risk for RPL (HR = 2.34: P < 0.05). The overall conclusion of the study was that altered miR-125a expression may cause deregulation in target genes LIFR, ERBB2 and STAT3 resulting in adverse consequence in the outcome of pregnancy.

Association between semen parameters and recurrent pregnancy loss: An umbrella review of meta-analyses.

Recurrent pregnancy loss (RPL) is a common clinical reproductive problem. With research advancements, an increasing number of studies have suggested that male factors play an important role in RPL. However, the evaluation results of male sperm quality in published meta-analyses are inconsistent. We aimed to summarize the evidence of the association between semen factors and RPL and evaluate the level and validity of the evidence. We searched PubMed, Cochrane Library, EMBASE, Web of Science, and Scopus databases for systematic reviews or meta-analyses to evaluate the association between male semen parameters and RPL. The methodological quality of the included meta-analyses was assessed, and data and evidence were re-synthesized and stratified using a random-effects model. Seven meta-analyses and nine semen parameters were included in the final analysis. The methodological quality of all publications was considered low or very low. There was highly suggestive evidence for the association between sperm DNA fragmentation (SDF), sperm progressive motility rate, and RPL (class II). The evidence level for the association between sperm concentration, normal sperm morphology, sperm deformity rate, total motility, and RPL was suggestive evidence (class III). The evidence level for the association between sperm volume and sperm count and RPL was weak (class IV). There was no significant association between sperm pH and RPL (class NS). Our results suggest level II evidence for the association between male SDF and RPL, while the evidence level for the association between conventional semen routine parameters and RPL was low (classes III and IV).

Neopterin and the anti-Mullerian hormone's role in Unexplained Recurrent Pregnancy Loss

Background: Recurrent pregnancy loss were prevalent complications in pregnancy. Immunological biomarkers as Neopterin used as a marker in recurrent pregnancy loss. Anti-Mullerian hormone is a reliable marker of the ovarian reserve. Therefore, was also considered as a marker to evaluate its role in the recurrent pregnancy losses. This study was conducted to assess whether neopterin and Anti- Mullerian hormone can be used as a marker in the condition of unexplained recurrent pregnancy loss. Material’s and Methods: A case control study was done in the Department of Obstetrics and Gynecology Salahdeen general hospital in Tikrit city from first January to thirty of August 2020. One hundred patient enrolled in the study. Study contains two groups:1- Group A as cases. 2- Group B as control group. The data collection done through:1-Designed closed and open-ended questionnaire, 2- Laboratory investigations of : Serum Neopterin and Anti-Mullerian hormone levels using enzyme-linked immunosorbent assay. Results: Mean Neopterin level was significantly higher among cases (24.3±10.7)nml/l than control group (2.9±1.1) nml/l. Mean Neopterin level increased with increasing abortion number. It was significantly higher among those with ≥5 abortions (30.02±10.04) nml/l than those with (3-4) and 2 abortions (28.7±15.29) nml/l and (20.1±0.03) nml/l. The mean Neopterin level increased with increasing age among cases and controls. Among cases was (28.7±12.03) nmol/L among those aged (31- 35) year, which was higher than those aged (18-25), and those aged (26-35) years Mean Anti-Mullerian hormone level was significantly lower among cases (0.8±0.6) ng/ml than control group (5.01±2.7) ng/ml. Mean Anti-Mullerian hormone level was significantly higher among those with 2 abortions (1.06±0.7) ng/ml than those with (3-4) and 5 abortions (0.6±0.56) ng/ml and (0.97±0.4) ng/ml respectively. Among case the mean AMH level was lower among those aged (31-35) years (0.5±0.31) ng/ml than those aged (26-30), and (18-25) years (0.8±0.7) ng/ml, (0.97±0.7) ng/ml, respectively Conclusions: A statistically significant high Mean Neopterin level among cases and its increased mean Neopterin level with increasing abortion number. A statistically significant low levels of mean Anti-Mullerian hormone level among cases, more reduction in its level with increasing abortion. This pioneer study in Iraq, determined the significant association between the unexplained recurrent pregnancy loss with the increased levels of Neopterin and its effect on Anti-Mullerian hormone that lead to recurrent pregnancy loss.

BBOX1-AS1 mediates trophoblast cells dysfunction via regulating hnRNPK/GADD45A axis†.

Recurrent pregnancy loss (RPL) is a common pathological problem during pregnancy, and its clinical etiology is complex and unclear. Dysfunction of trophoblasts may cause a series of pregnancy complications, including preeclampsia, fetal growth restriction, and RPL. Recently, lncRNAs have been found to be closely related to the occurrence and regulation of pregnancy-related diseases, but few studies have focused on their role in RPL. In this study, we identified a novel lncRNA BBOX1-AS1 that was significantly upregulated in villous tissues and serum of RPL patients. Functionally, BBOX1-AS1 inhibited proliferation, migration, invasion, tube formation and promoted apoptosis of trophoblast cells. Mechanistically, overexpression of BBOX1-AS1 activated the p38 and JNK MAPK signaling pathways by upregulating GADD45A expression. Further studies indicated that BBOX1-AS1 could increase the stability of GADD45A mRNA by binding hnRNPK and ultimately cause abnormal trophoblast function. Collectively, our study highlights that the BBOX1-AS1/hnRNPK/GADD45A axis plays an important role in trophoblast-induced RPL and that BBOX1-AS1 may serve as a potential target for the diagnosis of RPL.

Molecular subtype identification and predictive power of N6-methyladenosine regulator in unexplained recurrent pregnancy loss.

The etiology of recurrent pregnancy loss (RPL) is complicated and effective clinical preventive measures are lacking. Identifying biomarkers for RPL has been challenging, and to date, little is known about the role of N6-methyladenosine (m6A) regulators in RPL. Expression data for m6A regulators in 29 patients with RPL and 29 healthy controls were downloaded from the Gene Expression Omnibus (GEO) database. To establish a diagnostic model for unexplained RPL, differential gene expression analysis was conducting for 36 m6A regulators using least absolute shrinkage and selection operator (LASSO) regression. Unsupervised cluster analysis was conducted on hub genes, and probable mechanisms were explored using gene set enrichment analysis (GSEA) and gene ontology (GO) analysis. Correlations between m6A-related differentially expressed genes and immune infiltration were analyzed using single-sample GSEA. A total of 18 m6A regulators showed significant differences in expression in RPL: 10 were upregulated and eight were downregulated. Fifteen m6A regulators were integrated and used to construct a diagnostic model for RPL that had good predictive efficiency and robustness in differentiating RPL from control samples, with an overall area under the curve (AUC) value of 0.994. Crosstalk was identified between 10 hub genes, miRNAs, and transcription factors (TFs). For example, YTHDF2 was targeted by mir-1-3p and interacted with embryonic development-related TFs such as FOXA1 and GATA2. YTHDF2 was also positively correlated with METTL14 (r = 0.5983, p < 0.001). Two RPL subtypes (Cluster-1 and Cluster-2) with distinct hub gene signatures were identified. GSEA and GO analysis revealed that the differentially expressed genes were mainly associated with immune processes and cell cycle signaling pathway (normalized enrichment score, NES = -1.626, p < 0.001). Immune infiltration was significantly higher in Cluster-1 than in Cluster-2 (p < 0.01). In conclusion, we demonstrated that m6A modification plays a critical role in RPL. We also developed and validated a diagnostic model for RPL prediction based on m6A regulators. Finally, we identified two distinct RPL subtypes with different biological processes and immune statuses.

Comparative proteome profiling of seminal components reveal impaired immune cell signalling as paternal contributors in recurrent pregnancy loss patients.

Recurrent pregnancy loss (RPL) is usually evaluated from a women's perspective, however, recent evidence implies involvement of male factors as paternally expressed genes predominate placenta. During fertilization, prior to implantation the immune system purposefully produces early pregnancy factors with potent immunomodulatory properties for adaptation to antigenically dissimilar embryo. Therefore, it is hypothesized that paternal immunological factors play a role in RPL. Comparative proteome profiling (label free liquid chromatography mass spectroscopy: LC-MS/MS) of the seminal extracellular vesicles (SEVs), extracellular vesicle free seminal plasma (EVF-SP) and spermatozoa was carried out in semen of RPL patients (n=21) and fertile donors (n=21). This was followed by pathway and protein-protein interaction analysis, and validation of key proteins' expression (western blot). A total of 68, 28 and 49 differentially expressed proteins in SEVs, EVF-SP and spermatozoa of RPL patients, respectively, were found to be involved in inflammatory response, immune cell signalling and apoptosis. In SEVs, underexpressed GDF-15 and overexpressed C3 imply distorted maternal immune response to paternal antigens leading to impaired decidualization. Dysregulated TGFβ signalling in EVF-SP surmises defective modulation of inflammatory response and induction of immune tolerance to seminal antigens in the female reproductive tract through generation of regulatory T cells. Retained histone variants in spermatozoa construe defective expression of early paternal genes, while underexpressed PTN may inflict defective angiogenesis resulting in expulsion of decidua. Impaired modulation of immune response and improper placental development due to altered cytokine levels in seminal components may be the contributing paternal factors in RPL.

HLA sharing and maternal HLA expression in couples with recurrent pregnancy loss in Taiwan

ObjectiveThe aim of this study is to investigate the frequency and distribution of human HLA sharing and maternal HLA allele expression in couples with recurrent pregnancy loss in Taiwan. Materials and methodsWe retrospectively reviewed couples experienced two or more pregnancy loss before 20th weeks of gestation from March 2014 to November 2020 having HLA determination. Fertile individuals with one or more live-birth offspring receiving HLA allele determination during the same period were included as the control group. The distribution and frequency of HLA sharing were analyzed and presented by descriptive statistics. Fisher Exact Test were used to analyze specific maternal and paternal HLA allele comparing individuals with RPL to fertile group. P-value < 0.05 was thought to be statistically significant. Results72 couples were enrolled from March 2014 to November 2020. Regarding the distribution of HLA sharing, HLA sharing between females and their male partners less and equal to 2 pairs were found in 40.3% of the couples. HLA sharing greater and equal to 3 pairs are found in 59.8% couples. HLA sharing was most frequently found in alleles HLA-A02, A11, DQ07, C07 and B60 in descending order. There was a significant lower expression of HLA-B13 in women with RPL compared to women who had successful pregnancy (p = 0.0234). Compared infertile men with fertile men cohort, the frequency of HLA-DR04 (p = 0.0438, OR 2.444, 95% CI 1.0251–5.8287), HLA-DR12 (p = 0.001, OR 30.85, 95% CI 4.0296–236.19) and HLA-15 (p = 0.0357, OR 9.354, 95% CI 1.1610–75.37) were found to be significantly higher in men with RPL. On the contrary, HLA-DR07 (p = 0.0085, OR 0.124, 95% CI 0.0264–0.587) and HLA-DR10 (p = 0.0395, OR 0.048, 95% CI 0.0027–0.8641) were found to be significantly lower in men with RPL. ConclusionWe found a tendency to recurrent pregnancy loss in couples with more than 2 pairs of HLA sharing. The similarity of HLA sharing, the expression of maternal HLA-B13 allele and paternal HLA-DR alleles in Taiwanese couples might play a role in recurrent pregnancy loss.

Acquired thrombophilia and fetal factors associated with recurrent pregnancy loss

Recurrent pregnancy loss is the most common complication of pregnancy, is the spontaneous loss of a pregnancy before the fetus has reached viability. Improvement of pregnancy outcome is considered as an important area of action for those concerned with the improvement of women’s health and pregnancy outcome. Several studies identified thrombophilia as the principal cause of recurrent pregnancy loss. However, reported studies often do not evaluate other causes of miscarriages in their inclusion and exclusion criteria. So, the aim of this review is to investigate the role of acquired thrombophilia and fetal factors and their role in recurrent pregnancy loss.

The impact of hereditary thrombophilias in recurrent pregnancy loss

Introduction: Recurrent pregnancy loss (RPL) is defined as two or more consecutive pregnancy loss which occurs before the 20th weeks of pregnancies for the last menstrual period. Hereditary cause of thrombophilic gene mutations and polymorphism may play an essential role in RPLs. Material and Method: 291 women with a history of two or more consecutive abortions as a study group and 61 women without the history of miscarriages as a control group were included in a study. In this study we analysed the effects of Factor II Prothrombin mutation ,FV Leiden mutation, MTHFR C677T, MTHFT A1298C, PAI-1, ?-fibrinogen, Factor XIIIA (V34L) and Glycoprotein IIIa (L33P) polymorphisms on RPL by using pyrosequencing. Chi-square and multiple regression analysis were used for statistical analysis. Results: FII prothrombin mutation, FV Leiden mutation, MTHFR C677T, MTHFR A1298C, PAI1 and Beta fibrinogen were found statistically significant in the chi-square test. Heterozygous FV G1691A (OR:8.092, CI: 1.280-51.165), homozygous MTHFR A1298C (OR:17.621, CI: 3.644 - 85.203), Heterozygous MTHFR C677T (OR: 2.921 CI: 0.811-10.515), Homozygous MTHFR C677T (OR: 3.619 CI: 1.647-7.954), heterozygous MTHFR A1298C (OR: 5.989, CI: 2.574-13.934), homozygous PAI1 (OR: 8.756, CI: 2.805 -27.334), heterozygous PAI1 ( OR: 7.114, CI: 3.145- 16.096) homozygous FibrinogenG455A (4.085, CI: 1.438-11.610) were found statistically significant in logistic regression analysis for RPL(p&lt;0.05). Discussion: This study indicated that there is a significant association between thrombophilias and RPL. Therefore, it is important to detect thrombophilic mutations in RPL.

Association of the single nucleotide polymorphism C1858T of the PTPN22 gene with unexplained recurrent pregnancy loss: A case-control study

Background Lymphoid-tyrosine-phosphatase which is encoded by the protein tyrosine phosphatase non-receptor 22 (PTPN22) gene plays a pivotal role in the regulation of immune responses by dephosphorylating several signaling intermediates of immune cells.ObjectiveSince a balanced immune response has been shown to be important during pregnancy, the purpose of this research was to compare the frequency of the PTPN22 C1858T polymorphism in women with unexplained recurrent pregnancy loss (URPL) vs. in a control group for the first time.Materials and MethodsGenomic DNA from 200 individuals with URPL and 200 individuals without URPL (the control group) at the infertility center in Yazd, Iran was isolated using the salting-out method. The PTPN22 C1858T polymorphism of the two groups was analyzed using polymerase chain reaction-restriction fragment length polymorphism. Genotype frequencies in the women with URPL and the fertile control group were compared using the Chi-square test.ResultsThere were significant differences in the frequency of the PTPN22 1858T polymorphism in the URPL individuals vs. the healthy controls, i.e. 32.0% and 21.5%, respectively (p = 0.01).ConclusionOur findings suggest that the PTPN22 1858T polymorphism could play a role in recurrent pregnancy loss. Therefore, genotyping of the mentioned polymorphism can help clinicians to predict the probable risk of URPL.

Investigating the role of EGF-CFC gene family in recurrent pregnancy loss through bioinformatics and molecular approaches

ABSTRACT Recurrent pregnancy loss (RPL) is the most common reproductive failure, reaching 1-5% of women throughout their lives, and having unknown etiology in 50% of the cases. In humans, EGF-CFC1 (Epidermal Growth Factors & Cripto/FRL-1/Cryptic) gene family is composed by TDGF1 and CFC1, two developmental genes. The aim of this study was to investigate the role of EGF-CFC on RPL. To this, multiple approaches were performed; we conducted an expression analysis of TDGF1 and CFC1 using publicly available data from Gene Omnibus Expression (GEO), systems biology analyses and functional prediction; and a molecular analysis carried out in a case-control study. Our GEO analysis showed a decrease in TDGF1 expression in the endometrium (p=0.049) and CFC1 expression in placenta (p=0.015) of women with RPL. Network analysis, gene ontology and literature pointed to a strong connection between EGF-CFC1 gene family to pathways that play key roles during pregnancy, including TGF-β, c-Src/MAPK/AKT, Notch, TNFα, IFNγ and IL-6. A pathogenicity score developed for this gene family showed that the c.-14+1429T>C (rs3806702) variant in the TDGF1 and the p.Arg47Gln (rs201431919) variant in CFC1 gene would be the ones with the highest deleterious effect for RPL. In the case-control study, which involved 149 women with RPL and 159 controls, no statistical difference was observed in the allele and genotype distributions of the variants studied in the two groups. In this study, we performed extensive bioinformatics analysis for biomarker prioritization followed by experimental validation of proposed selected markers. Although there is no statistical difference in the frequencies of these variants between RPL and controls, the expression analysis results suggest that TDGF1 and CFC1 genes might play a role in RPL. In addition, systems biology analyzes raise the hypothesis that genes in other signaling pathways that may be related to RPL as good candidates for future studies. Abbreviations RPL: recurrent pregnancy loss; EGF-CFC1: Epidermal Growth Factors – Cripto/FRL-1; GEO: Gene Omnibus Expression; KEGG: Kyoto Encyclopedia of Genes and Genomes

P–424 Evaluating the relationship between endoplasmic reticulum stress and recurrent pregnancy loss in a young population cohort

Abstract Study question Does endoplasmic reticulum (ER) stress evaluated by X-box binding protein 1 (XBP–1) among patients aged 18–30 years play a role in recurrent pregnancy loss (RPL)? Summary answer High levels of XBP–1 observed in patients with known RPL indicate that ER stress plays an important role in RPL. What is known already Female reproductive tract undergoes dynamic changes during oogenesis which require protein synthesis, folding, maturation, which take place in the ER. ER is also responsible for destruction of unfolded/misfolded proteins. Excess accumulation of these faulty proteins leads to ER stress, which actives unfolded protein response (UPR). XBP–1 is a transcription factor involved in UPR and regulates ER stress-mediated apoptosis. Regulation of ER homeostasis is important in folliculogenesis, oocyte maturation and embryogenesis. It is also known that ER stress has a positive correlation with age and it is associated with age-related diseases. Study design, size, duration This prospective case-controlled study was conducted at University of Health Sciences Turkey, Istanbul Kanuni Sultan Suleyman Training and Research Hospital Department of Obstetrics and Gynecology between March 2020 – September 2020. A total of 70 subjects were included in the study. All patients gave their written informed consent before their enrollment in the study. Participants/materials, setting, methods 38 patients aged 18–30 years with a history of RPL were included in the study. Patients who had miscarriages due to fetal abnormalities, patients with infections, endocrine or genetic disorders, smokers, alcohol and/or drug abusers, with acute/chronic inflammatory diseases, patients using steroids, anti-inflammatory and antioxidant medications were excluded from the study. Age-matched 32 healthy subjects without RPL were included in the control group. XBP–1 levels were determined using Human XBP–1 ELISA Kit (Elabscience Co., USA). Main results and the role of chance The mean age in the control group and in the study group were 25.21±3.3 and 25.26±2.6, respectively and they were statistically similar (p = 0.324). When groups were compared according to thyroid stimulating hormone (TSH) levels and body mass index (BMI), which are additional risk factors of RPL both groups were statistically similar (p = 0.642 and 0.942, respectively). As expected gravidity and abortus numbers were significantly higher in the study group (p &amp;lt; 0.001). A mean XBP–1 level of 1233.41± 3902.97 was determined in the control group. The mean value of the study group was calculated to be 2251.49± 9621.12. Mean XBP–1 level in the study group was significantly high (p &amp;lt; 0.001). A receiver operating curve (ROC) analysis was conducted in the study group. The area under the curve was found to be 87% (95% CI: 79% - 95%). The specificity was 75%, sensitivity was 89%, positive LR was 3.5, negative LR was 0.15, positive predictive value was 80% and negative predictive value was 87% for the cut-off XBP–1 level at 1364.68 pg/mL. Limitations, reasons for caution Small sample size is an important limitation of this study. In addition, evaluating XBP–1 only in serum samples does not let us drive any conclusions on the local changes of ER stress. Studies with larger samples sizes and studying XBP–1 levels in tissue samples of endometrial material is needed. Wider implications of the findings: The significantly high levels of XBP–1 in RPL patients younger than 30 years, indicate higher ER stress in this group even when age dependent increase in ER stress is calculated out of the equation. XBP–1 can be a promising marker in evaluating patients with a fertility wish for RPL risk. Trial registration number NCT04455256

Association of M55L and Q192R polymorphisms of paraoxonase 1 gene (PON1) with recurrent pregnancy loss risk: A case-control study.

Background Recurrent pregnancy loss (RPL) refers to the incidence of two or more abortions before the first half of pregnancy. Oxidative stress has been hypothesized to play a central role in RPL.Objective To investigate the relationship between Q192R and L55M polymorphisms of PON1 as antioxidant enzyme and the risk of RPL.Materials and MethodsIn this case–control study, 110 women with RPL (case) and 110 healthy fertile women (control) referred to the Research and Clinical Center for Infertility, Shiraz, Iran were enrolled. Genomic DNA was extracted from the peripheral blood in all participants. Polymorphisms were genotyped by polymerase chain reaction-restriction fragment length polymorphism method.ResultsStatistical analysis of Q192R polymorphism showed a significant difference for the RR genotype between the case and control group (OR = 11, CI = 1.39–86.87, p = 0.005) but none for the QR and QQ genotypes. No significant association was observed between the R and Q allelic frequency in the RPL participants compared to the control group (p = 0.53). Also, statistical analysis of the L55M polymorphism for MM genotype in the case group compared with the control group showed a significant difference (OR = 3.59, CI = 0.97–13.30, p = 0.042), but none for the LM and LL genotypes.ConclusionThe findings showed a significant correlation between the Q192R polymorphisms and the L55M PON1 enzyme and RPL in this study population.

Genetic Testing for Aneuploidy in Patients Who Have Had Multiple Miscarriages: A Review of Current Literature.

Recurrent pregnancy loss (RPL) is an obstetrical complication that affects about 3% of reproductive age couples. Genetic and non-genetic causes of RPL are multiple; however, aneuploidy is the most common obstetrical complication that can explain single and recurrent pregnancy loss (present in about 60% of recognized clinical pregnancies which result in a miscarriage). Parental karyotyping will only be of potential benefit for 2 to 5 percentage of RPL couples who are translocation carriers. Products of conception (POC) karyotype analysis has been used to direct management in RPL and has been shown to be cost-effective, but the technique has many limitations including high culture failure rate and maternal cell contamination. These limitations can be significantly reduced using POC chromosomal microarray (CMA) technology. We believe that POC genetic testing should be performed after the second and subsequent pregnancy loss using CMA. Although the results will not generally alter the course of treatment, the knowledge of the reason for the loss is of great emotional comfort to many patients. In addition, POC CMA performed in conjunction with a regular complete maternal RPL work-up will identify the group of truly unexplained RPL. Thus, only 10% of patients with RPL will complete an evaluation having a euploid loss and an otherwise normal work-up. This group of “truly unexplained RPL” would be ideal for new research trials and therapies. Pre-implantation genetic testing (PGT) technology has improved recently with day 5 trophectoderm biopsy as compared to biopsy on day 3 as well as with the addition of CMA and next-generation sequencing technologies. The most recent studies on PGT-SR (PGT-Structural rearrangement) show improved clinical and live birth rates per pregnancy, as well as decreased miscarriage rate for translocation carriers. PGT-A (PGT-aneuploidy) may have a limited role in RPL in cases with documented recurrent POC aneuploidy.

Expression of ADAMTS13 in human endometrium and its potential role in recurrent pregnancy loss

IntroductionThe mechanisms underlying the pathogenesis of recurrent pregnancy loss (RPL) and the effective approaches to treat this disease still remain vague and absent. Proteinases of ADAMTS family play important roles in embryonic growth and development. Our previous study suggest a role of ADAMTS13 during pregnancy. Current Study was to determine the expression of ADAMTS13 in human endometrium and its association with RPL.Material and methodsThe spatiotemporal expression of ADAMTS13 in human endometrium was examined by immunohistochemistry. real-time PCR sand western blot were then employed to determine the mRNA and protein expression levels of ADAMTS13 in human endometrium. Proteolytic cleavage of FRETS-VWF73 were performed to determine the activity of ADAMTS13 in plasma and that secreted by human endometrium. ELISA was carried out to measure plasma VWF antigen.ResultsWe show that proteolytically active ADAMTS13 is expressed in human endometrium throughout the menstrual cycle and pregnancy. The decidual expression levels of mRNA and protein in women with RPL were significantly lower compared with women with uncomplicated pregnancies (P&lt;0.01, P&lt;0.05, respectively). Furthermore, significantly reduced plasma ADAMTS13 activity (median [range] 69.09 [65.2–93.7]% versus 93.62 [88.1–115.6]%, P&lt;0.001) and elevated plasma VWF antigen levels (median [range] of 125.5 [54.2–262.8]% versus 91.9[80.4–138.7]%, P &lt; 0.01) were detected in RPL patients compared with the control group.ConclusionsThese findings suggest that ADAMTS13 may play a role in embryo implantation and the pathogenesis of recurrent pregnancy loss. Further investigation on ADAMTS13 gene knockout animal models is necessary for understanding the molecular mechanisms of the biological roles of ADAMTS13 during gestation.

Genome wide methylation analysis to uncover genes related to recurrent pregnancy loss.

Recurrent pregnancy loss (RPL) refers to two or more consecutive spontaneous abortion before 24weeks of gestation, representing 1% of couples of childbearing age. Epigenetic factors including dysregulation of DNA methylation of some genes may play a role in RPL. To identify RPL related genes modulated by DNA methylation expressed in decidua and blood. Three decidua samples each from RPL patients and normal controls were recruited to perform genome-wide bisulfite sequencing (GWBS) and transcriptome sequencing. Based on the above results, 22.52kb of differential methylation regions (DMRs) from 17 genes were verified by bisulfite sequencing PCR at specific region (Hi-MethylSeq) in another 15 decidua (7RPL vs. 8 Controls) and 13 blood (5RPL vs. 8 Controls) samples. 23 genes showed significantly differential cytosine methylation status and distinct expression level between PRL patients and healthy controls synergistically. Three signaling pathways were found to be shared between genes with both hypomethylated differential methylation regions (DMR) and upregulated differential gene expression (DGE). The results from Hi-MethylSeq showed that the hypermethylation of SGK1 in both blood and decidua samples in RPL patients, which was consistent to its lower expression in endometrium reported earlier. SGK3 and CREB5 also showed modulated methylation level in RPL decidua. Our finding supported that aberrant methylation of SGK1 and CREB5 could be a cause of the dysregulation of these gens in the endometrium, which is one of cause of reproductive failure. The function of SGK3 in reproduction system deserves further investigation.

Maternal killer-cell immunoglobulin-like receptors and paternal human leukocyte antigen ligands in recurrent pregnancy loss cases in Turkey.

Objective The survival of a semi-allogeneic fetus depends on several immunological mechanisms, and it has been suggested that recurrent pregnancy loss (RPL) could develop as a result of one or more immunological abnormalities.Methods Compatibility between partners for human leukocyte antigen (HLA) genotypes and the relationships between maternal killer-cell immunoglobulin-like receptor (KIR) and paternal HLA-Bw4/Bw6 and HLA-C1/C2 supra-groups were investigated in 25 couples with RPL in comparison to healthy couples with children. HLA and KIR genotyping was performed using polymerase chain reaction with sequence-specific primers and/or sequence-specific oligonucleotides.Results HLA class I incompatibility between partners, especially in HLA-B alleles, was more common in the RPL group (p= 0.01). HLA-C2 homozygosity was more frequent in the male partners of RPL couples than in other groups (p= 0.03). The KIR2DL5 gene frequency was significantly higher in both the female and male partners of RPL couples, whereas the KIR2DS3 gene frequency in male partners of RPL couples was significantly reduced (p= 0.03). The presence of KIR2DL3 in women with RPL was correlated with the presence of HLA-C2 alleles in their spouses (p= 0.03).Conclusion Our data from a Turkish population suggest that male HLA-C2 homozygosity may play an important role in RPL. Additionally, an incidental match between male HLA-C2 and female HLA-C1 ligand KIR receptors might perturb the balance between activatory and inhibitory KIR-ligand interactions during pregnancy in couples affected by RPL. The roles of orphan KIR2DL5 and orphan KIR2DS3 in RPL remain obscure.

Protective Effects of Antioxidant Supplements on Sperm Parameters, Sperm DNA Damage and Level of Seminal ROS in RPL Patients: A Clinical Trial Study

Background: Recurrent Pregnancy Loss (RPL) is an important medical problem comprising approximately 15% of all pregnancies. Sperm DNA damage and level of seminal ROS may have a role in RPL. Objectives: The main aim of the present study was to appraise the efficacy of antioxidants for improving these parameters in RPL patients. Methods: In a clinical trial, 90 couples suffering from RPL were divided into two groups from April 2014 to December 2015 at the Yazd Research and Clinical Center for Infertility in Iran. Group I received supplemental vitamin E (400 µg) daily in combination with Se (200 µg) and the other group received vitamin E plus zinc for three months. Sperm parameters, sperm DNA damage, and seminal ROS were evaluated before and after treatment and compared between the two groups. Data were analyzed by SPSS and P values of ≤ 0.05 were considered statistically significant. Results: Our results revealed normal sperm parameters in RPL patients. Sperm parameters substantially improved in the two treated groups (P ≤ 0.05). The ROS level dramatically reduced in the two groups (P = 0.01). The evaluation of chromatin integrity with AB staining did not show any remarkable changes in group 1 (P = 0.3), but it showed a considerable rate reduction in group 2 (P = 0.04). Also, it showed significant decreases with TB, CMA3, and TUNEL assays in both groups (P = 0.001). However, we did not find any significant difference between the two groups. Conclusions: Supplemental vitamin E in combination with selenium or zinc may improve semen quality by decreasing the level of seminal ROS and sperm DNA damage in RPL patients. We advocate their use for improving the quality of the men’s sperm DNA.

FOXD1 mutations are related to repeated implantation failure, intra-uterine growth restriction and preeclampsia

BackgroundHuman reproductive disorders consist of frequently occurring dysfunctions including a broad range of phenotypes affecting fertility and women’s health during pregnancy. Several female-related diseases have been associated with hypofertility/infertility phenotypes, such as recurrent pregnancy loss (RPL). Other occurring diseases may be life-threatening for the mother and foetus, such as preeclampsia (PE) and intra-uterine growth restriction (IUGR). FOXD1 was defined as a major molecule involved in embryo implantation in mice and humans by regulating endometrial/placental genes. FOXD1 mutations in human species have been functionally linked to RPL’s origin.MethodsFOXD1 gene mutation screening, in 158 patients affected by PE, IUGR, RPL and repeated implantation failure (RIF), by direct sequencing and bioinformatics analysis. Plasmid constructs including FOXD1 mutations were used to perform in vitro gene reporter assays.ResultsNine non-synonymous sequence variants were identified. Functional experiments revealed that p.His267Tyr and p.Arg57del led to disturbances of promoter transcriptional activity (C3 and PlGF genes). The FOXD1 p.Ala356Gly and p.Ile364Met deleterious mutations (previously found in RPL patients) have been identified in the present work in women suffering PE and IUGR.ConclusionsOur results argue in favour of FOXD1 mutations’ central role in RPL, RIF, IUGR and PE pathogenesis via C3 and PlGF regulation and they describe, for the first time, a functional link between FOXD1 and implantation/placental diseases. FOXD1 could therefore be used in clinical environments as a molecular biomarker for these diseases in the near future.KeywordsRecurrent pregnancy loss, Preeclampsia, Intra-uterine growth restriction, FOXD1

Role of TRF2 and TPP1 regulation in idiopathic recurrent pregnancy loss

Telomeres are the tandem repeats (TTAGGG) present at the ends of the chromosomes that ensure chromosome stability and protect chromosomes from degradation. Telomeres in somatic human cells shorten after every cellular division and are linked to the cellular senescence. In this study we have investigated telomere length and expression of shelterin genes in aborted fetus material from idiopathic recurrent pregnancy losses. Telomere length was measured using Telomere Restriction Fragment Length (TRF) analysis. The gene expression levels for important shelterin complex proteins (TRF1, TRF2, POT1, and TPP1) were determined by Real-time Quantitative Reverse Transcriptase PCR (qRT-PCR). Our results demonstrated down regulation of TRF2 and TPP1 and a strong decline in average telomere length in abort material from women suffering from idiopathic recurrent pregnancy loss. We suggest that shorter telomere length and downregulation of the major shelterin components TRF2 and TPP1 leading to “telomere uncapping”, might play a critical role in recurrent pregnancy loss.