Trophoblast PR-SET7 dysfunction induces viral mimicry response and necroptosis associated with recurrent miscarriage.

Recurrent miscarriage (RM) is a distressing pregnancy complication. While the etiology of RM remains unclear, growing evidence has indicated the relevance of trophoblast impairment to the pathogenesis of RM. PR-SET7 is the sole enzyme catalyzing monomethylation of H4K20 (H4K20me1) and has been implicated in many pathophysiological processes. However, how PR-SET7 functions in trophoblasts and its relevance to RM remain unknown. Here, we found that trophoblast-specific loss of Pr-set7 in mice led to defective trophoblasts, resulting in early embryonic loss. Mechanistic analysis revealed that PR-SET7 deficiency in trophoblasts derepressed endogenous retroviruses (ERVs), leading to double-stranded RNA stress and subsequent viral mimicry, which drove overwhelming interferon response and necroptosis. Further examination discovered that H4K20me1 and H4K20me3 mediated the inhibition of cell-intrinsic expression of ERVs. Importantly, dysregulation of PR-SET7 expression and the corresponding aberrant epigenetic modifications were observed in the placentas of RM. Collectively, our results demonstrate that PR-SET7 acts as an epigenetic transcriptional modulator essential for repressing ERVs in trophoblasts, ensuring normal pregnancy and fetal survival, which sheds new light on potential epigenetic causes contributing to RM.

ENO1 promotes trophoblast invasion regulated by E2F8 in recurrent miscarriage.

Recurrent miscarriage (RM) is related to the dysfunction of extravillous trophoblast cells (EVTs), but the comprehensive mechanisms remain largely unexplored. We analyzed single-cell RNA sequencing (scRNA-seq), bulk RNA sequencing and microarray datasets obtained from Gene Expression Omnibus (GEO) database to explore the hub genes in the mechanisms of RM. We identified 1724 differentially expressed genes (DEGs) in EVTs from the RM, and they were all expressed along the trajectory of EVTs. These DEGs were associated with hypoxia and glucose metabolism. Single-cell Regulatory Network Inference and Clustering (SCENIC) analysis revealed that E2F transcription factor (E2F) 8 (E2F8) was a key transcription factor for these DEGs. And the expression of ENO1 can be positively regulated by E2F8 via RNA sequencing analysis. Subsequently, we performed immunofluorescence assay (IF), plasmid transfection, western blotting, chromatin immunoprecipitation (ChIP), real-time quantitative polymerase chain reaction (qRT-PCR), and transwell assays for validation experiments. We found that the expression of alpha-Enolase 1 (ENO1) was lower in the placentas of RM. Importantly, E2F8 can transcriptionally regulate the expression of ENO1 to promote the invasion of trophoblast cells by inhibiting secreted frizzled-related protein 1/4 (SFRP1/4) to activate Wnt signaling pathway. Our results suggest that ENO1 can promote trophoblast invasion via an E2F8-dependent manner, highlighting a potential novel target for the physiological mechanisms of RM.

Pyroptosis is involved in the immune microenvironment regulation of unexplained recurrent miscarriage.

Unexplained recurrent miscarriage (URM) is a common pregnancy complication with few effective therapies. Moreover, little is known regarding the role of pyroptosis in the regulation of the URM immune microenvironment. To address this issue, gene expression profiles of publicly available placental datasets GSE22490 and GSE76862 were downloaded from the Gene Expression Omnibus database. Pyroptosis-related differentially expressed genes were identified and a total of 16 differentially expressed genes associated with pyroptosis were detected, among which 1 was upregulated and 15 were downregulated. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses indicated that the functionally enriched modules and pathways of these genes are closely related to immune and inflammatory responses. Four hub genes were identified: BTK, TLR8, NLRC4, and TNFSF13B. BTK, TLR8, and TNFSF13B were highly connected with immune cells, according to the correlation analysis of four hub genes and 20 different types of immune cells (p < 0.05). The four hub genes were used as research objects to construct the interaction networks. Chorionic villus tissue was used for quantitative real-time polymerase chain reaction and western blot to confirm the expression levels of hub genes, and the results showed that the expression of the four hub genes was significantly decreased in the chorionic villus tissue in the URM group. Collectively, the present study indicates that perhaps pyroptosis is essential to the diversity and complexity of the URM immune microenvironment, and provides a theoretical basis and research ideas for subsequent target gene verification and mechanism research.

Downregulation of FHL2 suppressed trophoblast migration, invasion and epithelial-mesenchymal transition in recurrent miscarriage.

RESEARCH QUESTION: Is four and a half LIM domain 2 (FHL2) involved in trophoblast migration, invasion and epithelial-mesenchymal transition (EMT) in recurrent miscarriage? DESIGN: Villus tissue was collected from 24 patients who had experienced recurrent miscarriage and 24 healthy controls. FHL2 mRNA and protein expression in villus specimens were observed by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Small interfering RNA and overexpression plasmid were used to change the FHL2 expression. JAR and HTR8/SVneo cell lines were used to conduct scratch-wound assay and transwell assay to detect trophoblast migration and invasion of FHL2. Downstream molecule expression of mRNA and protein and EMT markers were verified by qRT-PCR and Western blot. RESULTS: Significantly lower FHL2 mRNA (P = 0.019) and protein (P = 0.0014) expression was found in trophoblasts from the recurrent miscarriage group compared with healthy controls. FHL2 knockdown repressed migration (P = 0.0046), invasion (P < 0.001) and EMT, as shown by significant differences in mRNA and protein expression of the EMT markers N-cadherin, E-cadherin, Vimentin and Snail (all P < 0.05) of extravillus trophoblasts. FHL2 overexpression enhanced migration (P = 0.025), invasion (P < 0.001) and EMT of extravillus trophoblasts (all EMT markers P < 0.05). The positive upstream factor FHL2 in the extracellular signal-related kinase pathway induced JunD expression, thereby promoting trophoblast migration and invasion via matrix metalloproteinase 2. CONCLUSIONS: FHL2 is involved in a regulatory pathway of trophoblast migration, invasion and EMT during early pregnancy, and may have a role in recurrent miscarriage pathogenesis, which can serve as a possible target for novel therapeutic development.

Unexplained recurrent pregnancy loss and markers of arterial vascular health: An experimental case-control study.

OBJECTIVE: To investigate whether women with unexplained recurrent pregnancy loss have impaired arterial vascular health compared with controls, and to evaluate whether this is modifiable by exercise. DESIGN: Experimental case-control pilot study. SETTING: University medical centre in the Netherlands. POPULATION: Twelve women with unexplained recurrent pregnancy loss, 11 nulliparous women and 19 primiparous women with a history of uncomplicated pregnancies. METHODS: In all three groups we measured baseline characteristics, metabolic components and arterial vascular health, and repeated this in women with unexplained recurrent pregnancy loss after 1 month of protocolled and supervised cycle training. MAIN OUTCOME MEASURES: Differences in arterial vascular health between women with unexplained recurrent pregnancy loss and controls, and the effect of exercise on arterial vascular health in women with unexplained recurrent pregnancy loss. RESULTS: Women with unexplained recurrent pregnancy loss have a significantly increased carotid intima media thickness in comparison with both controls (both P < 0.01), a significantly decreased brachial endothelial dependent flow-mediated vasodilation in comparison with both controls (nulliparous: P < 0.01; primiparous: P = 0.05) and a significantly decreased femoral endothelial dependent flow-mediated vasodilation in comparison with primiparous women (P = 0.01). The endothelium independent glyceryl trinitrate response was similar in all groups. With 1 month of exercise, the carotid intima media thickness decreased significantly by 7% (P = 0.05) and the femoral FMD increased significantly by 10% (P = 0.01) in women with unexplained recurrent pregnancy loss. CONCLUSIONS: Women with unexplained recurrent pregnancy loss have an impaired vascular health in comparison with controls. This impaired arterial vascular health can be improved by exercise.

Association of inherited thrombophilia mutations and their combinations among palestinian women with unexplained recurrent miscarriage.

BACKGROUND: Inherited thrombophilia (IT) has a complex pathophysiology and is associated with recurrent miscarriage (RM) by causing placental insufficiency and inhibiting fetal development. However, thrombophilia screening in unexplained RM cases is still questionable. This study aimed to investigate the association between the common eight IT mutations and their combinations among Palestinian women with unexplained RM. METHODS: This is an unmatched case-control study with 200 women (100 unexplained RM cases, 100 controls). Eight common IT mutations namely Factor V Leiden (FVL), prothrombin gene (FII) G202120A, Methylenetetrahydrofolate Reductase (MTHFR) gene (C677T and A1298C), B-fibrinogen gene - 455G > A, FV HR2 A4070G, Plasminogen activator inhibitor 1 (PAI1) 5G/4G and Factor XIIIA (FXIIIA) V34L; were analyzed. The first five mutations were analyzed by Restriction Fragment Length Polymorphism PCR and the other three mutations were analyzed using Amplification Refractory Mutation System PCR. RESULTS: The prevalence of the eight IT mutations among the control group was in the order PAI1 5G/4G (69%), MTHFR C677T (53%) and A1298C (47%), BFG - 455G > A (35%), FVL and FV HR2 (each 18%), FXIIIA V34L (16%) and FII G20210A (3%). Patients had a higher percentage of MTHFR A1298C (heterozygotes and mutant homozygote) compared to controls (p = 0.016). Frequencies of mutant alleles MTHFR A1298C (p < 0.001) and FXIIIA V34L (p = 0.009) were higher among patients compared to controls. No significant differences were observed for all other mutations or mutant alleles. Most patients (75%) and controls (75%) have 2-4 mutant alleles out of 8 mutant alleles studied, while 1% of patients and 2% of controls have zero mutant alleles. None of the combinations of the most often studied mutations (FVL, FII G20210A, MTHFR C1677T, and MTHFR A1298C) showed a significant difference between patients and controls. CONCLUSIONS: There was a significant association between unexplained RM and the mutant alleles of MTHFR A1298C and FXIIIA V34L. No significant association was observed between unexplained RM and the combination of both mutant alleles for the mutations studied. This study is the first Palestinian report that evaluates eight inherited thrombophilia mutations and their alleles' combinations in unexplained RM cases.

Association of histidine-rich glycoprotein C633T single nucleotide polymorphism and recurrent miscarriage in Iranian women.

BACKGROUND: Recurrent miscarriage (RM) is defined as the occurrence of at least two or three subsequent miscarriages within the 20th -24th weeks of pregnancy. The primary objective of this study was to investigate whether histidine-rich glycoprotein C633T single nucleotide polymorphism (HRG C633T SNP) statistically correlates with the occurrence of RM among Iranian women. METHODS AND RESULTS: Blood samples from 200 women were taken at the outset of the study. Then, the blood samples of 100 women who had a record of RM (case group) were compared with the other 100 women's blood samples who had no record of RM (control group). Following DNA extraction, the polymorphism of histidine-rich glycoprotein C633T (HRG C633T) for every case was specified and all women were genotyped by the amplification-refractory mutation system (ARMS) method. The results of the study revealed that there was a statistically significant difference between T/T genotype (OR = 3.5, CI (1.39-8.77), p = 0.007), and C/T genotype (OR = 1.83, CI (0.99-3.37), p = 0.05) in the case and control groups. Also, a statistically significant association was observed in T allelic frequency in the RM participants compared to the control group (OR = 2.01, CI (1.31-3.09), p = 0.01). CONCLUSIONS: The present study determined that there was a statistically significant relationship between HRG C633T SNP and increased RM regarding allelic and genotypical aspects. Moreover, it became apparent that women with homozygous T/T genotype were more susceptible to the risk of RM.

Peroxiredoxin2 regulates trophoblast proliferation and migration through SPIB-HDAC2 pathway.

Adequate proliferation and migration of placental trophoblasts is the prerequisite of a successful pregnancy. Peroxiredoxin2 (Prdx2) is a multi-functional gene involved in various signal events to maintain essential biological functions and normal cellular homeostasis. In this study, substantially lower Prdx2 levels were found in the first trimester cytotrophoblasts of women who suffered from recurrent miscarriage (RM). Prdx2 downregulation inhibited trophoblast proliferation and migration. We demonstrated that histone deacetylase2 (HDAC2) acts downstream of Prdx2 in regulating trophoblast proliferation and migration. HDAC2 deacetylates histone-3-lysine-9 in E-cadherin (E-cad) promoter and reduces the transcription of E-cad epigenetically, whereas it promotes the expression of Slug and Snail genes. These molecular changes may contribute to the trophoblast epithelial-mesenchymal transition. We further verified whether Prdx2 modulated the expression of HDAC2 through SPIB. SPIB could bind to the HDAC2 promoter PU-box region and induce HDAC2 expression. In RM, down-regulated Prdx2 suppresses SPIB-HDAC2 pathway, leading to increased E-cad and decreased Slug and Snail, and eventually restrains trophoblast proliferation and migration. Our study unveils the role of Prdx2-regulated SPIB-HDAC2 pathway in the pathology of RM and provides diagnostic and therapeutic targets for RM as well as other "great obstetrical syndromes" including preeclampsia and intrauterine growth restriction.

Novel Insights into Prokineticin 1 Role in Pregnancy-related Diseases.

Prokineticin 1 (PROK1) is a secreted protein involved in a range of physiological activities such as cell proliferation, migration, angiogenesis, and neuronal cell proliferation. Emerging evidences show that PROK1/PROK receptors (PROKRs) are expressed by trophoblasts, and decidual stroma cells at the maternal-fetal interface. PROK1 plays a critical role in successful pregnancy establishment by regulating the decidualization, implantation and placental development. Dysregulation of prokineticin signaling has been described in certain pathological states associated with pregnancy, including pre-eclampsia, recurrent miscarriage and fetal growth restriction. In this review, the expression and pleiotropic roles of PROK1 under physiological and pathological pregnancy conditions are discussed.

Progestogens in women with threatened miscarriage or recurrent miscarriage: A meta-analysis.

INTRODUCTION: Clinical practice guidelines provide inconsistent recommendations regarding progestogen supplementation for threatened and recurrent miscarriage. We conducted a systematic review and meta-analysis to assess the effectiveness and safety of progestogens for these patients. MATERIAL AND METHODS: We searched Medline, Embase, and Cochrane Central Registry of Controlled Trials up to October 6, 2023 for randomized control trials (RCTs) comparing progestogen supplementation to placebo or no treatment for pregnant women with threatened or recurrent miscarriage. We assessed the risk of bias using a modified version of the Cochrane risk-of-bias tool and the certainty of evidence using the GRADE approach. RESULTS: Of 15 RCTs (6616 pregnancies) reporting on threatened or recurrent miscarriage, 12 (5610 pregnancies) reported on threatened miscarriage with or without a prior history of miscarriage. Results indicated that progesterone probably increases live births (relative risk (RR) 1.04, 95% confidence interval (CI) 0.99-1.10, absolute increase 3.1%, moderate certainty). Of these RCTs, three (1973 pregnancies) reporting on threatened miscarriage with a prior history of miscarriage indicated that progesterone possibly increases live births (RR 1.06, 95% CI: 0.97-1.16, absolute increase 4.4%; low certainty), while four (2540 pregnancies) reporting on threatened miscarriage and no prior miscarriage left the effect very uncertain (RR 1.02, 95% CI: 0.96-1.10, absolute increase 1.7%; very low certainty). Three trials reporting on 1006 patients with a history of two or more prior miscarriages indicated progesterone probably increases live births (RR 1.08, 95% CI: 0.98-1.19, absolute increase 5.7%, moderate certainty). Six RCTs that reported on 2979 patients with at least one prior miscarriage indicated that progesterone probably increases live births (RR 1.07, 95% CI: 1.01-1.13, absolute increase 5.0%; moderate certainty). Progesterone probably has little or no effect on congenital anomalies (RR 1.06, 95% CI: 0.76-1.48, absolute increase 0.1%; moderate certainty), and other serious adverse pregnancy events (RR 1.07, 95% CI: 0.83-1.40, absolute increase 0.2%, moderate certainty). CONCLUSIONS: In women at increased risk of pregnancy loss, progestogens probably increase live births without increasing adverse maternal and neonatal events. It remains possible that the benefit is restricted to those with prior miscarriages.

Fondaparinux Results in Similar Pregnancy Outcomes with Lower Adverse Reaction Rates Compared to Low Molecular Weight Heparin in Chinese Recurrent Miscarriage Women.

Fondaparinux inhibits thrombin generation by inactivating factor Xa, which has the potential to treat recurrent miscarriage (RM). However, more clinical evidence is required to support its application in Chinese women with RM. This research aimed to compare the live birth rate, gestational weeks at delivery, birth weight, Apgar score of newborns, and adverse reaction rates between fondaparinux and low molecular weight heparin (LMWH) in Chinese women with RM. Totally, 132 women with RM treated with fondaparinux or LMWH were included in this retrospective study. According to the corresponding treatment, women with RM were divided into the fondaparinux cohort (N = 45) and LMWH cohort (N = 87). The live birth rate was 68.9% in the fondaparinux cohort and 56.3% in the LMWH cohort, which was not different between the two cohorts (P = 0.161). Multivariable logistics regression analysis suggested that only previous miscarriage times (≥ 4 times vs. < 4 times) were independently related to a lower possibility of live birth in women with RM (odds ratio = 0.431, P = 0.036). It was also observed that gestational weeks at delivery (38.1 ± 1.4 vs. 37.7 ± 1.7 weeks) (P = 0.258), birth weight (2,923.7 ± 355.0 vs. 2,807.8 ± 334.0 g) (P = 0.144), and Apgar score of newborns (9.8 ± 0.5 vs. 9.6 ± 0.8) (P = 0.175) were not different between the fondaparinux cohort and LMWH cohort. Inspiringly, the total adverse reaction rate was reduced in the fondaparinux cohort vs. the LMWH cohort (20.0% vs. 37.9%) (P = 0.036). Fondaparinux results in similar pregnancy outcomes with lower adverse reaction rates compared to LMWH in Chinese women with RM.

Effect of anti-thyroid antibodies on recurrent miscarriage: A meta-analysis.

SETTING: Previous studies addressed the association between anti-thyroid antibodies and recurrent miscarriage (RM), however, the role of anti-thyroid antibodies in RM patients is debatable. OBJECTIVES: Therefore, we conducted this meta-analysis and the aim of this current study was to assess whether anti-thyroid peroxidase (anti-TPO) and/or anti-thyroglobulin (anti-TG) antibody positivity was associated with RM. DESIGN: A meta-analysis was conducted. PARTICIPANTS: Recurrent miscarriage patients. METHODS: STATA 12.0 software were applied to compute odds ratios (ORs)/relative risks (RRs) and 95% CIs regarding association between anti-TPO and anti-TG antibodies and the prevalence of RM. RESULTS: N = 28 studies (8875 participants) explored effect of anti-thyroid antibodies on RM. Analysis of the 28 studies revealed significant association between anti-TPO, anti-TG antibodies and the prevalence of RM with a random effects model (OR/RR = 2.02; 95% CI: 1.63-2.51, p < 0.001; I2 = 44.3%, p value for Q test = 0.004). Analysis of the 20 studies revealed significant association between anti-TPO antibodies and the prevalence of RM with a random effects model (OR/RR = 1.59; 95% CI: 1.25-2.03, p < 0.001; I2 = 43.1%, p value for Q test = 0.022). Analysis of the 14 studies revealed significant association between anti-TG antibodies and the prevalence of RM with a random effects model (OR/RR = 2.25; 95% CI: 1.56-3.23, p < 0.001; I2 = 49.2%, p value for Q test = 0.019). CONCLUSIONS: Based on the currently available analysis, our findings suggest that women with anti-TPO and/or anti-TG antibodies have a higher risk of RM than that in negative antibody women. Further investigation is needed to better clarify the exact role of the anti-thyroid antibodies in RM and whether treatment is of benefit. LIMITATIONS: First, differences from various detection methods and reagents used in different studies may affect the diagnostic interpretation of anti-thyroid antibodies, which might influence the accuracy of this meta-analysis. Second, positive anti-thyroid antibodies seem likely to be part of a more general disorder of maternal immune system, due to restrictions of funding and condition, a complete autoantibody screening investigation is hardly to conduct in all participants, and this could be a possible limitation of all included studies. Third, there is no mention of thyroxine therapy on RM, making the meta-analysis even more limited.

Adverse outcomes in subsequent pregnancies in women with history of recurrent spontaneous abortion: A meta-analysis.

OBJECTIVE: This study aimed to investigate the association between a history of recurrent spontaneous abortion (RSA) and adverse outcomes in women with spontaneous conception. METHODS: A search strategy from the inception to March 3, 2023 was run in PubMed, Embase, Cochrane Library, and Web of Science databases. The odds ratio (OR), and the 95% confidence interval (CI) or point estimation were used as the evaluation indexes. Each outcome measure tested was assessed for heterogeneity using the Cochran Q test. Sensitivity analyses were performed to test the credibility of the meta-analysis results. RESULTS: Fifteen studies involving 1 475 389 pregnant women were included. A history of RSA was associated with gestational diabetes (OR: 2.21, 95% CI: 1.70-2.87, p < 0.001), preeclampsia (OR: 2.06, 95% CI: 1.49-2.86, p < 0.001), placenta previa (OR: 1.82, 95% CI: 1.09-3.02, p = 0.021), placental abruption (OR: 1.67, 95% CI: 1.36-2.06, p < 0.001), miscarriage (OR: 6.37, 95% CI: 3.83-10.57, p < 0.001), preterm birth (OR: 1.80, 95% CI: 1.36-2.37, p < 0.001), cesarean section (OR: 1.47, 95% CI: (1.13-1.91, p = 0.004), perinatal death (OR: 2.24, 95% CI: 1.39-3.60, p = 0.001), and neonatal intensive care unit admission (OR: 1.39, 95% CI: 1.01-1.92, p = 0.047). However, the associations of a history of RSA with gestational hypertension, small for gestational age, fetal anomalies, fetal growth restriction, and postpartum hemorrhage were not observed. CONCLUSION: This meta-analysis indicates a history of RSA was associated with increased risks of several adverse outcomes in pregnant women with spontaneous conception.

Levothyroxine for subclinical hypothyroidism during pregnancy: an updated systematic review and meta-analysis of randomized controlled trials.

PURPOSE: We aimed to perform a systematic review and meta-analysis addressing the efficacy of levothyroxine therapy in pregnant women with subclinical hypothyroidism considering most recent evidence and subgroups of interest for clinical practice. METHODS: PubMed, Embase, and Cochrane Central were searched from inception for randomized controlled trials (RCTs) comparing levothyroxine with placebo or no intervention in pregnant women with subclinical hypothyroidism. We used a random-effects model and conducted subgroup analyses based on thyroid peroxidase antibody status, thyroid stimulating hormone levels, fertility treatment, and recurrent miscarriage. RESULTS: We included 11 RCTs comprising 2,749 pregnant women with subclinical hypothyroidism. Patients treated with levothyroxine (1,439; 52.3%) had significantly lower risk of pregnancy loss (risk ratio 0.69; 95% confidence interval 0.52-0.91; p < 0.01; 6 studies). However, there was no significant association between levothyroxine and live birth (risk ratio 1.01; 95% confidence interval 0.99-1.03; p = 0.29; 8 studies). No statistically significant interaction was observed across subgroups (p > 0.05). CONCLUSION: Levothyroxine replacement therapy for subclinical hypothyroidism during pregnancy may decrease pregnancy loss when early prescribed. Nevertheless, further investigation is needed in patients with thyroid stimulating hormone above four milliunits per liter, especially when associated with recurrent miscarriage or infertility.

Australasian Recurrent Pregnancy Loss Clinical Management Guideline 2024 Part I.

Guidelines for the investigation and management of recurrent pregnancy loss (RPL) have been developed in Europe, USA and UK, but there is currently no Australasian guideline. The Australasian Certificate of Reproductive Endocrinology and Infertility Consensus Expert Panel on Trial Evidence group has prepared a two-part guideline to provide guidance on the management of RPL. In Part I chromosomal, anatomical, and endocrine factors are outlined along with relevant recommendations for clinical management, levels of evidence and grades of consensus. In Part II thrombophilia, autoimmune factors, infective, inflammatory, and endometrial causes, environmental and lifestyle factors, male factor and unexplained causes will be outlined.

Australasian recurrent pregnancy loss clinical management guideline 2024, part II.

Part II of the Australasian guideline for the investigation and management of recurrent pregnancy loss (RPL) provides evidence-based guidance on the management of RPL provided. The implications of inherited and acquired thrombophilia with respect to RPL and suggestions for clinical management are provided. Autoimmune factors, including human leukocyte antigen, cytokines, antinuclear antibodies and coeliac antibodies, and guidance for management are discussed. Infective, inflammatory and endometrial causes of RPL are discussed in detail. Environmental and lifestyle factors, male factor and unexplained causes are outlined. Levels of evidence and grades of consensus are provided for all evidence-based statements.

Clinical Relevance of Vaginal and Endometrial Microbiome Investigation in Women with Repeated Implantation Failure and Recurrent Pregnancy Loss.

Recent studies have investigated if and how the vaginal and endometrial microbiome might affect endometrial receptivity and reproductive health. Although there is no consensus on the existence of a core uterine microbiome yet, evidence shows that the dominance of Lactobacillus spp. in the female reproductive tract is generally associated with eubiosis and improved chances of successful implantation and an ongoing pregnancy. Conversely, vaginal and endometrial dysbiosis can cause local inflammation and an increase of pro-inflammatory cytokines, compromising the integrity and receptivity of the endometrial mucosa and potentially hampering successful embryonic implantation. This review provides a critical appraisal of the influence of the vaginal and endometrial microbiome as parts of the female reproductive tract on fertility outcomes, focusing on repeated implantation failure (RIF) and recurrent pregnancy loss (RPL). It seems that RIF as well as RPL are both associated with an increase in microbiome diversity and a loss of Lactobacillus dominance in the lower female reproductive system.

Decreased Expression of Placental Proteins in Recurrent Pregnancy Loss: Functional Relevance and Diagnostic Value.

Miscarriages affect 50-70% of all conceptions and 15-20% of clinically recognized pregnancies. Recurrent pregnancy loss (RPL, ≥2 miscarriages) affects 1-5% of recognized pregnancies. Nevertheless, our knowledge about the etiologies and pathophysiology of RPL is incomplete, and thus, reliable diagnostic/preventive tools are not yet available. Here, we aimed to define the diagnostic value of three placental proteins for RPL: human chorionic gonadotropin free beta-subunit (free-ß-hCG), pregnancy-associated plasma protein-A (PAPP-A), and placental growth factor (PlGF). Blood samples were collected from women with RPL (n = 14) and controls undergoing elective termination of pregnancy (n = 30) at the time of surgery. Maternal serum protein concentrations were measured by BRAHMS KRYPTOR Analyzer. Daily multiple of median (dMoM) values were calculated for gestational age-specific normalization. To obtain classifiers, logistic regression analysis was performed, and ROC curves were calculated. There were differences in changes of maternal serum protein concentrations with advancing healthy gestation. Between 6 and 13 weeks, women with RPL had lower concentrations and dMoMs of free ß-hCG, PAPP-A, and PlGF than controls. PAPP-A dMoM had the best discriminative properties (AUC = 0.880). Between 9 and 13 weeks, discriminative properties of all protein dMoMs were excellent (free ß-hCG: AUC = 0.975; PAPP-A: AUC = 0.998; PlGF: AUC = 0.924). In conclusion, free-ß-hCG and PAPP-A are valuable biomarkers for RPL, especially between 9 and 13 weeks. Their decreased concentrations indicate the deterioration of placental functions, while lower PlGF levels indicate problems with placental angiogenesis after 9 weeks.

Decreased B7-H3 promotes unexplained recurrent miscarriage via RhoA/ROCK2 signaling pathway and regulates the secretion of decidual NK cells†.

The cause for at least 50% of recurrent miscarriages is unclear, which is defined as unexplained recurrent miscarriages. The B7-H1 (PD-L1), a molecule of the B7 family, promotes tumor development by modulating immune evasion, and recent researchers have also attached importance to the role of B7-H3, another molecule of B7 family, in tumor. Based on the similarity between growth and immune response in tumors and pregnancy, we first explored the role of B7-H3 in unexplained recurrent miscarriages. We found reduced levels of B7-H3 in the villus tissue of unexplained recurrent miscarriage patients, and it was mainly expressed on the cell membrane of extravillous trophoblasts. Further, the HTR-8/SVneo and JEG-3 cells were selected to explore the role of B7-H3 in proliferation, apoptosis, tube formation, migration, and invasion. We found that B7-H3 regulated trophoblast migration and invasion via RhoA/ROCK2 signaling pathway. Inflammatory cytokines were detected through enzyme-linked immunosorbent assay after co-culturing with decidual natural killer cells and B7-H3-knockout JEG-3. Results showed that B7-H3 inhibited IL-8 and IP-10 secretion from the decidual natural killer cells. In a CBA/J × DBA/2 abortion-prone mice model, treatment with B7-H3-Fc protein successfully reduced the rate of embryo resorption. In conclusion, our results revealed a possible mechanism by which decreased B7-H3 on trophoblasts of unexplained recurrent miscarriages inhibited trophoblast migration and invasion and increased IL-8 and IP-10 secretion from the decidual natural killer cells. Furthermore, B7-H3 may be a promising new therapeutic target in unexplained recurrent miscarriage patients.

Preconceptual thyroid peroxidase antibody positivity in women with recurrent pregnancy losses may contribute to an increased risk for another miscarriage.

OBJECTIVE: To investigate preconceptual thyroid peroxidase antibody (TPO-ab) positivity and/or thyroid stimulating hormone (TSH) levels in the upper range of normal as risk factors for recurrent unexplained first-trimester miscarriage. DESIGN: A post-hoc study of a randomized trial, in which acetylsalicylic acid did not affect the risk of a new miscarriage. PATIENTS: Women (n = 483) with at least three unexplained recurrent first-trimester miscarriages investigated at a Swedish secondary referral center. MEASUREMENTS: The levels of TPO-ab and TSH were determined before pregnancy. The occurrence of a new first-trimester miscarriage was analyzed by logistic regression with adjustments when applicable, for age, number of previous miscarriages, obesity and the investigated covariates levels of TPO-ab and TSH. RESULTS: Including all first trimester miscarriages, odds ratio (OR) according to presence of TPO-ab was 1.60 (95% confidence interval [CI]; 0.99-2.57), after adjustment 1.54 (95% CI; 0.94-2.53). Very early (biochemical) pregnancy losses occurred more often in women with than without preconceptual TPO-ab (6.8% vs. 2.0%), OR 3.51 (95% CI; 1.15-10.71), after adjustment 2.91 (95% CI; 0.91-9.29). There was no association between TSH in the upper range of normal and a new miscarriage, adjusted OR 0.76 (95% CI; 0.32-1.83). A prediction model for a new miscarriage included number of previous miscarriages, woman's age and presence of TPO-ab. CONCLUSION: In women with at least three recurrent unexplained pregnancy losses, the presence of TPO-ab may contribute to an increased risk of a first-trimester miscarriage, possibly more pronounced in very early pregnancy. TSH levels 2.5-4.0 mU/L do not seem to increase the miscarriage risk.