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-β-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 β-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 β-hCG: AUC = 0.975; PAPP-A: AUC = 0.998; PlGF: AUC = 0.924). In conclusion, free-β-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.

The prevention of pre-eclampsia is difficult due to the syndromic nature and multiple underlying mechanisms of this severe complication of pregnancy. The current clinical distinction between early- and late-onset disease, although clinically useful, does not reflect the true nature and complexity of the pathologic processes leading to pre-eclampsia. The current gaps in knowledge on the heterogeneous molecular pathways of this syndrome and the lack of adequate, specific diagnostic methods are major obstacles to early screening and tailored preventive strategies. The development of novel diagnostic tools for detecting the activation of the identified disease pathways would enable early, accurate screening and personalized preventive therapies. We implemented a holistic approach that includes the utilization of different proteomic profiling methods of maternal plasma samples collected from various ethnic populations and the application of systems biology analysis to plasma proteomic, maternal demographic, clinical characteristic, and placental histopathologic data. This approach enabled the identification of four molecular subclasses of pre-eclampsia in which distinct and shared disease mechanisms are activated. The current review summarizes the results and conclusions from these studies and the research and clinical implications of our findings.

Prematurity, which occurs in about 12% of pregnancies worldwide, continues to be one of the leading causes of perinatal morbidity and mortality worldwide. Preterm birth (PTB) can still be considered a syndrome with a variety of causes and underlying factors, which results in mostly unnoticed contraction of the uterus and changes in the cervix. Despite considerable effort aimed at decreasing the incidence of spontaneous PTB, PTB remains the leading cause of perinatal morbidity and mortality. In light of the available data, screening strategies for PTB are deficient. Approaches used to identify women considered by historical factors to be low risk for preterm delivery, as well as those at high risk for PTB, continue to evolve. Ultrasound evaluation of the cervix during pregnancy has been the focus of much research during the past decades. Cervical measurement by transvaginal sonography (TVS) has been shown to be a good predictive test for spontaneous PTB in high-and low-risk singleton pregnancy. Cervical shortening, which is often detected on ultrasound examination before it can be appreciated on physical examination, is one of the first steps in the processes leading to labor and can precede labor by several weeks. This is likewise true for funneling and loss of cervical gland area, which cannot be assessed with the physical examination. Therefore, all of these markers, especially if they are used together, can be useful to predict PTB and start adequate therapy as soon as possible to prevent spontaneous preterm delivery.

Preeclampsia is a syndromic disease of the mother, fetus, and placenta. The main limitation in early and accurate diagnosis of preeclampsia is rooted in the heterogeneity of this syndrome as reflected by diverse molecular pathways, symptoms, and clinical outcomes. Gaps in our knowledge preclude successful early diagnosis, personalized treatment, and prevention. The advent of “omics” technologies and systems biology approaches addresses this problem by identifying the molecular pathways associated with the underlying mechanisms and clinical phenotypes of preeclampsia. Here, we provide a brief overview on how the field has progressed, focusing on studies utilizing state-of-the-art transcriptomics and proteomics methods. Moreover, we summarize our systems biology studies involving maternal blood proteomics and placental transcriptomics, which identified early maternal and placental disease pathways and showed that their interaction influences the clinical presentation of preeclampsia. We also present an analysis of maternal blood proteomics data which revealed distinct molecular subclasses of preeclampsia and their molecular mechanisms. Maternal and placental disease pathways behind these subclasses are similar to those recently reported in studies on the placental transcriptome. These findings may promote the development of novel diagnostic tools for the distinct subtypes of preeclampsia syndrome, enabling early detection and personalized follow-up and tailored care of patients.

Spontaneous preterm birth remains a major cause of neonatal morbidity and mortality across the world. Hence, there is an urgent need to find and implement diagnostic methods and interventions that can reduce this public health treat. The ultrasonographic assessment of the cervix is one tool that can be utilized to identify women at increased risk who may be candidates for preventive interventions. There are three main characteristics of the cervix, which can be evaluated during the ultrasound examination of the cervix: cervical length (CL), funneling and cervical gland area. Cervical shortening is one of the first steps in the processes leading to labor and can precede labor by several weeks. Because shortening begins at the internal cervical os and progresses caudally, it is often detected on ultrasound examination before it can be appreciated on physical examination. This is equally true for funneling and cervical gland area (CGA), which cannot be assessed with the physical examination. Based on previous experiences, the timing and frequency of ultrasonographic assessment of the cervix is primarily based on the patient’s prior obstetric history (low-risk women are screened once at 18–24 weeks of gestation; high-risk population usually begins screening at about 16 weeks of gestation and the frequency depends on the measurement result). Classically the diagnosis of short cervix is defined when the CL is less than or equal to 25 mm at these gestational weeks, with the best prediction for PTB obtained at 16–24 weeks of gestation. The CL measurement, evaluation of funneling and CGA together increased the sensitivity of cervical screening for PTB and appeared to be powerful predictor of PTB before 32 weeks gestation. Generally, the importance of positive test is to try to recognize cervical changes on time, to plane the adequate therapy, to prepare for sufficient intrauterine transport, and to administered course of antenatal corticosteroid therapy to women at risk for PTB reduced the incidence and severity of respiratory distress syndrome (RDS) and mortality in offspring. Many interventions (bed rest, lifestyle intervention, cervical cerclage, pessary, progesteron, indomethacin, antibiotics, etc.) have been proposed in an attempt to prevent PTB depending on risk classification.

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Introduction: Miscarriages affect 50-70% of all gestations and 15-20% of clinically recognized pregnancies. Recurrent pregnancy loss (RPL) occurs in 1-5% of clinical pregnancies and has an enormous demographic impact. However, the etiologies and molecular pathways of RPL are scarcely understood, and therefore, reliable diagnostic and preventive methods are not yet available. Here, we aimed to discover novel biomarkers for RPL using next-generation proteomics technology to help develop early and effective diagnostic tools. Methods: First-trimester blood samples were collected from women with RPL (n=11) and controls with elective termination of pregnancy (n=11) between 6–13 weeks of gestation. After immunodepleting 14 highly abundant proteins, plasma samples were reduced, alkylated, and trypsin digested. For the separation of peptides, nano-flow reversed-phase chromatography was applied, and then mass spectrometric analysis was performed. Differentially abundant (DA) proteins were identified using strict criteria and analyzed by protein network and Gene Ontology (GO) enrichment analyses, and two biomarker candidates (CGB and PAPPA) were validated by immunoassay. Biomarker predictive properties were demonstrated using Receiver Operating Characteristic (ROC) curves. Assessments were performed for all cases and then for two gestational age groups, before and after the start of placental circulation [“early RPL”: gestational weeks (GW) 6–9, “late RPL”: GW 9–13]. Results: Altogether, 651 proteins were identified and quantified across all samples. When comparing “early control” and “late control” samples, 60 proteins [11 predominantly placenta-expressed (PPE)] were DA. When analyzing all cases, 50 DA proteins were found in RPL (top 3 down: PZP, PSG9, CGB; top 3 up: C4BPA, HBA, HBB), among which 11 PPE proteins were found, all downregulated. Enriched GO terms included ‘placental function’, ‘oxidative processes’, ‘immune function’, and ‘blood coagulation’ related biological processes. When cases were split into early and late RPL groups, 40 DA proteins were identified in early RPL (top 3 down: SHBG, CGB, CGA; top 3 up: C4BPA, SAMP, C4BPB) and 90 in late RPL (top 3 down: PZP, PAPPA, PSG9; top 3 up: THBS1, ECM1, HBB), among which only 15 were shared by both RPL groups. In early RPL, only ‘placental function’ and ‘immune function’ related biological processes were enriched, while in late RPL the top enriched GO terms included ‘placental function’, ‘oxidative processes’, ‘immune function’, ‘blood coagulation’, ‘angiogenesis’, ‘cell migration’, and ‘blood circulation’ related biological processes. Among GO terms, only ‘placental function’ related biological processes were enriched when early- and late RPL DA proteins were analyzed together. Furthermore, the areas under the ROC curves were >0.9 for two protein candidates in all RPL, for five proteins in early RPL, and for ten proteins in late RPL. Among these candidates, CGB and PAPPA were validated by immunoassay which showed a good correlation with MS data (RCGB=0.795 and RPAPPA=0.965). Conclusion: We discovered distinct as well as shared molecular pathways associated with RPL pathogenesis before and after the start of placental circulation and identified novel biomarkers for these pathways which have outstanding discriminative properties. Our results may facilitate a better understanding of the molecular pathways of RPL. However, larger clinical studies are needed to investigate whether the identified biomarkers also have predictive power for RPL before pregnancies fail and to test drugs for the modulation of the identified disease pathways and the prevention of RPL. Our findings highlight the importance of the maternal immune system in maintaining successful pregnancy and suggest that targeting immune pathways may offer novel therapeutic approaches for RPL.

AbstractThe concept of the Great Obstetrical Syndromes was introduced to explain the unique nature of obstetrical disease, which differs fundamentally from disorders in other areas of medicine. These syndromes, including preeclampsia, fetal growth restriction, fetal death, and spontaneous preterm birth, represent clinical endpoints rather than single diseases and share defining characteristics: they arise from multiple etiologies, have a prolonged subclinical phase, involve the fetus as an active participant, are adaptive in nature, and result from complex genetic and environmental interactions between the mother and fetus. Among the diverse mechanisms leading to these syndromes, abnormalities of the maternal supply line to the placenta constitute one major etiology and are often caused by vascular disorders affecting the maternal cardiovascular system and uterine spiral arteries, resulting in placental lesions of maternal vascular malperfusion. The most severe spiral artery lesion is atherosis, which closely resembles atherosclerosis and links obstetrical syndromes to maternal vascular disease. Disorders of deep placentation associated with maternal vascular malperfusion are accompanied by characteristic alterations in angiogenic balance, and the ratio of placental growth factor to soluble fms-like tyrosine kinase-1 in the maternal circulation serves as a biomarker of this pathophysiologic process. Importantly, each obstetrical syndrome is associated with a stereotypic temporal pattern of angiogenic imbalance that reflects differences in disease burden, timing, and clinical expression. While substantial progress has been made in the prediction and prevention of preeclampsia, these concepts extend to other obstetrical syndromes, including fetal growth restriction, fetal death, and spontaneous preterm labor, supporting a unified biologic framework for early risk assessment and personalized prevention.