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.

An Ultra-Rare Manifestation of an X-Linked Recessive Disorder: Duchenne Muscular Dystrophy in a Female Patient.

Duchenne muscular dystrophy (DMD) is the most common inherited muscle dystrophy. Patients are characterized by muscle weakness, gross motor delay, and elevated serum creatinine kinase (CK) levels. The disease is caused by mutations in the DMD gene located on the X chromosome. Due to the X-linked recessive inheritance pattern, DMD most commonly affects males, who are generally diagnosed between the age of 3-5 years. Here we present an ultra-rare manifestation of DMD in a female patient. Cytogenetic examination showed that she has a t(X;10)(p21.1;p12.1) translocation, which turned out to affect the DMD gene with one of the breakpoints located in exon 54 (detected by genome sequencing). The X-inactivation test revealed skewed X-inactivation (ratio 99:1). Muscle histology and dystrophin immunohistochemistry showed severe dystrophic changes and highly reduced dystrophin expression, respectively. These results, in accordance with the clinical picture and a highly elevated serum CK, led to the diagnosis of DMD. In conclusion, although in very rare cases, DMD can manifest in female patients as well. In this case, a balanced X-autosome reciprocal translocation disrupts the DMD gene and skewed X-inactivation leads to the manifestation of the DMD phenotype.

Analysis of nationwide reflection of rising cesarean section rate in neonatal outcomes of Hungary.

AIM: The authors investigated the effect of rising cesarean rates on maternal and perinatal mortality and in neonatal morbidity in Hungary. METHODS: The authors searched the validated Hungarian National Health database of neonatal discharge records for ICD-10 codes: P0590 (growth restriction); P2010 (fetal hypoxia in labor); P2090 (intrauterine hypoxia); P2200 (respiratory distress syndrome); P2390 (congenital pneumonia); P2400 (meconium aspiration); P2850 (respiratory insufficiency); P3990 (neonatal infection); P9130 (cerebral irritability) and P9141 (cerebral depression) collected between 2006 and 2015. County-specific number of deliveries, perinatal deaths, and data on the route of delivery from the national database were also collected. Controlling for regional disparities with Human Development Index was also performed. Logistic regression analysis was performed to identify determinants of early neonatal mortality. RESULTS: While cesarean section-rate rose from 28.6% to 38.2% perinatal mortality stayed within the range between 7.8‰ and 6.1‰. Early (0-6 days) neonatal mortality (NMR) of the whole country in 2006 was 2.8‰ and showed a significant decrease over the years to reach 1.7‰ in 2015. The rate of most neonatal morbidities decreased significantly. Controlled regression analysis proved that cesarean rates negatively influenced neonatal mortality rates. CONCLUSION: The rise of the cesarean section rate in Hungary between 2006 and 2015 was associated with a detectable reduction of perinatal mortality and morbidity; however, the proportion of improvement in neonatal morbidity is far less than the increment in cesarean rate.

Interfering effect of maternal cell contamination on invasive prenatal molecular genetic testing.

OBJECTIVE: Fetal samples obtained by invasive techniques are prone to maternal cell contamination (MCC), which may lead to false genotyping results. Our aim was to determine 3 molecular genetic tests' sensitivity to MCC. METHOD: By mixing experiments, 1%, 5%, 10%, 20%, 30%, and 40% MCC was simulated, and significant MCC levels were determined for Sanger DNA sequencing, multiplex ligation-dependent probe amplification (MLPA), and pyrosequencing, a next-generation sequencing method. RESULTS: For Sanger sequencing, the limit of sensitivity to MCC was 5% to 30%. For MLPA, a higher proportion of MCC (≥40%) was shown to lead to diagnostic uncertainty. In contrast, pyrosequencing proved to be very sensitive to MCC, detecting a proportion as low as 1%. CONCLUSION: In the case of Sanger sequencing, sensitivity to MCC was variable, while for MLPA, only high levels of MCC proved to be significant. Although the next-generation sequencing method was sensitive to low-level MCC, if MCC level is determined in parallel, accurate quantification of allelic ratios can help to interpret the diagnostic results. Knowledge of significant MCC levels allows correct prenatal diagnosis even if samples are not purely of fetal origin and repeated sampling can be avoided in many of the cases.

Thyroglobulin level at week 16 of pregnancy is superior to urinary iodine concentration in revealing preconceptual and first trimester iodine supply.

Pregnant women are prone to iodine deficiency due to the increased need for iodine during gestation. Progress has recently occurred in establishing serum thyroglobulin (Tg) as an iodine status biomarker, but there is no accepted reference range for iodine sufficiency during pregnancy. An observational study was conducted in 164 pregnant women. At week 16 of gestation urinary iodine concentration (UIC), serum Tg, and thyroid functions were measured, and information on the type of iodine supplementation and smoking were recorded. The parameters of those who started iodine supplementation (≥150 µg/day) at least 4 weeks before pregnancy (n = 27), who started at the detection of pregnancy (n = 51), and who had no iodine supplementation (n = 74) were compared. Sufficient iodine supply was found in the studied population based on median UIC (162 µg/L). Iodine supplementation ≥150 µg/day resulted in higher median UIC regardless of its duration (nonusers: 130 µg/L vs. prepregnancy iodine starters: 240 µg/L, and pregnancy iodine starters: 205 µg/L, p < .001, and p = .023, respectively). Median Tg value of pregnancy starters was identical to that of nonusers (14.5 vs. 14.6 µg/L), whereas prepregnancy starters had lower median Tg (9.1 µg/L, p = .018). Serum Tg concentration at week 16 of pregnancy showed negative relationship (p = .010) with duration of iodine supplementation and positive relationship (p = .008) with smoking, a known interfering factor of iodine metabolism, by multiple regression analysis. Serum Tg at week 16 of pregnancy may be a promising biomarker of preconceptual and first trimester maternal iodine status, the critical early phase of foetal brain development.

[Inborn error of cholesterol biosynthesis: Smith-Lemli-Opitz syndrome]. / A koleszterin-bioszintézis veleszületett zavara: a Smith-Lemli-Opitz-szindróma.

Smith-Lemli-Opitz syndrome is an autosomal recessive mental retardation and multiple malformation syndrome caused by deficiency of the 7-dehydrocholesterol reductase, the enzyme catalyzing the last step in cholesterol biosynthesis. The authors summarize the pathophysiology, epidemiology, clinical picture, diagnostics and therapy of the disease based on a review of the international literature. Since 2004, fourteen patients have been diagnosed with Smith-Lemli-Opitz syndrome in Hungary, which suggests an underdiagnosis of the disease based upon estimated incidence data. Due to deficiency of the 7-dehydrocholesterol reductase, serum cholesterol concentration is low and 7-dehydrocholesterol concentration is elevated in blood and tissues; the latter being highly specific for the syndrome. Detection of disease-causing mutations makes the prenatal diagnosis possible. The clinical spectrum is wide, the most common symptom is syndactyly of the second and third toes. Standard therapy is cholesterol supplementation. Recent publications suggest that oxidative compounds of 7-dehydrocholesterol may play a role in the pathophysiology of the disease as well.

[Update on preimplantation genetic diagnosis and screening]. / A preimplantációs genetikai diagnózis és szurés idoszeru kérdései.

Recent advancement in both human embryology and genomics has created a completely new situation for practical and widespread application of preimplantation genetic diagnosis and screening with a dramatic effect on assisted reproduction. The mapping of the first human genome and the advancement in sequencing technology and bioinformatics has led to the discovery of the exact genetic background of exponentially increasing number of diseases. In parallel, methods for culturing human embryos have also radically improved, enabling the late transfer, and the procedure of vitrification the safe cryopreservation. In consequence, refined genetic analyses have become available from blastocyst biopsy followed by the application of novel genomic methods. Furthermore, some studies suggest that by the selection of aneuploid embryos the pregnancy- and birth-rates can be increased. The amount and the depth of information obtainable from the embryos raise several technical and ethical questions that can be answered by further prospective randomized trials.

Case report: Complex evaluation of coagulation, fibrinolysis and inflammatory cytokines in a SARS-CoV-2 infected pregnant woman with fetal loss.

Background: SARS-CoV-2 infection during pregnancy increases the risk of severe obstetrical complications. Detailed evaluation of COVID-19-associated coagulopathy in a pregnancy with stillbirth hasn't been described so far. Besides knowledge gaps in the pathomechanism leading to stillbirth in COVID-19 pregnancies, currently, no prognostic biomarker is available to identify pregnant patients who are at imminent risk of COVID-19-associated maternal and fetal complications, requiring immediate medical attention. Case: Here we report the case of a 28-year-old SARS-CoV-2 infected pregnant patient, admitted to our hospital at 28 weeks of gestation with intrauterine fetal loss. The presence of SARS-CoV-2 placentitis was confirmed by immunohistological evaluation of the placenta. She had only mild upper respiratory symptoms and her vital signs were within reference throughout labor and postpartum. The stillborn infant was delivered per vias naturales. Fibrinogen concentrate was administered before and after labor due to markedly decreased fibrinogen levels (1.49 g/l) at admission and excessive bleeding during and after delivery. Although coagulation screening tests were not alarming at admission, the balance of hemostasis was strikingly distorted in the patient. As compared to healthy age- and gestational age-matched pregnant controls, increased D-dimer, low FVIII activity, low FXIII level, marked hypocoagulability as demonstrated by the thrombin generation assay, together with shortened clot lysis and decreased levels of fibrinolytic proteins were observed. These alterations most likely have contributed to the increased bleeding observed during labor and in the early postpartum period. Interestingly, at the same time, only moderately altered inflammatory cytokine levels were found at admission. Serum ACE2 activity did not differ in the patient from that of age- and gestational age-matched healthy controls, suggesting that despite previous speculations in the literature, ACE2 may not be used as a potential biomarker for the prediction of COVID-19 placentitis and threatening fetal loss in SARS-CoV-2-infected pregnancies. Conclusions: Although based on this case report no prognostic biomarker could be identified for use in pregnant patients with imminent risk of fetal loss associated with COVID-19 placentitis, the above-described hemostasis alterations warrant awareness of postpartum hemorrhagic complications and could be helpful to identify patients requiring intensified medical attention.

Classification of preeclampsia according to molecular clusters with the goal of achieving personalized prevention.

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.

Risk Estimation of Gestational Diabetes Mellitus in the First Trimester.

CONTEXT: There is no early, first-trimester risk estimation available to predict later (gestational week 24-28) gestational diabetes mellitus (GDM); however, it would be beneficial to start an early treatment to prevent the development of complications. OBJECTIVE: We aimed to identify early, first-trimester prediction markers for GDM. METHODS: The present case-control study is based on the study cohort of a Hungarian biobank containing biological samples and follow-up data from 2545 pregnant women. Oxidative-nitrative stress-related parameters, steroid hormone, and metabolite levels were measured in the serum/plasma samples collected at the end of the first trimester from 55 randomly selected control and 55 women who developed GDM later. RESULTS: Pregnant women who developed GDM later during the pregnancy were older and had higher body mass index. The following parameters showed higher concentration in their serum/plasma samples: fructosamine, total antioxidant capacity, testosterone, cortisone, 21-deoxycortisol; soluble urokinase plasminogen activator receptor, dehydroepiandrosterone sulfate, dihydrotestosterone, cortisol, and 11-deoxycorticosterone levels were lower. Analyzing these variables using a forward stepwise multivariate logistic regression model, we established a GDM prediction model with a specificity of 96.6% and sensitivity of 97.5% (included variables: fructosamine, cortisol, cortisone, 11-deoxycorticosterone, SuPAR). CONCLUSION: Based on these measurements, we accurately predict the development of later-onset GDM (24th-28th weeks of pregnancy). Early risk estimation provides the opportunity for targeted prevention and the timely treatment of GDM. Prevention and slowing the progression of GDM result in a lower lifelong metabolic risk for both mother and offspring.

The role of bile acid measurement in the management of intrahepatic cholestasis of pregnancy / Az epesavmérés szerepe a súlyos terhességi cholestasis szülészeti ellátásában

Introduction: Intrahepatic cholestasis of pregnancy complicates 1% of pregnancies. It increases the risk of severe fetal complications significantly, including preterm delivery and stillbirth. Objective: To summarize our experience with serum total bile acid level measurement that has recently become available for clinical routine in Hungary, and to present the way of gestational cholestasis care at our university. Patients and method: In a retrospective case series, we analyse the data of 12 patients suffering from severe cholestasis of pregnancy treated between September 2020 and September 2021 at the Department of Obstetrics and Gynecology, University of Debrecen. We also determine the statistical correlation between bile acid, transaminase and bilirubin levels in severe cholestasis. Results: 1258 serum samples of 758 patients were measured. 5 of them (0.7% of all cases, 6.4% of cholestasis cases) had severe (total bile acid 40-99 mu mol/L), 7 (0.9% of all cases and 9.0% of cholestasis cases) had very severe (total bile acid >= 100 mu mol/L) disease. The average age of the 12 cases was 30.6 (21-43) years, 7 of them were primigravid. 5 of the patients had a predisposing disease in their history. 6/12 patients received ursodeoxycholic acid treatment, resulting in significant decrease in the bile acid concentrations. Bile acid and GOT (R-2 = 0,14) and bile acid and GPT (R-2 = 0,17) correlations were found to be week in severe cholestasis (n = 45). Postpartum bile acid levels showed rapid improvement. So far, 11 of the patients have delivered and 13 neonates were born, 2/12 were multiple pregnancies. Average gestational age at delivery was 37 (33-40) weeks. 3/11 (27%) were preterm deliveries. 7/8 (88%) of term deliveries were induced. Elective cesarean delivery was not indicated in any of the cases, and in only 2/11 (18%) of the cases did emergency cesarean sections become necessary during labour. No stillbirth occurred. Conclusion: Serum total bile acid measurement is an effective tool in the diagnosis and follow-up of intrahepatic cholestasis of pregnancy, and is inevitable for the protocoll-based obstetrical management of patients. We also present the local protocol of our Department for the management of obstetrical cholestasis.

Early pathways, biomarkers, and four distinct molecular subclasses of preeclampsia: The intersection of clinical, pathological, and high-dimensional biology studies.

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.

How the cesarean delivery rate decreased at the Department of Obstetrics and Gynecology, University of Debrecen during the COVID-19 pandemic / Hogyan csökkent a császármetszések aránya a debreceni Szülészeti Klinikán a koronavírus-pandémia alatt?

Összefoglaló. Bevezetés: Az új koronavírus (SARS-CoV-2) okozta pandémia számos változást eredményezett életünk minden területén, így a debreceni Szülészeti és Nogyógyászati Klinikán is. Célkituzések: A koronavírus-járvány többek között a terminus körüli elektív szülésindukciók gyakorlatának megváltoztatását igényelte. A bevezetett új szakmai eljárásrendeknek, járványügyi intézkedéseknek, a megyei kórházi funkciót ellátó debreceni Kenézy Kórház Szülészeti Osztálya és a Szülészeti Klinika összevonásának, illetve a megváltozott jogi környezetnek a szülészeti ellátásra kifejtett együttes hatásait elemezzük. Módszerek: Helyi protokollokat és részletes eljárásrendeket készítettünk és vezettünk be. 1. Szülésindukció. 2. Szülésindukció cervixérlelést igénylo esetekben. 3. A szülés módjának megválasztása elozményi császármetszés után. 4. A császármetszés utáni hüvelyi szülést támogató, terminus körüli gondozási protokoll. A protokollok bevezetését követo elso 9 hónap szülészeti mutatóit (szülésszám, császármetszések aránya, perinatalis kimenetel, szülésindukciók aránya és sikeressége) vizsgáltuk és hasonlítottuk össze a megelozo idoszak debreceni adataival, illetve az országos szülészeti mutatókkal. Eredmények: Klinikánkon a császármetszés aránya 2020-ban 33,5%-ra csökkent, a protokollok bevezetése óta pedig 30,7%, miközben a 2020. évi magyarországi arány 40,3%. Az összes szülés 20%-a volt 2020-ban szülésindukció, melyek közül 74% végzodött hüvelyi szüléssel, míg ugyanez 2019-ben 11% indukcióból 47%-nak adódott. A havi szülésszám 2020. január és 2021. január között folyamatosan emelkedett (250-rol 450-re), a havi császármetszési arány pedig 41%-ról 25%-ra csökkent. Mindezen változások mellett 2019-rol 2020-ra csökkent mind a perinatalis mortalitás (6,3‰-rol 4,2‰-re), mind a szülés után a Neonatalis Intenzív Centrumba történo felvételek aránya (14,8%-ról 13,5%-ra). Következtetések: A bemutatott tényezok együttes hatásaként - elsodlegesen a megfelelo és következetesen betartott protokolloknak köszönhetoen - a szülésindukciók aránya és sikeressége jelentosen növekedett, a császármetszések aránya szignifikánsan csökkent, javuló perinatalis morbiditási és mortalitási mutatók mellett. Orv Hetil. 2021; 162(21): 811-823. INTRODUCTION: The pandemic caused by the new coronavirus (SARS-CoV-2) has catalized several changes in many fields of our lives, and also at the Department of Obstetrics and Gynecology of the University of Debrecen, Hungary. OBJECTIVES: We wanted to analyse the compound effect of our new local protocols regarding elective labour inductions at term, the coronavirus pandemic and the resulting infection control measures, the merging of the Obstetrics and Gynecology Ward of the Kenézy County Hospital of Debrecen and the University Department of Obstetrics and Gynecology, and also the change of the legal environment. METHODS: Local protocols were introduced: 1. Labour induction. 2. Cervical ripening in labour induction. 3. Choosing the route of delivery after cesarean. 4. Management of pregnancy around term in the case of planned trial of labour after cesarean. We compared the obstetrical data (number of deliveries, cesarean section rate, perinatal outcome and the rate and success rate of labour inductions) before and after the implementation of the protocols. The results were also compared to the Hungarian national database. RESULTS: The annual cesarean rate at our department dropped to 33.5% in 2020. In the first 9-month period, after the introduction of the new reforms, the cesarean rate decreased to 30.7%, whereas the Hungarian national rate was 40.3% in 2020. At our department, 20% of all the deliveries were induced and 74% of them led to vaginal deliveries in 2020, while in 2019 only 11% of deliveries were labour inductions, and 47% of these cases were vaginal deliveries. The monthly number of deliveries was rising constantly between January 2020 (250 deliveries) and January 2021 (450 deliveries), and the monthly cesarean rate decreased from 41% to 25%. Comparing the data of 2019 and 2020, the annual perinatal mortality rate dropped from 6.3‰ in 2019 to 4.2‰ in 2020. Neonatal morbidity, as measured by admissions to the neonatal intensive care unit, also decreased (14.8% in 2019 and 13.5% in 2020). CONCLUSIONS: As a compound result of the described factors, but mainly due to the new protocols, both the rate and the success rate of labour inductions increased significantly, while the cesarean rate decreased with improving perinatal mortality and morbidity. Orv Hetil. 2021; 162(21): 811-823.

Large fetal weight alone in Robson-1 parturients doesn't translate into a risk of Caesarean delivery higher then that of a vaginal birth.

OBJECTIVE: The authors analysed the Caesarean section rate as a function of birth weight among Robson-1 parturients and compared with that among the unselected obstetric population. STUDY DESIGN: A retrospective analysis of birth weight, maternal height and the route of delivery was carried out in an unselected obstetric population of 26,012 parturients. The authors compared birth weight centile distributions of vaginally, and that of abdominally delivered fetuses between Robson-1 parturients as well as those of the total obstetric population. RESULTS: The 90th birth weight centile of fetuses delivered at 37, 38, 39, 40, 41, and 42 weeks gestation were 3960 g, 3960 g, 4000 g, 3950 g, 4000 g and 3820 g, respectively. Among Robson-1 parturients, 677 fetuses weighed >4000 g, and 448 patients (66%) were delivered vaginally. Maternal height did not influence either the birth-weight-percentiles or the Caesarean-rates substantially. Above the birth weight of 4000 g, the Caesarean-rate among Robson-1 parturient rose similarly to that of the total obstetric population. In the knowledge of the most accurately estimated fetal weight, the odds of Caesarean delivery among Robson-1 parturients was not different from that of the total obstetric population. Among pregnancies with fetuses weighing less than 5000 g, the Caesarean-rate was below 50% in both Robson-1 parturients and the total obstetric population of 10 years. CONCLUSION: Even the best possible estimation of fetal weight cannot give a valid reason to downplay the intent of vaginal birth based on the fetal size above 3900 g that would be associated with increased odds of Caesarean delivery.

Screening for preeclampsia in the first trimester of pregnancy in routine clinical practice in Hungary.

We aimed to evaluate the contribution of different factors in the Fetal Medicine Foundation algorithms for preeclampsia (PE) risk calculation during first-trimester screening in Hungary. We selected subjects for the nested case-control study from a prospective cohort of 2545 low-risk pregnancies. Eighty-two patients with PE and 82 gestational age-matched controls were included. Individual PE risk was calculated using two risk-assessing softwares. Using Astraia 2.3.1, considering maternal characteristics and biophysical parameters only, detection rates (DR) were 63.6% for early-PE and 67.6% for late-PE. When we added placenta associated plasma protein A (PAPP-A) to the risk calculation, DRs decreased to 54.5% and 64.8% respectively. Using Astraia 2.8.2 with maternal characteristics and biophysical parameters resulted in the DRs of 63.6% (early-PE) and 56.3% (late-PE). If we added PAPP-A to the risk calculation, DRs improved to 72.7% and 54.9%. The addition of placental growth factor (PlGF) did not increase detection rates in either calculation. In conclusion, using maternal characteristics, biophysical parameters, and PAPP-A, an acceptable screening efficacy could be achieved for early-PE during first-trimester screening. Since PlGF did not improve efficacy in our study, we suggest setting new standard curves for PlGF in Eastern European pregnant women, and the evaluation of novel biochemical markers.

Integrated Systems Biology Approach Identifies Novel Maternal and Placental Pathways of Preeclampsia.

Preeclampsia is a disease of the mother, fetus, and placenta, and the gaps in our understanding of the complex interactions among their respective disease pathways preclude successful treatment and prevention. The placenta has a key role in the pathogenesis of the terminal pathway characterized by exaggerated maternal systemic inflammation, generalized endothelial damage, hypertension, and proteinuria. This sine qua non of preeclampsia may be triggered by distinct underlying mechanisms that occur at early stages of pregnancy and induce different phenotypes. To gain insights into these molecular pathways, we employed a systems biology approach and integrated different "omics," clinical, placental, and functional data from patients with distinct phenotypes of preeclampsia. First trimester maternal blood proteomics uncovered an altered abundance of proteins of the renin-angiotensin and immune systems, complement, and coagulation cascades in patients with term or preterm preeclampsia. Moreover, first trimester maternal blood from preterm preeclamptic patients in vitro dysregulated trophoblastic gene expression. Placental transcriptomics of women with preterm preeclampsia identified distinct gene modules associated with maternal or fetal disease. Placental "virtual" liquid biopsy showed that the dysregulation of these disease gene modules originates during the first trimester. In vitro experiments on hub transcription factors of these gene modules demonstrated that DNA hypermethylation in the regulatory region of ZNF554 leads to gene down-regulation and impaired trophoblast invasion, while BCL6 and ARNT2 up-regulation sensitizes the trophoblast to ischemia, hallmarks of preterm preeclampsia. In summary, our data suggest that there are distinct maternal and placental disease pathways, and their interaction influences the clinical presentation of preeclampsia. The activation of maternal disease pathways can be detected in all phenotypes of preeclampsia earlier and upstream of placental dysfunction, not only downstream as described before, and distinct placental disease pathways are superimposed on these maternal pathways. This is a paradigm shift, which, in agreement with epidemiological studies, warrants for the central pathologic role of preexisting maternal diseases or perturbed maternal-fetal-placental immune interactions in preeclampsia. The description of these novel pathways in the "molecular phase" of preeclampsia and the identification of their hub molecules may enable timely molecular characterization of patients with distinct preeclampsia phenotypes.

Characterization of a new disease-causing mutation of SH2D1A in a family with X-linked lymphoproliferative disease.

Males with an expressed mutation in the SH2D1A gene that encodes an SH2 domain protein named SH2D1A or SAP (NP_002342; signaling lymphocyte activating molecule [SLAM]-associated protein), have an X-linked syndrome characterized by an increased vulnerability to infection with Epstein-Barr virus (EBV). We evaluated two related male patients with fatal infectious mononucleosis (FIM) and mutation in the SH2D1A gene. Sequence analysis revealed a hemizygous c.47G>A mutation in one of the patients, and heterozygosity for this mutation in the genomic DNA from his mother and maternal grandmother. This mutation resulted in p.G16D amino acid change in the sequence of the SAP protein. To analyze the effect of this missense mutation on protein function cDNA was generated by site-directed mutagenesis and expressed in COS cells. We found that half-life of the p.G16D protein was comparable to that of wild type SAP. However, the mutant protein was defective in binding to its physiological ligands SLAM and 2B4. These results suggest that a defect in ligand binding contributes to the loss of function of the SAP protein in patients carrying p.G16D mutation.

Measuring expression levels of small regulatory RNA molecules from body fluids and formalin-fixed, paraffin-embedded samples.

MicroRNAs are involved in the regulation of various pathophysiological processes such as immune regulation and cancer. Next-generation sequencing methods enable us to monitor their presence in various types of samples but we need flexible methods for validating datasets generated by high-throughput methods. Here we describe the detailed protocols to be used with our MiRNA Primer Design Tool assay design system. The presented methods allow the flexible design of the oligonucleotides needed for the RT-qPCR detection of any variant of small regulatory RNA molecules from virtually any species. This method can be used to measure miRNA levels from formalin-fixed, paraffin-embedded (FFPE) samples and various body fluids. As an example, we show the results of the hsa-miR-515-3p, hsa-miR-325, and hsa-miR-155 quantification using a specific UPL probe (Universal Probe Library) and a stem-loop RT-qPCR assay. The small nucleolar RNA RNU43 is used as endogenous control for normalization of the results. Urine from healthy pregnant women and FFPE samples from patients diagnosed with colorectal cancer and treated with antibody-based anti-EGFR monotherapy were used as samples.

In utero incarceration of congenital diaphragmatic hernia.

In utero diagnosis of incarcerated congenital diaphragmatic hernia has never been reported. In our case, congenital diaphragmatic hernia presented at 34 weeks of gestation with dilated bowel loops, pleural effusion, and ascites on fetal ultrasound. Preterm delivery and emergency exploration revealed a tight posterolateral diaphragmatic defect with extensive bowel infarction.