Biomarker development for presymptomatic molecular diagnosis of preeclampsia: Feasible, useful or even unnecessary?

The past decade saw the advent of a number of promising biomarkers to detect pregnancies at risk for preeclampsia (PE), the foremost being those associated with an imbalance of angiogenic factors. In late pregnancy, these are useful for the detection of imminent cases of PE, while earlier they were more predictive for early- than late-onset PE. This suggests that there may be fundamental differences between the underlying pathology of these two PE forms. Therefore, it is possible that such a biological premise may limit the development of biomarkers that will permit the efficacious detection of both early- and late-onset PE via an analysis of first-trimester maternal blood samples. Consequently, a significant increase in our understanding of the underlying pathology of PE, using a variety of approaches ranging from systems biology to animal models, will be necessary in order to overcome this obstacle. © Informa UK, Ltd

Exosomal small RNA profiling in first-trimester maternal blood explores early molecular pathways of preterm preeclampsia

IntroductionPreeclampsia (PE) is a severe obstetrical syndrome characterized by new-onset hypertension and proteinuria and it is often associated with fetal intrauterine growth restriction (IUGR). PE leads to long-term health complications, so early diagnosis would be crucial for timely prevention. There are multiple etiologies and subtypes of PE, and this heterogeneity has hindered accurate identification in the presymptomatic phase. Recent investigations have pointed to the potential role of small regulatory RNAs in PE, and these species, which travel in extracellular vesicles (EVs) in the circulation, have raised the possibility of non-invasive diagnostics. The aim of this study was to investigate the behavior of exosomal regulatory small RNAs in the most severe subtype of PE with IUGR.MethodsWe isolated exosomal EVs from first-trimester peripheral blood plasma samples of women who later developed preterm PE with IUGR (n=6) and gestational age-matched healthy controls (n=14). The small RNA content of EVs and their differential expression were determined by next-generation sequencing and further validated by quantitative real-time PCR. We also applied the rigorous exceRpt bioinformatics pipeline for small RNA identification, followed by target verification and Gene Ontology analysis.ResultsOverall, >2700 small RNAs were identified in all samples and, of interest, the majority belonged to the RNA interference (RNAi) pathways. Among the RNAi species, 16 differentially expressed microRNAs were up-regulated in PE, whereas up-regulated and down-regulated members were equally found among the six identified Piwi-associated RNAs. Gene ontology analysis of the predicted small RNA targets showed enrichment of genes in pathways related to immune processes involved in decidualization, placentation and embryonic development, indicating that dysregulation of the induced small RNAs is connected to the impairment of immune pathways in preeclampsia development. Finally, the subsequent validation experiments revealed that the hsa_piR_016658 piRNA is a promising biomarker candidate for preterm PE associated with IUGR.DiscussionOur rigorously designed study in a homogeneous group of patients unraveled small RNAs in circulating maternal exosomes that act on physiological pathways dysregulated in preterm PE with IUGR. Therefore, our small RNA hits are not only suitable biomarker candidates, but the revealed biological pathways may further inform us about the complex pathology of this severe PE subtype

Characteristic Changes in Decidual Gene Expression Signature in Spontaneous Term Parturition

Background: The decidua has been implicated in the "terminal pathway" of human term parturition, which is characterized by the activation of pro-inflammatory pathways in gestational tissues. However, the transcriptomic changes in the decidua leading to terminal pathway activation have not been systematically explored. This study aimed to compare the decidual expression of developmental signaling and inflammation-related genes before and after spontaneous term labor in order to reveal their involvement in this process. Materials and Methods: Chorioamniotic membranes were obtained from normal pregnant women who delivered at term with spontaneous labor (TIL, n=14) or without labor (TNL, n=15). Decidual cells were isolated from snap-frozen chorioamniotic membranes with laser microdissection. The expression of 46 genes involved in decidual development, sex steroid and prostaglandin signaling, as well as pro- and anti-inflammatory pathways was analyzed using high-throughput quantitative real-time polymerase chain reaction (qRT-PCR). Chorioamniotic membrane sections were immunostained and then semi-quantified for five proteins, and immunoassays for three chemokines were performed on maternal plasma samples. Results: The genes with the highest expression in the decidua at term gestation included insulin-like growth factor-binding protein 1 (IGFBP1), galectin-1 (LGALS1), and progestogen-associated endometrial protein (PAEP); the expression of estrogen receptor 1 (ESR1), homeobox A11 (HOXA11), interleukin 1beta (IL1B), IL8, progesterone receptor membrane component 2 (PGRMC2), and prostaglandin E synthase (PTGES) was higher in TIL than in TNL cases; the expression of chemokine C-C motif ligand 2 (CCL2), CCL5, LGALS1, LGALS3, and PAEP was lower in TIL than in TNL cases; immunostaining confirmed qRT-PCR data for IL-8, CCL2, galectin-1, galectin-3, and PAEP; and no correlations between the decidual gene expression and the maternal plasma protein concentrations of CCL2, CCL5, and IL-8 were found. Conclusion: Our data suggests that with the initiation of parturition, the decidual expression of anti-inflammatory mediators decreases, while the expression of pro-inflammatory mediators and steroid receptors increases. This shift may affect downstream signaling pathways that can lead to parturition

The role of neutrophil activation in determining the outcome of pregnancy and modulation by hormones and/or cytokines

Neutrophils are often exclusively considered as first line innate immune defence, able to rapidly kill or trap pathogens and causing in case of over activation tissue damage. In the female reproductive tract, however, the presence and activity of neutrophils seem to be tightly regulated. Major players in orchestrating this regulation are cyclical steroid sex hormones present during the menstrual cycle and pregnancy. This review describes the role of sex hormones in regulating directly or indirectly the functionality of neutrophils, the role of neutrophils during fertilisation and pregnancy and in controlling viral, fungal and bacterial infection. This review also discusses the consequence of overt neutrophil activation in pregnancy pathologies. This article is protected by copyright. All rights reserved

Placental galectins regulate innate and adaptive immune responses in pregnancy

IntroductionGalectins are master regulators of maternal immune responses and placentation in pregnancy. Galectin-13 (gal-13) and galectin-14 (gal-14) are expressed solely by the placenta and contribute to maternal-fetal immune tolerance by inducing the apoptosis of activated T lymphocytes and the polarization of neutrophils toward an immune-regulatory phenotype.Furthermore, their decreased placental expression is associated with pregnancy complications, such as preeclampsia and miscarriage. Yet, our knowledge of the immunoregulatory role of placental galectins is incomplete.MethodsThis study aimed to investigate the effects of recombinant gal-13 and gal-14 on cell viability, apoptosis, and cytokine production of peripheral blood mononuclear cells (PBMCs) and the signaling pathways involved.ResultsHerein, we show that gal-13 and gal-14 bind to the surface of non-activated PBMCs (monocytes, natural killer cells, B cells, and T cells) and increase their viability while decreasing the rate of their apoptosis without promoting cell proliferation. We also demonstrate that gal-13 and gal-14 induce the production of interleukin (IL)-8, IL-10, and interferon-gamma cytokines in a concentration-dependent manner in PBMCs. The parallel activation of Erk1/2, p38, and NF-ĸB signaling evidenced by kinase phosphorylation in PBMCs suggests the involvement of these pathways in the regulation of the galectin-affected immune cell functions.DiscussionThese findings provide further evidence on how placenta-specific galectins assist in the establishment and maintenance of a proper immune environment during a healthy pregnancy

Feto-Maternal Microchimerism: The Pre-eclampsia Conundrum

Feto-maternal microchimerism (FMM) involves bidirectional cross-placental trafficking during pregnancy, leading to a micro-chimeric state that can persist for decades. In this manner a pregnant woman will harbor cells from her mother, as well as, cells from her child. Historically, eclampsia, a severe disorder of pregnancy provided the basis for FMM following the detection of trophoblast cells in the lungs of deceased women. Bi-directional cell trafficking between mother and fetus is also altered in pre-eclampsia and has been suggested to contribute to the underlying etiology. FMM has been implicated in tolerance promotion, remission of auto-inflammatory disorders during pregnancy, or the development of autoimmune conditions post-partum. The underlying mechanism whereby the host immune system is modulated is unclear but appears to involve HLA class II molecules, in that incompatibility between mother and fetus promotes remission of rheumatoid arthritis, whereas feto-maternal HLA compatibility may assist in the post-partum initiation of scleroderma. Couples having a high degree of HLA class II compatibility have an increased risk for pre-eclampsia, while the occurrence of scleroderma and rheumatoid arthritis is greater in pre-eclamptic cases than in women with normal pregnancies, suggesting a long term autoimmune predisposition. Since pregnant women with pre-eclampsia exhibit significantly lower levels of maternally-derived micro-chimerism, the question arises whether pre-eclampsia and post-partum development of autoimmune conditions occur due to the failure of the grandmothers cells to adequately regulate an inappropriate micro-chimeric constellation

The prediction of early preeclampsia

To identify maternal plasma protein markers for early preeclampsia (delivery <34 weeks of gestation) and to determine whether the prediction performance is affected by disease severity and presence of placental lesions consistent with maternal vascular malperfusion (MVM) among cases.This longitudinal case-control study included 90 patients with a normal pregnancy and 33 patients with early preeclampsia. Two to six maternal plasma samples were collected throughout gestation from each woman. The abundance of 1,125 proteins was measured using high-affinity aptamer-based proteomic assays, and data were modeled using linear mixed-effects models. After data transformation into multiples of the mean values for gestational age, parsimonious linear discriminant analysis risk models were fit for each gestational-age interval (8-16, 16.1-22, 22.1-28, 28.1-32 weeks). Proteomic profiles of early preeclampsia cases were also compared to those of a combined set of controls and late preeclampsia cases (n = 76) reported previously. Prediction performance was estimated via bootstrap.We found that 1) multi-protein models at 16.1-22 weeks of gestation predicted early preeclampsia with a sensitivity of 71% at a false-positive rate (FPR) of 10%. High abundance of matrix metalloproteinase-7 and glycoprotein IIbIIIa complex were the most reliable predictors at this gestational age; 2) at 22.1-28 weeks of gestation, lower abundance of placental growth factor (PlGF) and vascular endothelial growth factor A, isoform 121 (VEGF-121), as well as elevated sialic acid binding immunoglobulin-like lectin 6 (siglec-6) and activin-A, were the best predictors of the subsequent development of early preeclampsia (81% sensitivity, FPR = 10%); 3) at 28.1-32 weeks of gestation, the sensitivity of multi-protein models was 85% (FPR = 10%) with the best predictors being activated leukocyte cell adhesion molecule, siglec-6, and VEGF-121; 4) the increase in siglec-6, activin-A, and VEGF-121 at 22.1-28 weeks of gestation differentiated women who subsequently developed early preeclampsia from those who had a normal pregnancy or developed late preeclampsia (sensitivity 77%, FPR = 10%); 5) the sensitivity of risk models was higher for early preeclampsia with placental MVM lesions than for the entire early preeclampsia group (90% versus 71% at 16.1-22 weeks; 87% versus 81% at 22.1-28 weeks; and 90% versus 85% at 28.1-32 weeks, all FPR = 10%); and 6) the sensitivity of prediction models was higher for severe early preeclampsia than for the entire early preeclampsia group (84% versus 71% at 16.1-22 weeks).We have presented herein a catalogue of proteome changes in maternal plasma proteome that precede the diagnosis of preeclampsia and can distinguish among early and late phenotypes. The sensitivity of maternal plasma protein models for early preeclampsia is higher in women with underlying vascular placental disease and in those with a severe phenotype