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INTRODUCTION: Oxidative stress as observed in fetal growth restriction (FGR) and preeclampsia (PE) can be identified by decreased levels of systemic free thiols (FT) and increased levels of plasma ischemia-modified albumin (IMA), which may serve as biomarkers in maternal blood for pregnancy complications. We evaluate the performance of oxidative stress-associated potential biomarkers for FGR and PE, and their relationship with clinical characteristics.METHODS: A prospective clinical pilot study was performed in healthy controls and women with pregnancies complicated by severe FGR with or without PE. Blood samples were taken directly after inclusion and analyzed for FT; IMA; soluble FMS-like tyrosine kinase-1 (sFlt-1); placenta growth factor (PlGF); and biomarkers like leptin and soluble receptors for advanced glycation end products (sRAGE). Placentas were examined microscopically. Descriptive statistics and receiver operating characteristics statistics were performed.RESULTS: Mothers with both severe FGR and PE had significantly reduced FT levels (p < 0.001) and PlGF levels (p < 0.001), and increased levels of plasma IMA (p < 0.05), sFlt (p < 0.001), leptin (p < 0.05) and sRAGE (p < 0.01) compared to women with FGR only. Systemic FT levels were significantly inversely associated with blood pressure (p < 0.01) and plasma IMA (p < 0.001), leptin (p = 0.01) and sRAGE (p < 0.001). Systemic FT and leptin showed significant discriminative ability to differentiate mothers with both FGR and PE from mothers with uncomplicated pregnancies or pregnancies complicated by FGR only.DISCUSSION: There is a significant discriminative capacity of FT, IMA, leptin and sRAGE that harbor potential as biomarkers of pregnancies complicated by combined FGR and PE.

Introduction: Placental pathology and pregnancy complications are associated with unfavorable regulation of the maternal immune system. Although much research has been performed towards the role of immune cells like macrophages and T cells in this context, little is known about the presence and function of mast cells (MC). MC can be sub classified in tryptase-positive (MCT) and tryptase- and chymase-positive (MCTC). This study investigates the presence of MC in the decidua of pregnancies complicated by fetal growth restriction (FGR) and stillbirth (SB).Methods: Placental tissue from FGR (n = 250), SB (n = 64) and healthy pregnancies (n = 42) was included. Histopathological lesions were classified according to the Amsterdam Placental Workshop Group criteria. Tissue sections were stained for tryptase and chymase. Decidual MC were counted manually, and the results were expressed as number of cells/mm2 decidual tissue.Results: A significant lower median number of MCTC was found in the decidua of FGR (0.40 per mm2; p < 0.001) and SB (0.51 per mm2; p < 0.05) compared to healthy controls (1.04 per mm2). No difference in MCT number (1.19 per mm2, 1.88 per mm2 and 1.37 per mm2 respectively) was seen between the groups. There was no difference in number of MCT and MCTC between placental pathological lesions.Discussion: Our findings suggest a shift in decidual MC balance towards MCT in pregnancy complications. No difference in numbers of MC subtypes was found to be related to histopathologic lesions.

PROBLEM: Women with a previous uncomplicated pregnancy have lower risks of immune-associated pregnancy disorders in a subsequent pregnancy. This could indicate a different maternal immune response in multigravid women compared to primigravid women. In a previous study, we showed persistent higher memory T cell proportions with higher CD69 expression after uncomplicated pregnancies. To our knowledge no studies have reported on immune cells in general, and immune memory cells and macrophages specifically in multigravid and primigravid women.METHOD OF STUDY: T cells and macrophages were isolated from term decidua parietalis and decidua basalis tissue from healthy primigravid women (n = 12) and multigravid women (n = 12). Using flow cytometry, different T cell populations including memory T cells and macrophages were analyzed. To analyze whether a different immune phenotype is already present in early pregnancy, decidual tissue from uncomplicated ongoing pregnancies between 9 and 12 weeks of gestation from multigravida and primigravid women was investigated using qRT-PCR.RESULTS: Nearly all T cell subsets analyzed in the decidua parietalis had significantly higher CD69+ proportions in multigravid women compared to primigravid women. A higher proportion of decidual (CD50- ) M2-like macrophages was found in the decidua parietalis in multigravid women compared to primigravid women. In first trimester decidual tissue higher FOXP3 mRNA expression was found in multigravid women compared to primigravid women.CONCLUSIONS: This study shows that decidual tissue from multigravid women has a more activated and immunoregulatory phenotype compared to decidual tissue from primigravid women in early pregnancy and at term which could suggest a more balanced immune adaptation towards pregnancy after earlier uncomplicated pregnancies. This article is protected by copyright. All rights reserved.

Pregnancy complications including fetal growth restriction, preeclampsia, and preterm birth, as well as gestational diabetes, affect one in every four to five pregnancies. Accumulating evidence indicates that increased production of reactive oxygen species accompanies these complications. Given that reactive oxygen species are cell stress-inducing agents, they may have a causal role in disease pathophysiology, although the exact mechanisms by which they contribute to pregnancy complications are not completely understood. Since many environmental and lifestyle factors and exposures are known to modulate reactive oxygen species production, the exposome of pregnant women could contribute to increased generation of reactive oxygen species. The objective of this review is to provide a comprehensive overview of the endogenous and exogenous exposome factors that regulate reactive species in healthy and complicated pregnancies. We also provide a description of dietary interventions aimed at the reduction of reactive species in order to attenuate adverse pregnancy outcome. Dietary interventions in general hold minimal risk in pregnancy and could therefore be considered a promising therapeutic approach.

Pregnancy after solid organ transplantation (SOT) has potential risks for the offspring. Most existing research focused on short-term pregnancy outcomes. The aim of this systematic review was to evaluate available data concerning longer term outcomes (>1 year) of these children. A systematic literature search, following PRISMA guidelines, of PubMed and Embase was performed from the earliest date of inception through to 6th April 2022. Publications on all types of (combined) SOT were eligible for inclusion. In total, 53 articles were included. The majority assessed offspring after kidney (78% of offspring) or liver transplantation (17% of offspring). 33 studies included offspring aged >4 years and five offspring aged >18 years. One study was included on fathers with SOT. The majority of the 1,664 included children after maternal SOT had normal intellectual, psychomotor, and behavioral development. Although prematurity and low birth weight were commonly present, regular growth after 1 year of age was described. No studies reported opportunistic or chronic infections or abnormal response to vaccinations. In general, pregnancy after SOT appears to have reassuring longer term outcomes for the offspring. However, existing information is predominantly limited to studies with young children. Longer prospective studies with follow-up into adulthood of these children are warranted.

Background There is increasing evidence that a history of preeclampsia is an important risk factor for future cardiovascular events. Awareness of this risk could provide opportunities for identification of women at risk, with opportunities for prevention and / or early intervention. A standardized follow-up has not yet been implemented in the north of the Netherlands. The objective of this qualitative study was to explore the opinions and wishes among women and physicians about the follow-up for women with a history of preeclampsia. Methods Semi-structured interviews with 15 women and 14 physicians (5 obstetricians, 4 general practitioners, 3 vascular medicine specialists and 2 cardiologists) were performed and addressed topics about knowledge on CVR, current - and future follow-up. Women were approached through the HELLP foundation and their physicians. Physicians were approached by email. The interviews were recorded, typed and coded using ATLAS.ti software. A theoretical-driven thematic analysis was performed. Results Women had some knowledge about the association between preeclampsia and the increased CVR, but missed information from their health care providers. Specialists were aware of the association, but the information and advice they provided to their patients was minimal and inconsistent according to themselves. Whereas some general practitioners regarded their own knowledge as limited. There was a clear desire among women for a more extensive follow-up with specific attention to both emotional and physical consequences of preeclampsia. Physicians indicated that they preferred to see a follow up program concerning the CVR at the general practitioner as part of the already existent cardiovascular risk management (CVRM) program. Conclusion Women and medical specialists consider it important to improve aftercare for women after a pregnancy complicated by preeclampsia. Introducing these women into the CVRM program at the general practitioner is regarded as a preferred first step. Further research is warranted to establish an evidence-based guideline for the follow-up of these women.

Immune cells are critically involved in placental development and functioning, and inadequate regulation of the maternal immune system is associated with placental pathology and pregnancy complications. This study aimed to explore numbers of decidual immune cells in pregnancies complicated with fetal growth restriction (FGR) and stillbirth (SB), and in placentas with histopathological lesions: maternal vascular malperfusion (MVM), fetal vascular malperfusion (FVM), delayed villous maturation (DVM), chorioamnionitis (CA), and villitis of unknown etiology (VUE). Placental tissue from FGR (n = 250), SB (n = 64), and healthy pregnancies (n = 42) was included. Histopathological lesions were classified according to criteria developed by the Amsterdam Placental Workshop Group. Tissue slides were stained for CD68 (macrophages), CD206 (M2-like macrophages), CD3 (T cells), FOXP3 [regulatory T (Treg) cells], and CD56 [natural killer (NK) cells]. Cell numbers were analyzed in the decidua basalis using computerized morphometry. The Mann-Whitney U-test and Kruskal Wallis test with the Dunn's as post-hoc test were used for statistical analysis. Numbers of CD68+ macrophages were higher in FGR compared to healthy pregnancies (p < 0.001), accompanied by lower CD206+/CD68+ ratios (p < 0.01). In addition, in FGR higher numbers of FOXP3+ Treg cells were seen (p < 0.01) with elevated FOXP3+/CD3+ ratios (p < 0.01). Similarly, in SB elevated FOXP3+ Treg cells were found (p < 0.05) with a higher FOXP3+/CD3+ ratio (p < 0.01). Furthermore, a trend toward higher numbers of CD68+ macrophages was found (p < 0.1) in SB. Numbers of CD3+ and FOXP3+ cells were higher in placentas with VUE compared to placentas without lesions (p < 0.01 and p < 0.001), accompanied by higher FOXP3+/CD3+ ratios (p < 0.01). Elevated numbers of macrophages with a lower M2/total macrophage ratio in FGR suggest a role for a macrophage surplus in its pathogenesis and could specifically indicate involvement of inflammatory macrophages. Higher numbers of FOXP3+ Treg cells with higher Treg/total T cell ratios in VUE may be associated with impaired maternal-fetal tolerance and a compensatory response of Treg cells. The abundant presence of placental lesions in the FGR and SB cohorts might explain the increase of Treg/total T cell ratios in these groups. More functionality studies of the observed altered immune cell subsets are needed.

Corticosteroids have been utilised in the assisted reproduction setting with the expectation of suppressing aberrant immune activation and improving fertility in women. However, the effects of corticosteroids on fertility, and on pregnancy and offspring outcomes, are unclear. In this study, mice were administered prednisolone (1 mg/kg) or PBS daily in the pre-implantation phase, and effects on the adaptive immune response, the implantation rate, fetal development and postnatal outcomes were investigated. Prednisolone disrupted the expected expansion of CD4+ T cells in early pregnancy, inhibiting generation of both regulatory T cells (Treg cells) and effector T cells and suppressing IFNG required for T cell functional competence. Prednisolone caused an 8–20% increase in the embryo implantation rate and increased the number of viable pups per litter. In late gestation, fetal and placental weights were reduced in a litter size-dependent manner, and the canonical inverse relationship between litter size and fetal weight was lost. The duration of pregnancy was extended by ~ 0.5 day and birth weight was reduced by ~ 5% after prednisolone treatment. Viability of prednisolone-exposed offspring was comparable to controls, but body weight was altered in adulthood, particularly in male offspring. Thus, while prednisolone given in the pre-implantation phase in mice increases maternal receptivity to implantation and resource investment in fetal growth, there is a trade-off in long-term consequences for fetal development, birth weight and offspring health. These effects are associated with, and likely caused by, prednisolone suppression of the adaptive immune response at the outset of pregnancy.

Problem: Preeclampsia is a major cause of fetal and maternal mortality and morbidity. Disturbed fetal-maternal immune tolerance, and therewith memory T cells, might be involved in its etiology. This study aims to give insight into memory T-cell populations and its associated cytokines in the decidual layers in early-onset preeclampsia (EO-PE) and late-onset preeclampsia (LO-PE). Method of Study: Lymphocytes were isolated from the decidua parietalis and basalis from EO-PE (n = 6), LO-PE (n = 8) and healthy (n = 15) pregnancies. CD4+ and CD8+ central- (CCR7+), effector- (CCR7−), tissue resident- (CD103+), and regulatory- (Foxp3+) memory cell (CD45RO+) populations and their activation status (CD69+) were analyzed using flow cytometry. qRT-PCR analysis was performed on decidua parietalis and basalis biopsies to detect mRNA expression of interferon-gamma, interleukin-1B, IL2, IL6, IL7, IL8, IL10, IL15, and IL23. Results: CD4+ central-memory (CM) cell proportions were lower in the decidua parietalis in LO-PE (P + memory (P + CM (P

Preeclampsia is one of the leading causes of maternal and neonatal mortality and morbidity worldwide, affecting 2–8% of all pregnancies. Studies suggest a link between complement activation and preeclampsia. The complement system plays an essential role in the innate immunity, leading to opsonization, inflammation, and elimination of potential pathogens. The complement system also provides a link between innate and adaptive immunity and clearance of immune complexes and apoptotic cells. During pregnancy there is increased activity of the complement system systemically. However, locally at the placenta, complement inhibition is crucial for the maintenance of a normal pregnancy. Inappropriate or excessive activation of the complement system at the placenta is likely involved in placental dysfunction, and is in turn associated with pregnancy complications like preeclampsia. Therefore, modulation of the complement system could be a potential therapeutic target to prevent pregnancy complications such as preeclampsia. This review, based on a systematic literature search, gives an overview of the complement system and its activation locally in the placenta and systemically during healthy pregnancies and during complicated pregnancies, with a focus on preeclampsia. Furthermore, this review describes results of animal and human studies with a focus on the complement system in pregnancy, and the role of the complement system in placental dysfunction. Various clinical and animal studies provide evidence that dysregulation of the complement system is associated with placental dysfunction and therefore with preeclampsia. Several drugs are used for prevention and treatment of preeclampsia in humans and animal models, and some of these drugs work through complement modulation. Therefore, this review further discusses these studies examining pharmaceutical interventions as treatment for preeclampsia. These observations will help direct research to generate new target options for prevention and treatment of preeclampsia, which include direct and indirect modulation of the complement system.

The development of microglia Microglia are the brain's immune cells, and they play important roles in health and neurodegenerative disease. Kracht et al. performed single-cell analysis of human microglial gene expression and chromatin accessibility and compared the results with those of other studies of human and mice microglial development. By using in situ validation, these data identify fetal microglial subsets that appear to be distinct from adult human microglia, suggesting functional differences between the developing and mature brain. Science , this issue p. 530

Disturbances in fetal neurodevelopment have extensively been related to neurodevelopmental disorders in early and later life. Fetal neurodevelopment is dependent on adequate functioning of the fetal immune system. During pregnancy, the maternal immune system is challenged to both tolerate the semi-allogenic fetus and to protect the mother and fetus from microbes. The fetal immune system is influenced by maternal immune disturbances; therefore, perturbations in maternal immunity likely do not only alter pregnancy outcome but also alter fetal neurodevelopment. A possible common pathway could be modulating the functioning of tissue macrophages in the placenta and brain.Maternal immune tolerance towards the fetus involves several complex adaptations. In this active maternal immune state, the fetus develops its own immunity. As cytokines and other players of the immune system-which can pass the placenta-are involved in neurodevelopment, disruptions in immune balance influence fetal neurodevelopment. Several studies reported an association between maternal immune activation, complications of pregnancy as preeclampsia, and altered neonatal neurodevelopment. A possible pathway involves dysfunctioning of microglia cells, the immune cells of the brain. Functionality of microglia cells during normal pregnancy is, however, poorly understood. The recent outbreak of ZIKA virus (ZKV), but also the literature on virus infections in general and its consequences on microglial cell function and fetal neurodevelopment show the devastating effects a virus infection during pregnancy can have.

Insufficient adaptations of the maternal immune system are associated with pregnancy complications such as preeclampsia. Memory T cells might be implicated in the pathophysiology of preeclampsia and its recurrence risk. Therefore, peripheral blood samples were taken from healthy pregnant women (n = 15), preeclamptic pregnant women (n = 15), formerly healthy pregnant women (n = 16), and formerly preeclamptic women (n = 15). CD4(+) and CD8(+) memory cells (CD45RO(+)), central-memory cells (CM, CD45RO(+) CCR7(+)), effector-memory cells (EM, CD45RO(+) CCR7(-)), and their activated (CD69(+)) proportions were analyzed using flow cytometry. A magnetic Luminex assay was performed on plasma samples from all groups to analyze 16 cytokines associated with memory T cells homeostasis and T cell differentiation. Proportions of CD4 + and CD8(+) memory cell populations did not differ between preeclamptic and healthy pregnant women or between formerly preeclamptic and formerly healthy pregnant women. However, activated proportions of the general CD4(+) memory, CD4(+) EM, and CD4(+) CM population in the peripheral blood were lower during preeclampsia compared to healthy pregnancies and were also lower postpartum in formerly preeclamptic compared to formerly healthy pregnant women. This was accompanied by lower IL2 concentrations in plasma from formerly preeclamptic compared to formerly healthy pregnant women. The lower activated proportions of memory T cells after a preeclamptic pregnancy were not associated with altered memory T cell associated cytokine plasma concentrations. These findings showed lower activation of memory CD4(+) T cell subsets in and after preeclampsia as compared with healthy pregnancies, which makes their implication in the preeclampsia recurrence risk likely.

Insufficient adaptations of the maternal immune system are associated with pregnancy complications such as preeclampsia. Memory T cells might be implicated in the pathophysiology of preeclampsia and its recurrence risk. Therefore, peripheral blood samples were taken from healthy pregnant women (n = 15), preeclamptic pregnant women (n = 15), formerly healthy pregnant women (n = 16), and formerly preeclamptic women (n = 15). CD4

Objective: Management of late fetal growth restriction (FGR) is limited to adequate fetal monitoring and optimal timing of delivery. The Disproportionate Intrauterine Growth Intervention Trial At Term (DIGITAT) trial compared induction of labor with expectant management in pregnancies at (near) term complicated by suspected FGR. Findings of the DIGITAT trial were that expectant monitoring prolonged pregnancy for 10 days and increased birth weight with only 130 grams. This resulted in more infants born below the 2.3rd percentile compared to induction of labor, respectively, 12.5% in induction of labor and 30.6% in expectant monitoring group. The main placental lesions associated with FGR are maternal vascular malperfusion, fetal vascular malperfusion, and villitis of unknown etiology. We investigated whether placentas of pregnancies complicated with FGR in the expectant monitoring group reveal more and more severe pathology due to pregnancy prolongation.Material and methods: The DIGITAT trial was a multicenter, randomized controlled trial with suspected FGR beyond 36 + 0 weeks. We now analyzed all available cases (n = 191) for placental pathology. The macroscopic details were collected and histological slides were recorded and classified by a single perinatal pathologist, blinded for pregnancy details and outcome. The different placental lesions were scored based on the latest international criteria for placental lesions as defined in the Amsterdam Placental Workshop Group Consensus Statement.Results: The presence of maternal vascular malperfusion and chorioamnionitis were higher in the expectant management group (0 Conclusion: Expectant management of late FGR is associated with increased maternal vascular malperfusion and chorioamnionitis. This may have implications for fetal and neonatal outcome, such as programming in the developing child influencing health outcomes later in life.

Adaptations of the maternal immune response are necessary for pregnancy success. Insufficient immune adaption is associated with pregnancy pathologies such as infertility, recurrent miscarriage, fetal growth restriction, spontaneous preterm birth, and preeclampsia. The maternal immune system is continuously exposed to paternal-fetal antigens; through semen exposure from before pregnancy, through fetal cell exposure in pregnancy, and through microchimerism after pregnancy. This results in the generation of paternal-fetal antigen specific memory T cells. Memory T cells have the ability to remember previously encountered antigens to elicit a quicker, more substantial and focused immune response upon antigen reencounter. Such fetal antigen specific memory T cells could be unfavorable in pregnancy as they could potentially drive fetal rejection. However, knowledge on memory T cells in pregnancy has shown that these cells might play a favorable role in fetal-maternal tolerance rather than rejection of the fetus. In recent years, various aspects of immunologic memory in pregnancy have been elucidated and the relevance and working mechanisms of paternal-fetal antigen specific memory T cells in pregnancy have been evaluated. The data indicate that a delicate balance of memory T cells seems necessary for reproductive success and that immunologic memory in reproduction might not be harmful for pregnancy. This review provides an overview of the different memory T cell subtypes and their function in the physiology and in complications of pregnancy. Current findings in the field and possible therapeutic targets are discussed. The findings of our review raise new research questions for further studies regarding the role of memory T cells in immune-associated pregnancy complications. These studies are needed for the identification of possible targets related to memory mechanisms for studies on preventive therapies.

Follicular fluid (FF) surrounds the granulosa cell–oocyte complex and is one of the mediating factors in the communication between the cells within the follicle. Literature reveals that human FF and its components are key factors to the success of natural fertilization. Among other substances, FF consists of multiple cytokines and immune cells, including interleukin 6 (IL6), IL12, sHLA-G, macrophages, NK cells and lymphocytes. Together, these cells and cytokines might influence the oocyte–granulosa–cell complex. Altered balances of immune content might be involved in changes on folliculogenesis, oocyte maturation, oocyte quality and ovulation. Furthermore, these altered balances are possibly involved in infertility associated with immune-mediated diseases such as endometriosis. The aim of this narrative review is to elaborate on the function and contents of FF and its immunological profile in patients with endometriosis. A comprehensive literature search was performed for the published literature on FF (immune) contents, FF function and FF content alterations in endometriosis patients. In FF of patients with endometriosis, elevated levels of macrophages and several cytokines have been reported. The role of specific immune cells in FF and a clarification of the biological mechanism in healthy women and endometriosis patients remain largely unknown. Future studies in this field will give us more insight in the role of FF immune cells and the effect of altered balances in patients with endometriosis.

Obesity is seen as a low grade inflammatory state, and is associated with adverse pregnancy outcomes. Disturbed macrophage characteristics might be essential in obesity associated pregnancy pathology via effects on the regulation of angiogenesis and placental development. This study aims to address the effects of maternal obesity on macrophage subsets in the decidua of women with term uncomplicated pregnancies. Macrophages were isolated from the decidua basalis and decidua parietalis of women with pre-gravid BMI 30 (obese). Macrophages were characterized and quantified using multi-color flow cytometry. Placentas of 10 obese and 10 control women after an uncomplicated term pregnancy were included. The decidua parietalis, but not decidua basalis, showed significantly lower levels of M1-type (HLA-DR+, CD163(-)) macrophages (p

IntroductionTo establish pregnancy, the maternal immune system must adapt to tolerate the semiallogenic fetus. Less than optimal adaptation of the maternal immune system during (early) pregnancy is implicated in several complications of pregnancy. The development of effective immune modulation interventions as preventive or therapeutic strategies for pregnancy complications holds promise. Several studies sought to evaluate the safety and effectiveness of various approaches. However, a limitation is the high variability in clinical and immune outcomes that are reported. We, therefore, aim to develop a core outcome set for application to studies of immune modulation in pregnancy (COSIMPREG).Methods and analysisWe will use a stepwise approach to develop a COSIMPREG. First, we will perform a systematic review to identify reported outcomes. For this review, Preferred Reporting Items for Systematic Reviews and Meta-Analyses guidelines will be followed. Second, we will use the Delphi method to develop a preliminary COSIMPREG. In three rounds, the outcomes of the systematic review will be scored. A panel comprising experts from relevant disciplines and diverse geographical locations will be assembled until a sufficient quality of the panel is reached. We will use predefined decision rules for outcomes. After each round outcomes, including scores, will be returned to the panel for further refinement. The outcomes not excluded after the third round will be taken to a consensus meeting. In this meeting, experts from all relevant disciplines will discuss and finalise the COSIMPREG.Ethics and disseminationFor this study ethical approval is not required. The systematic review will be published in an appropriate open access reproductive immunology journal. Once the COSIMPREG is finalised, it will be published in an open access reproductive immunology journal, and disseminated at appropriate international meetings, as well as through relevant research and scientific societies. Experts involved in the Delphi study will be asked to give informed consent.

Pregnancies with a male fetus are associated with higher risks of pregnancy complications through maladaptation of the maternal immune system. The pathophysiology of this phenomenon is unknown. A possible pathway could be a fetal sex-dependent maternal immune response, since males have a Y chromosome encoding specific allogenic proteins, possibly contributing to a different response and higher complication risks. To analyze whether fetal sex affects mRNA expression of maternal immune genes in early pregnancy, real-time PCR quantification was performed in the decidual tissue from primigravid pregnancies (n=20) between 10 and 12 weeks with uncomplicated term outcomes. Early-pregnancy decidual mRNA expression of the regulatory T-cell marker, FOXP3, was sixfold lower (p<0.01) in pregnancies with a male fetus compared to pregnancies with a female fetus. Additionally, mRNA expression of IFNγ was sixfold (p<0.05) lower in pregnancies with a male fetus. The present data imply maternal immunologic differences between pregnancies with male and female fetuses which could be involved in different pregnancy pathophysiologic outcomes. Moreover, this study indicates that researchers in reproductive immunology should always consider fetal sex bias.

Pregnancy is an immune challenge to the maternal immune system. The effects of pregnancy on maternal immunity and particularly on memory T cells during and after pregnancy are not fully known. This observational study aims to show the short term and the long term effects of pregnancy on the constitution, size and activation status of peripheral human memory T-lymphocyte populations. Effector memory (EM) and central memory (CM) T-lymphocytes were analyzed using flow cytometry of peripheral blood from 14 nulligravid, 12 primigravid and 15 parous women that were on average 18 months postpartum. The short term effects were shown by the significantly higher CD4+ EM cell and activated CD4+ memory cell proportions in primigravid women compared to nulligravid women. The persistent effects found in this study were the significantly higher proportions of CD4+ EM, CD4+ CM and activated memory T cells in parous women compared to nulligravid women. In contrast to CD4+ cells, activation status of CD8+ memory cells did not differ between the groups. This study shows that pregnancy persistently affects the pre-pregnancy CD4+ memory cell pool in human peripheral blood. During pregnancy, CD4+ T-Iymphocytes might differentiate into EM cells followed by persistent higher proportions of CD4+ CM and EM cells postpartum. The persistent effects of pregnancy on memory T cells found in this study support the hypothesis that memory T cells are generated during pregnancy and that these cells could be involved in the lower complication risks in multiparous pregnancies in humans. (C) 2016 The Authors. Published by Elsevier Ireland Ltd. This is an open access article under the CC BY license.

Recurrent miscarriage is a reproductive disorder affecting many couples. Although several factors are associated with recurrent miscarriage, in more than 50% of the cases the cause is unknown. Maladaptation of the maternal immune system is associated with recurrent miscarriage and could explain part of its pathophysiology. Modulating the maternal immune system toward pregnancy tolerance could benefit pregnancy outcome. Although there is a clear scientific rationale that modulating the maternal immune system could benefit recurrent miscarriage, only a few studies suggest possible beneficial effects of immune modulators as a therapy for recurrent miscarriage. Therapies skewing the maternal immune response to a tolerating regulatory T cell rich environment seem especially promising; however, more research is needed to find effective and safe maternal immune modulators for reproductive pathologies as recurrent miscarriage. Moreover, the possible side effects on maternal, fetal, and neonatal immune function are essentially unknown, and its elucidation is crucial before any possible therapeutic strategies could be clinically implemented. (C) 2014 Elsevier Ireland Ltd. All rights reserved.

Introduction Previous studies indicate that pregnancy persistently affects memory T lymphocyte populations. However, their function and relevance in fetal-maternal immune tolerance and therewith immune related complications of pregnancy are not known. This study aims to show differences in memory T lymphocyte populations in decidual tissue of pre-eclamptic versus healthy controls. Methods Decidua basalis and decidua parietalis from placentas from pre-eclamptic (n = 6) and healthy uncomplicated (n = 13) pregnancies were collected. In the decidual tissue, CD4 effector- (CCR7-), central- (CCR7+), tissue resident-(CD103+), and regulatory (FOXP3+) memory cell (CD45RO+) populations were analyzed using flow cytometry. Comparison was performed between placentas from pre-eclamptic patients and healthy controls using Mann–Whitney U test, p 0.05. Results The total memory T cell proportion (CD45RO+) of CD4 + T lymphocytes was significantly lower in the decidua parietalis from pre-eclamptic pregnancies compared to decidua parietalis of healthy controls. A trend (p 0.1) towards lower T effector memory and lower T central memory cells was observed in decidua basalis and parietalis from pre-eclamptic pregnancies compared to healthy controls. In the decidua basalis from pre-eclamptic patients, significantly lower expression of the activation marker CD69 was observed on all different memory cell subset that were analyzed. Discussion We show that CD4 + memory T lymphocytes at the fetal maternal interface are present at lower levels and have a lower activation status in decidual tissue of pre-eclamptic pregnancies compared to healthy uncomplicated pregnancies. These findings suppose reduced imprinted memory with possible consequences for the current and subsequent pregnancy.

T regulatory (Treg) cells are essential mediators of the maternal immune adaptation necessary for embryo implantation. In mice, insufficient Treg cell activity results in implantation failure, or constrains placental function and fetal growth. In women, Treg cell deficiency is linked with unexplained infertility, miscarriage, and pre-eclampsia. To devise strategies to improve Treg cell function, it is essential to define the origin of the Treg cells in gestational tissues, and the regulators that control their functional competence and recruitment. Male seminal fluid is a potent source of the Treg cell-inducing agents TGFβ and prostaglandin E, and coitus is one key factor involved in expanding the pool of inducible Treg cells that react with paternal alloantigens shared by conceptus tissues. In mice, coitus initiates a sequence of events whereby female dendritic cells cross-present seminal fluid antigens and activate T cells, which in turn circulate via the blood to be sequestered into the endometrium. Similar events may occur in the human genital tract, where seminal fluid induces immune cell changes that appear competent to prime Treg cells. Improved understanding of how seminal fluid influences Treg cells in women should ultimately assist in the development of new therapies for immune-mediated pathologies of pregnancy.

Regulatory T (Treg) cells facilitate maternal immune tolerance of the semiallogeneic conceptus in early pregnancy, but the origin and regulation of these cells at embryo implantation is unclear. During the preimplantation period, factors in the seminal fluid delivered at coitus cause expansion of a CD4(+)CD25(+) putative Treg cell population in the para-aortic lymph nodes draining the uterus. Using flow cytometry, immunohistochemistry, and real-time quantitative PCR (qPCR) for the signature Treg cell transcription factor FOXP3, we confirmed the identity of the expanded lymph node population as FOXP3(+) Treg cells and showed that this is accompanied by a comparable increase in the uterus of FOXP3(+) Treg cells and expression of Foxp3 mRNA by Day 3.5 postcoitum. Seminal plasma was necessary for uterine Treg cell accumulation, as mating with seminal vesicle-deficient males failed to elicit an increase in uterine Treg cells. Furthermore seminal fluid induced expression of mRNA encoding the Treg chemokine CCL19 (MIP3beta), which acts through the CCR7 receptor to regulate Treg cell recruitment and retention in peripheral tissues. Glandular and luminal epithelial cells were identified as the major cellular origins of uterine CCL19, and exposure to both seminal plasma and sperm was required for maximum expression. Together, these results indicate that Treg cells accumulate in the uterus prior to embryo implantation and that seminal fluid is a key regulator of the uterine Treg cell population, operating by both increasing the pool of available Treg cells and promoting their CCL19-mediated recruitment from the circulation into the implantation site.

Introduction During pregnancy, tolerance towards the fetus is induced by adaptations in the maternal immune system. Maladaptation of the maternal immune system is associated with pregnancy complications like preeclampsia and fetal growth restriction. Memory T-cells might play an important role in fetal-maternal tolerance. It is at the fetal-maternal interface where the maternal immune system especially needs to create tolerance towards the fetus. Decidua basalis is the maternal part of the basal plate whereas decidua parietalis is the maternal part of the fetal membranes. Decidual distribution pattern of memory T-cells during pregnancy is not known. Objective Since it is unknown how memory T-cells are distributed in decidual tissue, memory T-cells in decidua basalis and decidua parietalis were evaluated. Methods Lymphocytes were isolated from decidua basalis and decidua parietalis from uncomplicated term pregnancies. Using flowcytometry, subsets of memory T-cells (central, effector, regulatory, and tissue resident) of uncomplicated term pregnancies were analysed (n = 27). To compare different subsets of memory T-cells between decidual tissues, a student’s t-test (p Results Levels of memory T-cells (CD45RO+) were higher in decidua parietalis compared to decidua basalis in both CD4+ and CD8+ cells. Moreover, CD8+ central memory cells (CD45RO+CCR7+) and CD8+ tissue resident cells (CD45RO+CD103+) were higher in decidua parietalis compared to decidua basalis. Lastly, activation status (CD69+) was higher for all memory subsets (except for CD4+ effector memory T-cells) in decidua parietalis compared to decidua basalis. Discussion This study shows that memory T-cell subsets are differently distributed in decidua parietalis and basalis, with higher percentages of memory T-cell subsets as well as more activated memory T-cell subsets found in decidua parietalis compared to decidua basalis. Since memory T-cells are found to be present at higher percentages in decidua parietalis, this could suggest that these cells are especially important for creating tolerance to the fetal membranes.

BACKGROUND Adaptation of the maternal immune response to accommodate the semi-allogeneic fetus is necessary for pregnancy success, and disturbances in maternal tolerance are implicated in infertility and reproductive pathologies. T regulatory (Treg) cells are a recently discovered subset of T-lymphocytes with potent suppressive activity and pivotal roles in curtailing destructive immune responses and preventing autoimmune disease. METHODS A systematic review was undertaken of the published literature on Treg cells in the ovary, testes, uterus and gestational tissues in pregnancy, and their link with infertility, miscarriage and pathologies of pregnancy. An overview of current knowledge on the generation, activation and modes of action of Treg cells in controlling immune responses is provided, and strategies for manipulating regulatory T-cells for potential applications in reproductive medicine are discussed. RESULTS Studies in mouse models show that Treg cells are essential for maternal tolerance of the conceptus, and that expansion of the Treg cell pool through antigen-specific and antigen non-specific pathways allows their suppressive actions to be exerted in the critical peri-implantation phase of pregnancy. In women, Treg cells accumulate in the decidua and are elevated in maternal blood from early in the first trimester. Inadequate numbers of Treg cells or their functional deficiency are linked with infertility, miscarriage and pre-eclampsia. CONCLUSIONS The potency and wide-ranging involvement of Treg cells in immune homeostasis and disease pathology indicates the considerable potential of these cells as therapeutic agents, raising the prospect of their utility in novel treatments for reproductive pathologies.

Objective: Immunological mechanisms are involved in the pathophysiology of preeclampsia. During pregnancy there is an increase in regulatory T (Treg) cells, which has an important role in regulating tolerance to the immunologically distinct fetus. We hypothesised that percentages of Treg cells are decreased in preeclamptic patients. Methods: Peripheral blood was obtained from 26 healthy pregnant controls and 18 preeclamptic patients. Treg cells were measured using flow-cytometry. Results: Women with pregnancies complicated by preeclampsia had significantly lower percentages of CD4(+)FOXP3(+) Treg cells. Conclusion: We conclude that a deficiency of regulatory T cells may play a role in the pathophysiology of preeclampsia.

Maternal interleukin (IL) 10 deficiency elevates susceptibility to fetal loss induced by the model Toll-like receptor agonist lipopolysaccharide, but the mechanisms are not well elucidated. Here, we show that Il10 null mutant (Il10(-/-)) mice exhibit altered local T cell responses in pregnancy, exhibiting pronounced hyperplasia in para-aortic lymph nodes draining the uterus with >6-fold increased CD4(+) and CD8(+) T cells compared with wild-type controls. Among these CD4(+) cells, Foxp3(+) T regulatory (Treg) cells were substantially enriched, with 11-fold higher numbers at Day 9.5 postcoitum. Lymph node hypertrophy in Il10(-/-) mice was associated with more activated phenotypes in dendritic cells and macrophages, with elevated expression of MHCII, scavenger receptor, and CD80. Affymetrix microarray revealed an altered transcriptional profile in Treg cells from pregnant Il10(-/-) mice, with elevated expression of Ctse (cathepsin E), Il1r1, Il12rb2, and Ifng. In vitro, Il10(-/-) Treg cells showed reduced steady-state Foxp3 expression, and polyclonal stimulation caused greater loss of Foxp3 and reduced capacity to suppress IL17 in CD4(+)Foxp3(-) T cells. We conclude that despite a substantially expanded Treg cell pool, the diminished stability of Treg cells, increased numbers of effector T cells, and altered phenotypes in dendritic cells and macrophages in pregnancy all potentially confer vulnerability to inflammation-induced fetal loss in Il10(-/-) mice. These findings suggest that IL10 has a pivotal role in facilitating robust immune protection of the fetus from inflammatory challenge and that IL10 deficiency could contribute to human gestational disorders in which altered T cell responses are implicated.

Maternal interleukin (IL) 10 deficiency elevates susceptibility to fetal loss induced by the model Toll-like receptor agonist lipopolysaccharide, but the mechanisms are not well elucidated. Here, we show that Il10 null mutant (Il10(-/-)) mice exhibit altered local T cell responses in pregnancy, exhibiting pronounced hyperplasia in para-aortic lymph nodes draining the uterus with6-fold increased CD4(+) and CD8(+) T cells compared with wild-type controls. Among these CD4(+) cells, Foxp3(+) T regulatory (Treg) cells were substantially enriched, with 11-fold higher numbers at Day 9.5 postcoitum. Lymph node hypertrophy in Il10(-/-) mice was associated with more activated phenotypes in dendritic cells and macrophages, with elevated expression of MHCII, scavenger receptor, and CD80. Affymetrix microarray revealed an altered transcriptional profile in Treg cells from pregnant Il10(-/-) mice, with elevated expression of Ctse (cathepsin E), Il1r1, Il12rb2, and Ifng. In vitro, Il10(-/-) Treg cells showed reduced steady-state Foxp3 expression, and polyclonal stimulation caused greater loss of Foxp3 and reduced capacity to suppress IL17 in CD4(+)Foxp3(-) T cells. We conclude that despite a substantially expanded Treg cell pool, the diminished stability of Treg cells, increased numbers of effector T cells, and altered phenotypes in dendritic cells and macrophages in pregnancy all potentially confer vulnerability to inflammation-induced fetal loss in Il10(-/-) mice. These findings suggest that IL10 has a pivotal role in facilitating robust immune protection of the fetus from inflammatory challenge and that IL10 deficiency could contribute to human gestational disorders in which altered T cell responses are implicated.

During pregnancy the maternal immune system has to coordinate uterine spiral-artery remodelling, trophoblast invasion, and acceptance of the semi-allogenic fetus simultaneously. As dysregulation of the immune system is associated with adverse pregnancy outcomes, we analysed first-trimester deciduas of pregnancies for immune parameters in later complicated pregnancies. Higher IL6 and macrophage mRNA expression, and lower ratios of regulatory macrophages were found in first-trimester deciduas of pregnancies later complicated with pregnancy-induced hypertension. Lower Gata3 (Th2) mRNA expression was found in deciduas of pregnancies with later foetal growth restriction. Our results suggest that adverse pregnancy outcomes are associated with immunological disturbances in first-trimester deciduas. (C) 2012 Elsevier Ltd. All rights reserved.

IL6 is a multifunctional cytokine with pivotal roles in the inflammatory response and in directing T cell differentiation in adaptive immunity. IL6 is widely expressed in the female reproductive tract and gestational tissues, and exerts regulatory functions in embryo implantation and placental development, as well as the immune adaptations required to tolerate pregnancy. Here, we summarise the current understanding of how membrane-bound and soluble receptors mediate IL6 signalling to regulate leukocytes and non-haemopoietic cells. We review the published literature regarding the expression and actions of IL6 in the uterus, decidua and placenta, and studies implicating this cytokine in pregnancy disorders. Elevated IL6 is frequently evident in the altered cytokine profiles characteristic of unexplained infertility, recurrent miscarriage, preeclampsia and preterm delivery. Notably, there is compelling evidence indicating altered systemic IL6 trans-signalling in women prone to recurrent miscarriage, with excessive IL6 bioavailability potentially inhibiting generation of CD4+ T regulatory cells required for pregnancy tolerance. Insufficient local IL6 may also contribute to fetal loss, since IL6 expression is reduced in the endometrium of women with recurrent miscarriage, and in the fetal-placental tissue of CBA x DBA/2 mice. Consistent with the role of 16 in key reproductive events, 116 null mutant mice exhibit elevated fetal resorption and delayed parturition. Investigation of the association between IL6 signalling components and T cell responses in pregnant women, as well as detailed analysis of the maternal immune response in IL6-deficient mice, is now required to define the mechanisms by which this cytokine exerts influence on reproductive success. (C) 2012 Elsevier Ireland Ltd. All rights reserved.

Regulatory T (Treg) cells facilitate maternal immune tolerance of the semiallogeneic conceptus in early pregnancy, but the origin and regulation of these cells at embryo implantation is unclear. During the preimplantation period, factors in the seminal fluid delivered at coitus cause expansion of a CD4(+)CD25(+) putative Treg cell population in the para-aortic lymph nodes draining the uterus. Using flow cytometry, immunohistochemistry, and real-time quantitative PCR (qPCR) for the signature Treg cell transcription factor FOXP3, we confirmed the identity of the expanded lymph node population as FOXP3(+) Treg cells and showed that this is accompanied by a comparable increase in the uterus of FOXP3(+) Treg cells and expression of Foxp3 mRNA by Day 3.5 postcoitum. Seminal plasma was necessary for uterine Treg cell accumulation, as mating with seminal vesicle-deficient males failed to elicit an increase in uterine Treg cells. Furthermore seminal fluid induced expression of mRNA encoding the Treg chemokine CCL19 (MIP3beta), which acts through the CCR7 receptor to regulate Treg cell recruitment and retention in peripheral tissues. Glandular and luminal epithelial cells were identified as the major cellular origins of uterine CCL19, and exposure to both seminal plasma and sperm was required for maximum expression. Together, these results indicate that Treg cells accumulate in the uterus prior to embryo implantation and that seminal fluid is a key regulator of the uterine Treg cell population, operating by both increasing the pool of available Treg cells and promoting their CCL19-mediated recruitment from the circulation into the implantation site.

OBJECTIVE: During pregnancy the maternal immune system has to adapt its response to accommodate the fetus. The objective of this study was to analyze the effects of smoking on the maternal immune system.STUDY DESIGN: First-trimester decidual tissue and peripheral blood of smoking and nonsmoking women were analyzed by real time reverse transcription-polymerase chain reaction (RT-PCR) and flow cytometry. A mouse model was used to further analyze the effects of smoking. Murine tissue was analyzed by flow cytometry, real-time RT-PCR, and immunohistochemistry.RESULTS: Smoking caused lower percentages of viable pups in mice and lower birthweights in humans. Smoking mothers, both mice and human, had more natural killer cells and inflammatory macrophages locally, whereas systemically they had lower percentages of regulatory T cells than nonsmoking controls.CONCLUSION: Maternal smoke exposure during pregnancy influences local and systemic immune responses in both women and mice. Such changes may be involved in adverse pregnancy outcomes in smoking individuals.