Regulatory T cells (Treg) expand during pregnancy and are present at the fetal-maternal interface at very early stages in pregnancy. The migration mechanisms of Treg to the pregnant uterus are still unclear. Human chorionic gonadotropin (hCG) is secreted by the blastocyst immediately after fertilization and has chemoattractant properties. Therefore, we sought to analyze whether hCG secreted by early trophoblasts attracts Treg to the uterus and hence contributes to maternal tolerance toward the fetus. Decidua and placenta tissue samples from patients having spontaneous abortions or ectopic pregnancies were employed to evaluate Treg and hCG levels. Age-matched samples from normal pregnant women served as controls. We further performed in vitro studies with primary first trimester trophoblast cells and a choriocarcinoma cell line (JEG-3) aiming to evaluate the ability of secreted hCG to attract Treg. Patients having miscarriages or ectopic pregnancy presented significantly decreased hCG mRNA and protein levels associated with decreased Foxp3, neuropilin-1, IL-10, and TGF-β mRNA levels as compared with normal pregnant women. Using migration assays we demonstrated that Treg were attracted by hCG-producing trophoblasts or choriocarcinoma cells. Treg migration toward cells transfected with hCG expression vectors confirmed the chemoattractant ability of hCG. Our data clearly show that hCG produced by trophoblasts attracts Treg to the fetal-maternal interface. High hCG levels at very early pregnancy stages ensure Treg to migrate to the site of contact between paternal Ags and maternal immune cells and to orchestrate immune tolerance toward the fetus.

SummaryT helper type 17 (Th17) and regulatory T (Treg) cells are active players in the establishment of tolerance and defence. These attributes of the immune system enmesh to guarantee the right level of protection. The healthy immune system, on the one hand, recognizes and eliminates dangerous non-self pathogens and, on the other hand, protects the healthy self. However, there are circumstances where this fine balance is disrupted. In fact, in situations such as in pregnancy, the foreign fetal antigens challenge the maternal immune system and Treg cells will dominate Th17 cells to guarantee fetal survival. In other situations such as autoimmunity, where the Th17 responses are often overwhelming, the immune system shifts towards an inflammatory profile and attacks the healthy tissue from the self. Interestingly, autoimmune patients have meliorating symptoms during pregnancy. This connects with the antagonist role of Th17 and Treg cells, and their specific profiles during these two immune challenging situations. In this review, we put into perspective the Th17/ Treg ratio during pregnancy and autoimmunity, as well as in pregnant women with autoimmune conditions. We further review existing systems biology approaches that study specific mechanisms of these immune cells using mathematical modelling and we point out possible future directions of investigation. Understanding what maintains or disrupts the balance between these two opponent yet reciprocal cells in healthy physiological settings, sheds light into the development of innovative pharmacological approaches to fight pregnancy loss and autoimmunity.

How the semi-allogeneic fetus is tolerated by the maternal immune system remains a fascinating phenomenon. Despite extensive research activity in this field, the mechanisms underlying fetal tolerance are still not well understood. However, there are growing evidences that immune–immune interactions as well as immune–endocrine interactions build up a complex network of immune regulation that ensures fetal survival within the maternal uterus. In the present review, we aim to summarize emerging research data from our and other laboratories on immune modulating properties of pregnancy hormones with a special focus on progesterone, estradiol, and human chorionic gonadotropin. These pregnancy hormones are critically involved in the successful establishment, maintenance, and termination of pregnancy. They suppress detrimental maternal alloresponses while promoting tolerance pathways. This includes the reduction of the antigen-presenting capacity of dendritic cells (DCs), monocytes, and macrophages as well as the blockage of natural killer cells, T and B cells. Pregnancy hormones also support the proliferation of pregnancy supporting uterine killer cells, retain tolerogenic DCs, and efficiently induce regulatory T (Treg) cells. Furthermore, they are involved in the recruitment of mast cells and Treg cells into the fetal–maternal interface contributing to a local accumulation of pregnancy-protective cells. These findings highlight the importance of endocrine factors for the tolerance induction during pregnancy and encourage further research in the field.

Various physiologically relevant processes are regulated by the interaction of the receptor tyrosine kinase (c-Kit) and its ligand stem cell factor (SCF), with SCF known to be the most important growth factor for mast cells (MCs). In spite of their traditional role in allergic disorders and innate immunity, MCs have lately emerged as versatile modulators of a variety of physiologic and pathologic processes. Here we show that MCs are critical for pregnancy success. Uterine MCs presented a unique phenotype, accumulated during receptivity and expanded upon pregnancy establishment. KitW-sh/W-sh mice, whose MC deficiency is based on restricted c-Kit gene expression, exhibited severely impaired implantation, which could be completely rescued by systemic or local transfer of wild-type bone marrow-derived MCs. Transferred wild-type MCs favored normal implantation, induced optimal spiral artery remodeling and promoted the expression of MC proteases, transforming growth factor-β and connective tissue growth factor. MCs contributed to trophoblast survival, placentation and fetal growth through secretion of the glycan-binding protein galectin-1. Our data unveil unrecognized roles for MCs at the fetomaternal interface with critical implications in reproductive medicine.

Normal pregnancy is characterized by an early expansion of regulatory T cells (Tregs), which is known to contribute to fetal tolerance. However, mechanisms and factors behind Treg expansion are not yet defined. Recently, we proposed that the pregnancy hormone human chorionic gonadotropin (hCG) efficiently attracts human Tregs to trophoblasts, favoring their accumulation locally. In this study, we hypothesized that hCG not only acts as a chemoattractant of Tregs but also plays a central role in pregnancy-induced immune tolerance. Virgin, normal pregnant, and abortion-prone female mice were treated either with 10 IU/ml hCG or PBS at days 0, 2, 4, and 6 of pregnancy. The hCG effect on Treg frequency and cytokine secretion was determined in Foxp3gfp females. hCG impact on Treg suppressive capacity was studied in vitro. In vivo, we investigated whether hCG enhances Treg suppressive capacity indirectly by modulating dendritic cell maturation in an established mouse model of disturbed fetal tolerance. Application of hCG increased Treg frequency in vivo and their suppressive activity in vitro. In females having spontaneous abortions, hCG provoked not only an augmentation of Treg numbers, but also normalized fetal abortion rates. hCG-generated Tregs were fully functional and could confer tolerance when adoptively transferred. hCG also retained dendritic cells in a tolerogenic state that is likely to contribute to both Treg expansion and prevention of abortion. Our results position hCG in a novel, so far unknown role as modulator of immune tolerance during pregnancy.

Breg emerge as important players in pregnancy; they suppress undesired immune responses from maternal T cells and are therefore important for tolerance acquisition.

Implantation of the fertilized egg into the maternal uterus depends on the fine balance between inflammatory and anti-inflammatory processes. Whilst regulatory T cells (Tregs) are reportedly involved in protection of allogeneic fetuses against rejection by the maternal immune system, their role for pregnancy to establish, e.g., blastocyst implantation, is not clear. By using 2-photon imaging we show that Foxp3+ cells accumulated in the mouse uterus during the receptive phase of the estrus cycle. Seminal fluid further fostered Treg expansion. Depletion of Tregs in two Foxp3.DTR-based models prior to pairing drastically impaired implantation and resulted in infiltration of activated T effector cells as well as in uterine inflammation and fibrosis in both allogeneic and syngeneic mating combinations. Genetic deletion of the homing receptor CCR7 interfered with accumulation of Tregs in the uterus and implantation indicating that homing of Tregs to the uterus was mediated by CCR7. Our results demonstrate that Tregs play a critical role in embryo implantation by preventing the development of a hostile uterine microenvironment.

Abstract-Heme Oxygenase-1 (HO-1) and its metabolite carbon monoxide (CO) promote implantation and placentation.Pregnancy disorders such as preeclampsia and intrauterine growth restriction are linked to both HO-1 diminution and impaired remodeling of maternal spiral arteries (SAs). Here, we investigated whether CO is able to prevent preeclampsia and intrauterine growth restriction through the modulation of uterine natural killer (uNK) cells that are necessary for initiation of SA remodeling. Hmox1 +/− or Hmox1 −/− implantations presented fewer uNK cell numbers and lower expression of uNK-related angiogeneic factors compared with Hmox1 +/+ sites. Quantitative histology revealed that Hmox1 +/− and Hmox1 −/− implantations had shallow SA development that was accompanied by intrauterine growth restriction and gestational hypertension. Application of CO at low dose during early to midgestation prevented intrauterine growth restriction in Hmox1 +/− mothers, this being associated with enhanced in situ proliferation of uNK cells and normalization of angiogenic parameters. Most importantly, CO improved SA remodeling and normalized blood pressure, ensuring a proper fetal growth. Thus, CO emerges as a key molecular player in pregnancy success by modulating uNK cells, which results in promotion of SA remodeling, adequate fetal support/growth, and prevention of hypertension. Linzke et al Targeting uNK Cells to Suppress Hypertension 581 Materials and Methods AnimalsFor analysis of implantation sites, Hmox1-competent or Hmox1-deficient BALB/c mice were used. These mice were initially provided by Dr Saw Feng-Yet and bred and maintained in our facility. The progeny obtained from breeding Hmox1 +/− female animals and Hmox1 +/− male animals were genotyped and recorded. C57BL/6 male animals were obtained from Charles River (Sulzfeld, Germany). The mice were maintained in our husbandry with a 12-hour light/ dark cycle and with food and water ad libitum. All experiments were approved by German authorities (Saxony-Anhalt's Ministry 2 -868).Hmox1 For all experiments, the detection of a vaginal plug indicated day 0 of pregnancy. Animals were euthanized on days 8, 10, 12, or 18 of pregnancy. Whole implantation sites for paraffin embedding were kept in 4% paraformaldehyde with 0.1% saccharose for 6 hours. For RNA isolation, tissue of MLAp and DB was carefully washed with cold sterile PBS (pH 7.4), snap frozen, and stored at −80°C. Tissue of MLAp and DB was stored in RPMI 1640 with 10% FBS and 1% penicillin and streptomycin for flow cytometry. CO InhalationFollowing our standard protocol, Hmox1 +/+ or Hmox1 +/− female animals were exposed to CO (50 ppm) day-long from day 3 to 8 of pregnancy. 5,6 Control animals inhaled the same air mixture without CO. The exposure of animals to mixed air or CO took place in cages placed in a 98-L Plexiglas animal chamber (A-Chamber; BioSpherix, NY). Duration of exposure and dosage was previously tested and established. Both time exposure and dose prevented fetal death without any toxic side effects. 5,6 Blo...