Clare Tower

Macrophages Infiltrate the Human and Rat Decidua During Term and Preterm Labor: Evidence That Decidual Inflammation Precedes Labor

ABSTRACT Preterm delivery is the leading cause of perinatal mortality and morbidity. Current tocolytics target myometrial contractions, a late step in the labor cascade. Identifying earlier events in parturition may lead to more effective therapeutic strategies. We hypothesized that inflammatory events in decidua (the maternal-fetal interface), characterized by leucocyte infiltration, are an early event during term and preterm labor (PTL). Leucocyte abundance in decidua of human pregnancies was quantified following term labor and PTL (idiopathic and infection associated), in conjunction with investigation of temporal inflammatory events in rat uterus during the perilabor period and in PTL induced by mifepristone. In human decidua, macrophage numbers were 4-fold higher in term labor (P < 0.01) and 2.5-fold higher in non-infection-associated PTL (P < 0.05) than in term nonlaboring samples. Neutrophil abundance was unchanged with labor but elevated in PTL with infection (5- to 53-fold increase; P < 0.01). T and NK cells were more abundant in idiopathic PTL than TL (P < 0.05). In rat, decidual macrophage infiltration increased 4.5-fold 12 h prior to labor and remained elevated during labor and early postpartum (P < 0.01). Decidual infiltration preceded that of the myometrium and was 4-fold higher (P < 0.01). In rat PTL, decidual macrophage numbers were also elevated (P < 0.01) and exceeded those of the myometrium (P < 0.05). These studies show for the first time that leucocytes infiltrate decidua during labor at term and preterm, supporting a role for leucocyte-derived inflammatory mediators in decidual activation. In the rat, this occurred prior to labor, suggesting it is an early event during parturition and thus a potential target for intervention.

Immune cell activation by trophoblast-derived microvesicles is mediated by syncytin 1

Envelope glycoproteins of human endogenous retrovirus (HERV), such as syncytin 1 (HERV‐W), are highly expressed in the placenta and some family members have immunomodulatory properties. Placental microvesicles (MV), which are shed into the maternal circulation during pregnancy, have been demonstrated to induce immune cell activation. Therefore, the aim of this study was to investigate the immunological properties of the highly expressed placental HERV‐W protein, syncytin 1, and its potential involvement in placental MV modulation of immune cell activity. The MV shed from first trimester, normal term and pre‐eclamptic term placentas, and from the BeWo trophoblast cell line, all contain syncytin 1. Recombinant syncytin 1 and syncytin 1‐positive BeWo trophoblast MV both induced peripheral blood mononuclear cell (PBMC) activation, indicated through production of cytokines and chemokines. Reducing syncytin 1 content in BeWo MV inhibited PBMC activation. Recombinant syncytin 1 and syncytin‐1‐positive BeWo MV dampened PBMC responses to lipopolysaccharide challenge. Our findings suggest that syncytin 1 is shed from the placenta into the maternal circulation in association with MV, and modulates immune cell activation and the responses of immune cells to subsequent lipopolysaccharide stimulation. These studies implicate placental MV‐associated HERV in fetal regulation of the maternal immune system.

Identification of Chemokines Associated with the Recruitment of Decidual Leukocytes in Human Labour: Potential Novel Targets for Preterm Labour

Current therapies for preterm labour (PTL) focus on arresting myometrial contractions but are largely ineffective, thus alternative therapeutic targets need to be identified. Leukocytes infiltrate the uterus around the time of labour, and are in particularly abundant in decidua (maternal-fetal interface). Moreover, decidual inflammation precedes labour in rat pregnancies and thus may contribute to initiation of labour. We hypothesized that chemokines mediate decidual leukocyte trafficking during preterm labour (PTL) and term labour (TL), thus representing potential targets for preventing PTL. Women were recruited into 4 groups: TL, term not in labour (TNL), idiopathic PTL and PTL with infection (PTLI). Choriodecidual RNA was subjected to a pathway-specific PCR array for chemokines. Differential expression of 12 candidate chemokines was validated by real time RT-PCR and Bioplex assay, with immunohistochemistry to confirm cellular origin. 25 chemokines were upregulated in choriodecidua from TL compared to TNL. A similar pattern was detected in PTL, however a distinct profile was observed in PTLI consistent with differences in leukocyte infiltration. Upregulation of CCL2, CCL4, CCL5, CXCL8 and CXCL10 mRNA and protein was confirmed in TL, with CCL8 upregulated in PTL. Significant correlations were detected between these chemokines and decidual leukocyte abundance previously assessed by immunohistochemical and image analysis. Chemokines were primarily expressed by decidual stromal cells. In addition, CXCL8 and CCL5 were significantly elevated in maternal plasma during labour, suggesting chemokines contribute to peripheral inflammatory events during labour. Differences in chemokine expression patterns between TL and idiopathic PTL may be attributable to suppression of chemokine expression by betamethasone administered to women in PTL; this was supported by in vitro evidence of chemokine downregulation by clinically relevant concentrations of the steroid. The current study provides compelling evidence that chemokines regulate decidual leukocyte recruitment during labour. The 6 chemokines identified represent potential novel therapeutic targets to block PTL.

Heightened Pro-Inflammatory Effect of Preeclamptic Placental Microvesicles on Peripheral Blood Immune Cells in Humans

ABSTRACT Normal pregnancy is associated with the presence of circulating placental microvesicles (MVs). Increased MV shedding and altered immune activation are seen in patients with preeclampsia, suggesting that placental MVs may play a role in the pathophysiology of this disease. Therefore, the aim of this study was to investigate the activation of peripheral blood mononuclear cells (PBMCs) by MVs shed by first-trimester, normal term, and preeclamptic term placenta. First-trimester and preeclamptic term, but not normal term, placental-derived MVs activated PBMCs, as evidenced by elevated IL1B. Significant changes were also seen with several other cytokines and chemokines, and in general when compared to normal term MVs, preeclamptic MVs induced a greater pro-inflammatory response in PBMCs. Pretreatment of PBMCs with first-trimester or normal term placental MVs resulted in a dampened IL1B response to a subsequent lipopolysaccharide (LPS) challenge. In contrast, treatment of PBMCs with preeclamptic term placental MVs exacerbated the LPS response. This was also the case for several other cytokines and chemokines. These studies suggest that placental MVs can modulate basal peripheral immune cell activation and responsiveness to LPS during normal pregnancy, and that in preeclampsia this effect is exacerbated.

SLE and pregnancy: the potential role for regulatory T cells

Systemic lupus erythematosus (SLE) is a multisystem autoimmune disorder that disproportionally affects women, especially in their reproductive years. SLE is associated with considerable pregnancy-related morbidity—including fetal loss, preterm birth, fetal growth restriction and pre-eclampsia. CD4+CD25+ regulatory T (TREG) cells have a potent immunosuppressive function and contribute to immunological self-tolerance. These cells might be essential for successful placental development by ensuring fetal tolerance. The numbers of TREG cells are augmented during normal pregnancy and, conversely, diminished numbers are associated with pregnancy loss and pre-eclampsia. Several studies have shown that patients with SLE have decreased numbers of TREG cells that might be functionally defective. This defective TREG cell functioning could predispose women with SLE to pregnancy complications. This article provides an overview of current knowledge of the role and function of TREG cells in SLE and pregnancy and how these cells might contribute to improving pregnancy-related outcomes in patients with SLE in the future.

Syncytin 1 in the human placenta

This study characterises HERV-W (syncytin 1) expression in normal and pathologic placenta and in BeWo cells. HERV-W mRNA levels were higher in the first trimester than at term, and similar patterns were observed with another retrovirally-derived mRNA species, ERV-3. N-glycosylated syncytin 1 precursor (73 kDa) is cleaved to surface-associated (SU) and transmembrane (TM) subunits. Both were evident in villous trophoblast, where perinuclear and punctate cytoplasmic deposits were observed, and linear TM subunit immunoreactivity was seen at the syncytial microvillous membrane. Punctate immunoreactivity was seen in BeWo cells with antibodies to SU and TM, and the two were co-localised. SU immunoreactivity was observed in association with fetal endothelium, and this effect was increased in tissue from pre-eclamptic placentas, which also showed a higher level of total SU protein. Absence of the TM subunit from endothelium suggests it is not a biosynthetic source. We suggest that SU is released from trophoblast into fetal circulation where it may bind vascular endothelium.

Thrombophilia and Pregnancy Complications.

There is a paucity of strong evidence associated with adverse pregnancy outcomes and thrombophilia in pregnancy. These problems include both early (recurrent miscarriage) and late placental vascular-mediated problems (fetal loss, pre-eclampsia, placental abruption and intra-uterine growth restriction). Due to poor quality case-control and cohort study designs, there is often an increase in the relative risk of these complications associated with thrombophilia, particularly recurrent early pregnancy loss, late fetal loss and pre-eclampsia, but the absolute risk remains very small. It appears that low-molecular weight heparin has other benefits on the placental vascular system besides its anticoagulant properties. Its use is in the context of antiphospholipid syndrome and recurrent pregnancy loss and also in women with implantation failure to improve live birth rates. There is currently no role for low-molecular weight heparin to prevent late placental-mediated complications in patients with inherited thrombophilia and this may be due to small patient numbers in the studies involved in summarising the evidence. There is potential for low-molecular weight heparin to improve pregnancy outcomes in women with prior severe vascular complications of pregnancy such as early-onset intra-uterine growth restriction and pre-eclampsia but further high quality randomised controlled trials are required to answer this question.

Detrimental Effects of Ethanol and Its Metabolite Acetaldehyde, on First Trimester Human Placental Cell Turnover and Function

Fetal alcohol spectrum disorder (FASD) describes developmental issues from high maternal alcohol intake, which commonly results in fetal growth restriction and long term morbidity. We aimed to investigate the effect of alcohol and acetaldehyde, on the first trimester placenta, the period essential for normal fetal organogenesis. Normal invasion and establishment of the placenta during this time are essential for sustaining fetal viability to term. We hypothesise that alcohol (ethanol) and acetaldehyde have detrimental effects on cytotrophoblast invasion, turnover and placental function. Taurine is an important amino acid for neuronal and physiological development, and so, its uptake was assayed in cells and placental explants exposed to alcohol or acetaldehyde. First trimester villous explants and BeWo cells were treated with 0, 10, 20, 40 mM ethanol or 0, 10, 20, 40 µM acetaldehyde. The invasive capacity of SGHPL4, a first trimester extravillous cytotrophoblast cell line, was unaffected by ethanol or acetaldehyde (p>0.05; N = 6). The cells in-cycle were estimated using immunostaining for Ki67. Proliferating trophoblast cells treated with ethanol were decreased in both experiments (explants: 40% at 20 mM and 40 mM, p<0.05, N = 8–9) (cell line: 5% at 20 mM and 40 mM, p<0.05, N = 6). Acetaldehyde also reduced Ki67-positive cells in both experiments (explants at 40 µM p<0.05; N = 6) (cell line at 10 µM and 40 µM; p<0.05; N = 7). Only in the cell line at 20 µM acetaldehyde demonstrated increased apoptosis (p<0.05; N = 6). Alcohol inhibited taurine transport in BeWo cells at 10 mM and 40 mM (p<0.05; N = 6), and in placenta at 40 mM (p<0.05; N = 7). Acetaldehyde did not affect taurine transport in either model (P<0.05; N = 6). Interestingly, system A amino acid transport in placental explants was increased at 10 µM and 40 µM acetaldehyde exposure (p<0.05; N = 6). Our results demonstrate that exposure to both genotoxins may contribute to the pathogenesis of FASD by reducing placental growth. Alcohol also reduces the transport of taurine, which is vital for developmental neurogenesis.

Regulatory T cells in Systemic Lupus Erythematosus and Pregnancy

Systemic lupus erythematosus (SLE) is a chronic multisystem autoimmune disorder that predominantly affects women of reproductive age. As clinical outcomes improve, pregnancy in these women is becoming more common. Although epidemiological data have documented an improvement in the prognosis of pregnancy in these women over recent years, they are still at significantly increased risk of pregnancy complications, such as miscarriage, stillbirth, pre‐eclampsia and impaired foetal growth. The pathogenesis of SLE involves marked immune dysfunction, and in particular, the function of immunosuppressive elements of the immune system is impaired, including regulatory T‐cell function. Because regulatory T cells are likely to be the key cell‐modulating feto‐maternal tolerance, this review overviews the possibility that regulatory T‐cell impairments contribute to pregnancy pathology in women with SLE and contribute to the clinical challenge of managing these women during pregnancy.

Differential expression of angiotensin II type 1 and type 2 receptors at the maternal–fetal interface: potential roles in early placental development

Angiotensin II (Ang II) is locally generated in the placenta and regulates syncytial transport, vascular contractility and trophoblast invasion. It acts through two receptor subtypes, AGTR1 and AGTR2 (AT1 and AT2), which typically mediate antagonising actions. The objectives of this study are to characterise the cellular distribution of AGTR1 and AGTR2 at the maternal-fetal interface and explore the effects on cytotrophoblast turnover. Low levels of AGTR2 mRNA were detected in first trimester placental homogenates using real-time PCR. Immunohistochemistry using polyclonal antibodies against AGTR1 and AGTR2 detected the receptors in first trimester placenta, decidua basalis and villous tip outgrowths in culture. Serial staining with cytokeratin-7 was used to identify extravillous trophoblasts (EVTs). AGTR1 was found in the syncytiotrophoblast microvillous membrane, in a subpopulation of villous cytotrophoblasts, and in Hofbauer cells. AGTR1 was strongly upregulated in cytotrophoblasts in cell columns and villous tip outgrowths, but was absent in interstitial and endovascular EVTs within the decidua. AGTR2 immunostaining was present in Hofbauer cells and villous cytotrophoblasts, but was absent from syncytiotrophoblast. Faint staining was detected in cell column cytotrophoblasts and villous outgrowths, but not in EVTs within the decidua. Both receptors were detected in placental homogenates by western blotting. Ang II significantly increased proliferation of cytotrophoblasts in both villous explants and villous tip outgrowths, but did not affect apoptosis. Blockade of AGTR1 and AGTR2 together abrogated this effect. This study shows specific expression patterns for AGTR1 and AGTR2 in distinct trophoblast populations at the maternal-fetal interface and suggests that Ang II plays a role in placental development and generation of EVTs.