Placental syncytiotrophoblast constitutes a major barrier to vertical transmission of Listeria monocytogenes.
ABSTRACT: Listeria monocytogenes is an important cause of maternal-fetal infections and serves as a model organism to study these important but poorly understood events. L. monocytogenes can infect non-phagocytic cells by two means: direct invasion and cell-to-cell spread. The relative contribution of each method to placental infection is controversial, as is the anatomical site of invasion. Here, we report for the first time the use of first trimester placental organ cultures to quantitatively analyze L. monocytogenes infection of the human placenta. Contrary to previous reports, we found that the syncytiotrophoblast, which constitutes most of the placental surface and is bathed in maternal blood, was highly resistant to L. monocytogenes infection by either internalin-mediated invasion or cell-to-cell spread. Instead, extravillous cytotrophoblasts-which anchor the placenta in the decidua (uterine lining) and abundantly express E-cadherin-served as the primary portal of entry for L. monocytogenes from both extracellular and intracellular compartments. Subsequent bacterial dissemination to the villous stroma, where fetal capillaries are found, was hampered by further cellular and histological barriers. Our study suggests the placenta has evolved multiple mechanisms to resist pathogen infection, especially from maternal blood. These findings provide a novel explanation why almost all placental pathogens have intracellular life cycles: they may need maternal cells to reach the decidua and infect the placenta.
Project description:Although Zika virus (ZIKV) infection in pregnant women can cause placental damage, intrauterine growth restriction, microcephaly, and fetal demise, these disease manifestations only became apparent in the context of a large epidemic in the Americas. We hypothesized that ZIKV is not unique among arboviruses in its ability to cause congenital infection. To evaluate this, we tested the capacity of four emerging arboviruses [West Nile virus (WNV), Powassan virus (POWV), chikungunya virus (CHIKV), and Mayaro virus (MAYV)] from related (flavivirus) and unrelated (alphavirus) genera to infect the placenta and fetus in immunocompetent, wild-type mice. Although all four viruses caused placental infection, only infection with the neurotropic flaviviruses (WNV and POWV) resulted in fetal demise. WNV and POWV also replicated efficiently in second-trimester human maternal (decidua) and fetal (chorionic villi and fetal membrane) explants, whereas CHIKV and MAYV replicated less efficiently. In mice, RNA in situ hybridization and histopathological analysis revealed that WNV infected the placenta and fetal central nervous system, causing injury to the developing brain. In comparison, CHIKV and MAYV did not cause substantive placental or fetal damage despite evidence of vertical transmission. On the basis of the susceptibility of human maternal and fetal tissue explants and pathogenesis experiments in immunocompetent mice, other emerging neurotropic flaviviruses may share with ZIKV the capacity for transplacental transmission, as well as subsequent infection and injury to the developing fetus.
Project description:Mammalian pregnancy requires protection against immunological rejection of the developing fetus bearing discordant paternal antigens. Immune evasion in this developmental context entails silenced expression of chemoattractant proteins (chemokines), thereby preventing harmful immune cells from penetrating the maternal-fetal interface. Here, we demonstrate that fetal wastage triggered by prenatal Listeria monocytogenes infection is driven by placental recruitment of CXCL9-producing inflammatory neutrophils and macrophages that promote infiltration of fetal-specific T cells into the decidua. Maternal CD8+ T cells with fetal specificity upregulated expression of the chemokine receptor CXCR3 and, together with neutrophils and macrophages, were essential for L. monocytogenes-induced fetal resorption. Conversely, decidual accumulation of maternal T cells with fetal specificity and fetal wastage were extinguished by CXCR3 blockade or in CXCR3-deficient mice. Remarkably, protection against fetal wastage and in utero L. monocytogenes invasion was maintained even when CXCR3 neutralization was initiated after infection, and this protective effect extended to fetal resorption triggered by partial ablation of immune-suppressive maternal Tregs, which expand during pregnancy to sustain fetal tolerance. Together, our results indicate that functionally overriding chemokine silencing at the maternal-fetal interface promotes the pathogenesis of prenatal infection and suggest that therapeutically reinforcing this pathway represents a universal approach for mitigating immune-mediated pregnancy complications.
Project description:Genes involved in invasion of trophoblast cells and angiogenesis are crucial in determining pregnancy outcome. We therefore studied expression profiles of these genes in both fetal and maternal tissues to enhance our understanding of feto-maternal dialogue. We investigated the expression of genes involved in trophoblast invasion, namely Kiss1, Kiss1 Receptor (Kiss1R) and MMP9 as well as the expression of angiogenic ligands Vascular Endothelial Growth Factor-A (VEGF-A) and Prokineticin-1 (PROK1) and their respective receptors (VEGFR1, VEGFR2 and PROK1R) across the feto-maternal interface of healthy human pregnancies. The placenta, placental bed and decidua parietalis were sampled at elective caesarean delivery. Real-time RT-PCR was used to investigate transcription, while immunohistochemistry and western blot analyses were utilized to study protein expression. We found that the expression of Kiss1 (p<0.001), Kiss1R (p<0.05) and MMP9 (p<0.01) were higher in the placenta compared to the placental bed and decidua parietalis. In contrast, the expression of VEGF-A was highest in the placental bed (p<0.001). While VEGFR1 expression was highest in the placenta (p<0.01), the expression of VEGFR2 was highest in the placental bed (p<0.001). Lastly, both PROK1 (p<0.001) and its receptor PROK1R (p<0.001) had highest expression in the placenta. Genes associated with trophoblast invasion were highly expressed in the placenta which could suggest that the influence on invasion capacity may largely be exercised at the fetal level. Furthermore, our findings on angiogenic gene expression profiles suggest that angiogenesis may be regulated by two distinct pathways with the PROK1/PROK1R system specifically mediating angiogenesis in the fetus and VEGFA/VEGFR2 ligand-receptor pair predominantly mediating maternal angiogenesis.
Project description:Listeria monocytogenes is a facultative intracellular bacterial pathogen that can infect the placenta, a chimeric organ made of maternal and fetal cells. Extravillous trophoblasts (EVT) are specialized fetal cells that invade the uterine implantation site, where they come into direct contact with maternal cells. We have shown previously that EVT are the preferred site of initial placental infection. In this report, we infected primary human EVT with L. monocytogenes. EVT eliminated ?80% of intracellular bacteria over 24-hours. Bacteria were unable to escape into the cytoplasm and remained confined to vacuolar compartments that became acidified and co-localized with LAMP1, consistent with bacterial degradation in lysosomes. In human placental organ cultures bacterial vacuolar escape rates differed between specific trophoblast subpopulations. The most invasive EVT-those that would be in direct contact with maternal cells in vivo-had lower escape rates than trophoblasts that were surrounded by fetal cells and tissues. Our results suggest that EVT present a bottleneck in the spread of L. monocytogenes from mother to fetus by inhibiting vacuolar escape, and thus intracellular bacterial growth. However, if L. monocytogenes is able to spread beyond EVT it can find a more hospitable environment. Our results elucidate a novel aspect of the maternal-fetal barrier.
Project description:Feto-placental infections represent a major cause of pregnancy complications, and yet the underlying molecular and cellular mechanisms of vertical transmission are poorly understood. Listeria monocytogenes, a facultative intracellular pathogen, is one of a group of pathogens that are known to cause feto-placental infections in humans and other mammals. The purpose of this study was to evaluate possible mechanisms of vertical transmission of L. monocytogenes. Humans and guinea pigs have a hemochorial placenta, where a single layer of fetally derived trophoblasts separates maternal from fetal circulation. We characterized L. monocytogenes infection of the feto-placental unit in a pregnant guinea pig model and in primary human trophoblasts and trophoblast-derived cell lines. The clinical manifestations of listeriosis in the pregnant guinea pigs and the tropism of L. monocytogenes to the guinea pig placenta resembled those in humans. Trophoblast cell culture systems were permissive for listerial growth and cell-to-cell spread and revealed that L. monocytogenes deficient in internalin A, a virulence factor that mediates invasion of nonphagocytic cells, was 100-fold defective in invasion. However, crossing of the feto-placental barrier in the guinea pig model was independent of internalin A, suggesting a negligible role for internalin-mediated direct invasion of trophoblasts in vivo. Further understanding of vertical transmission of L. monocytogenes will help in designing more effective means of treatment and disease prevention.
Project description:To explore the interactions between the range of maternal and fetal placental cell types present, we profiled the transcriptomes of more than 50,000 single cells from matched first trimester samples of maternal blood and decidua, as well as fetal cells from the placenta itself. This part contains RNA-seq of Decidua cells using Smartseq 2
Project description:Structural and physiological changes in the female reproductive system underlie the origins of pregnancy in multiple vertebrate lineages. In mammals, the glandular portion of the lower reproductive tract has transformed into a structure specialized for supporting fetal development. These specializations range from relatively simple maternal nutrient provisioning in egg-laying monotremes to an elaborate suite of traits that support intimate maternal-fetal interactions in Eutherians. Among these traits are the maternal decidua and fetal component of the placenta, but there is considerable uncertainty about how these structures evolved. Previously, we showed that changes in uterine gene expression contributes to several evolutionary innovations during the origins of pregnancy (Mika et al., 2021b). Here, we reconstruct the evolution of entire transcriptomes ('ancestral transcriptome reconstruction') and show that maternal gene expression profiles are correlated with degree of placental invasion. These results indicate that an epitheliochorial-like placenta evolved early in the mammalian stem-lineage and that the ancestor of Eutherians had a hemochorial placenta, and suggest maternal control of placental invasiveness. These data resolve major transitions in the evolution of pregnancy and indicate that ancestral transcriptome reconstruction can be used to study the function of ancestral cell, tissue, and organ systems.
Project description:The bacterial pathogen <i>Listeria monocytogenes</i> causes foodborne systemic disease in pregnant women, which can lead to preterm labor, stillbirth, or severe neonatal disease. Colonization of the maternal decidua appears to be an initial step in the maternal component of the disease as well as bacterial transmission to the placenta and fetus. Host-pathogen interactions in the decidua during this early stage of infection remain poorly understood. Here, we assessed the dynamics of <i>L. monocytogenes</i> infection in primary human decidual organ cultures and in the murine decidua <i>in vivo</i> A high inoculum was necessary to infect both human and mouse deciduas, and the data support the existence of a barrier to initial colonization of the murine decidua. If successful, however, colonization in both species was followed by significant bacterial expansion associated with an inability of the decidua to mount appropriate innate cellular immune responses. The innate immune deficits included the failure of bacterial foci to attract macrophages and NK cells, cell types known to be important for early defenses against <i>L. monocytogenes</i> in the spleen, as well as a decrease in the tissue density of inflammatory Ly6C<sup>hi</sup> monocytes <i>in vivo</i> These results suggest that the infectivity of the decidua is not the result of an enhanced recruitment of <i>L. monocytogenes</i> to the gestational uterus but rather is due to compromised local innate cellular immune responses.
Project description:Intrauterine infection is a major detriment for maternal-child health and occurs despite local mechanisms that protect the maternal-fetal interface from a wide variety of pathogens. The bacterial pathogen Listeria monocytogenes causes spontaneous abortion, stillbirth, and preterm labor in humans and serves as a model for placental pathogenesis. Given the unique immunological environment of the maternal-fetal interface, we hypothesized that virulence determinants with placental tropism are required for infection of this tissue. We performed a genomic screen in pregnant guinea pigs that led to the identification of 201 listerial genes important for infection of the placenta but not maternal liver. Among these genes was lmrg1778 (lmo2470), here named inlP, predicted to encode a secreted protein that belongs to the internalin family. InlP is conserved in virulent L. monocytogenes strains but absent in Listeria species that are nonpathogenic for humans. The intracellular life cycle of L. monocytogenes deficient in inlP (?inlP) was not impaired. In guinea pigs and mice, InlP increased the placental bacterial burden by a factor of 3 log<sub>10</sub> while having only a minor role in other maternal organs. Furthermore, the ?inlP strain was attenuated in intracellular growth in primary human placental organ cultures and trophoblasts. InlP is a novel virulence factor for listeriosis with a strong tropism for the placenta. This virulence factor represents a tool for the development of new modalities to prevent and treat infection-related pregnancy complications.
Project description:Zika virus (ZIKV) infection during pregnancy leads to an increased risk of fetal growth restriction and fetal central nervous system malformations, which are outcomes broadly referred to as the Congenital Zika Syndrome (CZS). Here we infect pregnant rhesus macaques and investigate the impact of persistent ZIKV infection on uteroplacental pathology, blood flow, and fetal growth and development. Despite seemingly normal fetal growth and persistent fetal-placenta-maternal infection, advanced non-invasive in vivo imaging studies reveal dramatic effects on placental oxygen reserve accompanied by significantly decreased oxygen permeability of the placental villi. The observation of abnormal oxygen transport within the placenta appears to be a consequence of uterine vasculitis and placental villous damage in ZIKV cases. In addition, we demonstrate a robust maternal-placental-fetal inflammatory response following ZIKV infection. This animal model reveals a potential relationship between ZIKV infection and uteroplacental pathology that appears to affect oxygen delivery to the fetus during development.