Elisabeth Menu

Dynamic Shift from CD85j/ILT-2 to NKG2D NK Receptor Expression Pattern on Human Decidual NK during the First Trimester of Pregnancy

During the first trimester of human pregnancy, Natural Killer (NK) cells of the maternal uterine mucosa (e.g. decidua) have a unique phenotype and are involved in crucial physiological processes during pregnancy. We investigated whether modifications of the NK receptor repertoire occur during the first trimester of pregnancy. We found significantly decreased expression of KIR2DL1/S1 and KIR2DL2/L3/S2 receptors, NKp30 and NKp44 activatory receptors, and the CD85j (ILT-2) inhibitory receptor. We also observed significantly increased expression of the NKG2D activatory receptor at the decidual NK cell surface. By flow cytometry, we further highlighted an evolution of NK subsets between 8 and 12 weeks of gestation, with a shift from the KIR2DL1/S1+/KIR2DL2/L3/S2+ subset towards the double negative subset, coupled with a decrease of the CD85j+/NKG2D− subset in favour of the CD85j−/NKG2D+ subset. Furthermore, cell surface expression of NK receptor ligands, including CD85j and NKG2D ligands, has been characterized by flow cytometry on decidual immune CD14+ and CD3+ cells. HLA-G, the high affinity ligand of CD85j, was detected on both cell types. In contrast, NKG2D ligands ULBP-2 ULBP-3 and MICA/B were not expressed on CD14+ and CD3+ cells, however a variable expression of ULBP-1 was observed. The ligand expression of KIR2DL1/S1 and KIR2DL2/L3/S2 was also analyzed: the HLA-C molecule was expressed at a low level on some CD14+ cells whereas it was not detected on CD3+ cell surface. NK receptor ligands are known to be also expressed on the invading placental trophoblast cells. Thus, the phenotypic evolutions of decidual NK cells described in this present study may preserve their activation/inhibition balance during the first trimester of pregnancy.

Characterization of the main placental cytokine profiles from HIV-1-infected pregnant women treated with anti-retroviral drugs in France

Cytokines are involved in regulating HIV‐1 infection. They are also placental environment major components. We assessed the potential impact of HIV‐1 infection and/or anti‐retroviral drugs on the placental cytokine profiles that may be involved in controlling HIV‐1 placental dissemination. Placental explants were obtained after elective caesarean section from anti‐retroviral‐treated HIV‐1‐infected pregnant women and from HIV‐1 non‐infected pregnant women. The main placental cytokines were assessed for protein secretion in the supernatants of 24‐h placental culture explants and/or in uncultured placental explants for mRNA expression levels. The cytokine profiles were different between the HIV‐1‐infected and the non‐infected groups. Higher medians of leukaemia inhibiting factor (LIF), tumour necrosis factor (TNF)‐α and interleukin (IL)‐8 secretion were found in the 24‐h culture supernatant of term placenta from HIV‐1‐infected women. High median levels of IL‐16 and regulated upon activation normal T cell expressed and secreted (RANTES) levels were found in both groups. The mRNA expression medians were lower for TNF‐α and IL‐8 and higher for stromal cell‐derived factor‐1 (SDF‐1) in uncultured placental explants from HIV‐1‐infected women. In the HIV‐1‐infected group, but not in the non‐infected group, the secretion levels of TNF‐α and IL‐8, as well as their mRNA expression levels, were highly positively correlated; furthermore, their secretion levels were correlated positively with LIF and IL‐10 secretion levels. We found no correlation between the cytokine levels and the immunovirological status of the HIV‐1‐infected mothers or the type or duration of treatment. These results highlight the potential impact of HIV‐1 and of the anti‐retroviral treatments on the placental cytokines pattern, independently of their anti‐viral activity.

Decidual soluble factors participate in the control of HIV-1 infection at the maternofetal interface

BackgroundMaternofetal transmission (MFT) of HIV-1 is relatively rare during the first trimester of pregnancy despite the permissivity of placental cells for cell-to-cell HIV-1 infection. Invasive placental cells interact directly with decidual cells of the uterine mucosa during the first months of pregnancy, but the role of the decidua in the control of HIV-1 transmission is unknown.ResultsWe found that decidual mononuclear cells naturally produce low levels of IL-10, IL-12, IL-15, TNF-α, IFN-α, IFN-γ and CXCL-12 (SDF-1), and large amounts of CCL-2 (MCP1), CCL-3 (MIP-1α), CCL-4 (MIP-1β), CCL-5 (Rantes), CXCL-10 (IP-10), IL-6 and IL-8. CCL-3 and CCL-4 levels were significantly upregulated by in vitro infection with R5 HIV-1 but not X4. Decidual CD14+ antigen presenting cells were the main CCL-3 and CCL-4 producers among decidual leukocytes. R5 and X4 HIV-1 infection was inhibited by decidual cell culture supernatants in vitro. Using HIV-1 pseudotypes, we found that inhibition of the HIV-1 entry step was inhibited by decidual soluble factors.ConclusionOur findings show that decidual innate immunity (soluble factors) is involved in the control of HIV-1 infection at the maternofetal interface. The decidua could thus serve as a mucosal model for identifying correlates of protection against HIV-1 infection.

Antigen-Presenting Cells Represent Targets for R5 HIV-1 Infection in the First Trimester Pregnancy Uterine Mucosa

Background During the first trimester of pregnancy, HIV-1 mother-to-child transmission is relatively rare despite the permissivity of placental cells to cell-to-cell HIV-1 infection. The placenta interacts directly with maternal uterine cells (decidual cells) but the physiological role of the decidua in the control of HIV-1 transmission and whether decidua could be a source of infected cells is unknown. Methodology/Principal Findings To answer to this question, decidual mononuclear cells were exposed to HIV-1 in vitro. Decidual cells were shown to be more susceptible to infection by an R5 HIV-1, as compared to an X4 HIV-1. Infected cells were identified by flow cytometry analysis. The results showed that CD14+ cells were the main targets of HIV-1 infection in the decidua. These infected CD14+ cells expressed DC-SIGN, CD11b, CD11c, the Fc gamma receptor CD16, CD32 and CD64, classical MHC class-I and class-II and maturation and activation molecules CD83, CD80 and CD86. The permissivity of decidual tissue was also evaluated by histoculture. Decidual tissue was not infected by X4 HIV-1 but was permissive to R5 HIV-1. Different profiles of infection were observed depending on tissue localization. Conclusions/Significance The presence of HIV-1 target cells in the decidua in vitro and the low rate of in utero mother-to-child transmission during the first trimester of pregnancy suggest that a natural control occurs in vivo limiting cell-to-cell infection of the placenta and consequently infection of the fetus.

Tumour necrosis factor-α stimulates HIV-1 replication in single-cycle infection of human term placental villi fragments in a time, viral dose and envelope dependent manner

BackgroundThe placenta plays an important role in the control of in utero HIV-1 mother-to-child transmission (MTCT). Proinflammatory cytokines in the placental environment are particularly implicated in this control. We thus investigated the effect of TNF-α on HIV-1 expression in human placental tissues in vitro.ResultsHuman placental chorionic villi fragments were infected with varying doses of luciferase reporter HIV-1 pseudotypes with the R5, X4-Env or the vesicular stomatitis virus protein G (VSV-G). Histocultures were then performed in the presence or absence of recombinant human TNF-α. Luciferase activity was measured at different time points in cell lysates or on whole fragments using ex vivo imaging systems.A significant increase in viral expression was detected in placental fragments infected with 0.2 ng of p24 antigen/fragment (P = 0.002) of VSV-G pseudotyped HIV-1 in the presence of TNF-α seen after 120 hours of culture. A time independent significant increase of viral expression by TNF-α was observed with higher doses of VSV-G pseudotyped HIV-1. When placental fragments were infected with R5-Env pseudotyped HIV-1, a low level of HIV expression at 168 hours of culture was detected for 3 of the 5 placentas tested, with no statistically significant enhancement by TNF-α. Infection with X4-Env pseudotyped HIV-1 did not lead to any detectable luciferase activity at any time point in the absence or in the presence of TNF-α.ConclusionTNF-α in the placental environment increases HIV-1 expression and could facilitate MTCT of HIV-1, particularly in an inflammatory context.

In Vitro and In Vivo Human Herpesvirus 8 Infection of Placenta

Herpesvirus infection of placenta may be harmful in pregnancy leading to disorders in fetal growth, premature delivery, miscarriage, or major congenital abnormalities. Although a correlation between human herpesvirus 8 (HHV-8) infection and abortion or low birth weight in children has been suggested, and rare cases of in utero or perinatal HHV-8 transmission have been documented, no direct evidence of HHV-8 infection of placenta has yet been reported. The aim of this study was to evaluate the in vitro and in vivo susceptibility of placental cells to HHV-8 infection. Short-term infection assays were performed on placental chorionic villi isolated from term placentae. Qualitative and quantitative HHV-8 detection were performed by PCR and real-time PCR, and HHV-8 proteins were analyzed by immunohistochemistry. Term placenta samples from HHV-8-seropositive women were analyzed for the presence of HHV-8 DNA and antigens. In vitro infected histocultures showed increasing amounts of HHV-8 DNA in tissues and supernatants; cyto- and syncitiotrophoblasts, as well as endothelial cells, expressed latent and lytic viral antigens. Increased apoptotic phenomena were visualized by the terminal deoxynucleotidyl transferase-mediated deoxyuridine nick end-labeling method in infected histocultures. Ex vivo, HHV-8 DNA and a latent viral antigen were detected in placenta samples from HHV-8-seropositive women. These findings demonstrate that HHV-8, like other human herpesviruses, may infect placental cells in vitro and in vivo, thus providing evidence that this phenomenon might influence vertical transmission and pregnancy outcome in HHV-8-infected women.

Human decidual macrophages and NK cells differentially express Toll-like receptors and display distinct cytokine profiles upon TLR stimulation.

Maternofetal pathogen transmission is partially controlled at the level of the maternal uterine mucosa at the fetal implantation site (the decidua basalis), where maternal and fetal cells are in close contact. Toll-like receptors (TLRs) may play an important role in initiating rapid immune responses against pathogens in the decidua basalis, however the tolerant microenvironment should be preserved in order to allow fetal development. Here we investigated the expression and functionality of TLRs expressed by decidual macrophages (dMs) and NK cells (dNKs), the major decidual immune cell populations. We report for the first time that both human dMs and dNK cells express mRNAs encoding TLRs 1-9, albeit with a higher expression level in dMs. TLR2, TLR3, and TLR4 protein expression checked by flow cytometry was positive for both dMs and dNK cells. In vitro treatment of primary dMs and dNK cells with specific TLR2, TLR3, TLR4, TLR7/8, and TLR9 agonists enhanced their secretion of pro- and anti-inflammatory cytokines, as well as cytokines and chemokines involved in immune cell crosstalk. Only dNK cells released IFN-γ, whereas only dMs released IL-1β, IL-10, and IL-12. TLR9 activation of dMs resulted in a distinct pattern of cytokine expression compared to the other TLRs. The cytokine profiles expressed by dMs and dNK cells upon TLR activation are compatible with maintenance of the fetotolerant immune environment during initiation of immune responses to pathogens at the maternofetal interface.

Plasmodium falciparum Infection Significantly Impairs Placental Cytokine Profile in HIV Infected Cameroonian Women

Background Placental cytokines play crucial roles in the establishment and maintenance of pregnancy as well as protecting the foetus from infections. Previous studies have suggested the implication of infections such as P. falciparum and HIV in the stimulation of placental cytokines. This study assessed the impact of P. falciparum on placental cytokine profiles between HIV-1 positive and negative women. Materials and Methods P. falciparum infection was checked in peripheral and placental blood of HIV-1 negative and positive women by the thick blood smear test. Cytokines proteins and messenger RNAs were quantified by ELISA and real time PCR, respectively. Non-parametric tests were used for statistical analyses. Results Placental and peripheral P. falciparum infections were not significantly associated with HIV-1 infection (OR: 1.4; 95% confidence interval (95%CI): 0.5–4.2; pu200a=u200a0.50 and OR: 0.6; 95%CI: 0.3–1.4; pu200a=u200a0.26, respectively). Conversely, placental P. falciparum parasitemia was significantly higher in the HIV-1 positive group (pu200a=u200a0.04). We observed an increase of TNF-α mRNA median levels (pu200a=u200a0.02) and a trend towards a decrease of IL-10 mRNA (pu200a=u200a0.07) in placenta from HIV-1 positive women compared to the HIV negative ones leading to a median TNF-α/IL-10 mRNA ratio significantly higher among HIV-1 positive than among HIV-1 negative placenta (pu200a=u200a0.004; 1.5 and 0.8, respectively). Significant decrease in median secreted cytokine levels were observed in placenta from HIV-1 positive women as compared to the HIV negative however these results are somewhat indicative since it appears that differences in cytokine levels (protein or mRNA) between HIV-1 positive and negative women depend greatly on P.falciparum infection. Within the HIV-1 positive group, TNF-α was the only cytokine significantly associated with clinical parameters linked with HIV-1 MTCT such as premature rupture of membranes, CD4 T-cell number, plasma viral load and delay of NVP intake before delivery. Conclusions These results show that P. falciparum infection profoundly modifies the placenta cytokine environment and acts as a confounding factor, masking the impact of HIV-1 in co-infected women. This interplay between the two infections might have implications in the in utero MTCT of HIV-1 in areas where HIV-1 and P. falciparum co-circulate.

Distinct Characteristics of Endometrial and Decidual Macrophages and Regulation of Their Permissivity to HIV-1 Infection by SAMHD1

ABSTRACT In order to develop strategies to prevent HIV-1 (human immunodeficiency virus type 1) transmission, it is crucial to better characterize HIV-1 target cells in the female reproductive tract (FRT) mucosae and to identify effective innate responses. Control of HIV-1 infection in the decidua (the uterine mucosa during pregnancy) can serve as a model to study natural mucosal protection. Macrophages are the main HIV-1 target cells in the decidua. Here we report that in vitro, macrophages and T cells are the main HIV-1 targets in the endometrium in nonpregnant women. As reported for decidual macrophages (dM), endometrial macrophages (eM) were found to have an M2-like phenotype (CD68+ CD163+ CD206+ IL-10high). However, eM and dM may belong to different subpopulations, as they differently express certain markers and secrete different amounts of proinflammatory and anti-inflammatory cytokines. We observed strong expression of the SAMHD1 restriction factor and weak expression of its inactive form (pSAMHD1, phosphorylated at residue Thr592) in both eM and dM. Infection of macrophages from both tissues was enhanced in the presence of the viral protein Vpx, suggesting a role for SAMHD1 in the restriction of HIV-1 infection. This study and further comparisons of the decidua with FRT mucosae in nonpregnant women should help to identify mechanisms of mucosal protection against HIV-1 infection. IMPORTANCE The female reproductive tract mucosae are major portals of HIV-1 entry into the body. The decidua (uterine mucosa during pregnancy) can serve as a model for studying natural mucosal protection against HIV-1 transmission. A comparison of target cells and innate responses in the decidua versus the endometrium in nonpregnant women could help to identify protective mechanisms. Here, we report for the first time that macrophages are one of the main HIV-1 target cells in the endometrium and that infection of macrophages from both the endometrium and the decidua is restricted by SAMHD1. These findings might have implications for the development of vaccines to prevent HIV-1 mucosal transmission.

Distinct efficacy of HIV-1 entry inhibitors to prevent cell-to-cell transfer of R5 and X4 viruses across a human placental trophoblast barrier in a reconstitution model in vitro

Background and methodsHIV-1 cell-to-cell transmission is more efficient than infection of permissive cells with cell-free particles. The potency of HIV-1 entry inhibitors to inhibit such transmission is not well known. Herein, we evaluated the efficacy of this new class of antiretrovirals to block cell-to-cell transmission of HIV-1 in a model of reconstitution of the human placental trophoblast barrier in vitro.ResultsOur data show that CCR5 antagonists and T20 inhibit the passage of the virus across the BeWo cell monolayer in contact with PBMCs infected with an R5 (Ba-L) and a dualtropic (A204) HIV-1 with IC50s in the range of 100 – 5,000 nM for TAK779; 90 to 15,000 nM for SCH-350581 and 3,000 to 20,000 nM for T20. The CXCR4 antagonist AMD3100 is also effective against X4 HIV-1 infected PBMCs in our model with IC50 comprised between 4 nM and 640 nM. HIV-1 entry inhibitors are less efficient to block cell-to-cell virus transmission than cell-free HIV-1 infection of PBMCs and CCR5 antagonists do not prevent PBMC infection by dual tropic HIV-1 in contrast to cell-to-cell infection in our model.Surprisingly, T20 (and C34) do not block cell-to-cell transmission of X4 HIV-1 but, rather, increase 80 to 140 fold, compared to control without drug, the passage of the virus across the trophoblast barrier. Additional experiments suggest that the effect of T20 on BeWo/PBMC-X4 HIV-1 is due to an increase of effector-target cells fusion.ConclusionOur results support further evaluation of HIV-1 coreceptor antagonists, alone or combined to other antiretrovirals, in a perspective of prevention but warn on the use of T20 in patients bearing X4 HIV-1 at risk of transmission.