First-trimester Human Decidua Research Articles

Tripeptidyl Peptidase 1 Regulates Human Trophoblast Cell Proliferation Implying a Role in Placentation.

Proper placentation in the first trimester is essential for a healthy pregnancy in humans. A recent proteomics study of human placental tissue has identified that tripeptidyl peptidase 1 (TPP1) production is reduced in the placenta in early-onset preeclampsia compared to uncomplicated pregnancy. However, it remains to be investigated if TPP1 plays a role in regulating trophoblast cell function during early pregnancy. In this study, immunohistochemistry was used to determine the production and localization of TPP1 in human placenta throughout gestation and the first-trimester decidua/implantation sites. TPP1 siRNA (20 nM) was transfected into a human trophoblast cell line (HTR8/SVneo) to knock down TPP1, and functional consequences on cell adhesion, proliferation, migration, and invasion were analyzed via xCELLigence real-time monitoring. The expression of TPP1 downstream targets was examined by qPCR. Our data show that TPP1 localized to the discrete foci in the cytoplasm in syncytiotrophoblast, cytotrophoblast, and decidual cells across all trimesters of pregnancy. In the first-trimester human decidua, TPP1 exhibited similar staining patterns in the cytotrophoblast cells based at the cell columns. However, minimal/no staining was identified in the HLA-G positive extravillous trophoblast cells (EVTs), especially in the EVTs that invaded in the decidua. Knockdown of TPP1 in HTR8/SVneo cells by 95% significantly impaired cell adhesion and proliferation without affecting cell migration and invasion. qPCR revealed that the expression of cell proliferation markers P21 and MKI67 and TPP1-related genes MRE11, CLN3, and CLN8 was significantly changed after TPP1 knockdown in HTR8/SVneo cells compared to control. Overall, our data demonstrate that TPP1 alters trophoblast cell line function suggesting that it may be involved in regulating human placentation in the first trimester via controlling trophoblast cell adhesion and proliferation.

Profiling the expression and function of oestrogen receptor isoform ER46 in human endometrial tissues and uterine natural killer cells.

STUDY QUESTIONDoes the oestrogen receptor isoform, ER46, contribute to regulation of endometrial function?SUMMARY ANSWERER46 is expressed in endometrial tissues, is the predominant ER isoform in first trimester decidua and is localised to the cell membrane of uterine natural killer (uNK) cells where activation of ER46 increases cell motility.WHAT IS KNOWN ALREADYOestrogens acting via their cognate receptors are essential regulators of endometrial function and play key roles in establishment of pregnancy. ER46 is a 46-kDa truncated isoform of full length ERα (ER66, encoded by ESR1) that contains both ligand- and DNA-binding domains. Expression of ER46 in the human endometrium has not been investigated previously. ER46 is located at the cell membrane of peripheral blood leukocytes and mediates rapid responses to oestrogens. uNK cells are a phenotypically distinct (CD56brightCD16−) population of tissue-resident immune cells that regulate vascular remodelling within the endometrium and decidua. We have shown that oestrogens stimulate rapid increases in uNK cell motility. Previous characterisation of uNK cells suggests they are ER66-negative, but expression of ER46 has not been characterised. We hypothesise that uNK cells express ER46 and that rapid responses to oestrogens are mediated via this receptor.STUDY DESIGN, SIZE, DURATIONThis laboratory-based study used primary human endometrial (n = 24) and decidual tissue biopsies (n = 30) as well as uNK cells which were freshly isolated from first trimester human decidua (n = 18).PARTICIPANTS/MATERIALS, SETTING, METHODSPrimary human endometrial and first trimester decidual tissue biopsies were collected using methods approved by the local institutional ethics committee (LREC/05/51104/12 and LREC/10/51402/59). The expression of ERs (ER66, ER46 and ERβ) was assessed by quantitative PCR, western blot and immunohistochemistry. uNK cells were isolated from first-trimester human decidua by magnetic bead sorting. Cell motility of uNK cells was measured by live cell imaging: cells were treated with 17β-oestradiol conjugated to bovine serum albumin (E2-BSA, 10 nM equivalent), the ERβ-selective agonist 2,3-bis(4-hydroxyphenyl)-propionitrile (DPN; 10 nM) or dimethylsulphoxide vehicle control.MAIN RESULTS AND THE ROLE OF CHANCEER46 was detected in proliferative and secretory phase tissues by western blot and was the predominant ER isoform in first-trimester decidua samples. Immunohistochemistry revealed that ER46 was co-localised with ER66 in cell nuclei during the proliferative phase but detected in both the cytoplasm and cell membrane of stromal cells in the secretory phase and in decidua. Triple immunofluorescence staining of decidua tissues identified expression of ER46 in the cell membrane of CD56-positive uNK cells which were otherwise ER66-negative. Profiling of isolated uNK cells confirmed expression of ER46 by quantitative PCR and western blot and localised ER46 protein to the cell membrane by immunocytochemistry. Functional analysis of isolated uNK cells using live cell imaging demonstrated that activation of ER46 with E2-BSA significantly increased uNK cell motility.LARGE SCALE DATAN/A.LIMITATIONS, REASONS FOR CAUTIONExpression pattern in endometrial tissue was only determined using samples from proliferative and secretory phases. Assessment of first trimester decidua samples was from a range of gestational ages, which may have precluded insights into gestation-specific changes in these tissues. Our results are based on in vitro responses of primary human cells and we cannot be certain that similar mechanisms occur in situ.WIDER IMPLICATIONS OF THE FINDINGSE2 is an essential regulator of reproductive competence. This study provides the first evidence for expression of ER46 in the human endometrium and decidua of early pregnancy. We describe a mechanism for regulating the function of human uNK cells via expression of ER46 and demonstrate that selective targeting with E2-BSA regulates uNK cell motility. These novel findings identify a role for ER46 in the human endometrium and provide unique insight into the importance of membrane-initiated signalling in modulating the impact of E2 on uNK cell function in women. Given the importance of uNK cells to regulating vascular remodelling in early pregnancy and the potential for selective targeting of ER46, this may be an attractive future therapeutic target in the treatment of reproductive disorders.STUDY FUNDING/COMPETING INTEREST(S)These studies were supported by Medical Research Council (MRC) Programme Grants G1100356/1 and MR/N024524/1 to PTKS. H.O.D.C. was supported by MRC grant G1002033. The authors declare no competing interests related to the published work.

Placental bed research: II. Functional and immunological investigations of the placental bed

Research on the placenta as the interface between the mother and the fetus has been undertaken for some 150 years, and in 2 subsequent reviews, we attempted to summarize the situation. In the first part, we described the discovery of unique physiological modifications of the uteroplacental spiral arteries, enabling them to cope with a major increase in blood flow necessary to ensure proper growth of the fetus. These consist of an invasion of the arterial walls by trophoblast and a progressive disappearance of its normal structure. Researchers then turned to the pathophysiology of the placental bed and in particular to its maternal vascular tree. This yielded vital information for a better understanding of the so-called great obstetrical syndromes (preeclampsia, fetal growth restriction, premature labor and delivery, placenta accreta). Systematic morphological investigations of the uteroplacental vasculature showed that preeclampsia is associated with decreased or failed transformation of spiral arteries and the persistence of endothelial and smooth muscle cells in segments of their myometrial portion. Here we report on recent functional investigations of the placental bed, including in situ biophysical studies of uteroplacental blood flow and vascular resistance, and manipulation of uteroplacental perfusion. These new methodologies have provided a novel way of identifying pregnancies in which remodeling is impaired. In animals it is now possible to manipulate uteroplacental blood flow, leading to an enhancement of fetal growth; this opens the way to trials in abnormal human pregnancies. In this second part, we explored a new, extremely important area of research that deals with the role of specific subsets of leukocytes and macrophages in the placental bed. The human first-trimester decidua is rich in leukocytes called uterine natural killer cells. Both macrophages and uterine natural killer cells increase in number from the secretory endometrium to early pregnancy and play a critical role in mediating the process of spiral artery transformation by inducing initial structural changes. It seems therefore that vascular remodeling of spiral arteries is initiated independently of trophoblast invasion. Dysregulation of the immune system may lead to reproductive failure or pregnancy complications, and in this respect, recent studies have advanced our understanding of the mechanisms regulating immunological tolerance during pregnancy, with several mechanisms being proposed for the development of tolerance to the semiallogeneic fetus. In particular, these include several strategies by which the trophoblast avoids maternal recognition. Finally, an important new dimension is being explored: the likelihood that pregnancy syndromes and impaired uteroplacental vascular remodeling may be linked to future maternal and even the child's cardiovascular disease risk. The functional evidence underlying these observations will be discussed.

Progesterone and cyclic adenosine monophosphate down-regulate CD90 in the stromal cells of human decidua. In vitro evidence and in situ findings.

CD90 is a glycoprotein involved in leukocyte relocation and cell differentiation. CD90 is expressed in endothelial and stromal cells in human endometrium; however, its role in the remodeling of the decidual tissue during pregnancy is poorly understood. Here, we investigate how CD90 expression in decidual stromal cells (DSCs) is regulated. The native CD90 receptor in stromal cells in decidua was investigated via histology. We further develop in vitro culture of DSCs which allows us to test the effects of hormones and paracrine signals on CD90 expression. Stromal cells in first-trimester human decidua display heterogeneous levels of CD90 expression. In vitro analyses reveal that progesterone, a factor normally secreted by trophoblast cells in the placenta, and extracellular cyclic adenosine monophosphate, a known downstream signaling messenger of progesterone, reduce CD90 expression in DSCs by ~30%. This reduction in CD90 expression correlates with a change toward a more highly differentiated cell state. DSCs in early pregnancy show different levels of CD90 expression, suggesting different DSC differentiation and selective interactions with cells during decidual morphogenesis.

IFN‐γ Inhibits Osteopontin Expression in Human Decidual Stromal Cells and can be Attenuated by 1α,25‐Dihydroxyvitamin D3

The effect of IFN-γ on the expression of osteopontin (OPN), in the presence or absence of 1α,25-dihydroxyvitamin D3 (1,25(OH)(2) D(3) ), in decidual stromal cells (DSC). Decidual stromal cells were isolated from women undergoing elective termination of pregnancy (gestational age, 6-9weeks). After characterization, they were treated with IFN-γ in the presence or absence of 1,25(OH)(2) D(3) . The uterus of pregnancy IFN-γ knockout mice were collected on gestation day (gd) 7.5, and the expression of OPN were examined. IFN-γ drastically decreased the expression of OPN in DSC, which was reverted by the addition of 1,25(OH)(2) D(3) to the IFN-γ-treated decidual cells. Moreover, the OPN expression in uterus of IFN-γ knockout mice was higher than that of wild-type counterparts. We demonstrated OPN was expressed in DSC in human first-trimester decidua and in the uterus in mice at 7.5gd. The OPN expression was closely correlated with regulation of IFN-γ and 1,25(OH)(2) D(3) in human early pregnancy. OPN expression in DSC was significantly decreased with the treatment of IFN-γ. 1,25(OH)(2) D(3) played an opposite role in IFN-γ-mediated inhibition of OPN expression in human DSC.

Altered expression of immune-associated genes in first-trimester human decidua of pregnancies later complicated with hypertension or foetal growth restriction

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.

CTGF expression is up-regulated by PROK1 in early pregnancy and influences HTR-8/Svneo cell adhesion and network formation

BACKGROUNDProkineticin-1 (PROK1) and connective tissue growth factor (CTGF) are expressed in human endometrium and first-trimester decidua and have individually been proposed to have roles in implantation and placentation. We have recently demonstrated that CTGF may be a target gene for PROK1 in gene array analysis of a prokineticin receptor-1 stably transfected Ishikawa endometrial epithelial cell line (PROKR1-Ishikawa). The first aim of the study was to determine the effect of PROK1 on CTGF expression in PROKR1-Ishikawa cells and first-trimester decidua samples. Secondly, the effect of CTGF on trophoblast-derived HTR-8/SVneo cell adhesion and network formation was investigated.METHODS AND RESULTSReal-time qPCR showed that CTGF expression is elevated in first-trimester decidua compared with non-pregnant endometrium. In decidua, CTGF co-localized with PROKR1 to the glandular epithelium and a subset of stromal cells. PROK1 increased CTGF mRNA and protein expression in PROKR1-Ishikawa cells and first-trimester human decidua (8–12 weeks gestation). Knock down of endogenous PROK1 using micro RNA constructs targeted at PROK1, resulted in decreased expression of CTGF mRNA and protein in decidua. Inhibitors of specific cell signalling molecules demonstrated that PROK1 regulates CTGF expression via the Gq, phospholipase C (PLC), cSrc, epidermal growth factor receptor (EGFR), mitogen-activated protein kinase/extracellular signal-regulated kinase (MAPK/ERK) kinase pathway activation. Treatment of trophoblast-derived HTR-8/Svneo cells with 1 µg/ml CTGF significantly increased adhesion to collagen IV, and differentiation of the cells into tube-like structures in matrigel.CONCLUSIONSCTGF expression in early pregnancy decidua is regulated by PROK1, via activation of the Gq, PLC, cSrc, EGFR, MAPK/ERK kinase pathway. CTGF in turn may contribute to the regulation of trophoblast conversion of maternal spiral arteries.

FOXP3+ Regulatory T Cells and T Helper 1, T Helper 2, and T Helper 17 Cells in Human Early Pregnancy Decidua1

In pregnancy, the decidua is infiltrated by leukocytes promoting fetal development without causing immunological rejection. Murine regulatory T (Treg) cells are known to be important immune regulators at this site. The aim of the study was to characterize the phenotype and origin of Treg cells and determine the quantitative relationship between Treg, T-helper type 1 (T(H)1), T(H)2, and T(H)17 cells in first-trimester human decidua. Blood and decidual CD4(+) T cells from 18 healthy first-trimester pregnant women were analyzed for expression of Treg-cell markers (CD25, FOXP3, CD127, CTLA4, and human leukocyte antigen-DR [HLA-DR]), chemokine receptors (CCR4, CCR6, and CXCR3), and the proliferation antigen MKI67 by six-color flow cytometry. Treg cells were significantly enriched in decidua and displayed a more homogenous suppressive phenotype with more frequent expression of FOXP3, HLA-DR, and CTLA4 than in blood. More decidual Treg cells expressed MKI67, possibly explaining their enrichment at the fetal-maternal interface. Using chemokine receptor expression profiles of CCR4, CCR6, and CXCR3 as markers for T(H)1, T(H)2, and T(H)17 cells, we showed that T(H)17 cells were nearly absent in decidua, whereas T(H)2-cell frequencies were similar in blood and decidua. CCR6(+) T(H)1 cells, reported to secrete high levels of interferon gamma (IFNG), were fewer, whereas the moderately IFNG-secreting CCR6(-) T(H)1 cells were more frequent in decidua compared with blood. Our results point toward local expansion of Treg cells and low occurrence of T(H)17 cells. Furthermore, local, moderate T(H)1 activity seems to be a part of normal early pregnancy, consistent with a mild inflammatory environment controlled by Treg cells.

First-trimester human decidua contains a population of mesenchymal stem cells

To determine whether first-trimester human decidua contains multipotent stromal cells capable of differentiating into other cell lines. In vitro-cultured decidual stromal cells were analyzed by flow cytometry and induced to differentiate into osteogenic and adipogenic lineages, endothelial cells, and PRL-secreting mature decidual cells. Research laboratory. Eight decidua samples were collected from healthy women aged 26-32 years undergoing elective vaginal surgical terminations of early pregnancy (8-10 gestational weeks). Cell suspensions from human decidual stromal cells were cultured at clonogenic concentrations and in bulk under differentiation conditions and analyzed for specific markers. Multipotent differentiation potential of decidual stromal cells. Decidual stromal cells express the surface markers specific to cells of mesenchymal origin as analyzed by flow cytometry. A pool of the decidual stromal cells can be induced to differentiate into mature PRL-secreting decidual cells and into osteogenic, adipogenic, and endothelial cells expressing the corresponding specific markers. It is demonstrated for the first time that first-trimester human decidua contains multipotent mesenchymal stem cells that can be grown in vitro for prolonged periods, have clonogenic properties, can differentiate into different cell lineages, and express surface markers specific to mesenchymal stem cells.

Angiogenic Growth Factors and Their Receptors in First-Trimester Human Decidua of Pregnancies Further Complicated By Preeclampsia or Fetal Growth Restriction

Disturbances in decidual and placental vascular development may play a role in the pathogenesis of pregnancy complications. This study focused on the role of angiogenic factors in the first trimester in the pathogenesis of preeclampsia (PE) and/or fetal growth restriction (FGR). First-trimester decidua was obtained during chorionic villous sampling.The expression of the angiogenic factors was determined by reverse transcriptase polymerase chain reaction and related to the pregnancy outcome. First-trimester decidua expressed all angiogenic factors.The differential expression of angiogenic factors appeared to be more prominent in FGR than in PE. These first-trimester samples provided a unique opportunity to obtain information regarding the onset of PE and FGR. First-trimester changes in angiogenic factor expression may well occur as a compensatory mechanism. This, in turn, may unintentionally set the stage for increased angiogenesis and altered decidual/placental vascular adaptation, which may be part of the pathogenesis of PE and/or FGR.

BDCA-1+, BDCA-2+ and BDCA-3+ dendritic cells in early human pregnancy decidua

Dendritic cells (DCs) can acquire unique features or phenotypes in different tissue microenvironments and decide whether immunity or tolerance develops. DCs observed within the decidua have been implicated in pregnancy maintenance. However, the precise distribution of decidual DC subsets and their phenotypic characteristics are largely unknown. Using flow cytometry, we identified three DC subsets in normal human first-trimester decidua: BDCA-1+ CD19- CD14(-) myeloid DC type 1 (MDC1), BDCA-3+ CD14- myeloid DC type 2 (MDC2) and BDCA-2+ CD123+ plasmacytoid DC (PDC). The percentage of MDC1 to mononuclear cells in the decidua was similar to that in the peripheral blood controls. The percentage of MDC2 in the decidua was significantly higher than that in the peripheral blood controls, whereas the percentage of PDC was significantly lower. Both MDC1 and MDC2 subsets expressed human leucocyte antigen D-related, CD86 and CD80 at low levels, suggesting a characteristic of immature myeloid DCs. Immunoglobulin-like transcript 3, suggested to be involved in immune tolerance induction, was also expressed on decidual MDC1 and MDC2 subsets. In addition, as gestational age increased from 6 to 9 weeks, the numbers of MDC1 decreased but MDC2 increased significantly. This is the first study to demonstrate the presence of three previously unidentified BDCA-1+, BDCA-3+ and BDCA-2+ DC subsets in human decidua, these decidual DCs might play important role in the maintenance of pregnancy.

Leukocyte Populations and Steroid Receptor Expression in Human First-Trimester Decidua; Regulation by Antiprogestin and Prostaglandin E Analog

Progesterone acting via its cognate receptor is critical to maintaining a viable endometrial environment for implantation and pregnancy. During medical termination of pregnancy, the biological effect of progesterone is pharmacologically withdrawn and prostaglandins administered exogenously. Leukocytes within the uterus are the effector cells of an inflammatory response and play important roles in both tissue breakdown and remodeling. The aim of this study was to identify the separate and combined effects of the antiprogestin Mifepristone (single dose, 200 mg) and the prostaglandin E (PGE) analog (gemeprost) on leukocyte populations and steroid receptor expression in human first-trimester decidua. Eighty women were recruited from the termination of pregnancy service with a gestational age of between 35 and 65 d at the time of surgical termination of pregnancy. Immunohistochemistry was used to measure macrophage (CD68 +ve), neutrophil (neutrophil elastase +ve), and uterine natural killer cell (CD56 +ve) populations and progesterone (PR(A) and PR(B)), estrogen (ERalpha and ERbeta), and androgen receptor (AR) expression. After administration of both antiprogestin and the PGE analog, macrophage and neutrophil numbers were significantly increased, whereas natural killer cell numbers were unchanged. Antiprogestin and PGE analog coadministration also significantly decreased PR and ERalpha immunoreactivity but had no effect on androgen receptor or ERbeta receptor expression. PGE analog alone was also capable of reducing PR expression. In this study, we demonstrate that the inflammatory response induced by antiprogestin in combination with PGE analog is accompanied by both increases in macrophages and neutrophils numbers and decreases in PR and ERalpha expression in human first-trimester decidua.

Cytokine microenvironments in human first trimester decidua are dependent on trophoblast cells

Objective To compare cytokine expression profiles of decidua basalis (containing trophoblast cells) and decidua parietalis (without trophoblast cells) for determination of microenvironments in human first trimester decidua. Design Retrospective study. Setting School of Medicine, RWTH University of Aachen, Aachen Germany, and Bourgognekliniek Maastricht, Maastricht, The Netherlands. Patient(s) Forty-six women who had undergone elective first-trimester termination of viable pregnancy at 5 to 12 weeks. Main outcome measure(s) Quantitative cytokine protein analysis in decidual tissues by enzyme-linked immunosorbent assay, qualitative cytokine messenger (m)RNA analysis in isolated decidual cell samples, and comparative mRNA and protein analysis in tissues of decidua basalis compared with decidua parietalis. Result(s) Interleukin-2, interferon-γ (Th-1), interleukin-4 (Th-2), and interleukin-1β proteins are expressed in the human first-trimester decidua. Interleukin-2, interferon-γ, and interleukin-4 mRNA mainly derive from the decidual tissue leukocytes. Interleukin-1β mRNA is expressed by all decidual cell types. Interferon-γ mRNA and protein is detected predominantly in the decidua basalis, which contains trophoblast cells. Conclusion(s) Microenvironments are established topographically by different expression of cytokines in decidua basalis and decidua parietalis. These locally specific patterns are indicative of fetomaternal cross-talk. Higher interferon-γ concentrations in decidua basalis may influence leukocyte differentiation (e.g., macrophage activation) and trophoblast invasion (e.g., by induction of expression of major histocompatibility complex).

Human first-trimester decidua vascular density: an immunohistochemical study using VE-cadherin and endoglin as endothelial cell markers.

Angiogenesis and vasculogenesis appear to be of critical importance for the success of pregnancy. Recent data have emphasized that pregnancy complications, such as abortion or pre-eclampsia, are linked with vascular pathologies. The aim of this study was to quantify human first-trimester decidua microvascular density, using two novel, highly specific endothelial cell markers, VE-cadherin and endoglin. We collected decidua from women undergoing termination of normal pregnancies. VE-cadherin and endoglin were localized by immunohistochemistry. The blood vessel densities detected by VE-cadherin or endoglin-stainings were microscopically quantified per mm2. Endothelial cells in first-trimester human decidua both express VE-cadherin and endoglin. The microvascular density detected by VE-cadherin-staining varied from 32.2 +/- 1.7 in decidua basalis, to 30 +/- 0.6 in decidua parietalis. For the endoglin-staining, the values varied from 37.5 +/- 3 in decidua basalis, and 26.7 +/- 1.2 in decidua parietalis. Our data shows that both VE-cadherin and endoglin are good candidates to highlight the decidual endothelial cells, and to quantify the blood vessels density of endometrium.

A subset of CD56+ large granular lymphocytes in first-trimester human decidua are proliferating cells

Objective: To determine whether the unusually high number of CD56+ large granular lymphocytes (LGL) in the decidua of early human pregnancy arises from selective migration or in situ proliferation. Design: Controlled clinical study. Setting: Academic research environment. Patient(s): Thirty healthy women undergoing therapeutic abortion of an intact pregnancy at 5–11 weeks’ gestation. Intervention(s): Decidua was obtained by suction curettage; tissue and isolated cells were subjected to immunohistochemical and flow cytometric investigation. Main Outcome Measure(s): Proliferation rate of LGL. Result(s): The proportion of CD56+ cells positive for the proliferation-associated Ki-67 antigen was found to be 7%–23.5% by three different methods of investigation. These findings are consistent with those of flow cytometric analysis of the nuclear phase, which revealed 6%–22% of the LGL nuclei to be in the phases S+G 2+M. Conclusion(s): The various methods of investigation revealed marked proliferative activity in the LGL of early pregnancy decidua. This finding suggests that in situ proliferation may be responsible for the high density of these cells in the decidua.

Distribution of cell adhesion molecules on CD56++, CD3-, CD16- large granular lymphocytes and endothelial cells in first-trimester human decidua.

Human decidua exhibits a unique infiltrate of large granular lymphocytes (LGL) with a natural killer (NK) cell phenotype (CD56++, CD16-, CD3-). The mechanisms underlying the binding of circulating LGL to vascular endothelium in the decidua and their migration into the decidual stroma were investigated immunohistochemically in first-trimester decidua with antibodies against endothelial adhesion molecules and their counter-receptors on leukocytes. Decidual and peripheral blood LGL were also investigated by flow cytometry. In the immunohistochemical investigations, moderate to large numbers of lymphoid cells in the decidua were found to express the alpha 4 and alpha L integrin subunits, platelet endothelial cell adhesion molecule (PECAM) and intercellular adhesion molecule-1 (ICAM-1). PECAM and ICAM-1 were found on the endothelium of large numbers of decidual blood vessels of all types. Vascular cell adhesion molecule (VCAM), however, was found on the endothelium of only small to moderate numbers of arterioles and venules and a few capillaries, the latter being the main site of migration of leukocytes into the stroma. Weak staining for endothelial leukocyte adhesion molecule (ELAM) was seen only in a moderate number of blood vessels. Flow cytometry revealed expression of the alpha L integrin subunit by 72 +/- 10% and 97 +/- 3% of decidual and peripheral blood CD56+ LGL, respectively, of the alpha 4 integrin subunit by 85 +/- 7% and 90 +/- 5%, of PECAM by 40 +/- 12% and 30 +/- 15%, and of ICAM-1 by 22 +/- 10% and 1 +/- 1%.(ABSTRACT TRUNCATED AT 250 WORDS)

Production of Prolactin by Cultures of Isolated Cells From Human First-Trimester Decidua

An enriched fraction of first-trimester decidual cells that synthesize and release prolactin (PRL) was obtained by discontinuous Percoll gradient (20-50%) centrifugation of collagenase type I- and deoxyribonuclease I-dispersed cells (3 mg/mL and 50 micrograms/mL, respectively). Centrifugation of the cell suspension yielded three major bands aggregating at the density interfaces. The fraction of the 30-40% Percoll interface contained enlarged decidual cells and constantly secreted significant amounts of PRL into the medium for at least 10 days. The fraction of the 40-50% Percoll interface contained fibroblastic cells and secreted a small amount of PRL into the medium. Cells in the other fractions did not attach to the plastic dishes in 48 hours. Under the influence of progesterone (100 ng/mL), the cultured decidual cells retained their capability of PRL production for at least 10 days because no decline of the secretion rate was observed. The culture system established by the present study is satisfactory for investigating decidual cell functions, including the regulatory mechanisms of PRL production.

Immunolocalization of extracellular matrix proteins and collagen synthesis in first‐trimester human decidua

The extracellular matrix (ECM) of first-trimester human decidua was examined with indirect immunofluorescence using affinity-purified antibodies to human collagen types I, III, IV, V, laminin, and fibronectin. In addition, the validity of the classification "mesenchymal-epithelioid" for differentiated decidual cells was addressed using antibodies to the intermediate filament proteins, vimentin, a mesenchymal marker, and keratin, an epithelial marker. Biosynthesis of extracellular matrix components was examined by radiolabeling of decidual explants in culture with 3H-proline, followed by immunoprecipitations of synthesized proteins with collagen type-specific antibodies. Immunofluorescence showed decidual cells are embedded in an extensive network of collagen types I and III, and intracytoplasmic staining suggested synthesis of these collagens by the decidual cells. Collagen type IV and laminin localized in the external lamina which surrounds the differentiated decidual cell, and some fluorescence was evident in the peripheral cytoplasm. Immunoreactive collagen type V was observed in close association with the external lamina and in the peridecidual matrix. Fibronectin localized throughout the decidual ECM and in fibrillar and punctuate patterns in the decidual cell cytoplasm. Differentiated decidual cells retained a "mesenchymal" intermediate filament cytoskeleton containing an abundance of vimentin filaments, but very few, if any, keratin filaments. Collagen types I, III, V, and to a lesser extent, IV, were immunoprecipitated from the medium of decidual explants after 24 hours of culture, demonstrating in vitro synthesis and secretion of these collagens by first trimester human decidua.