Trimester Decidua Research Articles

Location of cell cycle regulators cyclin B1, cyclin A, PCNA, Ki67 and cell cycle inhibitors p21, p27 and p57 in human first trimester placenta and deciduas

Although placental development and implantation depend on the coordination of trophoblast proliferation, differentiation and invasion, little is known about the cell cycle regulators that govern the control of these events. The hypothesis that the coordinated expression of cell cycle progression and inhibition factors will determine whether cytotrophoblasts proliferate or undergo cell cycle arrest or cell cycle exit allowing subsequent differentiation was tested. The cell cycle promotors cyclin A, cyclin B1, PCNA, Ki67 and the cell cycle inhibitors p21, p27 and p57 were immunolocalized in tissue sections of first trimester pregnancies (weeks 6 and 9-12). Double staining with cytokeratin 7 allowed unambiguous identification of extravillous cytotrophoblast (EVT) in the decidua. Villous cytotrophoblasts were immunolabelled for Ki67 and cyclin A but only few were stained with anti-cyclin B1. The syncytiotrophoblast was devoid of immunoreactivity for any of the cell cycle progression factors. It expressed especially p21, whereas p27 and p57 were predominantly found in villous cytotrophoblasts. PCNA, Ki67, cyclin A and cyclin B1 were immunolocalized in proximal and distal EVTs of anchoring villi and in EVT which had invaded the upper decidual segments. All EVTs strongly expressed p27 and p57, but not p21. These data clearly suggest different functions for p21, p27 and p57 in placental development with distinct roles for p21 and p57 in syncytiotrophoblast and EVT differentiation, respectively. p27 appears to be involved in both the processes. The results may also challenge the concept of differential mitotic activity in the proximal and distal parts of the first trimester cytotrophoblast cell column, but more functional studies are clearly needed. The presence of p27 and p57 in EVT cells, which invade the deciduas deeply, may account for the loss of mitogenic potential of these cells.

Regulation of Monocyte Chemoattractant Protein-1 Expression by Tumor Necrosis Factor-α and Interleukin-1β in First Trimester Human Decidual Cells: Implications for Preeclampsia

The current study describes a statistically significant increase in macrophages (CD68-positive cells) in the decidua of preeclamptic patients. To elucidate the regulation of this monocyte infiltration, expression of monocyte chemoattractant protein-1 (MCP-1) was assessed in leukocyte-free first trimester decidual cells. Confluent decidual cells were primed for 7 days in either estradiol or estradiol plus medroxyprogesterone acetate to mimic the decidualizing steroidal milieu of the luteal phase and early pregnancy. The medium was exchanged for a serum-free defined medium containing corresponding steroids +/- tumor necrosis factor (TNF)-alpha or interleukin (IL)-1beta. After 24 hours, enzyme-linked immunosorbent assay measurements indicated that the addition of medroxyprogesterone acetate did not affect MCP-1 output, whereas 10 ng/ml of TNF-alpha or IL-1beta increased output by 83.5-fold +/- 20.6 and 103.1-fold +/- 14.7, respectively (mean +/- SEM, n = 8, P < 0.05). Concentration-response comparisons revealed that even 0.01 ng/ml of TNF-alpha or IL-1beta elevated MCP-1 output by more than 15-fold. Western blotting confirmed the enzyme-linked immunosorbent assay results, and quantitative reverse transcriptase-polymerase chain reaction confirmed corresponding effects on MCP-1 mRNA levels. The current study demonstrates that TNF-alpha and IL-1beta enhance MCP-1 in first trimester decidua. This finding suggests a mechanism by which recruitment of excess macrophages to the decidua impairs endovascular trophoblast invasion, the primary placental defect of preeclampsia.

11β-Hydroxysteroid dehydrogenases in human endometrium

Key reproductive events, such as menstruation and implantation, are considered to be inflammatory processes and glucocorticoids act as anti-inflammatory agents. The balance of expression of types 1 and 2 11β-hydroxysteroid dehydrogenases (11βHSD) controls the availability of cortisol to bind to the glucocorticoid receptor (GR) and mineralocorticoid receptor (MR). Expression profiles of glucocorticoid-metabolising enzymes and their cognate receptors have been characterized in the reproductive tract. We propose that factors that peripherally promote glucocorticoid action are part of an anti-inflammatory response to tissue remodelling in human endometrium. Protein and mRNA expression in endometrium were investigated using immunohistochemistry and quantitative real-time PCR. There was up-regulated expression of 11βHSD-1 at menstruation and in first trimester decidua. 11βHSD-2 and GR were expressed across the cycle. The MR expression pattern across the cycle and in decidua implies progesterone may also play a regulatory role. The precise roles and interactions of these proteins require further investigation.

Perforin and Fas/FasL Cytolytic Pathways at the Maternal–Fetal Interface

The immunogenetic enigma of maternal acceptance of the fetal semiallograft has been termed an immunological paradox. The first trimester decidua is heavily infiltrated with CD56(bright) CD16- uterine natural killer (uNK) cells which must be prepared to respond to potential pathogen challenges and still be able to control immune responses that allow the development of the fetus. The significant presence of cytolytic mediators, perforin and Fas/Fas ligand (FasL), at the maternal-fetal interface raises a question of their role(s) in the immunological interrelations between maternal tissues and trophoblast cells. As uNK cells in vitro lyse target cell lines (K562, P815 and P815Fas) using these effector molecules, it seems that, although immunocompetent, their cytotoxicity is not directed against trophoblast during normal pregnancy. Therefore, it is generally believed that the hormonal and Th1/Th2 cytokine balance plays an important role in the tolerance and maintenance of pregnancy. This paper gives an overview of the recent findings on the complex immunological events that occur at the maternal-fetal interface.

A novel protein Depp, which is induced by progesterone in human endometrial stromal cells activates Elk-1 transcription factor

Decidualization of the endometrial stromal cells (ESC), considered to be stimulated by progesterone and/or cAMP, is crucial for embryo implantation and placentation. In this study, we isolated a novel clone encoding decidual protein induced by progesterone (Depp) from a human ESC cDNA library enriched with progesterone-inducible genes. Depp mRNA was expressed in various human tissues including placenta, ovary and kidney. Increased expression of Depp was observed in endometria during mid- and late-secretory phases and 1st trimester deciduas. In vitro, Depp mRNA was induced in ESC within 30 min of progesterone treatment, which was inhibited by the antiprogestin RU486. Androgen alone also induced Depp expression. Depp increased the level of phosphorylated Erk and activated the Elk-1 transcription factor in human embryonal kidney 293 cells, suggesting that Depp modulates the effects of progesterone during decidualization and in the decidua by affecting gene expression. Elucidation of the biological function of Depp in the endometrium will facilitate our understanding of the molecular mechanisms of decidualization and placental development.

Inflammatory Cytokine and Thrombin Regulation of Interleukin-8 and Intercellular Adhesion Molecule-1 Expression in First Trimester Human Decidua

Decidual neutrophil infiltration complicates spontaneous abortions associated with inflammation and hemorrhage. Transendothelial neutrophil migration into inflamed tissues involves IL-8-mediated chemoattraction, then neutrophil attachment to endothelial cell-expressed intercellular adhesion molecule-1 (ICAM-1). The aim of this study was to assess IL-8 and ICAM-1 regulation in decidual inflammation and hemorrhage; the effects of the proinflammatory cytokines, TNFalpha, IL-1beta and the hemostatic, proinflammatory cytokine thrombin were measured on IL-8 expression in first trimester decidual cells (DCs), and ICAM-1 immunostaining was compared in normal, inflamed, and hemorrhagic first trimester decidua. Immunohistochemistry of human decidua and in vitro treatment of human decidual cells were performed. The study was conducted at the Academic Medical Center. . DCs were passaged until they were leukocyte-free (CD45 free by FACS), then were incubated with estradiol or medroxyprogesterone acetate alone or with TNFalpha or IL-1beta or thrombin. Normal, inflamed, and hemorrhagic decidua were immunostained for ICAM-1 and the neutrophil marker CD14. ICAM-1 immunostaining was performed in decidua. IL-8 levels in DC-conditioned media were assessed by ELISA and immunoblotting; IL-8 mRNA levels were measured by quantitative RT-PCR. Endothelial cell ICAM-1 immunostaining was similar in normal and inflamed or hemorrhagic decidua. In cultured DCs, optimal concentrations of IL-1beta, TNFalpha, and thrombin elevated secreted IL-8 levels by 1083 +/- 261-, 370 +/- 77-, and 45 +/- 15-fold, respectively (mean +/- SEM; P < 0.05; n = 8). The effects were dose dependent and were unaffected by medroxyprogesterone acetate. Western blotting confirmed the ELISA results, and corresponding effects on IL-8 mRNA levels were observed. Cytokine/thrombin-enhanced DC IL-8 expression interacts with constitutively expressed ICAM-1 in decidual endothelium to modulate neutrophil trafficking into hemorrhagic and inflamed first trimester decidua.

240. Spatial and temporal expression pattern of furin in the human endometrium

Furin is a proprotein convertase (PC) implicated in the endoproteolytic maturation of inactive protein precursors of growth factors, hormones, receptors, and viral envelope glycoproteins.1 Two functionally active forms of furin, one membrane-bound containing a C-terminal transmembrane domain (TD) and a cytoplasmic tail (CT), and one soluble without the TD and CT, have been characterised. We have previously shown that PC6, one of the PCs closely related to furin, is expressed in the human endometrium and is closely associated with decidualization of stromal cells during implantation.2 Although furin is ubiquitously expressed, its expression in the human endometrium is unknown. In this study, we investigated the spatial and temporal expression pattern of furin in the human endometrium using RT-PCR and immunohistochemistry. While furin expression is detected throughout the menstrual cycle and during early pregnancy, lowest mRNA levels are seen during the proliferative phase. Using an antibody directed against the C-terminus of the membrane bound form, furin is detected in the stroma, glandular and luminal epithelium, as well as in endothelia and neutrophils throughout the menstrual cycle and during early pregnancy. In the stroma, highest levels of furin are present during menstruation (n = 3), they are also high during the proliferative phase (n = 4), but significantly lower levels are detected during the secretory phase (n = 10, P &lt; 0.05, Tukey HSD). In the first trimester decidua, furin is present in well decidualised stromal cells. The overall expression pattern of furin is different to that of PC6; in particular, furin expression is associated only with well decidualized stromal cells whereas PC6 is involved in the initial stages of decidualization. These data suggest that furin and PC6 play different roles in the human endometrium, especially during embryo implantation. (1)Nakayama K. (1997). Biochem. J. 327, 625–635.(2)Nie et al. (2005). Biol. Reprod. 72, 1029–1036.

Effect of steroid hormones on uPA and PAI-1 expression in term decidual stromal cells: Implications for the onset of labor

DECIDUAL STROMAL CELLS: IMPLICATIONS FOR THE ONSET OF LABOR NASTARAN FOYOUZI, LYNN BUCHWALDER, FREDERICK SCHATZ, EDMUND FUNAI, CHARLES LOCKWOOD, ERROL NORWITZ, Yale University, OB/GYN, New Haven, Connecticut OBJECTIVE: Labor is associated with decidual activation. Decidual proteolysis and extracellular matrix degradation is mediated in part by a balance between urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-1 (PAI-1). The effects of sex steroids on uPA and PAI-1 production by human decidual stromal cells is investigated. STUDY DESIGN: Stromal cells were isolated and purified from first trimester and term decidua, and treated with estradiol (10 M [E2]), medroxyprogesterone acetate (10 M [MPA]), both, or vehicle for 7 days. After 24h incubation, uPA and PAI-1 levels in conditioned supernatant were measured by ELISA. Measurements were corrected for protein content, and expressed as meanGSEM. RESULTS: Although unaffected by hormone treatment, basal uPA production was higher by term versus first trimester decidual stromal cells (32.4 G 13.9 vs 4.6 G 2.4 fg/mg protein [P ! .05]). PAI-1 secretion by term decidual stromal cells was higher than by first trimester cells (4.7G 2.1 vs 1.7G 0.2 pg/mg protein [P ! .05]). MPA (not E2) increased PAI-1 production in both populations (3.5and 2.0-fold, respectively). E2 augmented MPA-stimulated PAI-1 production by first trimester (6-fold) but not term decidual cells (1.1-fold). CONCLUSION: Sex steroids exert a differential effect on uPA and PAI-1 production in first trimester and term decidual stromal cells, which may be important in promoting decidual activation and spontaneous labor at term. MPA stimulation of PAI-1 by term decidual cells may account for its pregnancy prolonging properties.

Localisation of Indoleamine 2,3-dioxygenase and Kynurenine Hydroxylase in the Human Placenta and Decidua: Implications for Role of the Kynurenine Pathway in Pregnancy

Indoleamine 2,3-dioxygenase (IDO) has been implicated in contributing to immunotolerance in early pregnancy, but the presence in the term placenta of mRNAs for enzymes that produce other biologically active kynurenine end-products suggests other functions for kynurenine pathway metabolites. The aim of this study was to investigate the localisation of two key enzymes - IDO and kynurenine hydroxylase (KYN-OHase) - in first trimester decidua and in the human placenta across pregnancy. Using immunocytochemistry, it was shown that there was strong expression of IDO and KYN-OHase in stromal and glandular epithelial cells of first trimester decidua. In first and second trimester placenta, IDO and KYN-OHase were localised to the syncytiotrophoblast, stroma and macrophages. IDO and KYN-OHase mRNAs were also identified, and the enzymes appear to be functional because kynurenine and 3-hydroxy-anthranilic acid (respective products of the activity of these enzyme) were released into the medium when first trimester placental explants were maintained in culture for 48h. In term placenta, both IDO and KYN-OHase immunoreactivities were confined mainly to vascular endothelial cells of villous blood vessels, and to macrophages within the fetal villus, whereas syncytial staining was very weak or absent. The shift of expression of these enzymes away from the syncytiotrophoblast to fetal endothelial cells in terminal villi suggests that the function of the enzymes may change from a role in immunosuppression at the maternal-fetal interface in early pregnancy, to one associated with regulation of fetoplacental blood flow or placental metabolism in late gestation.

The Placental Immunomodulatory Cytokine Regeneration and Tolerance Factor is also Expressed by Both Human Cycling and Early Pregnant Endometrium

Regeneration and tolerance factor (RTF) has been recently suggested to contribute to the control of fetal-ablating immunity at the maternal-fetal interface through the induction of T helper 2 (Th2)-dominated response. The protein consists of a membrane-associated domain and an extracellular portion which is proteolitically cleaved to yield a soluble peptide. In humans, it has been shown to be expressed by invading cytotrophoblasts and decidual lymphoid cells, to be increased on peripheral blood B lymphocytes during a normal gestation and on circulating natural killer cells during unsuccessful pregnancies. However, the expression of RTF in other cell types and, specifically, in non-hematopoietic maternal cells of the human uterus has not been characterized in detail. Thus, we have specifically studied the expression and modulation of the cytokine in human endometrium obtained in different phases of the cycle and in early pregnancy. The 20 kDa extracellular domain of RTF has been localized by immunohistochemical method and Western blot analysis. Levels of RTF messenger RNA (mRNA) in basal and stimulated conditions have been evaluated by semiquantitative reverse transcription-polymerase chain reaction. The extracellular domain of RTF could be detected in both the glandular epithelium and stroma with diffuse distribution in both cycling endometrium and first trimester decidua. Both cycling and pregnant endometrium expressed the gene for RTF but mRNA levels resulted significantly increased in secretory phase-endometrial stromal cells when compared to proliferative phase samples. Inflammatory cytokines, interleukin-1beta and tumour necrosis factor alpha were able to directly increase endometrial RTF mRNA expression. These results indicate that RTF is constitutively expressed at endometrial and decidual level and its up-regulation during the secretory phase of the cycle may be relevant in mediating some immune-related aspects of uterine physiology.

The cytokine receptor gp130 and its soluble form are under hormonal control in human endometrium and decidua.

The transmembrane protein gp130 plays a central role in cytokine action as a signal transducing receptor subunit common to all interleukin-6 type cytokines. Endometrial tissue obtained from women with a normal menstrual cycle and decidua obtained from women in the first or second trimester of pregnancy were assessed for gp130 by western blotting, immunohistochemistry and enzyme-linked immunosorbent assay (ELISA) analysis. By immunoblotting, two forms of gp130 were detected: one-the soluble form-of approximately 100 kDa and a larger membrane-bound form of approximately 150 kDa. The latter became clearly visible in the mid to late secretory phase and was more pronounced in decidual tissue of second trimester compared to first trimester. Immunohistochemically, gp130 was located in glandular epithelial cells during the mid to late secretory phase, whereas staining in the proliferative phase was rather weak. In first and second trimester decidua, glandular cells were also positively stained. In addition, the invading trophoblast cells were gp130 positive. Soluble gp130 release was measured in the supernatants from primary endometrial and decidual cell cultures by ELISA and reached maximum values in cell cultures without addition of hormones. In cultured endometrial epithelial cells obtained during the proliferative phase of the cycle, the soluble gp130 release increased significantly under combined estradiol/progesterone supplementation which mimics the secretory phase conditions compared to estradiol supplementation alone. In cultured epithelial cells derived from decidual tissue of first trimester of pregnancy, similar effects of hormonal regulation were observed. Our results suggest that the balance between soluble gp130 and its membrane-bound form may play an important role in regulating cytokine action necessary for blastocyst implantation and for further interaction between the decidualized endometrium and the invading trophoblast.

Macrophages of human first trimester decidua express markers associated to alternative activation.

Depending on the type of their activation, macrophages may promote TH1- or TH2-type of immune responses. To date, not much is known about the activation phenotype of decidua macrophages, which - together with NK cells - constitute the majority of bone marrow derived cells at this location. The study was based on analysis of healthy first trimester decidua by immunohistochemistry and flow cytometry. We analyzed expression of markers characteristic for alternatively activated macrophages (Mphi2). The markers MS-1 (stabilin-1) and coagulation factor XIIIa were found expressed in the interior of decidua macrophages (DMphi). In contrast, indoleamine 2,3-dioxygenase (IDO), an enzyme induced in macrophages by IFNgamma, was not present in DMphi. First trimester DMphi display phenotypic markers associated to alternatively activated macrophages. In addition, absence of IDO indicates that DMphi are not under a predominant influence of IFNgamma.

In-vivo concentrations of inhibins, activin A and follistatin in human early pregnancy

The aims of this study were to investigate the relationship between inhibins, activin A and follistatin in first trimester fetal fluids, maternal serum, placenta and decidua, and to investigate if these hormones are present in the circulation of the early second trimester human fetus. Amniotic and coelomic fluid, maternal serum, placental villi and decidual tissue were obtained from normal pregnancies at 8–12 weeks. Fetal blood by cardiocentesis and maternal blood were collected at 14–16 weeks gestation. Placental extracts had higher concentrations of inhibins, activin A and follistatin compared with decidual extracts. In the second trimester, inhibins and follistatin were detectable in fetal blood at 14–16 weeks gestation. Maternal serum concentrations of inhibin A ( P < 0.001) and follistatin ( P < 0.05) were significantly higher than fetal serum whereas inhibin B ( P < 0.01) and pro-alpha C concentrations ( P < 0.001) were higher in fetal serum. Inhibin B concentrations were also higher in male fetal serum samples that had higher concentrations of testosterone. The presence of all molecular forms of inhibins, activin A and follistatin in the first trimester fetal fluids, placental and decidual extracts in the first trimester confirms other reports. In the second trimester, high concentrations of inhibin B with testosterone in the fetal circulation indicate that these hormones may interact in the development of the male fetal gonads.

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).

Lymphocyte subset distribution and cytokine secretion in third trimester decidua in normal pregnancy and preeclampsia

Background: Excessive Th1 activity in peripheral blood plays a probable role in the pathogenesis of preeclampsia. The aim of the study was to investigate whether disturbed local immune reactions are also present in decidua. Methods: Flow cytometric analysis of CD3, CD19, CD56/CD16, CD4, CD8, CD4/CD29, CD4/CD45RA, CD4/CD45RO, CD8/CD28, CD3/CD69 lymphocyte subsets isolated from third trimester decidua of pregnants with preeclampsia ( n=21) and pregnant controls ( n=11) subjected to elective caesarean sections. Spontaneous and phytohemaglutynine stimulated “in vitro” secretion of IL-2, IL-4, IL-6, IL-10, IL-12, IFN-γ and TGF-β by decidual lymphocytes was studied by ELISA. For the statistical significance of differences between the groups the U Mann–Whitney test was performed (confidence interval P<0.05). Results: Preeclamptic patients were characterized with an increased percentage of the CD3−/CD56+CD16+, CD8+/CD28+ and decreased percentage of CD3+, CD19+, CD4+/CD45RA+ lymphocytes. The profile of secreted cytokines shifts in favor of Th1 activity (extremely high IFN-γ and low IL-6 and IL-10 secretion). Decidual IL-12 secretion in preeclamptic patients is decreased compared to controls. Conclusion: Changes in NK and T lymphocyte subsets followed with Th1 cytokine IFN-γ over-activity, could affect local immunoregulatory mechanisms in third trimester decidua of preeclamptic patients.

Characterisation of antigen presenting cells in the human first trimester decidua

Characterisation of antigen presenting cells in the human first trimester decidua

Immunoregulatory properties of decidua macrophages

About 15–20% of bone marrow derived cells in the endometrium and the decidua are CD14‐positive macrophages coexpressing the scavenger receptor CD163. The costimulatory molecule CD86 (B7‐2) but not CD80 (B7‐1) is found expressed on first trimester and term decidua macrophages (D‐Mph). The number of D‐Mph increases at the implantation site in the basal decidua, where they are in contact with the HLA‐G‐positive invading extravillous cytotrophoblast. As D‐Mph express the inhibitory receptors CD85d (ILT4, LIR‐2) and CD85e (ILT2, LIR‐1), they are potentially susceptible to inhibition by class Ia and class Ib molecules. D‐Mph may have specific functions relating to successful pregnancy, and phenotypic and functional properties may differ between early and term pregnancy. A number of data make their molecular repertoire comparable to those of alternatively activated macrophages which are immunosuppressive. For instance, first trimester decidua macrophages express IL‐10 and the MS‐1 high molecular weight antigen on the cell surface fibronectin and F XIIIa are detectable. Functional data are required to complement these studies.

Absence of HLA-G expression in macrophages of human decidua.

Macrophages - together with natural killer (NK) cells - constitute the majority of bone marrow derived infiltrating cells in the decidua. As interferon-gamma (IFN-gamma), a cytokine produced by NK cells, has been reported to induce expression of human leukocyte antigen (HLA-G) in monocytic cells, suggesting expression of HLA-G on decidua macrophages potentially stimulated by IFN-gamma, the question arises whether decidua macrophages in normal pregnancy express HLA-G. The study was based on immunohistochemistry and flow cytometry. In order to exclude that potentially elusive soluble HLA-G was not detected by immunohistochemistry, we performed in addition RT-PCR of flow-sorted decidua macrophages. Our findings indicate that HLA-G is not present on macrophages of first trimester or term decidua in either membrane-bound or soluble form. Transcripts for soluble HLA-G1 and -G2 were not detected. We exclude a role of HLA-G on the surface of decidua macrophages or of soluble HLA-G1 or -G2 as a secretory product of decidua macrophages with regard to interaction with HLA-G receptors present in or outside the decidua.

The Ontogeny of 25-Hydroxyvitamin D 3 1α-Hydroxylase Expression in Human Placenta and Decidua

In addition to its classical calciotropic effects, the active form of vitamin D, 1,25-dihydroxyvitamin D(3) (1,25(OH)(2)D(3)) is a potent anti-proliferative/immunomodulatory secosteroid. The enzyme that catalyzes the synthesis of 1,25(OH)(2)D(3), 1alpha-hydroxylase (1alpha-OHase), is expressed in many human tissues, highlighting its possible role as an autocrine/paracrine activator of vitamin D. Immunohistochemical and RNA analyses were used to characterize the ontogeny of 1alpha-OHase expression in human placenta and decidua. Protein for 1alpha-OHase was detectable in trophoblast and decidua; the latter being stronger in decidualized stromal cells than macrophages, with no staining of lymphocytes. Quantitative reverse transcriptase-polymerase chain reaction was used to assess changes in mRNA expression for 1alpha-OHase at different gestations: first (mean, 9.1 +/- 1.5 weeks); second (mean, 14 +/- 1.8 weeks), and third trimester (mean, 39.3 +/- 2.5 weeks). 1alpha-OHase expression in decidua was approximately 1000-fold higher in first (95% confidence limits, 611 to 1376) and second (95% confidence limits, 633 to 1623) trimester biopsies when compared with the third trimester (95% confidence limits, 0.36 to 2.81) (both P < 0.001). In placenta, 1alpha-OHase expression was 80-fold higher in the first (range, 42 to 137) and second (range, 30 to 199) trimester when compared with third trimester biopsies (0.6 to 1.6) (both P < 0.001). Similar results were obtained by semiquantitative IHC. Parallel analysis of the receptor for 1,25(OH)(2)D(3) (vitamin D receptor) indicated that, as with 1alpha-OHase, highest levels of expression occurred in first trimester decidua. However, changes in vitamin D receptor mRNA expression across gestation were less pronounced than 1alpha-OHase. These spatiotemporal data emphasize the potential importance of 1alpha-OHase during early fetoplacental life and, in particular, suggest an autocrine/paracrine immunomodulatory function for the enzyme.

IL-15 Regulation in Human Endometrial Stromal Cells

A greater knowledge of IL-5 regulation within human endometrium is important in understanding key reproductive events and uterine Natural Killer cell function. In the present study, expression of IL-15 mRNA was shown to be up-regulated by both PGE(2) and IFN-gamma in cultures of human endometrial stromal cells (ESC). Release of IL-15 protein was also shown to be under the control of PGE(2) and IFN-gamma using an enzyme-linked immunosorbent assay for IL-15. In addition, 8-Bromo cAMP was able to increase IL-15 release from ESCs (P < 0.005) implying the actions of PGE(2) may be via this second messenger. Addition of a progestin appeared to enhance these effects. Real-time quantitative PCR has demonstrated an up-regulation in IL-15 mRNA expression in the late secretory phase of the menstrual cycle (P < 0.005) and a progressive rise in IFN-gamma expression throughout the secretory phase and into first trimester decidua. These results suggest that IL-15 regulation in the human endometrium is complex and that hormonal control may be indirect.