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Experten-Interview | Mit einem Bluttest können Schwangere seit 2012 feststellen lassen, ob ihr Kind das Down-Syndrom haben wird. Der nicht invasive Pränataltest ist risikoärmer und kann früher erfolgen als eine Fruchtwasseruntersuchung. Der Gemeinsame Bundesausschuss hat am 19. September 2019 beschlossen, dass ein NIPT in begründeten Einzelfällen nach ärztlicher Beratung unter Verwendung einer noch zu entwickelnden Versicherteninformation als Kassenleistung eingesetzt werden kann. Pränataldiagnostiker Dr. Stefan Hutter beschreibt die Untersuchungsmethode.

The objective of this study was to analyze the expression of the glucocorticoid receptor (GR) subtypes GRα and GRβ in placentas affected by intrauterine growth restriction (IUGR). We analyzed the sex-specific placental expression of GRα and GRβ in 23 IUGR and 40 control placentas using immunohistochemistry and immunofluorescence. The GR gene, also known as nuclear receptor subfamily 3 group C member 1 (NR3C1), mRNA production in trophoblast-like cell line BeWo after stimulation with prednisolone was analyzed using quantitative polymerase chain reaction (qPCR) and on the protein level using western blot analysis. GR subtypes showed a sex-specific upregulation in placentas from IUGR compared to control placentas. An increased expression of GRα was detectable in female placental tissue, whereas GRβ was increased in males. Our data support previous findings suggesting that the glucocorticoid metabolism plays a role in the pathophysiology of IUGR. Furthermore, the data suggest that the underlying molecular mechanisms differ between male and female cases.

Serious hepatic complications, although rare, are one of the leading causes of maternofetal morbidity and mortality in hypertensive pregnancy disorders. A 28-year-old primigravida was transferred to our hospital complaining of refractory epigastric pain in the 29th week of pregnancy and was subsequently admitted due to superimposed pre-eclampsia and hemolysis, elevated liver enzyme levels, and low platelet count syndrome. Following a pathological cardiotocogram, a cesarean section was performed. The intra-abdominal situs presented with 1000 mL of blood and a bleeding rupture of the left lobe of the liver. The trauma to the liver was surgically repaired with a suture and the patient's state was stabilized. Following the surgical procedures and neonatal intensive care, mother and newborn both recovered without residues. In order to avoid unnecessary maternal morbidity, we therefore recommend an abdominal ultrasound, beyond an obstetric focus, as an additional and sensible means of diagnostic imaging in cases of hemolysis, elevated liver enzyme levels, and low platelet count syndrome.

Gestational diabetes mellitus (GDM) is known to increase the risk for feto-maternal complications during pregnancy. A state of low-grade inflammation, with elevated levels of proinflammatory molecules, similar to patients with obesity or diabetes mellitus type 2 has also been partly described in GDM. The placenta, as unique interface between mother and fetus, is not only passively affected by changes in one of these organisms, but also acts as a modulator by expressing hormones and cytokines. This study aimed to investigate the expression of the proinflammatory cytokines Interleukin (IL) 7, 8 and 15 in GDM in placental tissue. A total number of 80 placentas were included (40 GDM/40 control group). The expression of IL-7, 8 and 15 was investigated in extravillous trophoblast (EVT) and syncytiotrophoblast (SCT) by immunohistochemistry and immunofluorescence double staining. The immunohistochemical staining was evaluated with the semiquanitfied immunoreactive score (IRS). While the expression IL-15 was significantly upregulated in EVTs of women with GDM. The expression of IL-8 was significantly decreased in EVT of the GDM group. Furthermore, significant fetal sex specific differences were detectable in all three cytokines. Our findings suggest an involvement of the investigated cytokines in the maintenance of a state of chronic low-grade inflammation on placental level in patients suffering from GDM.

Thyroid hormones are essential for development of trophoblasts and the fetus. They also regulate a wide range of metabolic processes. We investigated the influence of maternal gestational diabetes mellitus (GDM) on thyroid hormone receptor (THR) isoforms THR&alpha, 1, THR&alpha, 2, THR&beta, 1 and THR&beta, 2 of the human placenta in a sex- and cell-type specific manner. Term placental tissue was obtained from women with (n = 40) or without GDM (control, n = 40). THRs levels were measured by semi-quantitative immunohistochemistry and real-time qRT-PCR. We localized THR immunostaining in syncytiotrophoblast (SCT), which was the tissue with the strongest signal. Double immunofluorescence identified THR in decidual cells in the stroma and in extravillous cytotrophoblasts. GDM did not change THR&alpha, 1 immunolabelling intensity in decidua, but was associated with a stronger immunolabelling in SCT compared to GDM (p &lt, 0.05). The SCT difference of GDM vs. control was strongest (p &lt, 0.01) in female placentas. THR&alpha, 2 was only weakly present and immunolabelling was weaker (p &lt, 0.05) in SCT of only male GDM placentas in comparison to male controls. THR&beta, 1/&beta, 2 immunostaining was weak in all cell types without changes in GDM. However, more THR&beta, 1/2 protein was present (p &lt, 0.001) in male than female placentas. All these protein changes were paralleled by changes of THR transcript levels. The data show that THR are expressed in term trophoblast in relation to fetal sex. Maternal GDM influences predominantly THR&alpha, 1 in SCT, with the strongest GDM effect in SCT of female placentas.

Gestational diabetes mellitus (GDM) is the most common pregnancy-associated metabolic disorder that negatively impacts on the health of both mothers and their offspring in the long-term. The molecular mechanisms involved are not fully understood. As in other states of insulin resistance, a disproportionate immune response in GDM leads to a state of chronic low-grade inflammation. Galectin-2 exerts regulatory effects on different immune cells. This study investigated galectin-2 expression in the placenta of 40 GDM patients and 40 controls, in a sex-specific manner. Immunohistochemistry was used for semi-quantitative analysis of expression strength. The phenotypes of galectin-2 expressing cells were characterized through double immunofluorescence. We found a significant up-regulation of galectin-2 in the fetal syncytiotrophoblast, as well as in the maternal decidua of GDM placentas. Double staining showed a strong galectin-2 expression in extra villous trophoblast cells and fetal endothelial cells in GDM. These findings present the first systematic investigation of galectin-2 in GDM. The findings contribute to the emerging understanding of the role of immunomodulation and inflammation in GDM and of galectin-2 itself. This might also have implications for the long-term cardiovascular health of the offspring.

Objectives l-dopa decarboxylase (DDC) is responsible for the synthesis of dopamine. Dopamine, which binds to the D2-dopamine receptor (D2R), plays an important role in the maintenance of pregnancy. Aim of our study was the analysis of DDC and D2R expression in placentas of spontaneous miscarriages (SMs) and recurrent miscarriages (RMs) in comparison to healthy controls. Methods Patients with SM (n = 15) and RM (n = 15) were compared with patients from healthy pregnancies (n = 15) (pregnancy weeks 7–13 each). Placental tissue has been collected from SMs and RMs from the first trimester (Department of Gynaecology and Obstetrics, LMU Munich) and from abruptions (private practice, Munich). Placental cell lines, BeWo- and JEG-3 cells, were stimulated with the trace amines T0AM and T1AM in vitro. Results Levels of DDC and D2R in trophoblasts and the decidua were lower in RMs in comparison to healthy controls. Stimulation of BeWo cells with T1AM significantly reduced DDC mRNA and protein levels. Via double-immunofluorescence, a DDC-positive cell type beneath decidual stromal cells and foetal EVT in the decidua could be detected. Conclusions Downregulation of DDC and D2R in trophoblasts of RMs reflects a reduced signal cascade of catecholamines on the foetal side.

This article contains errors in Figs. 5 and 6, for which we apologize. In Fig. 5f, the image ‘E12.5 tail’ was inadvertently replaced with a duplicate of the image ‘E12.5 trunk’ from the same panel. In Figure 6d, the image ‘E9.5/OH-TAM E8.5, embryo’ was inadvertently replaced with a duplicate of the image ‘E10.5/ OH-TAM E8.5, embryo’ from Fig. 6b. The corrected versions of these figures appear in the Author Correction associated with this Article.

Despite the ever-rising incidence of Gestational Diabetes Mellitus (GDM) and its implications for long-term health of mothers and offspring, the underlying molecular mechanisms remain to be elucidated. To contribute to this, the present study’s objectives are to conduct a sex-specific analysis of active histone modifications in placentas affected by GDM and to investigate the effect of calcitriol on trophoblast cell’s transcriptional status. The expression of Histone H3 lysine 9 acetylation (H3K9ac) and Histone H3 lysine 4 trimethylation (H3K4me3) was evaluated in 40 control and 40 GDM (20 male and 20 female each) placentas using immunohistochemistry and immunofluorescence. The choriocarcinoma cell line BeWo and primary human villous trophoblast cells were treated with calcitriol (48 h). Thereafter, western blots were used to quantify concentrations of H3K9ac and the transcription factor FOXO1. H3K9ac expression was downregulated in GDM placentas, while H3K4me3 expression was not significantly different. Cell culture experiments showed a slight downregulation of H3K9ac after calcitriol stimulation at the highest concentration. FOXO1 expression showed a dose-dependent increase. Our data supports previous research suggesting that epigenetic dysregulations play a key role in gestational diabetes mellitus. Insufficient transcriptional activity may be part of its pathophysiology and this cannot be rescued by calcitriol.

Glycosylation is one of the most important posttranslational modifications of proteins and lipids that contributes to the structural diversity of cellular molecules. Enzymes of the glycosyltransferase class are responsible for altering glycosylation patterns by adding carbohydrate chains to the respective acceptor molecules. It is well known that glycosylation is commonly altered in cancerous tissue. Therefore, the present study aimed to determine the incidence of N‑acetylgalactosaminyltransferase 6 (GALNT6), a prominent member of the glycosyltransferase class, in breast cancer tissue of different developmental stages by immunohistochemistry. Although no correlation was identified between tumour characteristics and GALNT6 staining intensity, to the best of our knowledge, this is the first study to demonstrate that tissue from carcinoma in situ‑tumours and metastases were more heavily stained than late‑stage breast cancers. This may indicate an important role of glycosylation aberration in escaping the immune system at early phases of tumour development. The present study also hypothesised that nascent or early metastasizing tumours are normally recognized by the immune system of the patient, but glycosylation pattern changes may facilitate tumor escape from immune recognition. In follow‑up studies, our group will aim to confirm and consolidate these results in a larger patient cohort that may give greater insight into breast cancer characterization as well as tumour treatment.

The nuclear receptor estrogen receptor α (ERα) is one of the key players in energy balance, insulin resistance, and trophoblast differentiation. We tested the hypothesis that gestational diabetes mellitus (GDM) alters expression of placental ERα in a cell type-specific manner and that this regulation may involve epigenetic changes.Expression of ERα was analyzed by immunohistochemistry using the semiquantitative immunoreactive score in 80 placentas (40 GDM/40 controls). Quantitative real-time polymerase chain reaction (PCR) measured ERα messenger RNA (mRNA) in decidual tissue. Methylation-specific PCR was performed to analyze cytosine-phosphatidyl-guanine-island methylation of the ERα promoter.Expression of ERα protein is upregulated (P = .011) in GDM in extravillous trophoblasts but not in syncytiotrophoblast. Gestational diabetes mellitus downregulated ERα in decidual vessels only in pregnancies with male but not female fetuses. Furthermore, mRNA of the ERα encoding gene estrogen receptor gene 1 (ESR1) was increased (+1.77 fold) in GDM decidua when compared to controls (P = .024). In parallel, the promoter of ESR1 was methylated only in decidua of healthy control individuals but not in GDM.Gestational diabetes mellitus affects expression of placental ERα in a cell type-dependent way, on epigenetic level. These data link GDM with epigenetic deregulations of ERα expression and open new insights into the intrauterine programming hypothesis of GDM.

Background The purpose of this study was to investigate the sex specific expression of histone protein modifications responsible for rapid gene expression in IUGR placentas. Patients and Methods We screened for fetal sex-specific expression of the histone proteins H3K4me3 and H3K9ac in 23 IUGR and 40 control placentas via immunohistochemistry. The trophoblast-like cell line BeWo was used in order to analyze a potential effect of stimulation with prednisolone on H3K4me3 and H3K9ac in vitro. Calculating regression models with additional adjustment for potential confounders were used. Results A significantly decreased level of H3K4me3 was detectable in female syncytiotrophoblasts, whereas H3K9ac was reduced predominantly in male extravillous throphoblast (EVT). No association to the gestational age existed. Conclusion Our data showed a reduced expression of the histone proteins H3K4me3 (female) and H3K9ac (male) in IUGR, furthermore elevated cortisol levels may lead to a sex-specific down-regulation of histone proteins in IUGR placentas.

The pathogenesis of type 2 diabetes (T2D) is still incompletely understood. In-depth phenotyping of young individuals at risk for T2D can contribute to the understanding of this process.The purpose of this study was to metabolically characterize women with recent gestational diabetes (GDM), an at-risk cohort for T2D.Participants were 147 women consecutively recruited 3 to 16 months after pregnancy: women who had GDM and women after a normoglycemic pregnancy (control subjects) in a 2:1 ratio.This was a monocenter cross-sectional analysis (Prediction, Prevention and Subclassification of Type 2 Diabetes Study [PPS-Diab]).A 5-point oral glucose tolerance test with calculation of the insulin sensitivity index and disposition index (validation by euglycemic clamp and intravenous glucose tolerance test) was performed. In addition, anthropometrics, medical and family history, clinical chemistry and biomarkers, statistical modeling, and a magnetic resonance imaging/magnetic resonance spectroscopy substudy (body fat distribution and liver and muscle fat; n = 66) were obtained.Compared with control subjects, women after GDM had a reduced disposition index, higher levels of plasma fetuin-A, and a lower insulin sensitivity index. A low insulin sensitivity index was also the major determinant of pathological glucose tolerance after GDM. The factors most strongly predictive of low insulin sensitivity were high plasma leptin, body mass index, triglycerides, and waist circumference. Ectopic lipids showed no body mass index-independent associations with having had GDM or low insulin sensitivity in a magnetic resonance imaging substudy.We found that β-cell function is already impaired in women with recent GDM, a young at-risk cohort for T2D. In addition, our data suggest that fetuin-A and leptin signaling may be important early contributors to the pathogenesis of T2D, at this disease stage equally or more relevant than ectopic lipids and low-grade inflammation.

Gestational diabetes mellitus (GDM) is often accompanied by low maternal vitamin D, that is, calcitriol (1,25[OH]2 vitamin D3), levels. Here, we tested the hypothesis that the placental vitamin D receptor (VDR) is regulated by calcitriol and altered in GDM with distinct changes in different placental cell types. Specifically, we aimed to localize VDR in human term placentas from normal and GDM pregnancies, to quantify its cellular expression and to study in vitro its regulation by its physiological agonist calcitriol.Placental tissue slides of 80 patients (40 with GDM/40 controls) were double stained for VDR and human leukocyte antigen G to identify extravillous trophoblasts (EVTs). Staining intensity was semiquantified. Quantitative real time-polymerase chain reaction and Western blotting measured VDR messenger RNA (mRNA) and protein in decidual tissue. The trophoblast cell line BeWo was used to study in vitro VDR regulation by calcitriol (0.01, 0.1, and 1 nmol/mL).Vitamin D receptor protein and mRNA levels are upregulated (P.05) in EVT (1.8-fold) as well as in placental endothelium (5.8-fold) of patients with GDM. Expression of VDR is regulated by calcitriol in a bimodal manner: high doses (0.1 and 1 nmol/mL) caused downregulation, whereas the low dose (0.01 nmol/mL) resulted in VDR upregulation.Vitamin D receptor is upregulated in EVT and endothelium of GDM placentas. This could be due to low maternal vitamin D levels in patients with GDM because in vitro low calcitriol doses upregulate VDR in trophoblast cells.

Aims: Peroxisome proliferator-activated receptor-gamma (PPARγ) plays an important role in insulin metabolism, trophoblast differentiation and anti-inflammatory circuits. The aim of this study was to investigate the expression of PPARγ in the placenta of patients with gestational diabetes mellitus (GDM) and the regulation of PPARγ by its agonists in trophoblast tumour cells BeWo in vitro. Methods: PPARγ expression in a total of 80 placentas (40 GDM/40 controls) was analysed by immunohistochemistry using the semi-quantitative immunoreactive score. Furthermore, a quantitative reverse transcription-polymerase chain reaction (PCR) was performed to determine the PPARγ mRNA-expression in both groups. We used a fused and a non-fused BeWo cell culture model for the stimulation with arachidonic acid and 15-Deoxy-Δ12,14-prostaglandin J2 (15d-PGJ2). Afterwards PPARγ mRNA-expression was analysed by quantitative real-time PCR (RT-PCR) (TaqMan). Results: Using immunohistochemistry we identified a decreased expression of PPARγ in the syncytiotrophoblast and the extravillous trophoblast of GDM placentas compared to normal controls. Furthermore, PPARγ mRNA-expression was reduced in GDM placentas. Stimulation of BeWo cells with arachidonic acid and 15d-PGJ2 caused a downregulation of PPARγ expression. Conclusion: As PPARγ is down regulated by arachidonic acid and 15d-PGJ2, the reduced PPARγ expression in GDM placentas may be due to an altered concentration of fatty acid derivates.

It is widely known that cells from epithelial tumors, e.g., breast cancer, detach from their primary tissue and enter blood circulation. We show that the presence of circulating tumor cells (CTCs) in samples of patients with primary and metastatic breast cancer can be detected with an array of selected tumor-marker-genes by reverse transcription real-time PCR. The focus of the presented work is on detecting differences in gene expression between healthy individuals and adjuvant and metastatic breast cancer patients, not an accurate quantification of these differences. Therefore, total RNA was isolated from blood samples of healthy donors and patients with primary or metastatic breast cancer after enrichment of mononuclear cells by density gradient centrifugation. After reverse transcription real-time PCR was carried out with a set of marker genes (BCSP, CK8, Her2, MGL, CK18, CK19). B2M and GAPDH were used as reference genes. Blood samples from patients with metastatic disease revealed increased cytokine gene levels in comparison to normal blood samples. Detection of a single gene was not sufficient to detect CTCs by reverse transcription real-time PCR. Markers used here were selected based on a recent study detecting cancer cells on different protein levels. The combination of such a marker array leads to higher and more specific discovery rates, predominantly in metastatic patients. Identification of CTCs by PCR methods may lead to better diagnosis and prognosis and could help to choose an adequate therapy.

Macrophages hold a key role in both regulating and executing the body's own immune response under various conditions. Hence, although endometriosis, preeclampsia and abortions are clinically different, all three are regarded to involve highly complex immunological processes. The aim of our current work was to evaluate the role of macrophages within these gynaecological disorders. Macrophages have been shown to invade endometriosis lesions and to mediate propagation of endometriotic cyst growth. However this is the first time that significant GPER up-regulation in macrophages is demonstrated. This highlights a potential alternative way through which estrogen may modulate immune response of macrophages in endometriosis. In addition, during spontaneous miscarriages the macrophage population increases significantly. This deregulation may possibly support an inflammatory scheme further triggering abortive procedures. Macrophage-mediated apoptosis of extravillous trophoblasts (EVT) has been associated with preeclampsia. Larger numbers of apoptotic EVT were detected in preeclamptic placentas compared with normal. In preeclamptic placentas, decidual macrophages were found to be Fas ligand (FasL)-positive. Our results highlight a new aspect of macrophage biology in endometriosis and pregnancy physiology and patho-physiology. Further studies with larger samples are needed to verify the current results and evaluate their clinical impact. Our data strongly indicate that macrophages hold key roles in various gynaecological disorders and might be crucial to further elucidate their patho-physiology.

Background: Circulating tumor cells (CTCs) are cells that detach from a primary tumor, circulate through the blood stream and lymphatic vessels, and are considered to be the main reason for remote metastasis. Due to their origin, tumor cells have different gene expression levels than the surrounding blood cells. Therefore, they might be detectable in blood samples from breast cancer patients by real-time quantitative polymerase chain reaction (RT-qPCR). Materials and Methods: Blood samples of healthy donors and adjuvant breast cancer patients were withdrawn and the cell fraction containing white blood cells and tumor cells was enriched by density gradient centrifugation. RNA was isolated and reverse transcribed to cDNA, which was then used in TaqMan real-time PCR against cytokeratin (CK)8, CK18 and CK19. 18S and GAPDH were used as controls. Results: All 3 CKs were, on average, found to be significantly higher expressed in adjuvant breast cancer samples compared to negative controls, probably due to the presence of CTCs. Unfortunately, gene expression levels could not be correlated to tumor characteristics. Conclusions: RT-qPCR could make up a new approach for the detection of CTCs from blood samples of breast cancer patients, but a correlation of the PCR data to gold standard methods in CTC detection would help to further improve the informative value of the qPCR results.

The occurrence of disseminated tumour cells in bone marrow of patients with breast cancer is linked to a worse prognosis. We present a method for DTC detection from bone marrow samples based on immunocytochemistry, using breast cancer-associated glycosylation molecules as markers for detection and characterization.A double immunofluorescence staining of a pan-cytokeratin (CK) marker and either Tn or O-Acetyl-GD3 was carried out in artificial and patient bone marrow samples.The sample in which most cells stained positive for CK/Tn and CK/O-AC-GD3, was obtained from a patient who certainly had remote metastases. All other bone marrow samples showed heterogenous staining, so no correlation to tumour characteristics could be revealed.A certain characterization of tumour cells can be achieved by a double staining of bone marrow samples with CK and a glycosylation marker. For future studies, analysis should be extended to a larger patient collective and further examination of more glycosylation markers should be carried out.

The detection of circulating tumour cells (CTCs) from peripheral blood of cancer patients can be carried out by real-time PCR approaches using different gene expression levels of tumour cells and surrounding blood cells.Potential marker genes were first analyzed in a model system and then applied to 20 blood samples of adjuvant breast cancer patients and gene expression levels were correlated to tumour characteristics.The mean of gene expression levels was found elevated for the four genes analyzed in the adjuvant breast cancer patient group in comparison to the samples of the group of healthy donors, but no correlation between gene expression and tumour characteristics could be detected as being statistically significant.The results demonstrated, that the employed methodology is functional, but has to be refined by certain approaches like simultaneously running a state-of-the-art system of CTC-detection comparing the results, and by an enlargement of patient collective and number of marker genes.

Endometrial adenocarcinoma is a frequently occurring cancer in women, accounting for 42,000 deaths every year. Despite treatment with standard therapy, occurrence of remote metastases and local recurrences is high. Through help of RT-qPCR minimal residual disease could be detected and characterized, facilitating therapeutic decision making.A number of marker genes were first tested in model systems and genes that performed best, were consequently used for the examination of 13 blood samples from endometrial carcinoma patients.Cytokeratin 19 and MIG7 were chosen for the analysis in patient samples. Both genes were found up-regulated in small tumours and in one large tumour, but no statistical correlations could be revealed between expression levels of these two genes and tumour characteristics.There seems to be a coherence between gene expression and the stage of tumorigenesis, but the number of samples is still too small, to be able to obtain statistical significant differences.

Galectins (gal) are members of the mammalian β-galactoside-binding proteins and recognize Galβ1-4GlcNAc and Galβ1-4GalNac (Thomsen-Friedenreich antigen (TF)) sequences of several cell surface oligosaccharides. In this study, gal-1, -2, -3 and -13 were investigated systematically in the trophoblast and decidua compartment of intrauterine growth restriction (IUGR) placentas and normal third trimester control placentas and stratified by fetal gender and gestational age. Within this study, 29 third trimester placentas after delivery were analyzed. Fetal gender was equally divided within both groups, and immunohistochemical staining was analyzed according to fetal gender and gestational age. Double immune-fluorescence with trophoblast-specific markers was used to identify galectin-expressing cells at the feto-maternal interface in the decidua. Gal-3 was significantly downregulated only in the extravillous trophoblast of IUGR placentas. In contrast, expressions of gal-2 and gal-13 were downregulated in both villous and extravillous trophoblast cells of IUGR placentas. In addition, gal-2 and gal-13 showed a highly correlated expression scheme in the placenta. There are significant gender-specific expression patterns for single prototype galectins with downregulation of gal-2 and gal-13 of male gender placentas in cases of IUGR. Gal-3 as the chimera type galectin shows only little gender-specific differences in expression, which disappear in IUGR cases.

Galectins are galactose binding proteins and, in addition, factors for a wide range of pathologies in pregnancy. We have analyzed the expression of prototype (gal-1, -2, -7, -10) and chimera-type (gal-3) galectins in the placenta in cases of spontaneous abortions (SPA) and recurrent abortions (RA) in the first trimester. Fifteen placental samples from healthy pregnancies were used as a control group. Nine placentas were examined for spontaneous abortions, and 12 placentas for recurrent abortions. For differentiation and evaluation of different cell types of galectin-expression in the decidua, immunofluorescence was used. For all investigated prototype galectins (gal-1, -2, -7, -10) in SPA and RA placenta trophoblast cells the expression is significantly decreased. In the decidua/extravillous trophoblast only gal-2 expression was significantly lowered, which could be connected to its role in angiogenesis. In trophoblasts in first-trimester placentas and in cases of SPA and RA, prototype galectins are altered in the same way. We suspect prototype galectins have a similar function in placental tissue because of their common biochemical structure. Expression of galectin 3 as a chimera type galectin was not found to be significantly altered in abortive placentas.

Galectins are members of the mammalian β-galactoside-binding proteins, which recognize Galβ1-4GlcNAc sequences of several cell surface oligosaccharides. Plenty of galectins are already described in human tissue, especially in placenta. Here, gal-1-4, 7-10 and gal-12 were investigated systematically in trophoblast and decidua cells of first trimester placentas.Within this study, 15 first trimester placentas after induced abortion (7th-14th week of gestation) were examined with immunohistology and immunofluorescence based on a scoring system. Moreover, isolated and cultivated trophoblast cells from the first trimester were analyzed and evaluated for expression of gal-1-4, gal-7-10 and gal-12 at mRNA and protein level with real-time RT-Polymerase chain Reaction/PCR (Taq-Man). Double immunofluorescence with trophoblast specific markers identified galectin expressing cells at the feto-maternal interface.We could detect immunohistochemical staining of galectins 1-4, 7-10 and 12 in first trimester placenta: all examined galectins were found in the cytotrophoblast (CTB) and syncytiotrophoblast (SCT). Gal-1, -2, -3, -4, -7, -8, -9, -10 and -12 were identified in extravillous trophoblast cells (EVT) in immunohistology and immunoflourescence. The expression of gal-1, -9, -10, and gal-12 increased after 96h incubation in vitro without stimulation at mRNA level, while gal-2, -3, -4, -7 and -8 were decreased.This study describes a systematic analysis of the expression of gal-1-4, gal-7-10 and gal-12 in first trimester placentas and isolated trophoblast cells. Expression levels at mRNA level and the change within 96h cultivation in vitro indicate a possible influence on syncytium building of trophoblast cell on expression of galectins. Therefore, an interaction of galectins in vitro in syncytium building is possible.

Preeclampsia (PE) and intrauterine growth retardation (IUGR) are rare but severe pregnancy complications that are associated with placental insufficiency often resulting in premature birth. The clinical pathologies are related to gross placental pathologies and trophoblastic deficiencies that might derive from inflammatory processes and oxidative stress injury. The mesenchymal core of placental villi has been identified as a possible niche for trophoblast progenitor cells that are called upon to replenish the injured syncytiotrophoblast layer. These progenitor cells are known to express trophoblast stem cell (CDX2) and pluripotency (SOX2, NANOG and OCT4A) markers, however only little data is available characterizing the expression of these transcription factors beyond the blastocyst stage. We aimed to describe the expression of these factors in healthy 1st and 3rd trimester placentae as well as PE, IUGR and combined PE+IUGR placentae. We analyzed 8 respective samples derived from 1st trimester (elective abortions), and 3rd trimester (healthy controls, PE, IUGR and combined PE+IUGR). We accomplished immunoperoxidase staining to detect the stem cell markers: CDX2 (trophectoderm), SOX2, NANOG and OCT4A (embryonal). Immunoreative scoring was used for objective analyses of staining patterns. All markers display clearly elevated signals in 1st trimester villous samples as compared to healthy 3rd trimester counterparts. Especially CDX2 and NANOG were specific to the cytotrophoblast layer and the mesenchymal core. Specific and differential expression patterns were visible in the villous/extravillous compartment of each placenta-associated pregnancy complication (PE: pan elevated expression; IUGR elevated SOX2 in basal plate; combined PE+IUGR pan loss of expression). Reduction of stem cell transcription factor expression in term placentae indicates temporal regulation, and probably a specific function which is yet to be elucidated. The differential expression patterns within placentae complicated with placenta-associated pregnancy complications indicate that PE, IUGR and combined PE+IUGR are separate entities. It is unclear whether the alterations are the cause or the effect of the clinical pathology.

Galectin-1 (gal-1), a member of the mammalian β-galactoside-binding proteins, exerts biological effects by recognition of glycan ligands, including those involved in cell adhesion and growth regulation. In previous studies, we demonstrated that gal-1 induces cell differentiation processes on the membrane of choriocarcinoma cells BeWo, including the receptor tyrosine kinases (RTKs) REarranged during Transfection (RET), Janus Kinase 2 (JAK2) and Vascular endothelial growth factor receptor 3 (VEGFR3). Furthermore, Mitogen-Activated Protein Kinases (MAPK) and serine/threonine kinases were phosphorylated by gal-1. In addition, gal-1 in trophoblast cells in vitro induced syncytium formation especially after concentration dependent stimulation of the cells with this galectin. This is in contrast to MAPK-inhibitor U0126 that reduced syncytium formation of BeWo cells. The aim of this study was to analyze the syncytium formation abilities of BeWo cells that were gal-1 silenced. We found a significantly reduced syncytium formation rate in gal-1 silenced BeWo cells. In addition, these cells show a different miRNA expression profile. In summary, we found that gal-1 is a major trigger for fusion processes in BeWo cells. This function is accompanied by different regulation of miRNA synthesis in the BeWo cell culture model.

Introduction The tandem-repeat type galectins, which comprise of gal-4, -6, -8, -9, and -12, form a sub-family of galectins. Gal-6 is expressed only in rodents, whereas the other four galectins, tandem-repeat galectins, are also detectable in human tissue. The placental expression of individual members of the tandem-repeat gal family is increasingly known, however, systematic, comparative analysis especially in the human placenta from normal or pathological pregnancies is still lacking. Material and Methods Within this study, third trimester placentas obtained at delivery (n = 14 IUGR, n = 15 controls, equally divided in placentas from male and female fetuses) were analyzed for the expression of gal-4, -8, -9 and -12 by immunohistology and immunofluorescence, data were obtained by using a semiquantitative scoring system. Double immune-fluorescence with trophoblast specific markers was used to identify co-expression in the decidua. Results We identified dysregulation of tandem repeat galectins in IUGR placentas with a strong connection to the fetal gender. We identified a significantly lower expression of gal-4 and gal-9 in villous trophoblast tissue of IUGR placentas with male fetuses and a downregulation of gal-4 and gal-8 in extravillous trophoblast (EVT) from IUGR and male fetuses. Conversely, expression of gal-9 and gal-12 was higher in EVT of IUGR cases in placentas with female fetuses. Double immunofluorescence using cytokeratin-7 confirmed the expression of tandem-repeat galectins in EVT. Discussion and Conclusion The human placenta expresses tandem-repeat type galectins in villous trophoblasts, EVT, endothelial cells and decidual stromal cells. Summarizing all effects, there is significant down-regulation of gal-4, -8 and gal-9 in the IUGR trophoblast of male fetuses. In contrast, in IUGR pregnancies with female fetus gal-9 and gal-12 are upregulated in the EVT and in endothelial cells in the cases of gal-12. Therefore we propose a fetal-gender specific action of tandem repeat galectins in IUGR placentas.

Altered glycosylation is a predominant feature of tumour cells; it serves for cell adhesion and detachment, respectively, and facilitates the immune escape of these cells. Therefore changes in the expression of glycosyltransferase genes could help to identify circulating tumour cells (CTCs) in the blood samples of cancer patients using a quantitative polymerase chain reaction (PCR) approach. Blood samples of healthy donors were inoculated with certain numbers of established breast cancer cell line cells, thus creating a model system. These samples were analysed by quantitative PCR for the expression of six different glycosyltransferase genes. The three genes with the best results in the model system were consecutively applied to samples from adjuvant breast cancer patients and of healthy donors. FUT3 and GALNT6 showed the highest increase in relative expression, while GALNT6 and ST3GAL3 were the first to reach statistically significant different ∆CT-values comparing the sample with and without addition of tumour cells. These three genes were applied to patient samples, but did not show any significant results that may suggest the presence of CTCs in the blood. Although the relative expression of some of the glycosyltransferase genes exhibited reasonable results in the model system, their application to breast cancer patient samples will have to be further improved, e.g. by co-analysis of patient blood samples by gold-standard methods.

Breast cancer is still the most frequent cause of cancer-related death in women worldwide. Often death is not caused only by the primary tumour itself, but also by metastatic lesions. Today it is largely accepted, that these remote metastases arise out of cells, which detach from the primary tumour, enter circulation, settle down at secondary sites in the body and are called Circulating Tumour Cells (CTCs). The occurrence of such minimal residual diseases in the blood of breast cancer patients is mostly linked to a worse prognosis for therapy outcome and overall survival. Due to their very low frequency, the detection of CTCs is, still a technical challenge. RT-qPCR as a highly sensitive method could be an approach for CTC-detection from peripheral blood of breast cancer patients. This assumption is based on the fact that CTCs are of epithelial origin and therefore express a different gene panel than surrounding blood cells. For the technical approach it is necessary to identify appropriate marker genes and to correlate their gene expression levels to the number of tumour cells within a sample in an in vitro approach. After that, samples from adjuvant and metastatic patients can be analysed. This approach may lead to new concepts in diagnosis and treatment.

Aim: Disseminated tumor cells are found in the bone marrow of patients with epithelial carcinoma and are correlated with a poor prognosis of the disease. Their detection is a technical challenge. This report describes a model system for the detection of cancer cells by co-immunostaining of Thomsen–Friedenreich and Her-2 antigens. Methods & results: Small numbers of cancer cells from different cancer cell lines were mixed with blood samples of healthy donors. Cytospins were prepared and double immunostaining against Thomsen–Friedenreich antigen and Her-2 was carried out by fluorochrome-coupled antibodies. Quantification of Thomsen–Friedenreich and/or Her-2-positive cells was performed with an epifluorescence microscope. On average, 83% of cancer cells were recovered by this method. Conclusion: Immunostaining is a useful method for the detection of cancer cells in blood samples. Results of this model system will be transferred to bone marrow patient samples to prove the benefits for detection of disseminated tumor cells.

The liver X receptors (LXRs) have been shown to be crucially involved in maternal-fetal cholesterol transport and placentation. The aim of this study was to investigate the expression pattern and frequency of LXR under normal physiological circumstances and in spontaneous abortion and/or recurrent miscarriage. A total of 29 (12 physiologic pregnancies/10 spontaneous abortions/7 recurrent miscarriages) human pregnancies in first trimester were analysed for LXR expression. Expression changes were evaluated by immunohistochemistry for receptor and quantitative RT-PCR (TaqMan) was performed to determine the level of LXR mRNA expression. We also stained for RXRαand PPARγas possible heterodimers of LXR. LXR expression was downregulated in the syncytiotrophoblast of spontaneous abortion placentas compared to normal pregnancy. In recurrent miscarriage there was a trend for a downregulation. Decidua showed an even stronger downregulation in both groups. In the syncytiotrophoblast we found a positive correlation for the combination of LXR/PPARγin abortions and a negative correlation for LXR/RXRα. In addition, double-immunofluorescence staining showed that LXR as well as RXRαand PPARγare expressed by the extravillous trophoblast. Finally, RXRαand LXR showed coexpression in the same extravillous trophoblast cells. To conclude, our data show that LXR expression is decreased in miscarriage.