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The process of embryo attachment and invasion through the endometrial epithelial cells and subsequent implantation into the decidualized endometrial stroma is the groundbreaking step for the establishment of a successful pregnancy. Necessary prerequisites are a receptive endometrium, a good-quality embryo and a well-orchestrated molecular dialog between embryo and maternal endometrium. The embryo-maternal dialog is conducted via a wide scope of factors, including secreted cytokines, chemokines, and growth factors in addition to the expression of corresponding receptors and co-receptors. Several embryonic proteins, including the aforementioned, are involved in the process of apoptosis, which necessarily needs to take place at the maternal endometrium to allow the embryo to invade. The endometrial epithelium is thereby disintegrated completely within a particular area, whereas the endometrial stroma seems to require a more depth-limited apoptosis. As of today, the exact mechanisms and factors mediating the apoptotic process involved in those apparently differently regulated incidents are not fully understood, particularly with regard to stromal cell apoptosis. There is evidence though, that cytokines and their respective receptors play a major role. A suggested important co-receptor for cytokines, which is highly upregulated in the receptive human endometrium, is the heparan sulfate proteoglycan syndecan-1. It is present on the cell surface and involved in the regulation of cell-cell-interaction, cell binding, cell signaling and cytoskeletal organization and therefore represents a possible mediator of apoptosis regulation in human endometrium. Herein, the literature on endometrial epithelial and stromal apoptosis in general, and in light of the influence of syndecan-1, is reviewed.

BACKGROUND Preeclampsia (PE) is a hypertensive disorder during pregnancy with endotheliosis leading to occlusion of renal perfusion with an impact on the glomerular filtration barrier. We therefore analyzed the role of intrarenal resistance indices in the renal interlobular arteries measured by Doppler ultrasound as a diagnosis of PE. METHODS Women with preeclampsia (n = 24; mean blood pressure/24 h = 145 ± 11/ 93 ± 7 mm Hg; mean proteinuria = 5.63 ± 1.0 g/24 h) were compared against a group of healthy pregnant women (n = 24). All patients underwent a Doppler ultrasound of the intrarenal arteries between the 24th week of gestation and the 5th week postpartum. Several risk factors for PE, as well as the arterial resistive indices of the Arteriae uterinae and the Arteria umbilicalis, were monitored in parallel. RESULTS The intrarenal resistive index (mean ± SD = 0.63 ± 0.05 in women with preeclampsia vs. 0.59 ± 0.056 in healthy pregnant women; P < 0.003), the pulsatile index (mean ± SD = 1.15 ± 0.19 in women with preeclampsia vs. 0.92 ± 0.13 in healthy pregnant women; P < 0.0001), and the end diastolic flow velocity (mean ± SD = 14.16 ± 4.75 cm/s in women with preeclampsia vs. 10.67 ± 2.68 cm/s in healthy pregnant women, P < 0.006) were elevated in patients with PE, as were the arterial resistive indices of the Aa. uterinae and A. umbilicalis. The intrarenal resistive indices correctly classified 84.2% of the women as having PE. CONCLUSIONS Intrarenal resistive indices are a significant classifier of PE, providing the possibility to predict nephropathy. They could be a prognostic tool for cardiovascular comorbidity in PE patients even after delivery.

Early molecular interaction between embryo and mother, involving chemoattractants, especially chemokine CXC-motif ligand 1 (CXCL1)(2), determines the pregnancy outcome. So far nothing is known about the signalling cascades of CXCL1 expression in human decidua. The aim of the study was to identify signalling cascades mediating the CXCL1 expression in human decidua incubated with IL-1β as a major secretion product of the embryo. Therefore, decidualised endometrial stromal cells were incubated with IL-1β in a concentration- and time-dependent manner. The specificity of the IL-1β induced CXCL1 expression was verified by application of the IL-1 receptor antagonist, which opposed the binding of IL-1β to its receptor, leading to a dose dependent diminished to complete CXCL1 elimination. IL-1β signalling was investigated using inhibitors for MAPK, STAT3 and JNKinase cascades. The CXCL1 secretion of decidualised endometrial cells was measured by ELISA. The MAPK signalling cascade was explored by western blot analysis of ERK and NFκB p65(3) as well as phospho ERK and pp65 activation by IL-1β in detail. A statistical significant increase in CXCL1 mRNA- and protein-expression after incubation with 0.1ng/ml IL-1β after 48h was detected. CXCL1 protein secretion could be completely prevented by IL-1 receptor antagonist treatment. Only inhibition of the MAPKinase pathway resulted in a statistically significant decrease of CXCL1 protein secretion. Initiation of the MAPK pathway depicted by phospho ERK activation started as early as 2min after coincubation of decidualised endometrial stromal cells with IL-1β. Activation of NFκB p65 could be measured within 15min of IL-1β incubation in decidualised endometrial stromal cells. CXCL1 is a target for the embryos' secretion product IL-1β in decidualised endometrial stromal cells during the peri-implantation period. IL-1β's rapid effect on CXCL1 synthesis is uniquely mediated via the MAPK-signalling cascade and the activation of CXCL1s' transcription factor NFκB p65.

Viele Frauen, die die Familienplanung immer weiter hinausschieben, verdrangen die Tatsache, dass ihre Fertilitat etwa ab dem 35. Lebensjahr drastisch sinkt. Was heute uber die „Biologie des reproduktiven Alters“ bekannt ist und mit welchen Methoden Frauen im kritischen Alter doch noch zu einem eigenen Kind verholfen werden kann, ist Thema des folgenden Beitrags.

The aim of this study was to estimate the rate of intrauterine adhesions and subsequent pregnancy outcome in patients with residual trophoblastic tissue treated with hysteroscopic resection versus ultrasound-guided dilation and evacuation (DE).Cohort study from 2 centers (Canadian Task Force classification II-2).Two surgical teams at the University of Duesseldorf Medical Center and the PAN Clinic in Cologne, Germany.Women with residual trophoblastic tissue after first- or second-trimester miscarriage or term delivery.Two techniques were used for the removal of residual trophoblastic tissue: ultrasound-guided evacuation with a curette (DE) and hysteroscopic resection of trophoblastic tissue (HR).We evaluated 95 patients who underwent secondary intervention for residual trophoblastic disease. A total of 42 patients underwent dilation of the cervix and ultrasound-guided curettage. In a second series of 53 patients, a resectoscope fitted with a 4-mm cutting loop was used for the removal of residual trophoblastic tissue used without application of current. Three months after the intervention, second-look office hysteroscopy was performed. Differences between both treatment groups were statistically significant. After HR, mild intrauterine adhesions were found in 2 patients (4.2%). After DE, 12 patients (30.8%) presented with intrauterine adhesions (mild intrauterine adhesions: n = 7 [17.9%]; single dense adhesions: n = 3 [7.7%]; and extensive endometrial fibrosis n = 1 [2.6%]). Eighty-two patients wanted to become pregnant. Conception rate of all patients examined was 68.8% (HR) and 59.9% (DE) (p.05). In patients younger than 35 years of age who underwent HR, the pregnancy rate was significantly (p.05) increased compared with patients who underwent DE (78.1% vs 66.6%). In addition, patients from the HR group demonstrated a significantly (p.05) shorter time to conception (11.5 month vs 14.5 month).The results of this study indicate that selective HR of residual trophoblastic tissue significantly reduces the incidence of intrauterine adhesions and increases pregnancy rates.

Objective Multidrug resistance gene 1 (MDR1) mediated resistance to chemotherapeutic agents is a major obstacle for the therapy of various cancer types. The use of conditionally replicating adenoviruses (CRAds) is dependent on molecular differences between tumor cells and non tumor cells. Transcriptional targeting of CRAd replication is an effective way to control replication regulation. The aim of this study was to evaluate the effect of a MDR1 targeted fiber-modified CRAd against chemotherapy resistant ovarian cancer. Methods MDR1 expression was evaluated in chemotherapy naive and pretreated ovarian cancer cells and various control cells. We constructed 2 variants of a fiber-modified CRAd, Ad5/3MDR1E1 and Ad5/3MDR1E1∆24 containing the MDR1 promoter to control viral replication via the E1A gene. The MDR promoter activity and cell killing efficacy were evaluated in vitro. Orthotopic murine models of peritoneally disseminated ovarian cancer were utilized to evaluate the preclinical efficacy of MDR targeted CRAds in vivo. To evaluate the liver toxicity of MDR1 targeted CRAds, we compared Ad5/3MDR1E1 with Ad5/3∆24, a CRAd that replicates in cancer cells inactive in the Rb/p16 pathway by use of an in vivo hepatotoxicity model. Results We demonstrate efficient oncolysis of Ad5/3MDR1E1 in both chemotherapy resistant ovarian cancer cell lines and in primary tumor cells from pretreated patients as well as therapeutic efficacy in an orthotopic mouse model. Ad5/3MDR1E1 demonstrated significantly decreased liver toxicity compared to other 5/3-fiber modified control vectors examined. Conclusions In summary, Ad5/3MDR1E1 is an efficient and safe gene therapy approach for specific targeting of chemotherapy resistant cancer cells.

Objectives Placental derived vasculogenic/angiogenic substances in maternal blood are dysregulated in pre-eclampsia. We hypothesized that CXCL12, a chemokine with vasculogenic actions, is amongst such molecules. Study design CXCL12, CXCL16, CXCR4, and CXCR6 immunolocalization in placental tissue was analyzed in pre-eclampsia (n = 8) in comparison to controls (n = 8). CXCL12, measured by ELISA in blood, in women diagnosed with pre-eclampsia (n = 14) and prior to the development of pre-eclampsia (at 20 weeks’ gestation, n = 20) was compared with CXCL12 concentrations in gestation-matched, healthy control subjects (n = 34). Results In placental tissue, syncytiotrophoblast staining for CXCL12 was increased in pre-eclampsia. Maternal serum CXCL12 was increased in pre-eclampsia [2000 (SD 402) vs 1484 (SD 261) pg/ml, P = 0.01] but not in plasma obtained at 20 weeks of gestation prior to the onset of pre-eclampsia [1183 (SD 336) vs 1036 (SD 144) pg/ml, P = 0.09]. Conclusion Our data suggest that the syncytiotrophoblast contributes to a pre-eclampsia-associated increase in CXCL12 levels in maternal blood. These findings support the hypothesis that an imbalance of angiogenic factors contributes to the pathogenesis of pre-eclampsia.

Fragestellung: Bei der Endometriose handelt es sich um eine benigne hormonabhangige Erkrankung der Frau im reproduktionsfahigen Alter. Ziel einer operativen Therapie ist die chirurgische Entfernung der Endometrioseherde. Medikamentose Therapieansatze zielen ab auf eine Suppression des Ostrogens auf ein postmenopausales Hormonniveau, was durch den Einsatz von Gonadotropin-Releasing-Hormon-(GnRH-)Analoga erreicht werden kann. Der therapeutische Nutzen einer systemischen Endometriosetherapie ist jedoch limitiert und mit signifikanten Nebenwirkungen vergesellschaftet. Aus diesem Grund besteht ein groser Bedarf an der Entwicklung einer spezifischen und zielgerichteten Therapie der Endometriose. Ziel dieser Untersuchung war die Identifikation geeigneter Zielstrukturen. Material und Methodik: Vergleichende Untersuchungen der Expression von VEGF, Midkine, Angiopoietin-2 und Heparanase erfolgten in Endometriose-Stroma und ‐drusenzellen, eutopem Endometrium und makroskopisch unauffalligen Peritonealgewebeproben mittels immunhistochemischer Untersuchungen und RT‐PCR. Anschliesend wurde die Expressionsstarke von VEGF und Heparanase in oberflachlichen peritonealen Endometrioseherden und tief infiltrierenden Endometrioseknoten immunhistochemisch untersucht. Zur Untersuchung des therapeutischen Einsatzes wurde ein VEGF-gesteuertes rekombinantes Adenovirus (AdVEGFE1) als Vektor eingesetzt. Untersucht wurden die spezifische Induktion von Apoptose in vitro sowie die Biodistribution in einem Mausmodell. Ergebnisse: VEGF und Heparanase werden in Endometriosezellen signifikant starker exprimiert als in eutopem Endometrium oder makroskopisch unauffalligen Peritonealgewebeproben. Weiterhin zeigen die korrespondierenden Promotoren beider Gene eine starke Expression in peritonealen Endometrioseherden. Eine Korrelation der Expressionsstarke mit dem Schweregrad der Erkrankung lies sich jedoch nur fur VEGF nachweisen. In Untersuchungen zur therapeutischen Anwendung eines VEGF-gesteuerten adenoviralen Vektors zeigt AdVEGFE1 in vitro eine Induktion von Apoptose in aufgereinigten kultivierten Endometriosezellen von Patientinnen. Das rekombinante konditional replizierende Adenovirus Ad5VEGFE1 zeigt eine effiziente Apoptoseinduktion in vitro und im Biodistribution-Mausmodell eine nur schwache Infektion des eutopen Endometriums und der Leber. Schlussfolgerung: VEGF ist eine geeignete Zielstruktur zur Entwicklung einer zielgerichteten Therapie der Endometriose.

Zwillingsschwangerschaften sind in Europa relativ haufig, Zwillingsgeburten kommen mit einer Frequenz von etwa 17 auf 1000 Geburten vor. Es existieren sowohl zeitlich als auch geographisch unterschiedliche Verteilungen. Der Grosteil der Zwillingsschwangerschaften ist dizygoten Ursprungs, der Anteil monozygoter Zwillingsschwangerschaften liegt bei etwa 4/1000 und ist sowohl zeitlich als auch geographisch konstant. Grunde fur die zunehmende Haufigkeit dizygoter Zwillingsschwangerschaften sind vor allem die Anwendung von Verfahren der assistierten Reproduktion und – bedingt durch steigende FSH-Spiegel und konsekutiv verursachte Polyovulationen – das zunehmende mutterliche Alter. Hohergradige Mehrlingsschwangerschaften entstehen hauptsachlich iatrogen, wobei die Rate an Drillingsschwangerschaften nach IVF oder ICSI in Deutschland erfreulicherweise rucklaufig ist. Eine weitere Reduktion der Mehrlingsraten nach IVF oder ICSI liese sich – wie Daten aus Skandinavien zeigen – durch die Einfuhrung eines elektiven „single-embryo-transfer“ erreichen. Hierfur ware aber eine Selektionsmoglichkeit in vitro notig, welche das Embryonenschutzgesetz derzeit ausschliest.

Despite advances in therapy, metastatic breast carcinoma maintains a disappointing 5-year survival rate of 20.4 %. Hence, there is a need for alternative and complementary therapeutic strategies. In this regard oncolytic virotherapy is a promising approach that uses mainly conditionally replicative viruses (CRAds) to selectively kill tumor cells by exploiting the lytic property of virus replication. Adenovirus-based vectors have emerged as leading candidates for in vivo virotherapy. However intravenously injected CRAds have shown only limited antitumor efficacy due mainly to low viral infectivity and insufficient viral delivery to the sites of tumor. Based on recent findings that systemically administered human mesenchymal stem cells (hMSCs) home to breast cancer metastasis of the lung, our investigation assesses the possibility of using hMSCs as cellular virus carriers to pulmonary metastases of breast cancer. HMSCs were transduced with CRAds. In a SCID mouse xenograft model we established breast cancer pulmonary metastatic disease to examine the effects of intravenously injected CRAd-loaded hMSCs versus CRAds alone. In mice treated with CRAd-loaded hMSCs the transduced stem cells homed to the metastatic lesions and led to reduced tumorgrowth and extended survival compared to mice treated with CRAds alone. We concluded that systemically administered hMSCs transduced with CRAds have the ability to home in on and kill metastatic breast cancer cells; these findings validate the feasibility of using hMSCs as cellular vehicles for the targeted therapy of metastatic breast cancer with CRAds.

Alternative and complementary therapeutic strategies need to be developed for metastatic breast cancer. Virotherapy is a novel therapeutic approach for the treatment of cancer in which the replicating virus itself is the anticancer agent. However, the success of virotherapy has been limited due to inefficient virus delivery to the tumor site. The present study addresses the utility of human mesenchymal stem cells (hMSCs) as intermediate carriers for conditionally replicating adenoviruses (CRAds) to target metastatic breast cancer in vivo. HMSC were transduced with CRAds. We used a SCID mouse xenograft model to examine the effects of systemically injected CRAd loaded hMSC or CRAd alone on the growth of MDA-MB-231 derived pulmonary metastases (experimental metastases model) in vivo and on overall survival. Intravenous injection of CRAd loaded hMSCs into mice with established MDA-MB-231 pulmonary metastatic disease homed to the tumor site and led to extended mouse survival compared to mice treated with CRAd alone. Injected hMSCs transduced with CRAds suppressed the growth of pulmonary metastases, presumably through viral amplification in the hMSCs. Thus, hMSCs may be an effective platform for the targeted delivery of CRAds to distant cancer sites such as metastatic breast cancer.

Background: While cyclosporine A (CsA) is an effective therapy for nephrotic syndrome, it has nephrotoxic side effects. We compared the anti-proteinuric effects and nephrotoxicity in rats with passive Heymann nephritis (PHN) of CsA and mycophenolate mofetil (MMF). Methods: PHN was induced in female Wistar rats. Two treatment groups consisting of 8 rats each received either 25 mg of CsA or 25 mg of MMF/kg body weight/day and were compared with untreated controls. Kidney function and proteinuria were monitored over 4 weeks. Western blots were used for densitometric analysis of renal cyclooxygenase-2 (COX-2) protein expression. Thromboxane B2 (TxB2) and 6-keto-PGF1α were determined by radioimmunoassays (RIAs) in renal tissue and urine. Results: Rats with PHN exhibited a marked proteinuria of 12.76 ± 4.42 vs. 0.73 ± 0.28 mg/24 h (p < 0.01) and showed increased glomerular concentrations of TxB2 and 6-keto-PGF1α (992.6 ± 216.9 and 1,187.0 ± 54.2 pg/mg protein, respectively) compared with healthy controls (595 ± 196.17 and 729 ± 297.84, respectively) and a strongly induced COX-2 protein expression. CsA and MMF treatment reduced PHN-related proteinuria to 2.10 ± 1.47 and 1.47 ± 7.2 mg/24 h, respectively. In rats with PHN, CsA induced a significant deterioration of renal function and enhanced urine excretion of thromboxane A2, paralleled by a significant, twofold increase in COX-2 protein expression and renal prostaglandins. By contrast, MMF treatment in rats with PHN was not nephrotoxic and had no effect on prostaglandin production. COX-2 protein expression under MMF was suppressed. Conclusion: While the antiproteinuric efficacy of MMF and CsA in PHN was comparable, the absence of nephrotoxicity might favor MMF in the treatment of nephrotic syndrome. The CsA-induced increase in COX-2 expression and COX-2-dependent prostacyclin may indicate a mechanism that compensates nephrotoxicity in the diseased and CsA-exposed kidney.

Angiogenesis, the growth of new capillaries from pre-existing blood vessels, is a physiological process involved in both normal menstrual cycling and implantation of the embryo. So far, very little is known about the expression of angiopoietins, growth factors involved in angiogenesis, in human endometrium. Both angiopoietin-1 (Ang-1) and angiopoietin-2 (Ang-2) are ligands for the endothelial cell-specific receptor tyrosine kinase Tie-2. In this study we determined the mRNA expression of Ang-1, Ang-2 and Tie-2 by quantitative competitive RT/(QC)-PCR (including specifically designed competitor cDNA) in biopsied human endometrium throughout the menstrual cycle. We detected the mRNA for the angiopoietins in 30 out of 32 endometrial biopsies (94%), covering early proliferative (n = 4), mid proliferative (n = 12), late proliferative (n = 3), early secretory (n = 3), mid secretory (n = 5) and late secretory (n = 3) phases. Analysis of the target/competitor ratios (QC-PCR) revealed that Ang-1 mRNA expression was significantly up-regulated (P = 0.027) during the secretory phase of the menstrual cycle. In contrast, the expression levels of both Ang-2 mRNA and Tie-2 mRNA showed only minor variations at different cycle stages. These findings were confirmed by the relative expression ratio of Ang-1 versus Ang-2 in a multiplex PCR. The expression of Ang-1, Ang-2 and Tie-2 mRNA was detected in both isolated endometrial epithelial and stromal cell fractions. Immunohistochemical localization of the proteins revealed qualitative differences in both cell type and cycle stage expression. In conclusion, the enhanced Ang-1 expression during the secretory phase might serve to stabilize the newly developed blood vessels.

Human chorionic gonadotropin (hCG) has long been associated with the initiation and maintenance of pregnancy, where angiogenesis plays an important role. However, the function of hCG in angiogenesis and the recruitment of vascular active cells are not fully understood. In this study, the role of hCG and its receptor in circulating angiogenic and human endothelial cells, including lymphatic, uterine microvascular, and umbilical vein endothelial cells, was examined. Immunohistochemistry and immunoblot analysis were used to detect LH/hCG receptor expression and the expression of hCG-induced angiogenic molecules. HIF-1α was determined via ELISA and downstream molecules, such as CXCL12 and CXCR4, via real-time PCR. Chemotaxis was analyzed using Boyden chambers. Our results show that the LH/hCG receptor was present in all tested cells. Furthermore, hCG was able to stimulate LH/hCG-receptor-specific migration in a dose-dependent fashion and induce key angiogenic molecules, including HIF-1α, CXCL12, and CXCR4. In conclusion, our findings underscore the importance of hCG as one of the first angiogenic molecules produced by the conceptus. hCG itself alters endothelial motility, recruitment, and expression of pro-angiogenic molecules and may therefore play an important role in vascular adaption during implantation and early placental formation.

Introduction Placental development involves the variation of oxygen supply due to vascular changes and cytotrophoblast invasion. Chemokines and their receptors play an important role during placental formation. Herein, the analysis of the chemokine/receptor pair CXCL12/CXCR4 and further chemokine receptors, such as CCR1, CCR7 and CXCR6 expression in human cytotrophoblasts was conducted. Methods Human cytotrophoblasts were examined directly after isolation or after incubation with different oxygen tensions and a chemical HIF-stimulator for 12 h with realtime PCR, immunoblot, immunohistochemistry. Conditioned media of placental villi, decidua, and endothelial cells was used for ELISA analysis of CXL12. Cytotrophoblast migration assays were conducted applying conditioned media of endothelial cells, a CXCL12 gradient, and different oxygen level. Endometrial and decidual tissue was stained for CXCL12 expression. Results An upregulation of CXCL12, CXCR4, CCR1, CCR7 and CXCR6 was observed after cytotrophoblast differentiation. Low oxygen supply upregulated CXCR4, CCR7 and CXCR6, but downregulated CXCL12 and CCR1. In contrast to the HIF associated upregulation of the aforementioned proteins, downregulation of CXCL12 and CCR1 seemed to be HIF independent. Cytotrophoblast migration was stimulated by low oxygen, the application of a CXCL12 gradient and endothelial cell conditioned media. CXCL12 was detected in endometrial vessels, glands and conditioned media of placental and decidual tissue, but not decidual vessels. Discussion/conclusion Taken together, oxygen supply and cytotrophoblast differentiation seem to be regulators of chemokine and receptor expression and function in human cytotrophoblasts. Therefore, this system seems to be involved in placental development, directed cytotrophoblast migration in the decidual compartment and a subsequent sufficient supply of the growing fetus.

Endometrial epithelial cells are known to undergo apoptosis during trophoblast invasion. We postulate that the cell surface molecule Syndecan-1 which is expressed on endometrial cells and syncytiotrophoblast is important for implantation in general and especially for induction of maternal cell apoptosis during trophoblast invasion because Syndecan-1's influence on apoptotic susceptibility of cancer cells is already described in the literature. Using the human endometrial epithelial cell line RL95-2, a new stable cell line with Syndecan-1 knockdown was generated. Via antibody array analysis, a significant decrease in the expression of anti-apoptotic proteins like inhibitors of apoptosis, Clusterin, heme oxygenase (HO-2), heat shock protein (HSP)27 and -70 and Survivin due to the Syndecan-1 knockdown was discovered. Correspondingly, active Caspase-3 as an indicator for apoptosis was increased more severely in these cells compared with unmodified RL95-2 after treatment with implantation-related stimuli, which are the cytokines interleukin-1β, interferon-γ, tumor necrosis factor-α and transforming growth factor-β1 and an anti-Fas antibody. Furthermore, a treatment with a combination of all factors caused a higher Caspase-3 induction compared with each single treatment. These results demonstrate that Syndecan-1 is involved in the control of apoptosis in RL95-2 cells and therefore may affect the fine tuning of apoptosis in endometrial epithelium regulating the embryo's invasion depth as a crucial step for regular implantation followed by successful pregnancy.

Purpose In early pregnancy the dialogue between maternal endometrium and embryo is a key process in establishing a receptive decidua and placental network. Decidual ISG15 induction is thought to promote pregnancy maintenance and development. ISG15 is involved in RNA splicing, cytoskeletal organization, stress response and further intracellular processes. Methods ISG15 expression was examined immunohistologically in paraffin-embedded human placental and decidual tissue samples of all pregnancy trimesters on adjacent sections (first trimester n = 5, second n = 5, third n = 3). Samples were processed using a protocol applying a rabbit polyclonal ISG15 antibody. A mouse monoclonal cytokeratin seven antibody was utilized to identify the different placental departments and decidual glands. Staining results and anatomical features were evaluated blindly with strict rating criteria. Results ISG15 expression was identified in first and second trimester tissue samples. ISG15 localized especially to the extravillous cytotrophoblasts in the maternal wall and in maternal blood vessel. Expression was detected in cytotrophoblast progenitor cells in the placental villi and the cell column with a maximum in the first trimester. The syncytial layer stained positive in first and second trimester samples. Third trimester samples showed no expression of ISG15 at all. Conclusions ISG15 abundance in the human placenta is an interesting finding, with implications for placental development, fetal growth and potential defense mechanism against infections. The maximal expression of ISG15 in the first and second trimester of pregnancy suggests that ISG function is needed when placental and embryo development is enormous and embryo susceptibility to external influences is high.

Background. The aim of this paper was to determine the influence of differentin vitroculture media on mRNA expression of Hedgehog genes,il-6,and important genes regarding reactive oxygen species in single mouse embryos.Methods. Reverse transcription of single embryos either culturedin vitrofrom day 0.5 until 3.5 (COOK’s Cleavage medium or Vitrolife’s G-1 PLUS medium) orin vivountil day 3.5post coitum. PCR was carried out forβ-actinfollowed by nested-PCR forshh, ihh, il-6, nox, gpx4, gpx1,andprdx2.Results. The number of murine blastocysts cultured in COOK medium which expressedil-6, gpx4, gpx1,andprdx2mRNA differed significantly compared to thein vivogroup. Except fornox, the mRNA profile of the Vitrolife media group embryos varied significantly from thein vivoones regarding the number of blastocysts expressing the mRNA ofshh, ihh, il-6, gpx4, gpx1andprdx2.Conclusions. The present study shows that differentin vitroculture media lead to different mRNA expression profiles during early development. Even the newly developedin vitroculture media are not able to mimic the female reproductive tract. The question of long-term consequences for children due to assisted reproduction techniques needs to be addressed in larger studies.

Fragestellung: Die rupturierte Extrauteringraviditat (EUG) ist nach wie vor eine der haufigsten Todesursachen junger Frauen. Fruhere Entzundungen des Unterleibs als Hauptursache fur die Entstehung ektoper Schwangerschaften pradisponieren uber postinfektiose, mechanische Hindernisse im Eileiter. In jungerer Zeit wird dazu noch evident, dass auch eine Kinderwunschbehandlung im Sinne einer in-vitro Fertilisation (IVF) das Risiko fur eine EUG erhoht. Entzundungsmediatoren wie CXCL8 (IL-8) lassen sich beim Menschen durch proinflammatorische Zytokine wie IL-1β induzieren, das nicht nur einen Infektionsmarker, sondern auch ein Hauptsekretionsprodukt des Embryos darstellt. Unsere Arbeitsgruppe konnte bereits zeigen, dass im menschlichen Eileiter der Sekretionsphase die Expression von CXCL8 im Gegensatz zur Dezidua herunterreguliert ist (Hess et al., 2011, in Begutachtung). Wir postulieren daher einen direkten hormonellen Einfluss der Chemokinexpression auf den regelrechten Transport und die Implantation des Embryos und vermuten daruber hinaus eine hormonelle Komponente bei der Entstehung der EUG. Da menschliche Eileiter fur Forschungszwecke kaum zur Verfugung stehen, ist es unser Ziel, in primaren Eileiter-Epithelzellen des Schweins die biochemischen Bedingungen einer EUG nachzuahmen und in diesem System mogliche hormonelle Auswirkungen einer IVF- Behandlung auf den Eileiter zu untersuchen. Methodik: Primare porcine Eileiter-Epithelzellen wurden aus Schlachthofmaterial isoliert und uber 5 Tage in Ham's F12-Medium kultiviert. Die Epithelzellen wurden nach Vorbehandlung mit einer im IVF-Verfahren ublichen Estradiolkonzentration von 3000 pg/mL uber 48 Stunden mit porcinem IL-1β inkubiert und anschliesend der Kulturuberstand gewonnen. Analog hierzu wurde mit hormonfrei kultivierten Kontrollen verfahren. Die CXCL8-Konzentration im Uberstand wurde durch einen kommerziell erhaltlichen ELISA quantifiziert. Ergebnisse: CXCL8 lies sich sowohl auf mRNA-Ebene als auch immunhistochemisch in den porcinen Eileiter-Epithelzellen nachweisen. Fur das sezernierte Chemokin konnte daruber hinaus eine signifikant gesteigerte Expression nach Inkubation mit IL-1β im Zellkulturuberstand mittels ELISA detektiert werden. Die Vorinkubation mit einer erhohten Estradiolkonzentration ergab jedoch keine weitere Steigerung der CXCL8-Expression gegenuber den hormonfrei kultivierten Kontrollen. Schlussfolgerung: Die signifikante Expressionssteigerung von CXCL8 in Eileiter-Epithelzellen des Schweins nach Inkubation mit IL-1β weist auf eine direkte Reaktion des Epithels auf die Anwesenheit des embryonalen Surrogatmarkers hin. Da die erhohte Estradiolkonzentration im Medium keine Auswirkung auf die CXCL8-Expression hatte, sind daruber hinaus weitere Versuche erforderlich, um die Ursachen einer erhohten Rate ektoper Schwangerschaften nach IVF-Behandlungen zu ergrunden. Die gute Verfugbarkeit von Eileitern des Schweins und der vergleichbare Sexualzyklus von Mensch und Schwein machen die primare Zellkultur von porcinem Eileiter-Epithel jedoch zu einem wertvollen Modell zur Erforschung tubarer Pathologien.

The placenta forms the interface between the mother and the fetus. During placental development cytotrophoblasts differentiate to form the syncytium or to invade the decidual wall to breach maternal vessels and establish the blood flow in the intervillous space. This process is still not well understood but it is proposed that chemokines and their receptors are involved in guiding cytotrophoblasts to the decidua and maternal vessels as well as attracting immunocompetent cells to the implantation site. CXCL12 is a chemokine expressed by cytotrophoblasts and is involved in cytotrophoblast invasion, differentiation and survival. One of its receptors, CXCR4, has been detected on cytotrophoblasts. Recent data show that CXCR7 and syndecan-4 might partially mediate CXCL12 function in other cell types. In this study, we examined CXCR7 and syndecan-4 expression at the maternal-fetal interface via immmunolocalization in placental tissue sections and in isolated cytotrophoblasts. We further used immunoblot analyses to confirm the data. We were able to show that cytotrophoblasts express both receptors and that upregulation occurs during the differentiation process of cytotrophoblasts towards the invasive phenotype. On a functional level CXCR7 seems not to be involved in JAR cell chemotaxis, suggesting a different function of this receptor. In conclusion, we propose that CXCL12 binds to CXCR4, but also to CXCR7 and syndecan-4. These three receptors could mediate different functions of CXCL12, such as cell migration, directed invasion, proliferation and survival. The latter molecules might also be involved in the development of placental pathologies, such as preeclampsia or choriocarcinoma growth.

A successful implantation of a mammalian embryo into the maternal endometrium depends on a highly synchronized fetal–maternal dialogue involving chemokines, growth factors, and matrix-modifying enzymes. A growing body of evidence suggests an important role for proteinases playing a role in matrix degeneration and enhancing the embryo's invasive capacity and influencing the mother's immunological status in favor of the conceptus. This study focused on the expression of cathepsin S (CTSS) and its inhibitors in the murine fetal–maternal interface as well as the detection of the cellular sources of either proteinase and inhibitors. Nested RT-PCR for detection of embryonic mRNAs, immunohistochemistry of maternal and fetal tissues in B6C3F1 mice, and FACS analysis for determination of immunocompetent cell population were applied. This study shows that the cysteine proteinase CTSS is upregulated in the stroma of the implantation site, and that pregnancy induces an influx of CTSS-positive uterine natural killer cells. Compared to maternal tissues, the CTSS inhibitors cystatin F and C, but not the proteinase itself, are expressed in blastocysts. In conclusion, CTSS underlies a hormonal regulation in the maternal tissue and therewith most likely supports the embryonic implantation. The invading embryo regulates the depth of its own invasion through the expression of the cathepsin inhibitors and furthermore, interleukin-6 to activate CTSS in maternal tissues. Additionally, the observed decrease in CD3+cells leads to the hypothesis that cells of the cytotoxic T-cell group are down-regulated in the decidua to support the implantation and ensure the survival of the embryo.

Objective To report the case of a Nigerian patient who suffered from sterility and underwent abdominal laparotomy to remove an adnexal tumor. Histologic analysis revealed Schistosoma haematobium ova. Design Case report. Setting Multidisciplinary group practice and teaching hospital. Patients One patient who underwent an abdominal laparotomy to remove an adnexal tumor. Intervention(s) In vitro fertilization and adnexal tumor resection via laparotomy. Main Outcome Measure(s) Treatment of the adnexal tumor affecting the reproductive health. Result(s) Histologic analysis performed on the tissue removed at surgery revealed fibrous patches around aggregates of calcified Schistosoma haematobium eggs in the fallopian tube wall. Conclusion(s) Schistosomiasis needs to be considered as a differential diagnosis of female infertility and sterility.

Based on the reported tocolytic action of the hormone relaxin (RLX) in rodents, locally produced in reproductive tissues and the corpus luteum in mammals, the present study aimed to evaluate the influence of RLX on contraction-mediating cyclooxygenases-1 and -2 (COX) and the contractile prostaglandin PGE(2) in human myometrial and decidual cells. Primary cultured cells were obtained from uteri and placentas of term and preterm women undergoing elective caesarean section.In vitro culture of primary myometrial and decidual cells, immunocytochemistry, reverse transcription and real-time PCR, Western blot, ELISA.We demonstrate for the first time an activating effect of RLX for human COX-1 and COX-2 in primary myometrial and decidual cells in vitro.These effects might potentially contribute to birth-associated induction of contractions in vivo.

Angiogenesis is a key process in the endometrium which undergoes dramatic changes during the menstrual cycle. Molecules such as vascular endothelial growth factor (VEGF), acting via two tyrosine kinase family receptors (VEGFR1 [Flt-1] and VEGFR2 [KDR/Flk-1]), are potent modulators of angiogenesis and vascular remodelling in the endometrium. Recently, neuropilin-1 (NRP-1) was shown to be expressed in endothelial cells binding VEGF(165) and therewith enhancing the binding of VEGF(165) to VEGFR2. This suggests that NRP-1, in addition to the known VEGF receptors, may play an important role in VEGF-induced angiogenesis. In this study, the expression of NRP-1 in the cycling human endometrium has been investigated by reverse transcription (RT)-polymerase chain reaction (RT-PCR), semi-quantitative competitive RT-PCR (RT-cPCR) and immunohistochemical staining. NRP-1 was expressed in all 32 endometrium samples throughout the menstrual cycle. However, samples from the proliferative phase showed significantly higher expression levels of NRP-1 mRNA compared to samples from the secretory phase (t/c-ratio 2.13 vs. 0.84, p0.05). Immunohistochemistry confirmed the results showing increased NRP-1 staining in vascular endothelium, glandular epithelium and stromal cells of the proliferative phase endometrium. This study demonstrates mRNA and protein expression of NRP-1 in human endometrium samples throughout the menstrual cycle. The enhanced expression of NRP-1 in the proliferative phase suggests that it may participate in hormonally regulated changes of endometrial angiogenesis, preparing the endometrium for the implantation of an embryo. NRP-1 expression might act as a co-factor for VEGF(165) enhancing the angiogenic stimulus.

Infertility and pregnancy wastage affect one of every nine couples in Western Europe and in the United States. The molecular events of embryonic attachment to the endometrial epithelium and subsequent invasion and nidation into the stroma have long been of interest, scientifically to reproductive biologists and clinically to couples with infertility or habitual abortion and to the physicians caring for them. In order to achieve a successful pregnancy in the human, two major conditions have to be fulfilled: during the 4–5 days of transport through the fallopian tube, the embryo must undergo a series of complex maturation processes and, in the same time, a receptive endometrium must have developed. Human endometrium undergoes characteristic cyclic changes of proliferation and secretion and, without embryonic implantation, the endometrium is shed and the menstrual bleeding occurs. Uterine endometrium therefore is the anatomic prerequisite for the continuation of our species and its main purpose during the reproductive age is to communicate with, receive, nourish and protect the implanting blastocyst.1

International audience; The human fallopian tube provides the environment for the first 5 days of embryonic development . The IL-1 system is involved in human embryo implantation. This study aimed to investigate IL-1β, IL-1ra and IL-1R tI expression within the length of the human fallopian tube on mRNA- and protein-level in samples from proliferative versus secretory phase, postmenopause samples (PMP) and samples from intra- (IUP) and extrauterine (EUP) pregnancies to examine possible spatial and hormonal induced changes (fimbrial, ampullary and isthmic tube segments). On mRNA-level, IL-1β was expressed in all samples except in PMP. IL-1R tI could be detected in all samples whereas IL-1ra was only expressed in secretory phase and the IUP sample. Immunohistochemically we could detect IL-1β and IL-1R t1 protein in all proliferative and secretory phase samples with maximum intensity in secretory phase samples whereas IL-1ra was expressed in secretory phase samples only. Overall no spatial but temporal differences possibly due to hormonal changes could be observed suggesting a precise regulation of the IL-1 system, especially for IL-1ra and moreover a stable molecular architecture within the full length of the fallopian tube.

Human embryo invasion and implantation into the inner wall of the maternal uterus, the endometrium, is the pivotal process for a successful pregnancy. Whereas disruption of the endometrial epithelial layer was already correlated with the programmed cell death, the role of apoptosis of the subjacent endometrial stromal cells during implantation is indistinct. The aim was to clarify whether apoptosis plays a role in the stromal invasion and to characterize if the apoptotic susceptibility of endometrial stromal cells to embryonic stimuli is influenced by decidualization and Syndecan-1. Therefore, the immortalized human endometrial stromal cell line St-T1 was used to first generate a new cell line with a stable Syndecan-1 knock down (KdS1), and second to further decidualize the cells with progesterone. As a replacement for the ethically inapplicable embryo all cells were treated with the embryonic factors and secretion products interleukin-1β, interferon-γ, tumor necrosis factor-α, transforming growth factor-β1 and anti-Fas antibody to mimic the embryo contact. Detection of apoptosis was verified via Caspase ELISAs, PARP cleavage and Annexin V staining. Apoptosis-related proteins were investigated via antibody arrays and underlying signaling pathways were analyzed by Western blot. Non-decidualized endometrial stromal cells showed a resistance towards apoptosis which was rescinded by decidualization and Syndecan-1 knock down independent of decidualization. This was correlated with an altered expression of several pro- and anti-apoptotic proteins and connected to a higher activation of pro-survival Akt in non-differentiated St-T1 as an upstream mediator of apoptotis-related proteins. This study provides insight into the largely elusive process of implantation, proposing an important role for stromal cell apoptosis to successfully establish a pregnancy. The impact of Syndecan-1 in attenuating the apoptotic signal is particularly interesting in the light of an already described influence on pregnancy disorders and therefore might provide a useful clinical tool in the future to prevent pregnancy complications provoked by inadequate implantation.

The oviduct and endometrium in primates have common embryologic origins. They are both highly sensitive to changes in circulating ovarian-derived steroid hormones, namely, estradiol (E 2 ) and progesterone, and their downstream effectors, although the cyclicity of the two organs is distinct. In the oviduct, remarkable changes occur in epithelial ciliogenesis and secretory cell development primarily in response to increasing levels of estradiol in the proliferative phase of the menstrual cycle. In the peri- and postovulatory period, under the influence of progesterone, the oviductal epithelium atrophies, regresses, and becomes quiescent, primarily through cellular apoptosis. The endometrium also undergoes intense cellular proliferation under the influence of estradiol. However, in contrast to the oviduct, after ovulation and under the influence of progesterone, it does not regress. Rather, it undergoes morphological and biochemical changes, preparing for implantation of the conceptus. The multitude of functions of the oviduct and the endometrium and the numerous processes and players are profound. The primates have been able to reproduce with a minimum difficulty and complications, although infections leading to tubal and endometrial damage are a testimony to the fragility of the reproductive process and its success.