I. Classen-Linke

The human placenta: a model for tenascin expression

SummaryTenascin is a large glycoprotein of the extracellular matrix. Previous reports have demonstrated that it is associated with epithelial-mesenchymal interfaces and is expressed during embryonic and tumour development, wound healing, cell proliferation and it may be involved in immunomodulation. The human placenta shows numerous features related to these aspects. We have investigated the presence of tenascin in the human placenta throughout pregnancy by immunohistochemistry. We used monoclonal (mAb) and polyclonal (pAb) antibodies to tenascin, a mAb to fibrin, a pAb to fibrinogen, and the mAb Ki-67 as proliferation marker. Tenascin was highly expressed in the mesenchymal villi which are considered the basis of growth and differentiation of the villous trees. Moreover, fibrinoid deposits at the surfaces of the villous trees were always separated from the fetal stroma by tenascin. The stroma of villi encased in fibrinoid was also positive for tenascin. This glycoprotein was also expressed in the villous stroma directly apposed to cell islands and cell columns. In the proximal portions of both epithelial structures, cytotrophoblast was Ki-67 positive. These data show that tenascin is expressed during the development of the placenta, particularly in the mesenchymal villi, cell islands and cell columns. These structures are considered to be the proliferating units of the villous trees. Tenascin underlying fibrinoid deposits suggests that it also participates in repair mechanisms. Thus, in the human placenta tenascin expression can be correlated with villous growth, cell proliferation, and fibrinoid deposition. Its role in immunoprotection of fetal tissues in areas where syncytiotrophoblast as barrier is missing or damaged is discussed.

Establishment of a human endometrial cell culture system and characterization of its polarized hormone responsive epithelial cells.

Abstract.Uterine epithelial cells from normal human endometrium were cultured as a primary cell culture in a dual-chambered system. The epithelial cells were isolated from endometrial tissue of the proliferative phase obtained by hysterectomy. The epithelial cells were seeded on Millicell CM filters coated with the extracellular matrix Matrigel. Depending on the culture conditions, the epithelial cells formed a polarized cell monolayer on Matrigel or gland-like structures in Matrigel. The epithelial cell polarity was maintained during culture, which could be proved by electron microscopy. The progesterone and estrogen receptors as typical marker molecules for physiologically intact endometrial epithelial cells could be detected immunohistochemically as well as by RT-PCR in vitro and were down-regulated by medroxyprogesterone acetate (MPA) used as progesterone analogue. As this cell culture system exhibits morphological and immunohistochemical characteristics, typical for the in vivo situation, and since it can be modulated by hormone treatment under the in vitro conditions described, it represents a valuable tool for investigating processes that are essential for endometrial differentiation and reproductive functions.

Effects of trophoblast invasion on the distribution of leukocytes in uterine and tubal implantation sites

OBJECTIVE To distinguish endocrine and paracrine influences on leukocyte subpopulations at uterine and tubal implantation sites. DESIGN Retrospective immunohistochemical study. SETTING Departments of Anatomy, and Obstetrics and Gynecology, School of Medicine, RWTH University of Aachen, Aachen, Germany. PATIENT(S) Ten women with a viable ectopic pregnancy (EP), 25 women who had undergone elective first-trimester termination of pregnancy, and 4 women who had undergone hysterectomy with adnexectomy. INTERVENTION(S) None. MAIN OUTCOME MEASURE(S) Quantitative analysis of leukocyte subpopulations at the implantation sites and their corresponding noninvaded tissues, decidual tissue from patients with EP, and tubal mucosa from normal menstrual cycle. RESULT(S) Similar numbers and characteristic distribution patterns of macrophages, T cells, and B cells were found at both normal intrauterine and tubal implantation sites. Natural killer (NK) cells were always absent from tubal mucosa. The number and distribution of leukocytes within decidual tissue from women with EP corresponded to those in the noninvaded decidual compartment in intrauterine pregnancy (IUP). CONCLUSION(S) Leukocyte populations present in the tubal and uterine mucosa are an intrinsic characteristic of these tissues. The distinct leukocyte distribution pattern at the implantation sites suggests that the invading trophoblast exerts a paracrine influence on endometrial and endosalpingeal leukocytes. The absence of natural killer cells from the tubal wall may be one reason for the higher degree of invasiveness of the trophoblast at the tubal implantation site.

Progestins, progesterone receptor modulators, and progesterone antagonists change VEGF release of endometrial cells in culture.

The influences of the synthetic progestin, medroxyprogesterone acetate (MPA), the progesterone receptor modulator J867, and the antagonist ZK137316 were studied in vitro on isolated endometrial epithelial cells, as well as endometrial fibroblasts. We evaluated the expression of estrogen receptor alpha (ER) and the progesterone receptor (PR) by RT-PCR. ER and PR were strongly expressed in the fibroblasts and epithelial cells under treatment with 10(-8) M 17beta-estradiol (E(2)). Treatment with 10(-6) M J867 or ZK137316 upregulated the PR expression as did E(2), in contrast to treatment with 10(-6) M MPA, which caused a downregulation of PR in epithelial cells, but not in fibroblasts. In addition, the vascular endothelial growth factor (VEGF) release into the cell culture medium was analyzed by a VEGF-ELISA. VEGF which plays an important role in angiogenesis, is regulated by steroid hormones as well as hypoxia. E(2) stimulates VEGF release into the medium in both cell types. MPA reduces VEGF release significantly in the fibroblast cell culture, but increases it in the epithelial cell culture. ZK137316, in the presence or absence of E(2), reduces VEGF release in fibroblast cell culture. J867 increases the VEGF production in fibroblasts only in the presence of E(2). Both compounds show no significant effects, compared to E(2), in epithelial cell culture. The different results for the epithelial cells and fibroblasts indicate that the pharmacological effects of PR modulators (PRMs) and progesterone antagonists (PAs) may be cell specific and depend on the presence or absence of partial progestagenic agonistic activities. This observation opens up new perspectives for various clinical applications.

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-gamma (Th-1), interleukin-4 (Th-2), and interleukin-1beta proteins are expressed in the human first-trimester decidua. Interleukin-2, interferon-gamma, and interleukin-4 mRNA mainly derive from the decidual tissue leukocytes. Interleukin-1beta mRNA is expressed by all decidual cell types. Interferon-gamma 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-gamma 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).

Redistribution of adhering junctions in human endometrial epithelial cells during the implantation window of the menstrual cycle

The human uterine epithelium is characterised by remarkable plasticity with cyclic changes in differentiation that are controlled by ovarian steroid hormones to optimise conditions for embryo implantation. To understand whether and how cell–cell adhesion is affected, the localisation of junction proteins was studied throughout the menstrual cycle. Expression patterns were examined by immunofluorescence in 36 human endometrial specimens of different cycle stages. Antibodies against the desmosomal proteins desmoplakin 1/2 (Dp 1/2) and desmoglein 2 (Dsg 2), the adherens junction proteins E-cadherin and β-catenin and also the common junctional linker protein plakoglobin showed a strong subapical staining during the proliferative phase until the early luteal phase (day 20). In the mid- to late luteal phase, however, these junctional proteins redistributed over the entire lateral plasma membranes. In contrast, tight junction proteins (ZO-1, claudin 4) remained at their characteristic subapical position throughout the menstrual cycle. mRNA levels of Dp 1/2, E-cadherin and ZO-1 obtained by real time RT-PCR were not significantly changed during the menstrual cycle. The observed redistribution of desmosomes and adherens junctions coincides with the onset of the so called implantation window of human endometrium. We propose that this change is controlled by ovarian steroids and prepares the endometrium for successful trophoblast invasion.

Apical plasma membrane-bound enzymes of rabbit uterine epithelium

SummaryIn order to monitor changes in the apical cell membrane of rabbit uterine epithelium which are postulated to be a precondition for trophoblast attachment, the marker enzymes: alkaline phosphatase, aminopeptidase M, γ-glutamyl transferase and dipeptidyl peptidase IV were investigated during the periimplantation phase. Endometrium of early pregnancy (implantation chamber, interblastocyst endometrium; 5–8 days post coitum, d p.c.) was compared with specimens obtained at hCG-induced pseudopregnancy (p. hCG) to distinguish between membrane changes regulated by maternal plasma steroid hormones and such which might be induced locally by blastocyst-derived signals.All enzymes tested showed their main activity at 5 d p.c./p. hCG. The weakest reaction in this series of stages was generally found at 8 d p.c. (interblastocyst segments) or at 8 d p. hCG. In contrast to the rest of the epithelium, the implantation chamber retained high activity of dipeptidyl peptidase IV, and the activity of alkaline phosphatase even raised here again at 7 and 8 d p.c. indicating a direct local influence of the blastocyst on the luminal epithelium. The results suggest that 1) considerable changes occur in the composition of the apical plasma membrane of the uterine epithelium when the endometrium enters the “receptive state”, 2) the overall trend is towards a loss of apical-type characteristics of this membrane domain and 3) the changes are modulated both systemically (by plasma steroid hormone levels) and locally by signals from the implanting blastocyst.

Preparation of Rabbit Uterine Epithelium for Trophoblast Attachment: Histochemical Changes in the Apical and Lateral Membrane Compartment

Embryo implantation is initiated by an interaction of the trophoblast with the uterine epithelium. In the preimplantation phase the uterine epithelium undergoes extensive changes that are detectable by morphological and histochemical methods. It is usually assumed that these changes ’are somehow related to the acquisition of a state of “receptivity” (Psychoyos, 1976) for trophoblast adhesion and invasion. The morphological transformation is already well documented in the rabbit, one of the most widely used models for implantation studies (Boving, 1963; Denker, 1970, 1977; Enders and Schlafke, 1971; Beier, 1973; Davies and Hoffman, 1973, 1975; Suzuki and Tsutsumi, 1980, 1981; Busch, 1982; Winterhager, 1985).

Uteroglobin expression and release in the human endometrium.

Initially, uteroglobin (UGL) was identified as the major protein component of rabbit uterine secretion and blastocyst fluid during the time before and at implantation.1,2 Recently, its progesterone-dependent expression was shown in the equine endometrium.3 Our research efforts have focused on the biological significance of UGL in human reproduction, particularly in the endometrium. An essential prerequisite to start with those functional studies is the exact cellular localization and studies on the expression of this protein throughout the menstrual cycle.4,5 To study the endocrine/paracrine regulation of UGL expression, we use a cell culture system of isolated human endometrial cells.6 By means of this primary cell culture system, we hope to elucidate the cell-cell interactions between epithelial and mesenchymal cells, the cytokine signaling on UGL expression, and its ligand binding dynamics since these studies are not permitted in vivo because of ethical reservations.

„Meet the AIX-PERTs...“

ZusammenfassungHintergrundDer berechtigte Anspruch, notwendige Fertigkeiten der Notfallversorgung von jedem Arzt erwarten zu können, und ein steigendes öffentliches Interesse an der Qualität ärztlicher Leistungen erfordern eine größere Gewichtung dieser Lerninhalte im Medizinstudium.MethodenIm Rahmen der seit dem 1.10.2003 gültigen neuen Approbationsordnung für Ärzte beschloss die medizinische Fakultät der RWTH Aachen zum Wintersemester 2003 einen „Modellstudiengang Medizin“ zu starten. Praktische Fertigkeiten der zukünftigen Mediziner sollen durch interdisziplinäre Vermittlung organsystembezogener Inhalte verbessert werden.ErgebnisUnter Einbindung von problemorientiertem Lernen wurde mit AIX-PERT (Aix-la-Chapelle Program for Emergency Medical Care and Resuscitation Training) ein multidisziplinäres Konzept zur Studiumseinführung entwickelt und evaluiert, das definierte Kernlernziele zur notfallmedizinischen Versorgung beinhaltet.SchlussfolgerungDie Evaluationsergebnisse demonstrieren die Akzeptanz von AIX-PERT und sind Basis longitudinaler Implementierung relevanter notfallmedizinischer Inhalte in das neue Curriculum.AbstractBackgroundExtensive knowledge and skills in the basics of emergency medical care are of paramount importance for every physician and should therefore be an integral part of medical education.MethodsRegulations for medical licensure in Germany were revised by the administrative authorities in 2002 and as a consequence the Medical Faculty of the University of Aachen (Germany) decided to start the Medical Reform Curriculum Aachen. A multidisciplinary, problem-oriented and organ-related approach to medical education replaces the classical discrimination between basic and clinical sciences.ResultsWith AIX-PERT (AIX-la-Chapelle Program for Emergency medical care and Resuscitation Training), a program consisting of problem-based learning sessions was developed for introduction to the first year students. Defined teaching objectives in emergency medicine are now incorporated in undergraduate medical education.ConclusionThe extremely positive evaluation of the new approach encouraged us to promote AIX-PERT further. In the future the effects of success of this approach will be assessed by longitudinal studies of skills and knowledge during the continuing curriculum.