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G Dohr

Publications and source records attributed to G Dohr.

At least 37 records · Page 2Linked to original sources
PubMed2000

Antibody reaction patterns in first trimester placenta: implications for trophoblast isolation and purity screening.

The aim of this immunohistochemical and cytochemical study was to select specific antibodies to establish an efficient purification protocol for first trimester trophoblast and for subsequent purity screening of isolated trophoblast cells. The reactivity of antibodies to various cytokeratin filaments, glycoprotein CD9, fibroblast specific antigen (FSA), common leukocyte antigen CD45RB and macrophage antigens CD163, CD68 and CD14 were studied on cryosections of placental tissue. Among the cytokeratins tested, cytokeratin 7 was the only keratin filament type, which was not expressed in placental mesenchymal cells, but in all trophoblast subpopulations. Since anti-CD9, in addition to mesenchymal cells, also strongly labels extravillous cytotrophoblast cells, whereas the antibody to FSA only reacts with mesenchymal cells, anti-FSA is suitable as a depletion antibody for mesenchymal cells. Among the macrophage markers anti-CD163 was the most specific for Hofbauer cells. CD45RB was expressed on maternal and fetal leukocytes as well as on Hofbauer cells. Isolated first trimester placental cell preparations that have been collected from a density gradient contained up to 45 per cent non-trophoblast cells. Immunocytochemistry using antibodies to CK7, FSA, vimentin, CD45RB and CD163 demonstrated that subsequent immunodepletion with antibodies to CD45RB and FSA increased the purity of the trophoblast preparation to greater than 98 per cent. According to this study trophoblasts from first trimester placentae should be identified by cytokeratin antibodies specific for the isoform 7. Purification of isolated trophoblasts by density gradient alone does not result in a sufficient degree of purity.

Antibodies↗
PubMed1999

Paracrine regulation of distinct trophoblast functions in vitro by placental macrophages.

In view of the accumulating evidence for paracrine mechanisms regulating trophoblast function, we tested the hypothesis that placental macrophages affect trophoblast activity in a paracrine fashion. Trophoblast was isolated from 17 term placentas (-IP). One aliquot of cells was further immunopurified (+IP) using an HLA class I antibody. This increased the proportion of trophoblast (+IP >97%; -IP approximately 70%) as identified by rigorous immunocytochemistry. Most (approximately 70%) non-trophoblast cells in -IP were macrophages. The cells were cultured for 5 days with a daily medium change. In addition, +IP cells from seven placentas were cultured with lipopolysaccharide (LPS)-stimulated or -unstimulated macrophage-conditioned media. The concentrations of lactate, trophoblast-specific hormones, human chorionic gonadotropin-beta (hCG-beta) and human placental lactogen (hPL), of several prostanoids and of endothelin-1 and angiotensin II were determined in the culture media. The accumulated amounts of substances released into the culture media, corrected for the greater proportion of trophoblast in +IP cultures, were on average two- to threefold higher (hCG-beta: 18-fold) in +IP than in -IP, with the exception of endothelin-1,2 (no change), angiotensin II (-70%) and 6-keto-prostaglandin-F1alpha (-40%). [3H]leucine incorporation into the trichloroacetic acid (TCA)-precipitable pool measured on day 5 was twofold higher in +IP than in -IP. Addition of conditioned media reverted these changes. The data demonstrate that placental macrophages in culture affect trophoblast biosynthetic activity in a paracrine fashion. We conclude that macrophages are important regulators of trophoblast activity.

Cell Communication↗
PubMed1999

HLA-G in the human thymus: a subpopulation of medullary epithelial but not CD83(+) dendritic cells expresses HLA-G as a membrane-bound and soluble protein.

The human MHC class Ib gene HLA-G is transcribed and translated in different placental cell subpopulations during pregnancy. In addition to this restricted tissue distribution, HLA-G proteins were also recently detected in the thymus of HLA-G transgenic mice, as well as in some human thymic epithelial cells (TEC). There was a need to further define the phenotype of the HLA-G-expressing cells in the human thymus as well as the type of translated forms that they produce. Using several HLA-G-specific mAb and immunohistochemistry performed on cryosections of human thymi at different ages, we found that the HLA-G-expressing cells are present on medullary cells exhibiting the epithelial morphological type 6. Co-localization experiments performed by double or triple immunofluorescence staining demonstrate that these HLA-G-expressing cells express various cytokeratins, epithelial cell markers but not the CD83 dendritic cell marker. We further show by ELISA measurements that a subset of primary cultured human TEC also expresses soluble HLA-G. Therefore, HLA-G protein tissue distribution is not restricted solely to placental cells. A subpopulation of medullary TEC also expresses HLA-G both at their cell surface and in secreted form, raising the question of the functional significance of such MHC class Ib molecules. Whether thymic soluble and/or membrane-bound HLA-G contribute to inhibit NK cells or to a negative selection of autoreactive T cells which could be harmful in case of pregnancy and/or to a positive selection of viral peptides/HLA-G-restricted CD8(+) T cells remains to be demonstrated.

Adolescent↗
PubMed1999

Fas and Fas-ligand are expressed in the uteroplacental unit of first-trimester pregnancy.

PROBLEM: Fas and Fas-ligand (FasL) are thought to provide a strategy for reducing graft rejection in immunologically 'privileged' tissues by controlling injurious lymphocyte reactions. As the uteroplacental unit is often defined as an immune-privileged site, we investigated the expression of Fas and FasL in this tissue in the first trimester of pregnancy. METHOD OF STUDY: Western blotting, immunohistochemistry, and double immunofluorescence were used for this examination. RESULTS: Western blotting with purified first-trimester trophoblast cells revealed one specific band for FasL. The presence of FasL on different trophoblast populations could be confirmed by immunohistochemistry and double immunofluorescence. In the villous part of the placenta, FasL is mostly located on cytotrophoblast cells with no access to maternal blood flow, whereas in trophoblast-invaded uterine tissue, interstitial trophoblast cells, which are in close contact with maternal leukocytes, revealed a strong signal for FasL, but no staining for Fas on these cells. However, Fas was found on CD45+ maternal leukocytes. CONCLUSION: Based on our experimental findings, we speculate that the abundant presence of FasL on trophoblast cells within the maternal decidua may play an important role in the maintenance of immune privilege in the pregnant uterus by endowing fetal trophoblast cells with a defense mechanism against activated maternal leukocytes, whereas in the villous part of the placenta, the Fas FasL system seems to be involved in the regulation of placental growth.

Apoptosis↗
PubMed1999

First trimester human endovascular trophoblast cells express both HLA-C and HLA-G.

PROBLEM: In human pregnancies, trophoblasts, in contrast to placental connective tissue and the fetus itself, come into direct contact with the maternal allorecognizing system at special sites. Villous syncytiotrophoblasts washed around by maternal blood lack HLA class I proteins, whereas extravillous trophoblasts, which deeply invade maternal uterine tissues, express high amounts of HLA-G and also HLA-C, the latter to a lesser degree, however. A subpopulation of extravillous trophoblasts, the endovascular trophoblast, enters maternal spiral artery lumen and, like syncytiotrophoblast, comes into direct contact with maternal blood. Less is known about HLA class I distribution on this endovascular trophoblast subpopulation. METHOD OF STUDY: A comparative immununohistochemical analysis was done on decidual cryo-sections containing trophoblast-invaded spiral arteries using different anti-HLA class I monoclonal antibodies (mAbs) and a peroxidase-labeled streptavidinbiotin detection system. RESULTS: MAbs W6/32 (anti-HLA-A, -B, -C, -G), HCA2 (anti-HLA-A, -G) G233 and 87G (both anti-HLA-G) resulted in strong positivity on endovascular trophoblasts. L31 (anti-HLA-C) and HC10 (anti-HLA-B, -C) revealed clear positivity, whereas TU149 (anti-HLA-B, -C, some -A) produced a heterogeneous staining pattern, faintly positive on some endovascular trophoblastic cells and negative on others. MAb LA45 (anti-HLA-A, -B) did not bind to any endovascular trophoblast, neither did BFL.1 (anti-HLA-G) nor 16G1 (anti-HLA-G, soluble). CONCLUSION: This study shows that trophoblastic cells belonging to the endovascular subpopulation express considerable amounts of HLA-G and slightly less HLA-C.

Antibodies, Monoclonal↗
PubMed1999

Placental glucose transporter expression is regulated by glucocorticoids.

Although glucocorticoids play important roles in development and fetal programming, they are widely used for treatment of a variety of diseases during pregnancy. In various tissues, glucocorticoids down-regulate glucose transport systems; however, their effects on glucose transporters in the placenta are unknown. In the present study, the glucose carrier proteins GLUT1 and GLUT3 were localized in the trophoblast and endothelium of the human, rat, and mouse placenta. Subsequently, it was investigated whether glucocorticoids affect messenger ribonucleic acid and protein expression of these molecules by Northern and Western blotting using 1) human term placental trophoblast cells cultured in the presence or absence of 0.5, 5, and 50 micromol/L triamcinolone; 2) placentas of rats that received a single i.p. dose of 0.38 mg/kg triamcinolone; and 3) placentas of transgenic mice bearing an antisense glucocorticoid receptor gene construct. In all of these systems, both glucose transporters were significantly down-regulated (P < 0.05), with the exception of increased GLUT3 messenger ribonucleic acid and protein levels in transgenic mice. The results demonstrate that triamcinolone is a potent regulator of placental GLUT1 and GLUT3 expression involving the glucocorticoid receptor. We speculate that impaired expression of placental glucose transporters after glucocorticoid administration might contribute to the adverse side-effects, the foremost of which is a growth-retarded fetus, of this treatment during pregnancy.

Animals↗
PubMed1999

Decidual macrophages are the population of decidual adherent cells which regulates perforin expression in cytolytic cells.

PROBLEM: We have shown that addition of decidual adherent cells (DAC) to the culture of decidual lymphocytes (DL) prevents the downregulation of perforin expression in these cells. Because DAC are a mixture of various cell populations, the aim is to analyze immunophenotypic characteristics of DAC and to determine which cell population is involved in the regulation of perforin expression. METHOD OF STUDY: First trimester pregnancy decidual cells were obtained by enzymatic tissue digestion. Decidual cells and peripheral blood lymphocytes (PBL) were centrifuged on Ficoll-Hypaque density gradient and cultured overnight to obtain adherent cells, which were analyzed by flow cytometry and immunocytochemically. RESULTS: Almost all peripheral blood adherent cells (PBAC) (ca 90%) expressed monocyte/macrophage markers but only 10-20% of DAC. The rest of DAC expressed markers of stromal cells. HLA-DR depleted population of DAC (stromal cells only) could not prevent downregulation of perforin expression in cultured DL and PBL. CONCLUSION: Decidual macrophages are involved in the regulation of perforin expression in DL.

Cell Adhesion↗
PubMed1999

Apoptotic nuclei within the uterine decidua of first trimester pregnancy arise from CD45 positive leukocytes.

PROBLEM: The Fas/Fas-ligand (FasL) system is discussed to be one of the main components responsible for the formation and maintenance of an immune privilege. As we were able to detect FasL on fetal trophoblast cells infiltrating the maternal uterine tissue and Fas on decidual leukocytes (Hammer et al., Am J Reprod Immunol, 1999; 41: 41-51), we investigated whether apoptosis occurs in the maternal decidua during the first trimester of pregnancy. METHOD OF STUDY: A TUNEL-assay was performed in combination with immunofluorescence to detect and characterize cells undergoing apoptosis. RESULTS: Apoptotic nuclei mainly arising from CD45+ leukocytes could be detected in the trophoblast-invaded decidua basalis as well as in the non-invaded decidua parietalis. However, while in the decidua parietalis the apoptotic leukocytes are scattered within the whole functional stroma, in the decidua basalis they are concentrated in the area around the so-called "necrotic foci" and especially in the primary feto-maternal contact zone. where trophoblast cells from the cell columns begin to invade the maternal tissue. CONCLUSION: Because of the irregular distribution of apoptotic CD45+ leukocytes in the decidua basalis we think that some of the maternal leukocytes are activated and killed by fetal trophoblast cells which express FasL.

Apoptosis↗
PubMed1998

HLA expression on immature and mature human germ cells.

Human leukocyte antigens (HLA) are membrane-bound glycoproteins encoded by the human major histocompatibility complex located on chromosome 6. They are known to function in immnunologic recognition and, with regard to reproduction, a number of non-immune functions have been proposed. Although the expression patterns of the major histocompatibility antigens have been extensively studied at the maternal fetal interface, there are still controversial reports on the expression of these molecules by human gametes and preimplantation stages. This brief review focuses on recent studies where the expression and distribution of HLA on human spermatogenic cells (spermatogonia, primary and secondary spermatocytes, spermatids, spermatozoa), primary and secondary oocytes, and preimplantation embryos have been investigated. These results, and their possible implications for the fertilization process and further embryonic development, will be presented.

Animals↗
PubMed1998

HLA expression at the maternal-fetal interface.

Pregnancy in the human presents an "immunological paradox," because of the unexpected willingness of mothers to accept genetically disparate tissues. The fact that the fetus can develop unharmed for nine months shows that protective mechanisms must exist to permit its survival. The conditions that permit the genetically dissimilar human fetus to evade rejection by its mother's immune system have been the subject of intense interest for several decades. As the placental cells, which are in contact with maternal blood or tissue, are devoid of HLA class II antigens, interest has focused on the expression of HLA class I molecules. Recent developments in the constitutive, transcriptional, and translational expression of HLA class I molecules on anatomically and morphologically different subpopulations of trophoblast cells will form the basis of this short review.

Female↗
PubMed1997

Endothelial cells in chorionic fetal vessels of first trimester placenta express HLA-G.

Using four different HLA-G-recognizing monoclonal antibodies (mAb), we investigated whether this nonclassical HLA class I molecule could be expressed in placental cell types other than extravillous cytotrophoblasts (evct) in which HLA-G has already been detected. Immunohistochemical analysis was performed on serial cryosections of first trimester placenta as well as on maternal decidual tissue. In addition to some proliferative evct, the recently described BFL.1 mAb also slightly stained some villous cytotrophoblast (vct) stem cells located near cell columns and cell islands, which until now have been considered as HLA-G negative. The same staining pattern was obtained with the 16G1 mAb raised against the soluble HLA-G isoform, whereas neither 87G nor HCA2 reacted with vct but did strongly label the invasive populations of evct, including interstitial and endovascular trophoblasts. Surprisingly, BFL.1 strongly and reproducibly stained endothelial cells in the fetal capillaries present in the mesenchymal core of the chorionic villi, whereas none of the other surrounding cellular components were stained. The same specific labeling was obtained, although with less intensity, with the three other HLA-G-recognizing mAb. In contrast, maternal endothelial cells present in spiral arteries of the decidua parietalis remained unstained. This unexpected cellular localization suggests that HLA-G may be present as a soluble form during the whole period of fetal vascularization and/or exert a nonimmunological function related to the endothelial cell type, in particular in the angiogenesis process which is highly active, until term, in chorionic villi.

Blood Vessels↗
PubMed1997

Cytokeratin 17 as an immunohistochemical marker for intramural cytotrophoblast in human first trimester uteroplacental arteries.

Trophoblast cells, as blastocyst-wall derivatives, are of epithelial origin and differentiate initially into syncytiotrophoblast and cytotrophoblast subpopulations. Cyto- and syncytiotrophoblasts are the two cell types present in the surface cell layers of placental villi. Cytotrophoblastic cells lie in contact with the basal lamina and are the proliferating stem cells that guarantee cytotrophoblast and syncytiotrophoblast persistence. Implantation and placenta formation are mainly based on these two cell types. Villous cytotrophoblasts are the stem cells for all extravillous trophoblast subpopulations, which exhibit strictly regulated invasiveness. One aspect of extravillous trophoblasts is that they invade maternal endometrial spiral arteries and dilate them in order to achieve sufficient fetal blood supply. During this process, trophoblast cells, which are located in the remodelled uteroplacental artery walls, are thus defined as intramural cytotrophoblasts. Trophoblast differentiation is accompanied and defined by alterations in, for example, the translation pattern for cytokeratin genes. In an immunohistochemical study, we have demonstrated that only intramural cytotrophoblasts, from all the trophoblast populations of the junctional zone, express cytokeratin 17. Furthermore, cell shape and vascular architecture indicate that, in human placenta, intra-arterial trophoblast cells reach their destination by migration through the endometrial interstitium with consecutive intravasation. Cytokeratin 17, in particular, can therefore be used as a specific immunohistochemical marker for the intramural trophoblast subpopulation.

Arteries↗
PubMed1997

Immunohistochemical examination of trophoblast populations in human first trimester and term placentae and of first trimester spiral arteries with the monoclonal antibody GZ 112.

This paper presents an immunohistochemical study with a monoclonal mouse antibody GZ 112, an IgG1 kappa, which is directed against an antigen expressed in first trimester placenta by all proliferative and invasive extravillous trophoblast populations including a population of Langhans cells that represent extravillous stem cells. Additionally, the GZ 112 antigen is associated with morphological changes of spiral arteries preceding local trophoblast invasion. In term placentae, GZ 112 also strongly reacts with all extravillous trophoblast populations, but additionally recognizes partly villous cytotrophoblast and syncytiotrophoblast too, displaying a heterogeneous staining pattern. GZ 112 is directed against a 42-KDa antigen. Intracytoplasmic network-like staining and cross-reactivity with various human surface and glandular epithelia indicate a cytokeratin intermediate filament or a cytokeratin intermediate filament associated molecule as antigen.

Animals↗
PubMed1997

Ultrastructural observations in human oocytes and preimplantation embryos after zona opening using an erbium-yttrium-aluminium-garnet (Er:YAG) laser.

For more than 3 years we have performed laser-assisted hatching prior to embryo transfer in patients with recurrent implantation failure using an erbium-yttrium-aluminium-garnet (Er:YAG) laser system that operates in the infrared region of the light spectrum. The laser beam is guided through a quartz fibre and is brought into direct contact with the zona pellucida. This study was undertaken to evaluate the ultrastructural effects of this laser on the zona pellucida and underlying cell membrane of unfertilized human oocytes and pathologically fertilized preimplantation embryos using light and scanning electron microscopy. The Er:YAG laser produces an almost circular zona opening in the shape of a truncated cone tapering off towards the inside, with a mean diameter of 18 mm. The exact diameter of the drilled site depends on the diameter of the fibre tip and the total number of pulses applied. After laser interaction, the zona matrix and the surface of the underlying ooplasm membrane showed no degenerative alterations. We conclude that the Er:YAG laser is an effective microsurgical tool for achieving reproducible, precise zona openings particularly suitable for purposes of assisted hatching because of their characteristic shape.

Aluminum↗
PubMed1997

Amnion epithelial cells, in contrast to trophoblast cells, express all classical HLA class I molecules together with HLA-G.

PROBLEM: The expression of the non-classical HLA-G gene has been shown at the protein level on trophoblast-derived embryonic tissue, like the extravillous cytotrophoblast. However, the presence of HLA-G on embryoblast-derived cells is currently controversial. The amnion epithelium is an embryoblast-derived cell layer covering the amnion cavity and is the main source for the amnion fluid. METHOD: The expression of HLA class I molecules was investigated by immunohistochemical, biochemical, and molecular biological methods in amnion membranes and amnion fluid. RESULTS: Immunohistochemically, HLA-C and occasionally also-B molecules as well as HLA-A and/or -G molecules have been identified on amnion epithelial cells. These results were extended by Western blotting with purified amnion epithelial cells where HLA-B and/or -C, HLA-A and HLA-G antigens have been detected. As expected HLA-G mRNA was detected in amino epithelial cells. Furthermore, classical HLA molecules as well as HLA-G were found in amnion fluid. CONCLUSION: These results show that the amnion epithelium frequently expresses classical HLA class I molecules as well as HLA-G. The expression of HLA-G antigens on amnion epithelial cells and their presence in the amnion fluid, which is continually ingested by the fetus, may be particularly relevant for the induction of peripheral tolerance.

Amnion↗
PubMed1997

HLA class I expression on the materno-fetal interface.

PROBLEM: The conditions that permit the genetically dissimilar (haplo-non-identical) human fetus to evade rejection by its mothers immune system have been the subject of intense interest for several years. As the placental cells, which are in contact with maternal blood or tissue, are devoid of HLA class II antigens, the interest has focused on the expression of HLA class I molecules. METHOD OF STUDY: Recent findings on the constitutive, transcriptional, and translational expression of HLA class I molecules on anatomically and morphologically different subpopulations of trophoblast cells will form the basis of this article. RESULTS: The expression of HLA class I molecules in the trophoblast cells, forming the materno-fetal junctional zone is inhomogeneous. It differs depending on the differentiation and location of trophoblast cells within the placenta and furthermore on the stage of gestation. On the transcriptional level HLA-A, -B, -C, -E, and -G could be detected on individual trophoblast populations, whereas only HLA-C and HLA-G seem to be translated to protein. CONCLUSIONS: The expression of HLA class I antigens by trophoblast cells is not simply suppressed. Instead, less polymorphic HLA-G and HLA-C antigens are carefully selected from the class I multigene family. This gives rise to the assumption that these two HLA class I molecules play an important role in the maintenance of pregnancy.

Female↗