Antigenicity of trophoblast and possible antigen-masking effects during pregnancy.

The interaction between cultured human trophoblast cells and materanl lymphocytes was used as an in vitro model to investigate trophoblast antigenicity. Cytotoxic effects in the trophoblast monolayer were apparent after 72 hr incubation and depended on the presence of non-lymphoid cell types in addition to lymphocytes. Lysis of trophoblast was preceded by blast cell formation and apparently involved close contact between the maternal cells and the trophoblast cells. The nature of the cytotoxic reaction suggested the presence of histocompatibility factors on trypsinized trophoblast cells. This manifestation could be due to removal of fibrinoid or enhancing antibody from the cultured cells. The ability of trypsinized trophoblast cells to synthesize mucoprotein was investigated by the Hale colloidal iron test and found to be unimpaired. The effect of maternal serum on the recognition of human trophoblast antigens by maternal lymphocytes was used as an in vitro model to investigate the occurrence of enhancing antibody in maternal serum. The cytotoxic effects of maternal lymphocytes on trophoblast were completely prevented by the presence of maternal serum, this protective effect being reduced significantly by removal of IgG from the maternal serum. A slight protective effect of allogeneic pregnancy serum was also observed. It is suggested that these findings support a role for immunological enhancement in maintaining the foetal allograft.     

Immunohistochemical localization of interferons in human placental tissues in normal, ectopic, and molar pregnancy

Interferon (IFN)alpha, beta, and gamma have been localized in normal and pathological human pregnancy using both polyclonal and monoclonal antibodies in immunohistochemical techniques. IFN alpha was localized to fetal chorionic villous syncytiotrophoblast throughout normal pregnancy, as well as to extravillous trophoblast in the placental bed and chorion lave. Maternal decidual leukocytes, as well as fetal Hofbauer cells in the villous mesenchyme, also contained IFN alpha, IFN gamma was detected in villous syncytiotrophoblast, while anti-IFN beta showed only patchy weak reactivity with syncytiotrophoblast. Reaction patterns on ectopic pregnancy tissues were similar to those in early intrauterine pregnancy. In molar pregnancy, reactivity for IFN alpha, beta, and gamma was observed in syncytiotrophoblast. Along with their potential anti-viral effects, placental interferons could play a role in local immunomodulation or in regulation of embryonic cell proliferation and differentiation.     

Role of placental alkaline phosphatase in the interaction between human placental trophoblast and Trypanosoma cruzi.

Congenital Chagas disease, due to the intracellular parasite Trypanosoma cruzi, is associated with premature labor, miscarriage, and placentitis. Human enzyme placental alkaline phosphatase (PLAP) (EC 3.1.3.1.) is membrane-anchored through glycosylphosphatidylinositol (GPI). PLAP is present in plasma in late pregnancy, 36 to 40 weeks; there are lower levels in maternal Chagas disease. Infants born to such mothers may have congenital Chagas disease. Human placental villi (PV) were treated with phospholipase-C (PL-C) and then cultured with T. cruzi to determine the effect of the parasites on PLAP activity as an in vitro model. There is less PLAP activity after treatment by PL-C and during culture with T. cruzi. Pretreatment of PV with PL-C before culture with T. cruzi yielded essentially normal specific activity of PLAP and prevented or greatly reduced infective penetration of villi by parasites. The results are consistent with a pathogenetic role for placental alkaline phosphatase in congenital Chagas disease. Receptor activation of membrane attachment to PLAP may be a device used by T. cruzi to enable parasite invasion of human trophoblast.     

Production of antibodies to human interferons in mice.

Neutralizing antibodies were raised in mice that had been inoculated repeatedly with moderate quantities of human leukocyte interferon highly purified by affinity chromatography on immobilized anti-interferon globulins. Interferon preparations of lesser purity sensitized the mice to subsequent inoculations of interferon and almost invariably caused death before anti-interferon titers developed. Antibody-purified interferon stabilized by sodium dodecyl sulfate was a superior antigen to interferon that had received mouse serum albumin as an additive. The amount of antibody could be augmented by experimental induction of ascites. The antibodies specifically neutralized leukocyte and lymphoblastoid interferons but not those interferons obtained cultures of human foreskin fibroblasts, embryonic kidney cells, and amnion cells.     

Immunology of human placental trophoblast membrane antigens

The identification of the mechanisms of immunological survival of the semiallogeneic conceptus in utero during viviparous pregnancy is central to current studies in pregnancy immunology, since any other unmatched intrauterine allograft would be expected to suffer tissue rejection. Elucidation of normal feto-maternal interactions in human uteroplacental tissues, as well as the characterization of fetal trophoblast cell surface antigen expression, has now offered insight into these processes; these have largely focussed on the lack of trophoblastic expression of classical class I MHC antigens and the specialized local immuno-regulatory response that may occur following maternal recognition of other particular trophoblast cell surface antigens. The role of trophoblastic expression of growth factor receptors, endogenous retroviral activity and cellular oncogene products in the growth and differentiation of trophoblast, and its interaction with the host maternal tissues in early pregnancy, remains to be further clarified.     

Onset of maternal arterial blood flow and placental oxidative stress. A possible factor in human early pregnancy failure

The aim was to measure changes in the oxygen tension within the human placenta associated with onset of the maternal arterial circulation at the end of the first trimester of pregnancy, and the impact on placental tissues. Using a multiparameter probe we established that the oxygen tension rises steeply from <20 mmHg at 8 weeks of gestation to >50 mmHg at 12 weeks. This rise coincides with morphological changes in the uterine arteries that allow free flow of maternal blood into the placenta, and is associated with increases in the mRNA concentrations and activities of the antioxidant enzymes catalase, glutathione peroxidase, and manganese and copper/zinc superoxide dismutase within placental tissues. Between 8 to 9 weeks there is a sharp peak of expression of the inducible form of heat shock protein 70, formation of nitrotyrosine residues, and derangement of the mitochondrial cristae within the syncytiotrophoblast. We conclude that a burst of oxidative stress occurs in the normal placenta as the maternal circulation is established. We speculate that this may serve a physiological role in stimulating normal placental differentiation, but may also be a factor in the pathogenesis of pre-eclampsia and early pregnancy failure if antioxidant defenses are depleted.     

New placental and pregnancy proteins as possible markers in oncology

Summary A number of placental and pregnancy proteins has been detected by immunochemical methods in extracts from human term placentae. Several of these proteins already have been isolated and characterized and are now investigated for their usefulness as markers in oncology.Several of these proteins so long tested appear to be useful as markers in monitoring tumour patients, i.e. in the early detection of tumour recurrence or metastases and in the control of therapy. In addition, we hope that a combination of several of these new markers with already established tumour markers will enable to better diagnose tumours. Also will the use of solitary tissue proteins aid in the differential diagnosis of cancers.     

An in vitro model of human placental trophoblast deportation/shedding

Deportation of trophoblast shed from the placenta into the maternal circulation was first described over 100 years ago. Despite this, little is known about the quantity or nature of the shed and deported trophoblasts. Neither do we have a clear understanding of the fate of deported trophoblasts nor do we have a clear understanding of their effects on the maternal physiology. This deficiency is largely due to the inaccessibility of deported trophoblasts in vivo. This study aimed to produce a model that would allow us to study deported trophoblasts. We devised a system for culturing placental explants of 12-week gestation in cell culture inserts with a stainless steel mesh bottom that allowed the ready harvesting of shed/deported trophoblasts. Immunohistochemical and morphologic investigations demonstrated that these in vitro shed/deported trophoblasts are similar to those found in vivo and that apoptotic, necrotic and viable trophoblasts were shed from the explants. Inhibiting caspases induced a change from predominantly apoptotic to predominantly necrotic trophoblast shedding. We have devised an in vitro model that allows the collection of shed/deported trophoblasts which will significantly enhance our ability to study these cells. Our preliminary investigations confirm that apoptosis plays an important role in trophoblast shedding/deportation.     

Identification of estrogen receptor beta-positive intraepithelial lymphocytes and their possible roles in normal and tubal pregnancy oviducts

BACKGROUND: Although intraepithelial lymphocytes (IELs) in human oviductal epithelium have been implicated in the regulation of local immunity, the precise kinetics and mechanism of steroid regulation of IEL are largely unknown. METHODS: We examined the localization of estrogen receptors (ERs) and progesterone receptors (PRs) in 41 human oviducts by immunohistochemistry. These tissues were obtained from various menstrual cycles, also from both post-menopausal women and tubal pregnancies. The expressions of ERbeta mRNA and membrane (m)PR mRNA were examined by in situ hybridization and RT-PCR, respectively. RESULTS: Most of the IEL expressed ERbeta at both mRNA and protein levels. The number of ERbeta-positive IEL, which were identified as CD8-positive T lymphocytes and also were mPR positive, was increased in the late proliferative, the mid-secretory and late secretory phases in normally cycling women (P < 0.05). Interestingly, in tubal pregnancy, ERbeta-positive IELs were consistently abundant. In addition, we found a high Ki-67-labelling index for IEL, although ERalpha was entirely absent in the tubal pregnancy oviducts. CONCLUSIONS: These results suggest that the number of IEL fluctuated because of estrogen and progesterone levels probably through ERbeta and mPR, respectively. ERbeta-positive IEL may be involved in regulating immune tolerance in tubal pregnancy oviducts.     

Identification of human trophoblast membrane antigens in maternal blood during pregnancy.

The development of an immunoradiometric assay for the detection of human trophoblast-specific membrane antigens is described. The test revealed for the first time circulating trophoblast-specific cell membrane antigens in the peripheral blood of pregnant women, but none in non-pregnant female or male controls. Comparison of the circulating levels of these trophoblast-specific proteins between normal and pre-eclamptic blood samples showed no significant differences, thus casting doubt on the role of differential trophoblast antigen deportation in the etiology of toxaemic pregnancy. Matched retroplacental cord blood from several normal and pre-eclamptic pregnancies were examined and found either negative or near the lower sensitivity limit of the assay, suggesting that deportation of trophoblast membrane antigens during gestation is limited to the maternal aspect of the placenta.     

Intrauterine growth restriction, human placental development and trophoblast cell death

Intrauterine growth restriction (IUGR) is a failure to achieve the growth potential of a fetus that is promised by the genetic constitution and environmental influences endogenous to the pregnancy. Optimal placental development and the ability of the placenta to compensate for stimulus-induced injury are central in promotion of normal fetal growth. In this review, we will overview placental development with a focus on how villous structure relates to function. We will also describe the differentiation and turnover of villous trophoblast while highlighting selected features of microscopic placental injury. Histopathological studies of the placenta in IUGR indicate that abnormalities of the maternal spiral arterioles, dysregulated villous vasculogenesis, and abundant fibrin deposition are characteristic of the injuries associated with this condition. We identify selected insults, including oxidative stress and complement activation, and key pathways that regulate apoptosis in villous trophoblast, including increased p53 activity, altered translation of AKT and mTOR proteins, and the stress response of the endoplasmic reticulum. We surmise that trophoblast dysregulation at a subcellular level and loss of functional mass of villous trophoblast via cell death pathways are key contributors to the suboptimal placental performance that yields IUGR. We predict that a better understanding of placental dysfunction in IUGR will lead to targeted therapeutic options for this important clinical condition.     

Expression of vasodilator-stimulated phosphoprotein in human placenta: possible implications in trophoblast invasion.

The vasodilator-stimulated phosphoprotein (VASP) is a 46 kDa protein present at the leading edge of migrating cells. Because trophoblastic cell migration and invasion are critical stages for the achievement of successful implantation and development of the placenta, we investigated VASP expression in different cell types of the human placenta throughout pregnancy by immunohistochemistry and Western blot analysis. We also studied the effect of leukaemia inhibitory factor (LIF) and transforming growth factor-beta1 (TGF-beta1) on the regulation of VASP expression in first trimester placental tissue explants. We found that VASP is expressed throughout pregnancy by a variety of cells in the human placenta. The strongest VASP immunoreactivity was observed in the first trimester. In these samples, the most intense immunoreactivity was in invasive trophoblasts, namely, extravillous cells of the anchoring villi, distal extravillous trophoblasts of cell columns, and also in cells of placental fibrinoids. We also found that LIF (but not TGF-beta1) has a stimulatory effect on VASP expression in placental explants. The strong VASP immunoreactivity in invasive trophoblasts suggests that this protein may be associated with trophoblastic cell motility and may have a role in implantation and trophoblastic cell invasion. We speculate that one of the effects of LIF in successful pregnancy may be its induction of VASP expression.     

Uterine haemodynamics as a possible driving force for endovascular trophoblast migration in the placental bed

Uteroplacental vascular adaptation during pregnancy depends on retrograde endovascular migration of trophoblast in the uterine spiral arteries and their subsequent incorporation into the vessel wall. In the human, this migration process occurs in a step-wise fashion, starting with plugging of the distal ends of the arteries, followed by migration into the decidual and, after several weeks' delay, into the myometrial segments. The hypothesis is put forward that haemodynamical forces play an important regulatory role in this process. A mechanical signal transduction system should then be present within the trophoblastic cells to trigger their rheotactic behaviour. Since the condition of preeclampsia is characterized by restricted colonization of spiral arteries by trophoblast, the implications of this proposed regulatory system on the pathogenesis of the disease are considered.     

Metalloproteinases and their roles in human cancer

It is now widely appreciated that members of the matrix metalloproteinase (MMP) family of enzymes play a key role in cancer development and progression along with many of the hallmarks associated with them. The activity of these enzymes has been directly implicated in extracellular matrix remodeling, the processing of growth factors and receptors, the modulation of cell migration, proliferation, and invasion, the epithelial to mesenchymal transition, the regulation of immune responses, and the control of angiogenesis. Certain MMP family members have been validated as biomarkers of a variety of human cancers including those of the breast, brain, pancreas, prostate, ovary, and others. The related metalloproteinases, the A disintegrin and metalloproteinases (ADAMs), share a number of these functions as well. Here, we explore these essential metalloproteinases and some of their disease-associated activities in detail as well as some of their complementary translational potential. Anat Rec, 2019. © 2019 Wiley Periodicals, Inc.     

Hyaluronan, CD44 and its variant exons in human trophoblast invasion and placental angiogenesis

Both hyaluronan and one of its receptors, CD44, can be demonstratedin the early human conceptus and in placental stroma. The variantsof CD44 resulting from variable exon splicing are found in metastasizinghuman malignancies and are also involved in hyaluronan uptakeand degradation. The resulting hyaluronan fragments are knownto be highly angiogenic. We postulated that the self-limitedprocess of trophoblast invasion of the uterine decidua resultsin part from the strategy of alternative splicing of CD44, similarto that used by invasive cancer cells in the course of metastaticspread and possibly angiogenesis. Monoclonal antibodies specificfor CD44s and for an exon expressed during metastatic tumourprogression, CD44v7, were used to examine this hypothesis. Inthis study we found human trophoblasts, for the first time,to express CD44. Intermediate trophoblasts of first and secondtrimester exhibited the standard form of CD44 while extravilloustrophoblasts, which are responsible for the invading characteristicsof the placenta, were positive for the alternatively splicedform, the CD44v7–8. Moreover, in the case of placentaaccreta there was a prominent membrane staining of the trophoblaststhat were embedded in the fibrin layer over the myometrium.The highly metastatic choriocarcinoma cells also expressed CD44v7–8.We propose, therefore, that the invading trophoblasts utilizethe alternatively splicing machinery. These cells retain theirinvasive capabilities through the permissive ECM by carryingthe CD44v7–8 isoform, which binds weakly to hyaluronanand thus prevents it from being degraded by intracellular hyaluronidase. CD44/hyaluronan/invasion/placentaArophoblast     

Production and secretion of thioredoxin from transformed human trophoblast cells

Thioredoxin is a redox active protein which has been implicated in reproductive processes. In this study we investigated the intracellular production and extracellular secretion of placental thioredoxin by human cytotrophoblast cell lines which were used as in-vitro model systems. Results clearly demonstrated that thioredoxin is not only synthesized by these cells but is also secreted and that while the intracellular thioredoxin is present only as a 12 kDa form, it would appear that the extracellular thioredoxin is present in two forms, a predominant 12 kDa form accompanied by a lower amount of a 10 kDa form. The observed localization and possible secretion of thioredoxin at the feto-maternal interface suggest important roles for this protein during pregnancy. Intracellular thioredoxin may be involved in the prevention of cellular damage due to oxidative stress whereas extracellular thioredoxin may act to integrate the actions of the cytokine network operating at the feto-maternal interface thereby assisting with implantation and the successful establishment of pregnancy.      

Distribution and characterization of anionic sites in trophoblast and capillary basal laminas of human placental villi

The distribution of anionic sites was studied in the trophoblastic and fetal capillary basal laminas of developing human placental villi with the cationic stain ruthenium red. At 7-12 weeks of gestation the trophoblastic basal lamina (TBL) contained ruthenium red-positive granules in a quasi-regular array throughout the lamina densa or sometimes concentrated at the interstitial surface of the lamina densa. The capillary basal lamina (CBL) (and anionic sites) were not present at this age. Anionic sites were also associated with collagen or reticular fibrils. At term, the TBL was largely devoid of anionic sites except for some distributed along its interstitial surface. The CBL was present in later gestation and sometimes had arrays of anionic sites. In order to characterize the anionic sites, minced pieces of villi were incubated in the presence or absence of either chondroitinase ABC, heparitinase, neuraminidase, or Streptomyces hyaluronidase in appropriate buffer systems. Incubation of early villi with heparitinase resulted in the disappearance of the TBL-associated sites. Chondroitinase ABC appeared to reduce staining of collagen-associated sites. In term villi, heparitinase removed those few sites still associated with the TBL but did not affect sites associated with the CBL or collagen. Chondroitinase ABC resulted in the disappearance of all anionic sites. In later gestation, a number of developmentally important macromolecules are transported across the trophoblast and enter the fetal capillaries. We conclude that the absence of an array of polyanionic sites from the term placenta TBL and the reduction in the amount of extracellular matrix intervening between the trophoblast and capillaries are adaptations to enhance the exchange of macromolecules across the placenta.     

Production of interferons during experimental African trypanosomiasis.

African trypanosomiasis is associated with profound changes in the function of the immune system. In this study we find that alpha/beta and gamma interferon (IFN) are released into the serum of mice infected with Trypanosoma brucei. The parasite-induced rise in serum IFNs is associated with a detectable parasitaemia, but the serum IFN peak precedes the peak parasitaemia in some cases. Unlike other protozoan interferon inducers, no parasite-dependent IFN production was observed in the pre-patent period of infection; while the most virulent clone induced very high IFN levels, no clear difference in stimulation was noted in the first waves of semi-acute and chronic T. brucei clones. However, subsequent IFN augmentation more closely reflected the host parasite load and virulence of infection. The nature of the stimulatory parasite component is as yet unknown, and the parasite surface glycoprotein had no effect on serum IFN. Injection of large quantities of lethally irradiated, but intact organisms did not stimulate IFN production; however this treatment significantly impaired antibody responses to the heterologous antigen SRBC. This suggests that the more severe effects of an actual trypanosome infection are required for induction of IFN synthesis, and that the presence of measurable serum IFN is not a prerequisite for parasite-induced suppression of host antibody responses.