The development of the baboon (Papio sp.) placenta during the fetal period of gestation

Baboon placentae ranging in age from 45 to 175 days were studied for progressive changes in their morphology. The baboon embryo implants superficially and develops a single, discoid, villous hemochorial placenta. The remainder of the chorion is membranous and a true decidua capsularis is not formed. A lobular structure is developed during the fetal period. At 45 days the amnion only partially fills the chorionic cavity but by 60 days has expanded and the two membranes are directly contiguous. Trophoblastic tissue is of two types: cytotrophoblast and syncytiotrophoblast. Cytotrophoblast is found primarily in the chorionic plate and cytotrophoblastic shell. The chorionic villi are originally composed of a double layer of trophoblast; cytotrophoblast internally and syncytiotrophoblast externally. The cellular layer gradually disappears so that by full-term the villous walls are formed by a single layer of syncytiotrophoblast. Hofbauer cells are common within the villous cores, diminishing in number toward term. Large amounts of collagenous connective tissue develop in the chorionic plate and in the villi, and fibrin and fibrinoid materials accumulate in the basal plate and anchoring villi. The endometrium is decidualized to its greatest extent by the beginning of the fetal period and undergoes only minor cytological alteration throughout the remainder of gestation.     

The Persistence of “Steroid-synthesizing Cell” Antigen and 3β-hydroxysteroid Dehydrogenase in Organ Cultures of Normal Human Placentas

The syncytiotrophoblast of chorionic villi of normal human mid-pregnancy placenta maintained in organ culture in a chemically-defined medium loses its various differentiated characters at different rates. Although certain oxidative and hydrolytic enzymes are retained for periods of 1-6 weeks and human placental lactogen persists in the cytoplasm for about 2 weeks, the enzyme 3β-hydroxysteroid dehydrogenase and the “steroid-synthesizing cell” antigen are lost after culture for 1 and 2 days respectively. These two characters reappear in cytotrophoblast that is newly differentiated during culture. It is concluded that organ cultures of this type will not be suitable for metabolic studies on the biosynthesis of steroid hormones by the placenta.     

Differentiation of the chorionic plate of the placenta: Cellular and extracellular matrix changes during development in the macaque

Background: The chorionic plate forms the fetal side of the placental disc, and its proper growth and development is important to the formation of a normal placenta. The development and structure of the chorionic plate has received little attention. Therefore, we have conducted a developmental and immunohistochemical study of the chorionic plate of the macaque placenta. Methods: Conventional light and transmission electron microscopy techniques were used to study macaque placental tissues collected from 22 days of gestation to near term. Standard immunoperoxidase methods were used to identify type IV collagen, laminin, and fibronectin in paraffin sections. Results: Early in gestation the chorionic plate trophoblast consisted of an outer layer of syncytiotrophoblast and a single underlying layer of cytotrophoblast. Beginning at about 100 days of gestation, the cytotrophoblast layer became stratified. The cytotrophoblast cells also became surrounded by variable amounts of extracellular matrix containing type IV collagen, laminin, and fibronectin. Ultrastructurally, the matrix contained abundant 10–12 nm diameter microfibrils. During later gestation the syncytiotrophoblast had a tendency to separate from the cytotrophoblast. Conclusions: The chorionic plate of the macaque placenta undergoes several distinctive morphological changes over the course of gestation. During the period of rapid diametrical growth of the disc, the chorionic plate trophoblast consists of a layer of syncytiotrophoblast and a single layer of cytotrophoblast. During later gestation the cytotrophoblast layer stratifies at a time coincident with that at which diametrical growth of the disc slows. The cytotrophoblast cells of later gestation appear synthetically active and at least some of their products are extracellular matrix components that encapsulate many of these cells. These components include type IV collagen, laminin, fibronectin, and microfibrils. © 1994 Wiley-Liss, Inc.     

Serum protein distribution in hydatidiform mole. An immunohistochemical study

In normal placentas during the first trimester of pregnancy, the syncytiotrophoblast appeared to be immunoreactive to alpha-antitrypsin (alpha 1-AT), alpha 1-antichymotrypsin, albumin, IgG, and transferrin. The underlying cytotrophoblast was negative for these same serum proteins. In the hydatidiform mole, these findings were profoundly different. The syncytiotrophoblast lost its immunoreactivity to albumin, IgG, transferrin, and, less frequently, to alpha 1-AT. Furthermore, the underlying cytotrophoblast became immunoreactive to albumin, alpha 1-AT, IgG, transferrin, ferritin, orosomucoid, and, sometimes, to alpha 1-antichymotrypsin. This altered immunohistochemical pattern suggested a notable change in the pinocytotic activity of the trophoblast in the placenta during molar degeneration. The absence of pinocytosis in the syncytiotrophoblast for several proteins can be explained by the partial loss of specific membrane receptors. The contemporaneous appearance of numerous serum proteins in the cytotrophoblast could indicate an activation, not only proliferative, but also functional, in the germinative cytotrophoblast. Diagnostically, this histochemical finding in the hydatidiform mole, which was quite different from that seen in normal placentas during the first trimester of pregnancy, could provide additional evidence concerning trophoblastic abnormalities in the chorionic villi during molar degeneration.     

The effects of mifepristone (RU486) on prostaglandin dehydrogenase in decidual and chorionic tissue in early pregnancy

Prostaglandin dehydrogenase is the main inactivating enzymefor prostaglandins and therefore controls local levels of prostaglandins.Since there is some evidence that the expression of this enzymeis under progesterone control it is reasonable that one of theeffects of antiprogestin is to reduce the concentration of thisenzyme and thus increase the effective concentration of prostaglandinwithin tissue. We have investigated the amount of enzyme activitywithin decidua and chorionic villi from women receiving theantigestagen mifepristone (RU486) 12, 24 and 36 h prior to surgicalabortion, and examined the effect on tissue concentrations ofprostaglandin dehydrogenase. Women receiving mifepristone inall groups had a significant reduction in concentration of prostaglandindehydrogenase enzyme in decidual tissue. There was also a markedreduction in prostaglandin dehydrogenase in decidual cells followingRU486, as demonstrated by immunochemical methods. At this stageof pregnancy, prostaglandin dehydrogenase was present in abundancein cytotrophoblast cells of chorionic villi but virtually absentfrom syncytiotrophoblast. In chorionic villi after RU486 administrationin vivo, there were no obvious differences in prostaglandindehydrogenase distribution or reactivity in the majority ofcases.     

解聚素和金属蛋白酶19在早孕绒毛中的表达及其与自然流产的相关性研究

目的通过对解聚素和金属蛋白酶19（ADAM19）在早孕自然流产和正常早孕绒毛中表达的研究,探讨其在妊娠过程中的作用及其与流产的关系。方法早期妊娠和难免流产的绒毛组织各30例,应用免疫组织化学链霉素抗生物素-过氧化物酶法,检测ADAM19的表达。结果在正常妊娠的绒毛中,ADAM19主要表达在细胞膜上,在细胞滋养细胞、细胞滋养层细胞柱中均有较强的表达,表达强度均强于合体滋养细胞;在流产组绒毛中,ADAM19在细胞滋养细胞、细胞滋养层细胞柱中的表达均明显低于正常绒毛组,二者比较差异有统计学意义（P〈0.01）。结论ADAM19在妊娠早期滋养细胞的侵入中有重要的作用,早期流产的发生与绒毛中ADAM19水平下降有关。     

Vasculogenesis in early human placental villi: an ultrastructural study

In this study we examined the chorionic villi of 5 normal human placentas at 12–14 weeks of gestation ultrastructurally with regard to differentiation of the vascular components. The aim of the present report is to discuss the factors influencing vasculogenesis (in situ formation of blood vessels) at the ultrastructural level.

Our observations have led us to think that the cytotrophoblast influences vasculogenesis in human chorionic villi. Mesenchymal-preendothelial cell groups were always found in very close association with the cytotrophoblast at the periphery of the villi, forming blood vessels. The cytotrophoblast probably attracts mesenchymal cells towards the margin of the villi by secreting vascular endothelial growth factor (VEGF). Once cells attach to the trophoblastic basement membrane they begin to differentiate into endothelial cells. This close structural relation between two cell types (cytotrophoblast and mesenchymal cells) may not be the only mechanism controlling vasculogenesis, but it seems to be one of the factors influencing the differentiation of mesenchymal cells into the endothelial cells of blood vessels in early human chorionic villi.     

Expression of merosin, a tissue-specific basement membrane protein, in the intermediate trophoblast cells of choriocarcinoma and placenta

Merosin is a novel tissue-specific basement membrane-associated protein found in basement membranes of trophoblast, striated muscle and Schwann cells. In placental extracts, the immuno-reactivity for merosin was detected in a protein band of 80 kilodaltons, and a 65 kilodalton polypeptide fragment of merosin could be isolated from proteolytic digests of placenta. In the present study, we describe the expression of merosin in human choriocarcinomas and normal placentas using immunoperoxidase staining of paraffin-embedded tissues. All five choriocarcinomas studied show immunoreactivity for merosin. Tumor cells, exhibiting the morphology typical of the intermediate trophoblast, stained distinctly for merosin. The cytotrophoblast and syncytiotrophoblast cells in these tumors showed negligible or no staining. In second and third trimester human placentas, merosin immunoreactivity was found in large extravillar mononuclear trophoblast cells of the basal plate as well as in the trophoblast basement membranes of the chorionic villi. The results indicate that merosin is mainly expressed in the intermediate trophoblast cells of both neoplastic and normal origin, whereas almost no expression is seen in cytotrophoblast and syncytiotrophoblast. Consequently, it is suggested that the intermediate trophoblast may represent a third, independently differentiated trophoblastic cell type.     

Expression of alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein and the 39-kd receptor-associated protein in human trophoblasts.

The alpha 2-macroglobulin receptor/low density lipoprotein receptor-related protein (alpha 2MR/LRP) and its 39-kd receptor-associated protein (RAP) were identified by indirect immunofluorescence in human extravillous and villous trophoblast cells at different stages of pregnancy. The alpha 2MR/LRP was detected in invading trophoblast cells and in some instances these invading cells did not express RAP. In chorionic villi of first and second trimester placenta, alpha 2MR/LRP and RAP were found in cytotrophoblast and syncytiotrophoblast. With advancing pregnancy alpha 2MR/LRP became primarily localized to the apical surface of the syncytiotrophoblast, while RAP was present in the cytoplasm. Villous cytotrophoblast cells lost both proteins by the third trimester. Isolated cytotrophoblast cells that undergo spontaneous differentiation into syncytiotrophoblast in culture increased expression of both alpha 2MR/LRP and RAP. With syncytium formation, alpha 2MR/LRP became localized to the plasma membrane in cup-like structures. Changes in the mRNAs for alpha 2MR/LRP and RAP paralleled the changes in relative abundance of the proteins assessed by immunofluorescence. cAMP treatment suppressed both alpha 2MR/LRP and RAP in the cultured trophoblasts, but alpha 2MR/LRP was reduced to a greater extent than RAP. We conclude that alpha 2MR/LRP and RAP are developmentally regulated in human trophoblast cells, that the temporal and spatial patterns of expression of these proteins can be dissociated, and that cAMP modulates both alpha 2MR/LRP and RAP in human trophoblast. The patterns of alpha 2MR/LRP and RAP expression in trophoblast cells are consistent with roles for the receptor in trophoblast invasion and transport of molecules across the syncytiotrophoblast.     

Morphological interactions of human first trimester placental villi co-cultured with decidual explants

Abnormalities of pregnancy such as pre-eclampsia and intrauterinegrowth retardation are characterized by shallow trophoblasticinvasion of the placental bed, the precise molecular pathophysiologyof which remains to be fully elucidated. An in-vitro model involvinga co-culture of first trimester placental villi and deciduaparietalis explants (of 8-12 weeks gestation) was developedand used to characterize the migration and local invasion oftrophoblast cells. Trophoblast proliferation (confirmed by Ki-67immunostaining), differentiation and loose attachment of placentalvilli to the underlying decidual epithelium or stroma occurredwithin the first 24 h of co-culture. This was followed by erosionof the syncytial layer of the placental villi and commencementof a progressive cytotrophoblast invasion after 48 h of co-culture,which continued until 120 h, when the experiments were terminated.E-cadherin was expressed at the interfaces between trophoblastcells within the villi, but expression of this adhesion moleculeseemed to be down-regulated in the invasive trophoblast cells.Our results suggest that the model could be useful in investigatingthe factors that control early human placentation and the feto-maternalinterface.     

Anaphase lag as the most likely mechanism for monosomy X in direct cytotrophoblasts but not in mesenchymal core cells from the same villi.

A 36 year old white female was referred for chorionic villus sampling for advanced maternal age. Direct (cytotrophoblast) preparations of chorionic villi were 45,X, but cultured mesenchymal core cells from the same villi were 46,XX. Study of embryonic and extraembryonic tissues showed the aneuploidy to be limited to cytotrophoblasts from specific placental sites. In aggregate, the cytogenetic findings can best be explained by anaphase lag during development of the cytotrophoblast, suggesting that this cytological mechanism and not non-disjunction is responsible for the common occurrence of monosomy X in villi.     

BCL-2 expression in the human placenta and its correlation with fibrin deposits

The human placenta performs numerous functions during its limited lifespan and its survival is a necessary prerequisite for fetal nutrition, even in unfavourable conditions. BCL-2 is a proto-oncogene implicated in the regulation of cell death and survival without affecting cell proliferation. An extracellular matrix molecule involved in the reparative and degenerative processes in the human placenta is fibrin. We have analysed by immunohistochemistry the expression of BCL- 2 and correlated it with fibrin deposits in placental tissues. In first and third trimester placentas BCL-2 was expressed in the syncytiotrophoblast. Only a few mesenchymal villi (first trimester) or terminal villi (third trimester) showed no staining in the syncytiotrophoblast. Villous cytotrophoblast, mesenchymal cells of the villous cores and extravillous cytotrophoblast of cell columns and cell islands were all negative for BCL-2. BCL-2 expression was enhanced in the syncytiotrophoblast overlying subtrophoblastic fibrin deposits. However, discontinuities and/or variations in intensity of BCL-2 expression characterized not only the villi showing perivillous fibrinoid but also those villi with a massive presence of fibrinoid in their cores. These data suggest that BCL-2 may be necessary for the preservation of the placenta during gestation as well as for the reparative processes of the trophoblast.      

Ultracytochemical localization of adenylate cyclase in early human placenta

A cytochemical study was carried out on adenylate cyclase (AC) activity in the early human placenta. Samples of placental villi were incubated in a medium containing adenylyl-imidodiphosphate (AMP-PNP) as specific substrate. No AC reaction product was encountered in placenta villi taken at 5 and 7 weeks of pregnancy. AC activity appeared at 9 weeks. At 9 and 10 weeks, AC reaction product was localized on the basal plasma membranes and on apposed plasma membranes of the Langhans cytotrophoblast. At 11 weeks AC activity was also clearly visible on Langhans cytotrophoblast and syncytiotrophoblast apposed plasma membranes. No AC reaction product was ever detected on the syncytiotrophoblast microvillar membrane. These results are in agreement with biochemical studies that localize AC on the villous trophoblast plasma membranes associated with the fetal circulation.     

The ultrastructure of early implantation in the marmoset monkey (Callithrix jacchus)

Summary The ultrastructural morphology of the initial stages of implantation in the marmoset monkey (Callithrix jacchus) was studied in pregnant monkeys at known time intervals after ovulation. The earliest samples, obtained 13 days after ovulation, displayed both cytotrophoblast and syncytiotrophoblast. The cytotrophoblast was restricted to the blastocoel, whilst syncytiotrophoblast intruded to the endometrial basal lamina. At later stages, days 16 and 19 after ovulation, both cytotrophoblast and syncytiotrophoblast had extended laterally around the uterus, and the syncytiotrophoblast also extended deeper into the maternal tissnes. The mesoderm layer was first discernible at 19 days after ovulation. At 23 days after ovulation the syncytiotrophoblast surrounded the maternal blood vessels entirely. In this study syncytiotrophoblast was not observed to breach the maternal blood vessels, even at 31 days after ovulation. Early cytotrophoblast columns could be seen at 31 days after ovulation. The endothelial cells lining the maternal blood vessels displayed hypertrophy from the earliest stages (day 13) onwards, although a true decidual response was only observed in samples of 23 and 31 days after ovulation.     

DEVELOPMENTAL REGULATION OF TISSUE TRANSGLUTAMINASE DURING HUMAN PLACENTATION AND EXPRESSION IN NEOPLASTIC TROPHOBLAST

The expression of tissue transglutaminase (tTG) was studied during the formation of the normal human placenta and in molar pregnancies and choriocarcinoma, in order to correlate its expression with the functional characteristics of the recognized trophoblast cell types. tTG expression was found to be developmentally regulated. Before 6–7 weeks' gestation, only the chorionic villous cytotrophoblast expresses tTG. Thereafter the overlying syncytiotrophoblast becomes positive. tTG expression is gradually downregulated in the intermediate trophoblast cells emerging from the tips of the chorionic villi invading the uterine tissue. In the decidual wall, the intermediate trophoblast does not express tTG, whereas scattered syncytial cells, the placental bed giant cells, express tTG. Villi from complete hydatidiform mole (CHM) show tTG expression in both the cyto- and the syncytiotrophoblast. The intermediate trophoblast cells from CHM show heterogeneous tTG expression, with a majority of negative cells, whereas extravillous syncytia always express tTG. In choriocarcinoma, the tumour cells show heterogeneous tTG expression, with a majority of positive cells. Analysis of tTG protein and mRNA in placental extracts by Western and Northern blotting did not provide evidence for expression of the truncated form of tTG found in some cell types. The regulated expression of tTG in the normal placenta suggests that the enzyme is involved in important trophoblastic functions and may participate in the control of invasion. © 1997 by John Wiley & Sons, Ltd.     

松驰素受体LGR7在人妊娠绒毛及胎盘组织中表达的研究

目的探讨松驰素受体LGR7在妊娠过程中的作用。方法应用免疫组织化学（免疫组化）链霉素抗生物素一过氧化物酶法和RT—PCR技术，对56例妊娠绒毛和胎盘组织中的LGR7检测。结果在早期妊娠的绒毛中，LGR7免疫反应呈现强阳性。主要分布于绒毛滋养层细胞的细胞膜上，在细胞滋养层细胞、合体滋养层细胞及滋养层细胞柱中都有较强的信号，并可见细胞滋养细胞的阳性反应明显强于合体滋养细胞；中、晚期妊娠的胎盘组织中LGR7呈现为强阳性或阳性，主要定位于合体滋养细胞的细胞膜上及毛细血管内皮细胞。免疫组化结果中，早期妊娠绒毛LGR7的强阳性率为90％，明显高于中晚期胎盘中的76．5％和73．7％，中、晚期妊娠分别与早期妊娠比较，差异均有统计学意义（P〈0．01）；但中期妊娠及晚期妊娠相比较，差异无统计学意义（P〉0．05）；早期妊娠的绒毛中LGR7mRNA的表达量为0．967±0．019，明显高于中晚期妊娠胎盘中的0．522±0．071和0．482±0．094，中、晚期妊娠分别与早期妊娠比较，差异均有统计学意义（P〈0．01）；中期妊娠及晚期妊娠比较，差异无统计学意义（P〉0．05）。结论LGR7可能是与妊娠早期滋养细胞侵袭有关的重要的因子，在妊娠的中晚期参与妊娠的维持。     

Expression of proliferation and apoptotic markers in human placenta during pregnancy

Trophoblast has unique properties in relation to its wide range of metabolic, endocrine and angiogenic functions. Trophoblastic cells invade endometrium and adjacent myometrium in a way that is imitated by malignant tumours. The aim of the present study was to analyse the expression of markers of proliferation and apoptosis in trophoblastic cells in normal human placenta during pregnancy. A total of 22 placentas, 12 of which were obtained from curettage and induced legal abortion and 10 placentas obtained from normal deliveries or caesarean sections were included in this study. Proliferation markers were strongly expressed in cytotrophoblast in early stages of gestation. In late term placentas, a distinct decrease in expression of these markers was observed. Syncytiotrophoblast was negative for proliferation markers in all placentas. Positive immunostaining for bcl-2, an anti-apoptotic marker, was observed only in syncytiotrophoblastic cells in first-trimester but also in third-trimester placentas. Cytotrophoblast and stromal mesenchymal cells of chorionic villi were negative for bcl-2. Expression of bcl-2 protein in syncytiotrophoblast may be one of the major factors preventing these structures from early cell death, which is indispensable for the maintenance of physiological pregnancy.     

Secretion of pro-apoptotic intron 4-retaining soluble HLA-G1 by human villous trophoblast

One major materno-fetal interface in the human placenta is constituted by the syncytiotrophoblast, in contact with maternal blood of the intervillous space, which derives from differentiation and fusion of the villous cytotrophoblast (vct). In the present work, we purified vct from term placenta by depleting HLA class I- and class II-positive cells. We found by RT-PCR that both soluble intron 4-retaining HLA-G1 (sHLA-G1) and HLA-G2 isoforms were transcribed in purified vct. Using different HLA-G-specific mAb, we demonstrated by intracellular flow cytometry, Western blotting and ELISA, that sHLA-G1 but no soluble HLA class Ia molecule was secreted by vct. We then purified sHLA-G1 from vct culture supernatant and found that it exhibited an unusual glycosylation pattern. Finally, we showed that such trophoblast-derived sHLA-G1 triggered specific apoptosis of activated CD8+ T cells. Taken together, these results demonstrated that vct did secrete functional sHLA-G1 in primary culture and suggested that, in vivo, sHLA-G1 might be an important immunomodulatory molecule controlling the activity of maternal immune effector CD8+ cells circulating in the blood that immerses chorionic villi.