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Hemomonochorial placentation occurs in diverse species. We have examined placental glycosylation in five widely separated mammals with this type of placentation--lesser hedgehog tenrec (Echinops telfairi), spotted hyena (Crocuta crocuta), nine-banded armadillo (Dasypus novemcinctus), human (Homo sapiens) and guinea pig (Cavia porcellus)--in order to assess whether evolutionary convergence to the hemomonochorial state is accompanied by a similar convergence of glycan expression. Placentae from 2 E. telfairi, 3 C. crocuta, 1 D. novemcinctus, 4 womenand 1 C. porcellus were fixed and processed into epoxy resin. Binding of twenty-three lectins was assessed using a semiquantitative ranking system. The trophoblast apical/microvillous membrane of all five species showed marked similarities in glycosylation. In the N-linked series, there were abundant bi/tri-antennary complex chains, while the non-bisected variants were much scarcer. All species had plentiful N-acetyl lactosamine sequences; at chain termini, binding to Galbeta1,4GlcNAc and Galbeta1,3GalNAc sequences was greatly enhanced after neuraminidase treatment. In all species, terminal NeuNAcalpha2,3 residues were detected. The tenrec had unusually abundant terminal N-acetyl galactosamine. The basal plasma membrane/basal lamina showed glycosylation patterns distinct from the microvillous membrane in each case, indicating chemical diversity of the two opposite faces of trophoblast. Similar classes of glycan at the hemochorial interface suggest conservation of function. The observed lectin binding patterns suggest broad similarities of glycosylation that may have arisen by convergent evolution., (Copyright 2007 S. Karger AG, Basel.)

The hyena placenta is unique among carnivores in being hemochorial. It also has areas of erythrocyte uptake that differ from those seen in more commonly studied carnivores. The availability of timed midterm pregnancies made it possible to examine the organization of the placenta, the distribution of regions of columnar trophoblast and the nature of the heterophagous and hemophagous regions in well-preserved material. The labyrinth of the placenta is orderly arranged with periodic primary villi that have a surface of syncytial trophoblast. Secondary projections from the primary villi anastomose extensively. The junctional extreme of each primary villus forms an expanded tip covered by columnar trophoblast cells. Marginal to the hemochorial placenta, the paraplacenta is exposed to varying amounts of extravasated maternal blood which the cytotrophoblast cells ingest and destroy. Columnar trophoblast cells at the tips of the primary villi, in the villi of paraplacental margins of the labyrinth, in isolated patches within the labyrinth and beneath the allantochorionic plate are all capable of and are variably involved in erythrophagocytosis. Variations in the structure and the lectin histochemistry of the columnar trophoblast cells indicate variation depending on the exposure of these cells to erythrocytes and/or other histotrophic materials. The widespread distribution of the hemophagous regions suggests a nutritive function in addition to an iron transfer function of the columnar cytotrophoblast.

The otter shrews are members of the subfamily Potamogalinae within the family Tenrecidae. No description of the ovaries of any member of this subfamily has been published previously. The lesser hedgehog tenrec, Echinops telfairi, is a member of the subfamily Tenrecinae of the same family and, although its ovaries have not been described, other members of this subfamily have been shown to have ovaries with non-antral follicles. Examination of these two species illustrated that non-antral follicles were characteristic of the ovaries of both species, as was clefting and lobulation of the ovaries. Juvenile otter shrews range from those with only small follicles in the cortex to those with 300- to 400-microm follicles similar to those seen in non-pregnant and pregnant adults. As in other species, most of the growth of the oocyte occurred when follicles had one to two layers of granulosa cells. When larger follicles became atretic in the Nimba otter shrew, hypertrophy of the theca interna produced nodules of glandular interstitial tissue. In the tenrec, the hypertrophying theca interna cells in most large follicles appeared to undergo degeneration. Both species had some follicular fluid in the intercellular spaces between the more peripheral granulosa cells. It is suggested that this fluid could aid in separation of the cumulus from the remaining granulosa at ovulation. The protruding follicles in lobules and absence of a tunica albuginea might also facilitate ovulation of non-antral follicles. Ovaries with a thin-absent tunica albuginea and follicles with small-absent antra are widespread within both the Eulipotyphla and in the Afrosoricida, suggesting that such features may represent a primitive condition in ovarian development. Lobulated and deeply crypted ovaries are found in both groups but are not as common in the Eulipotyphla making inclusion of this feature as primitive more speculative., (Copyright 2005 S. Karger AG, Basel.)

A specialized subset of invasive embryonic cytotrophoblast cells gains access to maternal uterine arteries early in the gestation of higher primates. These cells continue to migrate extensively within the lumina of spiral arteries, converting them to the highly modified uteroplacental arteries of pregnancy. Although trophoblast cell-mediated modifications are considered critical to the progress of normal pregnancy, few studies have addressed the cellular interactions between maternal arteries and embryonic cells in situ. Macaque placentas and endometrial tissues were collected from 12 animals from day 14 of gestation (blastocyst implantation begins on day 9) to day 49. Standard indirect immunoperoxidase methods were used to identify matrix metalloproteinases (MMP-1, MMP-3, MMP-9), cathepsin B, cathepsin D, platelet-endothelial cell adhesion molecule, cytokeratins, smooth muscle actin, CD68, and factor VIII-related antigen. Cytotrophoblast cells were located deep within spiral arteries in each of the specimens examined. In some examples tightly packed clusters of cytotrophoblast occluded the lumina of invaded arteries. Initial penetration of arterial tunica intima was revealed by discontinuities in the staining pattern for factor VIII and cytotrophoblast intrusion was indicated by cytokeratin staining of the trophoblast cells. Continued cytotrophoblast intrusion into the tunica media was apparent by gaps in the smooth muscle. MMP-1, MMP-3, and MMP-9 were localized within intraluminal and intramural cytotrophoblast. By contrast, neither cathepsin B nor cathepsin D were present, although both were seen in uterine macrophages and stromal cells. Upon reaching the surrounding uterine stroma the cytotrophoblast cells ceased migration. As cytotrophoblast accumulated in the arterial wall the vascular lumen expanded. Evidence of cell death was rarely encountered in associated maternal or embryonic tissues. We conclude that intra-arterial cytotrophoblast cells express several proteinases with substrate specificities sufficient to permit independent remodeling of the extracellular matrix comprising uterine artery walls. The remodeling of the arteries, which involves extensive displacement of maternal endothelium and smooth muscle, in addition to degradation and synthesis of extracellular matrix, is accomplished with little evidence of cell death or loss of the integrity of the arteries. This process provides an interesting example of cooperation between different types of interacting tissues from genetically distinct individuals.

Fetal trophoblast cells invade endometrial blood vessels and gain access to maternal blood within two days after the onset of blastocyst implantation in macaques. Soon thereafter, cytotrophoblast cells migrate well into the lumina of arteries and subsequently invade arterial walls. Using electron microscopy and light microscopy we investigated the interactions between invasive cytotrophoblast cells and the cellular and extracellular components in the walls of endometrial arteries. The placentas and adjacent endometrium of 22 macaques (GD 17 to term) were examined. Spiral arteries containing migratory cytokeratin-labeled cytotrophoblast cells were identified at all stages examined. Early modification of each artery showed that a plug of intraluminal cytotrophoblast cells temporarily filled the arterial lumen in the vicinity of the trophoblastic shell. Distal to this plug the group of cells tapered as a continuous mass, filling only a portion of the lumen. Endothelial cells were displaced from their basal lamina by closely apposed cytotrophoblast cell processes. Soon thereafter these processes penetrated the basal lamina and achieved contact with smooth muscle cells of the tunica media. As cytotrophoblast cells infiltrated the arterial wall they hypertrophied and secreted extracellular matrix, thereby differentiating into intramural cytotrophoblast. The patent lumen of the artery was reestablished concomitant with the migration of intraluminal cytotrophoblast cells through the arterial tunica intima and into the tunica media. The presence of clusters of cytotrophoblast cells in the arterial wall results in discontinuity of the tunica media and dispersion of the smooth muscle. The combined changes result in expanded circumferences of invaded arteries as well as diminished ability to contract. In portions of arteries adjacent to the trophoblastic shell cytotrophoblast usually occupied the entire perimeter and thickness of the artery wall, while in areas distal only a portion of the wall was invaded. Despite extensive arterial modification, evidence of cell death among the fetal and maternal tissues involved was rare. By later gestation only a few intraluminal cytotrophoblast cells were seen. Intramural cells were surrounded by a thick layer of matrix, but maintained contact with adjacent cells through cytoplasmic processes, some of which formed gap junctions. Maternal cellular and connective tissue elements were excluded from the cytotrophoblast-matrix pads and the cytotrophoblast cells retained attributes of glycoprotein producing cells to term. Spiral arteries were modified well into the spongiosum layer of the endometrium, and some were modified into the myometrium.

The interaction of cytotrophoblast with maternal endometrium, especially endometrial blood vessels, was examined in macaque gestational stages between 2 and 8 days after the onset of implantation. Serial sectioning of these early implantation sites allowed immunostaining of consecutive sections with a number of different antibodies, facilitating cell identification. In the earliest implantation site, immunostaining showed that antibody to cytokeratin stained cytotrophoblast, syncytial trophoblast, epithelial plaque and endometrial gland cells. However, only those cytotrophoblast cells near the maternal-fetal border and within vessels showed surface staining for neural cell adhesion molecules and only syncytial trophoblast showed SP1 reactivity. Even at this early stage cytotrophoblast filled the lumen of superficial arterioles, whereas dilated venules contained only a few cytotrophoblast cells. In later stages endovascular cytotrophoblast not only plugged many spiral arterioles but also migrated into the walls of these arterioles, and progressed into deeper coils. Displacement of endothelial cells and disruption of vessel walls were illustrated with antibody to factor VIII, TGF alpha, and desmin. Clusters of cytotrophoblast cells at the fetal-maternal interface tended to bypass clusters of epithelial plaque cells and larger clusters of maternal fibroblasts, but readily entered all vascular spaces. Consequently the vascular system constituted a major pathway of invasion, although the arterioles were the only component substantially invaded beyond the trophoblastic-shell/endometrial border.

Adhesion of trophoblast of the blastocyst to and penetration into the uterine epithelium, invasion into the maternal vessels and endometrial stroma, and establishment of the basic organization of the placenta all occur within the first week following the initiation of implantation in the human, macaques and several other primates. The cellular rearrangements and interactions of trophoblast with endometrial epithelial cells and stroma were studied during this preiimplantation stage in macaques. At the early lacunar stage, 1-2 days after the initiation of implantation, both cytotrophoblast and syncytial trophoblast can be found at the maternal surface of lacunae. As the lacunar stage advances, both syncytial trophoblast and cytotrophoblast are found throughout the implantation site including in septae partitioning lacunae, but syncytial trophoblast lines most of the lacunae and forms the confluence with the maternal vessels. Indentations of fetal mesenchyme and accumulation of cytotrophoblast cells within the septae occur rapidly. Over a period of about 2 days, clusters of cytotrophoblast cells pass beyond the syncytial trophoblast at the maternal surface as the anchoring villi, establishing the trophoblastic shell. The bypassing of epithelial plaque cells by cytotrophoblast cells and elimination of stromal matrix between such clusters verifies that this is a progressive invasion rather than simply superficial growth. Concomitant with establishment of the trophoblastic shell there is extravasation of blood, necrosis of plaque cells, and massive invasion of vessels by cytotrophoblast resulting in a necrotic zone between the trophoblastic shell and endometrium. It is concluded that only in the brief period of time of establishment of the trophoblastic shell is the full invasive potential of cytotrophoblast realized and that only at this stage does cytotrophoblast demonstrate the type of invasive and migratory behavior which has been achieved with isolated human cytotrophoblast cells in vitro.

Placentation in the green monkey (Cercopithecus aethiops) from gestational ages of 18-30 days was studied by light and electron microscopy. The placentae show many features in common with that of the rhesus monkey and other related macaques. Implantation is superficial and bidiscoid, and the placenta is classified as villous and hemomonochorial. Epithelial plaque formation and a moderate stromal reaction occur. Plaque cells are large, with massive deposits of glycogen and marginal pleomorphic dense bodies. Observations further support the concept that chorionic blood vessels, angioblastic and mesenchymal cells are not derived by delamination from cytotrophoblast. Capillary formation is by attachment of one or more mesenchymal cells and/or their processes. Embryonic blood was not observed in the villi until after day 22.

In the near-term paraplacental extravasate zone of the domestic cat, which includes paraplacental junctional areas and marginal hematomas, the uterine epithelium becomes highly proliferative, especially on the folds or protrusions, This hyperplastic surface and the adjacent connective tissue become symplasmic, degenerate and eroded, eventually resulting in maternal vascular breakdown. The uterine lumen thus contains maternal blood in addition to sloughed uterine cells. The adjacent cytotrophoblast apparently phagocytoses and ingests this material. Various stages of phagolysosomes containing erythrocytes of variable density are seen in these cells, as are myelin-like structures, ferritin, glucogen and lipid droplets. Crystalloid iron deposits are seen in the basal lamina and connective tissue subjacent to the hemophagous cytotrophoblast.

The syncytial trophoblast has previously been shown to have minimal intrasyncytial galactosyltransferase activity at term. The biochemical and autoradiographic study reported here shows that the microvillous surface of term human placental syncytial trophoblast has a galactosyltransferase activity capable of transferring 3H-galactose from uridine diphosphate-D-galactose-1-3H to trichloroacetic-acid precipitable, endogenous acceptors. This capability of resynthesizing cleaved galactose moieties in the glycocalyx, without dependence on cytoplasmically located galactosyltransferases, would allow for reinstatement of the original surface molecular configuration without requiring synthesis and insertion of a completely new membrane molecule. It is suggested that the surface galactosyltransferase might function to repair damage to syncytial trophoblast glycocalyx induced by the enzymes in maternal blood.

The entire chorioallantoic paraplacenta of the near-term domestic cat shows cytological evidence of maternal-fetal substance transfer, but there are probably qualitative and quantitative differences and increased transfer progressing from the poles to the placental border. In the interfetal polar zone, the cytotrophoblast remains proliferative, is squamous to columnar and shows only a small amount of pinocytotic activity. However, in the adjacent ring of the free polar zone, where the trophoblast is cuboidal or columnar and loosely apposed to the endometrium, material of uterine origin (histotroph) appears to be transferred from the columnar uterine surface epithelium to the trophoblast. The remaining paraplacenta extends from the polar zone to the girdle, forming an extravasate zone that is composed of hematomal areas containing extravasates of maternal blood, and interposed relatively unmodified paraplacental junctional areas. The trophoblast in this hematomal area is columnar and constitutes chorionic villi which face a surface formed by maternal symplasma, degenerative uterine tissue, and disintegrating blood, thus providing 'histolytic' and 'hemolytic' embryotroph to this hemophagous cytotrophoblast. When the junctional area is just forming, a cuboidal cytotrophoblast consisting of resting cells, opposite to the endometrial folds, appears to have both the capacity for digestion and transformation. However, during the recessive junctional stage, this cytotrophoblast is proliferative and absorptive only. Both sources of cuboidal trophoblast may give rise to syncytial trophoblast, which is presumably the invasive form that erodes the uterine tissue, including the interstitial layer and maternal endothelium. As long as there is firm junction of the fetal and maternal tissues, there appears to be a transfer of secretion products to form embryotroph. However, with the regression of the junctional area more products of cell breakdown (histolytic products) are absorbed by the cuboidal cytotrophoblast in the chorionic fossae.