Ultrastructure of the pre-implantation shark yolk sac placenta

During ontogeny, the yolk sac of viviparous sharks differentiates into a yolk sac placenta which functions in gas exchange and hematrophic nutrient transport. The pre-implantation yolk sac functions in respiration and yolk absorption. In a 10.0 cm embryo, the yolk sac consists of six layers, viz. (1) somatic ectoderm; (2) somatic mesoderm; (3) extraembryonic coelom; (4) capillaries; (5) endoderm; and (6) yolk syncytium. The epithelial ectoderm is a simple cuboidal epithelium possessing the normal complement of cytoplasmic organelles. The endoplasmic cisternae are dilated and vesicular. The epithelium rests upon a basal lamina below which is a collagenous stroma that contains dense bodies of varying diameter. They have a dense marginal zone, a less dense core, and a dense center. The squamous mesoderm has many pinocytotic caveolae. The capillary endothelium is adjacent to the mesoderm and is delimited by a basal lamina. The endoderm contains yolk degradation vesicles whose contents range from pale to dense. The yolk syncytium contains many morphologically diverse yolk granules in all phases of degradation. Concentric membrane lamellae form around yolk bodies as the main yolk granules begin to be degraded. During degradation, yolk platelets exhibit a vesicular configuration.     

Experimental manipulation of the rodent visceral yolk sac

The visceral yolk sac (VYS) is an especially important placental organ in the rodent because it is the primary source of exchange between the embryo and mother during early organogenesis before the chorioallantoic placenta circulation is established. The VYS is involved with nutritional, endocrine, metabolic, immunologic, secretory, excretory, and hematopoietic functions. The VYS also plays a role in steroid metabolism and interacts with a variety of blood-borne factors: parathyroid hormone, glucocorticoids, insulin, and vitamin D metabolites. The importance of the VYS during development is emphasized by the embryotoxicity resulting from exposure to agents which cause VYS dysfunction when administered to the pregnant animal during organogenesis. Several experimental procedures have provided useful information concerning a variety of VYS functions from early organogenesis to term: Culture of the Embryo, Fetal Incubation, Culture of the Fetus, Giant Yolk Sac, Short- and Long-Term Culture of the Yolk Sac, Modified Ussing's Chamber, Single or Double Diffusion Chamber, and the use of Heterologous Rodent Visceral Yolk Sac Antibodies. Since human yolk sac pathology has been associated with developmental toxicity and spontaneous abortion, it is important to discover whether there are some common functional roles among different mammalian species and to determine if other experimental animal models can be used to study the possible contribution of human yolk sac dysfunction to some human reproductive problems.     

Mineral metabolism in the developing turkey embryo--II. The role of the yolk sac.

1. Turkey embryos were incubated in ovo or in long-term shell-less culture (ex ovo) for 14, 18, 22 or 26 days, at which time the concentrations of zinc, copper, iron, manganese and calcium in yolk and yolk sac membrane were determined. 2. Yolk manganese and calcium concentrations increased during incubation in ovo while the concentrations of zinc, copper and iron declined. The concentrations of zinc, copper and iron in yolk from ex ovo embryos did not decline. Yolk calcium concentration increased during incubation ex ovo, although to a much lesser extent than that observed in ovo. 3. The concentration of zinc, copper and iron declined in yolk sac tissue during incubation in ovo whereas no decline was observed for yolk sac tissue from ex ovo embryos. Yolk sac calcium and manganese concentrations increased during incubation in ovo and ex ovo, although the increase in calcium concentration for ex ovo yolk sac was much smaller than that observed in ovo. 4. A peak corresponding to metallothionein (MT) which bound both zinc and copper was isolated from yolk sac cytosol on day 14 of incubation in ovo using gel-permeation column chromatography. 5. Further fractionation of the MT peak by anion exchange chromatography revealed three metal-binding peaks designated MT-1, MT-2a and MT-2b. The majority of the zinc was bound to MT-2a and MT-2b whereas most of the copper was bound to a single peak (MT-2b). 6. The concentrations of zinc and copper in yolk sac cytosol reached a maximum on day 14 of incubation in ovo and declined through to day 28 (hatching).(ABSTRACT TRUNCATED AT 250 WORDS)     

Oogenesis in the annelid Enchytraeus albidus with special reference to the origin and cytochemistry of yolk

In the annelid Enchytraeus albidus the ovary is composed of packets containing eight synchronously developing oocytes. Each oocyte in the packet is connected, via a bridge, to a common cytoplasmic mass. Developmental synchrony of oocytes within individual packets is probably related to the ooplasmic continuity. The young previtellogenic oocyte contains many polysomes, a few cisternae of smooth and rough endoplasmic reticulum, small Golgi complexes, and mitochondria. Many of the mitochondria are dumbbell-shaped and may thus represent division stages. Vitellogenesis is marked by the appearance of peripherally located lipid yolk and small, densely staining granules scattered throughout the ooplasm. There is an increase of smooth endoplasmic reticulum, mitochondria, and enlarged Golgi elements. Small multivesicular-like bodies, the early stages of developing yolk, are derived from the Golgi complex. The mature yolk sphere is bipartite and consists of (a) a variable number of dense spheres, the core bodies, which are produced in the ooplasm by the Golgi complex and which become embedded in (b) a dense matrix. The electron opaque tracer, horseradish peroxidase is incorporated into the oocyte and deposited in the matrix suggesting that this component of the yolk sphere is obtained by micropinocytosis. Enzyme digestions and various cytochemical techniques suggest that the core bodies are rich in carbohydrate, probably as glyco- or mucoproteins, and that the matrix is rich in lipid.     

Ultrastructural Peculiarities of the Embryogenesis of the Brittle Star Amphipholis kochii (Lutken, 1972)

This paper addresses morphogenetic processes and cell differentiation during embryogenesis of the brittle star Amphipholis kochii at the ultrastructural level. The radial cleavage is not strictly determined. Embryos are covered with a thick hyaline envelope and contain numerous yolk granules and small lipid drops. Blastulae feature a thick blastoderm with extensive intercellular cavities, which are retained in the crest epithelium of late gastrulae. Embryonic cells have single cilia with long cross-striated rootlets associated with the Golgi apparatus. Depolarized cells of the primary mesenchyme with a well-developed rough endoplasmic reticulum differentiate into sclerenchyme syncytium. Gastrulation occurs by invagination. Secondary mesenchymal cells emigrate from the archenteron tip to differentiate into amebocytes, which contain a well-developed Golgi apparatus and numerous mitochondria. The endoderm is formed of cubic cells with numerous yolk granules and rare microvilli. Flattened cells of the dorsal and ventral ectoderm contain a small amount of yolk. Yolk utilization during embryogenesis occurs by intracellular lysosomal digestion with selective exocytosis of toposomes.Original Russian Text Copyright © 2005 by Biologiya Morya, Gliznutsa, Dautov.     

Ultrastructural studies on the formation of yolk granules in the statoblast of a fresh-water bryozoan,Pectinatella gelatinosa

The formation of protein-carbohydrate yolk in the statoblast of a fresh-water bryozoan, Pectinatella gelatinosa, was studied by electron microscopy. Two types (I and II) of yolk cells were distinguished. The type I yolk cells are mononucleate and comprise a large majority of the yolk cells. The type II yolk cells are small in number; they become multinucleate by fusion of cells at an early stage of vitellogenesis. In both types of yolk cells, electron-dense granules (dense bodies) are formed in Golgi or condensing vacuoles, which are then called yolk granules. For the formation of yolk granules, the following processes are considered: 1. Yolk protein is synthesized in the rough-surfaced endoplasmic reticulum (RER) of the yolk cells. 2. The synthesized protein condenses in the cisternal space of the RER and is packaged into small oval swellings, which are then released from the RER as small vesicles (Golgi vesicles, 300-600 A in diameter). 3. The small vesicles fuse with one another to form condensing vacuoles, or with pre-existing growing yolk granules. 4. In the matrix of the condensing vacuoles or growing yolk granules, electron-dense fibers are fabricated and then arranged in a paracrystalline pattern to form the dense body. 5. After the dense body reaches its full size, excess membrane is removed and eventually the yolk granules come to mature. Toward the end of vitellogenesis of the yolk cells, the cytoplasmic organelles are ingested by autophagosomes derived from multivesicular bodies and disappear.     

Insufficient VEGFA activity in yolk sac endoderm compromises haematopoietic and endothelial differentiation

Vascular endothelial growth factor A (VEGFA) plays a pivotal role in the first steps of endothelial and haematopoietic development in the yolk sac, as well as in the establishment of the cardiovascular system of the embryo. At the onset of gastrulation, VEGFA is primarily expressed in the yolk sac visceral endoderm and in the yolk sac mesothelium. We report the generation and analysis of a Vegf hypomorphic allele, Vegf(lo). Animals heterozygous for the targeted mutation are viable. Homozygous embryos, however, die at 9.0 dpc because of severe abnormalities in the yolk sac vasculature and deficiencies in the development of the dorsal aortae. We find that providing 'Vegf wild-type' visceral endoderm to the hypomorphic embryos restores normal blood and endothelial differentiation in the yolk sac, but does not rescue the phenotype in the embryo proper. In the opposite situation, however, when Vegf hypomorphic visceral endoderm is provided to a wild-type embryo, the 'Vegf wild-type' yolk sac mesoderm is not sufficient to support proper vessel formation and haematopoietic differentiation in this extra-embryonic membrane. These findings demonstrate that VEGFA expression in the visceral endoderm is absolutely required for the normal expansion and organisation of both the endothelial and haematopoietic lineages in the early sites of vessel and blood formation. However, normal VEGFA expression in the yolk sac mesoderm alone is not sufficient for supporting the proper development of the early vascular and haematopoietic system.     

Characterization of a rat yolk sac carcinoma cell line

A continuous cell line was established from an experimentally induced rat yolk sac carcinoma. In the early passages both visceral and parietal yolk sac carcinoma were present (designated L1). When the cell line was reestablished in culture after serial transplantations in rats, only parietal yolk sac carcinoma could be identified (designated L2). This cell line expresses parietal yolk sac endoderm characteristics in that it synthesizes basement membrane components, in particular, laminin, but also entactin, collagen IV, and heparan sulfate proteoglycan. In addition, a noncartilage chondrotin sulfate proteoglycan is synthesized. This rat yolk sac carcinoma cell line L2 will be a valuable model for the study of basement membrane components.     

The yolk sac in late embryonic development of the stick insect Carausius morosus (Br.)

Differentiation of the yolk sac was examined ultrastructurally and cytochemically in late embryonic development of the stick insect Carausius morosus. During migration along the yolk sac, endodermal cells form a discontinuous cell epithelium, leaving wide intercellular channels between neighbouring cell clusters. Within the same cell cluster, cells are all joined by septate junctions. In the proximity of the proctodeum region, intercellular channels are filled with numerous cell debris which are shown to derive from vitellophages undergoing cell lysis. Yolk sacs resolved by gel electrophoresis are shown to release a number of vitellin polypeptides into the culture medium. These are equivalent in molecular weight to those present in the vitellophage yolk granules This observation is consistent with the evidence that the basement lamina may act as a course physical filter, retaining particles larger than colloidal thorium dioxide and allowing free percolation of peroxidase. Differentiating endodermal cells form a microvillar striated border along the apical plasma membrane. A number of vesicular criptae were frequently seen in these differentiating endodermal cells. Electron dense granules released by endodermal cells are suggested to play a role in vitellophage lysis and vitellin release from the enclosed yolk granules.     

Isolation and characterization of the Fc receptor from the fetal yolk sac of the rat

The yolk sac of the fetal rat and the proximal small intestine of the neonatal rat selectively transport maternal IgG. IgG-Fc receptors are thought to mediate transport across the epithelium of both tissues. We used a mouse mAb (MC-39) against the 45-54-kD component of the Fc receptor of the neonatal intestine to find an antigenically related protein that might function as an Fc receptor in fetal yolk sac. In immunoblots of yolk sac, MC-39 recognized a protein band with apparent molecular mass of 54-58 kD. MC-39 bound to the endoderm of yolk sac in immunofluorescence studies. In immunogold-labeling experiments MC-39 was associated mainly with small vesicles in the apical cytoplasm and in the region near the basolateral membrane of endodermal cells. The MC- 39 cross-reactive protein and beta 2-microglobulin, a component of the intestinal Fc receptor, were copurified from detergent-solubilized yolk sac by an affinity purification that selected for proteins which, like the intestinal receptor, bound to IgG at pH 6.0 and eluted at pH 8.0. In summary, the data suggest that we have isolated the Fc receptor of the yolk sac and that this receptor is structurally and functionally related to the Fc receptor of the neonatal intestine. An unexpected finding is that, unlike the intestinal receptor which binds maternal IgG on the apical cell surface, the yolk sac receptor appears to bind IgG only within apical compartments which we suggest represent the endosomal complex.     

Synthesis of serum proteins by the posthaematopoietic feline yolk sac

The feline yolk sac persists even after the end of its haematopoietic phase with prominent ER-cisternae in the endoderm suggesting biosynthetic capacity. Therefore, yolk sac explants from the 54th day and 57th day were incubated with [3H]-L-leucine in order to study its protein biosynthesis. Newly synthesized proteins were discovered in sliced SDS-polyacrylamide gels by the use of scintillation technique and identified by molecular weight determination and isoelectrofocusing, also using stained electropherograms of unlabeled tissue, serum, and marker proteins. The highest radioactive incorporations were found in 69,000--70,000 dalton proteins and interpreted as serum albumin and alpha-fetoprotein. The autoradiography revealed that the cytoplasm of the endoderm is the site of the most active biosynthesis of proteins which were obviously stored in the ER-cisternae for some time. The yolk sac fluid proteins are almost exclusively serum proteins, although in a very low concentration. We regard a large-scale formation of serum proteins in the yolk sac endoderm as the cause of this organ's very late regression in the cat.     

The pig yolk sac I

Summary Yolk sacs from pig embryos ranging between 18 mm and 55 mm in length were investigated by scanning electron microscopy (SEM), transmission electron microscopy (TEM), and histochemistry. The organ was no longer present in embryos of 70 mm length. The endoderm proliferates in embryos of about 20 mm length with gland-like endodermal cell columns and finally becomes stratified, representing over 90% of the yolk sac mass. The endodermal cells show a high activity of oxidoreductases and lysosomal enzymes; their luminal surface bears few absorptive specializations. The mesothelium is inert, as judged from its surface ultrastructure, organelle composition and enzyme content. TEM reveals the endodermal cells to be polarized even in stratified areas. They resemble liver parenchymal cells with respect to their basal villi, which are exposed to capillaries with discontinuous or fenestrated endothelium. Giant mitochondria with crystalline inclusions in the mature endodermal cytoplasm are outnumbered by large stacks of the rough ER, which can amount to 60% of the cytoplasm. This conspicuous RER is suspected to be the production site of serum proteins which are discharged into the vascular bed. Close to the time of the organ's regression, an unusual storage of material in terminal buds of the ER was found. Intercellular canaliculi and the endocytic apparatus of the endoderm are thought to serve regression.     

Characteristics of ceruloplasmin synthesis in mammalian embryogenesis. 2. The yolk sac as the site of the primary expression of the ceruloplasmin gene in rats

It was shown that the omphaloid placenta and, first of all, visceral wall of yolk sac is the site of primary synthesis of ceruloplasmin (CP), whereas the activation of CP synthesis in the liver cells is secondary and is revealed from the 12th day of embryo-genesis. The CP synthesis in the yolk sac cells proved by selective CP localization in the cells of the yolk sac visceral wall and, first of all, in the cells of visceral endoderm on sections stained by the method of indirect immunofluorescence and using the reaction of soluble peroxidase-antiperoxidase complex. A specific CP-mRNA has been revealed in the yolk sac cells which is actively translated in the polyribosomes isolated from the yolk sac and in the cell-free translation system from the rabbit reticulocytes. on the 14th day of embryogenesis CP amounts to ca. 4% of all polypeptides secreted by the yolk sac cells. As the embryogenesis proceeds, the relative rate of CP synthesis progressively decreases in the yolk sac and increases in the liver cells. CP synthesized by the yolk sac cells has a molecular mass of ca. 122 kD. Possible causes of differences between the "embryonic" and "adult" rat CPs are discussed. A suggestion has been put forward that the time of activation of CP synthesis coincides with the yolk sac formation (8-9th days of embryogenesis) and the cells of visceral endoderm are the site of primary expression of the CP gene.     

Growth and proliferation in mouse parietal yolk sac during whole embryo culture

Use of the culture techniques for postimplantation rodent embryos, modified by explanting Day 9 or Day 10 embryos with the trophoblast cells removed but the remainder of the parietal yolk sac left intact, resulted in significant expansion of Reichert's membrane, accompanied by a marked increase in the numbers of the adherent parietal endoderm cells which synthesize the membrane. The surface area of the parietal endoderm/Reichert's membrane complex roughly doubled during culture, and the combined protein content of the cells and their basement membrane was also raised after incubation. Parietal endoderm cell numbers, calculated from area and cell density measurements on flattened membranes, increased by 54-190%, depending on the age of the embryo.     

An in vitro morphological study of the mouse visceral yolk sac and possible yolk sac immunocyte precursors

Summary Mouse visceral yolk sac has been organ cultured from 9 days of gestation, a time prior to the thymus being lymphoid, until 12 days of gestation, a time after which the thymus is lymphoid. During the culture period the endodermal epithelial cells survived well, erythropoiesis diminished, endothelial-lined cavities formed in the mesodermal mass, and cells developed which have been classified as large, medium and small immunocyte precursors. The cytoplasm of the immunocyte precursors contains polysomes, spherical mitochondria, a few profiles of rough endoplasmic reticulum, occasional granules and a large Golgi complex. This study offers morphological support for the yolk sac origin of immunocyte precursors in the mouse which may seed the thymus and liver.Supported by NIH Grant AI 13486-01     

Apolipoprotein B-related gene expression and ultrastructural characteristics of lipoprotein secretion in mouse yolk sac during embryonic development.

In mice, the yolk sac appears to play a crucial role in nourishing the developing embryo, especially during embryonic days (E) 7;-10. Lipoprotein synthesis and secretion may be essential for this function: embryos lacking apolipoprotein (apo) B or microsomal triglyceride transfer protein (MTP), both of which participate in the assembly of triglyceride-rich lipoproteins, are apparently defective in their ability to export lipoproteins from yolk sac endoderm cells and die during mid-gestation. We therefore analyzed the embryonic expression of apoB, MTP, and alpha-tocopherol transfer protein (alpha-TTP), which have been associated with the assembly and secretion of apoB-containing lipoproteins in the adult liver, at different developmental time points. MTP expression or activity was found in the yolk sac and fetal liver, and low levels of activity were detected in E18.5 placentas. alpha-TTP mRNA and protein were detectable in the fetal liver, but not in the yolk sac or placenta. Ultrastructural analysis of yolk sac visceral endoderm cells demonstrated nascent VLDL within the luminal spaces of the rough endoplasmic reticulum and Golgi apparatus at E7.5 and E8.5. The particles were reduced in diameter at E13.5 and reduced in number at E18.5;-19.The data support the hypothesis that the yolk sac plays a vital role in providing lipids and lipid-soluble nutrients to embryos during the early phases (E7;-10) of mouse development. secretion in mouse yolk sac during embryonic development.     

Simultaneous localisation of human γ-globulin I125 and ferritin during transport across the rabbit yolk sac splanchnopleur

Summary The transport of human γ-globulin labelled with I¹²⁵, and ferritin, across the rabbit yolk sac splanchnopleur, has been followed using electronmicroscopy combined with autoradiography. Both ferritin, and human γ-globulin I¹²⁵ as indicated by grains, became similarly localised in the same absorptive vesicles present in the yolk sac endoderm. Only human γ-globulin I¹²⁵ could be convincingly demonstrated in the basement membrane of the yolk sac endoderm, in the vascular mesenchyme, and in the vitelline vessels. These findings are discussed in the light of other studies using fluorescent protein tracing to locate simultaneously transmitted and non-transmitted proteins, and in the light of suggested mechanisms, to explain selective transmission. Supported by an award from the Science Research Council, to whom grateful acknowledgement is made.     

Structure of the endodermal epithelium of the chick yolk sac during early stages of development.

The structure of the areas pellucida and vasculosa of the early chick embryo (stages 11-29) was examined by light, transmission and scanning electron microscopy. The most striking feature of the endodermal cells of these areas is the presence of large intracellular yolk drops which are characteristic of the regions in which they are found; lipid-like homogeneous drops in the area pellucida, heterogeneously composed pleomorphic drops in the mid-region of the area vasculosa and granular drops at the periphery of the area vasculosa in the region of the sinus terminalis. On morphological criteria it is postulated that granular drops may arise by direct engulfment of extracellular yolk, but this does not appear to be true for pleomorphic or homogeneous drops. Since the apical junctions between endodermal cells across the yolk sac are tight, they seal off the extraembryonic compartment from the vitelline circulation and presumably prevent intercellular passage of the yolk constituents. Thus the endodermal epithelium must mediate the transport of nutrients from the yolk mass to the developing embryo. Endodermal cells exhibit a variation across the yolk sac in the presence and number of structures associated with uptake of extracellular materials. The mid-region of the area vasculosa appears to be the most endocytotically active region with an abundance of microvilli, bristle-coated pits and vesicles and apical canaliculi and vacuoles. There is a close association between the endoderm and vitelline blood vessels and this association is maintained, as the yolk sac develops, by the formation of small vessels juxtaposed between the vascular surface of the endoderm and the walls of the large vitelline vessels.     

An electron-microscopic study of syncytium formation during early embryonic development of the freshwater planarian Bdellocephala brunnea

To substantiate the assumption that the egg cell and blastomeres in planarian embryos influence surrounding yolk cells to form a syncytium, embryos at 1- to 8-cell stages were examined by electron microscopy. Within special areas of the endoplasmic reticulum both in the egg cell and in the blastomeres, a large number of vacuoles of various sizes formed and then disappeared at least four times over the period from egg-laying through the 8-cell stage as if their contents were being secreted. These activities diminished markedly at the 8-cell stage. Yolk cells surrounding the egg cell and blastomeres were aggregated in close contact with one another in a small clump shortly after egg-laying, and then, late in the 4-cell stage, became fused, forming a syncytium. The correlation between release of vacuoles by the egg cell and blastomeres and the formation of a syncytium by the yolk cells indicate that the cell fusion could be induced by a factor contained in the vacuoles.     

Vitamin D and chick embryonic yolk calcium mobilization: identification and regulation of expression of vitamin D-dependent Ca2(+)-binding protein, calbindin-D28K, in the yolk sac

The developing chick embryo acquires calcium from two sources. Until about Day 10 of incubation, the yolk is the only source; thereafter, calcium is also mobilized from the eggshell. We have previously shown that during normal chick embryonic development, vitamin D is involved in regulating yolk calcium mobilization, whereas vitamin K is required for eggshell calcium translocation by the chorioallantoic membrane. We have studied here the biochemical action of 1,25-dihydroxy vitamin D3 in the yolk sac by examining the expression and regulation of the cytosolic vitamin D-dependent calcium-binding protein, calbindin-D28K. Two types of embryos are used for this study, normal embryos developing in ovo and embryos maintained in long-term shell-less culture ex ovo, the latter being dependent solely on the yolk as their calcium source. Our findings are (1) calbindin-D28K is expressed in the embryonic yolk sac, detectable at incubation Days 9 and 14; (2) the embryonic yolk sac calbindin-D28K resembles that of the adult duodenum in both molecular weight (Mr 28,000) and isoelectric point, as well as the presence of E-F hand Ca2(+)-binding structural domains; (3) systemic calcium deficiency caused by shell-less culture of chick embryos results in enhanced expression of calbindin-D28K in the yolk sac during late development; (4) yolk sac calbindin-D28K expression is inducible by 1,25-dihydroxy vitamin D3 treatment in vivo and in vitro; and (5) immunohistochemistry revealed that yolk sac calbindin-D28K is localized exclusively to the cytoplasm of the yolk sac endoderm. These findings indicate that the chick embryonic yolk sac is a genuine target tissue of 1,25-dihydroxy vitamin D3.