Allantoic Epithelium Research Articles

Temporal and spatial expression of aquaporins 1, 5, 8, and 9: Potential transport of water across the endometrium and chorioallantois of pigs

IntroductionThe uterus and placenta transport water during pregnancy recognition signaling, conceptus implantation, and placental development/placentation. This is likely influenced by aquaporins (AQPs) in the reproductive tract. This study determined mRNA and cell-type specific expression of AQP 1, 5, 8, and 9 proteins in the porcine uterus and placenta. MethodsPorcine uteri and Chorioallantois were subjected to real-time PCR and immunofluorescence microscopy. ResultsAQP1 mRNA was maximal by Day 25 in endometrium and remained stable thereafter. AQP1 mRNA did not change in chorioallantois. AQP1 protein localized to erythrocytes and endothelium of the endometrium and allantois, and to smooth muscle of the myometrium. AQP5 protein localized to apical and lateral surfaces of the chorionic epithelia of areolae, but mRNA did not change in chorioallantois. AQP8 mRNA was high in the endometrium from Days 15 through 60 of gestation, and protein localized to multiple cell types within the endometrium and chorioallantois. AQP9 mRNA was highest in the endometrium on Days 10, 12 and 25, but did not change in the chorioallantois. AQP9 protein localized to the apical surface of endometrial luminal epithelial cells during early pregnancy, with a shift towards the basal surface later. AQP9 protein was observed in the allantoic epithelium. DiscussionResults reveal pigs can potentially use AQP1, AQP5, AQP8, and AQP9 to transport water from the endometrial bloodstream to the allantoic bloodstream or allantoic fluid. The reverse is also possible and may explain the mechanism for changing volumes of allantoic fluid and hydration of allantoic connective tissues during pregnancy.

Kinetics of the chromosome 14 microRNA cluster ortholog and its potential role during placental development in the pregnant mare

BackgroundThe human chromosome 14 microRNA cluster (C14MC) is a conserved microRNA (miRNA) cluster across eutherian mammals, reported to play an important role in placental development. However, the expression kinetics and function of this cluster in the mammalian placenta are poorly understood. Here, we evaluated the expression kinetics of the equine C24MC, ortholog to the human C14MC, in the chorioallantoic membrane during the course of gestation.ResultsWe demonstrated that C24MC-associated miRNAs presented a higher expression level during early stages of pregnancy, followed by a decline later in gestation. Evaluation of one member of C24MC (miR-409-3p) by in situ hybridization demonstrated that its cellular localization predominantly involved the chorion and allantoic epithelium and vascular endothelium. Additionally, expression of predicted target transcripts for C24MC-associated miRNAs was evaluated by RNA sequencing. Expression analysis of a subset of predicted mRNA targets showed a negative correlation with C24MC-associated miRNAs expression levels during gestation, suggesting the reciprocal control of these target transcripts by this miRNA cluster. Predicted functional analysis of these target mRNAs indicated enrichment of biological pathways related to embryonic development, endothelial cell migration and angiogenesis. Expression patterns of selected target mRNAs involved in angiogenesis were confirmed by RT-qPCR.ConclusionThis is the first report evaluating C24MC kinetics during pregnancy. The findings presented herein suggest that the C24MC may modulate angiogenic transcriptional profiles during placental development in the horse.

Expression of Toll-like receptors 2, 4 and 6 in the equine chorioallantois

In mares, placental diseases are a common cause of pregnancy failure and they can have an economic impact on the horse breeding industry. To our knowledge no published data on TLR expression in the equine placenta exist. This study examined the expression of TLR 2, 4 and 6 as transcript and protein in the placenta (chorioallantois) of 14 foals born alive. By PCR, all examined placental samples contained TLR 2, 4 and 6 transcripts. Using immunohistochemistry, trophoblasts and allantoic epithelium were immunopositive for TLR 2, 4 and 6 in all placental samples. The majority of placental samples contained TLR 4 and 6 positive stromal cells and vascular smooth muscle cells. Since these results confirm the expression of TLR 2, 4 and 6 in different cell populations of the equine placenta, they are the basis for studies into the pathogenesis of TLR-associated placental diseases in mares.

Fetal Membrane Ultrastructure and Development in the Oviparous Milksnake Lampropeltis triangulum (Colubridae) with Reference to Function and Evolution in Snakes.

In eggs of oviparous reptiles, fetal membranes maintain developing embryos through the exchange of respiratory gases and provision of water and calcium. As part of a survey of reptilian fetal membranes, we used scanning electron microscopy to study fetal membrane morphology in the oviparous Pueblan milksnake, Lampropeltis triangulum campbelli. The chorioallantois initially is an avascular structure lined by enlarged chorionic and allantoic epithelia. Upon vascularization, the chorionic epithelium becomes greatly attenuated, enhancing the potential for gas exchange; the allantoic epithelium also flattens. The bilaminar omphalopleure of the yolk sac lacks blood vessels, but it becomes vascularized by allantoic capillaries and transformed into an omphalallantois. Upon regression of the isolated yolk mass, this membrane is converted to chorioallantois, equipping it for gas exchange. Allantoic fluid serves as a water reservoir, and we postulate that it facilitates water uptake by establishing an osmotic gradient. Early in development, epithelia of both the chorion and the omphalopleure show apical microvilli that greatly increase the cell surface area available for water uptake. However, these features are incompatible with gas exchange and are lost as oxygen needs take precedence. A comparison of the fetal membranes to those of other squamate species (both oviparous and viviparous) reveals characteristics that are probably ancestral for snakes, some of which are plesiomorphic for Squamata. The widespread phylogenetic distribution of these features reflects their utility as adaptations that serve functional requirements of squamate embryos.

Immunolocalization of Leptin Receptor and mRNA Expression of Leptin and Estrogen Receptors as well as Caspases in the Chorioallantoic Membrane (CAM) of the Chicken Embryo.

The chicken chorioallantoic membrane (CAM) is used as a model in tests of angiogenesis, the biocompatibility of materials as well as tumor invasive potential. To assess the properties of CAM tissue, the localization of leptin receptor in the CAM, and the mRNA expression of two leptin receptor isoforms, estrogen receptors (ERα and ERβ) and caspases (-1 and -3) in the CAM on embryonic days 12 (E12), 15 (E15) and 18 (E18) were investigated. The leptin receptor was immunolocalized in each structure of the CAM (chorionic epithelium, allantoic epithelium, mesodermal layer and the walls of blood vessels) and did not change among analyzed stages of embryonic development (E12, E15 and E18) and between sexes. Expression of mRNA of genes encoding leptin and estrogen receptors as well as caspases was detected in the CAM of female and male chicken embryos at all three analysed stages of development. The relative mRNA expression of the long form of leptin receptor exceeded that of its short isoform. The mRNA expression of ERβ was significantly higher than ERα as well as caspase-3 in comparison with caspase-1. There were no differences in mRNA expression of these genes between sexes and among analyzed developmental days. The results indicate that the CAM is a target tissue for leptin as well as for estrogens and that CAM development is partially regulated by caspase-1 and caspase-3 dependent cell death. These results should be taken into consideration in studies in which the CAM is used as an experimental model.

Ultrastructure of the fetal membranes of the oviparous kingsnake, Lampropeltis getula (Colubridae) as revealed by scanning electron microscopy

ABSTRACTIn reptilian sauropsids, fetal (extraembryonic) membranes that line the eggshell sustain developing embryos by providing for gas exchange and uptake of water and eggshell calcium. However, a scarcity of morphological studies hinders an understanding of functional specializations and their evolution. In kingsnakes (Lampropeltis getula), scanning electron microscopy reveals two major fetal membranes: the chorioallantois and yolk sac omphalopleure. In early development, the chorioallantois contains tall chorionic epithelial cells, avascular connective tissue, and enlarged allantoic epithelial cells. During its maturation, the chorionic and allantoic epithelia thin dramatically and become underlain by a rich network of allantoic capillaries, yielding a membrane ideally suited for respiratory gas exchange. Yolk sac development initially is like that of typical lizards and snakes, forming an avascular omphalopleure, isolated yolk mass (IYM), and yolk cleft. However, unlike the situation in most squamates studied, the omphalopleure becomes transformed into a “secondary chorioallantois” via three asynchronous events: flattening of the epithelium, regression of the IYM, and vascularization by the allantois. Progressive expansion of chorioallantois parallels growing embryonic needs for gas exchange. In early through mid‐development, external surfaces of both the chorionic and omphalopleure epithelium show an abundance of irregular surface protrusions that possibly increase surface area for water absorption. We postulate that the hypertrophied allantoic epithelial cells produce allantoic fluid, a viscous substance that facilitates water uptake and storage. Our findings are consistent with a previous study on the corn snake Pantherophis guttatus, but include new observations and novel functional hypotheses relevant to a reconstruction of basal squamate patterns. J. Morphol. 276:1467–1481, 2015. © 2015 Wiley Periodicals, Inc.

Ultrastructure of the fetal membranes of the oviparous kingsnake, Lampropeltis getula (Colubridae) as revealed by scanning electron microscopy

In reptilian sauropsids, fetal (extraembryonic) membranes that line the eggshell sustain developing embryos by providing for gas exchange and uptake of water and eggshell calcium. However, a scarcity of morphological studies hinders an understanding of functional specializations and their evolution. In kingsnakes (Lampropeltis getula), scanning electron microscopy reveals two major fetal membranes: the chorioallantois and yolk sac omphalopleure. In early development, the chorioallantois contains tall chorionic epithelial cells, avascular connective tissue, and enlarged allantoic epithelial cells. During its maturation, the chorionic and allantoic epithelia thin dramatically and become underlain by a rich network of allantoic capillaries, yielding a membrane ideally suited for respiratory gas exchange. Yolk sac development initially is like that of typical lizards and snakes, forming an avascular omphalopleure, isolated yolk mass (IYM), and yolk cleft. However, unlike the situation in most squamates studied, the omphalopleure becomes transformed into a "secondary chorioallantois" via three asynchronous events: flattening of the epithelium, regression of the IYM, and vascularization by the allantois. Progressive expansion of chorioallantois parallels growing embryonic needs for gas exchange. In early through mid-development, external surfaces of both the chorionic and omphalopleure epithelium show an abundance of irregular surface protrusions that possibly increase surface area for water absorption. We postulate that the hypertrophied allantoic epithelial cells produce allantoic fluid, a viscous substance that facilitates water uptake and storage. Our findings are consistent with a previous study on the corn snake Pantherophis guttatus, but include new observations and novel functional hypotheses relevant to a reconstruction of basal squamate patterns.

Developmental changes in the ENaC channel of the chick chorioallantoic membrane

Confined to the closed system of the egg, the chick embryo deposits its renal wastes into the chorioallantoic sac, an outgrowth of its hindgut. The allantoic fluid undergoes changes in both volume and ion composition throughout the development of the embryo. These changes are most likely the result of Na+ reabsorption from the allantoic fluid. In previous experiments we demonstrated that short‐circuit current across the epithelia increases five‐fold between embryonic days 9–19. This current is inhibitable by ouabain and amiloride, indicating that Na+‐K+‐ATPase and ENaC channels play a role. Aldosterone increased the amiloride‐inhibitable current from day 9 to day 18, suggesting that the ENaC channels may be up‐regulated by the hormone. The slow (100 min) aldosterone response appears to be the classic gene‐regulated response to a steroid hormone. In this study we are investigating the role of the epithelial sodium channel ENaC in allantoic short‐circuit current. We used RT‐PCR to measure normal developmental changes in the RNA message of the three ENaC subunits in the allantoic epithelium. The RNA message of all subunits increased two‐fold from day 9 to day 11, and then slowly decreased from days 11 to 17. These data were confirmed using Western blots. Subsequent experiments will investigate the role of aldosterone in inducing ENaC protein synthesis.

The Chick Chorioallantoic Membrane: A Model of Molecular, Structural, and Functional Adaptation to Transepithelial Ion Transport and Barrier Function during Embryonic Development

The chick chorioallantoic membrane is a very simple extraembryonic membrane which serves multiple functions during embryo development; it is the site of exchange of respiratory gases, calcium transport from the eggshell, acid-base homeostasis in the embryo, and ion and H2O reabsorption from the allantoic fluid. All these functions are accomplished by its epithelia, the chorionic and the allantoic epithelium, by differentiation of a wide range of structural and molecular peculiarities which make them highly specialized, ion transporting epithelia. Studying the different aspects of such a developmental strategy emphasizes the functional potential of the epithelium and offers an excellent model system to gain insights into questions partly still unresolved.

Initial Development of Bovine Placentation (Bos indicus) from the Point of View of the Allantois and Amnion

The aim of this study was to perform a morphological characterization of the initial bovine placental development, between 20 and 70 days post-insemination (p.i.), with emphasis on the differentiation of the allantois and amnion. After collection, the conceptuses were dissected, macroscopically measured and photographically documented. The extraembryonic membranes were cut into fragments measuring 5 cm(2), and then fixed in 4% paraformaldehyde for analysis using light microscopy, and in 2.5% glutaraldehyde for use in scanning and transmission electron microscopy. The extraembryonic and fetal membranes presented variable degrees of development throughout the periods analysed. The macroscopic appearance of vascularization of the allantois, its attempt to merge with the chorium and the effective appearance of the first cotyledons in development were the events observed from 30 to 40 days of pregnancy. The measurements of the amnion increased gradually as gestation developed. The allantoic epithelia presented cellular dimorphism from 20 to 25 days of pregnancy, but was shown to be immature from 60 to 70 days of pregnancy.

Annexins as cell-type-specific markers in the developing chicken chorionallantoic membrane

Between day E8 and E12 of embryonic development, the chicken chorioallantoic membrane (CAM) undergoes massive structural rearrangement enabling calcium-uptake from the eggshell to supply the growing embryo. However, the contribution of the various cell types of the chorionic epithelium including the capillary covering (CC) cells, villus cavity (VC) cells, endothelial-like cells, and basal cells to this developmental program is largely unknown. In order to obtain markers for the different cell types in the chorionic epithelium, we determined the expression patterns of various calcium-binding annexins in the developing chicken CAM. By reverse transcription/polymerase chain reaction with primers deduced from nucleotide sequences available in various databases, the presence of annexin (anx)-1, anx-2, anx-5, and anx-6 was demonstrated at days E8 and E12. Quantitative immunoblotting with novel antibodies raised against the recombinant proteins revealed that anx-1 and anx-5 were significantly up-regulated at day E12, whereas anx-2 and anx-6 expression remained almost unchanged in comparison to levels at day E8. Immunohistochemistry of paraffin-embedded sections of E12 CAM revealed anx-1 in CC cells and VC cells. Anx-2 was localized in capillaries in the chorionic epithelium and in basal cells of the allantoic epithelium, whereas anx-6 was detected in basal cells or endothelial-like cells of the chorionic epithelium and in the media of larger vessels in the mesenchyme. A 2-day exposure of the CAM to a tumor cell spheroid resulted in strong proliferation of anx-1-expressing CC cells suggesting that these cells participate in the embryonic response to experimental intervention. Thus, annexins exhibit complementary expression patterns and represent appropriate cell markers for the further characterization of CAM development and the interpretation of results obtained when using CAM as an experimental model.

Fissura placentária de gatas SRD, Felis catus - Linnaeus, 1758: aspectos macro e microscópicos

This research consists of study of shematic macroscopy cat's placenta and placental characterization as its type, zonary placenta, that 62,5% of cases presents a fissure at distal area of umbilical cord. This is formed by a joustfetal area, joustplacental area and middle area, histological findings: two arteries, a vein, two vitelline pedicule and two allantois pedicule. In the fissure, an allantoic epithelium covering this area at 10% of the cases and 90% found a trophoblast decrease compared with others placental areas out of the fissure. So, the feline placenta, in its maternal/fetal relationship shows a imcomplete zonary placenta, different of that occurs in other carnivores.

Cell type-specific and developmentally regulated expression of the AE1 anion exchanger in the chicken chorioallantoic membrane

Antibodies specific for the chicken AE1 anion exchanger have been used to determine the cell-type specific pattern of expression of this electroneutral transporter in the chick chorioallantoic membrane (CAM) during embryonic development. Immunolocalisation analyses demonstrated that the AE1 anion exchanger accumulated in the basolateral membrane of a subset of cells in both the chorionic and allantoic epithelial layers. Double immunostaining indicated that the AE1-positive cells in the chorionic and allantoic epithelia were also positive for the carbonic anhydrase isoform, CAII, which serves as a marker for the villus cavity (VC) cells of the chorionic epithelium and the mitochondria-rich cells of the allantoic epithelium. Immunoelectron microscopy revealed that AE1 accumulated in extensive projections that extended from the lateral membrane of VC cells towards the adjacent capillary covering cells. These results represent the first demonstration of anion exchanger expression in the chick CAM, and they suggest a role for basolateral AE1 in bicarbonate reabsorption that is required in the embryo for maintaining acid-base balance during development.

Developmental expression of glycocomponents in the chick chorioallantoic membrane.

Widespread interest has focused on the research of the chorioallantoic membrane (CAM) and its functional contribution to gaseous exchange, calcium reabsorption, water and electrolyte transport during chick embryogenesis. Nevertheless, very little information is available on the glycoconjugate components of this extra-embryonic structure. In the present study, we investigated by lectin histochemistry, the glycosylation pattern expressed in the CAM epithelia during embryonic development. Occurrence of sialic acid-associated glycoproteins was detailed by either specific lectins, which discriminate alpha2,3 and alpha2,6 sialoderivatives, or sialidase digestion combined with appropriate lectins to identify the sialic acid acceptor sugars. Lectin affinities proved to depend greatly on differentiation of the CAM epithelia which showed highest expression of binding sites during the second half of incubation up to hatching. Differences emerged between the chorionic and the allantoic epithelium, regarding qualitative, quantitative and temporal expression of sugar moieties. A cell type-specific distribution of glycocomponents was found in the chorionic epithelium where lectin binding sites were specifically located in the villus cavity cells. In the allantoic epithelium, high and heterogeneous occurrence of sialoglycoconjugates as well as specific presence of fucose residues were evidenced mostly in the granule cells. We conclude from these findings that various glycoconjugates in the CAM could participate in different physiological functions characteristic of the chorionic and the allantoic epithelium.

Endogenous origin of the lymphatics in the avian chorioallantoic membrane.

The lymphatics of the intestinal organs have important functions in transporting chyle toward the jugulosubclavian junction, but the lymphangiogenic potential of the splanchnic mesoderm has not yet been tested. Therefore, we studied the allantoic bud of chick and quail embryos. It is made up of endoderm and splanchnic mesoderm and fuses with the chorion to form the chorioallantoic membrane (CAM) containing both blood vessels and lymphatics. In day 3 embryos (stage 18 of Hamburger and Hamilton [HH]), the allantoic mesoderm consists of mesenchymal cells that form blood islands during stage 19 (HH). The endothelial network of the allantoic bud, some intraluminal and some mesenchymal cells express the hemangiopoietic marker QH1. The QH1-positive endothelial cells also express the vascular endothelial growth factor receptor-3 (VEGFR-3), whereas the integrating angioblasts and the round hematopoietic cells are QH1-positive/VEGFR-3-negative. The ligand, VEGF-C, is expressed ubiquitously in the allantoic bud, and later predominantly in the allantoic epithelium and the wall of larger blood vessels. Allantoic buds of stage 17-18 (HH) quail embryos were grafted homotopically into chick embryos and reincubated until day 13. In the chimeric CAMs, quail endothelial cells are present in blood vessels and lymphatics, the latter being QH1 and VEGFR-3 double-positive. QH1-positive hematopoietic cells are found at many extra- and intraembryonic sites, whereas endothelial cells are confined to the grafting site. Our results show that the early allantoic bud contains hemangioblasts and lymphangioblasts. The latter can be identified with Prox1 antibodies and mRNA probes in the allantoic mesoderm of day 4 embryos (stage 21 HH). Prox1 is a specific marker of the lymphatic endothelium throughout CAM development.

Histology and ultrastructure of the chorioallantoic membrane of the mallard duck (Anas platyrhynchos).

The histology and fine structure of the chorioallantoic membrane of the mallard duck (Anas platyrhynchos), and the density of vessels per millimeter of membrane were assessed between days 12 and 24 of incubation. Light and transmission electron microscopy of the chorioallantoic membrane of the mallard duck after various days of incubation was carried out. Blood vessels within the mesoderm were counted per millimeter of membrane by light microscopy (40x). The chorioallantoic membrane had three distinct layers from day 12 to 24 of incubation, the chorionic epithelium, the mesoderm, and the allantoic epithelium. After day 12, chorionic epithelium consisted of two layers of flattened, elongated epithelial cells interfaced by numerous desmosomes, and separated from the underlying mesoderm by a basement membrane. At this stage, the allantoic epithelium consisted of a single layer of flattened, overlapping cells. Blood capillaries were observed in the mesoderm close to the chorionic epithelium on days 12 and 13; by day 14, these capillaries were located within the chorionic epithelium, forming a capillary sinus. Between days 14 and 16, the chorion underwent cellular and cytological differentiation into three cell types: capillary covering cells, villus cavity cells, and less differentiated basal cells. The mesoderm was composed of a loose matrix of mesenchymal cells and collagen fibrils through which coursed blood and lymphatic vessels. The vascular density in the mesoderm increased rapidly from 4.2+/-0.6 vessels per mm (n = 12) on day 12 to a maximum of 9.4+/-0.3 vessels per mm (n = 15) by day 16. From day 16, the allantoic epithelium had two to three layers of elongated and overlapping cells. The luminal layer of allantoic epithelial cells had microvillus projections and varying numbers of membrane-bound dense vesicles at all stages from day 12 onward. The histologic and ultrastructural features of mallard duck chorioallantoic membrane from day 12 to 24 of incubation were very similar to those described in the chorioallantoic membrane of the chicken (Gallus gallus) from day 8 to 20 of incubation. Much of the information available concerning the CAM of the chicken also may apply to the CAM of the mallard, with timing adjusted to match the developmental time-frame recorded here.

Ovine amniotic and allantoic epithelia across gestation

Ovine amniotic and allantoic epithelia across gestation

Ovine amniotic and allantoic epithelia across gestation.

Extensive studies on the regulation of the volume and composition of amniotic and allantoic fluid in the sheep have suggested that the amniotic and allantoic membranes must play an active role in these processes. Little is known of the functional morphology of the sheep amnion and allantois beyond the presence of an epithelium overlying connective tissue. The ovine amnion and allantois were characterized at a range of gestational ages (27-140 days of gestation, where term is 145-150 days) by electron microscopy (SEM and TEM) and the presence of transporting ATPases examined by use of immunohistochemistry (Ca++-ATPase) and in situ hybridization (Na,K-ATPase). With increasing gestational age, the cell height of epithelium of the membranes increased, as did the number of apical microvilli and the length of zonulae occludentes. Epithelial cell cytoplasm increased in complexity, and cell shape changed from flattened to cuboidal. Proliferation of cells occurred until close to term. Immunoreactivity to Ca++-ATPase was present in the basolateral membranes at all stages of gestation examined, but hybridization with the alpha and beta subunits of Na,K-ATPase was present only at or after 100 days of gestation. The epithelia of the sheep amnion and allantois display characteristics typical of transporting epithelia. As the epithelia mature, changes related to increased capacity for solute and fluid transport regulation occur.

Studies into aromatase activity associated with fetal allantochorionic and maternal endometrial tissues of equine placenta. Identification of metabolites by gas chromatography mass spectrometry.

Maternal endometrial and fetal allantochorionic tissues were separated manually from the placentae of seven healthy thoroughbred and three pony mares, ranging in gestational age from 100 to 318 days. The homogeneity of subcellular fractions prepared from these tissues was assessed initially using the marker enzymes, succinate dehydrogenase, NADPH cytochrome C reductase and lactate dehydrogenase for the mitochondrial, microsomal and cytosolic fractions, respectively. Light microscopy and histochemical analysis demonstrated that the separated fetal allantochorionic membrane, which is made up of allantoic and chorionic epithelia, contained no significant contamination of maternal tissues. The maternal endometrium, however, was found to contain appreciable amounts of fetal chorion torn off during the separation process. Tissue homogenates and subcellular fractions were incubated with testosterone together with [4-(14)C] and [(2)H5 or (2)H3] labelled analogues in either an NADPH (1 mM) or a NADPH-regenerating environment; control experiments (without additional cofactor) were also performed. After extraction of the tissue homogenates, neutral and phenolic (oestrogen) unconjugated steroids were separated by column chromatography. Radiolabelled studies revealed that in allantochorionic tissue incubations 67-77% of testosterone was converted to oestrogenic material, subcellular fractionation indicating that oestrogen production was largely confined to the microsomal fraction and time-course studies showing that the rate of formation appeared to be linear up to 90 min. In contrast, only 5-25% conversion occurred using maternal endometrial tissues, which could be accounted for by the contaminating presence of fetal chorion. No oestrogen production was detected in control incubations. These radiolabelled studies demonstrate that aromatase activity is located on the fetal allantochorionic surface and, together with the histochemical data, further delineate this activity to the chorion in mature equine placenta. Gas chromatographic-mass spectrometric (GC-MS) analysis of the phenolic extracts from allantochorionic tissue homogenate incubations indicated the presence of substrate-derived oestradiol-17beta (E2), 6-oxo-oestradiol-17beta (6-oxo-E2) and 6beta-hydroxyoestradiol-17beta (6beta-OH-E2). Whereas all three oestrogens were identified as metabolites from testosterone in incubations performed using allantochorionic tissue homogenates and post-mitochondrial suspensions (PMS), only E2 was identified from incubations performed using microsomal fractions prepared from this tissue. We conclude that both the microsomal and cytosol fractions are required for the conversion of E2 to the 6-oxygenated species in vitro. Using stable isotope-labelled substrates and GC-MS analysis the mechanism of formation of these metabolites from these in vitro incubation studies may be inferred. GC-MS analysis of the neutral extracts from allantochorionic tissue homogenate incubations confirmed the presence of small quantities of substrate-derived 5(10)-oestrenediols. No substrate-derived 5(10)-oestrene-3,17-diols were detected in extracts from microsomal preparations incubated in the absence of cytosol. These data suggest that demethylation of C19 steroids to produce C18 neutral steroids may require the synergistic action of enzymic activities that appear to reside both in the microsomal and cytosolic fractions of equine allantochorionic tissues.

Cytokeratin Expression in the Stomach Epithelia of the Chicken Embryo is Regulated by Epithelial-Mesenchymal Interactions

Abstract Regulatory mechanisms of cytokeratin expression in the stomach (the proventriculus and the gizzard) epithelia of chicken embryos were examined by epithelial-mesenchymal recombination experiments. In the proventriculus, antigens detected by monoclonal antibodies (PKK1 and AE3) against cytokeratins invariably decreased in gland cells which synthesize pepsinogen, while luminal and duct epithelial cells were positive to these antibodies. When recombined and cultured with the proventricular mesenchyme, the esophageal, proventricular and gizzard epithelia formed proventriculus-type, pepsinogen-expressing glands and the expression patterns of PKK1 and AE3 antigens were identical to that in the normal proventriculus. Under the same experimental conditions, the small-intestinal and allantoic epithelia also formed complex glands but did not express pepsinogen, and PKK1 and AE3 antigens were actively expressed throughout the epithelia. These results indicate that the proventricular mesenchyme can regulate c...