Fuijkschot, J.; Wijnen, R.M.H.; Gerrits, G.P.; Dubois, S.V.; Rieu, P.N.M.A.
Neonatal umbilical anomalies usually represent remains of the vitelline duct or the allantois. We describe a case of an umbilical appendix in a neonate. The vermiform appendix was found to be positioned in the umbilical cord. In a brief literature review we found eight other reports concerning
Lawson, Kirstie A.; Dunn, N. Ray; Roelen, Bernard A.J.; Zeinstra, Laura M.; Davis, Angela M.; Wright, Christopher V.E.; Korving, Jeroen P.W.F.M.; Hogan, Brigid L.M.
In many organisms the allocation of primordial germ cells (PGCs) is determined by the inheritance of maternal factors deposited in the egg. However, in mammals, inductive cell interactions are required around gastrulation to establish the germ line. Here, we show that Bmp4 homozygous null embryos contain no PGCs. They also lack an allantois, an extraembryonic mesodermal tissue derived, like the PGCs, from precursors in the proximal epiblast. Heterozygotes have fewer PGCs than normal, due to a...
Uchida, Keiko; Nakazawa, Maki; Yamagishi, Chihiro; Mikoshiba, Katsuhiko; Yamagishi, Hiroyuki
The embryonic-maternal interface of the placental labyrinth, allantois, and yolk sac are vital during embryogenesis; however, the precise mechanism underlying the vascularization of these structures remains unknown. Herein we focus on the role of inositol 1,4,5-trisphosphate (IP3) receptors (IP3R), which are intracellular Ca(2+) release channels, in placentation. Double knockout (DKO) of type 1 and 3 IP3Rs (IP3R1 and IP3R3, respectively) in mice resulted in embryonic lethality around embryonic day (E) 11.5. Because IP3R1 and IP3R3 were co-expressed in endothelial cells in the labyrinth, allantois, and yolk sac, we investigated extra-embryonic vascular development in IP3R1- and IP3R3-DKO mice. The formation of chorionic plates and yolk sac vessels seemed dysregulated around the timing of the chorio-allantoic attachment, immediately followed by the disorganization of allantoic vessels, the decreased expression of the spongiotrophoblast cell marker Tpbpa and the growth retardation of the embryos in DKO mice. Fluorescent immunohistochemistry demonstrated downregulation of a vascular endothelial marker, CD31, in labyrinth embryonic vessels and poor elongation of extra-embryonic mesoderm into the labyrinth layer in DKO placenta, whereas the branching of the DKO chorionic trophoblast was initiated. In addition, allantoic and yolk sac vessels in extra-embryonic tissues were less remodeled in DKO mice. In vitro endothelial cord formation and migration activities of cultured vascular endothelial cells derived from human umbilical vein were downregulated under the inhibition of IP3R. Our results suggest that IP3R1 and IP3R3 are required for extra-embryonic vascularization in the placenta, allantois, and yolk sac. This is the first demonstration of the essential role of IP3/IP3Rs signaling in the development of the vasculature at the embryonic-maternal interface. Copyright © 2016 Elsevier Inc. All rights reserved.
Wolfe, Adam D; Rodriguez, Adriana M; Downs, Karen M
The allantois-derived umbilical component of the chorio-allantoic placenta shuttles fetal blood to and from the chorion, thereby ensuring fetal-maternal exchange. The progenitor populations that establish and supply the fetal-umbilical interface lie, in part, within the base of the allantois, where the germ line is claimed to segregate from the soma. Results of recent studies in the mouse have reported that STELLA (DPPA-3, PGC7) co-localizes with PRDM1 (BLIMP1), the bimolecular signature of putative primordial germ cells (PGCs) throughout the fetal-placental interface. Thus, if PGCs form extragonadally within the posterior region of the mammal, they cannot be distinguished from the soma on the basis of these proteins. We used immunohistochemistry, immunofluorescence, and confocal microscopy of the mouse gastrula to co-localize STELLA with a variety of gene products, including pluripotency factor OCT-3/4, mesendoderm-associated T and MIXl1, mesendoderm- and endoderm-associated FOXa2 and hematopoietic factor Runx1. While a subpopulation of cells localizing OCT-3/4 was always found independently of STELLA, STELLA always co-localized with OCT-3/4. Despite previous reports that T is involved in specification of the germ line, co-localization of STELLA and T was detected only in a small subset of cells in the base of the allantois. Slightly later in the hindgut lip, STELLA+/(OCT-3/4+) co-localized with FOXa2, as well as with RUNX1, indicative of definitive endoderm and hemangioblasts, respectively. STELLA was never found with MIXl1. On the basis of these and previous results, we conclude that STELLA identifies at least five distinct cell subpopulations within the allantois and hindgut, where they may be involved in mesendodermal differentiation and hematopoiesis at the posterior embryonic-extraembryonic interface. These data provide a new point of departure for understanding STELLA's potential roles in building the fetal-placental connection. Copyright © 2017 Elsevier
Kewal Arunkumar Mistry
Full Text Available Urachus or the median umbilical ligament is a fibrous strand connecting umbilicus to bladder, representing embryologic remnant of cloaca and allantois. Urachal anomalies are infrequent in adult population. Moreover they have a different course in adults than pediatric age group in which they are more common, frequently involute and have a benign course. These remnants are prone to infection and development of malignancy. A proper diagnostic workup by clinical and imaging tools is required. We present a case report of a urachal sinus complicated with abscess in an adult with brief review of the literature.
Full Text Available Angiogenesis is the generation of mature vascular networks from pre-existing vessels. Angiogenesis is crucial during the organism' development, for wound healing and for the female reproductive cycle. Several murine experimental systems are well suited for studying developmental and pathological angiogenesis. They include the embryonic hindbrain, the post-natal retina and allantois explants. In these systems vascular networks are visualised by appropriate staining procedures followed by microscopical analysis. Nevertheless, quantitative assessment of angiogenesis is hampered by the lack of readily available, standardized metrics and software analysis tools. Non-automated protocols are being used widely and they are, in general, time--and labour intensive, prone to human error and do not permit computation of complex spatial metrics. We have developed a light-weight, user friendly software, AngioTool, which allows for quick, hands-off and reproducible quantification of vascular networks in microscopic images. AngioTool computes several morphological and spatial parameters including the area covered by a vascular network, the number of vessels, vessel length, vascular density and lacunarity. In addition, AngioTool calculates the so-called "branching index" (branch points/unit area, providing a measurement of the sprouting activity of a specimen of interest. We have validated AngioTool using images of embryonic murine hindbrains, post-natal retinas and allantois explants. AngioTool is open source and can be downloaded free of charge.
Full Text Available Abstract Background Urachal carcinoma is an uncommon neoplasm associated with poor prognosis. Case presentation A 45-year-old man was admitted with complaints of abdominal pain and pollakisuria. A soft mass was palpable under his navel. TC-scan revealed a 11 × 6 cm tumor, which was composed of a cystic lesion arising from the urachus and a solid mass component at the urinary bladder dome. The tumor was removed surgically. Histological examination detected poor-differentiated adenocarcinoma, which had invaded the urinary bladder. The patient has been followed up without recurrence for 6 months. Conclusion The urachus is the embryological remnant of urogenital sinus and allantois. Involution usually happens before birth and urachus is present as a median umbilical ligament. The pathogenesis of urachal tumours is not fully understood. Surgery is the treatment of choice and role of adjuvant treatment is not clearly understood.
Rodriguez, Adriana M; Jin, Dexter X; Wolfe, Adam D; Mikedis, Maria M; Wierenga, Lauren; Hashmi, Maleka P; Viebahn, Christoph; Downs, Karen M
How the fetal-placental arterial connection is made and positioned relative to the embryonic body axis, thereby ensuring efficient and directed blood flow to and from the mother during gestation, is not known. Here we use a combination of genetics, timed pharmacological inhibition in living mouse embryos, and three-dimensional modeling to link two novel architectural features that, at present, have no status in embryological atlases. The allantoic core domain (ACD) is the extraembryonic extension of the primitive streak into the allantois, or pre-umbilical tissue; the vessel of confluence (VOC), situated adjacent to the ACD, is an extraembryonic vessel that marks the site of fetal-placental arterial union. We show that genesis of the fetal-placental connection involves the ACD and VOC in a series of steps, each one dependent upon the last. In the first, Brachyury (T) ensures adequate extension of the primitive streak into the allantois, which in turn designates the allantoic-yolk sac junction. Next, the streak-derived ACD organizes allantoic angioblasts to the axial junction; upon signaling from Fibroblast Growth Factor Receptor-1 (FGFR1), these endothelialize and branch, forming a sprouting VOC that unites the umbilical and omphalomesenteric arteries with the fetal dorsal aortae. Arterial union is followed by the appearance of the medial umbilical roots within the VOC, which in turn designate the correct axial placement of the lateral umbilical roots/common iliac arteries. In addition, we show that the ACD and VOC are conserved across Placentalia, including humans, underscoring their fundamental importance in mammalian biology. We conclude that T is required for correct axial positioning of the VOC via the primitive streak/ACD, while FGFR1, through its role in endothelialization and branching, further patterns it. Together, these genetic, molecular and structural elements safeguard the fetus against adverse outcomes that can result from vascular mispatterning of
Mikedis, Maria M; Downs, Karen M
The developmental relationship between the posterior embryonic and extraembryonic regions of the mammalian gastrula is poorly understood. Although many different cell types are deployed within this region, only the primordial germ cells (PGCs) have been closely studied. Recent evidence has suggested that the allantois, within which the PGCs temporarily take up residence, contains a pool of cells, called the Allantoic Core Domain (ACD), critical for allantoic elongation to the chorion. Here, we have asked whether the STELLA-positive cells found within this region, thought to be specified PGCs, are actually part of the ACD and to what extent they, and other ACD cells, contribute to the allantois and fetal tissues. To address these hypotheses, STELLA was immunolocalized to the mouse gastrula between Early Streak (ES) and 12-somite pair (-s) stages (~6.75-9.0 days post coitum, dpc) in histological sections. STELLA was found in both the nucleus and cytoplasm in a variety of cell types, both within and outside of the putative PGC trajectory. Fate-mapping the headfold-stage (~7.75-8.0 dpc) posterior region, by which time PGCs are thought to be segregated into a distinct lineage, revealed that the STELLA-positive proximal ACD and intraembryonic posterior primitive streak (IPS) contributed to a wide range of somatic tissues that encompassed derivatives of the three primary germ layers. This contribution included STELLA-positive cells localizing to tissues both within and outside of the putative PGC trajectory. Thus, while STELLA may identify a subpopulation of cells destined for the PGC lineage, our findings reveal that it may be part of a broader niche that encompasses the ACD and through which the STELLA population may contribute cells to a wide variety of posterior tissues of the mouse gastrula. Copyright © 2011 Elsevier Inc. All rights reserved.
Cooley Marion A
Full Text Available Abstract Background Cubilin is a peripheral membrane protein that interacts with the integral membrane proteins megalin and amnionless to mediate ligand endocytosis by absorptive epithelia such as the extraembryonic visceral endoderm (VE. Results Here we report the effects of the genetic deletion of cubilin on mouse embryonic development. Cubilin gene deletion is homozygous embryonic lethal with death occurring between 7.5–13.5 days post coitum (dpc. Cubilin-deficient embryos display developmental retardation and do not advance morphologically beyond the gross appearance of wild-type 8–8.5 dpc embryos. While mesodermal structures such as the allantois and the heart are formed in cubilin mutants, other mesoderm-derived tissues are anomalous or absent. Yolk sac blood islands are formed in cubilin mutants but are unusually large, and the yolk sac blood vessels fail to undergo remodeling. Furthermore, somite formation does not occur in cubilin mutants. Morphological abnormalities of endoderm occur in cubilin mutants and include a stratified epithelium in place of the normally simple columnar VE epithelium and a stratified cuboidal epithelium in place of the normally simple squamous epithelium of the definitive endoderm. Cubilin-deficient VE is also functionally defective, unable to mediate uptake of maternally derived high-density lipoprotein (HDL. Conclusion In summary, cubilin is required for embryonic development and is essential for the formation of somites, definitive endoderm and VE and for the absorptive function of VE including the process of maternal-embryo transport of HDL.
Deteksi Bovine Herpesvirus-1 Secara Immunohistokimia pada Membran Korioallantois Telur Ayam Berembrio (IMMUNOHISTOCHEMISTRY DETECTION OF BOVINE HERPESVIRUS-1 IN CORIOALLANTOIC MEMBRANE OF CHICKEN EMBRYONATED EGG
Yuli Purwandari Kristianingrum
Full Text Available Infectious Bovine Rhinotracheitis (IBR is caused by Bovine Herpes virus-1 in the cattle. The clinicalsigns demonstrate depression, anorexia, swelling of the vulva, redness of the vestibule, pustule and ulceron the vaginal mucosal. Based on previous research, IBR virus from the nasal swab could be grown inchorio-allantoic membrane of embryonated chicken eggs. This study aim was to confirm whether IBR virusin cattle could be grown in embryonated chicken eggs as a substitute for cell culture. A total of five nasalswab samples from the cows that were positive for IBR infection (diagnosed by Polymerase Chain Reactionand cell culture were inoculated on the chorio-allantois membrane of embryonated chicken eggs.Observation of lesions performed at 3-5 days after inoculation. Re-inoculation (passage was done threetimes. Pock characteristic lesions were observed on the corioallantoic membrane with the size of 5-7 mm,rounded shape, opaque edge, with necrosis in the central area. Furthermore, pock lesions were processedfor hematoxylin and eosin staining and immuno-histochemistry. The result of hematoxylin and eosinstaining showed that the formation of intranuclear inclusion bodies and vacuolization of the epithelial cellof membrane was observed. Immuno-histochemistry staining showed positive reaction for antibodiesagainst BHV-1 in the epithelial cells membrane. In conclusion, embryonated chicken eggs could be usedas a medium for detection of IBR.
Han, Jiayin; Liang, Aihu; Yi, Yan
To investigate the fetotoxicity of retrorsine. Mouse whole embryo culture (WEC) was applied. Post-implantation (8.5 d) mouse embryos were isolated from their mothers and put into the medium of immediately centrifuged serum (ICS) prepared from rats. Different concentrations of retrorsine (12.5, 25, 50, 100 mg x L(-1)) were added into the WEC culture. Development (yolk sac diameter, crown-rump length, head length, somite number) and organic morphodifferentiation (yolk sac circulation, allantois, embryonic flexion, heart, brain, optic-otic-olfactory organ, branchial arch, maxillary, mandible, bud) of embryos were observed at 48 h after treatment. Obvious fetotoxicity could be observed in various retrorsine treatment groups in a dose-dependent manner. Development of embryos was delayed significantly at dose 12.5-100 mg x L(-1). Malformations were shown in all organic morphodifferentiation indexes, especially in otic-olfactory organ, branchial arch, maxillary, mandible, bud. Retrorsine had obvious fetotoxicity in vitro WEC culture, indicating that exposure of pregnant mice to retrorsine may have potential risk on fetals.
Han, Jiayin; Liang, Aihua; Yi, Yan; Gao, Shuangrong; Nilsen, Odd Georg
To investigate the fetotoxicity of monocrotaline. Mouse whole embryo culture (WEC) was applied. Post-implantation (8.5 d) mouse embryos were isolated from their mothers and put into the medium of immediately centrifuged serum (ICS) prepared from rats. Different concentrations of monocrotaline (100, 50, 25, 12.5 mg x L(-1)) were added into the WEC. Development (yolk sac diameter, crown-rump length, head length, somite number) and organic morphodifferentiation (yolk sac circulation, allantois, embryonic flexion, heart, brain, optic-otic-olfactory organ, branchial arch, maxillary, mandible, bud) of embryos were observed at 48 h after treatment. Obvious fetotoxicity could be observed in various monocrotaline treatment groups in a dose-dependent manner. Development of embryos was delayed significantly at dose 12.5-100 mg x L(-1). Malformations were shown in all organic morphodifferentiation indice, especially in opti-otic organ, mandible and bud. Monocrotaline had obvious fetotoxicity in vitro WEC, indicating that exposure of pregnant mice to monocrotaline may have potential risk on fetus.
Lange-Consiglio, Anna; Corradetti, Bruna; Bertani, Sabrina; Notarstefano, Valentina; Perrini, Claudia; Marini, Maria Giovanna; Arrighi, Silvana; Bosi, Giampaolo; Belloli, Angelo; Pravettoni, Davide; Locatelli, Valentina; Cremonesi, Fausto; Bizzaro, Davide
The aim of this work was to provide, for the first time, a protocol for isolation and characterization of stem cells from porcine amniotic membrane in view of their potential uses in regenerative medicine. From three samples of allanto-amnion recovered at delivery, the amniotic membrane was stripped from overlying allantois and digested with trypsin and collagenase to isolate epithelial (amniotic epithelial cells [AECs]) and mesenchymal cells, respectively. Proliferation, differentiation, and characterization studies by molecular biology and flow cytometry were performed. Histological examination revealed very few mesenchymal cells in the stromal layer, and a cellular yield of AECs of 10 × 10(6)/gram of digested tissue was achieved. AECs readily attached to plastic culture dishes displaying typical cuboidal morphology and, although their proliferative capacity decreased to the fifth passage, AECs showed a mean doubling time of 24.77 ± 6 h and a mean frequency of one fibroblast colony-forming unit (CFU-F) for every 116.75 plated cells. AECs expressed mesenchymal stem cell (MSC) mRNA markers (CD29, CD166, CD90, CD73, CD117) and pluripotent markers (Nanog and Oct 4), whereas they were negative for CD34 and MHCII. Mesodermic, ectodermic, and endodermic differentiation was confirmed by staining and expression of specific markers. We conclude that porcine amniotic membrane can provide an attractive source of stem cells that may be a useful tool for biomedical research.
Bonnet, Xavier; Naulleau, Guy; Shine, Richard
The parchment-shelled eggs of squamate reptiles take up substantial water from the nest environment, enabling the conversion of yolk into neonatal tissue and buffering the embryo against the possibility of subsequent dry weather. During development, increasing amounts of water are stored in the embryonic sacs (i.e., membranes around the embryo: amnion, allantois, and chorion). The evolution of viviparity (prolonged uterine retention of developing embryos) means that embryonic-sac fluid storage now imposes a cost (increased maternal burdening), confers less benefit (because the mother buffers fetal water balance), and introduces a potential conflict among uterine siblings (for access to finite water supplies). Our data on nine species of squamate reptiles and published information on three species show that the embryonic-sac fluids comprise around 33% of neonatal mass in viviparous species versus 94% in full-term eggs of oviparous squamates. Data on parturition in 149 vipers (Vipera aspis, a viviparous species) show that larger offspring store more fluids in their fetal sacs and that an increase in litter size is associated with a decrease in fluid-sac mass per offspring. Overall, the evolutionary transition from oviparity to viviparity may have substantially altered selective forces on offspring packaging and created competition among offspring for access to water reserves during embryonic development.
Sartori, M R; Taylor, E W; Abe, A S
Development within the cleidoic egg of birds and reptiles presents the embryo with the problem of accumulation of wastes from nitrogen metabolism. Ammonia derived from protein catabolism is converted into the less toxic product urea or relatively insoluble uric acid. The pattern of nitrogen excretion of the green iguana, Iguana iguana, was determined during embryonic development using samples from allantoic fluid and from the whole homogenized egg, and in hatchlings and adults using samples of blood plasma. Urea was the major excretory product over the course of embryonic development. It was found in higher concentrations in the allantoic sac, suggesting that there is a mechanism present on the allantoic membrane enabling the concentration of urea. The newly hatched iguana still produced urea while adults produced uric acid. The time course of this shift in the type of nitrogen waste was not determined but the change is likely to be related to the water relations associated with the terrestrial habit of the adult. The green iguana produces parchment-shelled eggs that double in mass during incubation due to water absorption; the eggs also accumulate 0.02 mM of urea, representing 82% of the total measured nitrogenous residues that accumulate inside the allantois. The increase in egg mass and urea concentration became significant after 55 days of incubation then were unchanged until hatching. Copyright © 2012 Elsevier Inc. All rights reserved.
Stewart, James R; Thompson, Michael B
The reptilian placenta is a composite structure formed by a functional interaction between extraembryonic membranes and the maternal uterus. Study of placental structure of squamate reptiles over the past century has established that each of the multiple independent origins of placentation, which characterize the reproductive diversity of squamates, has resulted from the evolutionary transformation of these homologous structures. Because each evolutionary transformation is an independent novel relationship between maternal and embryonic tissues, the resulting placentae are not homologous, even though the individual components may be. The evolution of reptilian placentation should reveal much about evolutionary patterns and mechanisms because similar structural-functional systems have been transformed along parallel trajectories on multiple occasions. We compared extraembryonic membrane and placental development and pattern of embryonic nutrition in thamnophiine snakes and Pseudemoia lizards in the context of recent hypotheses of phylogenetic relationships. Two primary types of placentation, chorioallantoic and yolk sac, evolved in each lineage. Smooth, highly vascular regions of chorioallantoic placentation are indistinguishable homoplasies that evolved in parallel, likely to facilitate respiratory exchange. The yolk sac placenta of each lineage is specialized for histotrophic nutrient transfer, yet composition of these structures differs because of variation in the ancestral snakes and lizards. In addition, the omphalopleure that contributes to yolk sac placentation persists to later embryonic stages compared to oviparous outgroups, but the two lineages have evolved different structures that prevent replacement of the omphalopleure by the allantois. Each lineage has also evolved unique structural specializations of the chorioallantoic placenta. Copyright 2003 Wiley-Liss, Inc.
Fidanza, Antonella; Toschi, Paola; Zacchini, Federica; Czernik, Marta; Palmieri, Chiara; Scapolo, PierAugusto; Modlinski, Jacek A; Loi, Pasqualino; Ptak, Grazyna E
To evaluate how assisted reproductive technologies (ART) affect vasculogenesis of the developing conceptus, we analyzed placental and fetal development of in vitro-produced (IVP) sheep embryos. Pregnancies produced by ART carry increased risk of low birth weight, though what causes this risk remains largely unknown. We recently reported that developmental arrest of sheep conceptuses obtained by ART is most pronounced when the cardiovascular system develops (Days 20-30 of development). A total of 86 IVP blastocysts (2-4 per ewe) were surgically transferred to 30 recipient sheep 6 days after estrus; 20 sheep were naturally mated (control). Conceptuses were recovered from sheep at Days 20, 22, 26, and 30 of gestation and morphologically evaluated. Then, the conceptuses and part of their placentae (chorion-allantois) were fixed for histological and immunohistochemical analysis and snap-frozen in liquid nitrogen for subsequent mRNA expression analysis. Results demonstrate that the cardiovascular systems of sheep IVP conceptuses were severely underdeveloped. Pericardial and placental hemorrhages were noted in a majority (5/7) of the dead embryos. In the surviving IVP embryos, the expression of angiogenetic factors was reduced at Day 20. The placental vessels were underdeveloped on Days 20 and 22 (P sheep embryos induced severely impaired vasculogenesis early in gestation. This may lead to developmental programing problems, such as intrauterine growth restriction of the fetus, resulting in long-term health consequences for the offspring, such as cardiovascular diseases. © 2014 by the Society for the Study of Reproduction, Inc.
Ramírez-Pinilla, Martha Patricia; Parker, Scott L; Murphy, Christopher R; Thompson, Michael B
We used immunofluorescent confocal microscopy and scanning electron microscopy to quantify uterine vascularity and to describe uterine surface morphology during gestation in pregnant females of the lecithotrophic lizard Niveoscincus coventryi. As uterine angiogenesis and epithelial cell morphology are thought to be under progesterone control, we studied the effect of a progesterone receptor antagonist (mifepristone) on uterine and chorioallantoic microvasculature and features of the uterine epithelial surfaces. Although intussuceptive angiogenesis was observed in both, uterine and chorioallantoic, vascular beds during gestation, the only significant increases were in the diameters of the uterine vessels. An ellipsoid vessel-dense area grows in the mesometrial hemisphere of the developing conceptus, which parallels the expansion of the allantois to form the chorioallantoic placenta. Uterine surface topography changed during gestation. In particular, uterine blood vessels bulge over the luminal surface to form marked ridges on the uterine embryonic hemisphere, especially during the last stage of pregnancy, and ciliated cells are maintained in the embryonic and abembryonic hemispheres but disappear in both the mesometrial and antimesometrial poles. This distinct regionalization of uterine ridges and ciliated cells in the uterine surface and in the shape of the epithelial component of the chorion might be related to the function of both chorioallantoic and yolk sac placentae during gestation. There was no significant difference between females treated with or without mifepristone, which may be related to the partial function of mifepristone as a progestin antagonist and/or with the function and time of action of progesterone in the uterus during gestation in N. coventryi. Differences in the pattern of angiogenesis and uterine surface morphology during gestation among squamates may be related to the functional diversity of the uterine component of the different
Cañete, Ana; Carmona, Rita; Ariza, Laura; Sánchez, María José; Rojas, Anabel; Muñoz-Chápuli, Ramón
GATA transcription factors are expressed in the mesoderm and endoderm during development. GATA1-3, but not GATA4, are critically involved in hematopoiesis. An enhancer (G2) of the mouse Gata4 gene directs its expression throughout the lateral mesoderm and the allantois, beginning at embryonic day 7.5, becoming restricted to the septum transversum by embryonic day 10.5, and disappearing by midgestation. We have studied the developmental fate of the G2-Gata4 cell lineage using a G2-Gata4 Cre ;R26R EYFP mouse line. We found a substantial number of YFP + hematopoietic cells of lymphoid, myeloid and erythroid lineages in embryos. Fetal CD41 + /cKit + /CD34 + and Lin - /cKit + /CD31 + YFP + hematopoietic progenitors were much more abundant in the placenta than in the aorta-gonad-mesonephros area. They were clonogenic in the MethoCult assay and fully reconstituted hematopoiesis in myeloablated mice. YFP + cells represented about 20% of the hematopoietic system of adult mice. Adult YFP + hematopoietic stem cells constituted a long-term repopulating, transplantable population. Thus, a lineage of adult hematopoietic stem cells is characterized by the expression of GATA4 in their embryonic progenitors and probably by its extraembryonic (placental) origin, although GATA4 appeared not to be required for hematopoietic stem cell differentiation. Both lineages basically showed similar physiological behavior in normal mice, but clinically relevant properties of this particular hematopoietic stem cell population should be checked in physiopathological conditions. Copyright© Ferrata Storti Foundation.
Full Text Available Steel factor, the protein product of the Steel locus in the mouse, is a multifunctional signal for the primordial germ cell population. We have shown previously that its expression accompanies the germ cells during migration to the gonads, forming a "travelling niche" that controls their survival, motility, and proliferation. Here we show that these functions are distributed between the alternatively spliced membrane-bound and soluble forms of Steel factor. The germ cells normally migrate as individuals from E7.5 to E11.5, when they aggregate together in the embryonic gonads. Movie analysis of Steel-dickie mutant embryos, which make only the soluble form, at E7.5, showed that the germ cells fail to migrate normally, and undergo "premature aggregation" in the base of the allantois. Survival and directionality of movement is not affected. Addition of excess soluble Steel factor to Steel-dickie embryos rescued germ cell motility, and addition of Steel factor to germ cells in vitro showed that a fourfold higher dose was required to increase motility, compared to survival. These data show that soluble Steel factor is sufficient for germ cell survival, and suggest that the membrane-bound form provides a higher local concentration of Steel factor that controls the balance between germ cell motility and aggregation. This hypothesis was tested by addition of excess soluble Steel factor to slice cultures of E11.5 embryos, when migration usually ceases, and the germ cells aggregate. This reversed the aggregation process, and caused increased motility of the germ cells. We conclude that the two forms of Steel factor control different aspects of germ cell behavior, and that membrane-bound Steel factor controls germ cell motility within a "motility niche" that moves through the embryo with the germ cells. Escape from this niche causes cessation of motility and death by apoptosis of the ectopic germ cells.
Yon, Jung-Min; Baek, In-Jeoung; Lee, Beom Jun; Yun, Young Won; Nam, Sang-Yoon
The balance between reactive oxygen species production and antioxidant defense enzymes in embryos is necessary for normal embryogenesis. To determine the dynamic expression profile of manganese superoxide dismutase (MnSOD) in embryos, which is an essential antioxidant enzyme in embryonic organogenesis, the expression level and distribution of MnSOD mRNA and protein were investigated in mouse embryos, as well as extraembryonic tissues on embryonic days (EDs) 7.5-18.5. MnSOD mRNA levels were remarkably high in extraembryonic tissues rather than in embryos during these periods. MnSOD protein levels were also higher in extraembryonic tissues than in embryos until ED 16.5, but the opposite trend was found after ED 17.5. MnSOD mRNA was observed in the chorion, allantois, amnion, ectoderm, ectoplacental cone and neural fold at ED 7.5 and in the neural fold, gut, ectoplacental cone, outer extraembryonic membranes and primitive heart at ED 8.5. After removing the extraembryonic tissues, the prominent expression of MnSOD mRNA in embryos was seen in the sensory organs, central nervous system and limbs on EDs 9.5-12.5 and in the ganglia, spinal cord, sensory organ epithelia, lung, blood cells and vessels, intestinal and skin epithelia, hepatocytes and thymus on EDs 13.5-18.5. Strong MnSOD immunoreactivity was observed in the choroid plexus, ganglia, myocardium, blood vessels, heapatocytes, pancreatic acinus, osteogenic tissues, brown adipose tissue, thymus and skin. These findings suggest that MnSOD is mainly produced from extraembryonic tissues and then may be utilized to protect the embryos against endogenous or exogenous oxidative stress during embryogenesis.
Full Text Available Protein Kinase C delta and epsilon are mediators of important cellular events, such as cell proliferation, migration or apoptosis. The formation of blood vessels, i.e., vasculo- and angiogenesis, is a process where these isoforms have also been shown to participate. However, mice deficient in either Protein Kinase C delta or epsilon are viable and therefore their individual contribution to the formation of the vasculature appeared so far dispensable. In this study, we show that double null mutation of Protein Kinase C delta and epsilon causes embryonic lethality at approximately E9.5. At this stage, whole mount staining of the endothelial marker CD31 in double null embryos revealed defective blood vessel formation. Moreover, culture of double deficient mouse allantois showed impaired endothelial cell organization, and analyses of double deficient embryo sections showed dilated vessels, decreased endothelial-specific adherent junctions, and decreased contact of endothelial cells with mural cells. Protein kinase C delta and epsilon also appeared essential for vascular smooth muscle cell differentiation, since α-smooth muscle actin, a classical marker for vascular smooth muscle cells, was almost undetectable in double deficient embryonic aorta at E9.5. Subsequent qPCR analyses showed decreased VE-cadherin, Vegfr2, Cd31, Cdh2, Ets1, and Fli-1, among other angiogenesis related transcripts in double deficient embryos. Taken together, these data suggest for the first time an in vivo redundant role between members of the novel Protein Kinase C subfamily that allows for mutual compensation during mouse embryonic development, with vasculogenesis/angiogenesis as an obvious common function of these two Protein Kinase Cs. Protein Kinase C delta and epsilon might therefore be useful targets for inhibiting vasculo- and/or angiogenesis.
Hirsch, Tobias; Laemmle, Christine; Behr, Bjoern; Lehnhardt, Marcus; Jacobsen, Frank; Hoefer, Dirk; Kueckelhaus, Maximilian
Adipose-derived stem cells represent an interesting option for soft tissue replacement as they are easy to procure and can generate their own blood supply through the production of angiogenic factors. We seeded adipose-derived stem cells on a bioresorbable, biocompatible polymer alginate scaffold to generate autologous soft tissue constructs for repair. We built and optimized an alginate scaffold and tested its biocompatibility using the MTT assay and its hydration capacity. We then isolated, characterized, and differentiated murine, porcine, and human adipose-derived stem cells. We characterized their angiogenic potential in vitro by VEGF ELISA and HUVEC tube formation assay in traditional cell culture substrate and in the actual three-dimensional scaffold. We assessed the angiogenic potential of adipose-derived stem cell-colonized scaffolds in ovo by chorion allantois membrane angiogenesis assay. Adipose-derived stem cells differentiated into adipocytes within the alginate scaffolds and demonstrated angiogenic activity. VEGF secretion by adipose-derived stem cells decreased significantly over the 21-day course of adipocyte differentiation in traditional cell culture substrate, but not in scaffolds. Adipose-derived stem cells differentiated for 21 days in scaffolds led to the longest HUVEC tube formation. Scaffolds colonized with adipose-derived stem cells resulted in significantly improved vascularization in ovo. We demonstrate the feasibility of implant production based on adipose-derived stem cell-colonized alginate scaffolds. The implants demonstrate biocompatibility and promote angiogenesis in vitro and in ovo. Therefore, they provide a combination of essential properties for an implant intended for soft tissue replacement. Copyright © 2017 British Association of Plastic, Reconstructive and Aesthetic Surgeons. Published by Elsevier Ltd. All rights reserved.
Murphy, Bridget F; Brandley, Matthew C; Murphy, Christopher R; Thompson, Michael B
Frequent evolutionary changes in reproductive mode have produced a wide range of placental structures in viviparous squamate reptiles. Closely related species with different placental structures and resolved phylogenetic relationships are particularly useful for reconstructing how placentae might have transformed during the evolutionary process. We used light microscopy to study placental morphology in mid- to late stage embryos of four closely related species of Eulamprus, a genus of viviparous scincid lizards that we had reason to suspect may display significant interspecific variation in placental morphology. Embryos from all four species possess a chorioallantoic placenta, an omphaloplacenta and an interomphalopleuric membrane, characteristics present in other viviparous skinks. However, unlike other viviparous skinks but characteristic of oviparous skinks, the allantois expands to surround the yolk sac in each species, supplanting the omphalopleure with a larger area of chorioallantois until a chorioallantoic placenta surrounds the entire egg in one specimen that is only a few days from birth. All four Eulamprus species share the same extraembryonic membrane morphology, but the cellular morphology of the uterine epithelium in the chorioallantoic placenta and omphaloplacenta varies between species. We determined that the interomphalopleuric membrane is a shared derived character of the Eulamprus quoyii species group. New phylogenetic information indicates that variation in the chorioallantoic placenta is a result of two independent transitions, but that variation in the omphaloplacenta can be explained using a single change within the species studied. Our results indicate that E. quoyii group skinks are a valuable model for investigating the evolution of viviparity, as extraembryonic membrane development in these species shows features characteristic of both oviparous and viviparous skinks. PMID:22420511
Kim, Young K; Blackburn, Daniel G
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. © 2015 Wiley Periodicals, Inc.
Concepcion, Daniel; Washkowitz, Andrew J; DeSantis, Akiko; Ogea, Phillip; Yang, Jason I; Douglas, Nataki C; Papaioannou, Virginia E
Tbx6 is a T-box transcription factor with multiple roles in embryonic development as evidenced by dramatic effects on mesoderm cell fate determination, left/right axis determination, and somite segmentation in mutant mice. The expression of Tbx6 is restricted to the primitive streak and presomitic mesoderm, but some of the phenotypic features of mutants are not easily explained by this expression pattern. We have used genetically-inducible fate mapping to trace the fate of Tbx6 -expressing cells in wild-type and mutant embryos to explain some of the puzzling features of the mutant phenotype. We created an inducible Tbx6-creERT2 transgenic mouse in which cre expression closely recapitulates endogenous Tbx6 expression both temporally and spatially. Using a lacZ-based Cre reporter and timed tamoxifen injections, we followed temporally overlapping cohorts of cells that had expressed Tbx6 and found contributions to virtually all mesodermally-derived embryonic structures as well as the extraembryonic allantois. Contribution to the endothelium of major blood vessels may account for the embryonic death of homozygous mutant embryos. In mutant embryos, Tbx6-creERT2-traced cells contributed to the abnormally segmented anterior somites and formed the characteristic ectopic neural tubes. Retention of cells in the mutant tail bud indicates a deficiency in migratory behavior of the mutant cells and the presence of Tbx6-creERT2-traced cells in the notochord, a node derivative provides a possible explanation for the heterotaxia seen in mutant embryos. © 2017. Published by The Company of Biologists Ltd.
Full Text Available Tbx6 is a T-box transcription factor with multiple roles in embryonic development as evidenced by dramatic effects on mesoderm cell fate determination, left/right axis determination, and somite segmentation in mutant mice. The expression of Tbx6 is restricted to the primitive streak and presomitic mesoderm, but some of the phenotypic features of mutants are not easily explained by this expression pattern. We have used genetically-inducible fate mapping to trace the fate of Tbx6-expressing cells in wild-type and mutant embryos to explain some of the puzzling features of the mutant phenotype. We created an inducible Tbx6-creERT2 transgenic mouse in which cre expression closely recapitulates endogenous Tbx6 expression both temporally and spatially. Using a lacZ-based Cre reporter and timed tamoxifen injections, we followed temporally overlapping cohorts of cells that had expressed Tbx6 and found contributions to virtually all mesodermally-derived embryonic structures as well as the extraembryonic allantois. Contribution to the endothelium of major blood vessels may account for the embryonic death of homozygous mutant embryos. In mutant embryos, Tbx6-creERT2-traced cells contributed to the abnormally segmented anterior somites and formed the characteristic ectopic neural tubes. Retention of cells in the mutant tail bud indicates a deficiency in migratory behavior of the mutant cells and the presence of Tbx6-creERT2-traced cells in the notochord, a node derivative provides a possible explanation for the heterotaxia seen in mutant embryos.
Ptak, Grazyna Ewa; D'Agostino, Antonella; Toschi, Paola; Fidanza, Antonella; Zacchini, Federica; Czernik, Marta; Monaco, Federica; Loi, Pasqualino
Is DNA methyltransferase 1 (DNMT1) dysfunction involved in epigenetic deregulation of placentae from embryos obtained by assisted reproduction technologies (ARTs)? DNMT1 expression in growing placentae of in vitro produced (IVP) embryos is compromised and associated with pregnancy loss. DNMT1 maintains the methylation profile of genes during cell division. The methylation status of genes involved in placenta development is altered in embryos obtained in vitro. Disturbances in the epigenetic regulation of gene expression during placentogenesis could be involved in the frequent developmental arrest and loss of IVP embryos. Forty sheep were naturally mated (Group 1, CTR). IVP blastocysts (2-4 per ewe) were surgically transferred to the remaining 46 recipient sheep 6 days after oestrus (Group 2). Twenty-one recipients from Group 1 and 27 recipients from Group 2 were allowed to deliver in order to compare embryo survival in both groups at term (150 days). From the remaining recipients (n = 38), fetuses and placentae of both groups were recovered by paramedian laparotomy at Days 20, 22, 24, 26 and 28 of gestation. Immediately after collection, early placental tissues (chorion-allantois) were snap frozen in liquid nitrogen and DNMT1 expression and activity was evaluated. mRNA levels (for DNMT1, HDAC2, PCNA, DMAP1, MEST, IGF2, CDKN1C, H19) and the methylation status of H19 were also analyzed. Furthermore, embryo size and survival rate were measured. Our study shows that DNMT1 expression was reduced in early placentae from sheep IVP embryos. This reduction was associated with growth arrest and subsequent death of the sheep embryos. Conversely, normal levels of DNMT1 and its cofactors were observed in placentae from IVP embryos that survived this developmental bottleneck. Although DNA methylation machinery was severely compromised in IVP placentae only up to Day 24, the low DNMT1 enzymatic activity that persisted after this stage in IVP placentae was not lethal for the
Montague, Idoreyin P.
The Microgravity Research Program is part of NASA's Office of Biological and Physical Research (OBPR). The mission of the Microgravity Fluid Physics research program is to facilitate and conduct the best possible fluid physics research using the space environment and make this knowledge available to the scientific community and the public at large. During the summer of 2004, I worked in this division with Dr. Patricia Parsons-Wingerter. Dr. Parsons was working on several projects that used the chorioallantoic membrane (CAM) of Japanese quail eggs. The CAM develops in the eggs of birds and reptiles and is a very vascular fetal membrane composed of the fused chorion and adjacent wall of the allantois. The CAM is formed on day 4 of incubation and its primary job is to mediate gas exchanges with the extra embryonic environment. The CAM of our Japanese quail eggs is easily identifiable to us because it is transparent and it sits on top of the yolk with the embryo in the center. The CAM is of interest because of its many applications in the field of medicine as it relates to vascular remodeling and angiogenesis. Angiogenesis is simply the growth or formation of new blood vessels and anti-angiogenesis is the inhibition of said vessels. Angiogenesis occurs naturally in a healthy body for healing wounds and for restoring blood flow to tissues after injury and in females during the monthly reproductive cycle. In many serious diseases, like several types of cancer and those that affect the heart and cardiovascular system, the body loses control over angiogenesis. These diseases, which are dependent on angiogenesis, result when new blood vessels either grow excessively or insufficiently. The chorioallantoic membrane of our Japanese quail eggs gives a good model of angiogenesis. We used angiogenic regulators to inhibit or stimulate vascular growth in the CAM in a healthy manner and they induced distinct vascular patterns in vivo. Certain dominant regulators can be recognized by
Full Text Available In addition to nourishing the embryo, extra-embryonic tissues (EETs contribute to early embryonic patterning, primitive hematopoiesis, and fetal health. These tissues are of major importance for human medicine, as well as for efforts to improve livestock efficiency, but they remain incompletely understood. In bovines, EETs are accessible easily, in large amounts, and prior to implantation. We took advantage of this system to describe, in vitro and in vivo, the cell types present in bovine EETs at Day 18 of development. Specifically, we characterized the gene expression patterns and phenotypes of bovine extra-embryonic ectoderm (or trophoblast; bTC, endoderm (bXEC, and mesoderm (bXMC cells in culture and compared them to their respective in vivo micro-dissected cells. After a week of culture, certain characteristics (e.g., gene expression of the in vitro cells were altered with respect to the in vivo cells, but we were able to identify "cores" of cell-type-specific (and substrate-independent genes that were shared between in vitro and in vivo samples. In addition, many cellular phenotypes were cell-type-specific with regard to extracellular adhesion. We evaluated the ability of individual bXMCs to migrate and spread on micro-patterns, and observed that they easily adapted to diverse environments, similar to in vivo EE mesoderm cells, which encounter different EE epithelia to form chorion, yolk sac, and allantois. With these tissue interactions, different functions arose that were detected in silico and corroborated in vivo at D21-D25. Moreover, analysis of bXMCs allowed us to identify the EE cell ring surrounding the embryonic disc (ED at D14-15 as mesoderm cells, which had been hypothesized but not shown prior to this study. We envision these data will serve as a major resource for the future in the analysis of peri-implanting phenotypes in response to the maternal metabolism and contribute to subsequent studies of placental/fetal development in
Administered into the yolk sac of eggs of Lacerta viridis as a single dose of 17 to 40 micrograms, cytosine-arabinoside (Ara-C) was compatible with survival of the embryo, from the sixth day of incubation, for at least 20 to 25 days. The LD50 was 40 to 50 micrograms per egg. Doses of 20 to 40 micrograms of Ara-C introduced in the yolk sac of eggs of the slow-worm (Anguis fragilis) cultured in vitro, at stages of the allantoid bud of 0,5 mm to 2,5 mm long, killed the embryo in 4 to 8 days (possibly due to alterations of capillary blood vessels of allantois and area vasculosa). In the two species, these doses caused cytotoxic effects on embryonic proliferating tissues, growth inhibition and a variety of developmental defects. In young embryos of Anguis fragilis, similar doses of 20 to 40 micrograms of Ara-C caused, in 2 to 4 days, death of many cells in the anlagen of growing organs: neural tube, sensory organs, bronchi, mesoderm of the limb bud, subcutaneous mesenchyme, anlage of dorsal skeletal structures, etc.; followed by growth inhibition and malformations. On the other hand, in the limb bud, the apical ridge was less retrogressed than in control embryos; the limb buds showed slightly better development in treated embryos than in controls, but, Ara-C induced severe damage in their mesoderm. In all embryos of Lacerta viridis, treated at the stage of 6 days or of 10 days of incubation by doses of 20 to 40 micrograms of Ara-C and killed 15 to 35 days later, there was a general reduction of size and of weight and external and internal malformations, more or less severe, were present: modifications of the form of the head, shortening of the lower jaw, labial clefts, microphthalmia, micromelia and other limbs defects, developmental defects of the tail. In some embryos, the only external defects observed were missing fingers and toes; in three of these embryos, the same digits were missing in the four limbs. Modifications of limb morphogenesis induced by Ara-C are
Meloni, Tea; Comin, Antonella; Rota, Alessandro; Peric, Tanja; Contri, Alberto; Veronesi, Maria Cristina
between amnion and allantois, but in both fetal fluids, higher NEFA levels were detected in samples belonging to small breeds when compared with medium and large. These data strongly indicated that, also in the dog, a relation between fetal fluids IGF-I and NEFA concentrations and breed size exists. Further research is needed to elucidate the possible role of IGF-I and NEFA in the pathologic conditions related to canine fetal growth. Copyright © 2014 Elsevier Inc. All rights reserved.
J. H. Khorajiya
-3 weeks and reduced with appearance of torticollis. Gross lesions were characterized by multifocal to diffuse hemorrhages around proventricular glands, necrotic (diphtheritic haemorrhagic ulcers throughout the intestine, disseminated multiple foci of necrosis and pin-point hemorrhages in the spleen parenchyma. The microscopic lesions include focal to diffuse hemorrhages, diffuse infiltration of mononuclear cells, necrosis, and degeneration in visceral organs. All the 13 farm samples (n=13 resulted in death of all the embryos following incubation up to 72 h post-inoculation. All the 13 allantois fluids from field samples along with F and R2B vaccine sample were found positive for HA activity, which was further confirmed by HI using known NDV serum. The values of ICPI were 2.0 which were indicative of velogenic nature of the field NDV strain. Conclusion: The study indicated that presently available live and attenuated vaccines which include Genotype-II NDV have failed in protecting the flocks against Genotype-XIII and resulted in outbreaks with mortality above 50%. ICPI score of 2.0 confirmed that the present outbreaks were due to Genotype-XIII NDV, which is velogenic in nature.