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PubMed2006

Endodermal cyst of the oculomotor nerve: case report.

OBJECTIVE: Endodermal cysts are commonly found in the anterior spinal cord. Reports of intracranial cases are uncommon. We report a case of endodermal cyst located on the left oculomotor nerve. CLINICAL PRESENTATION: A 16-year-old female experienced diplopia followed by headache. Neurological examinations revealed left oculomotor nerve palsy. Magnetic resonance imaging scans showed a well-defined multilocular cystic mass in the proximity of the left oculomotor nerve at its exit from the midbrain. INTERVENTION: This patient underwent biopsy and drainage of the cyst. At surgery, the cyst seemed to be intrinsic to the oculomotor nerve and could not be totally resected. Pathological examination confirmed the diagnosis of endodermal cyst. CONCLUSION: Intracranial endodermal cysts are rare, particularly when associated with the cranial nerves. To our knowledge, this is the second case of endodermal cyst located on the oculomotor nerve. A detailed discussion and review of these case reports are presented.

Adolescent↗
PubMed1982

Developmental expression of murine extra-embryonic endodermal cytoskeletal proteins.

Murine extra-embryonic endodermal cell lines derived from either teratocarcinomas or mouse embryos contain a cytoskeletal protein (Endo A) of Mr = 55,000. Endo A was immunoprecipitated from [35S]methionine-labeled lysates of three parietal endodermal cell lines, A presumptive visceral endodermal cell line, and a fetal hepatoma cell line, but not from fibroblasts, myoblasts, erythroleukemic cells, neuroblastoma cells, keratinocytes, or embryonal carcinoma cells. Embryonal carcinoma cells induced to differentiate by exposure to retinoic acid synthesized increased amounts of Endo A approximately 48 h after exposure to the inducer. Two-dimensional gel analysis of immunoprecipitated samples confirmed that Endo A is distinct from vimentin and murine keratinocyte proteins recognized by two different keratin antisera. Comparison by two-dimensional gel electrophoresis of immunoprecipitated Endo A labeled with either [35S]methionine or [32P]orthophosphate indicated that the multiple forms of Endo A resolved by isoelectric focusing were due, at least in part, to phosphorylation. Serine was identified as the phosphorylated amino acid. Endo A was the only major antigenic protein found in a parietal endodermal cell line which was recognized by a monoclonal antibody prepared by other investigators against trophoblast cytoskeletons. The results indicate that Endo A, like the previously described Endo B protein, is distinct from other cytoskeletal proteins and will be useful as a marker of the differentiation of murine embryonal carcinoma cells to extra-embryonic endoderm.

Animals↗
PubMed1983

Characterization of cytosolic calcium-activated phospholipid-dependent protein kinase activity in embryonal carcinoma cells. Effect of retinoc acid-induced differentiation of F9 cells to parietal endoderm.

We have addressed the question of the possible presence of calcium-activated phospholipid-dependent protein kinase (Ca2+-PL protein kinase) activity in undifferentiated embryonal carcinoma cells, and if this activity might be altered during differentiation to a parietal endoderm cell type. Undifferentiated nullipotent F9 embryonal carcinoma cells, as well as differentiated parietal endoderm cells (PYS-2), were utilized. Using an in vitro assay with histone H1 as phosphate acceptor, Ca2+-PL protein kinase activity could not be found in the 100,000 X g supernatant prepared from either cell type. However, passage of 100,000 X g supernatant from PYS cells over a DEAE-cellulose column revealed Ca2+-PL protein kinase activity which eluted with 0.045 M NaCl. The partially purified PYS enzyme has an approximate Mr = 70,000 as determined by Sephadex G-150 gel filtration, and exhibits an apparent Ka for Ca2+ of 32 microM. The PYS Ca2+-PL protein kinase also exhibits a requirement for Mg2+, with maximal activity noted at 10 mM Mg2+. This enzyme is stimulated by acidic phospholipids, while neutral phospholipids such as phosphatidylcholine have little effect. Diacylglycerol markedly increased histone H1 phosphorylation in the presence of Ca2+ and phospholipid. Unlike that of PYS cells, when the 100,000 X g supernatant prepared from F9 cells was passed over a DEAE-cellulose column no Ca2+-PL protein kinase activity could be found in the eluted fractions. Previously it has been reported that exposure of F9 cells to all-trans-retinoic acid induces differentiation to a parietal endoderm cell type. Treatment of F9 cells with 0.1 microM retinoic acid provoked a time-dependent increase in cytosolic Ca2+-PL protein kinase activity as measured after DEAE-cellulose chromatography of the 100,000 X g supernatant. This increase in Ca2+-PL protein kinase activity correlates with differentiation to the parietal endoderm cell type. These findings indicate that cytosolic Ca2+-PL protein kinase activity is very low, or nonexistent, in undifferentiated embryonal carcinoma stem cells. With differentiation to a parietal endoderm cell type there is a marked increase in soluble Ca2+-PL protein kinase activity which exhibits properties similar to those described for this enzyme in other differentiated tissues.

Animals↗
PubMed2025

Enhancer remodeling by OTX2 directs specification and patterning of mammalian definitive endoderm.

The molecular mechanisms that drive essential patterning events in the mammalian embryo remain poorly understood. Analysis of transcription factor expression kinetics at peri-gastrulation stages of development suggest Otx2 as a candidate regulator of the definitive endoderm, the precursor of all gut-derived organs. Accordingly, timed OTX2 depletion in gastruloids or during directed differentiation results in abnormal definitive endoderm specification in mouse and human, characterized by altered expression of components and transcriptional targets of the canonical WNT signaling pathway, perturbed adhesion and migration programs, and de-repression of regulators of other lineages. These defects cumulate in impaired foregut formation. Mechanistically, OTX2 is required to activate a subset of endoderm-specific enhancers and to suppress select enhancers of other lineages, allowing timely exit from the primitive streak and correct specification of anterior endoderm. Our results establish OTX2 as an early gut regulator and suggest molecular principles underlying spatiotemporal cell identity conserved across germ layers and species.

Otx Transcription Factors↗
PubMed1999

Domains of differential cell proliferation suggest hinged folding in avian gut endoderm.

A profile of proliferative growth assessed with tritium autoradiograms from White Leghorn embryo stages Hamburger-Hamilton 6-21 labeled in ovo presents evidence of hinged folding driven by localized differential cell proliferation in endoderm. There is a significant, bilateral pattern, and differences are most pronounced in axial levels that are folding and rotating. Highest proliferation is in cells producing folds; lowest proliferation is in median cells. Localized changes in cell shape are lacking, as are TUNEL markers and cell morphology that would suggest involvement of apoptosis. Folding endoderm to form gut tube appears to be a process that is driven by domains of high cell proliferation flanking a domain of significantly lower proliferation. When considered in the context of an epithelium attached to subjacent mesoderm, these differentials could produce a forced and directed buckling of endoderm into lateral folds that join and enclose a tube. Patterns suggest that endoderm folds about a median hinge. In the light of this new information, we suggest it is more precise to refine the term, median hinge point (MHP), to neural hinge point (NHP) and gut hinge point (GHP).

Analysis of Variance↗
PubMed2000

Endoderm and heart development.

Since the first half of the 20th century, experimental embryologists have noted a relationship between endoderm cells and the development of cardiac tissue from mesoderm. During the past decade, the accumulation of evidence for an obligatory interaction between endoderm and mesoderm during the specification and terminal differentiation of myocardial, and more recently endocardial, cells has markedly accelerated. Moreover, the endoderm-derived molecules that may regulate these processes are being identified. It now appears that endoderm-derived growth factors regulate the formation of both myocardial and endocardial cells during specification, terminal differentiation, and perhaps morphogenesis of cells in the developing embryonic heart.

Animals↗
PubMed1994

Anterior endoderm is a specific effector of terminal cardiac myocyte differentiation of cells from the embryonic heart forming region.

The ability of anterior lateral plate mesoderm cells in the heart-forming region (HFR) of stage 6 chicken embryos to respond to cardiogenic stimuli from cells in adjacent germ layers has been investigated using explants cultured under defined conditions. Two types of explantation were evaluated: those in which two germ layers were explanted in contiguity, and those in which germ layers were isolated and co-cultured. Two parameters--contractility and expression of sarcomeric alpha-actin--were monitored to evaluate the terminal differentiation of cardiac myocytes. Contiguously explanted anterior endoderm/mesoderm became multilayered and underwent terminal differentiation within 2 days. By contrast, although contiguous anterior ectoderm/mesoderm or posterior endoderm/mesoderm co-explants also became multilayered, these explants did not differentiate, up to 5 days. To ascertain the cardiogenic potential of cells from different regions of the embryo, individual germ layers were isolated and co-cultured by placing the explants in separate areas of the culture chamber. These determinations demonstrated that anterior, but not posterior, endoderm effected differentiation of anterior mesoderm. As before, mesoderm in both types of co-culture survived and became multilayered; by contrast, mesoderm did not survive when cultured in isolation. These experiments provide evidence that anterior endoderm regulates the terminal differentiation, as opposed to growth, of presumptive cardiac myocytes in mesoderm cells from the anterior lateral plate. Finally, anterior endoderm was co-cultured with mesoderm from the posterior half of the embryo, which does not contain an HFR. The failure of these co-cultured explants to differentiate infers that pre-cardiac myoblasts in stage 6 anterior mesoderm are previously specified to respond to the terminal cardiogenic effects of endoderm.

Actins↗
PubMed1993

Scanning electron microscopy of developing primary endoderm during first 6 hours of incubation in the chick.

Development of primary endoderm in the domestic fowl (Gallus domesticus) is described in scanning electron microscopy (SEM) supplemented by transmission electron microscopy (TEM). Although complicated by great variability, the ventral surface of the blastoderm reveals this process during the first 6 hours of incubation. Primary endoderm arises (1) from the hypoblast, (2) from the margin of the area pellucida, and (3) from intervening portions of the area pellucida. The early hypoblast becomes several cells thick while individual cells are still spherical. TEM reveals a variety of immature cell junctions. During subsequent flattening of these cells into primary endodermal epithelium, numerous filopodia arise from their surfaces. These are 0.20-0.25 microns in diameter. They become long and branched, attaching to each other and to other cell bodies. Similar filopodial processes are present less conspicuously among cells in the margin of the area pellucida. Here, there is pseudopodial evidence that cells or cell sheets creep along the ventral surface of the epiblast. The filopodia disappear as cell flattening proceeds. The ventral surface of the exposed epiblast delaminates cells that become free after their exploratory filopodia and lamellipodia are put forth. Lateral contacts among cell bodies from the above three sources increase until a continuous epithelium is formed. The primary endoderm of the embryo, a simple squamous epithelium that separates the connective tissue space above from the gastrocoele below, is generated by these developmental events.

Animals↗
PubMed1995

Glycoconjugate expression in the chick embryonic chorioallantoic membrane: comparisons of the chorionic ectoderm and allantoic endoderm.

BACKGROUND: The chorioallantoic membrane (CAM) of the chick embryo expands during embryogenesis to meet the increased oxygen demands during growth and differentiation. Temporal and spatial glycosylation patterns of CAM ectodermal and endodermal proteins likely contribute to differentiation of the functional attributes of the CAM. METHODS: Using lectins for light and electron microscopic observations, we studied the patterns of glycoconjugate expression on the ectoderm and endoderm of the chorioallantoic membrane (CAM) of the chick at days 4.5, 5.0, 5.5, 6.0, and 10 of morphogenesis. For light microscopy, samples of unfixed CAM were incubated with the following FITC lectins: Con A, DBA, GSA-I, GSA-II, PNA, SBA, UEA-I, and WGA. RESULTS: All lectins, except GSA-I and -II, gave positive results. The positive lectins, labeled with HRP, served to ultrastructurally localize PNA, SBA, and WGA, but not DBA binding to the luminal surface of the endoderm. UEA-I and Con A bound similarly except on day 10 when UEA-I no longer bound. On the ectodermal surface, only WGA bound at all times studied. PNA and SBA binding were present from days 5.0 to 6.0 but absent at days 4.5 and 10. DBA binding occurred through day 5.0 but was absent thereafter. UEA-I bound to the ectoderm at days 4.5, 5.0, and 10 but not days 5.5 and 6.0. Con A bound only on days 5.0 and 10. CONCLUSION: That the ultrastructurally similar ectoderm and endoderm of the CAM display functional differences conforms to the hypothesis that differential expression of glycoconjugate microdomains likely contributes to such functional specialization.

Allantois↗
PubMed2003

Molecular components of the endoderm specification pathway in Xenopus tropicalis.

Xenopus laevis has been instrumental in elucidating a conserved molecular pathway that regulates vertebrate endoderm specification. However, loss-of-function analysis is required to resolve the precise function of the genes involved. For such analysis, antisense oligos and possibly forward genetics are likely to be more effective in the diploid species Xenopus tropicalis than in the pseudotetraploid Xenopus laevis. Here we have isolated most of the tropicalis genes in the endoderm specification pathway, specifically, tVegT, tMixer, tMix, tBix, tGata6, tSox17alpha, tSox17beta, tFoxA1, tHex, and tCerberus, which lack the redundant copies that are found in laevis. In situ hybridization analysis has revealed identical expression patterns between the orthologous tropicalis and laevis endoderm genes, thus suggesting conserved genetic functions. Furthermore, we noted that the smaller tropicalis embryos gave better probe penetration than in laevis whole-mount in situ hybridizations-allowing us to visualize transcripts in the deep endoderm in tropicalis, which is difficult in laevis. This study illustrates how an entire genetic pathway can be quickly transferred from laevis to tropicalis due to high sequence conservation between the sister species and the large number of tropicalis-expressed sequence tags that are now available.

Animals↗
PubMed2006

An amylase/Cre transgene marks the whole endoderm but the primordia of liver and ventral pancreas.

Mice bearing a Cre-encoding transgene driven by a compound [SV40 small t antigen/mousealpha-amylase-2] promoter expressed the recombinase at early developmental stages broadly in the embryonic endoderm before the pancreas and lungs begin to outgrow, but not in other germ layers, as determined indirectly by beta-galactosidase and YFP reporter activity, indicating that the transgene is in fact an endodermic marker. Interestingly, the liver and ventral pancreas were excluded from this expression pattern, denoting that the chimerical alpha-amylase-2 promoter was not active in the anterior leading edge of the endoderm (the presumptive region from which liver and ventral pancreas form). These transgenics thus confirm, among other findings, that dorsal and ventral pancreatic primordia have different intrinsic transcriptional capabilities. In conclusion, we have generated a new transgenic mouse that should be useful to target endoderm at early stages, without affecting the liver or ventral pancreas before embryonic day E12.5.

Animals↗
PubMed1988

Differentiation-dependent production of a platelet-derived growth factor-like mitoattractant by endoderm cells derived from embryonal carcinoma cells.

Retinoic acid-induced differentiation of F9 embryonal carcinoma cells to endoderm provokes the secretion of a protein factor that acts as both a chemoattractant and mitogen for smooth muscle cells. Undifferentiated F9 cells and PSA-5E (visceral endodermlike) cells produced little of this factor. However, PYS-2 (parietal endodermlike) and Dif 5 endoderm cells were found to produce significant amounts of endoderm-derived mitoattractant (EDM) activity. The activity secreted by the Dif 5 cells was partially purified using gel filtration chromatography using chemotaxis and mitogenic assays as markers for biological activity. The partially purified activity competes with [125I]iodo-platelet-derived growth factor (PDGF) for binding to target cells, and the biological activity is neutralized with anti-PDGF IgG, suggesting shared domains in the two molecules. However, the factor appears to be different from PDGF, based on its thermal stability, molecular weight, and charge. The differentiated endoderm cells including retinoic acid (RA)-treated F9, Dif 5, PSA-5E, and PYS-2 cells also exhibit specific [125I]iodo-PDGF binding, and the PSA-5E cells respond to PDGF as a chemoattractant. Conceivably, such a PDGF-like factor may contribute to the regulation of cell growth and migration during the early stages of embryogenesis.

Animals↗
PubMed1988

Effects of teratogenic antibodies on cultured visceral yolk-sac endodermal cells: cytotoxicity and morphological studies.

Primary cultures of visceral yolk-sac (VYS) endodermal cells were used to assess the effects of teratogenic and nonteratogenic antibodies. When assessed by cytotoxicity assay, teratogenic antibodies appeared to be lethal to the cultured cells at high concentrations (1.25-5 mg of antibodies per ml of culture medium). At a nonlethal dosage, the teratogenic antibodies induced morphological changes, including retraction and rounding up of living cells. The cytotoxic effect as well as the effect on cell morphology appeared to be dose-dependent and specific to VYS endodermal cells. The mechanisms of cell killing were not the same as those attributed to complement-mediated cell lysis. The nonteratogenic antibodies did not have any cytotoxic effect nor did they cause any cell morphological alterations. The results of this investigation, when interpreted by correlating the dose-dependent effects of the teratogenic antibodies on cultured endodermal cells with the in vivo teratogenic effect, suggest that teratogenic antibodies when given at a teratogenic dose cause congenital abnormalities without killing the VYS endodermal cells.

Animals↗
PubMed1993

Homeobox-containing genes in teratocarcinoma embryoid bodies: a possible role for Hox-D12 (Hox-4.7) in establishing the extraembryonic endoderm lineage in the mouse.

We wish to identify genes involved in mediating early lineage decisions in the mouse embryo. F9 teratocarcinoma cells treated with retinoic acid (RA) in suspension culture develop into embryoid bodies (EBs) with an outer layer of visceral endoderm. In order to identify genes that are involved in establishing this extraembryonic endoderm lineage we have employed a PCR-based approach using cDNAs from early EBs as templates. PCR reactions were performed with degenerate oligonucleotide primers coding for the highly conserved regions of the homeodomains of the Drosophila Antennapedia, bicoid, and zerknüllt proteins. Among the PCR products were representatives of previously identified mouse genes, including Hox-A5 (1.3), A1 (1.6), A9 (1.7), B8 (2.4), B2 (2.8), C8 (3.1), and D12 (4.7). Whole mount in situ hybridization analysis, performed to examine the temporal and spatial distribution of transcripts, suggests a possible role for the Hox-D12 gene during endoderm differentiation in F9 EBs. Whereas the expression patterns of several other homeobox genes are essentially uniform throughout the aggregates, Hox-D12 expression is restricted to the outer surface of early EBs at a time when lineage decisions may be occurring. In order to establish the relationship between the Hox-D12 expression pattern and the role of RA in inducing F9 EB differentiation, we examined PSA-1 EBs that differentiate in the absence of added RA. PSA-1 EBs show similar temporal and spatial localization of Hox-D12 when compared to F9 EBs. These data suggest that the pattern of Hox-D12 expression correlates with endoderm differentiation and not with RA treatment and point to a possible role for homeobox-containing genes during the early stages of mouse embryogenesis.

Amino Acid Sequence↗
PubMed2000

Anterior endoderm is sufficient to rescue foregut apoptosis and heart tube morphogenesis in an embryo lacking retinoic acid.

The vitamin A deficient (VAD) quail embryo lacks active retinoids, fails to express normally GATA-4, and develops a nonlooping heart tube morphogenetic defect that is a model for congenital cardiomyopathy. VAD quail embryos, or chick embryos depleted specifically for GATA factors, show in addition abnormal foregut development, characterized by apoptosis of the endoderm cells associated with presumptive myocardium during the process of heart tube formation. Exogenous retinoic acid or transplantation of normal chick embryo anterior endoderm is sufficient to rescue apoptosis as well as GATA-4 expression and results in normal development and heart tube morphogenesis. Normal posterior endoderm also contains retinoids but is unable to rescue the VAD defect. Our results indicate that a retinoid-dependent transcriptional program, mediated at least in part by GATA factors, is critical in presumptive foregut endoderm for normal heart tube morphogenesis.

Animals↗
PubMed1993

Transient gene expression by SV40 promoter characterizes sequential differentiation of embryonal carcinoma F9 cells into primitive and visceral endoderm.

Transient expression of the chloramphenicol acetyltransferase (CAT) gene under the control of simian virus 40 (SV40)y Moloney murine leukemia virus, human T cell leukemia virus, and cytomegalovirus promoters was stimulated by the differentiation of F9 stem cells into primitive endoderm, but repressed again by further differentiation into visceral endoderm. Deletion mutants of the SV40 enhancer showed that a similar set of motifs is critical for CAT expression at all stages of F9 differentiation, but differentiation dependency was observed even in their absence. The stability of transient gene expression under the control of the SV40 promoter was markedly dependent on F9 differentiation. Appreciable expression was detected even in undifferentiated F9 cells immediately after gene transfection, was maximal at 12 h and declined rapidly thereafter. On the other hand, expression in primitive endoderm increased until 72 h. The decline was accelerated again in visceral endoderm. This shift was somewhat specific to the virus promoter since CAT expression in undifferentiated F9 cells under the control of the elongation factor 1 alpha promoter was more stable than for virus promoters tested. Thus, the change in stability of expression is important for differentiation-dependent virus promoter activity.

Animals↗
PubMed1996

Endoderm-specific gene expression in embryonic stem cells differentiated to embryoid bodies.

Mouse embryonic stem cells can differentiate into various cell types within cell aggregates called embryoid bodies (EBs). This structure consists of ectodermal, mesodermal, and endodermal tissues, which resemble the embryo of egg-cylinder stage. After 8-10 days in culture, about half of the EBs expand into large cystic structures homologous to visceral yolk sac of postimplantation embryos. To study endoderm differentiation at molecular level, we examined expression of endoderm marker genes during the processes of EB development. alpha-Fetoprotein (AFP) and transthyretin (TTR) transcripts increased at the stage when embryoid bodies began to form yolk-sac-like structures and were expressed strongly thereafter. Expression of hepatocyte nuclear factor (HNF) 4, a variant form of HNF1 (also called HNF1beta), and HNF3beta started before the onset of AFP and TTR expression. HNF1 (also called HNF1alpha) expression began a few days after the onset of the expression of the transcription factors described above. Serum albumin (ALB) transcript was only found in late large cystic EBs. Also, AFP gene expression preceded ALB gene expression. These results suggest that the patterns of endoderm gene expression during EB development reflect the order found during mouse development in vivo, and EB formation may serve as an in vitro system to study the differentiation process.

Animals↗
PubMed1998

Spontaneous calcium oscillations in embryonic stem cell-derived primitive endodermal cells.

In vitro differentiation of mouse embryonic stem cells within three-dimensional cell aggregates called embryoid bodies parallels the development of postimplantation embryos at the egg cylinder stage, where visceral and parietal endoderm diverge from the primitive endoderm. We have investigated spontaneous [Ca2+]i oscillations by means of confocal laser-scanning microscopy in primitive endodermal cell layers of embryoid bodies during their differentiation to parietal and visceral endoderm. The frequency of [Ca2+]i oscillations increased from day 4 to day 19 of development, whereas their duration decreased from day 3 to days 16-17. Oscillations depended on both extracellular Ca2+ and Ca2+ release from intracellular stores as they were abolished in Ca(2+)-free solution and in the prescence of Ni2+ and thapsigargin. Signal transduction operated via the phospholipase C (PLC)-mediated inositol 1,4,5-triphosphate (InsP3) pathway with a negative feedback loop via protein kinase C (PKC) as U73,122, a blocker of PLC; bisindolylmaleimide 1, staurosporine, and H-7, blockers of PKC; and 10 mM caffeine totally inhibited [Ca2+]i spiking. Thimerosal, which hypersensitizes the InsP3 receptor, as well as vasopressin and bradykinin, which act via the InsP3 pathway, increased the frequency of [Ca2+]i spikes. In the prescence of brefeldin A (50 microM) or monensin (20 microM), which both inhibit endo/exocytotic vesicle pathways, an immediate transient increase in spiking activity was followed by a decline within 1 to 2 h. In the presence of brefeldin A or thapsigargin or in the absence of extracellular Ca2+, endocytotic vesicles were absent, suggesting that oscillating [Ca2+]i transients are involved in the exo/endocytotic vesicle shuttle.

Animals↗