Differentiation of blood islands in chick blastoderms in association with Morris hepatoma grafts and its relationship to specific chromosomal changes in the hepatomas.
Explore the source record for details and available documents.
Animals↗
SEARCH · Search PubMed
Results for “germ layer specification”
Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.
Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.
A dual fluorescence technique for visualization of Staphylococcus epidermidis biofilm using scanning confocal laser microscopy.
A new dual fluorescence technique is described which, when combined with scanning confocal laser microscopy (SCLM), can be used to visualize the components of biofilm produced by Staphylococcus epidermidis. Chemostat cultures of RP62A (a well-characterized slime-producing strain of S. epidermidis) were used to produce mature biofilm on polyvinylchloride (PVC) disks immobilized in a modified Robbins device using a 'seed' and 'feed' model system. Serial horizontal and vertical optical thin sections, as well as three-dimensional computer reconstructions, were obtained on in situ biofilm using the dual fluorescence procedure. Bacteria were visualized by green autofluorescence excited at 488 nm with an Argon laser. Cell-associated and exocellular matrix material (slime) was visualized by red fluorescence excited at 568 nm with a Krypton laser after interaction of the biofilm with Texas Red-labeled wheat germ agglutinin which is a slime-specific lectin marker. Structural analysis revealed that the cocci grew in slime-embedded cell clusters forming distinct conical-shaped microcolonies. Interspersed open channels served to connect the bulk liquid with the deepest layers of the mature, hydrated biofilm which increased overall surface area and likely facilitated the exchange of nutrients and waste products throughout the biofilm. The combined dual fluorescence technique and SCLM is potentially useful as a specific noninvasive tool for studying the effect of antimicrobial agents on the process of biofilm formation and for the characterization of the architecture of S. epidermidis biofilm formed in in vivo and in vitro on medical grade virgin or modified inert polymer surfaces.
Biofilms↗
Human embryonic stem cells derived without feeder cells.
BACKGROUND: Human embryonic stem cells are likely to play an important role in the future of regenerative medicine. However, exposure of existing human embryonic stem-cell lines to live animal cells and serum risks contamination with pathogens that could lead to human health risks. We aimed to derive an embryonic stem-cell line without exposure to cells or serum. METHODS: Frozen cleavage-stage embryos were thawed and cultured to the blastocyst stage. Inner cell masses were isolated by immunosurgery and plated onto extracellular-matrix-coated plates that can be easily sterilised. Six established human embryonic stem-cell lines were also maintained with this serum and feeder free culture system. FINDINGS: A new stem-cell line was derived from human embryos under completely cell and serum free conditions. The cells maintained normal karyotype and markers of pluripotency, including octamer binding protein 4 (Oct-4), stage-specific embryonic antigen (SSEA)-3, SSEA-4, tumour-rejection antigen (TRA)-1-60, TRA-1-81, and alkaline phosphatase. After more than 6 months of undifferentiated proliferation, these cells retained the potential to form derivatives of all three embryonic germ layers both in vitro and in teratomas. These properties were also successfully maintained (for more than 30 passages) with the established stem-cell lines. INTERPRETATION: This system eliminates exposure of human embryonic stem cells and their progeny to animal and human feeder layers, and thus the risk of contamination with pathogenic agents capable of transmitting diseases to patients.
Animals↗
Determination of blastoderm cells in Drosophila melanogaster.
A method for culturing blastoderm cells of Drosophila in vivo has been developed that allows these cells to differentiate into larval or adult structures. By intermixture of genetically marked cells from bisected and whole embryos, it was shown that blastoderm cells are restricted in their potential for forming adult epidermal structures. Cells isolated from anterior-half embryos are determined for forming head and thoracic structures, whereas cells from posterior-half embryos are determined for forming thoracic and abdominal structures. The specificity of determination and the localization of determinative factors is discussed.
Animals↗
Identification of a retinoic acid-sensitive period during primary axis formation in Xenopus laevis.
Retinoic acid (RA) is able to profoundly alter patterning of the primary body axis in embryos of the frog Xenopus laevis. The response to RA is dose-dependent, and leads to progressive truncation of the anteroposterior axis, with anterior structures most sensitive. Both mesodermal and ectodermal tissues are affected, and in vitro assays demonstrate that induced dorsal ectoderm is one direct target of RA. RA represses expression of anterior-specific genes and concomitantly induces expression of at least one posterior-specific gene. Resistance to RA is acquired gradually, during gastrula and early neurula stages, with posterior structures becoming resistant before anterior structures. These data demarcate in the embryo an anterior "domain," which may define the head rudiment and which transcends germ layers. RA can alter the axial pattern after its initial induction; thus, RA sensitivity defines a labile intermediate that occurs during axial patterning. These data suggest a possible role for RA in normal axis formation.
Animals↗
Genomic organization, splice variants and expression of CGM1, a CD66-related member of the carcinoembryonic antigen gene family.
The tumor marker carcinoembryonic antigen (CEA) belongs to a family of proteins which are composed of one immunoglobulin variable domain and a varying number of immunoglobulin constant-like domains. Most of the membrane-bound members, which are anchored either by a glycosylphosphatidylinositol moiety or a transmembrane domain, have been shown to convey cell adhesion in vitro. Here we describe two splice variants of CGM1, a transmembrane member of the CEA family without immunoglobulin constant-like domains. CGM1a and CGM1c contain cytoplasmic domains of 71 and 31 amino acids, respectively. The cytoplasmic region of CGM1a is encoded by four exons (Cyt1-Cyt4). Differential splicing of the Cyt1 exon (53 bp) leads to the formation of CGM1c. The presence or absence of potential protein kinase phosphorylation sites in the cytoplasmic domains and a sequence consensus motif involved in signal transduction in multichain immune recognition receptors indicates that this splice event is of functional importance. CGM1a mRNA, the predominant CGM1 transcript, was found in the granulocytic lineage, but not in monocytes, lymphocytes nor in a number of tumors derived from all three germ layers. Weak staining using monoclonal antibodies Tu2 and 73 in fluorescence-activated cell scan analyses indicate low concentrations of CGM1 protein on the surface of granulocytes. The CGM1 protein is also recognized by CD66 antibodies. Therefore, the granulocyte-specific CD66 epitope is present on at least four CEA family members: CGM1, CEA, NCA-50/90 and NCA-160.
Amino Acid Sequence↗
Autonomous and nonautonomous Notch functions for embryonic muscle and epidermis development in Drosophila.
The Notch (N) gene encodes a cell signaling protein that mediates neuronal and epidermal determination in Drosophila embryos. N also regulates several aspects of myogenic development; embryos lacking N function have too many muscle founder cells and fail to properly differentiate somatic muscle. To identify cell-autonomous requirements for Notch function during muscle development, we expressed a Notch minigene in the mesoderm, but not in the ectoderm, of amorphic N-embryos. In these embryos, muscle founder hypertrophy is rescued, indicating that Notch is autonomously required by mesoderm cells to regulate the proper number of muscle founders. However, somatic muscle differentiation is only partially normalized, suggesting that Notch is also required in the ectoderm for proper muscle development. Additionally, mesodermal expression of Notch partially rescues epidermal development in overlying neurogenic ectoderm. This is unexpected, since previous studies suggest that Notch is autonomously required by proneural ectoderm cells for epidermal development. Mesodermal expression of a truncated Notch protein lacking the extracellular domain does not rescue ventral epidermis, suggesting that the extra-cellular domain of Notch can non-autonomously rescue epidermal development across germ layers.
Animals↗
Defects in specific associations between astroglia and neurons occur in microcultures of weaver mouse cerebellar cells.
The neurological mutation weaver is characterized by defects in granule cell migration along Bergmann glial processes and by subsequent death and disposition of granule cells. Immunocytochemical localization of antisera raised against purified glial filament protein (AbGF) and transmission electron microscopy were used to visualize specific associations between granule neurons and astroglia in microcultures of cerebellar cells dissociated from normal (+/+), heterozygous (+/wv), and homozygous (wv/wv) B6CBA-w mouse cerebella. In microcultures of cells dissociated from normal B6CBA-Aw-J-wv (+/+) cerebella, staining with AbGF closely resembled results previously reported for cells taken from C57BL/6J (+/+) tissue. Two forms of stained astroglia were seen, one with a larger perikaryon and shorter processes, among which 12 to 20 unstained cells nestled; and another with a smaller cell soma and longer processes, along which a few unstained cells were seen. The first resembled astrocytes of the internal granular layer and white matter, the second Bergmann astroglia. In microcultures of heterozygous animals (+/wv), the number of granule cells was reduced slightly. Many stained astroglia resembled those from +/+ cerebella, but others had thickened processes and enlarged terminal "endfeet." Granule cells associated with both forms of stained astroglia. Cultures from homozygous weaver (wv/wv) cerebella contained very few, if any, granule cells and did not exhibit specific neuronal/glial interactions characteristic of +/+ cells. Both forms of stained wv/wv glial cells had enlarged cell somata giving rise to stunted processes, suggesting that both Bergmann glia and astrocytes are affected by the weaver gene. By both immunocytochemical staining and electron microscopy, accumulation and tangling of glial filaments were seen. Immunocytochemical staining of weaver cerebellar tissue with AbGF also revealed abnormalities in astrocytes of the internal granular layer and white matter in addition to previously reported defects in Bergmann glia. Agglutination with five plant lectins, each with a different carbohydrate-binding specificity, revealed that postnatal weaver cerebellar cells agglutinate with the lectins concanavalin A and the wheat germ agglutinin, suggesting the persistence postnatally of embryonic cell surface elements on postnatal weaver cells.
Animals↗
Localization of alpha-galactomannan and of wheat germ agglutinin receptors in Schizosaccharomyces pombe.
The location of galactomannan on the surface of Schizosaccharomyces pombe cells was reexamined by scanning electron microscopy by an indirect but specific method using gold markers. The polysaccharide was found on the cell surface and at the end beginning to grow but not on the wall established by division. Galactomannan was also localized on S. pombe thin sections by transmission electron microscopy using the same method. The polysaccharide was found deposited in two layers in the cell wall, i.e. at the periphery of the wall and near the plasmalemma. The septum was also marked but mainly near the plasmalemma. These results indicated that the polysaccharide is elaborated onto the outside of the wall during extension but not during septum formation. When thin sections of S. pombe were marked with gold granules labeled with wheat germ agglutinin, marking was found in vacuoles but not in the cell wall. This confirmed that S. pombe cell wall is devoid of chitin.
Ascomycota↗
Adult stem cells--perspectives in treatment of metabolic diseases.
There is currently great excitement and expectation concerning the differential potential of adult stem cells or adult cells with capacity of differentiation. As the body of work concerning transdifferentiation of somatic stem cells and bone marrow derived stem cells grows, the number of critics increases steadily questioning the reliability of reported findings. So scientists are now challenged more and more to prove that resulting differentiated somatic cells originated from somatic adult stem cell through a transdifferentiation process. Phenomenons such as fusion of cells have to be ruled out and the origin of the differentiated cell has to be determined by specific techniques i.e. in situ hybridisation. Cellular mimicry through uptake of specific factors out of the medium is questioned to be the reason for cells staining positive for Insulin. Some multipotent adult stem cells can cross lineage boundaries and differentiate into somatic cells of other lineages after being relocated. Bone marrow cells have been described to have the greatest plasticity among adult stem cells regenerating damaged liver or myocardium. It has been proposed that the differentiation of bone marrow derived adult stem cells occurs naturally even in healthy organs as a physiologic process of tissue-regeneration. Others believe that organ damage is essential to induce transdifferentiation by release of organ specific microenvironmental factors. We here try to constitute necessary data which should be demonstrated to give substantial evidence for transdifferentiation of newly characterized cells including exclusion of fusion, phagocytosis or DNA uptake, description of the outset cell, differentiation into all three germ layers and functional parameters.
Adult↗
Distribution of toxigenic Fusarium spp. and mycotoxin production in milling fractions of durum wheat.
A reliable and sensitive PCR assay to specifically detect trichothecene-producing Fusarium spp. in milling fractions and kernel tissue of naturally infected durum wheat is reported. Assays were based on a combination of primers derived from the trichodiene synthase and the beta-tubulin genes. The occurrence of toxigenic Fusarium spp. in semolina and wheat tissue (grain ends, crease, pericarp, aleurone layer, germ and albumen) was detected, even for a weakly contaminated wheat sample. Penetration of toxigenic Fusarium spp. into the interior of durum wheat kernel was demonstrated for the Nefer variety, indicating that none of the tissue structures within the wheat kernel acted as an effective barrier to fungal invasion. Moreover, after inoculation by toxigenic Fusarium strains, semolina was shown to allow high yields of trichothecenes, while bran was demonstrated to contain biochemical inhibitors able to significantly reduce trichothecene production. These results will be useful in improving breeding strategies to control trichothecene contamination of durum wheat kernels.
DNA, Fungal↗
Zebrafish organizer development and germ-layer formation require nodal-related signals.
The vertebrate body plan is established during gastrulation, when cells move inwards to form the mesodermal and endodermal germ layers. Signals from a region of dorsal mesoderm, which is termed the organizer, pattern the body axis by specifying the fates of neighbouring cells. The organizer is itself induced by earlier signals. Although members of the transforming growth factor-beta (TGF-beta) and Wnt families have been implicated in the formation of the organizer, no endogenous signalling molecule is known to be required for this process. Here we report that the zebrafish squint (sqt) and cyclops (cyc) genes have essential, although partly redundant, functions in organizer development and also in the formation of mesoderm and endoderm. We show that the sqt gene encodes a member of the TGF-beta superfamily that is related to mouse nodal. cyc encodes another nodal-related proteins, which is consistent with our genetic evidence that sqt and cyc have overlapping functions. The sqt gene is expressed in a dorsal region of the blastula that includes the extraembryonic yolk syncytial layer (YSL). The YSL has been implicated as a source of signals that induce organizer development and mesendoderm formation. Misexpression of sqt RNA within the embryo or specifically in the YSL induces expanded or ectopic dorsal mesoderm. These results establish an essential role for nodal-related signals in organizer development and mesendoderm formation.
Amino Acid Sequence↗
Secreted modular calcium-binding protein-1 localization during mouse embryogenesis.
BM-40 is an extracellular matrix-associated protein and is characterized by an extracellular calcium-binding domain as well as a follistatin-like domain. Secreted modular calcium-binding protein-1 (SMOC-1) is a new member of the BM-40 family. It consists of two thyroglobulin-like domains, a follistatin-like domain and a new domain without known homologues and is expressed ubiquitously in many adult murine tissues. Immunofluorescence studies, as well as immunogold electron microscopy, have confirmed the localization of SMOC-1 in or around basement membranes of adult murine skin, blood vessels, brain, kidney, skeletal muscle, and the zona pellucida surrounding the oocyte. In the present work, light microscopic immunohistochemistry has revealed that SMOC-1 is localized in the early mouse embryo day 7 throughout the entire endodermal basement membrane zone of the embryo proper. SMOC-1 mRNA is synthesized, even in early stages of mouse development, by mesenchymal as well as epithelial cells deriving from all three germ layers. In embryonic stage day 12, and fetal stages day 14, 16, and 18, the protein is present in the basement membrane zones of brain, blood vessels, skin, skeletal muscle, lung, heart, liver, pancreas, intestine, and kidney. This broad and organ-specific distribution suggests multifunctional roles of SMOC-1 during mouse embryogenesis.
Animals↗
Cardiovascular therapeutic aspects of cell therapy and stem cells.
The recent advancements in stem cell biology, molecular and cell biology, and tissue engineering have paved the way to the development of a new biomedical discipline: regenerative medicine. The heart represents an attractive candidate for this emerging discipline since these emerging technologies could be used to potentially treat a variety of myocardial disorders. Here we describe our efforts in using stem cell and cell therapy strategies to restore the myocardial electromechanical properties. Specifically, our research has focused on the potential role of human embryonic stem cells (hESC) for myocardial regeneration (for the treatment of heart failure) and on using genetically engineered cell grafts to modify the myocardial electrophysiological properties (for the treatment of cardiac arrhythmias). The recently described hESC lines are unique pluripotent cell lines that can be propagated in the undifferentiated state in culture and coaxed to differentiate into cell derivatives of all three germ layers, including cardiomyocytes. The current article describes this unique cardiomyocyte differentiating system and details the molecular, ultrastructural, and functional properties of the generated hESC-derived cardiomyocytes (hESC-CMs). The ability of the hESC-CMs to integrate structurally and functionally with host cardiomyocytes in both in vitro and in vivo studies will be described as well as their ability to restore the myocardial electromechanical function in animal models of diseased hearts. We will next present detailed in vitro, in vivo, and computer simulation studies performed in our laboratory testing the hypothesis that cell grafts, engineered to express specific ion channels, can be used to modify the myocardial electrophysiological properties of cardiac tissue. The potential and drawbacks of this novel approach for the treatment of both tachyarrhythmias (using cell grafts expressing potassium channels) and bradyarrhythmias (using hESC coaxed to differentiate into pacemaking cells or conducting tissue) will be described.
Animals↗
Xenopus endo B is a keratin preferentially expressed in the embryonic notochord.
Screening of a cDNA library from neurula stage Xenopus laevis for notochord-specific sequences led to the isolation of a cDNA clone, XK endo B (Xenopus keratin endo B), which encodes a nonepidermal type I keratin. In situ and Northern blot hybridizations indicate that expression of XK endo B RNA is concentrated in the notochord, whereas expression in the endoderm is 5-10 times lower. XK endo B mRNA is present in the oocyte and increases from late gastrula. Accumulation peaks by late neurula and is greatly reduced by the tadpole stage; in the adult, a low level of XK endo B RNA is present in the liver. XK endo B shows sequence homology to mouse endo B; genomic Southern blots show that XK endo B is the most similar sequence to mouse endo B in the Xenopus genome, and vice versa, indicating that XK endo B and mouse endo B are homologs. The use of endo B as a marker and the germ layer derivation of the notochord are discussed in light of these results.
Amino Acid Sequence↗
Feeder-free growth of undifferentiated human embryonic stem cells.
Previous studies have shown that maintenance of undifferentiated human embryonic stem (hES) cells requires culture on mouse embryonic fibroblast (MEF) feeders. Here we demonstrate a successful feeder-free hES culture system in which undifferentiated cells can be maintained for at least 130 population doublings. In this system, hES cells are cultured on Matrigel or laminin in medium conditioned by MEF. The hES cells maintained on feeders or off feeders express integrin alpha6 and beta1, which may form a laminin-specific receptor. The hES cell populations in feeder-free conditions maintained a normal karyotype, stable proliferation rate, and high telomerase activity. Similar to cells cultured on feeders, hES cells maintained under feeder-free conditions expressed OCT-4, hTERT, alkaline phosphatase, and surface markers including SSEA-4, Tra 1-60, and Tra 1-81. In addition, hES cells maintained without direct feeder contact formed teratomas in SCID/beige mice and differentiated in vitro into cells from all three germ layers. Thus, the cells retain fundamental characteristics of hES cells in this culture system and are suitable for scaleup production.
Cell Culture Techniques↗
Limited left-right cell migration across the midline of the gastrulating avian embryo.
During avian development the earliest phase in which the avian embryo expresses axial features of a left-right axis is at the primitive streak stage. Until the stage of definitive primitive streak (streak 4 H&H), the axis seems to possess morphological bilateral symmetry. Morphological asymmetry begins only during the next few hours of incubation, with development of overt morphological and molecular asymmetry within Hensen's node (stage 5 H&H). In this report, we present an experimental study aimed at following the pattern of cell movements during primitive streak formation and gastrulation of specific left-right regions from earlier stages of the avian embryo. To determine the origin of cells contributing to each side of the primitive streak, we applied the dye Lysinated-Rodamine-Dextran (LRD) to one half, either left or right, of the pre-streak blastoderm (stages X-XIII, EG&K). We tried to estimate the relative cell contribution to primitive streak formation, and to the three germ layers evolving during gastrulation in the context of the left-right axis. Moreover, we asked whether the midline serves as a border, that is, as a physiological barrier preventing cell passing during gastrulation. Our results demonstrate that on each side of the axis, either the right or the left, most of the cells originate from the same half of a pre-streak blastoderm, populate the same half of the PS and contribute to tissues largely confined to that particular side. However, along the primitive streak, a few cells were detected on the opposite side of the midline. Moreover, variation in the number of cells crossing the midline at specific regions along the primitive streak was found. Most crossing cells were located near the mid rostrocaudal extent of the primitive streak, from 25-85% of its length. At the posterior end of the primitive streak, fewer crossing cells were detected. At the anterior region of the PS, that is, within Hensen's node, cells do not cross the midline. These results suggest that differences occur in the process of ingression along the rostrocaudal extent of the PS.
Animals↗
Molecular indices of cell lineage specification in sea urchin embryos.
The origins of several of the differentiated cell lineages of the advanced sea urchin embryo are well defined. Cytological application of molecular probes to three lineages, those responsible for the formation of the skeleton, the gut, and the aboral ectodermal wall of the late embryo, has demonstrated expression of lineage-specific genes long before overt morphological differentiation. These observations lead to useful generalizations regarding the processes of gene regulation that underlie the molecular biology of cell lineage specification in the embryo.
Actins↗