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At least 613 records · Page 34Linked to original sources
PubMed2004

Adult-derived liver stem cells acquire a cardiomyocyte structural and functional phenotype ex vivo.

We examined the differentiation potential of an adult liver stem cell line (WB F344) in a cardiac microenvironment, ex vivo. WB F344 cells were established from a single cloned nonparenchymal epithelial cell isolated from a normal male adult rat liver. Genetically modified, WB F344 cells that express beta-galactosidase and green fluorescent protein or only beta-galactosidase were co-cultured with dissociated rat or mouse neonatal cardiac cells. After 4 to 14 days, WB F344-derived cardiomyocytes expressed cardiac-specific proteins and exhibited myofibrils, sarcomeres, and a nascent sarcoplasmic reticulum. Further, rhythmically beating WB F344-derived cardiomyocytes displayed calcium transients. Fluorescent recovery after photobleaching demonstrated that WB F344-derived cardiomyocytes were coupled with adjacent neonatal cardiomyocytes and other WB F344-derived cardiomyocytes. Fluorescence in situ hybridization experiments suggested that fusion between WB F344 cells and neonatal mouse cardiomyocytes did not take place. Collectively, these results support the conclusion that these adult-derived liver stem cells respond to signals generated in a cardiac microenvironment ex vivo acquiring a cardiomyocyte phenotype and function. The identification ex vivo of microenvironmental signals that appear to cross germ layer and species specificities should prove valuable in understanding the molecular basis of adult stem cell differentiation and phenotypic plasticity.

Animals↗
PubMed2002

Regulation of Xenopus embryonic cell adhesion by the small GTPase, rac.

TGF-beta family signalling pathways are important for germ layer formation and gastrulation in vertebrate embryos and have been studied extensively using embryos of Xenopus laevis. Activin causes changes in cell movements and cell adhesion in Xenopus animal caps and dispersed animal cap cells. Rho family GTPases, including rac, mediate growth factor-induced changes in the actin cytoskeleton, and consequently, in cell adhesion and motility, in a number of different cell types. Ectopic expression of mutant rac isoforms in Xenopus embryos was combined with animal cap adhesion assays and a biochemical assay for rac activity to investigate the role of rac in activin-induced changes in cell adhesion. The results indicate that (1) the perturbation of rac signalling disrupts embryonic cell-cell adhesion, (2) that rac activity is required for activin-induced changes in cell adhesive behavior on fibronectin, and (3) that activin increases endogenous rac activity in animal cap explants.

Activins↗
PubMed2003

Differentiation of rhesus embryonic stem cells to neural progenitors and neurons.

Embryonic stem (ES) cells are pluripotent cells capable of differentiating into cell lineages derived from all primary germ layers including neural cells. In this study we describe an efficient method for differentiating rhesus monkey ES cells to neural lineages and the subsequent isolation of an enriched population of Nestin and Musashi positive neural progenitor (NP) cells. Upon differentiation, these cells exhibit electrophysiological characteristics resembling cultured primary neurons. Embryoid bodies (EBs) were formed in ES growth medium supplemented with 50% MEDII. After 7 days in suspension culture, EBs were transferred to adherent culture and either differentiated in serum containing medium or expanded in serum free medium. Immunocytochemistry on differentiating cells derived from EBs revealed large networks of MAP-2 and NF200 positive neurons. DAPI staining showed that the center of the MEDII-treated EBs was filled with rosettes. NPs isolated from adherent EB cultures expanded in serum free medium were passaged and maintained in an undifferentiated state by culture in serum free N2 with 50% MEDII and bFGF. Differentiating neurons derived from NPs fired action potentials in response to depolarizing current injection and expressed functional ionotropic receptors for the neurotransmitters glutamate and gamma-aminobutyric acid (GABA). NPs derived in this way could serve as models for cellular replacement therapy in primate models of neurodegenerative disease, a source of neural cells for toxicity and drug testing, and as a model of the developing primate nervous system.

Action Potentials↗
PubMed1999

Opioid growth factor and organ development in rat and human embryos.

In addition to neurotransmission, the native opioid peptide, [Met5]enkephalin, is a tonically active inhibitory growth molecule that is termed opioid growth factor (OGF). OGF interacts with the zeta (zeta) opioid receptor to influence cell proliferation and tissue organization. We now identify OGF and the zeta receptor in embryonic derivatives including ectoderm, mesoderm, and endoderm of the rat on gestation day 20. Messenger RNA for preproenkephalin (PPE), the precursor of OGF, was detected in the developing cells, suggesting an autocrine production of this peptide. Acute exposure of the pregnant female to OGF resulted in a decrease in DNA synthesis in cells of organs representing all three germ layers, and did so in a receptor-mediated fashion. The influence of OGF was direct, as evidenced in organ culture studies. Blockade of endogenous opioid interaction using naltrexone (NTX) produced an increase in DNA synthesis, indicating the constitutive and functional nature of opioid activity on growth during prenatal life. Human fetal cells contained OGF and the zeta receptor. These data support the hypothesis that endogenous opioid modulation of organ development is a fundamental principle of mammalian embryogenesis, and that OGF has a profound influence on ontogeny. Irregularities in the role of opioids as growth regulators in relationship to the more than 500,000 newborns suffering from birth defects each year in the US needs to be examined.

Animals↗
PubMed2003

Twenty one years of P19 cells: what an embryonal carcinoma cell line taught us about cardiomyocyte differentiation.

Many different stem cell types have been shown to differentiate into cardiac muscle cells in vitro but P19 embryonal carcinoma (EC) cells were one of the first examples described and have been the most extensively characterized. P19 EC cells, isolated from an experimental embryo-derived teratocarcinoma in mice, are multipotent and can differentiate into cell types of all three germ layers. Because of their capacity to form cardiomyocytes however, they have been used to dissect the role of cardiac-specific transcription factors and upstream signalling pathways in cardiac cell differentiation. Furthermore, they have shed light on unique aspects of cardiac cell physiology during heart cell differentiation, including regulation of the proteins underlying the electrical and contractile systems. Here, we review studies on different subclones of P19 cells, and what they have taught us about cardiac differentiation and physiology.

Animals↗
PubMed2003

Regulation of nodal and BMP signaling by tomoregulin-1 (X7365) through novel mechanisms.

During early vertebrate development, members of the transforming growth factor beta (TGFbeta) family play important roles in a variety of processes, including germ layer specification, patterning, cell differentiation, migration, and organogenesis. The activities of TGFbetas need to be tightly controlled to ensure their function at the right time and place. Despite identification of multiple regulators of Bone Morphogenetic Protein (BMP) subfamily ligands, modulators of the activin/nodal class of TGFbeta ligands are limited, and include follistatin, Cerberus, and Lefty. Recently, a membrane protein, tomoregulin-1 (TMEFF1, originally named X7365), was isolated and found to contain two follistatin modules in addition to an Epidermal Growth Factor (EGF) domain, suggesting that TMEFF1 may participate in regulation of TGFbeta function. Here, we show that, unlike follistatin and follistatin-related gene (FLRG), TMEFF1 inhibits nodal but not activin in Xenopus. Interestingly, both the follistatin modules and the EGF motif contribute to nodal inhibition. A soluble protein containing the follistatin and the EGF domains, however, is not sufficient for nodal inhibition; the location of TMEFF1 at the membrane is essential for its function. These results suggest that TMEFF1 inhibits nodal through a novel mechanism. TMEFF1 also blocks mesodermal, but not epidermal induction by BMP2. Unlike nodal inhibition, regulation of BMP activities by TMEFF1 requires the latter's cytoplasmic tail, while deletion of either the follistatin modules or the EGF motif does not interfere with the BMP inhibitory function of TMEFF1. These results imply that TMEFF1 may employ different mechanisms in the regulation of nodal and BMP signals. In Xenopus, TMEFF1 is expressed from midgastrula stages onward and is enriched in neural tissue derivatives. This expression pattern suggests that TMEFF1 may modulate nodal and BMP activities during neural patterning. In summary, our data demonstrate that tomoregulin-1 is a novel regulator of nodal and BMP signaling during early vertebrate embryogenesis.

Activins↗
PubMed2003

Signals from lateral plate mesoderm instruct endoderm toward a pancreatic fate.

During embryonic development, organs arise along the gut tube as a series of buds in a stereotyped anterior-posterior (A-P) pattern. Using chick-quail chimeras and in vitro tissue recombination, we studied the interactions governing the induction and maintenance of endodermal organ identify focusing on the pancreas. Though several permissive signals in pancreatic development have been previously identified, here we provide evidence that lateral plate mesoderm sends instructive signals to the endoderm, signals that induce expression of the pancreatic genes Pdx1, p48, Nkx6.1, glucagon, and insulin. Moreover, this instructive signal directs cells to form ectopic insulin-positive islet-like clusters in endoderm that would otherwise form more rostral organs. Once generated, endocrine cells no longer require interaction with mesoderm, but nonendocrine cells continue to require permissive signals from the mesoderm. Stimulation of activin, BMP, or retinoic acid signaling is sufficient to induce Pdx1 expression in endoderm anterior to the pancreas. Lateral plate mesoderm appears to pattern the endoderm in a posterior-dominant fashion as first noted in the patterning of the neural tube at the same embryonic stage. These findings argue for a central role of the mesoderm in coordinating the A-P pattern of all three primary germ layers.

Activins↗
PubMed2003

MyoR is expressed in nonmyogenic cells and can inhibit their differentiation.

The development of skeletal muscle in mammals is promoted by the muscle-specific basic helix-loop-helix transcription factors of the MyoD family. Evidence also suggests that there are basic helix-loop-helix proteins that specifically inhibit skeletal myogenesis, including Mtwist, Mist1, and the most recently described, MyoR. It has been suggested that MyoR expression is limited to the precursors of the skeletal muscle lineage and acts as a transcriptional repressor of the muscle differentiation program. However, our results demonstrate that MyoR is expressed in several different, nonmuscle adult tissues. Furthermore, MyoR is expressed in the embryonic ectoderm of blastocyst stage mouse embryos, well before skeletal muscle specification and even before delineation of the mesodermal germ layer. Using embryonic ectoderm analogous stem cells, we demonstrate that in these nonmuscle cells, as in skeletal muscle precursor cells, expression of MyoR is inversely correlated with the extent of cellular differentiation as induced by retinoic acid. Our preliminary results indicate that overexpression of exogenous MyoR inhibits retinoic-acid-induced differentiation in EC cells and is lethal to early mouse embryos. Our results suggest a much broader role for MyoR in the repression and/or determination of embryonic cell differentiation.

Animals↗
PubMed2002

A molecular view on pluripotent stem cells.

Pluripotent stem cells are undifferentiated cells that are capable of differentiating to all three embryonic germ layers and their differentiated derivatives. They are transiently found during embryogenesis, in preimplantation embryos and fetal gonads, or as established cell lines. These unique cell types are distinguished by their wide developmental potential and by their ability to be propagated in culture indefinitely, without loosing their undifferentiated phenotype. This short review intends to give a general overview on the pluripotent nature of embryo-derived stem cells with a focus on human embryonic stem cells.

Animals↗
PubMed2003

Developmental ontogeny of prolactin and prolactin receptor in the sea bream (Sparus aurata).

The expression of PRL and its receptor (PRLR) were characterised during sea bream embryonic and larval development, by semi-quantitative and quantitative RT-PCR, respectively, until 46 days post-hatch (DPH). Immunocytochemistry with antisera specific for sea bream PRLR was carried out with larval sections from hatching up to 46 DPH. A single transcript of PRL (1.35 Kb) and PRLR (2.8 Kb) identical to the transcripts previously characterised in adult tissue, are present in sea bream embryos and larvae. PRL expression is first detectable at neurula and in all samples collected thereafter. The lowest levels of PRL mRNA are detected in sea bream embryos up until neurula when expression starts to increase. The maximal levels of PRL expression were detected at 24 DPH. PRLR transcripts first appear at 12h post-fertilisation (0.002 rho mol/microg total larvae RNA) (blastula) and increase significantly during gastrulation (0.245 rho mol/microg total larvae RNA) reaching a maximum at 2 DPH (0.281 rho mol/microg total larvae RNA). After hatching a significant reduction in PRLR expression is observed which reaches a minimum at 4 DPH (0.103 rho mol/microg total larvae RNA), gradually increasing thereafter. Immunocytochemistry revealed the presence of PRLR in early post-hatching stages of larvae in tissues derived from all three germ layers.

Aging↗
PubMed1997

Developmental genetics of deleted mtDNA in mitochondrial oculomyopathy.

Heteroplasmic populations of mtDNA, consisting of normal mtDNA and mtDNA with large deletions, are found in the skeletal muscle and other tissues of certain patients with mitochondrial respiratory chain deficiencies, particularly in those with the CPEO (chronic progressive external ophthalmoplegia) phenotype. To study the developmental genetics of this mitochondrial disorder, the distribution of the deleted mtDNA in a wide range of tissues of different embryonic origins (total 34 samples from 27 tissues obtained at autopsy) was investigated in a patient with the CPEO syndrome. Three species of partially deleted mtDNA were observed, with deletions of 2.3 kb, 5.0 kb and 6.4 kb. Their tissue distribution suggests that the mtDNA deletions have occurred very early during embryonic development, prior to the differentiation events that lead to the formation of the three primary embryonic germ layers, and that the partially deleted mtDNA species were segregated during development mainly to the skeletal muscle and to tissues of the central nervous system.

Base Sequence↗
PubMed2001

Translational control in vertebrate development.

Translational control plays a large role in vertebrate oocyte maturation and contributes to the induction of the germ layers. Translational regulation is also observed in the regulation of cell proliferation and differentiation. The features of an mRNA that mediate translational control are found both in the 5' and in the 3' untranslated regions (UTRs). In the 5' UTR, secondary structure, the binding of proteins, and the presence of upstream open reading frames can interfere with the association of initiation factors with the cap, or with scanning of the initiation complex. The 3' UTR can mediate translational activation by directing cytoplasmic polyadenylation and can confer translational repression by interference with the assembly of initiation complexes. Besides mRNA-specific translational control elements, the nonspecific RNA-binding proteins contribute to the modulation of translation in development. This review discusses examples of translational control and their relevance for developmental regulation.

Animals↗
PubMed1996

Distribution and functions of parathyroid hormone-related protein in vertebrate cells.

Parathyroid hormone-related protein (PTHrP) was isolated from tumors and identified as the agent of humoral hypercalcemia of malignancy (HHM) in 1987. Since then its gene structure in several mammalian and an avian species has been analyzed and its gene expression demonstrated in many adult and embryonic tissues derived from all three germ layers. The composition and structure of PTHrP peptide depends on both differential gene splicing and posttranslational processing, which result in a range of peptides of potentially diverse functions. This chapter describes the distribution of PTHrP in both normal and neoplastic adult and embryonic tissues. PTHrP is of fundamental importance to cell survival because the absence of the gene is fatal; this aspect of PTHrP function in cell physiology becomes overwhelmingly important in neoplasia. Intracrine or paracrine actions for PTHrP seem to be most likely in mammalian and avian physiology, but in fishes high circulating levels suggest classic endocrine functions as well. Much remains to be learned of the biology of this fascinating protein.

Amino Acid Sequence↗
PubMed2006

Embryonic stem cells from parthenotes.

While human embryonic stem cells (hESCs) hold tremendous therapeutic potential, they also create societal and ethical dilemmas. Adult and placental stem cells represent two alternatives to the hESC, but may have technical limitations. An additional alternative is the stem cell derived from parthenogenesis. Parthenogenesis is a reproductive mechanism that is common in lower organisms and produces a live birth from an oocyte activated in the absence of sperm. However, parthenogenetic embryos will develop to the blastocyst stage and so can serve as a source of embryonic stem cells. Parthenogenetic ESCs (pESCs) have been shown to have the properties of self-renewal and the capacity to generate cell derivatives from the three germ layers, confirmed by contributions to chimeric animals and/or teratoma formation when injected into SCID mice. Therefore, this mechanism for generating stem cells has the ethical advantage of not involving the destruction of viable embryos. Moreover, the cells do not involve the union of male and female and so genetic material will be derived exclusively from the female oocyte donor (with the attendant potential immunological advantages). This chapter describes the biology underlying parthenogenesis, as well as provides detailed technical considerations for the production of pESCs.

Animals↗
PubMed2006

Vascular cells.

Embryonic stem (ES) cells are cells derived from the inner cell mass of a blastocyst stage embryo. These self-renewing multipotent cells are able to differentiate to the three embryonic germ layers, the endoderm, ectoderm, and mesoderm, and are thus able to produce virtually all cell types. The ES cell capacity to generate various cell types has been studied extensively, and exploitation of ES cell characteristics allowed the production of several differentiated cell types of multiple tissues. Moreover, the process of ES cell differentiation provides a unique opportunity to observe early embryonic developmental events that are unattainable in the embryo itself. This chapter addresses the in vitro differentiation procedure of endothelial and vascular smooth muscle cells from human ES cells, with reference to similar studies performed in mouse and nonhuman primate ES cells, and provides several tools for the detailed characterization of differentiated cells.

Animals↗
PubMed2006

Amniotic fluid and placental stem cells.

Human amniotic fluid has been used in prenatal diagnosis for more than 70 years. It has proven to be a safe, reliable, and simple screening tool for a wide variety of developmental and genetic diseases. However, there is now evidence that amniotic fluid may have more use than only as a diagnostic tool and may be the source of a powerful therapy for a multitude of congenital and adult disorders. A subset of cells found in amniotic fluid and placenta has been isolated and found to be capable of maintaining prolonged undifferentiated proliferation as well as able to differentiate into multiple tissue types encompassing the three germ layers. It is possible that in the near future, we will see the development of therapies using progenitor cells isolated from amniotic fluid and placenta for the treatment of newborns with congenital malformations as well as of adults, using cryopreserved amniotic fluid and placental stem cells. In this chapter, we describe a number of experiments that have isolated and characterized pluripotent progenitor cells from amniotic fluid and placenta. We also discuss various cell lines derived from amniotic fluid and placenta and future directions for this area of research.

Adult Stem Cells↗
PubMed1999

Confocal microscopic analysis of morphogenetic movements.

Confocal microscopy is an excellent means of imaging cellular dynamics within living zebrafish embryos because it provides a means of optically sectioning tissues that have been labeled with specific fluorescent probe molecules. In order to study genetically encoded patterns of cell behavior that are involved in the formation of germ layers and various organ primordia, it is possible to vitally stain an entire zebrafish embryo with one or more fluorescent probe molecules and then examine morphogenetic behaviors within specific cell populations of interest using time-lapse confocal microscopy. There are two major advantages to this "bulk-labeling" approach: (1) the applied fluorescent probe (a contrast-enhancing agent) allows all of the cells within an intact zebrafish embryo to be rapidly stained; (2) the morphogenetic movements and shape changes of hundreds of cells can then be examined simultaneously in vivo using time-lapse confocal microscopy. The neutral fluorophore Bodipy 505/515 and its sphingolipid-derivative Bodipy-C5-ceramide are particularly useful, nonteratogenic vital stains for imaging cellular dynamics in living zebrafish embryos. These photostable fluorescent probes (when applied with 2% DMSO) percolate through the enveloping layer epithelium of the embryo, and localize in yolk-containing cytoplasm and interstitial space, respectively, owing to their different physiochemical characteristics. Bodipy-ceramide, for instance, remains highly localized to interstitial fluid once it accumulates within a zebrafish embryo, allowing the boundaries of deep cells to be clearly discerned throughout the entire embryo. Through the use of either of these fluorescent vital stains, it is possible to rapidly convert a developing zebrafish embryo into a strongly fluorescent specimen that is ideally suited for time-lapse confocal imaging. For zebrafish embryos whose deep cells have been intentionally "scatter-labeled" with fluorescent lineage tracers (e.g., fluorescent dextrans), sequential confocal z-series (i.e., focus-throughs) of the embryo can be rendered into uniquely informative 3D time-lapse movies using readily available image-processing programs. Similar time-lapse imaging, combined with rapidly advancing computer-assisted visualization techniques, may soon be applied to study the dynamics of GFP-fusion proteins in vivo, as well as other types of synthetic probe molecules designed to reveal the cytological processes associated with the patterning and morphological transformations of the zebrafish's embryonic tissues.

Animals↗
PubMed1999

The EGF-CFC protein one-eyed pinhead is essential for nodal signaling.

The zebrafish EGF-CFC gene one-eyed pinhead (oep) is required zygotically for the formation of the ventral neuroectoderm, endoderm, and prechordal plate. Here we report that embryos lacking both maternal and zygotic Oep activity are defective in germ layer formation, organizer development, and the positioning of the anterior-posterior axis. An identical phenotype is displayed by double mutants for the nodal-related genes squint and cyclops. Mutations in oep eliminate the response to Squint and Cyclops overexpression but are suppressed by expression of Activin and activated forms of the type I receptor ActRIB and Smad2. Expression of the murine EGF-CFC gene cripto rescues oep mutants. These results suggest a conserved role for EGF-CFC proteins as essential extracellular cofactors for Nodal signaling during vertebrate development.

Activins↗