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PubMed1996

Development of cardiac musculature in the cranial vena cava of rat embryos.

Development of cardiac musculature in the rat cranial vena cava (common cardinal vein or duct of Cuvier) was examined by immunohistochemistry and transmission electron microscopy. Undifferentiated cardiac myocytes were detected in the cranial vena cava wall of rat embryos after 12.5 days post-coitum (dpc). The tunica media of the cranial vena cava was composed of cardiac myocytes after formation of the endothelium. Therefore, the cranial vena cava may be not only a part of the venous system but also of the heart. Myocytes in the cranial vena cava contained developing myofibrils, mitochondria and intercalated discs similar to those found in the myocytes in heart. Striated myofibrils began to differentiate as soon as myocytes appeared in the vena cava wall, and myocytes with differentiating myofibrils occur in the wall as the first component of the tunica media at 12.5 dpc. We concluded that the cardiac musculature in the vena cava is not a secondary extension into the tunica media after birth only in the rat, but a basic structure formed in all mammals during early embryonic development.

Animals↗
PubMed1985

Fate of DNA injected into early Drosophila embryos.

Plasmid DNA injected into early Drosophila embryos becomes enclosed in nuclei or nucleus-like structures where it remains at least throughout embryonic development. The fraction remaining in the cytoplasm is gradually degraded. The nuclear fraction is converted to a high-molecular-weight form consisting largely but not exclusively of tandem oligomers. Some of it, however, can occasionally become integrated in the genome. Extensive DNA replication takes place but few, if any, molecules are able to initiate a second round of replication.

Animals↗
PubMed1983

Spontaneous formation of nucleus-like structures around bacteriophage DNA microinjected into Xenopus eggs.

We have found that injection of bacteriophage lambda DNA into unfertilized Xenopus eggs causes the assembly around the DNA of structures resembling typical eucaryotic cell nuclei. These spherical structures begin to form 60-90 min after injection. They contain lambda DNA and are bounded by a phase-dense envelope. Immunofluorescent staining of the lambda-DNA-containing structures with anti-lamin antibody reveals the presence of the lamin nuclear proteins at the periphery of the structure, a pattern identical to that of embryonic nuclei. Electron microscopy reveals that the injected DNA is surrounded by a double bilayer nuclear membrane containing nuclear pore complexes. The "nuclei" containing lambda DNA respond to modulators of the Xenopus cell cycle in a manner that mimics the response of embryonic nuclei to these modulators during mitosis. These results suggest that nuclear reassembly and breakdown occur independently of specific DNA sequence information.

Animals↗
PubMed1984

Connective tissue metabolism in muscular dystrophy. Levels of collagen and mucopolysaccharides in embryonic chickens with genetic muscular dystrophy.

There were marked differences between the levels of collagen (measured as hydroxyproline) and mucopolysaccharides (measured as hexosamine) found in embryonic chicks with genetic muscular dystrophy and their normal controls. The chief differences were that the dystrophic tissues (gastrocnemius muscle and tendon, pectoralis major and skin) had: (a) greater amounts of hexosamine early in embryonic development; (b) hydroxyproline levels that rose at a faster rate, yielding different slopes than their normal controls; (c) relatively greater amounts of hydroxyproline than hexosamine later in embryonic life (day 20). Connective tissue systems in muscles were preferentially affected. The connective tissue system associated with dystrophic tissues appeared to lag behind the normal rhythm pattern of embryological development. The changes in connective tissue metabolism observed in dystrophic chicks suggested that the collagen from dystrophic embryonic chicks may be of a different structure or composition than that found in the normals.

Animals↗
PubMed2004

Spatial and temporal alterations in the collagen fibrillar array during the onset of transparency in the avian cornea.

In the latter stages of development, the embryonic avian cornea undergoes significant changes in structure, composition and transparency. The rearrangement of stromal collagen fibrils at this time is important because it is believed to play a key role in the acquisition of corneal transparency. Here, we investigate spatial alterations in the internal fine structure of the avian cornea during development. Chicken corneas at developmental days 14, 16 and 18 were examined by transmission electron microscopy and quantitative image analysis. For anterior and posterior regions we determined fibril number densities, two-dimensional distribution functions, and, where appropriate, radial distribution functions. Stromal collagen fibrils became more closely spaced over the developmental range studied here. Changes in fibril number density indicated that fibrils became compacted first in the anterior stroma, and later (i.e. after day 16) in the posterior stroma. By day 18 collagen fibril number densities were essentially the same in superficial and deep tissue regions. At day 14, two-dimensional distribution functions of collagen fibrils in the posterior stroma pointed to a fibrillar array that was unlike that in the anterior stroma because there was no clear radial symmetry. Rather, in the deep stroma at day 14 there was evidence of different nearest neighbour spacings in two orthogonal directions. By day 18, fibril distributions in the anterior and posterior stroma were spatially homogeneous and radially symmetric, with radial distribution functions typical of those ordinarily found in mature cornea. Corneal transparency requires the stromal matrix to have some degree of regularity in the arrangement of its uniformly thin collagen fibrils. The chicken cornea becomes progressively transparent between days 14 and 18 of development as the stroma dehydrates and thins. We show that over this time period collagen fibrils in the anterior stroma become configured in advance of fibrils in deeper stromal regions, leading to questions over the potential roles of sulphated proteoglycans in different regions of the corneal stroma during morphogenesis.

Animals↗
PubMed2006

Mechanistic insight into how Shh patterns the vertebrate limb.

The hands and feet of a newborn baby are a beautiful reminder of the complexity of embryonic patterning. Classical studies on how these structures form have led to a theoretical framework for understanding, in general, how discrete groups of cells can instruct differential fates across a wider field through the action of long-range signals. The discovery just more than a decade ago that localized expression of Sonic hedgehog (Shh) differentially patterns structures across the limb field, resulting in digits with unique characteristics, provided a starting point for readdressing these models at a molecular level. Current research has revealed unexpected complexity in how a gradient of Shh activity is both established and received, prompting re-evaluation of the nature of patterning mechanisms within the limb.

Animals↗
PubMed2000

Thalidomide inhibits angiogenesis in embryoid bodies by the generation of hydroxyl radicals.

Thalidomide is a teratogen with anti-angiogenic properties and causes stunted limb growth (dysmelia) during human embryogenesis. The molecular mechanisms of thalidomide action in embryopathy are currently unknown. Using the endothelial-specific antigen platelet endothelial cell adhesion molecule-1 and confocal laser scanning microscopy we have demonstrated that thalidomide exerts anti-angiogenic effects on the development of capillary structures in embryoid bodies differentiated from murine embryonic stem cells. Consequently, in thalidomide-treated embryoid bodies the diffusion properties of the tissue were deteriorated. Thalidomide raised reactive oxygen species (ROS), as revealed using 2'7'-dichlorodihydrofluorescein diacetate (H(2)DCF-DA) as an indicator. A comparable ROS generation was achieved with the thalidomide hydrolysis product phthaloyl glutamic acid (PGA), but not with phthalimide (PI), the major component of thalidomide. ROS formation by thalidomide was inhibited by the hydroxyl radical scavengers mannitol and 2-mercaptoethanol. After coadministration of either 2-mercaptoethanol or mannitol with thalidomide the anti-angiogenic effects of thalidomide were abolished and the diffusion properties of the tissue were restored to the control values. In summary, our data suggest that thalidomide exerts its anti-angiogenic properties via the generation of toxic hydroxyl radicals, which impair vasculogenesis and angiogenesis during embryoid body development.

Angiogenesis Inhibitors↗
PubMed2003

Signal-induced ubiquitination of p57(Kip2) is independent of the C-terminal consensus Cdk phosphorylation site.

The cyclin-dependent kinase inhibitor p57(Kip2) is required for normal mouse embryonic development. p57(Kip2) consists of four structurally distinct domains in which the conserved C-terminal nuclear targeting domain contains a putative Cdk phosphorylation site (Thr(342)) that shares a great similitude in the adjacent sequences with p27(Kip1) but not with p21(Cip1). Phosphorylation on Thr(187) has been shown to promote degradation of p27(Kip1). Although there is sequence homology between the C-terminal part of p27(Kip1) and p57(Kip2), we show that the ubiquitination and degradation of p57(Kip2) are independent of Thr(342). In contrast a destabilizing element located in the N-terminal is implicated in p57(Kip2) destabilization.

Amino Acid Sequence↗
PubMed1998

VEGF gene delivery to muscle: potential role for vasculogenesis in adults.

Constitutive expression of VEGF after implantation of genetically engineered myoblasts into non-ischemic muscle led to an increase in vascular structures. Previously, effects of VEGF delivery to adult muscle have only been reported in ischemic tissues. The resulting vascular structures were reminiscent of those formed during embryonic vasculogenesis, rather than angiogenesis, sprouting from preexisting vessels. Initially, VEGF caused an accumulation of endothelial cells and macrophages, followed by networks of vascular channels and hemangiomas with locally high serum VEGF levels. No effects were evident in adjacent tissue or contralateral legs, where low serum VEGF was detected. These data suggest that the induction by VEGF of angiogenesis or vasculogenesis may be dose-dependent. Furthermore, VEGF expression must be carefully modulated, as overexpression is deleterious.

Acetyltransferases↗
PubMed1997

Sequence characterization of gamma-crystallins from lip shark (Chiloscyllium colax): existence of two cDNAs encoding gamma-crystallins of mammalian and teleostean classes.

gamma-Crystallin is a common lens protein of most vertebrate eye lenses and the major protein component in lenses of fishes and in many mammalian species during embryonic and neonatal stages. To facilitate the structural characterization of gamma-crystallin possessing extensive charge heterogeneity, a cDNA mixture was constructed from the poly(A)+ mRNA isolated from shark eye lenses, and amplification by polymerase chain reaction (PCR) was carried out to obtain cDNAs encoding multiple shark gamma-crystallins. Sequencing analysis of multiple positive clones containing PCR-amplified inserts revealed the presence of a multiplicity of isoforms in the gamma-crystallin class of this cartilaginous fish. It was of interest to find that two shark cDNA sequences coexist, one encoding gamma-crystallin (gamma M1) of high methionine content (15.5%) and the other encoding one (gamma M2) of low methionine content (5.1%), each corresponding to the major teleostean and mammalian gamma-crystallins, respectively. Comparison of protein sequences encoded by these two shark cDNAs with published sequences of gamma-crystallins from mouse, bovine, human, frog, and carp lenses indicated that there is about 61-80% sequence homology between different species of the piscine class, whereas only 47-66% is found between mammals and shark. A phylogenetic tree constructed on the basis of sequence divergence among various gamma-crystallin cDNAs revealed the close relatedness between shark gamma M2-crystallin and mammalian gamma-crystallins and that between shark gamma M1 and teleostean gamma-crystallins. The results pointed to the fact that ancestral precursors of gamma-crystallins were present in the sharp lens long before the appearance of modern-day mammalian and teleostean gamma-crystallins.

Amino Acid Sequence↗
PubMed2000

Ciliated foregut cyst of the gallbladder: a case report and review of the literature.

A case is presented of a ciliated cyst of the gallbladder in a 36-year-old Korean woman which was incidentally found on ultrasonographic study. A cystic mass measuring 1.5 x 1 x 1 cm was found in the fundus of the gallbladder. The cyst was unilocular and intramural without communication to the lumen. Microscopically, the cyst wall was lined by a single layer of pseudostratified, ciliated, columnar epithelium and goblet cells with underlying smooth muscle layers. This was considered to be the cyst arising from the embryonic foregut and showing differentiation toward respiratory structures. The term 'ciliated foregut cyst of the gallbladder' is suggested here.

Adult↗
PubMed1993

Immunoelectron microscopic localization of small nuclear ribonucleoproteins and interchromatin granules in the 2-cell mouse embryo.

The distribution of U1, U2, U4, U5 and U6 small nuclear ribonucleoproteins (snRNPs) and interchromatin granules (IGs) was studied by electron-microscopic immunocytochemistry (EMI) in early 2-cell mouse embryos at the onset of embryonal transcription. The localization of these antigen structures was evaluated with respect to nucleoplasmic ribonucleoprotein (RPN) regions consisting of interchromatin and perichromatin areas. SnRNP structures of maternal origin (labelled with anti-Sm antibody) were widely distributed throughout the nucleoplasm. Specifically labelled IGs were detected by gold particle clusters distributed in the interchromatin regions of the nucleoplasm. Both immunodetections were negative in nucleolar precursor bodies (NPBs). In addition, the labelling of condensed chromatin blocks with anti-DNA antibody showed heterochromatin topology at this developmental stage. Small condensed chromatin blocks were distributed throughout the nucleus and also appeared in association with the NPB rim. The observed status quo represents a transient state of nuclear structure rearrangement.

Animals↗
PubMed2003

An experimental platform for studying growth and invasiveness of tumor cells within teratomas derived from human embryonic stem cells.

There is currently no available experimental system wherein human cancer cells can be grown in the context of a mixed population of normal differentiated human cells for testing biological aspects of cancer cell growth (e.g., tumor cell invasion and angiogenesis) or response to anti-cancer therapies. When implanted into immunocompromised mice, human embryonic stem cells develop teratomas containing complex structures comprising differentiated cell types representing the major germ line-derived lineages. We sought to determine whether human cancer cells would grow within such teratomas and display properties associated with malignancy, such as invasiveness and recruitment of blood vessels. HEY ovarian cancer cells stably expressing an H2A-GFP fusion protein (HEY-GFP) injected into mature teratomas developed into tumors, which allowed tracking of tumor cell invasion and recruitment of human teratoma-derived blood vessels. This provides a straightforward and powerful approach to studying the biological properties of cancer cells within the microenvironment of normal differentiated human cells.

Animals↗
PubMed1996

Differential 9-cis-retinoic acid-dependent transcriptional activation by murine retinoid X receptor alpha (RXR alpha) and RXR beta. Role of cell type and RXR domains.

The 9-cis-retinoic acid (9cRA)-inducible enhancer of the rat cellular retinol-binding protein type II gene (CRBP II) was shown to be differentially regulated by the murine retinoid X receptor alpha (RXR alpha) as compared with RXR beta. Transient transfection assays performed in NIH 3T3 fibroblast cells demonstrated that RXR alpha yielded a high level of 9cRA-dependent transcription of a reporter gene linked to the CRBP II enhancer, when compared with RXR beta. This effect was cell type-dependent, since both receptors elicited comparable transcriptional activation of the same reporter in P19 embryonal carcinoma cells. To further explore the structural determinants responsible for the differences between these two receptors, a series of chimeric receptor constructs were made. Co-transfection assays utilizing these chimeras demonstrated that both the N terminus and the hinge region connecting the DNA binding domain with the ligand binding domain of RXR alpha were responsible for the high level of 9cRA-dependent transcription observed in NIH 3T3 cells, Furthermore, the hinge region of RXR alpha was shown to be necessary to repress, in the absence of hormone, the transcriptional activation function located in the N-terminal domain of RXR alpha. These results stress the importance of functional links between different RXR domains and suggest an RXR subtype and cell type-dependent specificity in the control of the 9cRA response.

3T3 Cells↗
PubMed2002

Congenital tumours and tumour-like lesions in domestic animals. 1. Cattle. A review.

The literature on congenital tumours and tumour-like lesions in calves was reviewed. Lesions were subdivided by their anatomical distribution and in addition also according to their histologic-pathogenetic nature. As a result of the latter method, four main groups were formed covering most of the lesions described so far: malignant lymphomas, mesotheliomas, hamartomas and embryonic tumours. Most lesions were of mesenchymal structure, carcinomas being extremely rare. Some findings may point to early genetic events, for instance twin calves both affected with malignant lymphoma and related calves with congenital facial neurofibromatosis. An external factor, asbestos, is suspected to play a role in the genesis of peritoneal mesotheliomatosis. The effects of congenital tumours on their hosts were often considerable: death by generalization (malignant lymphomas), ascites (mesotheliomas) or the growth of large abdominal tumours (nephroblastomas, mixed tumours). The latter two conditions often caused dystocia.

Animals↗
PubMed1995

Thrombospondin-4, an extracellular matrix protein expressed in the developing and adult nervous system promotes neurite outgrowth.

Extracellular matrix (ECM) molecules are involved in multiple aspects of cell-to-cell signaling during development and in the adult. In nervous system development, specific recognition processes, e.g., during axonal pathfinding and synaptogenesis involve modulation and signaling by ECM components. Much less is known about their presence and possible roles in the adult nervous system. We now report that thrombospondin-4 (TSP-4), a recently discovered member of the TSP gene family is expressed by neurons, promotes neurite outgrowth, and accumulates at the neuromuscular junction and at certain synapse-rich structures in the adult. To search for muscle genes that may be involved in neuromuscular signaling, we isolated cDNAs induced in adult skeletal muscle by denervation. One of these cDNAs coded for the rat homologue of TSP-4. In skeletal muscle, it was expressed by muscle interstitial cells. The transcript was further detected in heart and in the developing and adult nervous system, where it was expressed by a wide range of neurons. An antiserum to the unique carboxyl-terminal end of the protein allowed to specifically detect TSP-4 in transfected cells in vitro and on cryostat sections in situ. TSP-4 associated with ECM structures in vitro and in vivo. In the adult, it accumulated at the neuromuscular junction and at synapse-rich structures in the cerebellum and retina. To analyze possible activities of TSP-4 towards neurons, we carried out coculture experiments with stably transfected COS cells and motor, sensory, or retina neurons. These experiments revealed that TSP-4 was a preferred substrate for these neurons, and promoted neurite outgrowth. The results establish TSP-4 as a neuronal ECM protein associated with certain synapse-rich structures in the adult. Its activity towards embryonic neurons in vitro and its distribution in vivo suggest that it may be involved in local signaling in the developing and adult nervous system.

3T3 Cells↗
PubMed1971

Morphogenesis of the collagenous stroma in the chick cornea.

The embryonic chick corneal epithelium produces a highly structured acellular matrix beneath its basal surface during early development. This matrix, which contains collagen, serves as a morphogenetic template for subsequent stromal development in that the three-dimensional architecture of the adult corneal stroma is initially established, in miniature, in this epithelially derived connective tissue. Examination of the early corneal epithelium and matrix in both the light and electron microscope suggests that self assembly of the matrix may be one of several important factors in the morphogenesis of this early connective tissue.

Animals↗
PubMed1972

A fine-structural analysis of the fusion of myogenic cells.

The fusion of myogenic cells has been examined on the fine-structural level in muscle cell cultures of embryonic Japanese Coturnix quail. Cells, selected by light microscopy, were serially sectioned normal to their long axis. In this plane, oblique sections of cell membranes are rare and plasmalemmal profiles are more easily traced between adjacent cells. In seven cases, pairs of cells, apparently fixed in the process of fusion, are joined by a single cytoplasmic bridge. Since obliquely sectioned membranes often suggest cytoplasmic confluence, tilting stage analysis was employed to resolve cell membranes in suspect cases. In contrast to such artifacts of superposition, however, the observed intercommunicating pores are contained within a pair of culs-de-sac formed by the fused membranes of both cells. These blind pouches can be traced back between the cells to the external space. The confluent regions are clearly demarcated and they are not simply areas between vesicular profiles. The results of this analysis suggest that (a) at no time is there any loss of integrity of the cellular envelope, and (b) fusion is most probably initiated at single sites between pairs of cells, the pore enlarging, leaving first vestiges and eventually no trace of the original intervening membranes.

Animals↗