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(1/8670) The homeobox gene Pitx2: mediator of asymmetric left-right signaling in vertebrate heart and gut looping.

Left-right asymmetry in vertebrates is controlled by activities emanating from the left lateral plate. How these signals get transmitted to the forming organs is not known. A candidate mediator in mouse, frog and zebrafish embryos is the homeobox gene Pitx2. It is asymmetrically expressed in the left lateral plate mesoderm, tubular heart and early gut tube. Localized Pitx2 expression continues when these organs undergo asymmetric looping morphogenesis. Ectopic expression of Xnr1 in the right lateral plate induces Pitx2 transcription in Xenopus. Misexpression of Pitx2 affects situs and morphology of organs. These experiments suggest a role for Pitx2 in promoting looping of the linear heart and gut.  (+info)

(2/8670) oko meduzy mutations affect neuronal patterning in the zebrafish retina and reveal cell-cell interactions of the retinal neuroepithelial sheet.

Mutations of the oko meduzy (ome) locus cause drastic neuronal patterning defect in the zebrafish retina. The precise, stratified appearance of the wild-type retina is absent in the mutants. Despite the lack of lamination, at least seven retinal cell types differentiate in oko meduzy. The ome phenotype is already expressed in the retinal neuroepithelium affecting morphology of the neuroepithelial cells. Our experiments indicate that previously unknown cell-cell interactions are involved in development of the retinal neuroepithelial sheet. In genetically mosaic animals, cell-cell interactions are sufficient to rescue the phenotype of oko meduzy retinal neuroepithelial cells. These cell-cell interactions may play a critical role in the patterning events that lead to differentiation of distinct neuronal laminae in the vertebrate retina.  (+info)

(3/8670) The Gab1 PH domain is required for localization of Gab1 at sites of cell-cell contact and epithelial morphogenesis downstream from the met receptor tyrosine kinase.

Stimulation of the hepatocyte growth factor (HGF) receptor tyrosine kinase, Met, induces mitogenesis, motility, invasion, and branching tubulogenesis of epithelial and endothelial cell lines in culture. We have previously shown that Gab1 is the major phosphorylated protein following stimulation of the Met receptor in epithelial cells that undergo a morphogenic program in response to HGF. Gab1 is a member of the family of IRS-1-like multisubstrate docking proteins and, like IRS-1, contains an amino-terminal pleckstrin homology domain, in addition to multiple tyrosine residues that are potential binding sites for proteins that contain SH2 or PTB domains. Following stimulation of epithelial cells with HGF, Gab1 associates with phosphatidylinositol 3-kinase and the tyrosine phosphatase SHP2. Met receptor mutants that are impaired in their association with Gab1 fail to induce branching tubulogenesis. Overexpression of Gab1 rescues the Met-dependent tubulogenic response in these cell lines. The ability of Gab1 to promote tubulogenesis is dependent on its pleckstrin homology domain. Whereas the wild-type Gab1 protein is localized to areas of cell-cell contact, a Gab1 protein lacking the pleckstrin homology domain is localized predominantly in the cytoplasm. Localization of Gab1 to areas of cell-cell contact is inhibited by LY294002, demonstrating that phosphatidylinositol 3-kinase activity is required. These data show that Gab1 is an important mediator of branching tubulogenesis downstream from the Met receptor and identify phosphatidylinositol 3-kinase and the Gab1 pleckstrin homology domain as crucial for subcellular localization of Gab1 and biological responses.  (+info)

(4/8670) p27 is involved in N-cadherin-mediated contact inhibition of cell growth and S-phase entry.

In this study the direct involvement of cadherins in adhesion-mediated growth inhibition was investigated. It is shown here that overexpression of N-cadherin in CHO cells significantly suppresses their growth rate. Interaction of these cells and two additional fibroblastic lines with synthetic beads coated with N-cadherin ligands (recombinant N-cadherin ectodomain or specific antibodies) leads to growth arrest at the G1 phase of the cell cycle. The cadherin-reactive beads inhibit the entry into S phase and the reduction in the levels of cyclin-dependent kinase (cdk) inhibitors p21 and p27, following serum-stimulation of starved cells. In exponentially growing cells these beads induce G1 arrest accompanied by elevation in p27 only. We propose that cadherin-mediated signaling is involved in contact inhibition of growth by inducing cell cycle arrest at the G1 phase and elevation of p27 levels.  (+info)

(5/8670) Endothelial cells modulate the proliferation of mural cell precursors via platelet-derived growth factor-BB and heterotypic cell contact.

Embryological data suggest that endothelial cells (ECs) direct the recruitment and differentiation of mural cell precursors. We have developed in vitro coculture systems to model some of these events and have shown that ECs direct the migration of undifferentiated mesenchymal cells (10T1/2 cells) and induce their differentiation toward a smooth muscle cell/pericyte lineage. The present study was undertaken to investigate cell proliferation in these cocultures. ECs and 10T1/2 cells were cocultured in an underagarose assay in the absence of contact. There was a 2-fold increase in bromodeoxyuridine labeling of 10T1/2 cells in response to ECs, which was completely inhibited by the inclusion of neutralizing antiserum against platelet-derived growth factor (PDGF)-B. Antisera against PDGF-A, basic fibroblast growth factor, or transforming growth factor (TGF)-beta had no effect on EC-stimulated 10T1/2 cell proliferation. EC proliferation was not influenced by coculture with 10T1/2 cells in the absence of contact. The cells were then cocultured so that contact was permitted. Double labeling and fluorescence-activated cell sorter analysis revealed that ECs and 10T1/2 cells were growth-inhibited by 43% and 47%, respectively. Conditioned media from contacting EC-10T1/2 cell cocultures inhibited the growth of both cell types by 61% and 48%, respectively. Although we have previously shown a role for TGF-beta in coculture-induced mural cell differentiation, growth inhibition resulting from contacting cocultures or conditioned media was not suppressed by the presence of neutralizing antiserum against TGF-beta. Furthermore, the decreased proliferation of 10T1/2 cells in the direct cocultures could not be attributed to downregulation of the PDGF-B in ECs or the PDGF receptor-beta in the 10T1/2 cells. Our data suggest that modulation of proliferation occurs during EC recruitment of mesenchymal cells and that heterotypic cell-cell contact and soluble factors play a role in growth control during vessel assembly.  (+info)

(6/8670) Cell surface sialic acid and the regulation of immune cell interactions: the neuraminidase effect reconsidered.

It has been known for over a decade that sialidase (neuraminidase) treatment could substantially enhance the capacity of resting B cells to stimulate the proliferation of allogeneic and antigen specific, syngeneic T cells. Thus, cell-surface sialic acid was implicated as a potential modulator of immune cell interaction. However, little progress has been made in either identifying explicit roles for sialic acid in this system or in hypothesizing mechanisms to explain the "neuraminidase effect." Here we show for the first time that cell surface sialic acid on medium incubated B cells blocks access to costimulatory molecules on the B cell surface, and that this is the most likely explanation for the neuraminidase effect. Further, we show that it is likely to be upregulation of ICAM-1 and its subsequent engagement of LFA-1 rather than loss of cell surface sialic acid that in part regulates access to CD86 and other costimulatory molecules. However, we cannot exclude a role for CD86-bound sialic acid on the B cell in modulating binding to T cell CD28. Because sialidase treatment of resting B cells but not resting T cells enables T cell activation, we suggest that sialidase treatment may still be an analogue for an authentic step in B cell activation, and show that for highly activated B cells (activated with polyclonal anti-IgM plus INF-gamma) there is specific loss 2, 6-linked sialic acid. Potential roles for sialic acid in modulating B cell/T cell collaboration are discussed.  (+info)

(7/8670) Glucocorticoid down-regulation of fascin protein expression is required for the steroid-induced formation of tight junctions and cell-cell interactions in rat mammary epithelial tumor cells.

Glucocorticoid hormones, which are physiological regulators of mammary epithelium development, induce the formation of tight junctions in rat Con8 mammary epithelial tumor cells. We have discovered that, as part of this process, the synthetic glucocorticoid dexamethasone strongly and reversibly down-regulated the expression of fascin, an actin-bundling protein that also interacts with the adherens junction component beta-catenin. Ectopic constitutive expression of full-length mouse fascin containing a Myc epitope tag (Myc-fascin) in Con8 cells inhibited the dexamethasone stimulation of transepithelial electrical resistance, disrupted the induced localization of the tight junction protein occludin and the adherens junction protein beta-catenin to the cell periphery, and prevented the rearrangement of the actin cytoskeleton. Ectopic expression of either the carboxyl-terminal 213 amino acids of fascin, which includes the actin and beta-catenin-binding sites, or the amino-terminal 313 amino acids of fascin failed to disrupt the glucocorticoid induction of tight junction formation. Mammary tumor cells expressing the full-length Myc-fascin remained generally glucocorticoid responsive and displayed no changes in the levels or protein-protein interactions of junctional proteins or the amount of cytoskeletal associated actin filaments. However, a cell aggregation assay demonstrated that the expression of Myc-fascin abrogated the dexamethasone induction of cell-cell adhesion. Our results implicate the down-regulation of fascin as a key intermediate step that directly links glucocorticoid receptor signaling to the coordinate control of junctional complex formation and cell-cell interactions in mammary tumor epithelial cells.  (+info)

(8/8670) Novel insights into human endometrial paracrinology and embryo-maternal communication by intrauterine microdialysis.

The regulation of human implantation is still unknown. Evidence from mice suggests an essential role for several paracrine mediators but species differences with implantation in the human preclude the extrapolation of these concepts to humans. An intrauterine microdialysis device (IUMD), consisting of microdialysis tubing glued into a balloon catheter on one side and into a polypropylene tube on the other, allows a dynamic and accurate in-vivo measurement of uterine paracrine interactions in humans. Inserted into the uterine cavity in the form of a loop, it can be continuously perfused with saline to reveal a number of relevant cytokines and growth factors in uterine effluents of non-pregnant women in both follicular and luteal phases. These included interleukin (IL)-1alpha, IL-1beta, IL-6, leukaemia inhibitory factor (LIF), macrophage colony-stimulating factor (M-CSF), epidermal growth factor, vascular endothelial growth factor (VEGF), insulin-like growth factor binding protein-1 (IGFBP-1), prolactin, and human chorionic gonadotrophin (HCG). The source of intrauterine HCG is unclear since endometrial mRNA for the HCG beta-subunit is not revealed using reverse transcriptase polymerase chain reaction analysis. Applying urinary HCG locally via the IUMD profoundly alters endometrial secretory parameters. Prolactin, IGFBP-1, and M-CSF are significantly inhibited and VEGF is regulated in a biphasic manner involving early stimulation followed by inhibition of intrauterine levels. Use of the IUMD has thus shown that the urinary HCG preparations routinely used for ovulation induction and luteal support may directly alter endometrial function.  (+info)