Biomedical subjects

Raymond B Runyan

Publications and source records attributed to Raymond B Runyan.

7 recordsLinked to original sources

PubMed2006

TGF beta-mediated RhoA expression is necessary for epithelial-mesenchymal transition in the embryonic chick heart.

Endothelia in the atrioventricular canal (AVC) of the embryonic heart undergo an epithelial-mesenchymal transition (EMT) and migrate into the underlying extracellular matrix. We explore here whether RhoA mediates this EMT. RhoA was detected in all cells of the chick heart during the stages studied. Expression was elevated when EMT was actively occurring. Explants treated with C3 exoenzyme in collagen gel cultures showed a significant decrease in mesenchymal cell numbers. siRNA was used to inhibit RhoA mRNA, and both activated endothelial and mesenchymal cells decreased significantly with treatment. Loss of RhoA produced a reduction of RhoB, cyclin-b2, and beta-catenin messages showing that these genes are regulated downstream of RhoA. In contrast, runx-2 was not reduced. Inhibition of TGFbeta3 or TGFbeta2 activity caused a large reduction of RhoA message. These data place RhoA in TGFbeta regulated pathways for both endothelial activation and mesenchymal invasion and demonstrate a functional requirement during EMT.

Animals↗

PubMed2006

Latrophilin-2 is a novel component of the epithelial-mesenchymal transition within the atrioventricular canal of the embryonic chicken heart.

Endothelial cells in the atrioventricular canal of the heart undergo an epithelial-mesenchymal transition (EMT) to form heart valves. We surveyed an on-line database (http://www.geisha.arizona.edu/) for clones expressed during gastrulation to identify novel EMT components. One gene, latrophilin-2, was identified as expressed in the heart and appeared to be functional in EMT. This molecule was chosen for further examination. In situ localization showed it to be expressed in both the myocardium and endothelium. Several antisense DNA probes and an siRNA for latrophilin-2 produced a loss of EMT in collagen gel cultures. Latrophilin-2 is a putative G-protein-coupled receptor and we previously identified a pertussis toxin-sensitive G-protein signal transduction pathway. Microarray experiments were performed to examine whether these molecules were related. After treatment with antisense DNA against latrophilin-2, expression of 1,385 genes and ESTs was altered. This represented approximately 12.5% of the microarray elements. In contrast, pertussis toxin altered only 103 (0.9%) elements of the array. There appears to be little overlap between the two signal transduction pathways. Latrophilin-2 is thus a novel component of EMT and provides a new avenue for investigation of this cellular process.

Animals↗

PubMed2005

Frzb modulates Wnt-9a-mediated beta-catenin signaling during avian atrioventricular cardiac cushion development.

Normal development of the cardiac atrioventricular (AV) endocardial cushions is essential for proper ventricular septation and morphogenesis of the mature mitral and tricuspid valves. In this study, we demonstrate spatially restricted expression of both Wnt-9a (formerly Wnt-14) and the secreted Wnt antagonist Frzb in AV endocardial cushions of the developing chicken heart. Wnt-9a expression is detected only in AV canal endocardial cells, while Frzb expression is detected in both endocardial and transformed mesenchymal cells of the developing AV cardiac cushions. We present evidence that Wnt-9a promotes cell proliferation in the AV canal and overexpression of Wnt-9a in ovo results in enlarged endocardial cushions and AV inlet obstruction. Wnt-9a stimulates beta-catenin-responsive transcription in AV canal cells, duplicates the embryonic axis upon ventral injections in Xenopus embryos and appears to regulate cell proliferation by activating a Wnt/beta-catenin signaling pathway. Additional functional studies reveal that Frzb inhibits Wnt-9a-mediated cell proliferation in cardiac cushions. Together, these data argue that Wnt-9a and Frzb regulate mesenchymal cell proliferation leading to proper AV canal cushion outgrowth and remodeling in the developing avian heart.

Amino Acid Sequence↗

PubMed2005

Cell biology of cardiac cushion development.

The valves of the heart develop in the embryo from precursor structures called endocardial cushions. After cardiac looping, endocardial cushion swellings form and become populated by valve precursor cells formed by an epithelial-mesenchymal transition (EMT). Endocardial cushions subsequently undergo directed growth and remodeling to form the valvular structures and the membranous septa of the mature heart. The developmental processes that mediate cushion formation include many prototypic cellular actions including adhesion, signaling, migration, secretion, replication, differentiation, and apoptosis. Cushion morphogenesis is unique in that these cellular possesses occur in a functioning organ where the cushions act as valves even while developing into definitive valvular structures. Cardiovascular defects are the most common congenital defects, and one of the most common causes of death during infancy. Thus, there is significant interest in understanding the mechanisms that underlie this complex developmental process. In this regard, substantial progress has been made by incorporating an understanding of cardiac morphology and cell biology with the rapidly expanding repertoire of molecular mechanisms gained through human genetics and research using animal models. This article reviews cardiac morphogenesis as it relates to heart valve formation and highlights selected growth factors, intracellular signaling mediators, and extracellular matrix components involved in the creation and remodeling of endocardial cushions into mature cardiac structures.

Emigration and Immigration↗

PubMed2004

TGF-beta1, -beta2 and -beta3 cooperate to facilitate tubulogenesis in the explanted quail heart.

BACKGROUND: Transforming growth factor-beta (TGF-beta) isoforms have been implicated as both pro- and anti-angiogenic modulators. In this study we addressed the roles of TGF-beta isoforms on coronary tubulogenesis. METHODS: Embryonic (E6) quail ventricular specimens were explanted onto collagen gels allowing endothelial cells to migrate and form vascular tubes. Growth factors and/or neutralizing growth factor antibodies were added to the cultures. Endothelial cells were identified using a quail endothelial cell marker, QH1. Image analysis was used to quantify aggregate tube length. RESULTS: Addition of any isoform (TGF-beta(1), TGF-beta(2) or TGF-beta(3)) virtually prevented tubulogenesis (>95% inhibition), while stimulation of tubulogenesis occurred by adding neutralizing antibodies to TGF-beta(3), but not to TGF-beta(1) or -beta(2). When all three isoforms were added, tubulogenesis was enhanced, indicating the key role of TGF-beta(3). Documentation of the inhibitory effect of TGF-beta isoforms on tubulogenesis is further supported by our experiments in which the marked enhancement of tube formation by bFGF and VEGF was negated when exogenous TGF-beta(1), -beta(2), or -beta(3) were added to the cultures. CONCLUSIONS: (1) TGF-beta(1), -beta(2) and -beta(3) each inhibits angiogenesis; (2) cooperation between the three TGF-beta isoforms and other angiogenic factors is essential for the regulation of normal tubulogenesis and (3) the stimulatory effect of VEGF or bFGF on tubulogenesis is negated by exogenous TGB-betas.

Animals↗

PubMed2003

Trichloroethylene effects on gene expression during cardiac development.

BACKGROUND: Halogenated hydrocarbon exposure is associated with changes in gene expression in adult and embryonic tissue. Our study was undertaken to identify differentially expressed mRNA transcripts in embryonic hearts from Sprague-Dawley rats exposed to trichloroethylene (TCE) or potential bio-transformation products dichloroethylene (DCE) and trichloroacetic acid (TCAA). METHODS: cDNA subtractive hybridization was used to selectively amplify expressed mRNA obtained from control or halogenated hydrocarbon exposed rat embryos. The doses used were 1100 and 110 ppm (8300 and 830 microM) TCE, 110 and 11 ppm (1100 and 110 microM) DCE, and 27.3 and 2.75 mg/ml (100 and 10 mM) TCAA. Control animals were given distilled drinking water throughout the period of experiments. RESULTS: Sequencing of over 100 clones derived from halogenated hydrocarbon exposed groups resulted in identification of numerous differentially regulated gene sequences. Up-regulated transcripts identified include genes associated with stress response (Hsp 70) and homeostasis (several ribosomal proteins). Down-regulated transcripts include extracellular matrix components (GPI-p137 and vimentin) and Ca(2+) responsive proteins (Serca-2 Ca(2+)-ATPase and beta-catenin). Two possible markers for fetal TCE exposure were identified: Serca-2 Ca(2+)-ATPase and GPI-p137, a GPI-linked protein of unknown function. Differential regulation of expression of both markers by TCE was confirmed by dot blot analysis and semi-quantitative RT-PCR with levels of TCE exposure between 100 and 250 ppb (0.76 and 1.9 microM) sufficient to decrease expression. CONCLUSIONS: Sequences down-regulated with TCE exposure appear to be those associated with cellular housekeeping, cell adhesion, and developmental processes, while TCE exposure up-regulates expression of numerous stress response and homeostatic genes.

Animals↗

PubMed2002

Temporal and distinct TGFbeta ligand requirements during mouse and avian endocardial cushion morphogenesis.

The formation of endocardial cushions in the atrioventricular (AV) canal of the rudimentary heart requires epithelial-to-mesenchymal cell transformation (EMT). This is a complex developmental process regulated by multiple extracellular signals and transduction pathways. A collagen gel assay, long used to examine endocardial cushion development in avian models, is now being employed to investigate genetically engineered mouse models with abnormal heart morphogenesis. In this study, we determine interspecies variations for avian and mouse cultured endocardial cushion explants. Considering these observed morphologic differences, we also define the temporal requirements for TGFbeta2 and TGFbeta3 during mouse endocardial cushion morphogenesis. TGFbeta2 and TGFbeta3 blocking antibodies inhibit endothelial cell activation and transformation, respectively, in avian explants. In contrast, neutralizing TGFbeta2 inhibits cell transformation in the mouse, while TGFbeta3 antibodies have no effect on activation or transformation events. This functional requirement for TGFbeta2 is concomitant with expression of TGFbeta2, but not TGFbeta3, within mouse endocardial cushions at a time coincident with transformation. Thus, both TGFbeta2 and TGFbeta3 appear necessary for the full morphogenetic program of EMT in the chick, but only TGFbeta2 is expressed and obligatory for mammalian endocardial cushion cell transformation.

Actins↗