Zebrafish Cardiovascular Developmental Genetics

Objectives

We are studying the embryonic zebrafish heart, a relatively simple organ compared with its mammalian counterpart, to understand the assembly of the early heart tube. We would like to understand: How do myocardium and endocardium communicate during cardiac morphogenesis? What determines the differentiation of endocardium into its different morphological derivatives? In collaboration with clinical researchers, we develop animal models for human cardiovascular diseases. Our long-term interest is to understand how the cellular mechanisms controlling zebrafish cardiogenesis shape the human heart and its associated blood vessels.

Research Focus

Background

Vertebrate organs are derived from epithelial or endothelial sheets of cells that undergo complex morphogenetic transformations. In our lab, we study the embryonic zebrafish heart, a relatively simple organ compared with its mammalian counterpart, to better understand the signaling events that instruct the assembly of the early heart tube. Initially the heart consists of only the outer myocardial and inner endocardial cell layers. The endocardium is a specialized population of endothelial cells that line the interior of the heart. We would like to understand: What are the signals that regulate the morphogenesis of myocardium and endocardium? To what extent do these two tissues communicate during cardiac jogging and looping morphogenesis? What determines the differentiation of endocardium into its different morphological derivatives such as cushion cells? In collaboration with clinical researchers, the group uses developmental genetics combined with cell biological and pharmacological approaches to develop animal models for human cardiovascular diseases. One particular focus of research is to characterize endocardial development. We use highly interdisciplinary approaches to analyze cardiac (both myocardial and endocardial) morphogenesis by combining expression analyses (deep sequencing; microarray studies), functional cell biological and genetic tools for tissue- or single cell level functional studies, in vivo high-resolution 4D-confocal imaging, systems biological approaches, in silico modeling of single cell behaviors during cardiac morphogenesis, and pharmacological studies.

Morphogen signaling during cardiac development

Signaling by morphogens (=form giving molecules) plays an important role in the regulation of cardiac progenitor cell behaviors. In a recent study, we were able to elucidate the molecular crosstalk between two Transforming Growth Factor-b (TGF-b morphogens, Nodals and Bone morphogenetic proteins (Bmps), and to characterize their impact on left/right asymmetric development of the heart. Our highly-interdisciplinary approach, which included detailed expression analyses combined with functional studies, high-resolution live imaging, and mathematical modeling of this process, suggested that Nodal, via modulating the extracellular matrix within the left cardiac field, dampens the efficiency of Bmp signaling on the left (Figure 1; Veerkamp et al., Dev. Cell 2013). Figure 1: Reversal of left/right asymmetric Bmp activity within the heart upon expression of Nodal within the right cardiac field. (A) The transgenic reporter line Tg(BRE:dmKO2)mw40 indicates that the activity of “Bmp”-Smads-1/5/8 is higher within the left compared with the right cardiac field upon misexpression of Nodal (Southpaw) within the right cardiac field (asterisks, GFP false-colored in white). Signaling intensities are indicated by color range. (B) Fluorescence two-color in situ hybridization reveals normal lefty1 (lft1, a target gene of Nodal signaling) cardiac expression in clones expressing a dominant-negative Bmp receptor (myl7:dnbmpr2a). L, left; R, right.

Figure 1.

Our findings suggest that minor left-right differences in Bmp activity within the cardiac field may determine bi-phasic states: cardiac progenitor cells within the right cardiac cone are slightly more adhesive, and cells on the left side exhibit a slightly more motile character. Thus, cardiac left-right asymmetry may be explained by Nodal modulating an anti-motogenic Bmp activity. In clonal misexpression studies, in which Bmp activity was clonally reduced within the right cardiac field, we observed an inversion of cardiac laterality. Moreover, reducing Bmp activity to below normal levels on the left side even enhanced cardiac jogging towards the left (Figure 2). One implication of these clonal studies is that changes in individual cell motility rates may impact the tissue displacement of larger coherent groups of cells.

Figure 2.

Figure 2: (A) Schematic diagram illustrating that the Nodal target Hyaluronan synthase 2 dampens Bmp activity within the left cardiac field which causes lower expression of non-muscle myosin II (NMII) and higher cardiac progenitor cell motility. (B) Cross section through the cardiac field (myocardial cells marked green; F-actin, red).

Molecular control of endocardial chamber morphogenesis

In another study, we have recently analyzed the process of chamber morphogenesis of the endocardium (Dietrich et al., Dev. Cell 2014). During heart development, the onset of heartbeat and blood flow coincides with a ballooning of the cardiac chambers (Figure 3). By combining functional manipulations, fate mapping studies, and high-resolution imaging, we showed that endocardial growth occurs without an influx of external cells. Instead, endocardial cell proliferation is regulated, both by blood flow and by Bmp signaling in a manner independent of Vegf signaling. We also found that during cardiac ballooning stages, endocardial cells obtain distinct chamber- and inner-versus-outer-curvature-specific surface area sizes.

Figure 3

Finally, we showed that the hemodynamic-sensitive transcription factor Krüppel-like factor 2a (Klf2a) is an important regulator of endocardial cell morphology. These findings establish the endocardium as the flow-sensitive tissue in the heart with a key role in adapting chamber growth in response to the mechanical stimulus of blood flow. Taken together, our findings are indicative of dynamic interactions between the myocardium and endocardium that constantly attune cellular sizes and shapes, and thus cardiac chamber dimensions in response to physiological changes, the most important of which is blood flow. Understanding this dynamic crosstalk during development may have implications for understanding how cardiac morphology changes later in life, in response to physiological adaptations or as pathophysiological conditions arise. Figure 3: Frontal view shows the two distinct endocardial chambers, atrium (A) and ventricle (V), during cardiac ballooning stages in a 48 hour old zebrafish embryo.The two endocardial chambers are visualized with the transgenic reporter lines [Tg(flt1:YFP), green; Tg(kdrl:mcherry), red].

Publications

2013 - ongoing

2018

 

Lisowska J, Rodel CJ, Manet S, Miroshnikova YA, Boyault C, Planus E, De Mets R, Lee HH, Destaing O, Mertani H, Boulday G, Tournier-Lasserve E, Balland M, Abdelilah-Seyfried S, Albiges-Rizo C, Faurobert E. Cerebral Cavernous Malformation 1/2 complex controls ROCK1 and ROCK2 complementary functions for endothelial integrity. J Cell Sci. 2018.

Paolini A, Abdelilah-Seyfried S. The mechanobiology of zebrafish cardiac valve leaflet formation. Curr Opin Cell Biol. 2018;55:52-8.

Donat S, Lourenço M, Paolini A, Otten C, Renz M, Abdelilah-Seyfried S. Heg1 and Ccm1/2 proteins control endocardial mechanosensitivity during zebrafish valvulogenesis. Elife. 2018. 7. pii: e28939.

Olmer R, Engels L, Usman A, Menke S, Malik MNH, Pessler F, Göhring G, Bornhorst D, Bolten S, Abdelilah-Seyfried S, Scheper T, Kempf H, Zweigerdt R, Martin U. Differentiation of Human Pluripotent Stem Cells into Functional Endothelial Cells in Scalable Suspension Culture. Stem Cell Reports. 2018. pii: S2213-6711(18)30144-9.

Merks A, Swinarski M, Meyer A, Müller N, Ozcan I, Donat S, Burger A, Gilbert S, Mosimann C, Abdelilah-Seyfried S, and Panakova D. Planar Cell Polarity signalling coordinates heart tube remodelling through tissue-scale polarisation of actomyosin activity. Nature communications. 2018. in press.

2016

Haack T, Abdelilah-Seyfried S. The force within: endocardial development, mechanotransduction and signalling during cardiac morphogenesis. Development. 2016;143(3):373-86.

Garcia de Vinuesa A, Abdelilah-Seyfried S, Knaus P, Zwijsen A, Bailly S. BMP signaling in vascular biology and dysfunction. Cytokine & growth factor reviews. 2016;27:65-79.

Cui H, Schlesinger J, Schoenhals S, Tonjes M, Dunkel I, Meierhofer D, Cano E, Schulz K, Berger MF, Haack T, Abdelilah-Seyfried S, Bulyk ML, Sauer S, Sperling SR. Phosphorylation of the chromatin remodeling factor DPF3a induces cardiac hypertrophy through releasing HEY repressors from DNA. Nucleic acids research. 2016;44(6):2538-53.

Dittmar F, Abdelilah-Seyfried S, Tschirner SK, Kaever V, Seifert R. Temporal and organ-specific detection of cNMPs including cUMP in the zebrafish. Biochemical and biophysical research communications. 2015;468(4):708-12.

Renz M, Otten C, Faurobert E, Rudolph F, Zhu Y, Boulday G, Duchene J, Mickoleit M, Dietrich AC, Ramspacher C, Steed E, Manet-Dupe S, Benz A, Hassel D, Vermot J, Huisken J, Tournier-Lasserve E, Felbor U, Sure U, Albiges-Rizo C, Abdelilah-Seyfried S. Regulation of Beta1 Integrin-Klf2-Mediated Angiogenesis by Ccm Proteins. Dev Cell. 2015;32(2):181-90.

Lombardo VA, Otten C, Abdelilah-Seyfried S. Large-Scale Zebrafish Embryonic Heart Dissection for Transcriptional Analysis. J Vis Exp. 2015(95):52087.

Dittmar F, Abdelilah-Seyfried S, Tschirner SK, Kaever V, Seifert R. Temporal and Organ-Specific Detection of Cnmps Including Cump in the Zebrafish. Biochem Biophys Res Commun. 2015. Epub 2015/11/10.

Cui H, Schlesinger J, Schoenhals S, Tonjes M, Dunkel I, Meierhofer D, Cano E, Schulz K, Berger MF, Haack T, Abdelilah-Seyfried S, Bulyk ML, Sauer S, Sperling SR. Phosphorylation of the Chromatin Remodeling Factor Dpf3a Induces Cardiac Hypertrophy through Releasing Hey Repressors from DNA. Nucleic Acids Res. 2015. Epub 2015/11/20.

2014

Dietrich AC, Lombardo VA, Veerkamp J, Priller F, Abdelilah-Seyfried S. Blood Flow and Bmp Signaling Control Endocardial Chamber Morphogenesis. Dev Cell. 2014;30(4):367-77.

Bennet M, Akiva A, Faivre D, Malkinson G, Yaniv K, Abdelilah-Seyfried S, Fratzl P, Masic A. Simultaneous Raman Microspectroscopy and Fluorescence Imaging of Bone Mineralization in Living Zebrafish Larvae. Biophys J. 2014;106(4):L17-9.

2013

Wieffer M, Cibrian Uhalte E, Posor Y, Otten C, Branz K, Schutz I, Mossinger J, Schu P, Abdelilah-Seyfried S, Krauss M, Haucke V. Pi4k2beta/Ap-1-Based Tgn-Endosomal Sorting Regulates Wnt Signaling. Curr Biol. 2013;23(21):2185-90.

Veerkamp J, Rudolph F, Cseresnyes Z, Priller F, Otten C, Renz M, Schaefer L, Abdelilah-Seyfried S. Unilateral Dampening of Bmp Activity by Nodal Generates Cardiac Left-Right Asymmetry. Dev Cell. 2013;24(6):660-7.

Tay HG, Schulze SK, Compagnon J, Foley FC, Heisenberg CP, Yost HJ, Abdelilah-Seyfried S, Amack JD. Lethal Giant Larvae 2 Regulates Development of the Ciliated Organ Kupffer's Vesicle. Development. 2013;140(7):1550-9.

Otten C, Abdelilah-Seyfried S. Laser-Inflicted Injury of Zebrafish Embryonic Skeletal Muscle. J Vis Exp. 2013(71):e4351.

2006 - 2012

2012

Zhang J, Liss M, Wolburg H, Blasig IE, Abdelilah-Seyfried S. Involvement of Claudins in Zebrafish Brain Ventricle Morphogenesis. Ann N Y Acad Sci. 2012;1257:193-8.

Otten C, van der Ven PF, Lewrenz I, Paul S, Steinhagen A, Busch-Nentwich E, Eichhorst J, Wiesner B, Stemple D, Strahle U, Furst DO, Abdelilah-Seyfried S. Xirp Proteins Mark Injured Skeletal Muscle in Zebrafish. PLoS One. 2012;7(2):e31041.

Lombardo VA, Sporbert A, Abdelilah-Seyfried S. Cell Tracking Using Photoconvertible Proteins During Zebrafish Development. J Vis Exp. 2012(67). Epub 2012/10/12.

de Pater E, Ciampricotti M, Priller F, Veerkamp J, Strate I, Smith K, Lagendijk AK, Schilling TF, Herzog W, Abdelilah-Seyfried S, Hammerschmidt M, Bakkers J. Bmp Signaling Exerts Opposite Effects on Cardiac Differentiation. Circ Res. 2012;110(4):578-87.

Cibrian Uhalte E, Kirchner M, Hellwig N, Allen JJ, Donat S, Shokat KM, Selbach M, Abdelilah-Seyfried S. In Vivo Conditions to Identify Prkci Phosphorylation Targets Using the Analog-Sensitive Kinase Method in Zebrafish. PLoS One. 2012;7(6):e40000. Epub 2012/07/07.

2011

Kur E, Christa A, Veth KN, Gajera CR, Andrade-Navarro MA, Zhang J, Willer JR, Gregg RG, Abdelilah-Seyfried S, Bachmann S, Link BA, Hammes A, Willnow TE. Loss of Lrp2 in Zebrafish Disrupts Pronephric Tubular Clearance but Not Forebrain Development. Dev Dyn. 2011;240(6):1567-77. Epub 2011/04/02.

Klein C, Mikutta J, Krueger J, Scholz K, Brinkmann J, Liu D, Veerkamp J, Siegel D, Abdelilah-Seyfried S, le Noble F. Neuron Navigator 3a Regulates Liver Organogenesis During Zebrafish Embryogenesis. Development. 2011;138(10):1935-45.

Ghani S, Riemke P, Schonheit J, Lenze D, Stumm J, Hoogenkamp M, Lagendijk A, Heinz S, Bonifer C, Bakkers J, Abdelilah-Seyfried S, Hummel M, Rosenbauer F. Macrophage Development from Hscs Requires Pu.1-Coordinated Microrna Expression. Blood. 2011;118(8):2275-84.

2010

Zhang J, Piontek J, Wolburg H, Piehl C, Liss M, Otten C, Christ A, Willnow TE, Blasig IE, Abdelilah-Seyfried S. Establishment of a Neuroepithelial Barrier by Claudin5a Is Essential for Zebrafish Brain Ventricular Lumen Expansion. Proc Natl Acad Sci U S A. 2010;107(4):1425-30. Epub 2010/01/19.

Abdelilah-Seyfried S. Claudin-5a in Developing Zebrafish Brain Barriers: Another Brick in the Wall. Bioessays. 2010;32(9):768-76. Epub 2010/07/24.

2009

Seipold S, Priller FC, Goldsmith P, Harris WA, Baier H, Abdelilah-Seyfried S. Non-Smc Condensin I Complex Proteins Control Chromosome Segregation and Survival of Proliferating Cells in the Zebrafish Neural Retina. BMC Dev Biol. 2009;9:40.

Nyholm MK, Abdelilah-Seyfried S, Grinblat Y. A Novel Genetic Mechanism Regulates Dorsolateral Hinge-Point Formation During Zebrafish Cranial Neurulation. J Cell Sci. 2009;122(Pt 12):2137-48.

Hava D, Forster U, Matsuda M, Cui S, Link BA, Eichhorst J, Wiesner B, Chitnis A, Abdelilah-Seyfried S. Apical Membrane Maturation and Cellular Rosette Formation During Morphogenesis of the Zebrafish Lateral Line. J Cell Sci. 2009;122(Pt 5):687-95.

Bakkers J, Verhoeven MC, Abdelilah-Seyfried S. Shaping the Zebrafish Heart: From Left-Right Axis Specification to Epithelial Tissue Morphogenesis. Dev Biol. 2009;330(2):213-20.

2008

Rohr S, Otten C, Abdelilah-Seyfried S. Asymmetric Involution of the Myocardial Field Drives Heart Tube Formation in Zebrafish. Circ Res. 2008;102(2):e12-9.

Munson C, Huisken J, Bit-Avragim N, Kuo T, Dong PD, Ober EA, Verkade H, Abdelilah-Seyfried S, Stainier DY. Regulation of Neurocoel Morphogenesis by Pard6 Gamma B. Dev Biol. 2008;324(1):41-54.

Lange M, Kaynak B, Forster UB, Tonjes M, Fischer JJ, Grimm C, Schlesinger J, Just S, Dunkel I, Krueger T, Mebus S, Lehrach H, Lurz R, Gobom J, Rottbauer W, Abdelilah-Seyfried S, Sperling S. Regulation of Muscle Development by Dpf3, a Novel Histone Acetylation and Methylation Reader of the Baf Chromatin Remodeling Complex. Genes Dev. 2008;22(17):2370-84.

Bit-Avragim N, Rohr S, Rudolph F, Van der Ven P, Furst D, Eichhorst J, Wiesner B, Abdelilah-Seyfried S. Nuclear Localization of the Zebrafish Tight Junction Protein Nagie Oko. Dev Dyn. 2008;237(1):83-90.

Bit-Avragim N, Hellwig N, Rudolph F, Munson C, Stainier DY, Abdelilah-Seyfried S. Divergent Polarization Mechanisms During Vertebrate Epithelial Development Mediated by the Crumbs Complex Protein Nagie Oko. J Cell Sci. 2008;121(Pt 15):2503-10.

2007

Cui S, Otten C, Rohr S, Abdelilah-Seyfried S, Link BA. Analysis of Apkclambda and Apkczeta Reveals Multiple and Redundant Functions During Vertebrate Retinogenesis. Mol Cell Neurosci. 2007;34(3):431-44.

Cibrian-Uhalte E, Langenbacher A, Shu X, Chen JN, Abdelilah-Seyfried S. Involvement of Zebrafish Na+,K+ Atpase in Myocardial Cell Junction Maintenance. J Cell Biol. 2007;176(2):223-30. Epub 2007/01/18.

2006

Anzenberger U, Bit-Avragim N, Rohr S, Rudolph F, Dehmel B, Willnow TE, Abdelilah-Seyfried S. Elucidation of Megalin/Lrp2-Dependent Endocytic Transport Processes in the Larval Zebrafish Pronephros. J Cell Sci. 2006;119(Pt 10):2127-37.

Rohr S, Bit-Avragim N, Abdelilah-Seyfried S. Heart and Soul/Prkci and Nagie Oko/Mpp5 Regulate Myocardial Coherence and Remodeling During Cardiac Morphogenesis. Development. 2006;133(1):107-15.

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