@article {72, title = {Wnt signaling is required for early development of zebrafish swimbladder.}, journal = {PLoS One}, volume = {6}, year = {2011}, month = {2011}, pages = {e18431}, abstract = {

BACKGROUND: Wnt signaling plays critical roles in mammalian lung development. However, Wnt signaling in the development of the zebrafish swimbladder, which is considered as a counterpart of mammalian lungs, have not been explored. To investigate the potential conservation of signaling events in early development of the lung and swimbladder, we wish to address the question whether Wnt signaling plays a role in swimbladder development.

METHODOLOGY/PRINCIPAL FINDINGS: For analysis of zebrafish swimbladder development, we first identified, by whole-mount in situ hybridization (WISH), has2 as a mesenchymal marker, sox2 as the earliest epithelial marker, as well as hprt1l and elovl1a as the earliest mesothelial markers. We also demonstrated that genes encoding Wnt signaling members Wnt5b, Fz2, Fz7b, Lef1, Tcf3 were expressed in different layers of swimbladder. Then we utilized the heat-shock inducible transgenic lines hs:Dkk1-GFP and hs:ΔTcf-GFP to temporarily block canonical Wnt signaling. Inhibition of canonical Wnt signaling at various time points disturbed precursor cells specification, organization, anterioposterior patterning, and smooth muscle differentiation in all three tissue layers of swimbladder. These observations were also confirmed by using a chemical inhibitor (IWR-1) of Wnt signaling. In addition, we found that Hedgehog (Hh) signaling was activated by canonical Wnt signaling and imposed a negative feedback on the latter.

SIGNIFICANCE/CONCLUSION: We first provided a new set of gene markers for the three tissue layers of swimbladder in zebrafish and demonstrated the expression of several key genes of Wnt signaling pathway in developing swimbladder. Our functional analysis data indicated that Wnt/β-catenin signaling is required for swimbladder early development and we also provided evidence for the crosstalk between Wnt and Hh signaling in early swimbladder development.

}, keywords = {Air Sacs, Animals, Animals, Genetically Modified, Apoptosis, Cell Differentiation, Cell Proliferation, Embryo, Nonmammalian, Epithelium, Gene Expression Regulation, Developmental, Gene Knockdown Techniques, Genetic Markers, Green Fluorescent Proteins, Heat-Shock Response, Hedgehog Proteins, Mesoderm, Models, Biological, Morphogenesis, Myocytes, Smooth Muscle, Recombinant Fusion Proteins, Reproducibility of Results, Signal Transduction, Wnt Proteins, Zebrafish, Zebrafish Proteins}, issn = {1932-6203}, doi = {10.1371/journal.pone.0018431}, author = {Yin, Ao and Korzh, Svitlana and Winata, Cecilia L and Korzh, Vladimir and Gong, Zhiyuan} } @article {68, title = {The role of vasculature and blood circulation in zebrafish swimbladder development.}, journal = {BMC Dev Biol}, volume = {10}, year = {2010}, month = {2010}, pages = {3}, abstract = {

BACKGROUND: Recently we have performed a detailed analysis of early development of zebrafish swimbladder, a homologous organ of tetrapod lung; however, the events of swimbladder development are still poorly characterized. Many studies have implicated the role of vascular system in development of many organs in vertebrates. As the swimbladder is lined with an intricate network of blood capillaries, it is of interest to investigate the role of the vascular system during early development of swimbladder.

RESULTS: To investigate the role of endothelial cells (ECs) and blood circulation during development of the swimbladder, phenotypes of swimbladder were analysed at three different stages (approximately 2, 3 and 5 dpf [day postfertilization]) in cloche (clo) mutant and Tnnt2 morphants, in the background of transgenic lines Et(krt4:EGFP)sq33-2 and Et(krt4:EGFP)sqet3 which express EGFP in the swimbladder epithelium and outer mesothelium respectively. Analyses of the three tissue layers of the swimbladder were performed using molecular markers hb9, fgf10a, acta2, and anxa5 to distinguish epithelium, mesenchyme, and outer mesothelium. We showed that the budding stage was independent of ECs and blood flow, while early epithelial growth, mesenchymal organization and its differentiation into smooth muscle, as well as outer mesothelial organization, were dependent on ECs. Blood circulation contributed to later stage of epithelial growth, smooth muscle differentiation, and organization of the outer mesothelium. Inflation of the swimbladder was also affected as a result of absence of ECs and blood flow.

CONCLUSION: Our data demonstrated that the vascular system, though not essential in swimbladder budding, plays an important role in the development of the swimbladder starting from the early growth stage, including mesenchyme organization and smooth muscle differentiation, and outer mesothelial organization, which in turn may be essential for the function of the swimbladder as reflected in its eventual inflation.

}, keywords = {Air Sacs, Animals, Animals, Genetically Modified, Blood Circulation, Endothelial Cells, Green Fluorescent Proteins, Troponin T, Zebrafish, Zebrafish Proteins}, issn = {1471-213X}, doi = {10.1186/1471-213X-10-3}, author = {Winata, Cecilia L and Korzh, Svetlana and Kondrychyn, Igor and Korzh, Vladimir and Gong, Zhiyuan} } @article {67, title = {Development of zebrafish swimbladder: The requirement of Hedgehog signaling in specification and organization of the three tissue layers.}, journal = {Dev Biol}, volume = {331}, year = {2009}, month = {2009 Jul 15}, pages = {222-36}, abstract = {

The swimbladder is a hydrostatic organ in fish postulated as a homolog of the tetrapod lung. While lung development has been well studied, the molecular mechanism of swimbladder development is essentially uncharacterized. In the present study, swimbladder development in zebrafish was analyzed by using several molecular markers: hb9 (epithelium), fgf10a and acta2 (mesenchyme), and anxa5 (mesothelium), as well as in vivo through enhancer trap transgenic lines Et(krt4:EGFP)(sq33-2) and Et(krt4:EGFP)(sqet3) that showed strong EGFP expression in the swimbladder epithelium and outer mesothelium respectively. We defined three phases of swimbladder development: epithelial budding between 36 and 48 hpf, growth with the formation of two additional mesodermal layers up to 4.5 dpf, and inflation of posterior and anterior chambers at 4.5 and 21 dpf respectively. Similar to those in early lung development, conserved expression of Hedgehog (Hh) genes, shha and ihha, in the epithelia, and Hh receptor genes, ptc1 and ptc2, as well as fgf10a in mesenchyme was observed. By analyzing several mutants affecting Hh signaling and Ihha morphants, we demonstrated an essential role of Hh signaling in swimbladder development. Furthermore, time-specific Hh inhibition by cyclopamine revealed different requirements of Hh signaling in the formation and organization of all three tissue layers of swimbladder.

}, keywords = {Air Sacs, Animals, Antigens, Differentiation, Body Patterning, Embryo, Nonmammalian, Hedgehog Proteins, Mutation, Signal Transduction, Zebrafish, Zebrafish Proteins}, issn = {1095-564X}, doi = {10.1016/j.ydbio.2009.04.035}, author = {Winata, Cecilia L and Korzh, Svetlana and Kondrychyn, Igor and Zheng, Weiling and Korzh, Vladimir and Gong, Zhiyuan} }