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1.
Development ; 150(7)2023 04 01.
Article in English | MEDLINE | ID: mdl-36938965

ABSTRACT

Blood vessels form elaborate networks that depend on tissue-specific signalling pathways and anatomical structures to guide their growth. However, it is not clear which morphogenetic principles organize the stepwise assembly of the vasculature. We therefore performed a longitudinal analysis of zebrafish caudal fin vascular assembly, revealing the existence of temporally and spatially distinct morphogenetic processes. Initially, vein-derived endothelial cells (ECs) generated arteries in a reiterative process requiring vascular endothelial growth factor (Vegf), Notch and cxcr4a signalling. Subsequently, veins produced veins in more proximal fin regions, transforming pre-existing artery-vein loops into a three-vessel pattern consisting of an artery and two veins. A distinct set of vascular plexuses formed at the base of the fin. They differed in their diameter, flow magnitude and marker gene expression. At later stages, intussusceptive angiogenesis occurred from veins in distal fin regions. In proximal fin regions, we observed new vein sprouts crossing the inter-ray tissue through sprouting angiogenesis. Together, our results reveal a surprising diversity among the mechanisms generating the mature fin vasculature and suggest that these might be driven by separate local cues.


Subject(s)
Endothelial Cells , Zebrafish , Animals , Zebrafish/genetics , Vascular Endothelial Growth Factor A/metabolism , Neovascularization, Physiologic , Veins/metabolism
2.
Sci Rep ; 12(1): 4795, 2022 03 21.
Article in English | MEDLINE | ID: mdl-35314737

ABSTRACT

Endothelial cells (ECs) lining blood vessels are exposed to mechanical forces, such as shear stress. These forces control many aspects of EC biology, including vascular tone, cell migration and proliferation. Despite a good understanding of the genes responding to shear stress, our insight into the transcriptional regulation of these genes is much more limited. Here, we set out to study alterations in the chromatin landscape of human umbilical vein endothelial cells (HUVEC) exposed to laminar shear stress. To do so, we performed ChIP-Seq for H3K27 acetylation, indicative of active enhancer elements and ATAC-Seq to mark regions of open chromatin in addition to RNA-Seq on HUVEC exposed to 6 h of laminar shear stress. Our results show a correlation of gained and lost enhancers with up and downregulated genes, respectively. DNA motif analysis revealed an over-representation of KLF transcription factor (TF) binding sites in gained enhancers, while lost enhancers contained more ETV/ETS motifs. We validated a subset of flow responsive enhancers using luciferase-based reporter constructs and CRISPR-Cas9 mediated genome editing. Lastly, we characterized the shear stress response in ECs of zebrafish embryos using RNA-Seq. Our results lay the groundwork for the exploration of shear stress responsive elements in controlling EC biology.


Subject(s)
Chromatin , Zebrafish , Animals , Binding Sites , Cells, Cultured , Chromatin/metabolism , Human Umbilical Vein Endothelial Cells/metabolism , Humans , Stress, Mechanical , Transcription Factors/genetics , Transcription Factors/metabolism , Zebrafish/genetics , Zebrafish/metabolism
3.
Development ; 149(7)2022 04 01.
Article in English | MEDLINE | ID: mdl-35297968

ABSTRACT

Vascular networks comprise endothelial cells and mural cells, which include pericytes and smooth muscle cells. To elucidate the mechanisms controlling mural cell recruitment during development and tissue regeneration, we studied zebrafish caudal fin arteries. Mural cells colonizing arteries proximal to the body wrapped around them, whereas those in more distal regions extended protrusions along the proximo-distal vascular axis. Both cell populations expressed platelet-derived growth factor receptor ß (pdgfrb) and the smooth muscle cell marker myosin heavy chain 11a (myh11a). Most wrapping cells in proximal locations additionally expressed actin alpha2, smooth muscle (acta2). Loss of Pdgfrb signalling specifically decreased mural cell numbers at the vascular front. Using lineage tracing, we demonstrate that precursor cells located in periarterial regions and expressing Pgdfrb can give rise to mural cells. Studying tissue regeneration, we did not find evidence that newly formed mural cells were derived from pre-existing cells. Together, our findings reveal conserved roles for Pdgfrb signalling in development and regeneration, and suggest a limited capacity of mural cells to self-renew or contribute to other cell types during tissue regeneration.


Subject(s)
Myocytes, Smooth Muscle , Pericytes , Receptor, Platelet-Derived Growth Factor beta , Zebrafish Proteins , Zebrafish , Animals , Endothelial Cells/metabolism , Myocytes, Smooth Muscle/metabolism , Pericytes/metabolism , Receptor, Platelet-Derived Growth Factor beta/genetics , Receptor, Platelet-Derived Growth Factor beta/metabolism , Zebrafish/metabolism , Zebrafish Proteins/genetics , Zebrafish Proteins/metabolism
4.
EMBO J ; 40(15): e107134, 2021 08 02.
Article in English | MEDLINE | ID: mdl-34180064

ABSTRACT

Long non-coding RNAs (lncRNAs) are emerging as key regulators of endothelial cell function. Here, we investigated the role of a novel vascular endothelial-associated lncRNA (VEAL2) in regulating endothelial permeability. Precise editing of veal2 loci in zebrafish (veal2gib005Δ8/+ ) induced cranial hemorrhage. In vitro and in vivo studies revealed that veal2 competes with diacylglycerol for interaction with protein kinase C beta-b (Prkcbb) and regulates its kinase activity. Using PRKCB2 as bait, we identified functional ortholog of veal2 in humans from HUVECs and named it as VEAL2. Overexpression and knockdown of VEAL2 affected tubulogenesis and permeability in HUVECs. VEAL2 was differentially expressed in choroid tissue in eye and blood from patients with diabetic retinopathy, a disease where PRKCB2 is known to be hyperactivated. Further, VEAL2 could rescue the effects of PRKCB2-mediated turnover of endothelial junctional proteins thus reducing hyperpermeability in hyperglycemic HUVEC model of diabetic retinopathy. Based on evidence from zebrafish and hyperglycemic HUVEC models and diabetic retinopathy patients, we report a hitherto unknown VEAL2 lncRNA-mediated regulation of PRKCB2, for modulating junctional dynamics and maintenance of endothelial permeability.


Subject(s)
Diabetic Retinopathy/genetics , Protein Kinase C beta/genetics , RNA, Long Noncoding/genetics , Zebrafish/genetics , Aged , Aged, 80 and over , Animals , Animals, Genetically Modified , Case-Control Studies , Diabetic Retinopathy/physiopathology , Embryo, Nonmammalian , Endothelium, Vascular , Gene Expression Regulation , Human Umbilical Vein Endothelial Cells , Humans , Middle Aged , Permeability , Protein Kinase C beta/metabolism , RNA, Long Noncoding/blood , Zebrafish/embryology , Zebrafish Proteins/genetics , Zebrafish Proteins/metabolism
5.
Nat Cell Biol ; 19(6): 653-665, 2017 Jun.
Article in English | MEDLINE | ID: mdl-28530658

ABSTRACT

The hierarchical organization of properly sized blood vessels ensures the correct distribution of blood to all organs of the body, and is controlled via haemodynamic cues. In current concepts, an endothelium-dependent shear stress set point causes blood vessel enlargement in response to higher flow rates, while lower flow would lead to blood vessel narrowing, thereby establishing homeostasis. We show that during zebrafish embryonic development increases in flow, after an initial expansion of blood vessel diameters, eventually lead to vessel contraction. This is mediated via endothelial cell shape changes. We identify the transforming growth factor beta co-receptor endoglin as an important player in this process. Endoglin mutant cells and blood vessels continue to enlarge in response to flow increases, thus exacerbating pre-existing embryonic arterial-venous shunts. Together, our data suggest that cell shape changes in response to biophysical cues act as an underlying principle allowing for the ordered patterning of tubular organs.


Subject(s)
Cell Shape , Endoglin/metabolism , Endothelial Cells/metabolism , Hemodynamics , Mechanotransduction, Cellular , Zebrafish Proteins/metabolism , Animals , Arteriovenous Malformations/genetics , Arteriovenous Malformations/metabolism , Arteriovenous Malformations/physiopathology , Endoglin/deficiency , Endoglin/genetics , Genetic Predisposition to Disease , Human Umbilical Vein Endothelial Cells/metabolism , Humans , Kruppel-Like Transcription Factors/genetics , Kruppel-Like Transcription Factors/metabolism , Mice, Knockout , Mutation , Neovascularization, Physiologic , Phenotype , Regional Blood Flow , Stress, Mechanical , Time Factors , Zebrafish/embryology , Zebrafish/genetics , Zebrafish/metabolism , Zebrafish Proteins/genetics
6.
Database (Oxford) ; 2014: bau011, 2014.
Article in English | MEDLINE | ID: mdl-24578356

ABSTRACT

A large repertoire of gene-centric data has been generated in the field of zebrafish biology. Although the bulk of these data are available in the public domain, most of them are not readily accessible or available in nonstandard formats. One major challenge is to unify and integrate these widely scattered data sources. We tested the hypothesis that active community participation could be a viable option to address this challenge. We present here our approach to create standards for assimilation and sharing of information and a system of open standards for database intercommunication. We have attempted to address this challenge by creating a community-centric solution for zebrafish gene annotation. The Zebrafish GenomeWiki is a 'wiki'-based resource, which aims to provide an altruistic shared environment for collective annotation of the zebrafish genes. The Zebrafish GenomeWiki has features that enable users to comment, annotate, edit and rate this gene-centric information. The credits for contributions can be tracked through a transparent microattribution system. In contrast to other wikis, the Zebrafish GenomeWiki is a 'structured wiki' or rather a 'semantic wiki'. The Zebrafish GenomeWiki implements a semantically linked data structure, which in the future would be amenable to semantic search. Database URL: http://genome.igib.res.in/twiki.


Subject(s)
Crowdsourcing/methods , Genome/genetics , Internet , Molecular Sequence Annotation/methods , Zebrafish/genetics , Animals , Databases, Genetic
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