title: Visualizing the cell-cycle progression of endothelial cells in zebrafish.
abstract: The formation of vascular structures requires precisely controlled proliferation of endothelial cells (ECs), which occurs through strict regulation of the cell cycle. However, the mechanism by which EC proliferation is coordinated during vascular formation remains largely unknown, since a method of analyzing cell-cycle progression of ECs in living animals has been lacking. Thus, we devised a novel system allowing the cell-cycle progression of ECs to be visualized in vivo. To achieve this aim, we generated a transgenic zebrafish line that expresses zFucci (zebrafish fluorescent ubiquitination-based cell cycle indicator) specifically in ECs (an EC-zFucci Tg line). We first assessed whether this system works by labeling the S phase ECs with EdU, then performing time-lapse imaging analyses and, finally, examining the effects of cell-cycle inhibitors. Employing the EC-zFucci Tg line, we analyzed the cell-cycle progression of ECs during vascular development in different regions and at different time points and found that ECs proliferate actively in the developing vasculature. The proliferation of ECs also contributes to the elongation of newly formed blood vessels. While ECs divide during elongation in intersegmental vessels, ECs proliferate in the primordial hindbrain channel to serve as an EC reservoir and migrate into basilar and central arteries, thereby contributing to new blood vessel formation. Furthermore, while EC proliferation is not essential for the formation of the basic framework structures of intersegmental and caudal vessels, it appears to be required for full maturation of these vessels. In addition, venous ECs mainly proliferate in the late stage of vascular development, whereas arterial ECs become quiescent at this stage. Thus, we anticipate that the EC-zFucci Tg line can serve as a tool for detailed studies of the proliferation of ECs in various forms of vascular development in vivo.
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan. Electronic address: fuku@ncvc.go.jp.
authors: Shigetomo
authors: S
authors: Fukuhara
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan.
authors: Jianghui
authors: J
authors: Zhang
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan.
authors: Shinya
authors: S
authors: Yuge
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan.
authors: Koji
authors: K
authors: Ando
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan.
authors: Yuki
authors: Y
authors: Wakayama
authors: Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako City, Saitama 351-0198, Japan; Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa, Wako City, Saitama 351-0198, Japan.
authors: Asako
authors: A
authors: Sakaue-Sawano
authors: Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako City, Saitama 351-0198, Japan; Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa, Wako City, Saitama 351-0198, Japan.
authors: Atsushi
authors: A
authors: Miyawaki
authors: Department of Cell Biology, National Cerebral and Cardiovascular Center Research Institute, Fujishirodai 5-7-1, Suita, Osaka 565-8565, Japan; JST-CREST, Tokyo, Japan. Electronic address: nmochizu@ri.ncvc.go.jp.
authors: Naoki
authors: N
authors: Mochizuki
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified / Q000196:embryology / Q000235:genetics; D002453:Cell Cycle / Q000235:genetics* / Q000502:physiology; D002455:Cell Division; D049109:Cell Proliferation; D050760:Cyclin-Dependent Kinase Inhibitor p27 / Q000235:genetics; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D055785:Gene Knockdown Techniques; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics; D008565:Membrane Proteins / Q000235:genetics; D060172:Morpholinos / Q000235:genetics; D018919:Neovascularization, Physiologic / Q000235:genetics*; D015027:Zebrafish / Q000235:genetics / Q000502:physiology*; D029961:Zebrafish Proteins / Q000096:biosynthesis
chemical_list: D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D060172:Morpholinos; D029961:Zebrafish Proteins; C064045:delta protein; D050760:Cyclin-Dependent Kinase Inhibitor p27
keywords: Angiogenesis; Cell cycle; Endothelial cell; Proliferation; Vascular development