PUBLICATION
The growth of endothelial-like cells in zebrafish embryoid body culture
- Authors
- Ibrahim, M., Xie, B., Richardson, M.K.
- ID
- ZDB-PUB-200501-6
- Date
- 2020
- Source
- Experimental cell research 392(2): 112032 (Journal)
- Registered Authors
- Richardson, Michael
- Keywords
- Blastocyst cells, Embryoid body, Endothelial differentiation, fli:GFP, kdrl:GFP
- MeSH Terms
-
- Animals
- Animals, Genetically Modified/embryology*
- Animals, Genetically Modified/physiology
- Blastocyst/cytology
- Blastocyst/drug effects
- Blastocyst/metabolism
- Cell Differentiation*
- Cells, Cultured
- Culture Media/pharmacology
- Embryo, Nonmammalian/cytology*
- Embryo, Nonmammalian/drug effects
- Embryo, Nonmammalian/metabolism
- Embryoid Bodies/cytology*
- Embryoid Bodies/drug effects
- Embryoid Bodies/metabolism
- Embryonic Stem Cells/cytology*
- Embryonic Stem Cells/drug effects
- Embryonic Stem Cells/metabolism
- Endothelial Cells/cytology*
- Endothelial Cells/drug effects
- Endothelial Cells/metabolism
- Green Fluorescent Proteins/metabolism
- Intercellular Signaling Peptides and Proteins/pharmacology
- Transcriptome
- Zebrafish/embryology*
- Zebrafish/physiology
- PubMed
- 32353375 Full text @ Exp. Cell Res.
Citation
Ibrahim, M., Xie, B., Richardson, M.K. (2020) The growth of endothelial-like cells in zebrafish embryoid body culture. Experimental cell research. 392(2):112032.
Abstract
There is increasing interest in the possibility of culturing organ-like tissues (organoids) in vitro for biomedical applications. The ability to culture organoids would be greatly enhanced by having a functional circulation in vitro. The endothelial cell is the most important cell type in this context. Endothelial cells can be derived from pluripotent embryonic blastocyst cells in aggregates called embryoid bodies. Here, we examine the yield of endothelial-like cells in embryoid bodies (EBs) developed from transgenic zebrafish fli:GFP and kdrl:GFP blastocyst embryos. The isolated blastocyst cells developed into EBs within the first 24 h of culture and contained fli:GFP+ (putative endothelial, hematopoietic and other cell types); or kdrl:GFP+ (endothelial) cells. The addition of endothelial growth supplements to the media and culture on collagen type-I substratum increased the percentages of fli:GFP+ and kdrl:GFP+ cells in culture. We found that EBs developed in hanging-drop cultures possessed a higher percentage of fli:GFP+ (45.0 ± 3.1%) and kdrl:GFP+ cells (8.7 ± 0.7%) than those developed on conventional substrata (34.5 ± 1.4% or 5.2 ± 0.4%, respectively). The transcriptome analysis showed a higher expression of VEGF and TGFβ genes in EB cultures compared to the adherent cultures. When transferred to conventional culture, the percentage of fli:GFP+ or kdrl:GFP+ cells declined significantly over subsequent days in the EBs. The fli:GFP+ cells formed a monolayer around the embryoid bodies, while the kdrl:GFP+ cells formed vascular network-like structures in the embryoid bodies. Differences were observed in the spreading of fli:GFP+ cells and network formation of kdrl:GFP+ cells on different substrates. The fli:GFP+ cells could be maintained in primary culture and sub-cultures. By contrast, kdrl:GFP+ cells were almost completely absent at 8d of primary culture. Our culture model allows real-time observation of fli:GFP+ and kdrl:GFP+ cells in culture. The results obtained from this study will be important for the development of vascular and endothelial cell culture using embryonic cells.
Genes / Markers
Expression
Phenotype
Mutations / Transgenics
Human Disease / Model
Sequence Targeting Reagents
Fish
Orthology
Engineered Foreign Genes
Mapping