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Updated: Jan 3, 2026
In Vitro Three-Dimensional Sprouting Assay of Angiogenesis Using Mouse Embryonic Stem Cells for Vascular Disease Modeling and Drug Testing
Published on: May 11, 2021
[Induced differentiation of mouse embryonic stem cell into endothelial cell in vitro]
Jixin Xiong1, Zhaoxuan Liu, Xiaochun Liu
1Department of Vascular Surgery, Second Affiliated Hospital of Nanchang University, Nanchang Jiangxi, 330006, PR China. liuzhaoxuan1981@163.com
Objective:
To explore an optional condition to induce mouse embryonic stem cell (ESC) to differentiate into endothelial cells so as to provide seed cells for tissue engineered vascular.
Methods:
The embryos from one pregnant 12.5-days mouse was harvested to culture the mouse embryonic fibroblasts (MEF). The ESC was reanimated by common method, and used to cultured into embryoid body (EB) in vitro. The EB which was used to induce into endothelial cells was divided into two groups. The EB was cultured in the EB medium with 3 ng/ml transforming growth factor beta1, 50 ng/ml vascular endothelial cell growth factor and 1 micromol/L potent and selective inhibitor of activin receptor-like kinase receptors in experimental group. The EB was cultured in the EB medium in the control group. After 14 days, RT-PCR and immunohistochemistry were used to detect vWF and CD34, to analyze the morphology and type of the differentiated cells from ESC.
Results:
The primary MEF had a high proliferation activity. At the 3rd day, the fusion rate of MEF was about 90% with a fusiform shape. The cells was fusiform shape and arranged compactly with fullness of nucleus and 2-3 entoblasts. The 3rd-5th generations EB was polygonal with fullness of cytoplasm and 3-4 entoblasts. ESC could maintain undifferentiated state, and the cells unit looked like bird nest with smooth margin; the cells was small at size and strong refractivity with high rate of nuclein and rapid proliferation. At 3 days of drop-culture, EB can seen grossly and at 3 days of suspension, large and transparent EB formed. EB was spread radiately with an intensive adhesion at the 2nd day. In experimental group, many round cells was differentiated around EB from the 4th day to the 7th day, and form tubular structures from the 10th day to the 14th day. The vWF and CD34 were expressed. In control group, EB could not form tubular structures, and the vWF and CD34 were not expressed.
Conclusion:
ESC can differentiate into endothelial cells under some conditions, and form vessel-like structure under condition culture, which can provide sources of seed cells for tissue engineered vessel.
