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Updated: Aug 5, 2026
A Simple Bioassay for the Evaluation of Vascular Endothelial Growth Factors
Published on: March 15, 2016
[Cloning, expression of soluble VEGFR2 fragment and its effect on tumor angiogenesis]
Bai-jun Kou1, Yu-lin Li, Li-hong Zhang
1Laboratory of Pathobiology, College of Basic Medicine, Jilin University 130021, China. koubaijun@yahoo.com
Objective:
To study the anti-tumor angiogenesis effect of soluble VEGF receptor fragment by blocking the combination of VEGF and its receptor in vivo and in vitro.
Methods:
RT-PCR technique was used to amplify Flk-1/KDR fragment from embryo mouse liver, which was recombinated to expression vector pET-28b(+) and retrovirus vector PLXSN, which was induced to be expressed, purified and identified with EcoR I and Hind III. Mouse endothelial cells were separated, cultured and identified by immunocytochemistrical staining using VIII factor-related antigen antibody. The expressed product was analyzed about its effect on endothelial cell's growth in vitro with MTT method. The retrovirus vector was transfected to tumor cell lines S180 and B16 by liposome method to observe the biological specificity in vitro after gene transfection.
Results:
1000 bp size sVEGFR fragment was amplify from E9, E11 embryo mouse liver tissues, which was recombinated to TA clone vector and identified by sequence analysis. This fragment was cloned to expression vector pET-28b(+), the expressed product was purified and identified correctly. The in vitro study showed this expressed product can effectively inhibit endothelial cell(s), growth and proliferation. The fragment was then cloned to retrovirus vector PLXSN and transfected to tumor cell lines S180 and B16 successfully with RT-PCR and SDS-PAGE. The experiments in vivo showed that the weight of tumor smaller, the size decreased significantly, the microvessel density was fewer and Flk1 protein expression were higher in the group of gene transfection than that of control.
Conclusion:
Soluble VEGFR fragment is a kind of effective gene engineer product for anti-tumor angiogenesis gene therapy and the development of anti-tumor drug.
Insights
This study demonstrates that a soluble VEGF receptor fragment effectively inhibits tumor angiogenesis by blocking VEGF signaling. This gene therapy approach shows promise for developing new anti-cancer drugs.
Area of Science:
- Molecular Biology
- Oncology
- Gene Therapy
Context:
- Tumor angiogenesis, the formation of new blood vessels that supply tumors, is crucial for cancer growth and metastasis.
- Vascular Endothelial Growth Factor (VEGF) and its receptor (VEGFR) play a critical role in regulating angiogenesis.
- Targeting the VEGF-VEGFR pathway is a validated strategy for anti-cancer therapy.
Purpose:
- To investigate the anti-tumor angiogenesis effect of a soluble VEGF receptor fragment.
- To block the interaction between VEGF and its receptor in vivo and in vitro.
- To assess the therapeutic potential of this fragment in gene therapy for cancer.
Summary:
- A soluble VEGFR fragment (sVEGFR) was successfully amplified, cloned, and expressed.
- In vitro studies confirmed that sVEGFR inhibits endothelial cell growth and proliferation.
- In vivo experiments showed that gene transfection with sVEGFR reduced tumor size, weight, and microvessel density, while increasing Flk1 expression.
Impact:
- The study validates sVEGFR as an effective agent for anti-tumor angiogenesis.
- This research supports the development of novel gene-engineered products for cancer therapy.
- The findings contribute to the advancement of anti-cancer drug development targeting angiogenesis.
