Adv Pharm Bull. 2014;4(4):323-328.
doi: 10.5681/apb.2014.047
PMID: 25436186
PMCID: PMC4137420
Scopus id: 84906258059
  Abstract View: 656
  PDF Download: 780

Original Research

Expression and Purification of Functional Human Vascular Endothelial Growth Factor-A121; the Most Important Angiogenesis Factor

Fatemeh kazemi-Lomedasht 1, Mahdi Behdani 1 * , Kamran Pooshang Bagheri 1, Mahdi Habibi Anbouhi 2, Mohsen Abolhassani 3, Hossein Khanahmad 1, Delavar Shahbazzadeh 1, Hassan Mirzahoseini 1

1 Biotechnology Research Center, Venom & Biotherapeutics Molecules Lab., Pasteur Institute of Iran, Tehran, Iran.
2 National Cell Bank of Iran, Pasteur Institute of Iran, Tehran, Iran.
3 Departmentof Immunology, Hybridoma Lab, Pasteur Institute of Iran, Tehran, Iran.

Abstract

Purpose: Angiogenesis or formation of new blood vessels is an essential process for tumor growth, invasion and metastasis. Vascular Endothelial Growth Factor (VEGF) and its receptors play an important role in angiogenesis-dependent tumors. VEGF-A is the most important factor in angiogenesis process. Human VEGF-A gene consists of eight exons that undergoes alternative exon splicing and produce five different proteins consisting of 121, 145, 165, 189 and 206 amino acids (named VEGF121, VEGF145, VEGF165, VEGF189, and VEGF206). Methods: In this study, VEGF121 gene synthesized and cloned into the pET-26b plasmid. The recombinant plasmid was transferred into appropriate expression strain of BL-21. Expression of VEGF121 induced by IPTG (Isopropyl β-D-1-thiogalactopyranoside) and confirmed by SDS-PAGE and Western-Blotting. Recombinant VEGF121 was purified by nickel affinity chromatography. HUVECs (Human Umbilical Vein Endothelia Cells) cells were isolated from umbilical vein and the effect of VEGF121 on tube formation of endothelial cells was investigated. Results: SDS-PAGE and Western-Blotting results verified the purification of VEGF121. The final yield of recombinant protein was about 5mg per liter. Endothelial cell tube formation assay results showed that VEGF121 leads to tube formation of endothelial cell on matrix and induces angiogenesis in vitro. Conclusion: Recombinant VEGF121 is important factor in tube formation of endothelial cell, so it could be used in different cancer researches and angiogenesis assay.
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