Fibrinogen stimulates in vitro angiogenesis by choroidal endothelial cells via autocrine VEGF

Satomi Shiose, Yasuaki Hata, Yoshihiro Noda, Yukio Sassa, Atsunobu Takeda, Hiroshi Yoshikawa, Kimihiko Fujisawa, Toshiaki Kubota, Tatsuro Ishibashi

Research output: Contribution to journalArticlepeer-review

33 Citations (Scopus)


Background: The purpose of this study is to investigate the effect of fibrinogen on angiogenesis in vitro formed by cultured bovine choroidal endothelial cells (BCECs) and the involvement of vascular endothelial growth factor (VEGF) in this mechanism. Methods: For in vitro tube formation assay, BCECs were seeded on collagen gel containing fibrinogen (0-1.5 mg/ml). After 3 days of cultivation, the total length of the tubular structure was measured using Macscope Analyzer. Total RNA and conditioned media were collected after fibrinogen treatment and subjected to Northern and Western blot analyses, respectively. Transcription factor HIF-1α was also analyzed by Western blot analysis using cytosolic and nuclear fraction of BCECs. Involvement of VEGF in fibrinogen-dependent in vitro tube formation was evaluated using anti-VEGF neutralizing antibody or VEGF receptor 2-selective inhibitor (SU5416). Results: Formation of the tubular structure was enhanced 20-50 times in fibrinogen-containing gel in a concentration-dependent manner. The treatment of BCECs with fibrinogen resulted in a significant increase in VEGF gene and protein expression. Accumulation of HIF-1α protein in the nuclear fraction was also detected after the treatment with fibrinogen. Finally, fibrinogen-induced tube formation was significantly inhibited in the presence of anti-VEGF-neutralizing antibody (52.0% inhibition at the concentration of 1 μg/ml, P<0.05) or SU5416 (54.8% inhibition at the concentration of 3 μM, P<0.05). Conclusions: Extravasated fibrinogen might play an important role in the development of choroidal neovascularization associated with age-related macular degeneration, at least in part, through the function of VEGF in an autocrine manner. Transcription factor HIF-1 appears to be involved in fibrinogen-induced VEGF expression.

Original languageEnglish
Pages (from-to)777-783
Number of pages7
JournalGraefe's Archive for Clinical and Experimental Ophthalmology
Issue number9
Publication statusPublished - Sept 2004

All Science Journal Classification (ASJC) codes

  • Ophthalmology
  • Sensory Systems
  • Cellular and Molecular Neuroscience


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