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Angiopoietin 2 displays a vascular endothelial growth factor dependent synergistic effect in hepatocellular carcinoma development in mice


Background: Orchestration of two major classes of angiogenic factors—namely, vascular endothelial growth factor (VEGF) and angiopoietin 2 (Ang-2)—has been shown to play a pivotal role in tumour angiogenesis, including hepatocellular carcinoma (HCC). However, few studies have focused on the direct interaction of these factors on in vivo tumour development and angiogenesis.

Aim: To examine the interaction between both factors in murine HCC.

Methods: We examined the combination effect of VEGF and Ang-2 overexpression by means of a combination of a retroviral tetracycline (tet) regulated gene manipulating system in vivo, by providing tet in the drinking water, and a conventional plasmid gene expression system.

Results: Neither Ang-2 nor VEGF overexpression induced proliferation of HCC cells in vitro. In vivo, although overexpression of Ang-2 did not increase tumour development, simultaneous expression of Ang-2 and VEGF synergistically augmented tumour growth and angiogenesis in murine HCC. Ang-2 plus VEGF induced tumour development was markedly attenuated by treatment with neutralising monoclonal antibodies against VEGF receptors. Ang-2 plus VEGF overexpression significantly increased the activities of matrix metalloproteinase (MMP)-2 and MMP-9 in the tumour. Suppression of intratumoral VEGF almost completely abolished this augmentation of MMPs.

Conclusions: These results suggest that Ang-2 synergistically augments VEGF mediated HCC development and angiogenesis. This proangiogenic activity was exerted only in the presence of VEGF, at least partly mediated via induction of MMP-2 and MMP-9 in the tumour.

  • α-SMA, α smooth muscle actin
  • Ang-1, Ang-2, angiopoietin 1 and 2, respectively
  • APMA, p-aminophenylmercuric acetate
  • bFGF, basic fibroblast growth factor
  • BNL-HCC, BNL 1.7R1 hepatocellular carcinoma
  • EC, endothelial cell
  • ECM, extracellular matrix
  • ELISA, enzyme linked immunosorbent assay
  • HCC, hepatocellular carcinoma
  • tet, tetracycline
  • mAb, monoclonal antibody
  • MMP, matrix metalloproteinase
  • MVD, microvessel density
  • PCR, polymerase chain reaction
  • VEGF, vascular endothelial growth factor
  • VEGFR-1, fms-like tyrosine kinase (flt-1)
  • VEGFR-2, kinase-insert domain containing receptor/fetal liver kinase-1 (KDR/Flk-1)
  • angiopoietin
  • vascular endothelial growth factor
  • hepatocellular carcinoma
  • angiogenesis
  • tumour growth

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