Abstract
Aims: RECK protein is involved in angiogenesis and matrix-metalloproteinase interaction. In common cancers RECK may exert tumour control, while in sarcomas, there is insufficient research to confirm this. Our study aims to investigate the role of RECK in osteo-sarcoma (OS) and chondrosarcoma (CS) angiogenesis, using clinical data and an in vitro angiogenesis assay.
Methods: Seven OS biopsy samples, 11 OS metastases and 24 samples of CS were stained for the RECK protein using immunohistochemistry. RECK expression in vessels supplying sarcoma tissue was semi-quantified using ImageProPlus software to determine staining density. RECK density in tumour vessels was compared with density in vessels of 18 normal tissues. The human endothelial cell line, HMEC-1, was cultured and then transfected with a plasmid containing the RECK gene. HMEC-1 cells over-expressing RECK were then distributed into wells containing 100% Matrigel. Formation of vessel-like structures was observed and photographed at 8 hours. Comparison was made with a non-transfected HMEC-1 group, and an empty vector transfected HMEC-1 group. Results: RECK is predominantly expressed in vessels supplying both sarcomas and less frequently in the tumour cells. RECK density in OS and CS primary tumour vessels was no different to normal vessels (p> 0.05). However there was a significantly higher RECK density in the vessels of OS metastasis tissue (0.76 ODU-optical density units), compared with primary OS vessels (0.57 ODU, p< 0.05), and normal vessels (0.56 ODU, p< 0.05).
The average number of tube formations in the RECK transfected HMEC-1 cells was 67.8, compared with 42 in the empty vector group (p=0.03) and 54.1 in the control group (p=0.03). The average maximal wall thickness of tube formations was 100.7um versus 77.4um in the empty vector group (p=0.04) and 83.0um in the control group (p=0.09).
Conclusions: RECK is upregulated in vessels supplying OS metastases compared to vessels in primary sarcoma and normal tissue, suggesting an active role in angiogenesis. Replication of these conditions in vitro using the HMEC-1 tube formation assay demonstrates the ability of RECK to increase vessel formation within a matrix while maintaining vessel wall thickness. In sarcoma RECK may have pro-angiogenic properties and could be an important part of the metastatic cascade.
The abstracts were prepared by David AF Morgan. Correspondence should be addressed to him at davidafmorgan@aoa.org.au
Declaration of interest: b
