Introduction Hyaline cartilage is a barrier to osteosarcoma invasion, however the mechanisms behind this resistance remain unclear. The aim of this study was to examine the temporo-spatial pattern of osteosarcoma growth and invasion of local tissue structures, including epiphyseal cartilage, and to investigate the molecular mechanisms behind the resistance of cartilage to malignant invasion.

Methods An in vivo mouse model of osteosarcoma was used, whereby osteosarcoma cells were orthotopically injected into the tibiae of nude mice. Animals were sacrificed at weekly timepoints. Control and tumour limbs were processed for histological examination of tumors at different stages of disease progression. Routine Haematoxylin & Eosin staining was used to examine morphology, and immunohistochemical staining using antibodies against proangiogenic vascular endothelial growth factor (VEGF) and anti-angiogenic pigment epithelium-derived factor (PEDF) was performed. PEDF from mouse liver was cloned into a mammalian expression vector in order to generate stably-transfected osteosarcoma cell lines.

Results Hyaline cartilage of the growth plate and articular surface was resistant to local invasion by osteosarcoma in all sections examined, despite increasing tumor size as well as extensive intra- and extra-osseous destruction. All tumours showed immunostaining for VEGF but not for PEDF. In the most advanced cases, only the lowermost layers of the hypertrophic zone of the growth plate were eroded. These layers displayed strong immunostaining for the potent angiogenic factor VEGF, and weak to absent immunostaining for PEDF. By contrast, the resting, proliferative and upper hypertrophic layers, which were resistant to osteosarcoma invasion in the cases studied, showed high expression levels of the potent anti-angiogenic factor PEDF.

Conclusions These results confirm that the balance of angiogenesis, influenced by pro and anti-angiogenic factors, determines tumour growth and invasion. Given the localization of PEDF specifically to the resistant cartilaginous layers and its exceptionally potent anti-angiogenic effects, there are exciting prospects for the use of PEDF in treatment for osteosarcoma as well as other cancers. To this end, we have established osteosarcoma cell lines that over-express PEDF and are currently characterizing these cells in vitro and assessing the propensity of PEDF to suppress tumour invasion in vivo. Growth plate cartilage is resistant to invasion by osteosarcoma. PEDF is likely to play an important role in this resistance. As such, it may have therapeutic applications in osteosarcoma as well as other malignancies.

In relation to the conduct of this study, one or more of the authors is in receipt of a research grant from a non-commercial source.