Gas6/Axl Inhibits Osteosarcoma Apoptosis Through Regulation of Apoptosis-Related Protein Bcl-2

Background: Dysregulation of the receptor tyrosine kinase Axl and its ligand Gas6 has been shown to promote the progression of osteosarcoma and correlates with poor prognosis. This study aimed to identify the role of Gas6/ Axl for anti-apoptosis as induced by cisplatin (DDP) and methotrexate (MTX) chemotherapy and to analyze the relationship between P-Axl and apoptosis-related proteins in osteosarcoma. Method: Cultured osteosarcoma cell lines MG63, 143B and U2OS were used for apoptosis assays, Axl siRNA transfection, cytotoxicity assays, cell cycle analysis, and other assessment methods. A total of 41 cases of osteosarcoma patients were included for immunohistochemistry staining and clinicopathological relative analysis. TUNEL assay was performed in ten pair’s cases for apoptosis detection and relative analysis of P-Axl. Results: Among the osteosarcoma cell lines, Gas6 could obviously protect tumor cells from apoptosis induced by DDP and MTX by binding to Axl (P<0.05). Axl siRNA transfection enhanced cell apoptosis, whereas Gas6 was unable to function upon previous knockdown of Axl. Among the 41 osteosarcoma cases, the positive rate of Bcl-2, Bax, and P-Axl was 70.7%, 36.6%, and 85.4%, respectively. In osteofibrous dysplasia, the positive rate of them was 22.2%, 11.1%, and 14.6%, respectively. The expression levels of these apoptosis-related factors were significantly higher in osteosarcoma than in osteofibrous dysplasia (P<0.05). Through clinico-pathological analysis, there were significant relationships between the survival status and Bcl-2 or Bax expression (P<0.05). TUNEL assay also demonstrated that P-Axl high expression inhibited apoptosis in osteosarcoma tissues. By Cox univariate analysis, Bcl-2 or Bax was correlated with the patients’ prognosis. Importantly, Pearson correlation analysis demonstrated that Bcl-2 was positively correlated to P-Axl with statistical significance (r=0.842, P<0.0001). Conclusion: Gas6/Axl protects osteosarcoma cells from the apoptosis induced by DDP and MTX chemotherapy and inhibits apoptosis in osteosarcoma tissue, possibly through the regulation of apoptosis-related protein Bcl-2.