, Volume 37, Issue 5, pp 2509-2515
Date: 22 Aug 2009

Arsenic trioxide inhibits the growth of Adriamycin resistant osteosarcoma cells through inducing apoptosis

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Abstract

Given that arsenic trioxide (As2O3) has been successfully used as a chemotherapeutic agent for refractory malignant tumors, this study is aimed at investigating the effect of As2O3 on human Adriamycin resistant osteosarcoma cell line Saos-2. The mechanism underlying multi drug resistance (MDR) in osteosarcoma cells and the anti-tumor effect of As2O3 on Adriamycin resistant osteosarcoma cells were analyzed. In our experiment, we first selected Adriamycin resistant osteosarcoma cell line by growing the classic osteosarcoma cell line Saos-2 in the medium with increasing drug concentrations. Then, we compared the IC50s of the osteosarcoma cells treated with different anticancer drugs by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Subsequently, we assessed the expression of classic MDR related molecules, Pgp, multidrug resistance-associated protein (MRP) and glutathione (GSH) activity in the wild type and Adriamycin resistant Saos-2 cells. Furthermore, the apoptosis was assessed by concerning DNA fragment and flow cytometry with Annexin-V staining. To elucidate the underlying mechanism of the apoptosis, related proteins Bcl-2, Bcl-xL, Bax, Bak, cleaved Caspase-3 and cleaved Caspase-9 were analyzed by western blotting. The data showed that the resistance to Adriamycin affected the sensitivity of osteosarcoma cell to other chemotherapeutic agents. The IC50s of Saos-2/ADM cells for methotrexate (1.74-fold), Cisplatin (1.43-fold) and As2O3 (1.21-fold) were increased compared with Saos-2 control cells. The expression of Pgp was upregulated comparing with the control cells. No significant difference was detected about the MRP and the glutathione-S-transferase activity and intracellular GSH concentration among different treated osteosarcoma cells. Apoptosis was observed and proved. The western blotting showed that the expression of Bcl-2 and Bcl-xL was downregulated. Meanwhile, the level of Bax, Bak, cleaved Caspase-3 and cleaved Caspase-9 was upregulated after treated with As2O3. The study suggests that Adriamycin resistant osteosarcoma cells have good response to As2O3-based chemotherapy in vitro, probably via the pathway of inducing apoptosis. And As2O3 might serve as an excellent alternative candidate for adjuvant chemotherapeutic agent on this incurable pediatric sarcoma.

Each author certifies that his or her institution has approved the experimental protocol for this investigation and that all investigations were conducted in conformity with ethical principles of research.