Preparation Of Selenium Doped Hydroxyapatite And The Experimental Study On Their Antitumor Effect

Hydroxyapatite (HA), because of its good biocompatibility and wide use in the biomedical field, has become a hotspot in recent years. As such in this paper we designed, synthesized and characterized selenium doped hydroxyapatite nanoparticles (SeHAN), hoping to apply the antitumor effect of selenium for the utility of repairing bone defect whilst inhibiting bone cancer recurrence. The research content of this paper mainly includes the following three aspects:(1) SeHAN with different Se/P mole ratios was synthesized by a co-precipitation method, using sodium selenite (Na2SeO3) as the Se source. X-ray diffraction (XRD), Fourier-transform infrared spectroscopy (FTIR), and transmission electron microscopy (TEM) etc. techniques revealed that, when the Se/P ratio was less than1.0, the product was hydroxyapatite phase with doped selenium, with increase in the amount of doped selenium, the lattice changed and the crystallinity decreased. When the Se/P ratio was0.3, amorphous flocculent substances existed in the products. When the Se/P ratio was more than10.0, calcium selenite hydrate appeared in the product. Sintering tests revealed that the powder with Se/P ratio of0.1had thermal stability at900℃for2hours. These results showed that Se/P ratio of0.1was the highest doping concentration of selenium for the preparation of SeHAN with HA phase.(2) Cell culture experiments revealed that, SeHAN improved bone mesenchymal stem cells (BMSCs) proliferation whilst inducing apoptosis of osteosarcoma cells (MG63). SeHAN could have entered the MG63cells through phagocytosis, and were found to be located in the cytoplasm or near to the nucleus, SeHAN decreased the proliferative activity of MG63cells by about40%; with the increase of the concentration of SeHAN used, the cell viability of MG63cells decreased; in case of the same concentration, the cell viability of MG63cells decreased with increase in amount of doped selenium. The results of cell staining exhibited that, early apoptosis appeared in MG63cells, and it was closely related with the increased reactive oxygen species (ROS) and decreased mitochondrial membrane potential (MMP). It was postulated that, SeHAN could have induced the MG63cell apoptosis by activating the mitochondria signal pathway. (3) The in vivo animal experiments revealed that, SeHAN exhibited antitumor effect on both osteosarcoma and hepatoma. Results about the in situ animal model of osteosarcoma showed that SeHAN suspension could retard the growth of osteosarcoma cells, and inhibit their metastasis. Compared to HAN, SeHAN showed a better biodegradable capability. Blood analysis showed that SeHAN could improve the function of liver, kidney and heart. Histopathology and Immunohistochemistry results showed that the injected SeHAN caused tissue necrosis occupying a large portion at the center of the tumor, and also resulted in a decrease in expression of MMP-9protein. As a model of hepatocellular carcinoma-bearing nude mice with high metastatic potentials, the injected SeHAN suspension dramatically prolonged the lifetime of nude mice. The overall survival rate of nude mice on the36th day in the control, HAN and SeHAN group was50.00%,76.92%and100.00%respectively. Blood biochemical studies confirmed that SeHAN group had significantly lower toxicities on the liver and kidney functions. Histopathological studies also revealed more conspicuous tissue necrosis and calcium depositions in the tumor following SeHAN treatment. Moreover, immunohistochemistry and Western blot assay showed a significant reduction in the expression of the Ki-67, VEGF and MMP-9protein after injection of SeHAN.In conclusion, SeHAN nanoparticles were applied, not only as a new nanomaterial in the treatment of osteosarcoma, but also as an antitumor agent in the treatment of hepatocellular carcinoma.