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目的探讨高同型半胱氨酸(HHcy)诱导人成骨肉瘤MG63细胞损伤的分子机制。方法体外培养的人成骨肉瘤MG63细胞经不同浓度(0.001mol/L、0.003mol/L和0.005mol/L)的Hcy处理不同时间后,采用2’,7’-二氯二氢荧光素二乙酸盐(H2 DCFDA)探针在荧光分光光度计下检测细胞内活性氧(ROS)变化水平;通过Hoechst 33342/碘化丙啶(PI)核双染法在荧光显微镜下观察计数凋亡和坏死细胞;采用Western blotting法检测促凋亡相关蛋白Caspase-3和Bax的表达变化。结果在高浓度的Hcy刺激下,以剂量和时间依赖的方式上调MG63细胞内ROS含量(P<0.05);同时,凋亡相关蛋白Caspase-3和Bax的表达明显增加,从而诱导MG63细胞的死亡。结论高浓度Hcy可通过增加MG63细胞内ROS的含量,上调凋亡相关蛋白Caspase-3和Bax的表达,导致MG63细胞的氧化损伤。
Objective To investigate the molecular mechanism of high homocysteine (HHcy) -induced human osteosarcoma MG63 cell injury. Methods Human osteosarcoma MG63 cells cultured in vitro were treated with Hcy at different concentrations (0.001 mol / L, 0.003 mol / L and 0.005 mol / L) for 2 h, then treated with 2 ’, 7’- dichlorodihydrofluorescin (H2 DCFDA) probe was used to detect the level of reactive oxygen species (ROS) in the cells under the fluorescence spectrophotometer. The apoptotic rates of the cells were observed under a fluorescence microscope with the Hoechst 33342 / propidium iodide (PI) Necrotic cells; Western blotting was used to detect the expression of apoptosis-related proteins Caspase-3 and Bax. Results In a dose-and time-dependent manner, the intracellular ROS level in MG63 cells was up-regulated (P <0.05) under high concentration of Hcy stimulation. At the same time, the expressions of Caspase-3 and Bax were significantly increased, leading to the death of MG63 cells . Conclusion High concentration of Hcy can up-regulate the expression of Caspase-3 and Bax in MG63 cells and increase the oxidative damage of MG63 cells.