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| 二苯乙烯苷对H2O2诱导的MC3T3-E1凋亡的保护作用及机制研究 |
| Protective effect of TSG on H2O2 induced cytotoxicity in MC3T3-E1 cells and the related mechanism |
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| DOI：10.3969/j.issn.1006-7108.2018.12.008 |
| 中文关键词: 二苯乙烯苷 氧化应激 成骨前体细胞 细胞凋亡 |
| 英文关键词:tetrahydroxystilbene glucoside oxidative stress MC3T3-E1 cells cell apoptosis |
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| 中文摘要: |
| 目的 研究二苯乙烯苷（tetrahydroxystilbene glucoside ，TSG）对氧化应激导致的成骨前体细胞MC3T3-E1凋亡的保护作用，并初步探讨相关机制。方法 不同浓度TSG预处理MC3T3-E1 24 h后，300 μmol/L H2O2作用24 h。实验分组：①空白对照组；②单纯H2O2处理组；③H2O2+TSG 0.1μmol/L处理组；④H2O2+TSG 1μmol/L处理组；⑤H2O2+TSG 10μmol/L处理组；⑥阳性对照抗氧化剂NAC 1mmol/L处理组，通过MTT法、Hoechst33258 染色来评价细胞凋亡情况及TSG对氧化损伤的保护作用； 利用荧光酶标仪和MDA检测试剂盒来检测细胞中ROS及MDA的水平来评价细胞的氧化应激状态；Western-blot和RT-PCR分别从蛋白水平和基因水平评价Bcl-2和Bax的表达水平。结果 单纯H2O2处理可以导致细胞死亡，显微镜下可见许多细胞死亡，脱壁；Hoechst33258染色可见有较多的细胞出现核固缩、核浓集现象；不同浓度的TSG预处理后，细胞凋亡现象得到改善。单独H2O2作用可以导致成骨前体细胞出现较大程度的凋亡，不同浓度TSG预处理后，细胞凋亡率有所降低；单纯H2O2作用可以导致细胞水平的ROS和脂质氧化产物MDA产生增加，TSG（1~10μmol/L）可以显著降低细胞水平ROS和MDA水平（P<0.05），改善细胞的氧化应激状态；Western-blotting和实时定量 RT-PCR结果TSG预处理可以减少H2O2处理后，促凋亡蛋白Bax的表达，增加抗凋亡蛋白Bcl-2的表达。结论 氧化应激可以导致成骨前体细胞MC3T3-E1凋亡增加，TSG可以通过降低氧化应激水平。其机理可能是TSG通过促凋亡蛋白Bax的表达及增加抗凋亡蛋白Bcl-2的表达起到保护作用。 |
| 英文摘要: |
| Objective The aim of this study was to explore the protective effect and related mechanism of tetrahydroxystilbene glucoside (TSG) on apoptosis of osteoblast precursor cells (MC3T3-E1) induced by H2O2. Methods MC3T3-E1 cells pretreated with different concentrations of TSG were treated with 300 μmol/L H2O2 for 24 h. The experiment consisted of 6 groups, including the control group, H2O2, H2O2+TSG (0.1 μmol/L), H2O2+TSG (1 μmol/L), H2O2+TSG (10 μmol/L) and H2O2+NAC (1 mmol/L) (anti-oxidant positive control group). We evaluated the protective effect of TSG on the apoptosis of MC3T3-E1 cells by MTT assay and Hoechst33258 staining. We tested the production of ROS and MDA by fluorescence microplate reader and MDA assay kit to investigate the oxidative stress status. We tested the production of Bcl-2 and Bax by Western-blotting and the gene expression of Bcl-2 and Bax by RT-PCR. Results Cell death induced by H2O2 alone was observed under microscope, and Hoechst33258 staining showed more cells with nuclear pyknosis and nuclear enrichment, which was improved after being pretreated with different concentrations of TSG. H2O2 alone could lead to a large degree of apoptosis in MC3T3-E1 cells, and the apoptosis rate decreased after pretreatment with different concentrations of TSG. At the same time, TSG (1-10 μmol/L) could significantly reduce the level of ROS and MDA (P<0.05) and improve the oxidative stress status in MC3T3-E1 cells. The results of Western-blotting and real-time quantitative RT-PCR demonstrated that pretreatment with TSG could decrease the expression of pro-apoptosis protein Bax and increase the expression of anti-apoptosis protein Bcl-2. Conclusion The apoptosis of MC3T3-E1 cells increased after induced by H2O2 and decreased by pretreatment with TSG. The mechanism of the protective effect could be through decreasing the expression of pro-apoptosis protein Bax and increasing the expression of anti-apoptosis protein Bcl-2. |
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