[1]冯敏,张睿锐,杨佩,等.Necroptosis与细胞凋亡在激素诱导成骨细胞死亡中的相互作用[J].南方医科大学学报,2019,(09):1030.[doi:10.12122/j.issn.1673-4254.2019.09.05]
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Necroptosis与细胞凋亡在激素诱导成骨细胞死亡中的相互作用()
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《南方医科大学学报》[ISSN:1673-4254/CN:44-1627/R]

卷:
期数:
2019年09期
页码:
1030
栏目:
出版日期:
2019-09-15

文章信息/Info

Title:
Interaction between necroptosis and apoptosis in MC3T3-E1 cell death induced by dexamethasone
作者:
冯敏张睿锐杨佩王坤正强辉
关键词:
糖皮质激素MC3T3-E1细胞细胞凋亡程序性坏死Necrostatin-1caspase
Keywords:
glucocorticoids MC3T3-E1 cells apoptosis necroptosis necrostatin-1 caspase
DOI:
10.12122/j.issn.1673-4254.2019.09.05
摘要:
目的探讨Necroptosis与凋亡在激素诱导成骨细胞死亡中的相互作用。方法向MC3T3-E1细胞内添加浓度为10-6 mol/L 的地塞米松诱导细胞死亡,随后在本组细胞中分别添加凋亡抑制剂z-VAD-fmk(40 μmol/L)和Necroptosis抑制剂Necrostatin-1 (40 μmol/L),作用2 h 后,AV/PI 双染观察细胞死亡的变化情况,并对细胞进行Hoechst 染色计算凋亡率,用透射电镜观察细 胞超微结构的改变,测定细胞线粒体膜电位和ATP水平,对Necroptosis 和凋亡通路的相关蛋白进行Western blot 检测。结果 10-6 mol/L地塞米松可以诱导MC3T3-E1细胞同时发生凋亡和Necroptosis。AV/PI双染的结果发现,当凋亡受到抑制后,更多的 细胞发生坏死(P<0.01),而透射电镜结果也证实细胞发生了坏死样改变,Hoechst 染色结果发现,凋亡细胞数明显减少(P< 0.01);当使用Necroptosis特异性抑制剂Necrostatin-1将这种作用阻断后,包括Hoechst染色和AV/PI双染的结果都证实凋亡的 细胞数明显增多(P<0.01),同时,在这一作用过程中伴随着MMP及ATP的明显变化(P<0.01)。结论在激素诱导成骨细胞死亡 过程中,Necroptosis和凋亡在一定条件下可以相互转化,这一转化过程中伴随着线粒体功能的明显改变。
Abstract:
Objective To investigate the relationship between necroptosis and apoptosis in MCET3-E1 cell death induced by glucocorticoids. Methods MC3T3-E1 cells were incubated with 10-6 mol/L dexamethasone followed by treatment with the apoptosis inhibitor z-VAD-fmk (40 μmol/L) or the necroptosis inhibitor necrostatin-1 (40 μmol/L) for 2 h. At 72 h after incubation with dexamethasone, the cells were harvested to determine the cell viability using WST-1 assay and the rate of necrotic cells using annexin V/PI double staining; the percentage of apoptotic cells was determined using Hoechst staining. The mitochondrial membrane potential and the level of ATP in the cells were also evaluated. Transmission electron microscopy was used to observe the microstructural changes of the cells. The expressions of RIP-1 and RIP-3 in the cells were detected by Western blotting. Results At a concentration of 10-6 mol/L, dexamethasone induced both apoptosis and necroptosis in MC3T3- E1 cells. Annexin V/PI double staining showed that inhibition of cell apoptosis caused an increase in cell necrosis manifested by such changes as mitochondrial swelling and plasma membrane disruption, as shown by electron microscopy; Hoechst staining showed that the percentage of apoptotic cells was significantly reduced. When necroptosis was inhibited by necrostatin-1, MC3T3-E1 cells showed significantly increased apoptosis as shown by both AV/PI and Hoechst staining, and such changes were accompanied by changes in mitochondrial membrane potential and ATP level in the cells. Conclusion In the process of dexamethasone-induced cell death, necroptosis and apoptosis can transform reciprocally accompanied by functional changes of the mitochondria.

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更新日期/Last Update: 1900-01-01