[Interaction between necroptosis and apoptosis in MC3T3-E1 cell death induced by dexamethasone].
[Interaction between necroptosis and apoptosis in MC3T3-E1 cell death induced by dexamethasone].
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DOI：
10.12122/j.issn.1673-4254.2019.09.05
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发表时间：
2019-09-30
影响因子：
--
通讯作者：
Qiang, Hui
中科院分区：
文献类型：
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作者：
Feng, Min;Zhang, Ruirui;Qiang, Hui
目的：目的探讨糖皮质激素诱导的MCET 3-E1细胞死亡中坏死性凋亡与细胞凋亡的关系。方法：将MC 3 T3-E1细胞与10-6 mol/L地塞米松孵育，然后用细胞凋亡抑制剂z-VAD-fmk（40 mumol/L）或坏死性凋亡抑制剂necrostatin-1（40 mumol/L）处理2 h。在与地塞米松孵育后72 h，收获细胞以使用WST-1测定法测定细胞活力，并使用膜联蛋白V/PI双染色测定坏死细胞的比率;使用Hoechst染色测定凋亡细胞的百分比。同时测定细胞线粒体膜电位和ATP含量。透射电镜观察细胞的显微结构变化。结果：10-6 mol/L地塞米松可诱导MC 3 T3- E1细胞凋亡和坏死性凋亡。Annexin V/PI双染显示，抑制细胞凋亡导致细胞坏死增加，电镜显示线粒体肿胀和质膜破裂等变化; Hoechst染色显示凋亡细胞百分比显著降低。结论：在地塞米松诱导的MC 3 T3-E1细胞死亡过程中，坏死性凋亡和细胞凋亡可转化为线粒体膜电位和ATP水平的改变，并伴有线粒体功能的改变。
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 mumol/L) or the necroptosis inhibitor necrostatin-1 (40 mumol/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.CONCLUSIONS: In the process of dexamethasone-induced cell death, necroptosis and apoptosis can transform reciprocally accompanied by functional changes of the mitochondria.