| 李 芬,周 琦,丁丽丽.Akt抑制剂MK-2206清除衰老神经母细胞瘤细胞的实验研究[J].肿瘤学杂志,2020,26(5):396-401. |
| Akt抑制剂MK-2206清除衰老神经母细胞瘤细胞的实验研究 |
| Elimination of Senescent Neuroblastoma Cells by Akt Inhibitor MK-2206 |
| 投稿时间：2019-10-08 |
| DOI：10.11735/j.issn.1671-170X.2020.05.B006 |
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| 中文关键词: Akt抑制剂 MK-2206 MLN8237 化疗诱导的细胞衰老 神经母细胞瘤 |
| 英文关键词:Akt inhibitor MK-2206 MLN8237 chemotherapy-induced senescent cells neuroblastoma |
| 基金项目:国家自然科学基金项目（81472706）；中央高校基本科研业务费专项资金(2015LC042) |
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| 中文摘要: |
| 摘 要：［目的］ 探讨Akt抑制剂MK-2206清除衰老神经母细胞瘤细胞IMR-32的效果及可能机制。［方法］ 极光激酶A抑制剂MLN8237处理神经母细胞瘤细胞系IMR-32，利用衰老相关β半乳糖苷酶（SA-β-gal）染色检测细胞状态、免疫荧光法检测组蛋白磷酸化和甲基化情况评估细胞衰老状况。CCK-8检测不同浓度MK-2206及MK-2206与MLN8237联合对IMR-32细胞增殖的影响，流式细胞术检测细胞凋亡状况。Western blot法检测MK-2206与MLN8237联合用药对IMR-32细胞 Akt、p-Akt、STAT3和N-Myc表达的影响。［结果］ MLN8237作用IMR-32细胞后，细胞核体积明显增大，细胞呈现大而扁平的衰老状态，细胞蓝染数显著增多；组氨酸 H3 Ser10磷酸化水平和组氨酸 H3 K9甲基化水平显著性升高。Western blot显示衰老IMR-32细胞内磷酸化Akt和STAT3蛋白显著性上调。10μmol/L MK-2206干预衰老细胞72h和96h后细胞存活率分别为（80.13±5.30）%、（74.29±4.06）%，较不干预衰老细胞组显著性下降。在MK-2206与MLN8237联合作用下细胞出现明显凋亡，凋亡率为（15.70±0.97）%，而不干预衰老细胞组细胞凋亡率为（8.81±1.33）%。同时两药联合组胞内p-Akt、STAT3和N-Myc蛋白显著性下调。［结论］ MK-2206可能通过抑制衰老神经母细胞瘤细胞内p-Akt，下调STAT3和N-Myc蛋白的机制，诱导细胞凋亡，从而达到清除MLN8237诱导的衰老细胞目的。 |
| 英文摘要: |
| Abstract：［Objective］ To investigate the effect and possible mechanism of Akt inhibitor MK-2206 on elimination of senescent neuroblastoma cells. ［Methods］ Neuroblastoma IMR-32 cells were treated with aurora A inhibitor MLN8237，and cell senescence was assessed by β galactosidase(SA-β-gal) activity assay and immunofluorescence assay for the detection of histone phosphorylation and methylation. CCK-8 assay was used to detect the effects of MK-2206 combined with MLN8237 on the proliferation of IMR-32 cells. Cell apoptosis was detected by flow cytometry. The expressions of Akt，p-Akt，STAT3 and N-Myc proteins were detected by Western blot analysis. ［Results］ MLN8237 treatment induced senescence of neuroblastoma IMR-32 cells，and the enlarged and flattened morphology with increased activity of SA-β-gal was observed in senescent IMR-32 cells. Immunofluorescence showed up-regulation of phosphorylation of histone H3Ser10 and methylation of histone H3K9 in MLN8237-treated IMR-32 cells. Western blot showed a significant up-regulation of phosphorylated Akt and STAT3 proteins in senescent IMR32 cells. The survival rate of senescent cells was(80.13±5.30)% and (74.29±4.06)% at 72h and 96h after 10mol/L MK-2206 intervention，which was significantly lower than that of the senescent cells without intervention. Treatment of MLN8237 combined with MK-2206 showed a significantly increased apoptosis rate(15.70±0.97) %，while that without intervention was(8.81±1.33)%. The expressions of p-Akt，STAT3 and N-Myc proteins in the combination group were also significantly down-regulated. ［Conclusion］ Akt inhibitor MK-2206 can enhance the apoptosis of MLN8237-induced senescent neuroblastoma cells，which may be associated with the inhibiting p-Akt，down-regulating STAT3 and N-Myc. |
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