Cisplatin-induced Senescence and Senescence-related Gene Expression Profiles in Human Nasopharyngeal Carcinoma Cells CNE2
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摘要: 目的 探讨顺铂体外诱导人鼻咽癌CNE2细胞衰老的效应及其分子机制。方法 采用 MTT 比色法观察顺铂对CNE2细胞增殖的抑制作用，衰老相关-β-半乳糖苷酶（SA-β-gal）活性评判细胞衰老，流式细胞术分析细胞周期分布，实时定量PCR法检测78个人类细胞衰老相关基因的表达。结 果 顺铂对CNE2细胞的增殖具有明显抑制作用；1.0 mg/L顺铂处理后第6和第8天检测SA-β-gal染色阳性细胞数分别达到67.63%和89.22%，细胞阻滞于G2 期；TP53、TP63、p16Ink4a，p27Kip1、PTEN、RB1、Cdc25C、TBX3、Gadd45a、IGF1R、PIK3CA、BMI-1、B2M、MORC3、MYC和SPARC等16个衰老相关基因的mRNA水平显著升高（P＜0.05或0.01），Cyclin A2、Cyclin B1、Cyclin E1、CDK6、ATM、ETS2、MDM2、E2F1、ETS1、FN1、IGFBP3、RBL2、SERPINB2及SIRT1等14个基因表达水平下降（P＜0.05 或 0.01）。结论 顺铂能够在体外诱导CNE2细胞衰老，其机制涉及p16和p27介导的p53-pRB和PTEN衰老信号调控网络。Abstract: Objective To investigate the effects of cisplatin induction in vitro on human nasopharyngeal carcinoma cells CNE2 and its possible molecular mechanism. Methods Proliferation of CNE2 cells treated with various concentrations(0.4-4.0 mg/L) of cisplatin was determined using MTT assay. The senescent cells were determined by a senescence-associated-β-galactosidase(SA-β-gal) activity assay and cell cycle distribution was analyzed by flow cytometry. A Sybe Green real-time polymerase chain reaction(RT-PCR) method was used to examine the changes of expression profiles of 78 human cellular senescence-related genes. Results Cisplatin inhibited the proliferation of CNE2 cells in a dose- and time-dependent manner, and significantly induced a cellular senescence in CNE2 cells, which respectively counted at 67.63% of senescent cells at day 6 and 89.22% at day 8, after treatment with 1.0 mg/L of cisplatin. A G2/M phase arrest was observed. The expression levels of 16 senescence-related genes including TP53, TP63, p16Ink4a, p27Kip1, PTEN, RB1, TBX3, Cdc25C, Gadd45a, IGF1R, PIK3CA, MI-1, B2M, MORC3, MYC and SPARC were increased(P＜0.05 or 0.01), while the 14 genes including Cyclin A2, Cyclin B1, Cyclin E1,CDK6, ATM, E2F1, ETS1, ETS2, MDM2, FN1, IGFBP3, RBL2, SERPINB2 and SIRT1 were decreased at mRNA levels(P＜0.05 or 0.01). Conclusion Cisplatin could induce G2/M phase arrest and cellular senescence in CNE2 cells in vitro, which may involve a omplicated network of p53-pRb signaling or/and PTEN signaling pathways mediated by p16 and p27.
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Key words:
- Cisplatin /
- Tumor cell senescence /
- Nasopharyngeal carcinoma /
- Cell cycle arrest
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