山东大学耳鼻喉眼学报 ›› 2020, Vol. 34 ›› Issue (4): 105-110.doi: 10.6040/j.issn.1673-3770.0.2019.503

• 论著 • 上一篇    下一篇

Notch受体调控上皮-间质转化对鼻咽癌细胞顺铂耐药的影响

韩继波,邹游,杨蕊,陶泽璋   

  1. 韩继波, 邹游, 杨蕊, 陶泽璋武汉大学人民医院 耳鼻咽喉头颈外科, 湖北 武汉 430060
  • 收稿日期:2019-10-09 出版日期:2020-07-20 发布日期:2020-08-28

Cisplatin resistance in nasopharyngeal carcinoma cells is affected by Notch receptors through the regulation of epithelial-mesenchymal transition rather than cell cycle controlHAN Jibo, ZOU You, YANG Rui, TAO Zezhang Department of Otolaryngology-Head and Neck Surgery, Renmin Hospital of Wuhan University, Wuhan 430060, Hubei, ChinaAbstract:Objective〓

The objective of this study was to analyze the effects and possible regulatory mechanisms that Notch receptors could have on cisplatin resistance, observed in nasopharyngeal carcinoma. MethodsWestern blot analysis was used for the detection of Notch receptor expression in nasopharyngeal carcinoma cells and cisplatin-resistant nasopharyngeal carcinoma cells(5-8F, 5-8F/CDDP). Flow cytometry was used to investigate how the combined treatment with 10 μM CDDP and different concentrations of γ-secretase inhibitor(DAPT)could affect apoptosis in 5-8F/CDDP cells. Flow cytometry was also used for the detection of cell cycle stages in DAPT-treated 5-8F / CDDP cells. Finally, western blotting was also used for the detection of drug resistance-related protein expression. All experiments were followed by statistical data analysis. ResultsWe observed significantly higher Notch1 and Notch4 receptor expression in 5-8F/CDDP cells than in 5-8F cells(P=0.003, P=0.004). Furthermore, we described that Notch signaling was inhibited by DAPT in 5-8F/CDDP cells, followed by a significant increase in the apoptosis rate and decrease in cell proliferation, induced by cisplatin in a dose-dependent manner(P<0.05). Moreover, after inhibiting the Notch signaling pathway in 5-8F/CDDP cells, DAPT treatment significantly decreased the expression of Cyclin E and CDK-2, proteins involved in cell cycle regulation, and contributed to blocking the cells in the G1/S phase(P<0.05). At the same time, the expression levels of both the EMT-related protein Slug and the DNA excision repair protein ERCC1 significantly decreased, while that of E-Cadherin was up-regulated. ConclusionUp-regulated expression of Notch1 and Notch4 receptors is associated with cisplatin resistance in nasopharyngeal carcinoma cells. The inactivation of the Notch signaling pathway might thus have the potential to enhance the efficiency of cisplatin chemotherapy in drug-resistant nasopharyngeal carcinoma cells by inhibiting EMT rather than blocking the G1/S cell cycle phase.   

  1. Key words: Notch receptors;
    Cell cycle;
    EMT;
    Cisplatin resistance of nasopharyngeal carcinoma
  • Received:2019-10-09 Online:2020-07-20 Published:2020-08-28

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摘要： 目的 研究不同Notch受体在调控鼻咽癌顺铂耐药中的作用及相关机制。 方法 蛋白免疫印迹检测鼻咽癌及顺铂耐药鼻咽癌细胞(5-8F,5-8F/CDDP)Notch受体表达;流式细胞术检测不同浓度DAPT联合顺铂对5-8F/CDDP细胞凋亡影响;流式细胞术检测DAPT抑制Notch信号通路后5-8F/CDDP细胞周期变化;蛋白免疫印迹检测肿瘤耐药调控相关蛋白表达变化,统计学处理数据。 结果 5-8F/CDDP细胞中Notch1和Notch4受体表达明显高于5-8F细胞(P=0.003,P=0.004);不同浓度DAPT作用5-8F/CDDP细胞后,顺铂诱导的细胞凋亡率显著上升,耐药细胞增殖受到抑制,且与DAPT浓度呈剂量效应关系(P<0.05);DAPT抑制5-8F/CDDP细胞Notch信号通路后,细胞周期调控相关蛋白CyclinE和CDK-2蛋白表达明显下降,且伴有细胞周期G1/S阻滞(P<0.05);同时EMT调控相关蛋白Slug及DNA切除修复蛋白ERCC1表达下降,E-Cadherin表达上调。 结论 Notch1和Notch4受体表达上调与鼻咽癌细胞顺铂耐药相关,抑制Notch信号通路激活,可通过抑制肿瘤细胞EMT过程,而非细胞周期G1/S阻滞,提高耐药鼻咽癌细胞顺铂化疗敏感性。

关键词: Notch受体, 细胞周期, 上皮-间质转化, 鼻咽癌顺铂耐药

Abstract: Objective The objective of this study was to analyze the effects and possible regulatory mechanisms that Notch receptors could have on cisplatin resistance, observed in nasopharyngeal carcinoma. Methods Western blot analysis was used for the detection of Notch receptor expression in nasopharyngeal carcinoma cells and cisplatin-resistant nasopharyngeal carcinoma cells(5-8F, 5-8F/CDDP). Flow cytometry was used to investigate how the combined treatment with 10 μM CDDP and different concentrations of γ-secretase inhibitor(DAPT)could affect apoptosis in 5-8F/CDDP cells. Flow cytometry was also used for the detection of cell cycle stages in DAPT-treated 5-8F / CDDP cells. Finally, western blotting was also used for the detection of drug resistance-related protein expression. All experiments were followed by statistical data analysis. Results We observed significantly higher Notch1 and Notch4 receptor expression in 5-8F/CDDP cells than in 5-8F cells(P=0.003, P=0.004). Furthermore, we described that Notch signaling was inhibited by DAPT in 5-8F/CDDP cells, followed by a significant increase in the apoptosis rate and decrease in cell proliferation, induced by cisplatin in a dose-dependent manner(P<0.05). Moreover, after inhibiting the Notch signaling pathway in 5-8F/CDDP cells, DAPT treatment significantly decreased the expression of Cyclin E and CDK-2, proteins involved in cell cycle regulation, and contributed to blocking the cells in the G1/S phase(P<0.05). At the same time, the expression levels of both the EMT-related protein Slug and the DNA excision repair protein ERCC1 significantly decreased, while that of E-Cadherin was up-regulated. Conclusion Up-regulated expression of Notch1 and Notch4 receptors is associated with cisplatin resistance in nasopharyngeal carcinoma cells. The inactivation of the Notch signaling pathway might thus have the potential to enhance the efficiency of cisplatin chemotherapy in drug-resistant nasopharyngeal carcinoma cells by inhibiting EMT rather than blocking the G1/S cell cycle phase.

Key words: Notch receptors, Cell cycle, EMT, Cisplatin resistance of nasopharyngeal carcinoma

中图分类号: 

  • R739.6
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