摘要
目的探讨Notchl对喉鳞状细胞癌细胞系Hep-2细胞周期、细胞凋亡及细胞迁移能力的影响，并研究其可能的分子作用机制。方法将细胞分为对照组、空载体组和Notchl组，通过CCK-8增殖实验和流式细胞术检测Notchl对细胞增殖、细胞周期和细胞凋亡的影响，采用Boyden小室体外迁移实验检测Notchl对Hep-2细胞迁移的影响，采用半定量逆转录聚合酶链反应（1it-PCR）和Westernblot法检测细胞增殖、细胞周期、细胞凋亡及细胞迁移相关分子表达的变化。结果与对照组和空载体组比较，Notchl组中Hep-2细胞增殖受到明显抑制（P〈0．05）。Notchl组中Go／G，期细胞的比例[（70．43±1．36）％]明显高于对照组[（46．39±1．19）％]和空载体组[（48．41±1．18）％]，差异有统计学意义（P=0．000）。Notchl组的细胞凋亡率[（22．464±0．90）％]明显高于对照组[（5．77±0．37）％]和空载体组[（6．09±0．20）％]，差异有统计学意义（P=0．000）。Notchl组的穿膜细胞数为（41．75±6．13）个，与对照组[（113．634-7．19）个]和空载体组[（110．89±6．37）个]相比，明显减少（P〈0．05）。半定量RT-PCR和Westernblot检测结果显示，细胞增殖抑制和细胞周期阻滞与细胞周期蛋白D1（cyclinD1）、cyclinE和细胞周期蛋白依赖性激酶2（CDK2）表达的下调以及p53表达的上调密切相关。Notchl表达上调介导的细胞凋亡与半胱天冬酶（caspase）3和caspase9表达上调和Bcl-2表达下调有极其密切的联系，而Hep-2迁移能力的降低与基质金属蛋白酶（MMP）-2和MMP-9表达下调明显相关。结论Notchl信号途径可能在喉鳞状细胞癌的发生发展中具有十分重要的作用，有望成为喉鳞状细胞痛治疗的分子靶点。
Objective To investigate the effect of Notchl on cell cycle, apoptosis and migration of laryngeal squamous cell carcinoma cell line Hep-2 and explore its possible molecular mechanism. Methods Hep-2 ceils were divided into three groups： untreated group, empty vector group and Notchl group. The effects of upregnlation of Notchl expression on cell proliferation, cell cycle and apoptosis were assessed by CCK-8 staining and flow cytometry, respectively, and effect of upregulation of Notchl expression on cell migration of Hep-2 cells was studied using Boyden chamber assay, and further expression changes of genes related to cell proliferation, cell cycle, apoptosis and migration were detected by semi-quantitative RT-PCR and Western blotting. Results Compared with the untreated group and empty vector group, cell proliferation of Hep-2 in the Notchl group was significantly inhibited （ P 〈 0.05 ）. The results of flow cytometry revealed that the percentage of ceils at G0/G1 phase in the Notchl group was （70.43±1.36）%, significantly higher than the （46.39±1.19） % in the untreated group or （48.41±1.18） % in the empty vector group, and there was a significant difference among the three groups （P = 0. 000）. In addition, the percentage of apoptotic cells in the Notchl group was （22.46±0.90） %, significantly higher than that in the untreated group [ （ 5.77±0.37 ） % ] or empty vector group [ （ 6.09± 0.20 ） % ], with a significant difference among the three groups （P = 0. 000）. The results of Boyden chamber assay demonstrated that the number of cells migrated through membrane in the Notchl group was evidently lower than that in the untreated group and empty vector group. Moreover, the results of semi-quantitative RT-PCR and Western blotting showed that cell proliferation inhibition and cell cycle arrest were closely associated with downregulation of cyclin D1, cyclin E and CDK2 expressions and upregnlation of p53 expression. Inaddition, upregulation of Notchl expression mediated cell apoptosis was tightly related to upregulation of caspase 3/9 expressions and downregulation of Bcl-2, while the decrease of Hep-2 cell migration ability was obviously correlated with downregulation of MMP-2/-9 expressions. Conclusions Notchl signaling pathway may play a pivotal role in the occurrence and development of laryngeal squamous cell carcinoma. Further study may elucidate that Notchl signaling pathway may become a molecular target for therapy of laryngeal squamous cell carcinoma.
出处
《中华肿瘤杂志》
CAS
CSCD
北大核心
2012年第2期104-109,共6页
Chinese Journal of Oncology
