文章摘要
赵孝通,李培华,刘方舟,钱亦淳,姚 瑶,张 园.siRNA沉默Gankyrin基因对喉鳞癌Hep-2细胞生物学行为的影响[J].南京医科大学学报,2016,(3):287~292
siRNA沉默Gankyrin基因对喉鳞癌Hep-2细胞生物学行为的影响
Effect of silencing Gankyrin by small interfering RNA on the biological behavior of laryngeal carcinoma cell lines Hep-2
投稿时间:2015-09-21  
DOI:10.7655/NYDXBNS20160307
中文关键词: 喉肿瘤  细胞增殖  细胞周期  细胞凋亡  Gankyrin
英文关键词: laryngeal neoplasms  cell proliferation  cell cycle  apoptosis  Gankyrin
基金项目:江苏省自然科学基金(BK20151561)
作者单位
赵孝通 徐州医学院研究生学院,江苏 徐州 221000 
李培华 徐州医学院附属医院耳鼻咽喉头颈外科,江苏 徐州 221000 
刘方舟 南京医科大学附属江苏省肿瘤医院头颈外科,江苏 南京 210009 
钱亦淳 南京医科大学附属江苏省肿瘤医院头颈外科,江苏 南京 210009 
姚 瑶 南京医科大学附属江苏省肿瘤医院头颈外科,江苏 南京 210009 
张 园 南京医科大学附属江苏省肿瘤医院头颈外科,江苏 南京 210009 
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中文摘要:
      目的:观察小干扰RNA(small interference RNA,siRNA)沉默Gankyrin基因对喉鳞癌Hep-2细胞的生物学行为影响。方法:用siRNA沉默喉鳞癌Hep-2细胞中的Gankyrin基因,运用实时定量反转录聚合酶链反应(qRT-PCR)及蛋白印迹法(Western blot)技术检测转染前后Gankyrin mRNA及蛋白的表达。采用CCK-8法检测细胞增殖能力,流式细胞仪检测细胞凋亡及细胞周期,细胞划痕损伤实验和Transwell小室实验检测细胞迁移能力,Matrigel侵袭实验检测细胞侵袭能力,Western blot检测Gankyrin下调后p53蛋白的表达。结果:qRT-PCR和Western blot结果显示,Gankyrin siRNA组Gankyrin mRNA和蛋白的相对表达量均低于对照siRNA组和未处理组(P < 0.001)。CCK-8法显示Gankyrin siRNA组细胞增殖速度明显下降(P < 0.001)。流式细胞仪检测结果表明,Gankyrin siRNA组细胞凋亡率[(7.70 ± 1.12)%]明显高于对照siRNA组[(2.34 ± 0.32)%]和未处理组[(1.82 ± 0.29)%],差异有统计学意义(P < 0.001);与两对照组相比,Gankyrin siRNA组G1期比例增加,S期比例减少,差异有统计学意义 (P < 0.001)。细胞划痕损伤实验和Transwell小室实验显示Gankyrin siRNA组的细胞迁移能力明显减弱(P < 0.01);Matrigel侵袭实验显示Gankyrin siRNA组细胞侵袭能力与对照siRNA组和未处理组比较,差异无统计学意义(P > 0.05)。Gankyrin表达下调增加了Hep-2细胞中p53蛋白的表达,差异有统计学意义(P < 0.001)。结论:Gankyrin表达下调能抑制Hep-2细胞的增殖和迁移,可能与细胞凋亡?细胞周期改变及p53的表达密切相关。
英文摘要:
      Objective:To investigate the effect of siRNA (small interference RNA) silencing Gankyrin on the behavior of human laryngeal carcinoma cell lines Hep-2. Methods:The expression of mRNA and protein levels of Gankyrin was detected by using qRT-PCR and Western blot before and after transfection,respectively. The cell proliferation was measured by CCK-8 assay. The apoptosis rate and cell cycle of Hep-2 cells were determined by flow cytometry. Cell migration was detected by wound healing assay and transwell assay. Matrigel assay was performed to observe cell invasive ability. The expression of p53 protein after down-regulation of Gankyrin was detected by Western blot. Results:Compared with the negative control group and the blank control group,the expressions of Gankyrin mRNA and protein were downregulated in the Gankyrin siRNA group,and the differences were statistically significant (P < 0.001). CCK-8 assay showed that the proliferation rate of the Gankyrin siRNA group was significantly decreased (P < 0.001). Flow cytometry showed that the apoptosis rate of the Gankyrin siRNA group[(7.70 ± 1.12)%]was significantly higher than that in the negative control group[(2.34 ± 0.32)%]and the blank control group[(1.82 ± 0.29)%],and the differences were statistically significant (P < 0.001). Compared with the two controls,G1 phase cells of the Gankyrin siRNA group were significantly increased,S phase cells were significantly decreased,and the difference was statistically significant (P < 0.001). Wound healing assay and transwell assay showed that the cell migration ability in the Gankyrin siRNA group was decreased significantly (P < 0.01). Matrigel assay showed that Gankyrin siRNA did not impact the invasive ability of Hep-2 cell (P > 0.05). The expression of p53 protein in the Gankyrin siRNA group was increased (P < 0.001). Conclusion:Down-regulation of Gankyrin inhibited the proliferation and migration ability of Hep-2 cell,which may be associated with the alteration of apoptosis,cell cycle and the expression level of p53.
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