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

作　　者：赵孝通 李培华[2] 刘方舟[3] 钱亦淳[3] 姚瑶[3] 张园[3] 

机构地区：[1]徐州医学院研究生学院,江苏徐州221000 [2]徐州医学院附属医院耳鼻咽喉头颈外科,江苏徐州221000 [3]南京医科大学附属江苏省肿瘤医院头颈外科,江苏南京210009

出　　处：《南京医科大学学报（自然科学版）》2016年第3期287-292,329,共7页Journal of Nanjing Medical University(Natural Sciences)

基　　金：江苏省自然科学基金(BK20151561)

摘　　要：目的 :观察小干扰RNA(small interference RNA,si RNA)沉默Gankyrin基因对喉鳞癌Hep-2细胞的生物学行为影响。方法:用si RNA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(q RT-PCR)及蛋白印迹法(Western blot)技术检测转染前后Gankyrin m RNA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:q RT-PCR和Western blot结果显示,Gankyrin si RNA组Gankyrin m RNA和蛋白的相对表达量均低于对照si RNA组和未处理组(P<0.001)。CCK-8法显示Gankyrin si RNA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 si RNA组细胞凋亡率[(7.70±1.12)%]明显高于对照si RNA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 si RNA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 si RNA组的细胞迁移能力明显减弱(P<0.01);Matrigel侵袭实验显示Gankyrin si RNA组细胞侵袭能力与对照si RNA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。Objective：To investigate the effect of si RNA（small interference RNA） silencing Gankyrin on the behavior of human laryngeal carcinoma cell lines Hep-2. Methods：The expression of m RNA and protein levels of Gankyrin was detected by using q RTPCR 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 m RNA and protein were downregulated in the Gankyrin si RNA group,and the differences were statistically significant（P〈0.001）. CCK-8 assay showed that the proliferation rate of the Gankyrin si RNA group was significantly decreased（P〈0.001）. Flow cytometry showed that the apoptosis rate of the Gankyrin si RNA 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 si RNA group were significantlyincreased,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 si RNA group was decreased significantly（P〈0.01）. Matrigel assay showed that Gankyrin si RNA did not impact the invasive ability of Hep-2 cell（P〈0.05）. The expression of p53 protein in the Gankyrin si RNA 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

关 键 词：喉肿瘤 细胞增殖 细胞周期 细胞凋亡 Gankyrin 

分 类 号：R739.65[医药卫生—肿瘤]