中华皮肤科杂志 ›› 2018, Vol. 51 ›› Issue (6): 447-450.doi: 10.3760/cma.j.issn.0412-4030.2018.06.013

• 研究报道 • 上一篇    下一篇

siRNA干扰Gadd45α表达对皮肤鳞状细胞癌A431细胞侵袭和迁移的影响

李小静1,李志锋2,韩昭2,燕霞2,李显平3,刘保国2,刘志军2   

  1. 1. 河北邯郸 河北工程大学附属医院
    2. 河北工程大学附属医院
    3. 河北邯郸 河北工程大学附属医院皮肤科
  • 收稿日期:2017-05-23 修回日期:2017-12-29 发布日期:2018-05-30
  • 通讯作者: 刘保国 E-mail:lbg66@163.com
  • 基金资助:
    河北省科技计划项目

Effects of of growth arrest and DNA damage inducible gene 45α interfered by small interference RNA on the invasion and migration of a cutaneous squamous cell carcinoma cell line A431

  • Received:2017-05-23 Revised:2017-12-29 Published:2018-05-30
  • Supported by:
    Science and Technology Planning Project of Hebei Province of China

PDF

25

摘要： 目的 探讨靶向沉默生长抑制和DNA损伤诱导45蛋白α（Gadd45α）对皮肤鳞状细胞癌（简称鳞癌）细胞株A431侵袭和迁移的影响。方法 采用实时荧光定量PCR、Western印迹检测A431和Colon16细胞Gadd45α基因和蛋白的表达，选取高表达Gadd45α的A431细胞株作为研究对象。将A431细胞分为实验组、阴性对照组和空白对照组：实验组转染Gadd45α?siRNA?1，阴性对照组转染阴性对照siRNA，空白对照组不做任何处理。采用实时荧光定量PCR和Western印迹法分别检测干预后Gadd45α mRNA和蛋白的表达， Transwell小室法及细胞划痕法检测干预对A431细胞株侵袭及迁移能力的影响。结果 A431细胞高表达Gadd45α基因和蛋白。干预后实验组A431细胞Gadd45α mRNA及蛋白相对表达量分别为0.286 ± 0.013、0.33 ± 0.007，明显低于阴性对照组组（1.028 ± 0.183、0.87 ± 0.002）以及空白对照组（1.001 ± 0.057、0.86 ± 0.004），差异均有统计学意义（F值分别为5 893.857、2 763.000，均P < 0.001）。实验组A431细胞株穿过聚碳酸酯膜的细胞数为66.33 ± 3.79，明显多于阴性对照组（26.00 ± 4.36）和空白对照组（28.33 ± 4.16），3组间差异有统计学意义（F = 20.084，P = 0.002）。划痕实验中，实验组每高倍视野下细胞迁移数为315.33 ± 6.66，明显多于阴性对照组（154.67 ± 2.08）和空白对照组（130.67 ± 3.51），3组间差异有统计学意义（F = 1 676.255，P < 0.001）。结论 A431细胞株高表达Gadd45α，靶向沉默Gadd45α表达能增强A431细胞株的侵袭和迁移能力。

关键词: 肿瘤, 鳞状细胞, RNA干扰, 肿瘤侵润, 细胞迁移分析, 生长抑制和DNA损伤诱导45蛋白α

Abstract: Li Xiaojing, Li Zhifeng, Han Zhao, Yan Xia, Li Xianping, Liu Baoguo, Liu Zhijun Depatment of Dermatology, Affiliated Hospital of Hebei University of Engineering, Handan 056002, Hebei, China Corresponding authors: Liu Baoguo, Email: LBG66@163.com; Liu Zhijun, Email: zlmdsh@126.com 【Abstract】 Objective To evaluate effects of targeted silencing of growth arrest and DNA damage inducible gene 45α （Gadd45α） by small interference RNA （siRNA） on the invasion and migration of a cutaneous squamous cell carcinoma cell line A431. Methods Real-time fluorescence-based quantitative PCR and Western blot analysis were performed to detect the mRNA and protein of A431 and Colon16 cells respectively, and then A431 cells with highly expressed Gadd45α served as a research object. Cultured A431 cells were divided into 3 groups: experimental group transfected with Gadd45α-siRNA-1, negative control group transfected with negative control siRNA, and blank control group receiving no treatment. After the RNA interference, the mRNA and protein of Gadd45α in the above 3 groups were measured by real-time fluorescence-based quantitative PCR and Western blot analysis, respectively, and the effect of the RNA interference on the invasion and migration abilities of A431 cells was evaluated by Transwell and wound scratch assays. Results Gadd45α mRNA and protein were highly expressed in the A431 cells. After the RNA interference, the experimental group showed markedly lower mRNA and protein s of Gadd45 in the A431 cells （0.286 ± 0.013, 0.33 ± 0.007, respectively） compared with the negative control group （1.028 ± 0.183, 0.87 ± 0.002, respectively）and blank control group （1.001 ± 0.057, 0.86 ± 0.004, respectively）, and there were significant differences in the mRNA and protein s of Gadd45 among the 3 groups （F = 5 893.857, 2 763.000, both P < 0.001）. The number of A431 cells crossing the polycarbonate membrane was higher in the experimental group （66.33 ± 3.79） than in the negative control group （26.00 ± 4.36） and the blank control group （28.33 ± 4.16）, and there was a significant difference among the 3 groups （F = 20.084, P = 0.002）. Moreover, the wound scratch assay showed that the number of migrating A431 cells per high-power field was significantly higher in the experimental group than in the negative control group and the blank control group （315.33 ± 6.66, 154.67 ± 2.08, 130.67 ± 3.51 respectively; F = 1 676.255, P < 0.001）. Conclusion Gadd45α is highly expressed in the cutaneous squamous cell carcinoma cell line A431, and targeted silencing of Gadd45α gene can enhance the in vitro invasion and migration abilities of A431 cells.

Key words: Neoplasms, squamous cell, RNA interference, Neoplasm invasiveness, Cell migration assays, Growth arrest and DNA damage-inducible 45α

引用本文

李小静 李志锋 韩昭 燕霞 李显平 刘保国 刘志军. siRNA干扰Gadd45α表达对皮肤鳞状细胞癌A431细胞侵袭和迁移的影响[J]. 中华皮肤科杂志, 2018,51(6):447-450. doi:10.3760/cma.j.issn.0412-4030.2018.06.013