中华皮肤科杂志 ›› 2014, Vol. 47 ›› Issue (6): 413-416.

• 论著 • 上一篇    下一篇

RNA干扰下调PLK1基因对恶性黑素瘤细胞侵袭和失巢凋亡的影响

丁克云,徐娟,满昌峰,范钰   

  1. 江苏大学附属人民医院
  • 收稿日期:2013-07-01 修回日期:2013-10-18 发布日期:2014-06-01
  • 通讯作者: 范钰 E-mail:zjfany36@gmail.com
  • 基金资助:
    江苏省镇江市社会发展基金

Impacts of RNA interference targeting the polo-like kinase-1 gene on the invasion of and anoikis in human malignant melanoma cells

  • Received:2013-07-01 Revised:2013-10-18 Published:2014-06-01
  • Contact: Yu Fan E-mail:zjfany36@gmail.com

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摘要： 目的 探讨RNA干扰技术下调PLK1基因对恶性黑素瘤细胞侵袭的影响和可能机制。 方法 用PLK1小干扰核糖核酸分子（siRNA）转染人恶性黑素瘤A375细胞后，分别用实时定量PCR和 Western 印迹法检测PLK1mRNA和蛋白表达，Transwell方法检测细胞侵袭能力，以平板集落形成方法了解癌细胞集落形成情况，分别用琼脂糖凝胶电泳和TUNEL方法检测癌细胞失巢凋亡情况。 结果 恶性黑素瘤A375细胞经PLK1 siRNA转染后，PLK1 mRNA和蛋白表达明显下调；siRNA转染组癌细胞生长明显受到抑制。与空白对照组和脂质体对照组比较，siRNA细胞转染组侵袭性明显下降。失巢凋亡检测发现，琼脂糖凝胶电泳出现明显的梯度图谱。TUNEL结果显示，PLK1 siRNA可诱导恶性黑素瘤A375细胞凋亡，siRNA细胞转染组较空白对照组和脂质体对照组凋亡指数明显升高［3.86% ± 0.35%（3.125 nmol/L siRNA组），7.35% ± 0.36%（6.250 nmol/L siRNA组），17.56% ± 0.38%（12.500 nmol/L siRNA组）比1.15% ± 0.25%和1.18% ± 0.22%），均P < 0.01］。 结论 PLK1 siRNA可抑制恶性黑素瘤细胞的侵袭，其机制可能与诱导失巢凋亡有关。

关键词: 黑色素瘤, RNA干扰, 基因，PLK1, 肿瘤浸润

Abstract: Ding Keyun, Xu Juan, Man Changfeng, Fan Yu. Cancer Institute, Affiliated People′s Hospital of Jiangsu University, Zhenjiang 212002, Jiangsu, China Corresponding author: Fan Yu, Email: yuf36@sina.com 【Abstract】 Objective To investigate the effects of down-regulation of polo-like kinase-1 （PLK1） gene by RNA interference （RNAi） on the invasion of a human malignant melanoma cell line A375 and their possible mechanisms. Methods Cultured A375 cells were classified into several groups: blank control group receiving no treatment, liposome group transfected with lipofectamine only, and three siRNA groups transfected with three concentrations of a small interference RNA （siRNA） targeting PLK1 respectively. After additional culture, real time quantitative PCR and Western blot analysis were performed to quantify the expressions of PLK1 mRNA and protein in A375 cells respectively, Transwell invasion assay to evaluate the invasive capacity of A375 cells, agarose gel electrophoresis and terminal deoxynucleotidyl-transferase-mediated dUTP-biotin nick end labelling（TUNEL） to detect anoikis in A375 cells. The colony-forming capacity was also evaluated for A375 cells. Statistical analysis was carried out by one-factor analysis of variance. Results There was a significant decrease in PLK1 mRNA and protein expressions as well as in colony-forming units in the siRNA groups compared with the blank control group （all P < 0.05）. The invasive capacity of A375 cells was significantly inhibited in the siRNA groups with the number of migrating cells in Transwell assay being 39 ± 5, 19 ± 5 and 9 ± 3 in A375 cells transfected with 3.125, 6.250 and 12.500 nmol/L siRNAs respectively, compared to 56 ± 5 in the blank control group （all P < 0.05）. A characteristic DNA ladder was observed on agarose gel electrophoresis in the siRNA （6.250 nmol/L） group. Compared with the blank control group and liposome group, the three siRNA groups showed increased apoptotic index （3.86% ± 0.35% （3.125 nmol/L siRNA）, 7.35% ± 0.36% （6.250 nmol/L siRNA） and 17.56% ± 0.38% （12.500 nmol/L siRNA） vs. 1.15% ± 0.25% （blank control group） and 1.18% ± 0.22% （liposome group）, all P < 0.05）. Conclusions PLK1 siRNA can inhibit the invasion of malignant melanoma cells, likely by inducing anoikis in these cells.

Key words: Melanoma, RNA interference, Genes, PLK1, Neoplasm invasiveness

引用本文

丁克云 徐娟 满昌峰 范钰. RNA干扰下调PLK1基因对恶性黑素瘤细胞侵袭和失巢凋亡的影响[J]. 中华皮肤科杂志, 2014,47(6):413-416. doi: