解剖学报

解剖学报 ›› 2019, Vol. 50 ›› Issue (4): 465-470.doi: 10.16098/j.issn.0529-1356.2019.04.010

• 肿瘤生物学 • 上一篇    下一篇

5-氮杂-2’-脱氧胞苷通过反转原钙黏蛋白10表达抑制人乳腺癌细胞系MDA-MB-231的侵袭和迁移

张微1,2 朱文斌1 岳丽玲1 刘立琨1*   

  1. 1.齐齐哈尔医学院医药科学研究院，黑龙江 齐齐哈尔 161006; 2.佳木斯大学药学院，黑龙江 佳木斯 154007
  • 收稿日期:2018-09-30 修回日期:2018-11-29 出版日期:2019-08-06 发布日期:2019-08-06
  • 通讯作者: 刘立琨 E-mail:llk_2008@126.com
  • 基金资助:
    乳腺癌全基因组甲基化差异分析及其交互作用通路研究;PCDH10基因启动子甲基化与乳腺癌发生发展的相关性研究;PCDH10基因及其甲基化状态对乳腺癌生长转移的调控机制研究

Effect of protocadherins 10 re-expression induced by 5-aza-2’-deoxycytidine on invasion and migration capacity of breast cancer cell line MDA-MB-231 and its mechanism

ZHANG Wei 1,2 ZHU Wen-bin1 YUE Li-ling1 LIU Li-kun 1*   

  1. 1.Research Institute of Medicine and Pharmacy, Qiqihar Medical University, Heilongjiang Qiqihar 161006, China; 2.Pharmacy School, Jiamusi University, Heilongjiang Jiamusi 154007, China
  • Received:2018-09-30 Revised:2018-11-29 Online:2019-08-06 Published:2019-08-06
  • Contact: LIU Li-kun E-mail:llk_2008@126.com

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摘要：

目的 探讨5-氮杂-2’-脱氧胞苷（5-Aza-CdR）诱导抑癌基因原钙黏蛋白10（PCDH10）重新表达对人乳腺癌细胞MDA-MB-231体外侵袭迁移能力的影响并初步探讨其机制。 方法 体外培养人乳腺癌细胞MDA-MB-231，设置对照组和5-Aza-CdR药物处理组，分别采用反转录聚合酶链反应（RT-PCR）检测PCDH10 mRNA 的表达水平；Transwell法和划痕实验检测细胞的侵袭迁移能力；Western blotting检测PCDH10、DNA甲基转移酶（DNMT）3A、DNMT3B、核因子（NF）-κB p65和基质金属蛋白酶（MMP）-2、MMP-9蛋白表达的变化。 结果 5-Aza-CdR能够反转PCDH10的mRNA和蛋白表达；PCDH10表达恢复后MDA-MB-231细胞的侵袭迁移能力受到抑制；Western blotting检测发现，MDA-MB-231细胞经5-Aza-CdR处理后DNMT3A、DNMT3B、NF-κB p65、MMP-2和MMP-9的表达下调。 结论 5-Aza-CdR可抑制MDA-MB-231细胞DNMT3A和DNMT3B的表达，使抑癌基因PCDH10表达恢复，从而通过阻滞NF-κB p65的活化，下调MMP-2和MMP-9表达而抑制乳腺癌细胞的侵袭转移。

关键词: 乳腺癌, 原钙黏蛋白10, 甲基化, 侵袭, 迁移, 反转录聚合酶链发应, 免疫印迹法, 人

Abstract:

Objective To investigate whether re-expression of protocadherin 10(PCDH10) induced by 5-aza-2’-deoxycytidine (5-Aza-CdR) could affect the invasion and migration of MDA-MB-231 cells, and to explore the possible mechanism. Methods Human breast cancer cell line MDA-MB-231 was cultured in vitro. Control group and 5-Aza-CdR treatment group were set up. PCDH10 mRNA expression in MDA-MB-231 cell line was determined by reverse transcription-polymerase chain reaction (RT-PCR); Transwell chamber and wound healing assay were performed to measure the invasion and migration capacity of the cells, and protein expression of PCDH10, DNA methyltransferase(DNMT)3A, DNMT3B, nuclear factor(NF)-κB p65, matrix metalloproteinases(MMP)-2 and MMP-9 were detec Western blotting. Results 5-Aza-CdR could reverse the methylation status of PCDH10 gene in MDA-MB-231 cells in a dose-dependent manner. Re-expression of PCDH10 significantly inhibited cell invasion and migration capacity in vitro. Western blottoing analysis revealed that the expression of DNMT3A, DNMT3B, NF-κB p65, MMP-2 and MMP-9 in MDA-MB-231 cells were down-regulated after exposure to 5-Aza-CdR. Conclusion Re-expression of PCDH10 significantly inhibits MDA-MB-231 invasion and migration capacity. The inhibitory effect is characterized that 5-Aza-CdR treatment down-regulates DNMT3A and DNMT3B levels, recovers the expression of anti-oncogene PCDH10, further blocks the activation of NF-κB p65, resultsing in a decrease in the secretion of MMP-2 and MMP-9.

Key words: Breast cancer, Protocadherin 10, Methylation, Invasion, Migration, RT-PCR, Western blotting, Human