天津医药

天津医药 ›› 2018, Vol. 46 ›› Issue (10): 1039-1044.doi: 10.11958/20180237

• 细胞与分子生物学 • 上一篇    下一篇

甲基转移酶SETDB2对肝癌侵袭迁移的影响及其机制研究

贾龙梅 1，殷香宝 2，曾磊 3，陈新 1，饶燕飞 4   

  1. 基金项目：江西省青年科学基金资助项目（2008GQY0050） 作者单位：1南昌大学第二附属医院核医学科（邮编330006），2肝胆外科，3病理科，4检验科 作者简介：贾龙梅（1976），女，本科，主管技师，主要从事消化系统疾病的诊断及防治研究
  • 收稿日期:2018-02-09 修回日期:2018-07-03 出版日期:2018-10-15 发布日期:2018-11-09
  • 通讯作者: 贾龙梅 E-mail:jialongmei111@126.com

The effect and mechanism of methyltransferase SETDB2 on the invasion and migration of hepatoma carcinoma

JIA Long-mei1, YIN Xiang-bao2, ZENG Lei3, CHEN Xin1, RAO Yan-fei4   

  1. 1 Department of Nuclear Medicine, 2 Department of Hepatobiliary Surgery, 3 Department of Pathology, 4 Department of Clinical Laboratory, the Second Affiliated Hospital of Nanchang University, Jiangxi 330006, China
  • Received:2018-02-09 Revised:2018-07-03 Published:2018-10-15 Online:2018-11-09
  • Contact: Long-Mei JIA E-mail:jialongmei111@126.com

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摘要： 摘要：目的 检测甲基转移酶SETDB2对肝癌侵袭转移的影响并探讨其机制。方法 应用Western blot、免疫组 化以及 Real-time PCR 等方法检测 37 例肝癌及对应癌旁组织标本中 SETDB2 蛋白及 mRNA 表达情况。将靶向 SETDB2 的短发卡 RNA（sh-SETDB2）和空载体（sh-NC）分别转染至人肝癌 MHCC97H 细胞构建稳转细胞系，应用 Tanswell侵袭实验、细胞划痕实验检测2组细胞的侵袭和迁移能力。通过基因芯片检测敲低SETDB2后表达有变化 的基因。敲低SETDB2后，运用Real-time PCR检测PTEN的mRNA的表达变化以及Western blot检测PTEN和组蛋白 H3第9位赖氨酸的三甲基化（H3K9me3）的表达变化，CHIP 检测H3K9me3在PTEN 的启动子区域的变化。在敲低 SETDB2的同时敲低PTEN的表达，Tanswell侵袭实验检测细胞侵袭能力的变化。结果 （1）Western blot和Real-time PCR的结果均表明肝癌组织中SETDB2蛋白及mRNA表达水平均高于癌旁组织；SETDB2表达的高低与肝癌组织学 分级和TNM分期有关。（2）转染sh-SETDB2的MHCC97H细胞的侵袭和迁移能力均明显低于sh-NC组。（3）基因芯 片、Western blot以及Real-time PCR等实验结果显示敲低SETDB2后PTEN表达上调。（4）在敲低SETDB2后H3K9me3 表达下调，PTEN启动子区域的H3K9me3减少。（5）与敲低SETDB1组相比，敲低SETDB2的同时敲低PTEN后细胞的 侵袭能力恢复。结论 SETDB2通过下调PTEN的表达促进肝癌细胞的侵袭迁移。

关键词: 癌, 肝细胞, 组蛋白赖氨酸N-甲基转移酶, 肿瘤侵润, 细胞迁移分析, PTEN磷酸水解酶, SETDB2

Abstract: bstract: Objective To investigate the effect of methyltransferase SETDB2 on the invasion and metastasis of live cancer and explore its mechanism. Methods Western blot, immunohistochemistry and Real-time PCR assays were performed to detect the expressions of SETDB2 protein and mRNA in liver cancer and the adjacent tissue specimens of 37 cases. The short hair pair RNA targeting SETDB2 (sh-SETDB2) and the corresponding empty vector (sh-NC) were transfected into MHCC97H cells. The invasion and migration ability of the two groups of cells were detected by Tanswell invasion assay and wound-healing assay. The gene chip was used to detect the expression of the altered genes after knocking down SETDB2. The mRNA expression of PTEN was detected by Real-time PCR. Western blot assay was used to detect the protein expressions of PTEN and H3K9me3. CHIP was used to detect the change of H3K9me3 in the promoter region of PTEN. The expression of PTEN was knocked down in shSETDB2-MHCC97H cells, and the invasion ability of cells was detected by the Tanswell invasion assay. Results (1) The results of Western blot and Real-time PCR assays showed that the expression levels of SETDB2 protein and mRNA were higher in the liver cancer tissues than those in the adjacent normal tissues. The expression level of SETDB2 was related to the histological grade and TNM stage of liver cancer. (2) The invasion and migration abilities were significantly lower in MHCC97H cells transfected with sh-SETDB2 than those in sh-NC group. (3) Microarray experiment, Western blot and Real-time PCR assays showed that PTEN was up-regulated after knocking down SETDB2. (4) The expression of H3K9me3 was decreased after knocking down SETDB2, and the appearance of H3K9me3 in PTEN promoter region was decreased as well. (5) Compared with the knockdown SETDB1, the invasive ability of cells was restored after both SETDB2 and PTEN were knocked down. Conclusion SETDB2 promotes the invasion and migration of hepatoma cells by downregulating the expression of PTEN.

Key words: carcinoma, hepatocellular, histone-lysine N-methyltransferase, neoplasm invasiveness, cell migration assays, PTEN phosphohydrolase, SETDB2