中国医科大学学报

中国医科大学学报
  • 中文核心期刊
  • 中国科技核心期刊
  • 中国高校百佳科技期刊
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中国医科大学学报 ›› 2019, Vol. 48 ›› Issue (3): 225-229.doi: 10.12007/j.issn.0258-4646.2019.03.007

• 论著 • 上一篇    下一篇

CNOT7基因敲减通过减少HepG2细胞TGF-β1分泌影响免疫微环境

郭舜1, 赵海潮2, 任晓静2, 任崇仁2, 贺杰峰2, 赵浩亮2   

  1. 1. 山西医科大学研究生院, 太原 030001;
    2. 山西医科大学附属大医院普通外科, 太原 030001
  • 收稿日期:2018-10-22 出版日期:2019-03-30 发布日期:2019-03-06
  • 通讯作者: 赵浩亮 E-mail:haoliangzhao@hotmail.com
  • 作者简介:郭舜(1990-),男,硕士研究生.
  • 基金资助:
    山西省自然科学基金(2015011116)

Effect of CNOT7 Gene Knockdown on the Immune Microenvironment of HepG2 Cells by Reduced TGF-β1 Secretion

GUO Shun1, ZHAO Haichao2, REN Xiaojing2, REN Chongren2, HE Jiefeng2, ZHAO Haoliang2   

  1. 1. Graduate School, Shanxi Medical University, Taiyuan 030001, China;
    2. Department of General Surgery, Shanxi Dayi Hospital Affiliated to Shanxi Medical University, Taiyuan 030001, China
  • Received:2018-10-22 Online:2019-03-30 Published:2019-03-06

摘要: 目的 研究人CCR4-NOT转录复合体亚基7(CNOT7)基因敲减对肝母细胞瘤细胞系HepG2免疫微环境的影响并探讨其意义。方法 设计细胞转染方案,将实验分为3组:靶向敲减CNOT7组、阴性对照组和过表达CNOT7组,获得各组细胞后,用倒置荧光显微镜观察细胞转染效率。Western blotting方法检测CNOT7、转化生长因子-β1(TGF-β1)及核转录因子-κB(NF-κB)p65蛋白的表达水平,ELISA法测定细胞培养上清液中TGF-β1水平。流式细胞术检测转染后肿瘤细胞对自然杀伤(NK)细胞杀伤功能的敏感性。结果 与阴性对照组相比,靶向敲减CNOT7组TGF-β1和NF-κB p65蛋白表达水平显著降低,过表达CNOT7组TGF-β1和NF-κB p65蛋白表达水平显著升高;靶向敲减CNOT7组细胞培养上清液中TGF-β1水平显著降低,过表达CNOT7组TGF-β1水平显著升高;NK细胞与肿瘤细胞共培养,靶向敲减CNOT7组细胞凋亡率显著升高,过表达CNOT7组细胞凋亡率显著降低。结论 CNOT7基因参与肝细胞癌免疫微环境的形成,靶向敲减CNOT7基因可减少TGF-β1分泌,增强NK细胞对肝癌细胞的杀伤功能。

关键词: 肝细胞癌, 免疫微环境, 转化生长因子-β1, CCR4-NOT转录复合体亚基7

Abstract: Objective To study the effect of human CCR4-NOT transcription complex subunit 7 (CNOT7) gene knockdown on the immune microenvironment of HepG2 cells and explore its significance. Methods We designed a cell transfection protocol and performed the experiment with three groups:CNOT7-targeted knockdown group,control group,and CNOT7 overexpression group. The transfection efficiency was assessed using inverted fluorescence microscopy,and the expression level of CNOT7,transforming growth factor-β1 (TGF-β1),and nuclear factor-kappa B (NF-κB) p65 proteins was determined by western blotting. The concentration of TGF-β1 secreted in the cell culture supernatant was measured by ELISA. The sensitivity of tumor cells to the killing function of natural killer (NK) cells was detected by flow cytometry. Results Compared with the control group,the expression level of TGF-β1 and NF-κB p65 proteins was significantly decreased in the CNOT7-targeted knockdown group,and the TGF-β1 concentration in the culture supernatant was also significantly reduced. However,in the CNOT7 overexpression group,the expression level of the two proteins and TGF-β1 concentration were significantly increased. NK cells were co-cultured with tumor cells,and the apoptosis rate of HepG2 cells transfected with CNOT7-specific shRNA was significantly increased. However,in the CNOT7 overexpression group,the apoptosis rate was significantly decreased. Conclusion CNOT7 forms the immune microenvironment of hepatocellular carcinoma. Targeted knockdown of CNOT7 can reduce TGF-β1 secretion and enhance the killing function of NK cells toward HepG2 cells.

Key words: hepatocellular carcinoma, immunomicroenvironment, transforming growth factor-β1, CCR4-NOT transcription complex subunit 7

中图分类号: 

  • R735.7
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