中国临床解剖学杂志 ›› 2018, Vol. 36 ›› Issue (3): 282-287.doi: 10.13418/j.issn.1001-165x.2018.03.013

• 实验研究 • 上一篇    下一篇

SETD4对脂多糖诱导的AML12细胞IL-6转录的调控作用

孙江, 李月, 牛世贤, 黄梦怡, 钟玙沄, 赵舒祺, 罗海华, 姜勇, 刘靖华   

  1. 南方医科大学基础医学院病理生理学教研室,广东省蛋白质组学重点实验室,  广州   510525
  • 收稿日期:2018-03-01 出版日期:2018-05-25 发布日期:2018-07-04
  • 通讯作者: 刘靖华,教授,博士生导师,Tel:(020)61648172-810,E-mail: liujhua@smu.edu.cn
  • 作者简介:孙江(1992-),在读硕士,主要从事脓毒症发生机制的研究,Tel:18565314585,E-mail:1181313426@qq.com
  • 基金资助:

    国家自然科学基金项目(81471901,81072425);广东省自然科学基金重点项目(2015A030311031)

Effect of SETD4 on LPS-induced IL-6 transcription in AML12 cells

SUN Jiang, LI Yue, NIU Shi-xian, HUANG Meng-yi, ZHONG Yu-yun, ZHAO Shu-qi, LUO Hai-hua, JIANG Yong, LIU Jing-hua   

  1. Key laboratory for Functional Proteomics of Guangdong Province,Department of Pathophysiology,School of Basic Medical Science, Southern Medical University,Guangzhou 510515
  • Received:2018-03-01 Online:2018-05-25 Published:2018-07-04

摘要:

目的 探讨SETD4(SET-domain containing protein 4)对脂多糖(LPS)诱导的AML12(小鼠肝脏细胞系)细胞IL-6的转录调控作用。  方法 构建SETD4表达质粒和IL-6 启动子荧光素酶报告基因质粒,共转染小鼠肝脏细胞系AML12,使用双荧光素酶报告基因技术检测LPS刺激后IL-6 启动子荧光素酶活性;单独转染SETD4表达质粒上调AML12细胞SETD4表达,检测LPS刺激后IL-6 mRNA水平变化。  结果 双酶切鉴定及核酸测序证实重组质粒pcDNA3.0/HA-SETD4、pGL3.0/IL-6 promoter的构建成功。pcDNA3.0/HA-SETD4与pGL3.0/IL-6 promoter共转染AML12细胞,LPS刺激后SETD4对IL-6 启动子荧光素酶活性没有影响;上调SETD4表达后,可以促进LPS诱导的IL-6 mRNA转录。  结论 成功构建SETD4真核表达质粒和IL-6启动子荧光素酶报告基因质粒;SETD4对LPS诱导的AML12细胞IL-6的转录发挥正调控作用。

关键词: SETD4,  LPS,  AML12,  IL-6 转录,  荧光素酶报告基因

Abstract:

Objective To explore the effect of SETD4 (SET-domain containing protein 4) in IL-6 transcription in AML12 cells (murine hepatocyte line) stimulated with LPS. Methods Recombination plasmids encoding SETD4 or IL-6 promoter luciferase were constructed and cotransfected into AML12 cells. After LPS stimulation, the luciferase activity of IL-6 promoter was detected by Dual-luciferase reporter assay system. The LPS-induced IL-6 mRNA in AML12 cells with SEDT4 over-expression was also measured. Results The plasmids of pcDNA3.0/HA-SETD4 and pGL3.0/IL-6 promoter were constructed successfully, evidenced by double restriction enzyme digestion identification and DNA sequencing. The results showed that SETD4 have no effect on luciferase activity when IL-6 promoter luciferase plasmid and SETD4 plasmid were cotransfected into the AML12 cells following LPS stimulation. Over-expression of SETD4 could up-regulate the LPS-induced IL-6 mRNA transcription in AML12 cells. Conclusion SETD4 expression plasmid and IL-6 promoter luciferase plasmid were constructed successfully. SETD4 may play a positive effect on LPS-induced IL-6 transcription in AML12 cells.

Key words:  , SETD4; LPS; AML12; IL-6 transcription; Luciferase reporter gene