上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

Ajuba对肝糖异生相关基因表达的影响

樊红延1，侯照远2，董莹1   

  1. 上海交通大学 1.医学院附属仁济医院内分泌科， 上海 200127； 2.基础医学院生化与分子细胞生物学系, 上海 200025
  • 出版日期:2015-05-28 发布日期:2015-06-04
  • 通讯作者: 董莹, 电子信箱: 13472519882@163.com。
  • 作者简介:樊红延（1990—）， 女， 硕士生; 电子信箱: hy.fan@163.com。
  • 基金资助:

    国家自然科学基金项目（81372028）；上海市科委医学引导项目（124119a5700）

Effects of Ajuba on expressions of genes relevant to gluconeogenesis

FAN Hong-yan1, HOU Zhao-yuan2, DONG Ying1   

  1. 1.Department of Endocrinology, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200127, China; 2.Department of Biochemistry and Molecular Cell Biology, Basic Medical College, Shanghai Jiao Tong University, Shanghai 200025, China
  • Online:2015-05-28 Published:2015-06-04
  • Supported by:

    National Natural Science Foundation of China, 81372028; Medical Guidance Project of Science and Technology Commission of Shanghai Municipality, 124119a5700

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摘要：

目的 观察Lim家族成员Ajuba对糖异生关键基因转录水平的影响，探讨Ajuba参与肝糖代谢调节的分子机制。方法 ①慢病毒感染人肝癌细胞系HepG2，筛选低表达和过表达Ajuba的稳转细胞，通过Western blotting鉴定Ajuba表达情况。②real-time PCR检测HepG2稳转细胞中糖异生关键酶磷酸烯醇式丙酮酸羧激酶（PEPCK）和葡萄糖-6-磷酸酶（G6P）基因mRNA表达。③分离培养C57BL/6小鼠原代肝细胞并以慢病毒感染法筛选低表达Ajuba的细胞，real-time PCR检测Pepck和G6p mRNA水平。结果 ①Western blotting结果显示，低表达和高表达Ajuba的HepG2细胞均构建成功。②与对照组相比，低表达Ajuba的HepG2细胞中PEPCK和G6P mRNA水平明显降低，而过表达细胞中则显著升高（P<0.05）。③低表达Ajuba的小鼠原代肝细胞中Pepck和G6p mRNA水平明显降低（P<0.05）。结论 Lim蛋白Ajuba能够通过促进肝糖异生关键基因PEPCK和G6P的表达参与糖代谢调节，且该作用不具有种属特异性。

关键词: Ajuba, 糖异生, HepG2, 小鼠原代肝细胞, 磷酸烯醇式丙酮酸羧激酶, 葡萄糖-6-磷酸酶

Abstract:

Objective To observe the effects of Lim protein Ajuba on transcriptional levels of key genes relevant to gluconeogenesis and explore the molecular mechanism of regulating the liver glycogen metabolism by Ajuba. Methods ①Human hepatoma cells HepG2 were infected with lentivirus and stable transfected cells with low/high expression of Ajuba were screened. The expression of Ajuba was detected by the Western blotting. ②Expressions of two key enzymes relevant to gluconeogenesis, i.e. phosphoenolpyruvate carboxykinase (PEPCK) and glucose-6-phosphatase (G6P), were detected by the real-time PCR. ③Primary hepatocytes of C57BL/6 mice were isolated and cells with low expression of Ajuba were screened by the lentivirus infection method. The mRNA expressions of Pepck and G6p were detected by the real-time PCR. Results ①The results of Western blotting showed that HepG2 cells with low and high expressions of Ajuba were successfully screened. ②Compared with controls, mRNA levels of PEPCK and G6P in HepG2 cells with low expression of Ajuba significantly decreased, while mRNA levels of PEPCK and G6P in HepG2 cells with high expression of Ajuba significantly increased (P<0.05). ③The mRNA levels of Pepck and G6p in mouse primary hepatocytes with low expression of Ajuba significantly decreased (P<0.05). Conclusion The Lim protein Ajuba can participate in the regulation of glucose metabolism by up-regulating the expressions of PEPCK and G6P, two key genes relevant to gluconeogenesis. This effect is not species-specific.

Key words: Ajuba, gluconeogenesis, HepG2, mouse primary hepatocytes, phosphoenolpyruvate carboxykinase, glucose-6-phosphatase