GIP/GIPR经Akt-TCF4-GIPR正反馈增强GIP效应

doi:10.11843/j.issn.0366-6964.2020.09.009

畜牧兽医学报 ›› 2020, Vol. 51 ›› Issue (9): 2120-2129.doi: 10.11843/j.issn.0366-6964.2020.09.009

GIP/GIPR经Akt-TCF4-GIPR正反馈增强GIP效应

叶华琼^1,2, 安翠平², 张凯艺², 谢宁², 毕延震³, 杨述林^2*, 李升和^1*

1. 安徽科技学院动物科学学院, 滁州 233100;
2. 中国农业科学院北京畜牧兽医研究所, 北京 100193;
3. 湖北省农业科学院畜牧兽医研究所, 武汉 430064

收稿日期:2020-03-02 出版日期:2020-09-25 发布日期:2020-09-25
通讯作者: 杨述林,主要从事小型猪糖尿病模型研究,E-mail:yangshulin@caas.cn;李升和,主要从事动物营养调控与健康研究,E-mail:lish@ahstu.edu.cn
作者简介:叶华琼(1997-),女,安徽合肥人,硕士生,主要从事小型猪糖尿病模型研究,E-mail:1964453906@qq.com;安翠平(1992-),女,河北保定人,硕士生,主要从事小型猪糖尿病模型研究,E-mail:ancuiping1992@163.com。
基金资助:
转基因生物新品种培育重大专项（2016ZX08010-003）；国家自然科学基金（81770832；31672502）；中国农业科学院科技创新工程（ASTIP-IAS05）；安徽省科技重大专项（201903a06020002）

GIP/GIPR Enhanced GIP Effect by Positive Feedback of Akt-TCF4-GIPR

YE Huaqiong^1,2, AN Cuiping², ZHANG Kaiyi², XIE Ning², BI Yanzhen³, YANG Shulin^2*, LI Shenghe^1*

1. College of Animal Science, Anhui Science and Technology University, Chuzhou 233100, China;
2. Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China;
3. Institute of Animal Sciences, Hubei Academy of Agricultural Sciences, Wuhan 430064, China

Received:2020-03-02 Online:2020-09-25 Published:2020-09-25

摘要/Abstract

摘要： 旨在从GIP/GIPR下游的Akt和PKA信号通路中筛选出调控GIPR表达的调控因子，并解析GIPR的表达调控机制。本研究以小鼠胰岛瘤细胞系Min6为试验材料，在Akt、PKA信号通路阻断的条件下，通过Western blot筛选出与GIPR表达相关的转录因子T细胞因子4（TCF4）；利用双荧光素酶报告系统确定TCF4对GIPR表达调控的影响，再通过敲除或过表达TCF4进一步验证两者之间的调控关系；采用CCK8法检测TCF4介导的促增殖作用，ELISA检测胰岛素分泌能力。结果显示，GIP可激活Akt磷酸化，并促进GIPR表达；在GIP激活及Akt、PKA信号通路阻断时，GIPR蛋白表达趋势与TCF4始终一致；TCF4可与GIPR核心启动子区结合，进而调控其表达；TCF4过表达时，GIPR的mRNA和蛋白表达上调，并促进β细胞增殖及胰岛素分泌；干扰TCF4显著降低GIP作用下GIPR的mRNA和蛋白表达，抑制β细胞增殖。综上，GIP结合GIPR后，经Akt信号通路上调TCF4进而增强GIPR表达，形成正反馈加强GIP信号，提高β细胞增殖和胰岛素分泌的功能，维持血糖稳态。因此，在胰岛素抵抗阻断Akt及上游信号通路时，经转录因子TCF4增强GIPR表达可作为改善胰岛β细胞功能障碍的靶点。

关键词: 葡萄糖依赖性促胰岛素多肽受体, T细胞因子4, Akt信号通路, Min6细胞

Abstract: The purpose of this study was to screen the regulatory factors that regulate the expression of GIPR from the downstream Akt and PKA signal pathways of GIP/GIPR, and to analyze the regulatory mechanism of GIPR expression. In this study, mouse insulinoma cell line Min6 was used as experimental material, under the condition of blocking Akt and PKA signal pathways, transcription factor T cytokine 4 (TCF4) related to GIPR expression was screened by Western blot, the effect of TCF4 on the regulation of GIPR expression was determined by double luciferase reporter system, and the relationship between TCF4 and GIPR was further verified by knockout or overexpression of TCF4. The proliferation-promoting effect mediated by TCF4 was detected by CCK8 method, and the ability of insulin secretion was detected by ELISA. The results showed that GIP could activate Akt phosphorylation and promote the expression of GIPR; when GIP was activated and Akt and PKA signal pathways were blocked, the expression trend of GIPR protein was consistent with that of TCF4; TCF4 could bind to the core promoter of GIPR and regulate its expression; when TCF4 was overexpressed, the mRNA and protein expression of GIPR were up-regulated, and β-cell proliferation and insulin secretion were promoted. Interfering with TCF4 significantly decreased the mRNA and protein expression of GIPR under the action of GIP, and inhibited the proliferation of β-cell. In conclusion, after GIP is combined with GIPR, TCF4 is upregulated through Akt signal pathway to enhance the expression of GIPR, which forms a positive feedback to enhance GIP signal, and enhance the function of β-cell proliferation and insulin secretion to maintain blood glucose homeostasis. Therefore, when insulin resistance blocks Akt and upstream signal pathway, the enhancement of GIPR expression by transcription factor TCF4 can be used as a target to improve islet β-cell dysfunction.

Key words: glucose dependent insulinotropic polypeptide receptor, T cell factor 4, Akt signal pathway, Min6 cell

中图分类号:

S852.2

叶华琼, 安翠平, 张凯艺, 谢宁, 毕延震, 杨述林, 李升和. GIP/GIPR经Akt-TCF4-GIPR正反馈增强GIP效应[J]. 畜牧兽医学报, 2020, 51(9): 2120-2129.

YE Huaqiong, AN Cuiping, ZHANG Kaiyi, XIE Ning, BI Yanzhen, YANG Shulin, LI Shenghe. GIP/GIPR Enhanced GIP Effect by Positive Feedback of Akt-TCF4-GIPR[J]. Acta Veterinaria et Zootechnica Sinica, 2020, 51(9): 2120-2129.

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GIP/GIPR经Akt-TCF4-GIPR正反馈增强GIP效应

GIP/GIPR Enhanced GIP Effect by Positive Feedback of Akt-TCF4-GIPR

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