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《天津医科大学学报》[ISSN:1006-8147/CN:12-1259/R]

卷:

26卷

期数:

2020年04期

页码:

317-323

栏目:

基础医学

出版日期:

2020-07-15

文章信息/Info

Title:

Study on the anti-diabetic effect and mechanism of the derivative Fla-CN

文章编号:

1006-8147(2020)04-0317-07

作者:

寇川; 梁艳; 张畅; 秦楠; 段宏泉; 陈莹

（天津医科大学药学院天然药物化学系，天津 300070）

Author(s):

KOU Chuan;  LIANG Yan;  ZHANG Chang;  QIN Nan;  DUAN Hong-quan;  CHEN Ying

（Department of Nature Pharmaceutical Chemistry, School of Pharmacy, Tianjin Medical University, Tianjin 300070, China）

关键词:

Fla-CN; 糖尿病; 糖摄取; 糖异生; AMPK通路

Keywords:

Fla-CN; diabetes mellitus;  glucose uptake; gluconeogenesis; AMPK signal pathway

分类号:

R589

DOI:

-

文献标志码:

A

摘要:

目的：体内外研究黄酮衍生物Fla-CN的抗糖尿病作用及其机制。方法：体外实验以人肝HepG2细胞为研究对象，不同浓度Fla-CN孵育后，利用荧光标记 2-脱氧葡萄糖（2-NBDG），通过检测HepG2 细胞内2-NBDG的荧光强度，观察Fla-CN对肝细胞葡萄糖摄取的影响；采用硫酸-蒽酮比色法检测HepG2细胞内糖原含量；通过检测一定时间内葡萄糖生成量，观察Fla-CN对肝细胞糖异生过程的影响。体内实验采用2型糖尿病模型db/db小鼠为研究对象，给予Fla-CN和二甲双胍干预4 周后，测定db/db小鼠空腹血糖、糖耐量以及胰岛素耐量，计算糖耐量、胰岛素耐量实验血糖-曲线下面积（AUC）。通过Western印迹法检测化合物干预后小鼠肝组织中磷酸化AMP活化蛋白激酶 (AMPK)、糖异生关键酶磷酸烯醇式丙酮酸羧激酶（PEPCK）、葡萄糖-6-磷酸酶（G6Pase）及转录调控因子过氧化物酶体增殖物激活受体γ协同刺激因子1α（PGC-1α）的表达。进一步通过Western 印迹检测AMPK抑制剂Compound C或AMPK激动剂AICAR 与Fla-CN联用后，HepG2细胞内PEPCK、G6Pase、PGC-1α的蛋白表达。结果：Fla-CN能够促进HepG2细胞对葡萄糖的摄取（F=33.50，P<0.01）和糖原合成（F=93.63，P<0.01），抑制糖异生（F=44.19,P<0.01）。同时，Fla-CN显著降低2型糖尿病db/db小鼠空腹血糖以及时间-血糖曲线下面积（F=98.40，P<0.05）。Western印迹显示Fla-CN能够激活db/db小鼠肝组织中 AMPK，抑制PEPCK、G6Pase 以及PGC-1α 的表达，并呈现一定剂量依赖性。Compound C能在一定程度上阻断Fla-CN对AMPK的激活和对PEPCK、G6Pase 以及PGC-1α的抑制作用。结论：黄酮衍生物Fla-CN通过激活AMPK，有效调节肝 HepG2 细胞糖代谢，抑制肝糖异生，降低2型糖尿病db/db小鼠空腹血糖，改善其受损的糖耐量，提高胰岛素敏感性。

Abstract:

Objective: To study the anti-diabetic effect and mechanism of the flavonoid derivative Fla-CN in vitro and in vivo. Methods: In vitro experiment, HepG2 cells were incubated with different concentrations of Fla-CN, and the fluorescence intensity of 2-NBDG in HepG2 cells was detected to observe the effect of Fla-CN on glucose uptake in hepatocytes. The glycogen content in HepG2 cells was measured by sulfuric acid anthronecolorimetry. The glucose production was measured to investigate the effect of Fla-CN on gluconeogenesis of hepatocytes. In vivo experiments, db/db mice model of type 2 diabetes were used as the subjects of study. After intervention with Fla-CN and metformin for 4 weeks, fasting blood glucose, glucose tolerance and insulin tolerance of db/db mice were investigated, and AUC of glucose tolerance and insulin tolerance were calculated. Western blotting was used to detect the expression of AMP dependent protein kinase (AMPK) phosphorylation, phosphoenolpyruvate carboxykinase(PEPCK), glucose-6-phosphatase(G6Pase) and peroxisome proliferator activated receptor γ coactivator-1 α(PGC-1α) in the liver tissue of mice. The protein expression of PEPCK, G6Pase and PGC-1α in HepG2 cells was further detected by Western blotting after an incubation of AMPK inhibitor Compound C or AMPK agonist AICAR combined with Fla-CN. Results: Fla-CN could promote glucose uptake (F=33.50, P<0.01), glycogen synthesis (F=93.63, P<0.01) and inhibit gluconeogenesis(F=44.19,P<0.01) in HepG2 cells. Meanwhile, Fla-CN significantly reduced fasting blood glucose and its AUC in db/db mice (F=98.40, P<0.05). Western blotting showed that Fla-CN could activate AMPK, inhibit the expression of PEPCK, G6Pase and PGC-1α in the liver of db/db mice in a dose-dependent manner. Compound C could impair the activation of AMPK and the inhibition of PEPCK, G6Pase and PGC-1α by Fla-CN. Conclusion: Fla-CN could effectively regulate the glucose metabolism of HepG2 cells, reduce the fasting blood glucose of db/db mice, and improve the impaired glucose tolerance and insulin sensitivity. Further studies have shown that Fla-CN promotes glucose uptake and inhibits glyconeogenesis by activating AMPK in liver.

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备注/Memo

备注/Memo:

基金项目 国家自然科学基金面上项目（81373297）
作者简介 寇川（1993-），男，硕士在读，研究方向：中药和天然药物活性物质的现代研究；
通信作者：陈莹，E-mail：chenyingweixiao37@163.com。

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更新日期/Last Update: 2020-07-15