Effect of Tangbikang Granules on PI3K Pathway in Type 2 Diabetic Mellitus Rats with Non-alcoholic Fatty Liver Disease

ZHANG Yaqi; QIN Lingling; ZHANG Chengfei; ZHANG Qiue; BAI Huizhong; LIU Gang; ZUO Xinwei; ZHAO Yi; LIU Tonghua; MU Xiaohong

doi:10.13422/j.cnki.syfjx.20230628

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Effect of Tangbikang Granules on PI3K Pathway in Type 2 Diabetic Mellitus Rats with Non-alcoholic Fatty Liver Disease

更新时间：2023-03-30

- Effect of Tangbikang Granules on PI3K Pathway in Type 2 Diabetic Mellitus Rats with Non-alcoholic Fatty Liver Disease
- Chinese Journal of Experimental Traditional Medical Formulae Vol. 29, Issue 9, Pages: 71-80(2023)
- 作者机构：
  
  1.北京中医药大学东直门医院，北京 100700
  2.北京中医药大学科技处，北京 100029
  3.教育部高等学校学科创新引智基地，北京 100029
  4.北京中医药大学生命科学学院，北京 100029
  5.北京中医药大学中医学院，北京 100029
  6.北京中医药大学教育部中医养生学重点实验室，北京 100029
  7.科技部中医药防治糖尿病国际联合研究中心，北京 100029
- 作者简介：
- 基金信息：
- DOI：10.13422/j.cnki.syfjx.20230628
  CLC： R2-0;R22;R242;R2-031;R285.5
- Received：19 September 2022，
  
  Published Online：03 February 2023，
  
  Published：05 May 2023
- 稿件说明：
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张亚奇,秦灵灵,张程斐等.糖痹康颗粒对2型糖尿病合并非酒精性脂肪肝大鼠PI3K通路的影响[J].中国实验方剂学杂志,2023,29(09):71-80.

ZHANG Yaqi,QIN Lingling,ZHANG Chengfei,et al.Effect of Tangbikang Granules on PI3K Pathway in Type 2 Diabetic Mellitus Rats with Non-alcoholic Fatty Liver Disease[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(09):71-80.
张亚奇,秦灵灵,张程斐等.糖痹康颗粒对2型糖尿病合并非酒精性脂肪肝大鼠PI3K通路的影响[J].中国实验方剂学杂志,2023,29(09):71-80. DOI： 10.13422/j.cnki.syfjx.20230628.

ZHANG Yaqi,QIN Lingling,ZHANG Chengfei,et al.Effect of Tangbikang Granules on PI3K Pathway in Type 2 Diabetic Mellitus Rats with Non-alcoholic Fatty Liver Disease[J].Chinese Journal of Experimental Traditional Medical Formulae,2023,29(09):71-80. DOI： 10.13422/j.cnki.syfjx.20230628.

摘要

目的

探讨糖痹康颗粒对2型糖尿病（T2DM）合并非酒精性脂肪肝（NAFLD）的治疗及阐明作用机制。

方法

采用自发性的雄性Zucker糖尿病脂肪（ZDF）大鼠，建立T2DM合并NAFLD模型。造模成功的大鼠进行糖痹康颗粒高、中、低剂量组（2.5、1.25、0.625 g·kg

-1

）持续灌胃治疗12周。治疗期间每4周记录空腹血糖（FBG）和体质量变化。取材前1周，检测给药治疗对大鼠进食量影响；并进行夜间禁食12 h后进行口服葡萄糖耐量实验（OGTT），曲线下面积（AUC）评价糖耐量，计算胰岛素抵抗指数（HOMA-IR）。腹主动脉取血和取肝脏，测定血糖和血脂代谢指标，包括空腹血清胰岛素（FINS）、血清总胆固醇（TC）、甘油三酯（TG）、低密度脂蛋白胆固醇（LDL-C）、高密度脂蛋白胆固醇（HDL-C）及游离脂肪酸（NEFA）。对肝脏进行称质量计算肝脏指数，并对其进行肝脏组织形态学苏木素-伊红（HE）染色、过碘酸-Schiff（PAS）染色观察分析。蛋白免疫印迹法（Western blot）检测胰岛素受体底物（IRS）、磷脂酰肌醇3-激酶（PI3K）、蛋白激酶B（Akt）和磷酸化IRS、Akt蛋白的表达水平。所有数据采用SPSS 20.0软件进行分析

。

结果

与正常组比较，模型组大鼠摄食量显著增多（

0.01）；与模型组比较，其余各给药组糖尿病大鼠摄食量减少（

0.05，

0.01）。与正常组比较，模型组大鼠8、12周体质量显著降低（

0.01）。与模型组比较，糖痹康颗粒组在一定程度上降低FBG水平，且呈浓度依赖性。与正常组比较，模型组OGTT血糖值和AUC显著增高（

0.01）；与模型组比较，糖痹康颗粒各组和二甲双胍组OGTT血糖值整体降低，AUC差异有统计学意义（

0.01）。与正常组比较，模型组大鼠血清FINS、HOMA-IR指数均显著升高（

0.01）；与模型组比较，糖痹康颗粒各组大鼠血清FINS水平、HOMA-IR指数均显著降低（

0.01）。与正常组比较，模型组大鼠血清TG、TC、HDL-C及NEFA水平均明显升高（

0.05，

0.01），LDL-C水平呈升高趋势，但差异无统计学意义。与模型组比较，糖痹康颗粒各组大鼠血清TG、TC、LDL-C及NEFA水平降低，且糖痹康颗粒各组TG、LDL-C和NEFA水平呈浓度依赖趋势，即糖痹康颗粒高剂量组最优。与模型组比较，糖痹康颗粒高剂量组大鼠HDL-C水平明显升高（

0.05）。与正常组比较，模型组大鼠肝脏指数显著升高（

0.01）；与模型组比较，给药各组大鼠肝脏指数呈降低趋势，但差异无统计学意义。肝组织形态HE染色观察，模型组大鼠肝小叶结构不清；大部分细胞肿大且大小不一，肝细胞脂肪化明显。糖痹康颗粒各组均可不同程度地减轻肝脏损伤，但以糖痹康颗粒高剂量组最为显著。肝组织形态PAS染色观察，与正常组比较，模型组肝脏细胞可见显著脂肪空泡，胞浆中紫红色糖原颗粒显著减少，颜色较浅。糖痹康颗粒高、中剂量组染色更接近正常组。Western blot检测肝组织蛋白表达，与正常组比较，模型组大鼠PI3K蛋白、p-IRS1/IRS1、p-Akt/Akt表达显著降低（

0.01）；与模型组比较，糖痹康颗粒高剂量组的PI3K蛋白、p-IRS1/IRS1、p-Akt/Akt表达显著提高（

0.01）。

结论

糖痹康颗粒治疗ZDF大鼠T2DM合并NAFLD，其机制可能与激活典型的PI3K信号通路相关。

Abstract

Objective

This study aims to investigate the therapeutic effect of Tangbikang granules（TBK） on type 2 diabetes mellitus （T2DM） complicated with non-alcoholic fatty liver disease （NAFLD） and to elucidate the underlying mechanism.

Method

T2DM and NAFLD were induced in ZDF rats， which were then respectively treated （

） with low-dose （0.625 g·kg

-1

）， medium-dose （1.25 g·kg

-1

）， and high-dose （2.5 g·kg

-1

） TBK for 12 weeks. Fasting blood glucose （FBG） and body mass were recorded every 4 weeks during the treatment. One week before sampling， the feed intake of rats was detected， and after 12 h night fasting， oral glucose tolerance test （OGTT） was performed. The area under the curve （AUC） was used to evaluate glucose tolerance， and the homeostatic model assessment for insulin resistance （HOMA-IR） was calculated. Blood in abdominal aorta and liver were collected for determination of blood glucose and lipid metabolism indexes： Fasting serum insulin （FINS）， serum total cholesterol （TC）， triglyceride （TG）， low density lipoprotein cholesterol （LDL-C）， high density lipoprotein cholesterol （HDL-C）， and nonesterified fatty acids （NEFA）. The liver was weighed to calculate the liver index， and the liver tissue morphology was observed and analyzed based on hematoxylin-eosin （HE） staining and periodic acid-Schiff （PAS） staining. The protein levels of insulin receptor substrate （IRS）， phosphatidylinositol 3-kinase （PI3K）， protein kinase B （Akt） and phosphorylated IRS and Akt were detected by Western blotting. All data were analyzed by SPSS 20.0.

Result

The feed intake of the model group was higher than that in the normal group （

0.01）， and the feed intake the administration groups was lower than that in the model group （

0.05，

0.01）. At the 8

and 12

week， the body mass in the model group was lower than that in the normal group （

0.01）. Compared with the model group， TBK reduced FBG in a concentration-dependent manner. The blood glucose level in OGTT and AUC in the model group were higher/larger than those in the normal group （

0.01）. The blood glucose value in OGTT was decreased in TBK groups and the metformin group compared with that in the model group， and AUC in the administration groups was significantly different from that in the model group （

0.01）. The serum level of FINS and HOMA-IR in the model group were higher than those in the normal group （

0.01）， and they were lower in the TBK groups than in the model group （

0.01）. Serum levels of TG， TC， HDL-C， NEFA （

0.05，

0.01）， and LDL-C were higher in the model group than in the normal group. Serum levels of TG， TC， LDL-C， and NEFA in the TBK groups were lower than those in the model group， and the levels of TG， LDL-C， and NEFA in TBK groups were concentration-dependent （lowest levels in high-dose TBK group）. Compared with the model group， high-dose TBK significantly increased the level of HDL-C （

0.05）. Liver index of the model group was higher than that in the normal group （

0.01）. The liver index of the administration groups showed a decreasing trend with no significant difference from that in the model group. As for the HE staining result of liver， the model group had unclear structure of liver lobule， enlarged cells of different sizes， and obvious steatosis of hepatocytes. TBK of all doses alleviated liver injury， particularly the high dose. For the PAS staining， compared with the normal group， the model group demonstrated significant fat vacuoles and significant reduction in purplish red glycogen granules in the cytoplasm. The staining results of high- and medium-dose groups of TBK were more similar to the normal group. Western blot was used to detect the protein expression of liver tissue. The expression of PI3K protein， p-IRS1/IRS1， and p-Akt/Akt in the model group were lower than those in the normal group （

0.01）， and they were higher in the high-dose TBK group than in the model group （

0.01）.

Conclusion

TBK exerts therapeutic effect on T2DM combined with NAFLD in ZDF rats by activating the typical PI3K signaling pathway.

关键词

Keywords

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