四君子汤干预二型糖尿病大鼠脂肪胰岛素抵抗的作用及其PI3K/AKT通路机制研究

Effects of Sijunzi Decoction on Adipose Insulin Resistance in T2DM Rats and Its Mechanism via the PI3K/AKT Pathway

关键词:  四君子汤  T2DM  PI3K/AKT通路  胰岛素抵抗  脂肪

Key words:  sijunzi decoction  Type 2 Diabetes Mellitus (T2DM)  PI3K/AKT pathway  Insulin Resistance (IR)  adipose tissue

摘要:

目的 探究四君子汤对2型糖尿病（T2DM）大鼠脂肪胰岛素抵抗（IR）的改善作用及其机制。方法造模采取SD大鼠高糖高脂饮食结合链脲佐菌素（STZ）干预，动物随机分为正常对照组、模型对照组、二甲双胍组、四君子汤组。灌胃给药，每天一次，持续4周。测定各组大鼠反应灵敏度、自主活动频率、摄食量、饮水量、体重的变化；测定口服葡萄糖耐量（OGTT）、胰岛素耐量（ITT）、空腹血糖（FBG）、空腹胰岛素（FINS），计算胰岛素抵抗指数（HOMA-IR）、胰岛 β 细胞功能指数（HOMA-β）；测定肝脏指数、血清总胆固醇（TC）、总甘油三酯（TG）、高密度脂蛋白胆固醇（HDL-C）、低密度脂蛋白胆固醇（LDL-C）；HE染色观察脂肪组织病理变化；实时荧光定量PCR（RT-qPCR）、Western blot（WB）技术检测脂肪中INSR、IRS-1、PI3K、AKT、GLUT4、GSK-3β、FOX-O1总mRNA和总蛋白表达量。结果 四君子汤能够显著改善T2DM大鼠反应灵敏度及自主活动频率（P<0.01），改善T2DM大鼠多饮多食症状，显著升高T2DM大鼠体重（P<0.01）；显著降低OGTT、ITT、FBG、HOMA-IR水平（P<0.01），明显降低FINS水平（P<0.05），显著升高HOMA-β 水平（P<0.01）；显著降低血清中TC含量（P<0.01），明显降低TG和LDL-C含量（P<0.05），明显升高 HDL-C 含量（P<0.05）；脂肪病理学观察显示四君子汤可以改善T2DM大鼠脂肪细胞的形态大小及变性，减少脂滴空泡等病理状况；RT-qPCR、WB结果显示四君子汤可以上调INSR、IRS-1、PI3K、AKT、GLUT4总mRNA表达量（P<0.01或P<0.05），下调GSK-3β、FOX-O1总mRNA表达量（P<0.01或P<0.05）；上调INSR、IRS-1、GLUT4并下调GSK-3β、FOX-O1总蛋白表达量（P<0.01或P<0.05），上调PI3K、AKT总蛋白表达量但无显著性差异（P>0.05）。结论 四君子汤可以通过调节 PI3K/AKT 信号通路中七个信号因子表达来改善T2DM大鼠脂肪部位胰岛素抵抗。

Abstract:

Objective To investigate the ameliorative effect of Sijunzi Decoction (SJZD) on adipose insulin resistance (IR) in type 2 diabetes mellitus (T2DM) rats and its underlying mechanism.Methods A T2DM model was established in SD rats using a high-sugar, high-fat diet combined with streptozotocin (STZ) intervention. Animals were randomly divided into: normal control group, model control group, metformin group, and SJZD group. Intragastric administration was performed once daily for 4 weeks. Changes in reaction sensitivity, spontaneous activity frequency, food intake, water intake, and body weight were measured in each group. oral glucose tolerance test (OGTT),Insulin Tolerance Test (ITT) fasting blood glucose (FBG), and fasting insulin (FINS) were measured in each group.Followed by the calculation of insulin resistance (HOMA-IR) and β-cell function (HOMA-β) using the homeostasis model assessment method. Serum levels of total cholesterol (TC), total triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) were measured. Pathological changes in adipose tissue were observed using hematoxylin-eosin (HE) staining. Total mRNA and total protein expression levels of INSR, IRS-1, PI3K, AKT, GLUT4, GSK-3β, and FOX-O1 in the liver were detected using real-time quantitative polymerase chain reaction (RT-qPCR) and Western blot (WB) techniques.Results SJZD significantly improved the response sensitivity and spontaneous activity frequency in T2DM rats (P<0.01), ameliorated symptoms of polydipsia and polyphagia, and significantly increased body weight (P<0.01). It significantly reduced OGTT, FBG, and HOMA-IR levels (P<0.01), significantly decreased FINS levels (P<0.05), and significantly increased HOMA-β levels (P<0.01). SJZD significantly reduced serum TC content (P<0.01), significantly decreased TG and LDL-C content (P<0.05), and significantly increased HDL-C content (P<0.05). Adipose histopathological observation showed that SJZD improved the morphology, size, and degeneration of adipocytes in T2DM rats and reduced pathological conditions such as lipid droplet vacuoles. RT-qPCR and WB results demonstrated that SJZD up-regulated the total mRNA expression of INSR, IRS-1, PI3K, AKT, and GLUT4 (P<0.01 or P<0.05), and down-regulated the total mRNA expression of GSK-3β and FOX-O1 (P<0.01 or P<0.05). SJZD up-regulated the total protein expression of INSR, IRS-1, and GLUT4 while down-regulating GSK-3β and FOX-O1 (P<0.01 or P<0.05). It also up-regulated PI3K and AKT total protein expression, although these increases were not statistically significant (P>0.05).Conclusion Sijunzi Decoction can ameliorate insulin resistance in the adipose tissue of T2DM rats by regulating the expression of seven signaling factors of the PI3K/AKT signaling pathway.