文献类型：期刊文献

中文题名：基于转录组学探讨糖止丸调控PI3K/Akt改善T2DM大鼠肝脏胰岛素抵抗的效应机制

英文题名：Mechanism Exploration of TG Regulating PI3K/Akt to Improve Insulin Resistance in Liver of T2DM Rats Based on Transcriptomics

作者：李钦[1,2];梁永林[1];史晓伟[3];刘轩[2];朱向东[4]

第一作者：李钦

机构：[1]甘肃中医药大学,兰州730000;[2]甘肃卫生职业学院,兰州730030;[3]甘肃省中医院,兰州730050;[4]宁夏医科大学,银川750001

第一机构：甘肃中医药大学

年份：2024

卷号：30

期号：2

起止页码：99

中文期刊名：中国实验方剂学杂志

外文期刊名：Chinese Journal of Experimental Traditional Medical Formulae

收录：CSTPCD;;Scopus;北大核心:【北大核心2023】；CSCD:【CSCD2023_2024】；

基金：甘肃省自然科学基金项目(22JR5RA810);甘肃省教育科技创新项目(2022A-249);宁夏自然科学基金重点项目(2023AAC02035);甘肃省中医药管理局中医药防治重大疾病科研课题(GZKZD-2018-01)。

语种：中文

中文关键词：糖止丸;磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt);2型糖尿病(T2DM);差异基因;胰岛素抵抗;转录组学

外文关键词：Tangzhi pills;phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt);type 2 diabetes(T2DM);differential gene;insulin resistance;transcriptomics

摘要：目的:探讨糖止丸基于差异基因调控磷脂酰肌醇3-激酶(PI3K)/蛋白激酶B(Akt)信号通路改善2型糖尿病(T2DM)肝脏胰岛素抵抗(IR)的效应及可能的分子机制。方法:采用高脂(HFD)饲料喂养联合链脲佐菌素(STZ)腹腔注射方法制备T2DM大鼠模型。实验分为空白组、模型组、盐酸二甲双胍组(0.18 g·kg^(-1))、糖止丸高(1.08 g·kg^(-1))、中(0.54 g·kg^(-1))、低(0.27 g·kg^(-1))剂量组。收集大鼠血清、肝脏和胰腺组织,苏木素-伊红(HE)染色肝脏、胰腺病理组织观察;检测空腹血糖值(FBG),进行口服葡萄糖耐量(OGTT)实验;酶联免疫吸附测定法(ELISA)检测大鼠空腹血清胰岛素(FINS)、糖化血红蛋白(GHb)水平;计算胰岛素抵抗稳态模型指数(HOMA-IR)、β细胞功能稳态指数(HOMA-β)、胰岛素敏感指数(ISI);生化法测定大鼠血清甘油三酯(TG)、总胆固醇(TC)、低密度脂蛋白胆固醇(LDL-C)、高密度脂蛋白胆固醇(HDL-C)含量。转录组学获得肝脏组织差异表达mRNA并富集差异表达通路。实时荧光定量聚合酶链式反应(Real-time PCR)检测肝脏组织环腺苷酸应答元件结合蛋白3样蛋白2抗体(CREB3l2)、B细胞淋巴瘤-2(Bcl-2)、Toll样受体2(TLR2)、周期蛋白依赖性激酶抑制剂1A(CDNK1A)、DNA损伤诱导转录因子4样蛋白(DDIT4)mRNA表达;蛋白免疫印迹法(Western blot)检测磷酸化(p)-PI3K、p-Akt、葡萄糖转运蛋白4(GLUT4)、胰岛素受体(INSR)、胰岛素受体底物2(IRS2)蛋白表达。结果:药效学实验结果显示,与模型组比较,糖止丸各剂量组不同程度修复肝脏和胰腺组织;降低血糖(P<0.01);促使血清FINS、GHb水平和HOMA-IR降低(P<0.05,P<0.01),HOMA-β、ISI升高(P<0.05,P<0.01);TC、TG、LDL-C水平降低(P<0.05,P<0.01),HDL-C水平升高(P<0.05,P<0.01)。转录组学实验结果确证PI3K/Akt信号通路在空白组与模型组及糖止丸高剂量组与模型组中均显著表达;CDNK1A、DDIT4、CREB3l2、Bcl-2、TLR2是糖止丸干预T2DM的显著差异表达mRNA。与模型比较,糖止丸各剂量组p-PI3K、p-Akt、GLUT4、INSR、IRS2蛋白表达显著升高(P<0.01);CREB3l2、Bcl-2、TLR2mRNA表达显著升高(P<0.01),CDNK1A、DDIT4 mRNA表达显著降低(P<0.01)。结论:糖止丸可能是基于CREB3l2、Bcl-2、TLR2、CDNK1A、DDIT4差异mRNA表达调控PI3K/Akt信号通路,从而发挥改善T2DM肝脏胰岛素抵抗。
Objective:To investigate the effect of Tangzhi pills on the improvement of insulin resistance(IR)in the liver with type 2 diabetes(T2DM)by regulating phosphatidylinositol 3-kinase(PI3K)/protein kinase B(Akt)signaling pathway based on differential genes and its possible molecular mechanism.Method:T2DM rat models were prepared by high fat(HFD)diet combined with streptozotocin(STZ)intraperitoneal injection.The experiment was divided into blank group,model group,metformin hydrochloride group(0.18 g·kg^(-1)),Tangzhi pills high(1.08 g·kg^(-1)),medium(0.54 g·kg^(-1))and low(0.27 g·kg^(-1))dose groups.Rat serum,liver,and pancreatic tissue were collected,and the pathological tissue of the liver and pancreas was observed using hematoxylin-eosin(HE)staining.The fasting blood glucose level(FBG)was detected,and oral glucose tolerance(OGTT)tests were conducted.Enzyme-linked immunosorbent assay(ELISA)was used to detect fasting serum insulin(FINS)and glycated hemoglobin(GHb)levels in rats.IR homeostasis model index(HOMA-IR),βcellular homeostasis index(HOMA-β),and insulin sensitivity index(ISI)were calculated.Biochemical methods were used to determine the levels of triglyceride(TG),total cholesterol(TC),low-density lipoprotein(LDL-C),and high-density lipoprotein(HDL-C)in rat serum.Transcriptomics obtained differentially expressed mRNA from liver tissue and enriched differentially expressed pathways.Real-time reverse transcriptase polymerase chain reaction(Real-time PCR)was used to detect the mRNA expression of cyclic adenylate responsive element binding protein 3-like protein 2 antibody(CREB3l2),B-lymphocyte tumor 2(Bcl-2),Toll-like receptor 2(TLR2),cyclin-dependent kinase inhibitor 1A(CDNK1A),and DNA damage induced transcription factor 4-like protein(DDIT4)in liver tissue.Western blot was used to detect the protein expression of phosphorylated phosphatidylinositol 3-kinase(p-PI3K),phosphorylated protein kinase B(p-Akt),glucose transporter 4(GLUT4),insulin receptor(INSR),and insulin receptor substrate 2(IRS2).Result:The pharmacodynamic experiment results showed that compared with model group,Tangzhi pills groups repaired liver and pancreatic tissue to varying degrees,reduced blood sugar(P<0.01),and promoted a decrease in serum FINS,GHb,and HOMA-IR(P<0.05,P<0.01).In addition,HOMA-βand ISI increased(P<0.05,P<0.01).The levels of TC,TG,and LDL-C decreased(P<0.05,P<0.01),while the levels of HDL-C increased(P<0.05,P<0.01).The transcriptomics experimental results confirmed that the PI3K/Akt signaling pathway was significantly expressed in both the blank group and model group,as well as in the high-dose Tangzhi pills group and model group.CDNK1A,DDIT4,CREB3l2,Bcl-2,and TLR2 were significantly differentially expressed mRNA during TG intervention in T2DM.Compared with the model group,the protein expression of p-PI3K,p-Akt,GLUT4,INSR,and IRS2 increased in all Tangzhi pills groups(P<0.01).The mRNA expression of CREB3l2,Bcl-2,and TLR2 increased(P<0.01),while that of CDNK1A and DDIT4 decreased(P<0.01).Conclusion:Tangzhi pills may regulate the PI3K/Akt signaling pathway based on the differential mRNA expression of CREB3l2,Bcl-2,TLR2,CDNK1A,and DDIT4,thereby improving IR in the liver with T2DM.