甘草查尔酮A缓解2型糖尿病所致异常糖异生及内质网应激分子机制研究  

Molecular mechanism underlying the effects of licochalcone A on abnormal gluconeogenesis and endoplasmic reticulum stress induced by type 2 diabetes mellitus

作　　者：许文镨 张佳瑜 汪逗逗 丁文文 陈姿伊 肖瑶[2] 刘颖 XU Wen-pu;ZHANG Jia-yu;WANG Dou-dou;DING Wen-wen;CHEN Zi-yi;XIAO Yao;LIU Ying(School of Life Sciences,Beijing University of Chinese Medicine,Beijing 102488,China;School of Chinese Pharmacy,Beijing University of Chinese Medicine,Beijing 102488,China)

机构地区：[1]北京中医药大学生命科学学院,北京102488 [2]北京中医药大学中药学院,北京102488

出　　处：《药学学报》2024年第12期3291-3303,共13页Acta Pharmaceutica Sinica

摘　　要：本文旨在探讨甘草查尔酮A(licochalcone A,LCA)缓解2型糖尿病(type 2 diabetes mellitus,T2DM)引起的异常糖异生及内质网应激的分子机制。体内研究采用8周龄雄性C57BL/6J小鼠,高脂高糖饲料喂养结合腹腔注射链脲佐菌素(streptozotocin,STZ)构建T2DM动物模型。腹腔注射LCA(5、10 mg·kg^(-1))治疗,以二甲双胍(metformin,MET)(200 mg·kg^(-1))为阳性对照,间隔3天给药,持续3周,动物实验方案经北京中医药大学实验动物伦理委员会审核并批准(批准号:BUCM-4-2021061701-2060)。体外实验以人肝癌细胞HepG2为实验细胞系,采用棕榈酸钠(sodium palmitate,SP)诱导胰岛素抵抗细胞模型,采用衣霉素(tunicamycin,TM)诱导内质网应激细胞模型。采用实时荧光定量PCR法(real-time quantitative polymerase chain reaction,RT-qPCR)、酶联免疫吸附法(enzymelinked immunosorbent assay,ELISA)和蛋白质印迹法(Western blot,WB)检测糖异生和内质网应激相关靶点的转录和蛋白水平;利用分子对接和分子动力学模拟对LCA和关键靶点的相互作用进行验证。结果表明,LCA可通过抑制磷酸烯醇式丙酮酸羧激酶(phosphoenolpyruvatecarboxykinase,PEPCK)和葡萄糖-6-磷酸酶(glucose-6-phosphatase,G6P)的转录和酶活,抑制丙酮酸羧化酶(pyruvate carboxylase,PC)的酶活,提高6-磷酸果糖激酶-2/果糖-2,6-二磷酸酶3(6-phosphofructokinase-2/fructose-2,6-bisphosphatase 3,PFKFB3)的转录和蛋白水平,抑制糖异生。同时,LCA可通过下调真核翻译起始因子2α(eukaryotic initiation factor 2 subunitα,eIF2α)、肌醇依赖酶1α(inositolrequiringenzyme1α,IRE1α)、X-框结合蛋白1(X-boxbindingprotein1,XBP1)、c-Jun氨基末端激酶1(c-JunNterminal kinase 1,JNK1)和活化转录因子6α(activating transcription factor 6α,ATF6α)的转录水平,降低葡萄糖调节蛋白78(glucose-regulated protein 78,GRP78)的转录和蛋白水平,抑制蛋白质激酶RNA样端激活因子(protein kinase RNA-like endoplasmic reticulum kinase,PERK)的转�The aim of this study is to investigate the molecular mechanism of licochalcone A(LCA) in alleviating abnormal gluconeogenesis and endoplasmic reticulum(ER) stress caused by type 2 diabetes mellitus(T2DM).In the in vivo study,8-week-old male C57BL/6J mice were fed with a high-fat and high-sugar diet and injected intraperitoneally with streptozotocin(STZ) to establish a T2DM model.LCA(5 and 10 mg·kg^(-1)) was administered at an interval of 3 days for 3 weeks with metformin(MET,200 mg·kg^(-1)) as a positive control drug.The animal experiment protocol was reviewed and approved by the Experimental Animal Ethics Committee of Beijing University of Chinese Medicine(approval number:BUCM-4-2021061701-2060).Human hepatoma cell line HepG2 was used as the experimental cell line for in vitro experiments.Sodium palmitate(SP) was used to induce the insulin resistance cell model and tunicamycin(TM) was applied to establish the ER stress cell model.Real-time quantitative polymerase chain reaction(RT-qPCR),enzyme-linked immunosorbent assay(ELISA) and Western blot(WB) were used to detect the mRNA and protein levels of gluconeogenesis and ER stress-related targets,respectively.Molecular docking and dynamics simulations were used to verify the interaction between LCA and key targets.The results showed that LCA inhibits gluconeogenesis by reducing phosphoenolpyruvate carboxykinase(PEPCK) and glucose-6-phosphatase(G6P) and increasing 6-phosphofructokinase-2/fructose-2,6-bisphosphatase 3(PFKFB3) at both the mRNA and protein levels,as well as suppressing the activity of pyruvate carboxylase(PC).Additionally,LCA alleviates ER stress by downregulating the transcription of eukaryotic initiation factor 2 subunit α(eIF2α),inositol-requiring enzyme 1α(IRE1α),X-box binding protein 1(XBP1),c-Jun N-terminal kinase 1(JNK1),and activating transcription factor 6α(ATF6α),inhibiting the transcription and protein expression of glucose-regulated protein 78(GRP78),and suppressing the phosphorylation of protein kinase RNA-like endoplasmic reticulu

关 键 词：甘草查尔酮A 2型糖尿病 糖异生 内质网应激 代谢紊乱 

分 类 号：R966[医药卫生—药理学]