转录组测序分析昆仑雪菊醇提物黄诺玛苷对胰岛素抵抗小鼠小肠类器官的影响  

Effect of Flavanomarein from Coreopsis tinctoria Ethanol Extract in Small Intestinal Organoids in Insulin-resistant Mice Based on Transcriptome Sequencing

作　　者：麦麦提亚森·多力昆 依米妮古丽·麦麦提 古丽那孜·别克达吾来提 陈龙 陈航宇 郑梦竹 李琳琳[1] 骆新[1] MAIMAITIYASEN Duolikun;YIMINIGULI Maimaiti;GULINAZI Biekedawulaiti;CHEN Long;CHEN Hangyu;ZHENG Mengzhu;LI Linlin;LUO Xin(College of Pharmacy,Xinjiang Medical University,Urumqi 830011,China;Peking University Third Hospital,Beijing 100191,China;Innovation Center for Genomics,Peking University,Beijing 100091,China)

机构地区：[1]新疆医科大学药学院,乌鲁木齐830011 [2]北京大学第三医院,北京100191 [3]北京大学基因组学创新中心,北京100091

出　　处：《中国实验方剂学杂志》2023年第10期142-151,共10页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：新疆维吾尔自治区高等学校科研计划项目(XJEDU2021I017)。

摘　　要：目的:探讨黄诺玛苷对胰岛素抵抗小鼠小肠类器官转录组的影响。方法:首先建立C57BL/6J和db/db小鼠小肠类器官;采用免疫荧光法检测细胞核核抗原(ki-67)和上皮细胞E-钙黏蛋白(E-cadherin)表达;将小肠类器官分为3组,分别是以C57BL/6J小鼠小肠类器官为正常组、db/db小鼠小肠类器官为模型组和db/db小鼠小肠类器官黄诺玛苷干预组为给药组(FM组);采用蛋白免疫印迹法(Western blot)检测3组小肠类器官胰高血糖素样肽-1(GLP-1)蛋白的表达;并对3组样本进行转录组测序。结果:小肠类器官培养第6天形成围绕管腔周围的环状结构,初步建立了小肠类器官培养模式;免疫荧光检测结果显示小肠类器官均表达ki-67和E-cadherin;Western blot结果显示黄诺玛苷可增加GLP-1蛋白的表达;差异基因表达分析中,模型组与正常组相比差异表达基因共有1862个,FM组与模型组相比差异表达基因共有2282个;通过蛋白质-蛋白质相互作用(PPI)网络分析2组共同表达的差异基因,得到Nr1i3、Cyp2c44、Ugt2b1、Gsta1、Gstm2、Ptgs1、Gstm4、Cyp2c38、Cyp4a32和Gpx310个Hub基因,这些基因在正常组中高表达,模型组中表达降低,黄诺玛苷干预后回调上述基因的表达。结论:黄诺玛苷可能通过逆转Nr1i3、Cyp2c44、Ugt2b1、Gsta1、Gstm2、Ptgs1、Gstm4、Cyp2c38、Cyp4a32和Gpx310个Hub基因的表达水平与相应的信号通路来发挥其改善胰岛素抵抗的作用。Objective:To investigate the effects of flavanomarein on the transcriptome of small intestinal organoids in insulin-resistant mice.Method:Firstly,small intestinal organoids of C57BL/6J and db/db mice were established.Ki-67 and E-cadherin expression was determined by immunofluorescence.Small intestinal organoids were divided into the following three groups:C57BL/6J mouse small intestinal organoids as the normal control group,db/db mouse small intestinal organoids as the model group(IR group),and db/db mouse small intestinal organoids treated with flavanomarein as the administration group(FM group).Western blot was used to detect the expression of glucagon-like peptide-1(GLP-1)protein on the small intestinal organoids of the three groups.Finally,transcriptome sequencing was performed on samples from the three groups.Result:On the 6th day of small intestine organoids culture,a cyclic structure was formed around the lumen,and a small intestine organoids culture model was preliminarily established.Immunofluorescence detection showed that ki-67 and E-cadherin were expressed in small intestinal organoids.Western blot results showed that the expression of GLP-1 protein was increased by flavanomarein.In the results of differential expressed gene(DEG)screening,there were 1862 DEGs in the IR group as compared with the normal control group,and 2282 DEGs in the FM group as compared with the IR group.Through protein-protein interaction(PPI)network analysis of the DEGs of the two groups,10 Hub genes,including Nr1i3,Cyp2c44,Ugt2b1,Gsta1,Gstm2,Ptgs1,Gstm4,Cyp2c38,Cyp4a32,and Gpx3,were obtained.These genes were highly expressed in the normal control group,and their expression was reduced in the IR group.After the intervention of flavanomarein,the expression of the above genes was reversed.Conclusion:Flavanomarein may play its role in improving insulin resistance by reversing the expression levels of 10 Hub genes,including Nr1i3,Cyp2c44,Ugt2b1,Gsta1,Gstm2,Ptgs1,Gstm4,Cyp2c38,Cyp4a32,and Gpx3.

关 键 词：胰岛素抵抗 黄诺玛苷 小肠类器官 转录组测序 差异表达基因 Hub基因 

分 类 号：R285[医药卫生—中药学] R965[医药卫生—中医学] R34