黄芩苷调控TUG1/PTBP1/NLRP3分子网络抑制巨噬细胞焦亡治疗溃疡性结肠炎的机制  

Mechanism of baicalin modulation of TUG1/PTBP1/NLRP3 molecular network to inhibit macrophage pyroptosis in treatment of ulcerative colitis

作　　者：李克亚[1] 肖金银 罗雯鹏 陆文洪[1] 贺荔枝[1] 潘燎 赵建政 成思宇 肖俐敏 王军文[1] 王真权[1] LI Keya;XIAO Jinyin;LUO Wenpeng;LU Wenhong;HE Lizhi;PAN Liao;ZHAO Jianzheng;CHENG Siyu;XIAO Limin;WANG Junwen;WANG Zhenquan(The Second Affiliated Hospital of Hunan University of Chinese Medicine,Changsha 410005,China)

机构地区：[1]湖南中医药大学第二附属医院,湖南长沙410005

出　　处：《中草药》2025年第5期1667-1681,共15页Chinese Traditional and Herbal Drugs

基　　金：2022年度湖南省自然科学基金青年基金项目(2022JJ40325);湖南省第一批中医药学科带头人(神农学者)(2100601);中医药人才培养重点学科(中医肛肠病学)(2023-01-16-006-011)。

摘　　要：目的探讨黄芩苷调控牛磺酸上调基因1(taurine up-regulated gene 1,TUG1)/多嘧啶束结合蛋白1(polypyrimidine tract binding protein 1,PTBP1)/核苷酸结合寡聚化结构域样受体蛋白3(nucleotide-binding oligomerization domain-like receptor protein 3,NLRP3)分子网络抑制巨噬细胞焦亡治疗溃疡性结肠炎(ulcerative colitis,UC)的作用机制。方法利用葡聚糖硫酸钠(dextran sulfate sodium salt,DSS)诱导构建UC小鼠模型,将小鼠随机分为对照组、模型组、美沙拉嗪(600 mg/kg)组和黄芩苷低、中、高剂量(25、50、100 mg/kg)组,连续ig给药处理7 d,观察小鼠日常活动变化,苏木素-伊红(hematoxylin eosin,HE)染色和透射电镜观察结肠组织的结构变化,生化检测血清中髓过氧化物酶(myeloperoxidase,MPO)活性。通过脂多糖(lipopolysaccharide,LPS)+三磷酸腺苷(adenosine triphosphate,ATP)诱导THP-1细胞分化构建UC体外模型,将其随机分为对照组、模型组、美沙拉嗪(100μmol/L)组、黄芩苷不同剂量组。酶联免疫吸附测定(enzyme linked immunosorbent assay,ELISA)检测血清和细胞中白细胞介素-1β(interleukin-1β,IL-1β)、IL-18表达;流式检测结肠组织和细胞中半胱氨酸天冬氨酸蛋白酶-1(cystein-asparate protease-1,Caspase-1)表达;Western blotting检测结肠组织和细胞中PTBP1、NLRP3、凋亡相关颗粒样蛋白(apoptosis-associated speck-like protein containing a CARD,ASC)、Caspase-1、GSDMD蛋白C端片段(gasdermin D-C,GSDMD-C)、GSDMD-N蛋白表达;逆转录实时定量聚合酶链式反应(reverse transcription quantitative polymerase chain reaction,RT-qPCR)检测TUGI、PTBP1、NLRP3表达。RNA pull-down和RNA结合蛋白免疫沉淀(RNA immunoprecipitation,RIP)验证TUG1与PTBP1的相互作用;挽救实验进一步验证TUG1-PTBP1在UC中调控巨噬细胞焦亡的作用。结果体内外研究表明,与模型组比较,黄芩苷能够显著降低UC小鼠的疾病活动指数(disease activity index,DAI)评分(P<0.001),抑制结肠缩短,减轻结Objective To explore the mechanism of action of baicalin in regulating the taurine up-regulated gene 1(TUG1)/polypyrimidine tract binding protein 1(PTBP1)/nucleotide-binding oligomerization domain-like receptor protein 3(NLRP3)molecular network to inhibit macrophage pyroptosis for the treatment of ulcerative colitis(UC).Methods UC mouse model was constructed using dextran sulfate sodium salt(DSS)induction,and the mice were randomly divided into control group,model group,mesalazine(600 mg/kg)group,and baicalin low,medium,and high dosage(25,50,and 100 mg/kg)groups,and treated with baicalin administered by consecutive gavage for 7 d,and the changes in the daily activities of mice were observed,and the structural changes in colonic tissues were observed by hematoxylin eosin(HE)staining and transmission electron microscopy,and biochemical detection was performed on the activity of myeloperoxidase(MPO)in serum.An in vitro model of UC was constructed by lipopolysaccharide(LPS)+adenosine triphosphate(ATP)-induced THP-1 cell differentiation,and the mice were randomly divided into control,model,mesalazine(100μmol/L),and baicalin intervention groups with different doses.The expression of interleukin-1β(IL-1β)and IL-18 in serum and cells was detected by enzyme-linked immunosorbent assay(ELISA),the expression of cystein-asparate protease-1(Caspase-1)in colon tissues and cells was detected by flow cytometry;the expression of PTBP1,NLRP3,apoptosis-associated speck-like protein containing a CARD(ASC),Caspase-1,gasdermin D-C(GSDMD-C),GSDMD-N protein in colon tissues and cells was detected byWestern blotting;reverse transcription quantitative polymerase chain reaction(RT-qPCR)was used to detect TUGI,PTBP1,NLRP3 expression.RNA pull-down and RNA immunoprecipitation(RIP)were used to verify the interaction between TUG1 and PTBP1;rescue experiments were used to further validate the effect of TUG1-PTBP1 in UC to regulate macrophage pyroptosis.Results In vitro and in vivo studies showed that baicalin significantly reduced disease acti

关 键 词：黄芩苷 溃疡性结肠炎 巨噬细胞 焦亡 TUG1/PTBP1/NLRP3分子网络 

分 类 号：R285.5[医药卫生—中药学]