中国药物警戒 ›› 2024, Vol. 21 ›› Issue (5): 535-539.
DOI: 10.19803/j.1672-8629.20230566

• 基础与临床研究 • 上一篇    下一篇

枯芩与条芩提取物对湿热型溃疡性结肠炎大鼠Th17/Treg免疫失衡的调节机制研究

黄伟1,2, 罗雅琴, 于新阳3, 董红敬2, 王晓2,*   

  1. 1山东省中医药研究院中药药理研究所药理实验中心,山东 济南 250014;
    2齐鲁工业大学（山东省科学院）山东省分析测试中心,山东 济南 250014;
    3山东中医药大学附属医院第一临床学院,山东 济南 250014
  • 收稿日期:2023-09-08 出版日期:2024-05-15 发布日期:2024-05-13
  • 通讯作者: *王晓,男,博士,研究员·博导,中药资源与药效物质基础研究。E-mail: wxjn1998@126.com
  • 作者简介:黄伟,男,博士,副研究员·硕导,中药药效物质基础及作用机理研究。△为并列第一作者。
  • 基金资助:
    中央本级重大增减支项目“名贵中药资源可持续利用能力建设项目”(2060302); 国家自然科学基金资助项目（81704029）; 山东省自然科学基金联合基金重点资助项目（ZR 2022LZY007）; 山东省中医药高层次人才培育项目（鲁卫人才字[2023] 37号）; 齐鲁卫生与健康杰出青年人才资助项目（鲁卫人才字[2020] 3号）; 山东省中医药科技项目（Z-2022098T、M-2022027）

Mechanisms of extracts of Pith-nodecayed and Pith-decayed of Scutellariae Radix against Th17/Treg immune imbalance in damp-heat ulcerative colitis rats

HUANG Wei1,2, LUO Yaqin, YU Xinyang3, DONG Hongjing2, WANG Xiao2,*   

  1. 1Shandong Academy of Chinese Medicine, Institute of Chinese Materia Medica Pharmacology (Pharmacological Experimental Center), Jinan Shandong 250014, China;
    2Qilu University of Technology (Shandong Academy of Sciences), Shandong Analysis and Test Center, Jinan Shandong 250014, China;
    3Affiliated Hospital of Shandong University of Chinese Medicine (First Clinical College), Jinan Shandong 250014, China
  • Received:2023-09-08 Online:2024-05-15 Published:2024-05-13

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摘要： 目的 比较枯芩、条芩提取物对湿热型溃疡性结肠炎（ulcerative colitis,UC）大鼠模型辅助性T细胞17（Th17）/调节性T细胞（Treg）细胞因子及转录因子维甲酸相关孤儿受体（ROR-γt）/叉头翼状螺旋转录因子3（Foxp3）表达的影响。方法 50只健康SD大鼠随机分为5组：正常组、模型组、枯芩组、条芩组和美莎拉嗪组,每组10只。采用高脂高糖饲料喂养+高度白酒灌胃+5%葡聚糖硫酸钠（dextran sulfate sodium, DSS）联合诱导法,建立湿热型UC大鼠模型;各组分别于造模第1天开始灌胃给药,枯芩组、条芩组给药剂量均为5.25 g·kg-1·d-1,美沙拉嗪组给药剂量为0.266 g·kg-1·d-1,正常组、模型组灌服等体积生理盐水。连续给药28 d。给药结束后,麻醉大鼠,腹主动脉取血,分离血清,Elisa法检测大鼠血清白介素-6（IL-6）、IL-17A、IL-10、转化生长因子-β1（TGF-β1）水平;分离脾脏和结肠组织,采用流式细胞术检测脾脏Th17、Treg细胞比例,计算Th17/Treg比值;采用实时荧光定量聚合酶链式反应（qRT-PCR）法检测结肠组织ROR-γt、Foxp3 mRNA表达水平;免疫蛋白印迹（western blot）法检测结肠组织ROR-γt、Foxp3蛋白表达水平。结果 与正常组比较,模型组大鼠血清IL-6、IL-17A、脾脏Th17细胞比例、Th17/Treg、结肠组织ROR-γt mRNA和蛋白表达水平均显著增加（P<0.01）,血清IL-10、IL-35、TGF-β1、脾脏Treg细胞比例、结肠组织Foxp3 mRNA和蛋白表达水平均显著降低（P<0.01）;给药28 d后,与模型组比较,各给药组大鼠血清IL-6、IL-17A、脾脏Th17细胞比例、Th17/Treg、结肠组织ROR-γt mRNA和蛋白表达水平出现不同程度的降低（P<0.05或P<0.01）,血清IL-10、IL-35、TGF-β1、脾脏Treg细胞比例、结肠组织Foxp3 mRNA和蛋白表达水平出现不同程度的升高（P<0.05或P<0.01）;与枯芩组比较,条芩组、美沙拉嗪组在降低IL-6、IL-17A,增加IL-10、IL-35、TGF-β1水平,调节ROR-γt mRNA和蛋白表达方面（P<0.05或P<0.01）均优于枯芩组。结论 枯芩、条芩可能通过调控转录因子ROR-γt、Foxp3表达,恢复Th17/Treg免疫失衡,抑制炎症反应,从而保护肠道黏膜屏障受损,发挥治疗湿热型UC的作用。

关键词: 黄芩, 枯芩, 条芩（或子芩）, 提取物, 湿热型溃疡性结肠炎, Th17/Treg失衡, 细胞因子, ROR-γt, Foxp3, 大鼠

Abstract: Objective To compare the effects of extracts from Pith-decayed and Pith-nodecayed products of Scutellariae Radix on expressions of helper T cell 17 (Th17)/regulatory T cell (Treg) cytokines and the transcription factor ROR-γt/ Foxp3 in rats with damp-heat ulcerative colitis (UC). Methods Fifty healthy SD rats were randomly divided into five groups: the normal group, model group, Pith-decayed products of Scutellariae Radix group, Pith-nodecayed products of Scutellariae Radix group and mesalazine group, with 10 rats in each. A damp-heat UC rat model was established via the combination of high-fat and high-sugar diet+high-liquor administration+5% dextran sodium sulfate (DSS). Each group was given intragastric administration on the first day of modeling, respectively. The dosage was 5.25 g·kg-1·d-1 for the Pith-decayed products group and Pith-nodecayed products group, but 0.266 g·kg-1·d-1 for the mesalazine group. The normal group and model group were given an equal volume of normal saline. The drug was administered for 28 consecutive days. After administration, the rats were anesthetized, blood was collected from the abdominal aorta, serum was separated, and serum levels of interleukin-6(IL-6), IL-17A, IL-10 and transforming growth factor-β1(TGF-β1) were detected by Elisa. Spleen and colon tissues were separated, the proportion of Th17 and Treg cells in spleens was detected by flow cytometry, and the Th17/Treg ratio was calculated. Real-time quantitative fluorescent polymerase chain reaction (qRT-PCR) was used to detect the mRNA expression levels of ROR-γt and Foxp3 in colon tissues. The expression levels of ROR-γt and Foxp3 in colon tissues were detected by Western blot. Results Compared with the normal group, serum levels of IL-6 and IL-17A, the proportion of Th17 cells in spleens, Th17/Treg, and mRNA and protein expression levels of ROR-γt in colon tissues of the model group were significantly increased (P<0.01) while serum levels of IL-10, IL-35, TGF-β1, the proportion of Treg cells in spleens and the expression of Foxp3 mRNA and protein in colon tissues were significantly decreased (P<0.01). After 28 days of administration, serum levels of IL-6, IL-17A, the Th17 cell ratio in spleens, Th17/Treg, and ROR-γt mRNA and protein expression levels in colon tissues of rats in each administration group were decreased to varying degrees compared with the model group (P<0.05 or P<0.01), while serum levels of IL-10, IL-35, TGF-β1, the proportion of Treg cells in spleens, and the expression of Foxp3 mRNA and protein in colon tissues were increased to different extents (P<0.05 or P<0.01). Compared with the Pith-decayed products of Scutellariae Radix group, Pith-nodecayed products of Scutellariae Radix group and mesalazine group were more effective in reducing levels of IL-6 and IL-17A, increasing levels of IL-10, IL-35 and TGF-β1, and regulating mRNA and protein expressions of ROR-γt (P<0.05 or P<0.01). Conclusion Pith-decayed and Pith-nodecayed products of Scutellariae Radix may help regulate the expressions of transcription factors ROR-γt and Foxp3, restore Th17 /Treg immune imbalance, inhibit inflammatory response, protect against intestinal mucosal barrier damage, and facilitate the treatment of damp-heat UC.

Key words: Scutellariae Radix, Pith-decayed, Pith-nodecayed, extracts, damp-heat ulcerative colitis, Th17/Treg imbalance, cytokine, ROR-γt, Foxp3, rats

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