基于TLR4/NF-κB/NLRP3和PI3K/Akt通路研究甘遂及其炮制品对水负荷小鼠的利尿作用及机制  

Diuretic effect and mechanism of Kansui Radix and its prepared products on water-loaded mice through TLR4/NF-κB/NLRP3 and PI3K/Akt signaling pathways

作　　者：宁晨旭 苏晓娟 任可乐 刘晓琴 孟祥龙 赵彩蓉 赵泓瑜 王娟 张朔生 NING Chen-xu;SU Xiao-juan;REN Ke-le;LIU Xiao-qin;MENG Xiang-long;ZHAO Cai-rong;ZHAO Hong-yu;WANG Juan;ZHANG Shuo-sheng(College of Chinese Medicine and Food Engineering,Shanxi University of Chinese Medicine,Jinzhong 030619,China;Key Laboratory of Chinese Medicine Processing in Shanxi Province,Shanxi University of Chinese Medicine,Jinzhong 030619,China)

机构地区：[1]山西中医药大学中药与食品工程学院,山西晋中030619 [2]山西中医药大学中药炮制山西省重点实验室,山西晋中030619

出　　处：《中草药》2022年第13期4007-4018,共12页Chinese Traditional and Herbal Drugs

基　　金：山西应用基础研究计划面上自然科学基金资助项目(201901D111342);山西省中医药大学中药炮制学科建设项目(2018-1);山西省中医药大学中药炮制与方药配伍研究科技创新团队(2018TD-006)。

摘　　要：目的研究甘遂及其炮制品对水负荷小鼠的利尿作用及机制。方法通过网络药理学探究甘遂的利尿作用机制。ICR小鼠给予甘遂及其炮制品后,ig大量生理盐水建立水负荷模型,测定水负荷差值及脏器指数;取十二指肠及空肠,观察病理变化;采用ELISA法测定小鼠血清中炎性因子指标白细胞介素-1β(interleukin-1β,IL-1β)、IL-18、IL-4、IL-10及小肠组织中氧化应激指标丙二醛(malondialdehyde,MDA)和还原型谷胱甘肽(glutathione,GSH)水平;采用Western blotting测定小鼠小肠组织中Toll样受体4(Toll-like receptor 4,TLR4)、核因子-κB(nuclear factor-κB,NF-κB)、NOD样受体蛋白3(NOD-like receptor protein 3,NLRP3)、磷脂肌醇-3-激酶(phosphatidylinositol-3-kinase,PI3K)和蛋白激酶B(protein kinase B,Akt)蛋白表达。结果通过构建“中药-成分-疾病-通路-靶点”网络及通路富集分析得到甘遂及其炮制品主要通过参与氧化应激反应及PI3K/Akt等信号通路调节水肿。体内实验结果显示,与模型组比较,各给药组小鼠水负荷差值在各个时间段均显著升高(P<0.01);血清中IL-1β、IL-18及小肠组织MDA水平均显著降低(P<0.05、0.01),血清中IL-4、IL-10及小肠组织GSH水平均显著升高(P<0.05、0.01);小肠组织TLR4、NLRP3、p-NF-κB/NF-κB、p-PI3K/PI3K及p-Akt/Akt蛋白表达水平均显著降低(P<0.01)。在各给药组中,1~6 h甘遂的利尿作用最强,醋甘遂和甘草制甘遂的利尿作用均有所缓解,说明甘遂经炮制后会缓和其刺激性而平衡其利尿作用。结论甘遂及其炮制品对水负荷小鼠均可以不同程度地抗水负荷并产生利尿作用,并可以通过抑制TLR4通道介导的NF-κB/NLRP3信号通路及PI3K/Akt信号通路从而抑制IL-1β和IL-18的表达。Objective To study the diuretic effect and mechanism of Gansui(Kansui Radix)and its prepared products on water-loaded mice.Methods Diuretic effects of Kansui Radix was explored by network pharmacology.After ICR mice were given Kansui Radix and its processed products,a large amount of normal saline was used to establish a water load model,water load difference and organ index were measured;Duodenum and jejunum were taken to observe the pathological changes;ELISA method was used to measure levels of inflammatory factor indicators interleukin-1β(IL-1β),IL-18,IL-4,IL-10 in serum and oxidative stress indicators malondialdehyde(MDA)and glutathione(GSH)in small intestinal tissue;Western blotting was used to measure protein expressions of Toll-like receptor 4(TLR4),nuclear factor-κB(NF-κB),NOD-like receptor protein 3(NLRP3),phosphatidylinositol-3-kinase(PI3K)and protein kinase B(Akt)in small intestinal tissue.Results Kansui Radix and its processed products mainly regulated edema by participating in oxidative stress response and PI3K/Akt and other signaling pathways through the construction of“traditional Chinese medicine-ingredient-disease-pathway-target”network and pathway enrichment analysis.The results of in vivo experiments showed that compared with model group,water load difference of mice in each administration group was significantly increased at each time period(P<0.01);Levels of IL-1β,IL-18 in serum and MDA in small intestinal tissue were significantly decreased(P<0.05,0.01),levels of IL-4,IL-10 in serum and GSH in small intestinal tissue were significantly increased(P<0.05,0.01);TLR4,NLRP3,p-NF-κB/NF-κB,p-PI3K/PI3K and p-Akt/Akt protein expressions in small intestinal tissue were significantly decreased(P<0.01).In each administration group,diuretic effect of Kansui Radix was the strongest at 1—6 h,and diuretic effect of vinegar Kansui Radix and Glycyrrhizae Radix et Rhizoma processed Kansui Radix were relieved,indicating that processed Kansui Radix would ease its irritation and balance its diuretic

关 键 词：甘遂 甘草制甘遂 醋甘遂 水负荷模型 炎性细胞因子 磷脂肌醇-3-激酶/蛋白激酶B信号通路 NOD样受体蛋白3 

分 类 号：R285.5[医药卫生—中药学]