基于TLR4/NF-κB/ICAM-1信号通路研究铁皮石斛水提物对高黏血症大鼠的作用  

Effect of Dendrobii Officinalis Caulis water extract on hyperviscosity rats based on TLR4/NF-κB/ICAM-1 signaling pathway

作　　者：陈紫妍 颜美秋 董英杰 苏洁[1] 俞静静 林琦伟 胡雪灵 傅萌 陈素红[2] 吕圭源[1] 陈建真[1] CHEN Zi-yan;YAN Mei-qiu;DONG Ying-jie;SU Jie;YU Jing-jing;LIN Qi-wei;HU Xue-ling;FU Meng;CHEN Su-hong;LYU Gui-yuan;CHEN Jian-zhen(School of Pharmaceutical Sciences,Zhejiang Chinese Medical University,Hangzhou 310053,China;Zhejiang University of Technology,Hangzhou 310014,China)

机构地区：[1]浙江中医药大学药学院,浙江杭州310053 [2]浙江工业大学,浙江杭州310014

出　　处：《中国中药杂志》2024年第19期5261-5272,共12页China Journal of Chinese Materia Medica

基　　金：国家重点研发计划项目(2017YFC1702203);浙江省自然科学基金项目(LY24H280005);浙江省重点实验室项目(2012E10002)。

摘　　要：研究铁皮石斛水提物对高糖、高盐、高脂饮食致高黏血症大鼠的作用与机制。将36只雄性SD大鼠随机分为正常组、模型组、复方丹参片组(0.5 g·kg^(-1))以及铁皮石斛水提物低、中、高剂量组(0.25、0.5、1.0 g·kg^(-1))。除正常组外,其余各组给予高盐、高脂饲料并定时灌胃糖水(20 g·kg^(-1)),连续12周,造模同时灌胃相应药物。实验期间测定抓力、自主活动、眩晕时间、痛阈值等一般体征,同时测定血流变、尾部微循环血流量;流式细胞术检测外周血细胞间黏附分子1(ICAM-1/CD54)、血管细胞黏附分子1(VCAM-1/CD106)阳性细胞率,Ca^(2+)荧光强度;酶联免疫吸附法(ELISA)检测ICAM-1、VCAM-1、血小板内皮细胞黏附分子1(PECAM-1/CD31)、内皮缩血管肽1(ET-1)、血栓素A2(TXA2)、大鼠前列环素I2(PGI2)、一氧化氮(NO)、白细胞介素-1β(IL-1β)、C反应蛋白(CRP)、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)、脂多糖(LPS)水平;苏木素-伊红(HE)、Masson、Gomori醛品红弹力纤维染色观察大鼠结肠、主动脉病理形态学变化;荧光定量PCR法检测主动脉组织中Toll样受体4(TLR4)、髓样分化因子88(MyD88)、核因子κB(NF-κB)、ICAM-1、VCAM-1 mRNA的表达;Western blot法检测结肠组织中紧密连接蛋白occludin、claudin-1和主动脉组织中ICAM-1、VCAM-1、IL-6蛋白的表达;免疫组化和免疫荧光法检测主动脉组织中TLR4、NF-κB的表达。结果显示,铁皮石斛水提物可明显改善高黏血大鼠相关表征,显著降低全血黏度、全血还原黏度和全血流阻,增加尾部微循环血流量,降低外周血CD54、CD106阳性细胞率,改善血行不畅;显著降低血清中ICAM-1、VCAM-1、PECAM-1、ET-1含量,升高PGI2和NO含量,改善血管内皮损伤;降低血清中IL-1β、TNF-α、IL-6、CRP含量,改善血管炎症损伤。此外,铁皮石斛水提物还可以改善结肠、主动脉组织形态,显著升高结肠occludin、claudin-1蛋白表达,显著降低�This study aims to reveal the effect and mechanism of Dendrobii Officinalis Caulis water extract on the rat model of hyperviscosity induced by a high-sugar,high-salt,and high-fat diet.Thirty-six male SD rats were randomized into normal,model,Compound Danshen Tablets(0.5 g·kg^(-1)),and low-,medium-,and high-dose(0.25,0.5,and 1.0 g·kg^(-1),respectively)Dendrobi Officinalis Caulis water extract groups.Except the normal group,the remaining groups were fed with a high-salt and high-fat diet and received regular gavage of sucrose solution(20 g·kg^(-1))for 12 consecutive weeks.The modeling was accompanied by gavage of corresponding drugs.During the experimental period,general signs(such as grip strength,automatic activity,vertigo time,and pain threshold),blood rheology,and blood flow in the tail microcirculation were measured.Flow cytometry was employed to detect the positive cells of intercellular cell adhesion molecule 1(ICAM-1/CD54)and vascular cell adhesion molecule 1(VCAM-1/CD106)in the peripheral blood and the Ca^(2+)fluorescence intensity.Enzyme-linked immunosorbent assay was employed to measure the levels of ICAM-1,VCAM-1,platelet endothelial cell adhesion molecule 1(PECAM-1/CD31),endothelin 1(ET-1),thromboxane A2(TXA2),prostaglandin I2(PGI2),nitric oxide(NO),interleukin-1β(IL-1β),C-reactive protein(CRP),interleukin-6(IL-6),tumor necrosis factor-α(TNF-α),and lipopolysaccharide(LPS).Hematoxylin-eosin,Masson,and Gomori′s aldehyde-fuchsin staining were used to observe the pathological and morphological changes in rat colon and aorta.Fluorescence quantitative PCR was conducted to determine the mRNA levels of Toll-like receptor 4(TLR4),myeloid differentiation factor 88(MyD88),nuclear factor-kappaB(NF-κB),ICAM-1,and VCAM-1 in the aortic tissue.Western blot was employed to determine the protein levels of occludin and claudin-1 in the colon tissue and ICAM-1,VCAM-1,and IL-6 in the aortic tissue.Immunohistochemistry and immunofluorescence were employed to detect the expression of TLR4 and NF-κB in the aortic t

关 键 词：铁皮石斛水提物 高黏血症 肠屏障 炎症 内皮损伤 黏附分子 

分 类 号：R285.5[医药卫生—中药学]