黄芩苷对脂多糖致内皮细胞炎症反应的保护作用及机制研究  

The protective effect of baicalin on inflammatory response of endothelial cells induced by lipopolysaccharide and its mechanism

作　　者：路青瑜 郭丽 张启云 杨付梅 李娇 孙黔云 LU Qing-yu;GUO Li;ZHANG Qi-yun;YANG Fu-mei;LI Jiao;SUN Qian-yun(State Key Lab of Function and Applications of Medicinal Plants, Guizhou Medical University,The Key Lab of Chemistry for Natural Products, Guizhou Province and Chinese Academy of Science, Guiyang 550014, China)

机构地区：[1]贵州医科大学省部共建药用植物功效与利用国家重点实验室,贵州省中国科学院天然产物化学重点实验室,贵州贵阳550014

出　　处：《中国药理学通报》2021年第2期251-257,共7页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No U1812403);贵州省科技计划及平台人才项目[黔科合人才(2016)4018号、黔科合平台人才(2016)5625号、黔科合平台人才(2019)5702号];贵州省科技计划项目[黔科合基础(2017)115号]。

摘　　要：目的研究黄芩苷对脂多糖(LPS)致人微血管内皮细胞(HMEC)炎症反应的保护作用及作用机制。方法采用LPS作用HMEC,建立炎症反应模型;以不同浓度的黄芩苷预处理细胞,然后将细胞暴露于LPS,ELISA检测炎症因子ICAM-1、IL-6和MCP-1的含量;荧光观察Ca^2+内流并采用流式细胞仪检测细胞内Ca^2+水平;免疫荧光法检测NF-κB p65入核情况;双荧光素酶报告基因方法检测NF-κB核内转录活性;Western blot检测NF-κB p65、p-NF-κB p65及TLR4的表达。结果HMEC暴露于LPS后,出现了明显的Ca^2+内流,NF-κB p65发生磷酸化,核内转录活性上调,ICAM-1、IL-6及MCP-1表达上调。不同浓度的黄芩苷均能抑制LPS刺激HMEC后产生的Ca^2+内流、NF-κB p65入核及核内转录活性上调,减少ICAM-1、IL-6、MCP-1的表达,下调NF-κB p65、p-NF-κB p65及TLR4的表达,且抑制作用呈现一定的量效关系。结论黄芩苷能抑制LPS诱导的HMEC炎症反应,其机制与抑制Ca^2+内流和NF-κB信号通路的活化,从而降低炎症反应的水平有关。Aim To investigate the protective effect of baicalin on inflammatory response of human microvascular endothelial cells(HMECs)induced by lipopolysaccharides(LPS)and its mechanism.Methods LPS was applied to establish inflammatory model on HMECs in this work.HMECs were pretreated with different concentrations of baicalin(1.0×10-6,1.0×10-7 and 1.0×10-8 mol·L^-1),and then exposed to LPS.The supernatant was removed and assayed for expression of the adhesion molecule ICAM-1,the inflamatory mediator IL-6 and MCP-1 by using ELISA reagent kits.The inward flow of calcium ion was observed by fluorescence microscope,and the intracellular calcium ion level was measured by flow cytometry.The fluorescence intensity of nucleus NF-κB p65 was detected by immunoflourescent technique.The nucleus transcriptional activity of NF-κB was measured by the dual luciferase reporter assay system.Moreover,the expression of protein NF-κB p65,phospo-NF-κB p65(p-p65)and Toll like receptor 4(TLR4)were detected by Western blot.Results The internal flow of calcium,the phosphorylation of NF-κB p65 and the transcription activity significantly increased,and the expression of ICAM-1,IL-6 and MCP-1 up-regulated after HMECs were exposed to LPS.Baicalin inhibited the inward flow of calcium ion,the nuclear translocation of NF-κB p65 and the up-regulation of transcriptional activity,decreased the expression of ICAM-1,IL-6 and MCP-1,and down-regulated the expression of NF-κB p65,p-p65 and TLR4 in a dose-dependent manner.Conclusions Baicalin possesses a protective effect on inflammatory response of endothelial cells induced by LPS,and its mechanism may be highly related to the inhibition of the internal flow of calcium and the activation of NF-κB signaling pathway,and thus reduce the level of inflammatory response.

关 键 词：黄芩苷 脂多糖 内皮细胞 NF-κB TLR4 炎症反应 钙离子内流 

分 类 号：R284.1[医药卫生—中药学] R322.12[医药卫生—中医学]