白藜芦醇衍生物TMS对脂多糖诱导人脐静脉内皮细胞中一氧化氮、血管细胞间黏附分子-1及核转录因子-κB表达的影响  

Effect of resveratrol derivative trans-3,5,4′-trimethoxystilbene on expressions of nitric oxide, vascular cell adhesion molecule-1 and nuclear factor-κB in human umbilical vein endothelial cells induced by lipopolysaccharide

作　　者：付海燕[1] 胡占升[1] 杜红阳[2] 周荣斌[3] 

机构地区：[1]辽宁医学院附属第一医院ICU,辽宁锦州121000 [2]辽宁医学院附属第一医院皮肤科,辽宁锦州121000 [3]北京军区总医院急诊科,北京100700

出　　处：《中国生物制品学杂志》2015年第8期808-813,共6页Chinese Journal of Biologicals

基　　金：辽宁省科技厅自然科学基金(2013022059);中国博士后科学基金(2015M572776);辽宁医学院青年科技启动基金(Y2011Z010)

摘　　要：目的探讨白藜芦醇衍生物TMS(trans-3,5,4′-trimethoxystilbene)对脂多糖(lipopolysaccharide,LPS)诱导人脐静脉内皮细胞(human umbilical vein endothelial cells,HUVECs)中一氧化氮(nitric oxide,NO)、血管细胞间黏附分子-1(vascular cell adhesion molecule-1,VCAM-1)及核转录因子-κB(nuclear factor-κB,NF-κB)表达的影响。方法取P3代HUVECs,采用CCK-8法检测不同TMS浓度(0、5、10、20、50、100μmol/L)对HUVECs细胞存活率的影响。将P6代HUVECs分为正常对照组、LPS组、TMS(高、中、低浓度)+LPS组及PDTC+LPS组,Griess法检测各组细胞产生NO浓度;Real-time PCR法检测各组细胞中VCAM-1、NF-κB p65基因m RNA的转录水平;Western blot法检测各组细胞中VCAM-1、NF-κB p65和IκBα蛋白的表达水平;免疫细胞化学染色法检测各组细胞中VCAM-1和NF-κB p65蛋白的表达水平。结果 5、10μmol/L浓度组TMS对细胞存活率影响较小,50、100μmol/L浓度组中细胞生存率明显下降(P<0.05),且呈时间、剂量依赖性。低、中、高浓度TMS+LPS组与PDTC+LPS组细胞中NO浓度显著下降(P<0.05);中浓度TMS+LPS组和PDTC+LPS组细胞中VCAM-1、NF-κB p65基因m RNA转录和蛋白表达水平均明显低于LPS组(P<0.05),IκBα蛋白表达水平明显高于LPS组(P<0.05);正常对照组细胞胞质中有弱VCAM-1蛋白荧光表达,未见NF-κB p65蛋白荧光表达,LPS组可见VCAM-1和NF-κB p65蛋白的强荧光表达,中浓度TMS+LPS组和PDTC+LPS组中VCAM-1和NF-κB p65蛋白荧光表达均弱于LPS组。结论白藜芦醇衍生物TMS可抑制LPS诱导HUVECs表达NO、VCAM-1和NF-κB p65,且其抑制VCAM-1效应可能是通过NF-κB细胞信号通路而发挥作用的,本研究为临床治疗血管内皮功能紊乱相关疾病提供了实验依据。Objective To investigate the effect of trans-3,5,4′-trimethoxystilbene（TMS）, a derivative of resveratrol, on the expressions of nitric oxide（NO）, vascular cell adhesion molecule-1（VCAM-1）and nuclear factor（NF）-κB in human umbilical vein endothelial cells（HUVECs） induced by lipopolysaccharide（LPS）. Methods The HUVECs of passage 3（P3）were treated with TMS at various concentrations（0, 5, 10, 20, 50 and 100 μmol / L）and determined for survival rate.The HUVECs of P6 were treated with LPS, TMS at high, moderate and low doses ＋ LPS and PDTC ＋ LPS respectively,using those untreated as normal control, and determined for NO concentration by Griess method, for transcription levels of VCAM-1 and NF-κB p65 m RNAs by real-time PCR, for expression levels of VCAM-1 and NF-κB p65 proteins by Western blot and immunocytochemetry assay, and for expression level of IκBα protein by Western blot. Results The TMS at concentrations of 5 and 10 μmol / L showed little effect on survival rate of cells. However, the survival rates of cells treated with 50 and 100 μmol / L TMS decreased significantly in time- and dose-dependent modes（P 〈 0. 05）. The NO concentrations in cells treated with low, moderate and high dose TMS ＋ LPS and PDTC ＋ LPS groups decreased significantly（P 〈 0. 05）. The m RNA transcription and protein expression levels of VCAM-1 and NF-κB p65 in cells in moderate dose TMS ＋ LPS and PDTC ＋ LPS groups were significantly lower, while the expression level of IκBα protein was significantly higher, than those in LPS group（each P 〈 0. 05）. Low expression of VCAM-1 was observed in cytoplasm of cells in normal control group, but no expression of NF-κB p65 protein. Both VCAM-1 and NF-κB p65 proteins were highly expressed in LPS group, while lowly expressed in moderate dose TMS ＋ LPS and PDC ＋ LPS groups. Conclusion TMS inhibited the expressions of NO, VCAM-1 and NF-κB p65 in HUVECs induced by LPS, of which the inhibitory mechanism to VCAM-1 might be NF-κ

关 键 词：白藜芦醇衍生物TMS 脂多糖 人脐静脉内皮细胞 一氧化氮 血管细胞间黏附分子-1 核转录因子-ΚB 

分 类 号：R765.21[医药卫生—耳鼻咽喉科]