【摘要】 目的:通过观察山莨菪碱(654-2)对激活的血管内皮细胞(VEC)中谷胱甘肽(GSH)含量、核因子-κB(NF-κB)活性和细胞间黏附分子(ICAM-1)表达的影响,探讨其对VEC损伤的保护机制。方法:建立人脐静脉内皮细胞(HUVEC)体外培养体系,经TNF-α刺激和合并应用654-2干预HUVEC后,采用荧光分光光度计法测定HUVEC胞内GSH含量,凝胶电泳迁移率改变分析法(EMSA)检测NF-κB活性和应用免疫组化方法检测HUVEC的ICAM-1表达。结果:HUVEC在TNF-α刺激后GSH含量显著降低(P<0.01),于3 h达低谷;NF-κB的活性显著增高(P<0.01),峰值在刺激后6 h;ICAM-1表达显著增强(P<0.01),并于12 h达峰值。TNF-α刺激并654-2干预后,相应时相点的GSH含量显著降低,但较单纯TNF-α刺激组显著增高(P<0.05),而NF-κB的活性、ICAM-1表达强度较单纯TNF-α刺激显著降低(P<0.05)。结论:654-2通过抑制TNF-α对HUVEC刺激所导致GSH水平的降低,减少了NF-κB活化核易位所启动ICAM-1基因高效表达,这可能是其防治全身炎症反应综合征/脓毒症介导MODS发生的作用环节。更多还原

【Abstract】 Objective:To investigate the mechanisms of anisodamine(654-2) in protection against the vascular endothe-lial cellular(VEC) damages.Methods:Anin vitroculture system of human umbilical venous endothelial cells(HUVEC) was established.After intervention by the combination of TNF-αstimulation and 654-2 treatment,the content of GSH in HUVEC was measured by fluorescence spectrophotometry,and the activity of NF-κB by electrophoretic mobility shift assay (EMSA) and expression of ICAM-1 in HUVEC by immunohistochemistry.Results:The GSHlevel in HUVEC obviously decreased after TNF-α stimulation(P<0.01),and reached the lowest level at 3 h.The activity of NF-κB and the expression of ICAM-1 increased significantly(P<0.01);their peak value was reached at 6 h and 12 h,respectively.The GSH level was negatively correlated with NF-κB activity(r=-0.315,P=0.012) and ICAM-1 expression(r=-0.389,P=0.001),while NF-κB activity was positively correlated with ICAM-1 expression(r=0.261,P=0.026).After TNF-α stimulation and 654-2 intervention,the GSHlevel was obviously reduced but was much higher than after TNF-αstimulation alone(P<0.05),while NF-κB activity and ICAM-1 expression(P<0.05) were lower than after TNF-α stimulation alone(P<0.05).Conclusions:The activation of NF-κB and excessive expression of ICAM-1 could be seen in TNF-α-stimulated VEC.654-2 plays a protective role in prevention of MODS induced by SIRS/sepsis byinhibiting GSH level reduction by TNF-αstimulation and downregulating the expression of ICAM-1 triggered by activation of NF-κB.更多还原