Verma Subodh, Badiwala Mitesh V, Weisel Richard D, Li Shu-Hong, Wang Chao-Hung, Fedak Paul W M, Li Ren-Ke, Mickle Donald A G
Division of Cardiac Surgery, Toronto General Hospital, University of Toronto, Ontario, Canada.
J Thorac Cardiovasc Surg. 2003 Dec;126(6):1886-91. doi: 10.1016/j.jtcvs.2003.07.026.
Elevated levels of C-reactive protein are one of the strongest prognostic factors in atherosclerosis. In addition to predicting vascular disease, C-reactive protein may directly facilitate the development of a proinflammatory and proatherosclerotic phenotype. Recent studies have demonstrated marked up-regulation of various adhesion molecules and inflammatory responses in endothelial cells subjected to C-reactive protein. The nuclear factor-kappaB signal transduction is known to play a key role in the expression of these proatherogenic entities. This study examines the direct effects of C-reactive protein on nuclear factor-kappaB activation and related mechanisms in saphenous vein endothelial cells.
The activation of nuclear factor-kappaB was determined by confocal microscopy assessing the nuclear localization of nuclear factor-kappaB in endothelial cells incubated with C-reactive protein (50 microg/mL) for 30 minutes and 3 hours. Cells not incubated with C-reactive protein were used as negative controls, and cells incubated with tumor necrosis factor-alpha (10 ng/mL) for 15 minutes were used as positive controls in all studies. The degradation of IkappaB-alpha and IkappaB-beta was assessed by Western blotting of the cell lysates obtained from cells incubated with human recombinant C-reactive protein (50 microg/mL) for 15 minutes, 30 minutes, and 1 hour.
Nuclear factor-kappaB nuclear translocation in endothelial cells increased significantly after 30 minutes of incubation with C-reactive protein (P <.01). Nuclear localization of nuclear factor-kappaB returned to baseline levels after 3 hours of incubation with C-reactive protein. Incubation with C-reactive protein resulted in degradation of IkappaB-alpha that was maximal at 30 minutes (P <.05). C-reactive protein showed no significant effect on IkappaB-beta degradation.
These data demonstrate, for the first time, that C-reactive protein activates the nuclear factor-kappaB signal transduction pathway in endothelial cells. Degradation of IkappaB-alpha, but not IkappaB-beta, seems to be the major pathway leading to nuclear factor-kappaB nuclear translocation and activation induced by C-reactive protein. These data support the concept that C-reactive protein, at concentrations known to predict diverse vascular insults, directly facilitates a proinflammatory and proatherosclerotic phenotype through activation of nuclear factor-kappaB. These data have important implications for saphenous vein atherosclerosis in patients with elevated C-reactive protein levels.
C反应蛋白水平升高是动脉粥样硬化最强的预后因素之一。除了预测血管疾病外，C反应蛋白可能直接促进促炎和促动脉粥样硬化表型的发展。最近的研究表明，在受到C反应蛋白作用的内皮细胞中，各种黏附分子和炎症反应显著上调。已知核因子-κB信号转导在这些促动脉粥样硬化因子的表达中起关键作用。本研究探讨C反应蛋白对大隐静脉内皮细胞核因子-κB激活的直接影响及其相关机制。
通过共聚焦显微镜评估与C反应蛋白（50μg/mL）孵育30分钟和3小时的内皮细胞中核因子-κB的核定位，来确定核因子-κB的激活情况。在所有研究中，未与C反应蛋白孵育的细胞用作阴性对照，与肿瘤坏死因子-α（10ng/mL）孵育15分钟的细胞用作阳性对照。通过对与人重组C反应蛋白（50μg/mL）孵育15分钟、30分钟和1小时的细胞所获得的细胞裂解物进行蛋白质印迹分析，评估IκB-α和IκB-β的降解情况。
与C反应蛋白孵育30分钟后，内皮细胞中核因子-κB的核转位显著增加（P<.01）。与C反应蛋白孵育3小时后，核因子-κB的核定位恢复到基线水平。与C反应蛋白孵育导致IκB-α降解，在30分钟时达到最大程度（P<.05）。C反应蛋白对IκB-β降解无显著影响。
这些数据首次证明，C反应蛋白在内皮细胞中激活核因子-κB信号转导途径。IκB-α而非IκB-β的降解似乎是导致C反应蛋白诱导核因子-κB核转位和激活的主要途径。这些数据支持以下概念，即已知能预测多种血管损伤的浓度的C反应蛋白，通过激活核因子-κB直接促进促炎和促动脉粥样硬化表型。这些数据对于C反应蛋白水平升高患者的大隐静脉动脉粥样硬化具有重要意义。
