Chen Jianfei, Huang Lan, Song Minbao, Yu Shiyong, Gao Pan, Jing Jun
Department of Cardiology, Xinqiao Hospital, Institute of Cardiovascular Disease of PLA, The Third Military Medical University, Chongqing, China.
J Cardiovasc Pharmacol. 2009 May;53(5):359-67. doi: 10.1097/FJC.0b013e31819b5438.
The receptor for advanced glycation end products (RAGE) may play an important role in inflammatory processes and endothelial activation, likely to accelerate the processes of atherosclerosis, especially in patients with diabetes. The factors that regulate the expression of RAGE are not completely clear. C-reactive protein (CRP) is identified as a key proinflammatory cytokine in patients with atherosclerosis. Atherosclerosis also induces reduction and dysfunction of endothelial progenitor cells (EPCs), a main cell type for vascular repair. Therefore, we tested the hypothesis that CRP can regulate RAGE expression and alter antioxidant defenses in EPCs.
EPCs, isolated from bone marrow, were cultured in the absence or presence of lipopolysaccharide-free CRP (5, 10, 15, 20, and 50 microg/mL). RAGE protein expression was measured by flow cytometric analysis and Western blot. RAGE messenger RNA expression was detected by polymerase chain reaction. Reactive oxygen species (ROS) were analyzed using the ROS assay kit. A spectrophotometer was used to assess superoxide dismutase and glutathione peroxidase activity, and polymerase chain reaction was used to test messenger RNA expression of superoxide dismutase and glutathione peroxidase. Apoptosis was evaluated by Annexin V immunostaining. Coculturing with CRP caused a significant upregulate expression of RAGE in EPCs, increased ROS production, altered antioxidant defenses, and induced EPC apoptosis. In addition, these effects were attenuated during blocking RAGE protein expression by small interfering RNA.
CRP at concentrations known to predict cardiovascular event may serve to impair EPC antioxidant defenses for upregulating the expression of RAGE and promote EPC sensitivity toward oxidative stress-mediated apoptosis. These data further support a direct role of CRP in the development and/or progression of atherosclerosis.
晚期糖基化终末产物受体（RAGE）可能在炎症过程和内皮细胞激活中起重要作用，可能会加速动脉粥样硬化进程，尤其是在糖尿病患者中。调节RAGE表达的因素尚不完全清楚。C反应蛋白（CRP）被认为是动脉粥样硬化患者的关键促炎细胞因子。动脉粥样硬化还会导致内皮祖细胞（EPCs）数量减少和功能障碍，而EPCs是血管修复的主要细胞类型。因此，我们验证了CRP可调节RAGE表达并改变EPCs抗氧化防御功能这一假说。
从骨髓中分离出EPCs，在无脂多糖CRP（5、10、15、20和50μg/mL）存在或不存在的情况下进行培养。通过流式细胞术分析和蛋白质印迹法测定RAGE蛋白表达。通过聚合酶链反应检测RAGE信使核糖核酸表达。使用活性氧检测试剂盒分析活性氧（ROS）。用分光光度计评估超氧化物歧化酶和谷胱甘肽过氧化物酶活性，并用聚合酶链反应检测超氧化物歧化酶和谷胱甘肽过氧化物酶的信使核糖核酸表达。通过膜联蛋白V免疫染色评估细胞凋亡。与CRP共培养导致EPCs中RAGE表达显著上调，ROS生成增加，抗氧化防御功能改变，并诱导EPCs凋亡。此外，在通过小干扰RNA阻断RAGE蛋白表达期间，这些作用减弱。
已知可预测心血管事件的浓度的CRP可能通过上调RAGE表达损害EPCs抗氧化防御功能，并促进EPCs对氧化应激介导的凋亡的敏感性。这些数据进一步支持了CRP在动脉粥样硬化发生和/或发展中的直接作用。
