Department of Medicine-Cardiology, German Heart Institute Berlin, Germany.
J Atheroscler Thromb. 2010 Feb 26;17(2):203-12. doi: 10.5551/jat.3004. Epub 2010 Feb 22.
C-reactive protein (CRP) is a pluripotent mediator of inflammation and is present at sites of vascular injury and in atherosclerotic lesions. CRP stimulates endothelial cell adhesion molecule expression and monocyte migration, thereby contributing to the development and progression of vascular lesion formation. In addition, chronic exposure to CRP is known to inhibit angiogenesis and endothelial cell (EC) proliferation.
Whether CRP also affects EC migration, however, has yet to be determined. The present study investigates how long-term exposure to CRP interacts with vascular endothelial growth factor (VEGF) -induced EC migration.
Using a Transwell chamber migration assay, VEGF (20 ng/mL, 5 h incubation)-induced migration of human umbilical vein EC was significantly inhibited in cells pretreated with CRP (10 microg/mL) for 24 h by more than 75%. EC migration in response to VEGF is known to require activation of the protein kinase B (Akt)/endothelial NO synthase (eNOS)- and the extracellular signal-regulated protein kinase 1/2 (ERK1/2) pathway. We therefore investigated the long-term effects of CRP on these signalling events. Immunoblotting with phosphospecific antibodies revealed rapid and transient activation/phosphorylation of the protein kinase Akt within 20 minutes after stimulation with VEGF, which was inhibited by 86% in EC pretreated with CRP (10 microg/mL, 24 h, p<0.05). Subsequent VEGF-induced phosphorylation of eNOS downstream of Akt was completely inhibited in CRP-treated EC. In contrast, CRP-pretreatment did not affect VEGF-induced phosphorylation of ERK1/2. Interestingly, stimulation of EC with CRP for 16-24 h induced marked expression of the phosphatase and tensin homolog (PTEN), which functions as a negative regulator of phosphatidylinositol 3 kinase (PI3K) -->Akt signalling.
The observed time course for CRP-mediated PTEN upregulation corresponds to the exposure time needed for inhibition of Akt phosphorylation and migration and may therefore constitute a potential mechanism by which CRP inhibits inducible Akt phosphorylation and EC migration.
C 反应蛋白 (CRP) 是一种多效炎症介质，存在于血管损伤部位和动脉粥样硬化病变处。CRP 可刺激内皮细胞黏附分子的表达和单核细胞的迁移，从而促进血管损伤形成的发展和进展。此外，长期暴露于 CRP 已知会抑制血管生成和内皮细胞 (EC) 的增殖。
然而，CRP 是否也会影响 EC 的迁移尚不清楚。本研究探讨了长期暴露于 CRP 如何与血管内皮生长因子 (VEGF) 诱导的 EC 迁移相互作用。
使用 Transwell 室迁移分析，发现 CRP(10μg/ml，预处理 24 小时)预处理可使 VEGF(20ng/ml，孵育 5 小时)诱导的人脐静脉 EC 迁移显著抑制超过 75%。已知 VEGF 诱导的 EC 迁移需要激活蛋白激酶 B (Akt)/内皮型一氧化氮合酶 (eNOS)和细胞外信号调节蛋白激酶 1/2 (ERK1/2)途径。因此，我们研究了 CRP 对这些信号事件的长期影响。用磷酸化特异性抗体进行免疫印迹显示，VEGF 刺激后 20 分钟内 Akt 迅速而短暂地被激活/磷酸化，而 CRP(10μg/ml，预处理 24 小时)可抑制 86%的磷酸化(p<0.05)。随后，CRP 处理的 EC 中 Akt 下游的 eNOS 磷酸化完全被抑制。相比之下，CRP 预处理不影响 VEGF 诱导的 ERK1/2 磷酸化。有趣的是，CRP 刺激 EC 16-24 小时可显著诱导磷酸酶和张力蛋白同系物 (PTEN)的表达，PTEN 作为磷脂酰肌醇 3 激酶 (PI3K)–Akt 信号的负调节剂。
观察到的 CRP 介导的 PTEN 上调时间过程与抑制 Akt 磷酸化和迁移所需的暴露时间相对应，因此可能构成 CRP 抑制诱导性 Akt 磷酸化和 EC 迁移的潜在机制。
