Role of NMDA receptor in homocysteine-induced activation of Mitogen-Activated Protein Kinase and Phosphatidyl Inositol 3-Kinase pathways in cultured human vascular smooth muscle cells
Role of NMDA receptor in homocysteine-induced activation of Mitogen-Activated Protein Kinase and Phosphatidyl Inositol 3-Kinase pathways in cultured human vascular smooth muscle cells
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DOI：
10.1016/j.thromres.2009.08.015
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发表时间：
2010-02-01
影响因子：
7.5
通讯作者：
Anfossi, Giovanni
中科院分区：
文献类型：
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作者：
Doronzo, Gabriella;Russo, Isabella;Anfossi, Giovanni
简介：血管平滑肌细胞（VSMC）暴露于与心血管事件风险增加相关的浓度的同型半胱氨酸，增强基质金属蛋白酶-2（MMP-2）的合成和分泌，其参与动脉粥样硬化斑块不稳定。这种作用被丝裂原活化蛋白激酶（MAPK）和磷脂酰肌醇3-激酶（PI 3-K）通路的抑制剂阻止，从而假设同型半胱氨酸可能通过受体介导的机制激活这两条通路。一种可能的受体是N-甲基-D-天冬氨酸受体（NMDAr），其在VSMC中表达并且参与其它细胞类型中的同型半胱氨酸效应。（两者均为100 μ mol/L; 5 min-8 h），通过测量进行研究：i）ERK 1/2、p38 MAPK的磷酸化（MAPK途径的信号分子）和Akt和p70 S6 K（PI 3-K途径的信号传导分子）; ii）MMP-2的合成和分泌（蛋白质印迹）; iii）MMP-2的活化（明胶色谱法）。为了评价NMDAr参与高半胱氨酸效应，在非竞争性NMDAr拮抗剂MK-801（50 μ mol/L）或L-甘氨酸（10 μ mol/L）存在下重复实验，所述非竞争性NMDAr拮抗剂MK-801或L-甘氨酸通过促进其内化来抑制NMDAr功能。结果如下：DL-同型半胱氨酸和NMDA时间依赖性地增加：i）ERK 1/2、p38 MAPK、Akt和p70 S6 K的磷酸化（ANOVA，p < 0.0001）; ii）MMP-2的合成、分泌和活化。DL-同型半胱氨酸和NMDA的影响被阻止VSMC预孵育MK-801或高浓度的L-甘氨酸concentration.Conclusions：在人VSMC同型半胱氨酸的浓度与心血管风险增加相关的激活MAPK和PI 3-K途径和MMP-2的合成和分泌，通过NMDA受体，一个潜在的机制，参与细胞内信号在响应同型半胱氨酸在VSMC。(C)2009爱思唯尔有限公司保留所有权利。
Introduction: Exposure of vascular smooth muscle cells (VSMC) to homocysteine, at concentrations associated with an increased risk of cardiovascular events, enhances synthesis and secretion of Matrix Metalloproteinase-2 (MMP-2), which is involved in atherosclerotic plaque instabilization. This effect was prevented by inhibitors of Mitogen Activated Protein Kinase (MAPK) and Phosphatidylinositol 3-Kinase (PI3-K) pathways, allowing to hypothesize that homocysteine activates both these pathways, likely via a receptor-mediated mechanism. One possible receptor is N-methyl-D-aspartate receptor (NMDAr), which is expressed in VSMC and is involved in homocysteine effects in other cell types.Materials and Methods: VSMC exposed to DL-homocysteine or NMDA (100 mu mol/L for both; 5 min-8 hours), were investigated by measuring: i) phosphorylation of ERK1/2, p38MAPK (signaling molecules of MAPK pathway) and Akt and p70S6K (signaling molecules of PI3-K pathway) by western blot; ii) synthesis and secretion of MMP-2 (western blot); iii) activation of MMP-2 (gelatin zimography). To evaluate NMDAr involvement in the homocysteine effects, the experiments were repeated in the presence of a non-competitive NMDAr-antagonist MK-801 (50 mu mol/L) or L-glycine (10 mu mol/L), which inhibits NMDAr function by promoting its internalization. Results: DL-homocysteine and NMDA time-dependently increased: i) the phosphorylation of ERK1/2, p38 MAPK, Akt and p70S6K (ANOVA, p < 0.0001); ii) the synthesis, secretion and activation of MMP-2. DL-homocysteine and NMDA effects were prevented by VSMC pre-incubation with MK-801 or high L-glycine concentrations.Conclusions: In human VSMC homocysteine-at concentrations associated with increased cardiovascular risk-activates MAPK and PI3-K pathways and MMP-2 synthesis and secretion through NMDA receptor, a potential mechanism involved in intracellular signaling in response to homocysteine in VSMC. (C) 2009 Elsevier Ltd. All rights reserved.