Tarín Carlos, Gomez Monica, Calvo Enrique, López Juan Antonio, Zaragoza Carlos
Fundación Centro Nacional de Investigaciones Cardiovasculares, Melchor Fernández Almagro 3, 28029 Madrid, Spain.
Arterioscler Thromb Vasc Biol. 2009 Jan;29(1):27-32. doi: 10.1161/ATVBAHA.108.169623. Epub 2008 Nov 6.
Lack of endothelial nitric oxide synthase worsens atherosclerosis at least by increasing monocyte adhesion to endothelial cells. The purpose of this study was to elucidate the molecular mechanism elicited by NO.
We evaluated atherosclerosis in apoE and NOS3/apoE-deficient mice fed with high-cholesterol diet. We found significant increase in aortic lesion size, and infiltration of macrophages in NOS3/apoE-null mice when compared to apoE-deficient animals. To test the relevance of cellular adhesion as well as extracellular matrix degradation, we evaluated ICAM-1, VCAM-1, PECAM-1, MMP-2, MMP-9, MMP-12, MT1-MMP, and MMP-13 levels in mouse aortas. Lack of NO inhibits MMP-13 and increases ICAM-1 levels in atherosclerosis as compared to apoE-null mice. Ectopically expression of ICAM-1 in eukaryotic cells revealed that extracellular domain of ICAM-1 harbors a substrate recognized by MMP-13. Incubation of COS-7 cells expressing ectopic ICAM-1 in the presence of active MMP-13 induced inhibition of RAW 264.7 cell adhesion to COS-7 monolayers. MALDI-TOF MS analysis combined to Liquid chromatography coupled to Ion Trap MS on ICAM-1 incubated with MMP-13 allowed us to determine the cleavage sites of MMP-13 at positions E61 and G98 of ICAM-1. G98 is part of a PDGQS moiety, which shows homology with the consensus PDGLS substrate located at the MMP-13 cleaved site of type II collagen I-alpha.
Taking together, these results point toward MMP-13 as a mechanism for the NO-mediated protection of atherosclerosis.
内皮型一氧化氮合酶的缺乏至少通过增加单核细胞与内皮细胞的黏附而使动脉粥样硬化恶化。本研究的目的是阐明一氧化氮引发的分子机制。
我们评估了喂食高胆固醇饮食的载脂蛋白E和一氧化氮合酶3/载脂蛋白E缺陷小鼠的动脉粥样硬化情况。我们发现，与载脂蛋白E缺陷动物相比，一氧化氮合酶3/载脂蛋白E基因敲除小鼠的主动脉病变大小显著增加，巨噬细胞浸润增多。为了测试细胞黏附以及细胞外基质降解的相关性，我们评估了小鼠主动脉中细胞间黏附分子-1、血管细胞黏附分子-1、血小板内皮细胞黏附分子-1、基质金属蛋白酶-2、基质金属蛋白酶-9、基质金属蛋白酶-12、膜型基质金属蛋白酶-1和基质金属蛋白酶-13的水平。与载脂蛋白E基因敲除小鼠相比，一氧化氮的缺乏在动脉粥样硬化中抑制基质金属蛋白酶-13并增加细胞间黏附分子-1水平。细胞间黏附分子-1在真核细胞中的异位表达表明，细胞间黏附分子-1的细胞外结构域含有基质金属蛋白酶-13识别的底物。在活性基质金属蛋白酶-13存在的情况下，对表达异位细胞间黏附分子-1的COS-7细胞进行孵育，可诱导RAW 264.7细胞对COS-7单层细胞的黏附受到抑制。将基质辅助激光解吸电离飞行时间质谱分析与液相色谱-离子阱质谱联用，对与基质金属蛋白酶-13孵育的细胞间黏附分子-1进行分析，使我们能够确定基质金属蛋白酶-13在细胞间黏附分子-1的E61和G98位点的切割位点。G98是PDGQS部分的一部分，该部分与位于II型胶原I-α的基质金属蛋白酶-13切割位点的共有PDGLS底物具有同源性。
综上所述，这些结果表明基质金属蛋白酶-13是一氧化氮介导的动脉粥样硬化保护机制。
