Fontaine Vincent, Jacob Marie-Paule, Houard Xavier, Rossignol Patrick, Plissonnier Didier, Angles-Cano Eduardo, Michel Jean-Baptiste
Institut National de la Santé et de la Recherche Médicale U460, Cardiovascular Remodeling, Hôpital Xavier Bichat, Paris, France.
Am J Pathol. 2002 Nov;161(5):1701-10. doi: 10.1016/S0002-9440(10)64447-1.
Acquired abdominal aortic aneurysms are usually associated with a mural thrombus through which blood continues to flow. Some early data suggest that aneurysmal evolution correlates with the biological activity of the thrombus. Our hypothesis was therefore that the thrombus could adsorb blood components and store, release, and participate in the activation of proteases involved in aneurysmal evolution. For this purpose, we have explored both the metalloproteinase and fibrinolytic systems in the thrombus and the wall of human aneurysms. We have first investigated blood clot formation and lysis in vitro. Spontaneous clotting induces a release of promatrix metalloproteinase (pro-MMP)-9 into the serum that was fourfold higher than in paired control plasma (P < 0.001). Fibrinolysis progressively released more MMP-9 in a time-dependent manner (P < 0.01). After selective isolation, we demonstrated that polymorphonuclear leukocytes are the main source of MMP-9 release during clot formation. Protease content was then analyzed in 35 mural thrombi and walls of human abdominal aortic aneurysms sampled during surgical repair. In 15 aneurysms, the liquid phase at the interface between the thrombus and the wall was sampled separately. Both thrombus and wall contained MMP-2 and MMP-9 but the ratio MMP-9/MMP-2 was higher in the thrombus than in the wall. The liquid interface also contained active MMP-9. Immunohistochemistry of the thrombus confirmed these findings, showing the presence of polymorphonuclear leukocytes at the luminal pole of the thrombus, co-localizing with MMP-9 storage. In contrast, MMP-3 and MMP-7 were only present in the aneurysmal wall. Plasminogen was present in the mural thrombus but plasmin activity was present in both thrombus and wall. In the liquid interface, plasmin-alpha(2)-anti-plasmin complexes were detected demonstrating in vivo the activation of plasminogen. In contrast, u-PA and t-PA were detectable only in the wall, suggesting that plasminogen present in the thrombus could be activated by factors secreted by the arterial wall. This was demonstrated in vitro, in which co-incubation of thrombus and wall extracts generated plasmin in the presence of a fibrin matrix and activated MMPs. In conclusion, our study strongly suggests that the mural thrombus, by trapping polymorphonuclear leukocytes and adsorbing plasma components could act as a source of proteases in aneurysms that may play a critical role in enlargement and rupture.
获得性腹主动脉瘤通常伴有附壁血栓，血液仍可通过血栓流动。一些早期数据表明，动脉瘤的演变与血栓的生物学活性相关。因此，我们的假设是，血栓可以吸附血液成分，并储存、释放和参与动脉瘤演变过程中蛋白酶的激活。为此，我们研究了人类动脉瘤血栓和瘤壁中的金属蛋白酶和纤溶系统。我们首先在体外研究了血凝块的形成和溶解。自发凝血会使血清中前基质金属蛋白酶（pro-MMP）-9的释放量比配对的对照血浆高出四倍（P < 0.001）。纤维蛋白溶解以时间依赖性方式逐渐释放更多的MMP-9（P < 0.01）。经过选择性分离，我们证明多形核白细胞是血凝块形成过程中MMP-9释放的主要来源。然后分析了手术修复过程中采集的35例人类腹主动脉瘤壁血栓和瘤壁中的蛋白酶含量。在15个动脉瘤中，分别采集了血栓与瘤壁界面处的液相样本。血栓和瘤壁均含有MMP-2和MMP-9，但血栓中MMP-9/MMP-2的比值高于瘤壁。液体界面也含有活性MMP-9。血栓的免疫组织化学证实了这些发现，显示血栓腔面极有多形核白细胞存在，与MMP-9储存共定位。相比之下，MMP-3和MMP-7仅存在于动脉瘤壁中。纤溶酶原存在于壁血栓中，但纤溶酶活性在血栓和瘤壁中均有。在液体界面检测到纤溶酶-α(2)-抗纤溶酶复合物，证明体内纤溶酶原被激活。相比之下，u-PA和t-PA仅在瘤壁中可检测到，这表明血栓中存在的纤溶酶原可能被动脉壁分泌的因子激活。这在体外得到了证实，在纤维蛋白基质存在的情况下，血栓和瘤壁提取物共同孵育可产生纤溶酶并激活MMPs。总之，我们的研究强烈表明，壁血栓通过捕获多形核白细胞和吸附血浆成分，可能成为动脉瘤中蛋白酶的来源，这可能在动脉瘤扩大和破裂中起关键作用。
