Proteomic Analysis of Intraluminal Thrombus Highlights Complement Activation in Human Abdominal Aortic Aneurysms

Abstract

Objective-To identify proteins related to intraluminal thrombus biological activities that could help to find novel pathological mechanisms and therapeutic targets for human abdominal aortic aneurysm (AAA). Approach and Results-Tissue-conditioned media from patients with AAA were analyzed by a mass spectrometry-based strategy using liquid chromatography coupled to tandem mass spectrometry. Global pathway analysis by Ingenuity software highlighted the presence of several circulating proteins, among them were proteins from the complement system. Complement C3 concentration and activation were assessed in plasma from AAA patients (small AAA, AAA diameter=3-5 cm and large AAA, AAA diameter >5 cm), showing decreased C3 levels and activation in large AAA patients. No association of a combination of single-nucleotide polymorphisms in complement genes between large and small AAA patients was observed. Intense extracellular C3 inmunostaining, along with C9, was observed in AAA thrombus. Analysis of C3 in AAA tissue homogenates and tissue-conditioned media showed increased levels of C3 in AAA thrombus, as well as proteolytic fragments (C3a/C3c/C3dg), suggesting its local deposition and activation. Finally, the functional role of local complement activation in polymorphonuclear (PMN) cell activation was tested, showing that C3 blockade by anti-C3 antibody was able to decrease thrombus-induced neutrophil chemotaxis and reactive oxygen species production. Conclusions-

Using Proteomics to Understand Abdominal Aortic Aneurysms: Where Are We?

“…To further explore the pathophysiology of ILT in human AAA, by increasing the number of identified proteins, Martinez-Pinna et al [26] analyzed ILT and wall conditioned media using liquid chromatography and tandem mass spectrometry. Global pathway analysis of identified peptides/proteins by Ingenuity software highlighted that complement system components were highly enriched in AAA tissue-conditioned media.…”

“…Global pathway analysis of identified peptides/proteins by Ingenuity software highlighted that complement system components were highly enriched in AAA tissue-conditioned media. In particular, they highlighted a decrease of systemic C3 concentration and activity in the later stages of AAA associated with local complement retention, consumption, and proteolysis in the thrombus that could induce polymorphonuclear (PMN) cell chemotaxis and activation, playing a detrimental role in AAA progression [26].…”

“…In the pie chart represented in Figure 2, it is shown the percentage of proteins identified by each author. Concerning the role of each paper in the identification of these secreted proteins, it is noteworthy that the study of Martinez-Pinna et al [26], provided about 83% of these identification. The high identification rate of this study [26] is probably due to the use of the two-dimensional liquid * UniProtKB/Swiss-Prot entry name (http://www.uniprot.org/) …”

“…The aim was to identify the key proteins involved in the pathogenesis of AAAs [28]. In this study we decided not to remove highly abundant proteins such as albumin and IgG since it would cause the removal of a broad range of other low mass Plasma protease C1 inhibitor IC1_HUMAN [25,26] and low abundant physiologically important regulatory and/or transient proteins. We confirmed a number of biomarkers associated with AAA that have been previously identified by various authors reported in this review [28].…”

Contribution of proteomics to the management of vascular disorders

“…Through this approach, proteins released into the blood from the vascular tissue can be discovered, constituting a potentially reliable source of disease biomarkers. In this sense, a characteristic secretome of atherosclerotic arteries [48], aortic valves with stenosis [49] and aortic intraluminal thrombi [50] has been reported, that is potentially relevant in the diagnosis of these pathologies and/or in the definition of novel therapeutic targets.…”