O-173 - DESTABILISING PROTEINS IN SYMPTOMATIC CAROTID PLAQUES FOR FUTURE RISK MARKERS, A PROTEOMIC STUDY

TOPIC:
Vascular Biology
AUTHORS:
Yeung K. (Rigshospitalet, Department of Vascular Surgery ~ Copenhagen ~ Denmark) , Sillesen H.H. (Rigshospitalet, Department of Vascular Surgery ~ Copenhagen ~ Denmark) , Eldrup N. (Rigshospitalet, Department of Vascular Surgery ~ Copenhagen ~ Denmark) , Eiberg J.P. (Rigshospitalet, Department of Vascular Surgery ~ Copenhagen ~ Denmark) , Davies M.J. (University of Copenhagen, Department of Biomedical Sciences ~ Copenhagen ~ Denmark) , Lorentzen L.G. (University of Copenhagen, Department of Biomedical Sciences ~ Copenhagen ~ Denmark)
Introduction:
Atherosclerotic plaque formation is caused of lipid accumulation in the arterial wall which is asymptomatic for decades. Destabilisation and rupture of plaques occur suddenly and give rise to acute thromboembolic event as myocardial infarction and stroke. Stabile plaques are hard lesions with fibrosis and calcifications while unstable plaques are softer lesions, vulnerable to rupture due to higher inflammatory activity, a thin fibrous cap, a large necrotic lipid core, and often intraplaque haemorrhage (1-3). Proteolytic degradation of the extracellular matrix (ECM) is hypothesised to drive plaque destabilisation, yet the exact mechanisms are not fully understood (4). The presented study aims to uncover mechanisms of ECM remodelling associated with the destabilisation of symptomatic carotid plaques using state-of-the-art proteomics.
Methods:
Patients operated for symptomatic carotid artery disease, due to recent stroke, transient ischaemic attack or ocular ischaemic event (acute permanent or temporary vision loss) were included. Atherosclerotic carotid plaques were removed in toto. 21 plaques were selected and macroscopically categorised as hard (7 plaques), soft (7 plaques) or mixed (7 plaques) by the respective surgeon and first author (KY). Plaques were solubilised to extract intra- and extracellular proteins (5). The proteome coverage was explored with mass spectrometry (6, 7), and proteolytic degradation was investigated by conducting N-terminal proteomics (8, 9).
Results:
More than 5000 proteins were identified and quantified from the carotid plaques, including 211 ECM proteins. The coverage of both the intra- and extracellular proteome across 21 samples were covered with high reproducibility. We identified 544 proteins with differential abundances between hard and soft lesions and observed an enrichment of proteins involved in inflammatory responses and ECM remodelling (Figure 1). Upregulation of a range of proteolytic enzymes with concomitant loss of ECM proteins in soft lesions was found. This was also observed in mixed plaques, albeit to a lesser extent. These data agreed well with results from N-terminal peptide proteomics analysis, in which we identified 2,752 N-terminal peptides. 652 N-terminal peptides were more abundant in soft plaques, and 557 of these were consistent with proteolytic cleavage. Structural ECM proteins, such as collagens and fibronectin, were among the most cleaved proteins (Figure 2).
Conclusion:
The data presented offer a unique insight into the inflammatory and proteolytic pathways of atherosclerotic plaque destabilisation. We identified novel targets of proteolytic ECM degradation proteins associated with soft lesions suggesting plaque destabilisation. Thus, the presented data provide a framework for the identification of novel blood borne biomarkers to distinguish stable from unstable plaques.
References:
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