Profiling of atherosclerotic lesions by gene and tissue microarrays reveals PCSK6 as a novel protease in unstable carotid atherosclerosis

Arterioscler Thromb Vasc Biol. 2013 Oct;33(10):2432-43. doi: 10.1161/ATVBAHA.113.301743. Epub 2013 Aug 1.


Objective: Carotid plaque instability is a major cause of ischemic stroke, but detailed knowledge about underlying molecular pathways is still lacking. Here, we evaluated large-scale transcriptomic and protein expression profiling in a biobank of carotid endarterectomies followed by characterization of identified candidates, as a platform for discovery of novel proteins differentially regulated in unstable carotid lesions.

Approach and results: Genes highly upregulated in symptomatic versus asymptomatic plaques were selected from Affymetrix microarray analyses (n=127 plaques), and tissue microarrays constructed from 34 lesions were assayed for 21 corresponding proteins by immunohistochemistry. Quantification of stainings demonstrated differential expression of CD36, CD137, and DOCK7 (P<0.05) in unstable versus stable lesions and the most significant upregulation of a proprotein convertase, PCSK6 (P<0.0001). Increased expression of PCSK6 in symptomatic lesions was verified by quantitative real-time polymerase chain reaction (n=233), and the protein was localized to smooth muscle α-actin positive cells and extracellular matrix of the fibrous cap by immunohistochemistry. PCSK6 expression positively correlated to genes associated with inflammation, matrix degradation, and mitogens in microarrays. Stimulation of human carotid smooth muscle cells in vitro with cytokines caused rapid induction of PCSK6 mRNA.

Conclusions: Using a combination of transcriptomic and tissue microarray profiling, we demonstrate a novel approach to identify proteins differentially expressed in unstable carotid atherosclerosis. The proprotein convertase PCSK6 was detected at increased levels in the fibrous cap of symptomatic carotid plaques, possibly associated with key processes in plaque rupture such as inflammation and extracellular matrix remodeling. Further studies are needed to clarify the role of PCSK6 in atherosclerosis.

Keywords: atherosclerosis; carotid stenosis; muscle, smooth; symptomatic carotid stenosis.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Asymptomatic Diseases
  • Carotid Stenosis / enzymology*
  • Carotid Stenosis / genetics*
  • Carotid Stenosis / immunology
  • Carotid Stenosis / pathology
  • Cells, Cultured
  • Cytokines / metabolism
  • Extracellular Matrix / metabolism
  • Fibrosis
  • Gene Expression Profiling / methods*
  • Humans
  • Immunohistochemistry
  • Inflammation Mediators / metabolism
  • Muscle, Smooth, Vascular / enzymology
  • Muscle, Smooth, Vascular / immunology
  • Myocytes, Smooth Muscle / enzymology
  • Myocytes, Smooth Muscle / immunology
  • Oligonucleotide Array Sequence Analysis*
  • Plaque, Atherosclerotic
  • Proprotein Convertases / genetics*
  • Proprotein Convertases / metabolism*
  • RNA, Messenger / metabolism
  • Real-Time Polymerase Chain Reaction
  • Reverse Transcriptase Polymerase Chain Reaction
  • Rupture, Spontaneous
  • Serine Endopeptidases / genetics*
  • Serine Endopeptidases / metabolism*
  • Tissue Array Analysis*
  • Up-Regulation


  • Cytokines
  • Inflammation Mediators
  • RNA, Messenger
  • PCSK6 protein, human
  • Proprotein Convertases
  • Serine Endopeptidases