Ljubica Perisic1, Erika Hedin, Anton Razuvaev, Mariette Lengquist, Cecilia Osterholm, Lasse Folkersen, Peter Gillgren, Gabrielle Paulsson-Berne, Fredrik Ponten, Jacob Odeberg, Ulf Hedin. 1. From the Department of Molecular Medicine and Surgery (L.P., E.H., A.R., M.L., C.O., U.H.), and Department of Medicine (G.P.-B., J.O.), Karolinska Institute, Stockholm, Sweden; Department of Molecular Genetics, Novo Nordisk, Copenhagen, Denmark (L.F.); Department of Surgery, Södersjukhuset, Stockholm, Sweden (P.G.); Department of Immunology, Genetics and Pathology, Uppsala University, Uppsala, Sweden (F.P.); and Department of Proteomics, Royal Institute of Technology, Stockholm, Sweden (J.O.).
Abstract
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.
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.
Authors: Vivek Nanda; Kelly P Downing; Jianqin Ye; Sophia Xiao; Yoko Kojima; Joshua M Spin; Daniel DiRenzo; Kevin T Nead; Andrew J Connolly; Sonny Dandona; Ljubica Perisic; Ulf Hedin; Lars Maegdefessel; Jessie Dalman; Liang Guo; XiaoQing Zhao; Frank D Kolodgie; Renu Virmani; Harry R Davis; Nicholas J Leeper Journal: Circ Res Date: 2015-11-23 Impact factor: 17.367
Authors: Chul-Hee Lee; Mi Jeong Kim; Hwan Hee Lee; Jin Chul Paeng; Young Joo Park; So Won Oh; Young Jun Chai; Young A Kim; Gi Jeong Cheon; Keon Wook Kang; Hyewon Youn; June-Key Chung Journal: Mol Imaging Biol Date: 2019-08 Impact factor: 3.488