Kindlin 3 (FERMT3) is associated with unstable atherosclerotic plaques, anti-inflammatory type II macrophages and upregulation of beta-2 integrins in all major arterial beds Niku Oksala a, b, c, * , Jenita P € arssinen d , Ilkka Sepp € al € a a , Norman Klopp h, i , Thomas Illig h, i , Reijo Laaksonen a, b , Mari Levula a , Emma Raitoharju a , Ivana Kholova e , Thanos Sioris f , Mika K € ah € onen b, c, g , Terho Lehtim € aki a, b, 1 , Vesa P. Hyt € onen d, 1 a Department of Clinical Chemistry, Fimlab Laboratories, Tampere University Hospital, Tampere, Finland b School of Medicine, University of Tampere, Finland c Division of Vascular Surgery, Department of Surgery, Tampere University Hospital, Finland d BioMediTech, University of Tampere, Tampere, Finland and Fimlab Laboratories, Tampere University Hospital, Tampere, Finland e Pathology, Fimlab Laboratories, Tampere University Hospital, Tampere, Finland f Heart Center, Tampere University Hospital, Tampere, Finland g Department of Clinical Physiology, Tampere University Hospital, Tampere, Finland h Research Unit of Molecular Epidemiology, Helmholtz Zentrum, German Research Center for Environmental Health, Munich, Germany i Hannover Unified Biobank, Hannover Medical School, Hannover, Germany article info Article history: Received 3 December 2014 Received in revised form 27 May 2015 Accepted 29 June 2015 Available online 13 July 2015 Keywords: Kindlin Integrin Immunohistochemistry Gene expression Atherosclerosis abstract Background: Kindlins (FERMT) are cytoplasmic proteins required for integrin (ITG) activation, leukocyte transmigration, platelet aggregation and thrombosis. Characterization of kindlins and their association with atherosclerotic plaques in human(s) is lacking. Methods and results: Exploratory microarray (MA) was first performed followed by selective quantitative validation of robustly expressed genes with qRT-PCR low-density array (LDA). In LDA, ITGA1 (1.30-fold, p ¼ 0.041) and ITGB3 (1.37-fold, p ¼ 0.036) were upregulated in whole blood samples of patients with coronary artery disease (CAD) compared to healthy controls. In arterial plaques, both robustly expressed transcript variants of FERMT3 (MA: 5.90- and 3.4-fold; LDA: 3.99-fold, p < 0.0001 for all) and ITGB2 (MA: 4.81- and 4.92-fold; LDA: 5.29-fold, p < 0.0001 for all) were upregulated while FERMT2 was down- regulated (MA: 1.61-fold; LDA: 2.88-fold, p < 0.0001 for both). The other integrins (ITGA1, ITGAV, ITGB3, ITGB5) were downregulated. All these results were replicated in at least one arterial bed. The latter FERMT3 transcript variant associated with unstable plaques (p ¼ 0.0004). FERMT3 correlated with M2 macrophage markers and in hierarchical cluster analysis clustered with inflammatory and macro- phage markers, while FERMT2 correlated with SMC-rich plaque markers and clustered with SMC markers. In confocal immunofluorescence analysis, FERMT3 protein colocalized with abundant CD68- positive cells of monocytic origin in the atherosclerotic plaques, while co-localization of FERMT3 with HHF35 indicative of smooth muscle cells was low. Conclusions: Kindlin-3 (FERMT3) is upregulated in atherosclerotic, especially unstable plaques, mainly in cells of monocytic origin and of M2 type. Simultaneous upregulation of ITGB2 suggests a synergistic effect on leukocyte adherence and transmigration into the vessel wall. © 2015 Elsevier Ireland Ltd. All rights reserved. 1. Introduction Endothelial dysfunction, infiltration of the arterial wall by macrophages (MF) and platelets, development of chronic inflam- mation, formation of foam cells, activation of smooth muscle cells (SMC), plaque neovascularization, hemorrhages and thrombosis are * Corresponding author. Tampere University Hospital, Fimlab Laboratories and Department of Surgery and Clinical Chemistry, Finn-Medi 2, 3rd, Floor, P.O. Box 2000, FI-33521 Tampere, Finland. E-mail address: nikuoksala@gmail.com (N. Oksala). 1 Equally contributed. Contents lists available at ScienceDirect Atherosclerosis journal homepage: www.elsevier.com/locate/atherosclerosis http://dx.doi.org/10.1016/j.atherosclerosis.2015.06.058 0021-9150/© 2015 Elsevier Ireland Ltd. All rights reserved. Atherosclerosis 242 (2015) 145e154