Aging-induced isoDGR-modified fibronectin activates monocytic and endothelial cells to promote atherosclerosis

Background and AimsAging is the primary risk factor for cardiovascular disease (CVD), but the mechanisms underlying age-linked atherosclerosis remain unclear. We previously observed that long-lived vascular matrix proteins can acquire gain-of-function isoDGR motifs that might play a role in atherosclerotic pathology. MethodsIsoDGR-specific mAb were generated and used for ELISA-based measurement of motif levels in plasma samples from patients with coronary artery diseases (CAD) and non-CAD controls. Functional consequences of isoDGR accumulation in age-damaged fibronectin were determined by bioassay for capacity to activate monocytes, macrophages, and endothelial cells (signalling activity, pro-inflammatory cytokine expression, and recruitment/adhesion potential). Mice deficient in the isoDGR repair enzyme PCMT1 were used to assess motif distribution and macrophage localisation in vivo. ResultsIsoDGR-modified fibronectin and fibrinogen levels in patient plasma were significantly enhanced in CAD and further associated with smoking status. Functional assays demonstrated that isoDGR-modified fibronectin activated both monocytes and macrophages via integrin receptor outside in signalling, triggering an ERK:AP-1 cascade and expression of pro-inflammatory cytokines MCP-1 and TNF to drive additional recruitment of circulating leukocytes. IsoDGR-modified fibronectin also induced endothelial cell expression of integrin {beta}1 to further enhance cellular adhesion and matrix deposition. Analysis of murine aortic tissues confirmed accumulation of isoDGR-modified proteins co-localised with CD68+ macrophages in vivo. ConclusionsAge-damaged fibronectin features isoDGR motifs that increase binding to integrins on the surface of monocytes, macrophages, and endothelial cells. Subsequent activation of outside-in signalling elicits a range of potent cytokines and chemokines that drive additional leukocyte recruitment to the developing atherosclerotic matrix. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=95 SRC="FIGDIR/small/21252419v1_ufig1.gif" ALT="Figure 1"> View larger version (20K): org.highwire.dtl.DTLVardef@190a620org.highwire.dtl.DTLVardef@18a9431org.highwire.dtl.DTLVardef@10749ddorg.highwire.dtl.DTLVardef@18272a6_HPS_FORMAT_FIGEXP M_FIG C_FIG HighlightsO_LIIsoDGR-modified plasma proteins are associated with CAD. C_LIO_LIIsoDGR binding to integrin receptors on monocytes, macrophage, and endothelial cells promotes outside-in signalling, monocyte infiltration, and endothelial binding. C_LIO_LIIsoDGR-modified fibronectin may initiate vascular inflammation in atherosclerotic CVD. C_LI