Collagens constitute a major portion of the extracellular matrix in the atherosclerotic plaque, where they contribute to the strength and integrity of the fibrous cap, and also modulate cellular responses via specific receptors and signaling pathways. This review focuses on the diverse roles that collagens play in atherosclerosis; regulating the infiltration and differentiation of smooth muscle cells and macrophages; controlling matrix remodeling through feedback signaling to proteinases; and influencing the development of atherosclerotic complications such as plaque rupture, aneurysm formation and calcification. Expanding our understanding of the pathways involved in cell-matrix interactions will provide new therapeutic targets and strategies for the diagnosis and treatment of atherosclerosis.

Abstract-Collagens are abundant within the atherosclerotic plaque, where they contribute to lesion volume and mechanical stability and influence cell signaling. The discoidin domain receptor 1 (DDR1), a receptor tyrosine kinase that binds to collagen, is expressed in blood vessels, but evidence for a functional role during atherogenesis is incomplete. In the present study, we generated Ddr1 ϩ/ϩ ;Ldlr Ϫ/Ϫ and Ddr1 Ϫ/Ϫ ;Ldlr Ϫ/Ϫ mice and fed them an atherogenic diet for 12 or 24 weeks. Targeted deletion of Ddr1 resulted in a 50% to 60% reduction in atherosclerotic lesion area in the descending aorta at both 12 and 24 weeks. Ddr1 Ϫ/Ϫ ;Ldlr Ϫ/Ϫ plaques exhibited accelerated deposition of fibrillar collagen and elastin at 12 weeks compared with Ddr1 ϩ/ϩ ;Ldlr Ϫ/Ϫ plaques. Expression analysis of laser microdissected lesions in vivo, and of Ddr1 Ϫ/Ϫ smooth muscle cells in vitro, revealed increased mRNA levels for procollagen ␣1(I) and ␣1(III) and tropoelastin, suggesting an enhancement of matrix synthesis in the absence of DDR1. Furthermore, whereas plaque smooth muscle cell content was unchanged, Ddr1 Ϫ/Ϫ ;Ldlr Ϫ/Ϫ plaques had a 49% decrease in macrophage content at 12 weeks, with a concomitant reduction of in situ gelatinolytic activity. Moreover, mRNA expression of both monocyte chemoattractant protein-1 and vascular cell adhesion molecule-1 was reduced in vivo, and Ddr1 Ϫ/Ϫ ;Ldlr Ϫ/Ϫ macrophages demonstrated impaired matrix metalloproteinase expression in vitro. These data suggest novel roles for DDR1 in macrophage recruitment and invasion during atherogenesis. In conclusion, our data support a role for DDR1 in the regulation of both inflammation and fibrosis early in plaque development. Deletion of DDR1 attenuated atherogenesis and resulted in the formation of matrix-rich plaques. Key Words: atherosclerosis Ⅲ discoidin domain receptor 1 Ⅲ collagen Ⅲ inflammation Ⅲ macrophage A therosclerosis is a fibroinflammatory disease of the arterial wall. The atherosclerotic plaque is home to multiple cell types, including endothelial cells, smooth muscle cells (SMCs), and bone marrow-derived monocyte/ macrophages, all interacting within a chronically inflamed, lipid-rich, and highly dynamic extracellular matrix microenvironment. Collagens are critical components of the extracellular matrix present within atherosclerotic plaques, where they contribute to lesion volume and can constitute up to 60% of total plaque protein. 1 Collagens also provide mechanical stability to the fibrous cap and protect against plaque rupture, a major cause of the clinical complications associated with atherosclerosis. 2 Furthermore, collagens stimulate diverse cellular responses that are central to plaque development. For example, collagen synthesis and degradation are important for smooth muscle cell migration, 3,4 and degraded type I collagen fragments stimulate the disassembly of focal adhesion complexes in SMCs. 5 By contrast, intact type I collagen inhibits SMC proliferation. 6 Additionally, type I collagen promotes monocyte differentiation ...

Key Words: atherosclerosis Ⅲ discoidin domain receptor Ⅲ extracellular matrix Ⅲ collagen Ⅲ smooth muscle cell T he accumulation of extracellular matrix regulates both growth and stability of the atherosclerotic plaque. Vascular smooth muscle cells (SMCs) undergo a phenotypic switch from contractile to synthetic as they proliferate and migrate into the lesion, elaborating a collagen-rich matrix. 1 Collagen accumulation has a multifaceted role in the etiology of atherosclerosis, whereas excess collagen can contribute to the expansion of lesion volume and vascular stenosis, maintenance of a thick collagen-rich fibrous cap is important for the prevention of plaque rupture. Inflammatory processes influence the turnover of collagens in the plaque. Plaque macrophages are a potent source of matrix metalloproteinases (MMPs), which destabilize the lesion by digesting collagen at the rupture prone plaque shoulders. 2 Thus, the balance between synthesis, remodeling, and degradation of collagens determines the content and organization of the atherosclerotic plaque matrix, influencing disease progression and clinical outcomes.The discoidin domain receptor (DDR)1 is a collagen receptor tyrosine kinase expressed on both SMCs and macrophages 3-7 that initiates signaling when bound by triple helical collagens. 8,9 Several studies have revealed that DDR1 plays important roles in controlling cell proliferation, migration, and matrix remodeling. 10 We have previously determined functional roles for DDR1 in atherogenesis using the hypercholesterolemic Ldlr Ϫ/Ϫ mouse model. Atherosclerotic plaques that formed in mice doubly deficient in DDR1 and LDLR (Ddr1 Ϫ/Ϫ ;Ldlr Ϫ/Ϫ ) were smaller in size and exhibited accelerated matrix deposition, decreased in situ MMP activity, and decreased macrophage content compared to mice deficient in LDLR only (Ddr1 ϩ/ϩ ;Ldlr Ϫ/Ϫ ). 4 DDR1 also mediates atherosclerotic plaque calcification by SMCs, a long-term complication of atherosclerosis. 11 We performed studies using bone marrow transplantation to delete DDR1 exclusively in bone marrow-derived cells of Ldlr Ϫ/Ϫ mice. 12 This re-

Intimal calcification is a feature of advanced atherosclerotic disease that predicts a two-to eightfold increase in the risk of coronary events. Type I collagen promotes vascular smooth muscle cell-mediated calcification, although the mechanism by which this occurs is unknown. The discoidin domain receptor 1 (DDR1) is a collagen receptor that is emerging as a critical mediator of atherosclerosis. To determine whether DDR1 is involved in intimal calcification, we fed male Ddr1 ؊/؊ ;Ldlr ؊/؊ and Ddr1 ؉/؉ ;Ldlr ؊/؊ mice an atherogenic diet for 6, 12, or 24 weeks. DDR1 deficiency significantly reduced the calcium content of the aortic arch, and microcomputed tomography demonstrated a significant decrease in hydroxyapatite deposition after 24 weeks of atherogenic diet. Reduced calcification was correlated with decreases in macrophage accumulation and tumor necrosis factor ␣ staining, suggesting that the reduction in calcification was in part due to decreased inflammation. The chondrogenic markers type II collagen, type X collagen , and Sox-9 were expressed within the mineralized foci. An in vitro assay performed with vascular smooth muscle cells revealed that DDR1 was required for cell-mediated calcification of the matrix , and Ddr1 ؉/؉ smooth muscle cells expressed more alkaline phosphatase activity, whereas Ddr1 ؊/؊ smooth muscle cells expressed elevated levels of mRNA for nucleotide pyrophosphatase phosphodiesterase 1, an inhibitor of tissue mineralization. Taken together, our results demonstrate that DDR1 mediates an important mechanism for atherosclerotic calcification. (Am J Pathol

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Intimal calcification is a serious complication of advanced atherosclerotic disease. Low density lipoprotein receptor‐deficient mice (Ldlr−/−) fed a high fat diet develop complicated atherosclerotic lesions. Here we report that Ldlr−/−mice develop foci of intimal calcification in the aortic arch, which are positive for Alizarin Red S (AR) and Von Kossa (VK), two histological stains for mineralized tissues. Cells within these foci are surrounded by a type II collagen‐rich matrix and express a chondrocyte‐specific transcription factor, Sox‐9. Atherosclerotic lesions are rich in collagens, which have been shown to promote smooth muscle cell‐mediated calcification. Thus, we hypothesized a role for the discoidin domain receptor 1 (DDR1), a collagen receptor, in intimal calcification. Male mice with a combined deficiency in LDLR and DDR1 (Ldlr−/−;Ddr1−/−) and controls (Ldlr−/−;Ddr1+/+) were fed a high fat diet for 12 weeks. DDR1‐deficiency attenuated the incidence of intimal calcification, measured by positive staining for AR and VK. Aortic arch calcium extraction confirmed a reduction in the extent of calcification in the Ldlr−/−;Ddr1−/− mice compared with control mice (45 ± 22 vs 123 ± 52 μmol/g dry weight). This study demonstrates regions of calcification within the intimal lesions of Ldlr−/− mice fed an atherogenic diet and provides novel evidence of a role for DDR1 in this process.