School of Biological Sciences, Nanyang Technological University, 60 Nanyang Drive, Singapore, 637551.
Diabetes Center, Khoo Teck Puat Hospital, Singapore; Saw Swee Hock School of Public Health, National University of Singapore, Singapore.
Atherosclerosis. 2021 May;324:58-68. doi: 10.1016/j.atherosclerosis.2021.03.020. Epub 2021 Mar 25.
Aging 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.
IsoDGR-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.
IsoDGR-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 TNFα to drive additional recruitment of circulating leukocytes. IsoDGR-modified fibronectin also induced endothelial cell expression of integrin β1 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.
Age-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.
衰老是心血管疾病（CVD）的主要危险因素，但与年龄相关的动脉粥样硬化相关的机制仍不清楚。我们之前观察到，长寿的血管基质蛋白可以获得“获得功能”的 isoDGR 基序，这些基序可能在动脉粥样硬化病理中发挥作用。
生成了 isoDGR 特异性 mAb，并用于基于 ELISA 的测定，以测量来自患有冠状动脉疾病（CAD）和非 CAD 对照的患者的血浆样本中的基序水平。通过测定激活单核细胞、巨噬细胞和内皮细胞的能力（信号活性、促炎细胞因子表达和募集/黏附潜能），来确定在年龄损伤的纤维连接蛋白中积累 isoDGR 的功能后果。使用缺乏 isoDGR 修复酶 PCMT1 的小鼠来评估体内基序分布和巨噬细胞定位。
患者血浆中的 isoDGR 修饰的纤维连接蛋白和纤维蛋白原水平在 CAD 中显著增强，并进一步与吸烟状况相关。功能测定表明，isoDGR 修饰的纤维连接蛋白通过整合素受体“外到内”信号激活单核细胞和巨噬细胞，触发 ERK:AP-1 级联反应和促炎细胞因子 MCP-1 和 TNFα 的表达，以驱动循环白细胞的额外募集。isoDGR 修饰的纤维连接蛋白还诱导内皮细胞表达整合素 β1，以进一步增强细胞黏附和基质沉积。对小鼠主动脉组织的分析证实了在体内与 CD68 巨噬细胞共定位的 isoDGR 修饰蛋白的积累。
年龄损伤的纤维连接蛋白具有 isoDGR 基序，这些基序增加了与单核细胞、巨噬细胞和内皮细胞表面整合素的结合。随后的“外到内”信号激活引发了一系列强效的细胞因子和趋化因子，这些因子驱动了更多的白细胞募集到正在发育的动脉粥样硬化基质中。
