Human monocyte-derived macrophages induce collagen breakdown in fibrous caps of atherosclerotic plaques. Potential role of matrix-degrading metalloproteinases and implications for plaque rupture.
Human monocyte-derived macrophages induce collagen breakdown in fibrous caps of atherosclerotic plaques. Potential role of matrix-degrading metalloproteinases and implications for plaque rupture.
复制标题
DOI：
--
复制
发表时间：
1995-09
期刊：
影响因子：
37.8
通讯作者：
P. Shah;E. Falk;J. Badimón;A. Fernández-Ortiz;A. Mailhac;G. Villareal-Levy;J. Fallon;J. Regnstrom;V. Fuster
中科院分区：
文献类型：
--
作者：
P. Shah;E. Falk;J. Badimón;A. Fernández-Ortiz;A. Mailhac;G. Villareal-Levy;J. Fallon;J. Regnstrom;V. Fuster
背景动脉粥样硬化斑块纤维帽破裂是导致冠状动脉血栓形成的关键事件，导致急性冠状动脉综合征。最近的研究表明，脆弱和破裂的动脉粥样硬化斑块的纤维帽具有与增加的巨噬细胞密度和减少的平滑肌细胞密度相关的胶原和糖胺聚糖含量减少。由于纤维帽中的胶原蛋白分解可能导致帽变薄和弱化，增加其破裂的脆弱性，我们测试了单核细胞衍生的巨噬细胞通过产生基质降解金属蛋白酶（MMPs）可以诱导人动脉粥样硬化纤维帽中胶原蛋白分解的假设。方法和结果通过Ficoll-Paque密度梯度从人血液中分离单核细胞，并使其在细胞培养物中生长，直至表型和染色特征表明转化为巨噬细胞（4至7天）。从人主动脉或颈动脉斑块上切下纤维帽，并在无血清培养基中与巨噬细胞一起孵育48小时，其中无MMP抑制剂（n = 21）和MMP抑制剂（n = 10）或仅无细胞和无血清培养基（n = 9）。通过分光光度法测量培养基中释放的羟脯氨酸，并用作纤维帽中胶原蛋白分解的证据。用特异性单克隆抗体免疫细胞化学法检测细胞培养液中MMP-1（间质胶原酶）和MMP-2（72-kD明胶酶）的表达，酶谱法检测培养上清中MMP活性。当纤维帽与巨噬细胞一起孵育时，释放的羟脯氨酸的量显著大于与无细胞培养基一起孵育时（0.4 +/- 0.16 μ g.mL-1.mg-1对0.02 +/- 0.03 μ g.mL-1.mg-1组织;通过Mann-Whitney检验，P <0.04）。当纤维帽与巨噬细胞在MMP抑制剂存在下孵育时，没有羟脯氨酸释放。免疫细胞化学显示MMP-1和MMP-2表达的巨噬细胞之间的第4和第7天，和酶谱证实存在的MMP-2活性的上清液中。结论：在这项研究中，人单核细胞源性巨噬细胞诱导人动脉粥样硬化斑块纤维帽中的胶原分解，与MMP活性的细胞表达和酶谱证据相关;在MMP抑制剂存在下，未发现胶原分解的证据。这些发现支持了这样的假设，即动脉粥样硬化斑块中增加的巨噬细胞密度和/或活化可能通过分泌MMP和可能的其他蛋白酶诱导纤维帽中的胶原分解，从而导致斑块破裂的脆弱性。
BACKGROUND Rupture of the fibrous cap of the atherosclerotic plaque is a key event that predisposes to coronary thrombosis, leading to acute coronary syndromes. Recent studies have shown that the fibrous caps of vulnerable and ruptured atherosclerotic plaques have reduced collagen and glycosaminoglycan content in association with an increased macrophage density and a reduced smooth muscle cell density. Since collagen breakdown in the fibrous caps may contribute to a thinning and weakening of the cap, increasing its vulnerability to rupture, we tested the hypothesis that monocyte-derived macrophages, by producing matrix-degrading metalloproteinases (MMPs), could induce collagen breakdown in human atherosclerotic fibrous caps. METHODS AND RESULTS Monocytes were isolated from human blood by Ficoll-Paque density gradient and allowed to grow in cell culture until phenotypic and staining characteristics indicated transformation into macrophages (4 to 7 days). Fibrous caps were dissected from human aortic or carotid plaques and incubated for 48 hours with macrophages in serum-free medium without (n = 21) and with (n = 10) an MMP inhibitor or with cell- and serum-free medium only (n = 9). Hydroxyproline released in the culture medium was measured by a spectrophotometric method and used as evidence of collagen breakdown in the fibrous caps. Immunocytochemistry with specific monoclonal antibodies was used to identify expression of MMP-1 (interstitial collagenase) and MMP-2 (72-kD gelatinase) in cell culture, and zymography was used to detect MMP activity in the culture supernatant. The amount of hydroxyproline released was significantly greater when fibrous caps were incubated with macrophages than when incubated with cell-free medium (0.4 +/- 0.16 micrograms.mL-1.mg-1 versus 0.02 +/- 0.03 micrograms.mL-1.mg-1 of tissue; P < .04 by Mann-Whitney test). There was no hydroxyproline release when fibrous caps were incubated with macrophages in the presence of an MMP inhibitor. Immunocytochemistry demonstrated MMP-1 and MMP-2 expression by macrophages between days 4 and 7, and zymography confirmed the presence of MMP-2 activity in the supernatant. CONCLUSIONS In this study, human monocyte-derived macrophages were shown to induce collagen breakdown in fibrous caps of human atherosclerotic plaques associated with cellular expression and zymographic evidence of MMP activity; no evidence of collagen breakdown was found in the presence of an MMP inhibitor. These findings support the hypothesis that increased macrophage density and/or activation in the atherosclerotic plaque may induce collagen breakdown in the fibrous cap by secreting MMPs and possibly other proteases, thus contributing to vulnerability to plaque rupture.