Department of Pathology, University of Washington School of Medicine, Seattle, WA 98104, USA.
J Immunol. 2013 Apr 15;190(8):4236-44. doi: 10.4049/jimmunol.1300046. Epub 2013 Mar 11.
Despite expanded definition of the leukocyte adhesion cascade and mechanisms underlying individual steps, very little is known about regulatory mechanisms controlling sequential shifts between steps. We tested the hypothesis that metalloproteinases provide a mechanism to rapidly transition monocytes between different steps. Our study identifies diapedesis as a step targeted by metalloproteinase activity. Time-lapse video microscopy shows that the presence of a metalloproteinase inhibitor results in a doubling of the time required for human monocytes to complete diapedesis on unactivated or inflamed human endothelium, under both static and physiological-flow conditions. Thus, diapedesis is promoted by metalloproteinase activity. In contrast, neither adhesion of monocytes nor their locomotion over the endothelium is altered by metalloproteinase inhibition. We further demonstrate that metalloproteinase inhibition significantly elevates monocyte cell surface levels of integrins CD11b/CD18 (Mac-1), specifically during transendothelial migration. Interestingly, such alterations are not detected for other endothelial- and monocyte-adhesion molecules that are presumed metalloproteinase substrates. Two major transmembrane metalloproteinases, a disintegrin and metalloproteinase (ADAM)17 and ADAM10, are identified as enzymes that control constitutive cleavage of Mac-1. We further establish that knockdown of monocyte ADAM17, but not endothelial ADAM10 or ADAM17 or monocyte ADAM10, reproduces the diapedesis delay observed with metalloproteinase inhibition. Therefore, we conclude that monocyte ADAM17 facilitates the completion of transendothelial migration by accelerating the rate of diapedesis. We propose that the progression of diapedesis may be regulated by spatial and temporal cleavage of Mac-1, which is triggered upon interaction with endothelium.
尽管白细胞黏附级联反应和各个步骤的机制得到了扩展定义，但对于控制步骤之间顺序转变的调节机制却知之甚少。我们检验了这样一个假设，即金属蛋白酶为单核细胞在不同步骤之间快速转换提供了一种机制。我们的研究确定出出芽是金属蛋白酶活性作用的一个靶点。延时视频显微镜显示，金属蛋白酶抑制剂的存在会导致人单核细胞在未激活或炎症状态的人内皮细胞上完成出芽所需的时间增加一倍，无论是在静态还是生理流动条件下。因此，出芽受到金属蛋白酶活性的促进。相比之下，单核细胞的黏附和在内皮上的迁移运动都不会因金属蛋白酶抑制而改变。我们进一步证明，金属蛋白酶抑制显著提高单核细胞表面整合素 CD11b/CD18（Mac-1）的水平，特别是在跨内皮迁移过程中。有趣的是，对于其他假定为金属蛋白酶底物的内皮和单核细胞黏附分子，不会检测到这种改变。两种主要的跨膜金属蛋白酶，解整合素金属蛋白酶（ADAM）17 和 ADAM10，被鉴定为控制 Mac-1 组成性裂解的酶。我们进一步证实，单核细胞 ADAM17 的敲低，但不是内皮细胞 ADAM10 或单核细胞 ADAM10 或 ADAM17，可重现金属蛋白酶抑制所观察到的出芽延迟。因此，我们得出结论，单核细胞 ADAM17 通过加速出芽速率来促进跨内皮迁移的完成。我们提出，出芽的进展可能受到 Mac-1 的空间和时间裂解的调节，这是与内皮相互作用触发的。
