Ionescu Carmen V, Cepinskas Gediminas, Savickiene Jurate, Sandig Martin, Kvietys Peter R
Vascular Biology/Inflammation Program, Lawson Health Research Institute, London, ON, Canada.
Microcirculation. 2003 Apr;10(2):205-20. doi: 10.1038/sj.mn.7800185.
In vitro studies have indicated that polymorphonuclear leukocytes (PMNs) traverse endothelial cell monolayers via the paracellular pathway (i.e., through endothelial cell-cell junctions. Herein, we assessed whether the adherens junctions (AJs) are disrupted during PMN transendothelial cell migration.
Human umbilical vein endothelial cells (HUVECs) were grown to confluence on porous membranes and activated with interleukin-1beta, and PMN transendothelial migration was facilitated by formyl-methionyl-leucyl-phenylalanine. Using dual immunofluorescence staining and laser scanning confocal microscopy, we assessed the effects of PMN-endothelial cell adhesive interactions (i.e., adhesion to and emigration across monolayers) on the AJ components vascular endothelial (VE)-cadherin, beta-catenin, alpha-catenin, and gamma-catenin.
In the AJ immediately adjacent to the adherent PMN, there was a loss of staining for some of the AJ components. AJ components further away from HUVEC-PMN adhesive interactions were unaffected. An iterative approach indicated that the four components were sequentially lost from the AJ. beta-catenin was lost first, followed by VE-cadherin, alpha-catenin, and, finally, gamma-catenin. In the absence of PMNs, the cross-linking of VE-cadherin, but not platelet endothelial cell adhesion molecule-1 or intercellular adhesion molecule-1, increased the cytoplasmic accumulation of beta-catenin. During PMN transendothelial migration, all of the junctional components under study were lost at the immediate site of monolayer penetration. Again, at regions removed from the actual site of PMN penetration of the monolayers, the AJ components were unaffected. PMN-induced disorganization of the AJs was partially prevented by an elastase inhibitor.
These findings suggest that adherent PMNs induce a localized, sequential disassembly of AJs, which is partially mediated by PMN-derived elastase and involves the initial loss of an intracellular component of AJs (i.e., beta-catenin).
体外研究表明，多形核白细胞（PMN）通过细胞旁途径（即通过内皮细胞间连接）穿过内皮细胞单层。在此，我们评估了在PMN跨内皮细胞迁移过程中黏附连接（AJ）是否被破坏。
人脐静脉内皮细胞（HUVEC）在多孔膜上生长至汇合，并用白细胞介素-1β激活，甲酰甲硫氨酰亮氨酰苯丙氨酸促进PMN跨内皮迁移。使用双重免疫荧光染色和激光扫描共聚焦显微镜，我们评估了PMN-内皮细胞黏附相互作用（即黏附于单层并穿过单层迁移）对AJ成分血管内皮（VE）-钙黏蛋白、β-连环蛋白、α-连环蛋白和γ-连环蛋白的影响。
在紧邻黏附的PMN的AJ中，一些AJ成分的染色缺失。远离HUVEC-PMN黏附相互作用的AJ成分未受影响。一种迭代方法表明，这四种成分从AJ中依次丢失。β-连环蛋白首先丢失，其次是VE-钙黏蛋白、α-连环蛋白，最后是γ-连环蛋白。在没有PMN的情况下，VE-钙黏蛋白的交联增加了β-连环蛋白的细胞质积累，但血小板内皮细胞黏附分子-1或细胞间黏附分子-1的交联未增加。在PMN跨内皮迁移过程中，所有研究的连接成分在单层穿透的直接部位丢失。同样，在远离单层PMN实际穿透部位的区域，AJ成分未受影响。弹性蛋白酶抑制剂部分阻止了PMN诱导的AJ紊乱。
这些发现表明，黏附的PMN诱导AJ的局部、顺序性解体，这部分由PMN衍生的弹性蛋白酶介导，并且涉及AJ的细胞内成分（即β-连环蛋白）的最初丢失。
