Regulatory Mechanisms of Chemokine-induced Eosinophil Adhesion and Transmigration with Pulmonary Microvascular Endothelial Cells

嗜酸性粒细胞通过α 4 β 1整合素/VCAM-1与内皮细胞相互作用被认为是选择性嗜酸性粒细胞募集到支气管哮喘气道的关键步骤。然而，这是可能的，通过这一途径的牢固粘附实际上抑制了随后的跨内皮迁移。本研究旨在确定诱导嗜酸性粒细胞在VCAM-1表达的滤器条件下迁移的因素。我们首先检测了一组嗜酸性粒细胞化学引诱物对嗜酸性粒细胞迁移穿过静息或TNF-α + IL-4处理的人肺微血管内皮细胞（HPMEC）单层的影响，因此VCAM-1强烈表达。嗜酸性粒细胞移行（PAF）（0.3 μ M），FMLP（0.1 μ M）、IL-5、GM-CSF无论HPMEC的处理条件如何，观察到C-C趋化因子RANTES、嗜酸性粒细胞趋化因子（eotaxin）和IL-8（10nM）的表达水平相似。当与静息HPMEC相比时，嗜酸性粒细胞趋化因子-2（3nM）、MCP-3或MCP-4（均为10nM）显著增加嗜酸性粒细胞穿过VCAM-1表达的HPMEC的迁移（p <0.05）。为了进一步证实这些发现，还测定了嗜酸性粒细胞穿过重组人（rh）-ICAM-1-或rh-VCAM-1-包被滤器的迁移。与FCS（对照）或rh-ICAM-1涂覆的过滤器相比，RANTES、嗜酸性粒细胞活化趋化因子、嗜酸性粒细胞活化趋化因子-2、MCP-3或MCP-4，而不是其他化学引诱剂，显著增加了嗜酸性粒细胞穿过rh-VCAM-1涂覆的过滤器的迁移（p <0.05）。抗β 2整合素和抗α 4整合素mAb分别显著抑制RANTES诱导的嗜酸性粒细胞穿过rh-ICAM-1和rh-VCAM-1涂层滤器的迁移。这些结果表明，通过α 4整合素/VCAM-1粘附于内皮细胞的嗜酸性粒细胞在C-C趋化因子存在下有效地进行迁移。

Eosinophil interaction with endothelial cells via α4β1 integrin/VCAM-1 is considered a key step for selective eosinophil recruitment to the airways of bronchial asthma. It is possible, however, that firm adhesion via this pathway actually inhibits subsequent transendothelial migration. This study was conducted to identify factor (s) that induces eosinophil transmigration across VCAM-1-expressed filter conditions. We first examined the effect of a panel of eosinophil chemoattractants on eosinophil migration across resting or TNF-α + IL-4-treated, hence strongly VCAM-1-expressed, human pulmonary microvascular endothelial cell (HPMEC) monolayers. Eosinophil transmigration by PAF (0.3μM), FMLP (0.1μM), IL-5, GM-CSF (both at 100pM) or IL-8 (10nM) was observed similar degree regardless of the treatment conditions of the HPMEC.Interestingly, C-C chemokines RANTES, eotaxin (both at 30nM), eotaxin-2 (3nM), MCP-3 or MCP-4 (both at 10nM) significantly increased eosinophil migration across VCAM-1-expressed HPMEC when compared with resting HPMEC (p<0.05). To further confirm these findings, eosinophil migration across either recombinant human (rh)-ICAM-1-or rh-VCAM-1-coated filter was also determined. RANTES, eotaxin, eotaxin-2, MCP-3 or MCP-4, but not other chemoattractants, significantly increased eosinophil migration across rh-VCAM-1-coated filters compared with FCS (control)- or rh-ICAM-1-coated filters (p<0.05). RANTES-induced eosinophil migration across rh-ICAM-1- and rh-VCAM-1-coated filters was significantly inhibited by anti-β2 integrin and anti α4 integrin mAb, respectively. These results suggest that eosinophils, which adhered to endothelial cells via α4 integrin/VCAM-1, undergo transmigration efficiently in the presence of C-C chemokines.