Eosinophil Transmigration across VCAM-1-Expressing Endothelial Cells Is Upregulated by Antigen-Stimulated Mononuclear Cells

Abstract

Background: Adhesion to and transmigration across endothelial cells expressing vascular cell adhesion molecule 1 (VCAM-1) may be key steps in the development of selective eosinophil accumulation at the allergic inflammation sites. There is evidence that cytokines/chemokines produced by CD4+ T cells play a prominent role in these processes. Objective: The objective of this study was to evaluate whether eosinophil migration across human pulmonary microvascular endothelial cells (HPMEC) expressing VCAM-1 is modulated by the supernatants of antigen-stimulated mononuclear cells obtained from atopic asthmatics. Methods: Peripheral blood mononuclear cells (PBMC) were isolated from Dermatophagoides farinae(Df)-sensitive asthmatic subjects and cultured for 96 h at 37°C in the presence or absence of 1 µg/ml Df antigen. Eosinophils were isolated from blood of healthy subjects and placed on the HPMEC monolayers cultured on a transwell filter (3-µm pore size) stimulated with IL-4 plus TNF-α (both at 100 pM, 24 h) The supernatants of PBMC were then applied to the lower compartment and the transmigration of eosinophils was examined. Results: The supernatants of PBMC stimulated with Df significantly enhanced the eosinophil transmigration across VCAM-1-expressing HPMEC (% migration: 7.6 ± 0.6 by the supernatants of PBMC cultured without Df vs. 12.3 ± 1.2 by the PBMC cultured with Df, p < 0.01, n = 8). The enhanced migration, but not spontaneous migration, was blocked by the anti-α₄ integrin antibody. Moreover, the enhanced transmigration was blocked by anti-CCR3 antibody. Conclusion: The antigen-stimulated PBMC from atopic asthmatics produce an activity to induce eosinophil migration across VCAM-1-expressing endothelial cells. This activity appears to involve CCR3 as an essential molecule.