Basophil Responses to Chemokines Are Regulated by Both Sequential and Cooperative Receptor Signaling

• Published in: The Journal of immunology (1950) 2000-12, Vol.165 (12), p.7224-7233
• Main Authors: Heinemann, Akos, Hartnell, Adele, Stubbs, Victoria E. L, Murakami, Kazuki, Soler, Dulce, LaRosa, Gregory, Askenase, Philip W, Williams, Timothy J, Sabroe, Ian
• Format: Article
• Language: English
• Subjects: Basophils - cytology; Basophils - immunology; Basophils - metabolism; CCR2 protein; CCR3 protein; Cell Size - immunology; Chemokine CCL2 - metabolism; Chemokine CCL2 - pharmacology; Chemokine CCL7; Chemokine CCL8; Chemokines - blood; Chemokines - pharmacology; Cytokines; Eosinophils - cytology; Eosinophils - metabolism; eotaxin; Flow Cytometry; Humans; Ion Channel Gating; Leukocytes, Mononuclear - cytology; Monocyte Chemoattractant Proteins - pharmacology; Monocytes - cytology; Monocytes - metabolism; Receptors, CCR2; Receptors, CCR3; Receptors, Chemokine - biosynthesis; Receptors, Chemokine - blood; Receptors, Chemokine - physiology; Scattering, Radiation; Signal Transduction - immunology
• ISSN: 0022-1767; 1550-6606
• DOI: 10.4049/jimmunol.165.12.7224

To investigate human basophil responses to chemokines, we have developed a sensitive assay that uses flow cytometry to measure leukocyte shape change as a marker of cell responsiveness. PBMC were isolated from the blood of volunteers. Basophils were identified as a single population of cells that stained positive for IL-3Ralpha (CDw123) and negative for HLA-DR, and their increase in forward scatter (as a result of cell shape change) in response to chemokines was measured. Shape change responses of basophils to chemokines were highly reproducible, with a rank order of potency: monocyte chemoattractant protein (MCP) 4 (peak at /= eotaxin-2 = eotaxin-3 >/= eotaxin > MCP-1 = MCP-3 > macrophage-inflammatory protein-1alpha > RANTES = MCP-2 = IL-8. The CCR4-selective ligand macrophage-derived chemokine did not elicit a response at concentrations up to 10 nM. Blocking mAbs to CCR2 and CCR3 demonstrated that responses to higher concentrations (>10 nM) of MCP-1 were mediated by CCR3 rather than CCR2, whereas MCP-4 exhibited a biphasic response consistent with sequential activation of CCR3 at lower concentrations and CCR2 at 10 nM MCP-4 and above. In contrast, responses to MCP-3 were blocked only in the presence of both mAbs, but not after pretreatment with either anti-CCR2 or anti-CCR3 mAb alone. These patterns of receptor usage were different from those seen for eosinophils and monocytes. We suggest that cooperation between CCRs might be a mechanism for preferential recruitment of basophils, as occurs in tissue hypersensitivity responses in vivo.