Biochemical and biological characterization of neutrophil chemotactic protein, a novel rabbit CXC chemokine from alveolar macrophages

The role of interleukin-8 (IL-8) and related CXC chemokines has been demons trated in many human diseases. However, more profound studies, e.g., by blo cking the effect of these inflammatory mediators, request animal models and hence the identification of all human counterparts for commonly used labor atory animals. In this study, we describe the identification of a novel neu trophil chemotactic protein (NCP) of the rabbit. Intact and NH2-terminally truncated NCP forms and IL-8 were isolated from LPS-stimulated rabbit alveo lar macrophages and purified to homogeneity by a four-step purification pro cedure. Determination of the complete primary structure of NCP by mass spec trometry and NH2-terminal sequencing of natural protein revealed high struc tural homology with human epithelial cell-derived neutrophil attractant-78 (ENA-78) and granulocyte chemotactic protein-2 (GCP-2), two related ELR+CXC chemokines, Intact NCP(1-76) was found to be 10-fold less potent than trun cated NCP(7,8-76) at inducing neutrophil chemotaxis. NCP(7,8-76) was equall y potent as intact rabbit IL-8 at chemoattracting human neutrophils and at inducing calcium fluxes in rabbit neutrophils, 1 ng/mL being the minimal ef fective concentration. However, like IL-8, NCP failed to induce monocyte or eosinophil migration at 300-fold higher concentrations. IL-8 desensitized the calcium increase induced by NCP and vice versa. Finally, intradermal in jection of NCP induced a dose-dependent and significant infiltration of neu trophils in mice skin, It can be concluded that NCP is a novel rabbit CXC c hemokine that is, like IL-8, implicated in animal models used to study vari ous human disorders in which neutrophils play an important role.