Binding of fluorescein-labeled anaphylatoxin C5a to human peripheral blood, spleen, and bone marrow leukocytes

Abstract

The expression of C5a receptors (C5aR) on human leukocytes was evaluated by flow cytometry using fluorescein-labeled human C5a (C5a- F). Granulocytes and CD14+ mononuclear cells (MNL) but not CD3+, CD20+, CD16+, CD56+, or CD11b+ lymphocytes in peripheral blood and spleen bound C5a-F. C5a-F binding was saturable and inhibitable by anti-C5a monoclonal antibody (MoAb) C17/5 or unlabeled C5a. During hemodialysis, which led to the generation of C5a, only granulocytes and monocytes increased their expression of the adhesion molecule CD11b (CR3). In vitro, C5a induced an increase of CR3 and p 150/95 (CD11c/CR4) only on myeloid cells. However, treatment of leukocytes with phorbol 12- myristate 13 acetate increased CR3 and CR4 expression on both myeloid cells and a lymphocyte subpopulation. Stimulation of MNL in mixed lymphocyte cultures or by treatment with conditioned medium or with IFN- gamma did not induce binding sites for C5aR on lymphocytes and reduced the binding of C5a-F to monocytes. The expression of C5aR on low- density bone marrow cells was analyzed by setting appropriate gates during flow cytometry. Cells that bound C5a-F were found in all populations that contained granulocyte and monocyte precursors, but not in lymphocyte precursor populations. All C5aR+ bone marrow cells were CD34 and expressed high levels of CR3, which suggests a late appearance of C5aR during myeloid cell maturation. Our results indicate that C5aR is exclusively expressed on myeloid cells within the hematopoetic cell population.

Expression and modulation of C5a receptor (CD88) on skin dendritic cells. Chemotactic effect of C5a on skin migratory dendritic cells

“…In fact, C5aR is absent in the immature myeloid cell lines KG-1, HL-60, TH-P1 and U937, and it is expressed lately during the maturation of granulocytes and monocytes in bone marrow. 28,29,40 The appearance of C5aR on fully mature myeloid cell is also supported by the observation that lines U937 and HL-60 differentiated with dibutyryl cyclic adenosine monophosphate (AMP) express C5aR. 26,28 According to phenotypic and ultrastructural studies by Romani et al, DC emigrated from epidermal/dermal explants comprised at least two different subsets: (i) epidermal rLC (CD1a high , CD36 negative, Lag negative, Birbeck granules abundant); and (ii) dermal DC (CD1a low , CD36 positive, Lag negative, Birbeck granules absent).…”

Role of complement C5a in mechanical inflammatory hypernociception: potential use of C5a receptor antagonists to control inflammatory pain

“…One of the main effector components of the complement system is the anaphylatoxin C5a. It is one of the most potent inflammatory peptide mediators, and its biological effects result in binding to its G-protein-coupled receptor (C5aR1) present in inflammatory cells, such as neutrophils, eosinophils and monocytes (Chenoweth and Hugli, 1978;Gerard et al, 1989;Werfel et al, 1992). C5a is a potent neutrophil chemoattractant and induces an increase in oxidative burst, phagocytosis and release of granule enzymes by these cells (Hetland et al, 1998).…”

Up‐regulation of C5a receptor expression and function on human monocyte derived dendritic cells by prostaglandin E2

“…1 It belongs to the superfamily of G-protein coupled receptors with seven hydrophobic domains and a serpentine transmembrane structure 2,3 and is expressed on cells of myeloid origin such as granulocytes and monocytes/macrophages. 4,5 Its ligand, C5a, is generated by cleavage of the ®fth component of complement (C5) upon activation of the classical or the alternative pathway of the complement system. It has several proin¯ammatory effects which may lead to changes in blood¯ow and impairment of vascular integrity associated with oedema.…”