The C3a molecule is an anaphylatoxin of the C system with a wide spectrum of proinflammatory effects predominantly on cells of myeloid origin. In this study we investigated the expression of the high affinity receptor for C3a (C3aR) in human T lymphocytes using receptor-specific mAb. C3aR expression was detected in CD4+ and CD8+ blood- or skin-derived T cell clones (TCC) from birch pollen-sensitized patients with atopic dermatitis. No significant difference in C3aR expression in CD4+ or CD8+ TCCs could be observed. In contrast to C3a(desArg), C3a led to a transient calcium flux in TCCs expressing the C3aR, whereas C3aR-negative TCCs were unreactive. Circulating T cells from patients suffering from severe inflammatory skin diseases expressed the C3aR, whereas no expression of C3aR could be found in unstimulated T lymphocytes from patients with mild inflammatory skin diseases or from healthy individuals. Type I IFNs, which are potent stimulators of cellular immunity, were identified as up-regulators of C3aR expression in vitro in freshly isolated or cloned T lymphocytes. Moreover, C3aR+ T cells were found at the sites of injection in IFN-β-treated patients with multiple sclerosis. These data provide direct evidence for the expression of C3aR on activated human T lymphocytes; this may point to a biological function of C3a in T cell-dependent diseases.

Varying results have been published in the past regarding the reactivity of different leucocyte subpopulations, including neutrophils, monocytes and B lymphocytes, to the anaphylatoxin C3a and its degradation product C3a(desArg). To better characterize the cellular distribution of C3a receptor (C3aR) expression, monoclonal antibodies against two different epitopes on the third extracellular domain of the human C3aR were generated. Quantification of C3aR as compared with C5aR densities was performed on peripheral blood leucocytes by quantitative indirect immunofluorescence. Eosinophils and basophils expressed similar numbers of C3aR and C5aR molecules/cell. On eosinophils 10 700±4500 (mean±SD) C3aR and 14 700±4100 C5aR were found, whereas basophils carried 8100±2100 C3aR and 13 500±3800 C5aR. Monocytes expressed approximately six times more C5aR than C3aR molecules on their surface (6000±2500 C3aR versus 34 100±9300 C5aR molecules) whereas on neutrophils, the expression of C5aR was more than 20 times higher than the expression of C3aR (3100±1000 C3aR versus 63 500±12 200 C5aR). No C3aR expression was detectable on peripheral blood‐derived B lymphocytes and on tonsillar B cells before and after stimulation with interleukin‐2/Staphylococcus aureus Cowan strain I. Our findings correspond well with the paucity of data on C3a‐induced functional activities in monocytes and neutrophils and suggest that eosinophilic and basophilic granulocytes represent the primary effector cells in the peripheral blood which can be stimulated by C3a.

SUMMARYDendritic cells (DC) are recruited to sites of in¯ammation for the initiation of immune responses. As the anaphylatoxins C5a and C3a are important mediators of in¯ammation, we investigated the expression of their receptors (C3aR and C5aR) on human DC. DC were isolated from human skin or generated from puri®ed blood monocytes and were identi®ed by their expression of CD1a or CD83. Freshly isolated or cultured dermal CD1a+ and CD83 + DC bound anti-C5aR and antiC3aR monoclonal antibodies (mAbs), as detected by¯ow cytometry. C5a induced calcium¯uxes in dermal CD1a+ and CD83 + DC, which could be inhibited by C17/5, an anti-C5a mAb. C3a did not induce calcium¯uxes in these cells. Anaphylatoxin receptor expression was down-regulated on dermal DC by adding tumour necrosis factor-a (TNF-a) to the culture medium. On CD1a + CD83 ± cells generated from isolated blood monocytes by culture with 6 . 25 ng/ml of granulocyte± macrophage colony-stimulating factor (GM-CSF) and 125 U/ml of interleukin-4 (IL-4), expression of both C5aR and C3aR was observed. In these cells, both C5a and C3a induced calcium¯uxes. After addition of TNF-a to the culture medium, the majority of the CD1a + cells expressed CD83 + . These cells ± expressing a phenotype of`mature DC' ± down-regulated the expression of the anaphylatoxin receptors and lost their reactivity to the respective ligands. Our results demonstrate the expression of the anaphylatoxin receptors C5aR and C3aR on human skin-derived DC and blood-derived cells expressing the DC-associated membrane molecule, CD1a. Furthermore, the expression of anaphylatoxin receptors on CD83 + dermal DC is indicative of an intermediate stage of maturation of these cells, which was not observed on in vitro-differentiated CD83 + cells.

International scoring system for evaluating prognosis in myelodysplastic syndromes. Blood 89:2079-88 Haralambieva E, Kleiverda K, Mason DY et al. (2002) Detection of three common translocation breakpoints in non-hodgkin's lymphomas by fluorescence in situ hybridization on routine paraffin-embedded tissue section.

Controversial results have been published in the past regarding the functional reactivity of monocytes (Mo) and macrophages (M phi) to the anaphylatoxin C3a and its degradation product C3a(desArg). In this study we performed binding and calcium mobilization experiments with recombinant human C3a (rC3a) and rC3a(desArg). Blood Mo displayed non-inhibitable binding of FITC-labeled rC3a (rC3aFITC) but responded to rC3a with a transient release of the intracellular calcium concentration ([Ca2+]i), whereas rC3a(desArg) was completely inactive. In contrast, binding of rC3aFITC to eosinophilic granulocytes and the mast cell line HMC-1 which have been shown previously to express C3a binding sites could be blocked by a monoclonal anti-C3a antibody. The rC3a-induced [Ca2+]i release in blood Mo was pertussis toxin (PTX)-sensitive suggesting the involvement of G-proteins in the signal transduction pathway. Skin-derived Mo/M phi reacted similarly to blood Mo as no specific binding of rC3aFITC to these cells could be demonstrated, whereas an intracellular release of calcium ions in response to the anaphylatoxin was observed. Homologous desensitization to rC3a but not heterologous desensitization to rC5a was detected in further experiments. The functional effect of C3a, but not the unspecific binding of rC3aFITC to blood Mo and skin-derived Mo/M phi could be blocked by the monoclonal anti-C3a antibody. These results suggest the expression of the recently cloned G-protein-coupled receptor for C3a on human blood Mo and skin-derived Mo/M phi. However, the total number of specific C3a binding sites on these cells is distinctly lower as compared to eosinophilic granulocytes and cells of the mast cell line HMC-1. The small number of C3a receptors on Mo/M phi may be masked by a pronounced non-inhibitable binding of rC3aFITC. This binding, however, may contribute to the recently described biological effects of C3a(desArg) on Mo.