EVIDENCE THAT THE RECEPTOR FOR C4A IS DISTINCT FROM THE C3A RECEPTOR

The cDNAs encoding the human (hC3aR) and mouse C3a receptors (mC3aR) w ere functionally expressed in RBL-2H3 cells. A calcium mobilization as say was utilized to assess the biologic activity of human anaphylatoxi ns, and C3a synthetic peptide agonists on hC3aR and mC3aR cells and th is activity was compared to the activity of the anaphylatoxins on huma n neutrophils. Both hC3aR and mC3aR cells responded in a concentration -dependent manner with a robust calcium mobilization response to C3a w ith 50% effective concentrations (EC50s) of 0.24 nM and 1.3 nM, respec tively. The response obtained with hC3aR cells was similar to the resp onse elicited by C3a on human neutrophils (EC50 0.77 nM). The potency of a C3a analogue synthetic peptide (WWGKKYRASKLGLAR), derived from th e fifteen carboxy-terminal residues (63-77) of C3a, relative to C3a, i n stimulating calcium mobilization differed on cells expressing the hu man vs, mouse receptors. While the peptide was approximately 10 fold l ess active than C3a in stimulating calcium mobilization on cells expre ssing the hC3aR (EC50 2.0 nM), the peptide was essentially equipotent to the native ligand when tested on cells expressing the mC3aR. Data o btained with C4a, purified from activated serum, were difficult to int erpret due to possible trace contamination of the C4a with C5a. Subseq uently, an alternative C4a isolation scheme was utilized, via cleavage in vitro of purified C4. Concentrations of this latter C4a preparatio n, of up to 3.3 mu M, had no effect on calcium mobilization in human n eutrophils or in cells stably expressing the cloned C3a receptors, an indication that C4a does not interact with the C3a receptor. (C) 1997 Elsevier Science B.V.