Characterization of synthetic C3a analog peptides on human eosinophils in comparison to the native complement component C3a

The C3a anaphylatoxin is a potent proinflammatory mediator derived from the complement system inducing biologic effects of human eosinophils like Ca2 transients and the activation of the respiratory burst. These findings sup port an important role for C3a in diseases typically associated with a peri pheral blood or tissue eosinophilia, Synthetic human C3a analogue peptides with variations at the C-terminal effector domain have been evaluated with respect to their binding affinity and signaling potency on human eosinophil s, Flow cytometrical analysis and RT-PCR revealed that the C3a receptor is constitutively expressed on human eosinophils, Peptides bearing an N-termin al 9-fluorenylmethoxycarbonyl and the 6-aminohexanoyl motif were the most p owerful peptides tested. Amino acid replacements in the conserved C-termina l pentapeptide decreased binding affinity and functional potency substantia lly. In addition, synthetic C3a analogue peptides induced C3aR internalizat ion, led to transient changes of intracellular Ca2+ concentration, and did release reactive oxygen species in human eosinophils indicating the in vivo relevance of C3a-related sequences. The tripeptide LAR was found to be ess ential for C3a receptor binding on human eosinophils. Moreover, the putativ e binding motif of C3a anaphylatoxin is also crucial for the induction of b iologic effects in the human system such as changes of intracellular Ca2+ c oncentration and the release of reactive oxygen species. This study demonst rates that the carboxyl terminus is important for the interaction with the C3aR and the biologic potency of C3a anaphylatoxin in the human system and plays a keg role in the activation process of human eosinophils.