EXPRESSION OF A FUNCTIONAL ANAPHYLATOXIN C3A RECEPTOR BY ASTROCYTES

Human astrocyte cell lines reportedly contain a specific receptor for the complement anaphylatoxin C3a based on ligand-binding studies, func tional responses, and RNA analysis by RT-PCR. Uptake of I-125-C3a by a strocytes was specific and reversible. Scatchard analysis indicated th e presence of two classes of binding sites. High-affinity binding site s were abundantly expressed (20,000-80,000 sites per cell) with an est imated KD of 1 - 2 nM. Low-affinity binding sites with a K-D of 209 nM were largely expressed (n greater than or equal to 4 x 10(6) sites pe r cell) and probably did not reflect a receptor-mediated binding, but rather an ionic interaction between C3a and the membrane. Analysis of astrocyte mRNA by RT-PCR with three different sets of primers covering 60% of the C3a receptor (C3aR) mRNA sequence indicated that glial C3a R was identical to the leukocytic one. Western blot analysis using a s pecific anti-C3aR evidenced a C3aR with a molecular mass of 60,000 Da, C3a and a superagonist peptide, E7, induced a transient increase of i ntracellular [Ca2+] in primary culture of astrocytes, Treatment of the ligands by carboxypeptidase B to eliminate the C-terminus Arg conside rably decreased the [Ca2+] response. Moreover, flow cytometry experime nts demonstrated the expression of C3aR on normal rat astrocyte membra ne. This report brings new insight for the role of the complement syst em in the brain inflammation response.