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, functional responses, and RNA analysis by RT-PCR. Uptake of 125I-C3a by astrocytes was specific and reversible. Scatchard analysis indicated the presence of two classes of binding sites. High-affinity binding sites were abundantly expressed (20,000-80,000 sites per cell) with an estimated K(D) of 1-2 nM. Low-affinity binding sites with a K(D) of 209 nM were largely expressed (n > or = 4 x 10(6) sites per 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 C3aR was identical to the leukocytic one. Western blot analysis using a specific anti-C3aR evidenced a C3aR with a molecular mass of 60,000 Da. C3a and a superagonist peptide, E7, induced a transient increase of intracellular [Ca2+] in primary culture of astrocytes. Treatment of the ligands by carboxypeptidase B to eliminate the C-terminus Arg considerably decreased the [Ca2+] response. Moreover, flow cytometry experiments demonstrated the expression of C3aR on normal rat astrocyte membrane. This report brings new insight for the role of the complement system in the brain inflammation response.
Ischenko
A
A
Research Institute of Highly Pure Biopreparations, St. Petersburg, Russia.
Sayah
S
S
Patte
C
C
Andreev
S
S
Gasque
P
P
Schouft
M T
MT
Vaudry
H
H
Fontaine
M
M
D000706:Anaphylatoxins / Q000378:metabolism*; D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D001665:Binding Sites / Q000502:physiology; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D015926:Complement C3a / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D007425:Intracellular Membranes / Q000378:metabolism; D009994:Osmolar Concentration; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D011951:Receptors, Complement / Q000235:genetics / Q000378:metabolism*; D012016:Reference Values
D000706:Anaphylatoxins; D012333:RNA, Messenger; D011951:Receptors, Complement; D015926:Complement C3a; D002118:Calcium