Uptake of human C5a by neutrophils was monitored in vitro using both I
-125-labeled and unlabeled C5a. The Ligand was internalized by the cel
ls in a dose-dependent manner and maximal binding/uptake was observed
after 5 min of incubation. Neutrophils were incubated with labeled C5a
and the cytosol and supernatant were analyzed by sodium dodecyl sulfa
te-polyacrylamide gel electrophoresis and autoradiography, C5a degrada
tion products were primarily observed in the supernatant, whereas most
df the protein remained intact in the cytosol even after 60 min of in
cubation. Cytosol from neutrophils incubated for 20 min with unlabeled
C5a was examined by radioimmunoassay and found to contain antigenical
ly intact C5a and retained the ability to induce a neutrophil (shape c
hange) response. The functional activity of C5a recovered from the cyt
osol was inhibited by antibodies to either C5a or the C5a receptor (CD
88). This data supports our hypothesis that although C5a is internaliz
ed it remains antigenically intact and functionally active inside the
cell and is primarily degraded extracellularly.