Human recombinant soluble decay accelerating factor inhibits complement activation in vitro and in vivo.
Complement plays a role in activating the inflammatory response and has been implicated in the pathogenesis of some inflammatory diseases. With a view toward controlling unwanted C activation, we evaluated the C regulator, human decay accelerating factor (DAF). Three forms of recombinant DAF were purified from transfected Chinese hamster ovary cells: glycophosphatidylinositol (GPI)-linked membrane DAF (mDAF) extracted from cell membranes; spontaneously shed soluble DAF (sDAF) derived from mDAF; and a novel secreted protein (seDAF), generated by deletion of the signal for GPI attachment. We show that all three molecules inhibit both the classical and alternative pathways of C activation. The following observations indicate that mDAF extracted from Chinese hamster ovary cells reincorporates into RBC membranes via its GPI anchor: 1) cells that are preincubated with mDAF and then washed remain fully protected from C-mediated hemolysis; 2) incubation with phosphatidylinositol-specific phospholipase C abolishes this protection; and 3) sDAF and seDAF, which lack a GPI anchor, do not associate with cell membranes. mDAF is a more potent inhibitor of C-mediated hemolysis than either sDAF or seDAF, suggesting that incorporation into cell membranes greatly enhances the efficiency with which DAF inhibits C activation on the cell surface. In contrast, C activation in the fluid phase is inhibited by sDAF and seDAF, but not by mDAF, possibly due to interference by serum lipoproteins. A reversed passive Arthus reaction in guinea pigs was used to evaluate the ability of recombinant seDAF to inhibit C activation in vivo. When administered at dermal sites, seDAF reduced the severity of immune complex-mediated inflammatory reactions induced by a reversed passive Arthus reaction, as judged by both gross and histologic examination. These data indicate that seDAF may be useful as an anti-inflammatory therapeutic.
Moran
P
P
Genentech, Inc., South San Francisco, CA 94080.
Beasley
H
H
Gorrell
A
A
Martin
E
E
Gribling
P
P
Fuchs
H
H
Gillett
N
N
Burton
L E
LE
Caras
I W
IW
D000818:Animals; D001183:Arthus Reaction / Q000517:prevention & control; D001483:Base Sequence; D018958:CD55 Antigens; D003167:Complement Activation / Q000187:drug effects*; D003169:Complement Inactivator Proteins / Q000494:pharmacology*; D006168:Guinea Pigs; D006801:Humans; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000494:pharmacology*; D051379:Mice; D008969:Molecular Sequence Data; D011817:Rabbits; D051381:Rats; D011994:Recombinant Proteins / Q000494:pharmacology; D013045:Species Specificity
D018958:CD55 Antigens; D003169:Complement Inactivator Proteins; D008565:Membrane Proteins; D011994:Recombinant Proteins