title: Requirement for human red blood cells in inactivation of C3b in immune complexes and enhancement of binding to spleen cells.
abstract: We showed previously that soluble immune complexes (IC) added to normal human serum and unseparated autologous blood cells bind predominantly to red blood cells (RBC) bearing C3b receptors. RBC-associated IC are processed both by C3b/C4b inactivator (I) cleavage of C3b and by alternative pathway-dependent C3b generation, and are returned to the serum in a modified form. In the present study, soluble IC were prepared by using 125I-BSA and guinea pig anti-BSA, and were treated with diluted human serum to optimize sustained C3b-dependent binding to human RBC. This binding to RBC could not be abrogated by preincubation of IC with heat-inactivated human serum or purified serum regulatory proteins, I and beta 1H (H), but after binding, pretreated IC dissociated. When IC were prepared with serum containing 125I-C3, treatment with I + H did not release labeled C3 from the complexes. Analysis by polyacrylamide gel electrophoresis in sodium dodecyl sulfate (SDS-PAGE) of 125I-C3b in IC showed similar labeled peaks whether IC were pretreated with buffer or with I + H or were bound to RBC without pretreatment. In contrast, when IC containing 125I-C3b were exposed to I + RBC, nearly 70% of the label was released from the IC. The released material analyzed by SDS-PAGE showed major 125I-C3 peaks corresponding in m.w. to free beta-chain and alpha 1 cleavage products of 30,000 to 33,000 daltons. IC in undiluted serum after incubation with RBC gave less binding to human polymorphonuclear leukocytes than after incubation without RBC, but notably more binding to guinea pig spleen cells and human lymphoblastoid cells. These findings argue that exposure to serum regulatory proteins does not prevent subsequent binding of experimental IC to RBC, that RBC binding promotes degradation of C3b in IC which proceeds beyond iC3b, and that this I-mediated breakdown renders IC reactive with spleen cells.
mesh_terms: D000818:Animals; D000936:Antigen-Antibody Complex / Q000378:metabolism*; D001665:Binding Sites; D002417:Cattle; D003180:Complement C3b Inactivator Proteins / Q000494:pharmacology* / Q000502:physiology; D017242:Complement Factor H; D004912:Erythrocytes / Q000378:metabolism*; D006168:Guinea Pigs; D006801:Humans; D009504:Neutrophils / Q000378:metabolism; D013154:Spleen / Q000166:cytology / Q000378:metabolism*
chemical_list: D000936:Antigen-Antibody Complex; C493495:CFH protein, human; D003180:Complement C3b Inactivator Proteins; D017242:Complement Factor H