We have previously shown that serum opsonization of immune complexes reduces their capacity to stimulate the secretion of pro-inflammatory cytokines fromleukocytes. These effects may be mediated either by the capacity of complement to reduce immune complex size (therefore limiting Fcγ-receptor cross-linking) or through inhibitory signals transduced through complement receptors. We tested each of these hypotheses using model immune complexes.

Unopsonized immune complexes consisted of BSA-anti-BSA formed at 2x antigen excess. Complexes were further purified by affinity chromatography on Protein A or subjected to gel filtration on Sephacryl-300. Complexes subjected to gel filtration were divided into 3 fractions of large (Fx1), medium (Fx2) and small (Fx3) molecular weight. C4 complexes consisted of model immune complexes incubated with purified human C4, which was cleaved in the fluid phase with activated C1s. Complexes were incubated with normal human peripheral blood mononuclear cells for 48 hours after which IL-1β was measured by ELISA in the cell culture supernates.

The capacity of unopsonized immune complexes to elicit IL-1β secretion from normal human PBMC's was directly related to their size. Fx1, the fraction of highest molecular weight, produced significantly more IL-1β (571± 451 pg/ml) than either Fx2 (142 ± 81 pg/ml) or Fx 3 (139± 71 pg/ml p<0.007). The binding of C4b to immune complexes significantly enhanced the capacity of the complexes to induce IL-1β secretion (884 ± 90 pg/ml) compared with unopsonized complexes (472± 100; p=0.001).

We conclude that complement binding per se does not reduce the capacity of immune complexes to induce cytokine secretion from leukocytes, and, indeed, enhances it. The reduced capacity of complement-opsonized immune complexes to induce IL-1β secretion from leukocytes is probably due to the capacity of complement to regulate immune complex size. Furthermore, these studies underline the phlogistic potential of immune complexes isolated from juvenile rheumatoid arthritis synovial fluids, which are both of high molecular weight and have bound C3b/C4b in vivo.