Figure 4. Annexin A2 binds factor H and increases complement activation in the fluid phase.

A) Factor H was purified from mouse plasma by heparin chromatography and Annexin A2 was detected in samples generated from wild-type serum and in a commercial preparation of factor H, but it was not seen in a sample generated from factor H-deficient (fH^−/−) serum. Bands from two different blots are shown. B) Factor H purified from plasma and recombinant annexin A2 were passed over a size exclusion column. Annexin A2 in the fractions was detected by Western Blot Analysis. Co-purified annexin A2 was detected in all of fractions of factor H purified from serum. When recombinant annexin A2 (rA2) was passed over the column in the absence of factor H, it did not elute in the higher molecular weight fractions (fractions 1 and 2; indicated with arrows). C) Purified annexin A2 was added to normal mouse serum and incubated at 37° C for 30 minutes, and C3a in the supernatant was measured by ELISA. The addition of annexin A2 to the reaction increased generation of C3a. The groups were compared by ANOVA. D) Western blot analysis of plasma from mice injected with recombinant annexin A2 revealed greater cleavage of plasma C3 compared to mice injected with PBS as a vehicle control.