Figure 3.

Binding of anti-dsDNA antibodies to annexin II on HMC mediates antibody internalization. (A and B) Cell surface staining of control human IgG (A) and anti-dsDNA antibodies (B) to HMC with time at either 4°C (○) or 37°C (●) as detected by flow cytometry. *P < 0.01 for 4°C versus 37°C. Reduced cell surface staining of anti-dsDNA antibodies on HMC was observed in cells incubated at 37°C but not at 4°C (B). (C) Representative images depicting colocalization of anti-dsDNA antibodies with cell surface annexin II expression at T = 0 (arrow, denoted as yellow staining in merged images), followed by internalization of anti-dsDNA antibodies to the nuclei within 30 minutes (depicted by asterisk). (D) Parallel studies were conducted in which anti-dsDNA antibodies were incubated with recombinant human annexin II before incubation with HMC. These representative images show no significant binding of these anti-dsDNA antibodies to cells and no antibody internalization after 30 minutes. (E) Representative images showing HMC incubated with control IgG. The nuclei are stained with DAPI. Magnification, ×400 for all.