Figure 1.
SIGN-R1 directly binds to apoptotic cells in vitro. (a) Jurkat T cells were treated with 3 μM of dexamethasone (Dexa) for 24 h, and apoptosis induction was confirmed with Annexin-V, followed by FACS analysis. Nonapoptotic and apoptotic Jurkat T cells were incubated with 2 μg of purified recombinant SIGN-R1 for 30 min at 37 °C and immunostained with a 22D1 anti-SIGN-R1 monoclonal antibody, which was followed by a FACS analysis. (b) Apoptotic thymocytes of C57BL/6 mice were prepared as in (a), and 1 × 106 nonapoptotic or apoptotic thymocytes were incubated with 2 μg of biotinylated transferrin (Tfe), efOVA, or recombinant purified SIGN-R1 for 30 min at 37 °C and immunostained with Alexa488-conjugated Streptavidin, which was followed by a FACS analysis. (c) As in (c), but thymocytes were incubated with 2 μg of Alexa568-conjugated efOVA or purified SIGN-R1 (red) and analyzed by fluorescent microscopy. (d) As in (b), but 1 × 106 nonapoptotic or apoptotic thymocytes (NA- or A-Thy, respectively) were incubated with 20 μg of whole cell lysates from DCEK_WT, or _SIGN-R1 cells for 1 h at 37 °C and immunostained with ananti-SIGN-R1 monoclonal antibody (Clone 22D1) or a nonreactive hamster IgG control