Figure 5. Blocking SIRPα signaling in DCs inhibits the clearance of engulfed mtDNA by activating NOX2.

(A) MC38 tumor cells were co-cultured with BMDCs or macrophages in the presence of 30µg/ml rat Ig or anti-CD47 mAb. Generation of phagosomal ROS from DCs or macrophages was measured. (B) MC38 tumor cells were co-cultured with bone marrow-derived DCs or macrophages from WT or gp91^phox−/− mice in the presence of 30µg/ml rat Ig or anti-CD47 mAb. Generation of phagosomal ROS from DCs or macrophages was measured and normalized to cells treated with rat Ig. (C) BMDCs were co-cultured with MC38 in the presence of rat Ig or anti-CD47 mAb for four hours. CD11c⁺ DCs were sorted by FACS and immunoprecipitated with antibody to p47^phox. Immunoblot analysis of tyrosine-phosphorylated proteins (p-Tyr) of p47^phox, or total p47^phox, SIRPα and SHP-1 was shown. (D) BMDCs or BMMs from WT or gp91^phox−/− mice were co-cultured with MC38 in the presence of rat Ig or anti-CD47 mAb for four hours. Cytosolic DNA was extracted and quantitated. Data are represented as mean ± SEM. *p< 0.05, **p< 0.01. ns, not significant (Two-tailed student’s t test). Data are representative of two independent experiments. See also Figure S4 and Figure S5.