Neutrophil-DC hybrids acquire functionality as APCs. (A) Ly6G⁺/CD11c⁻/MHC II⁻ neutrophils, Ly6G⁺/CD11c⁺/MHC II⁺ neutrophil-DC hybrids, and Ly6G⁻/CD11c⁺/MHC II⁺ traditional DCs purified from GM-CSF–supplemented BM culture (d 6) were examined for TLR mRNA expression profiles by quantitative polymerase chain reaction (means ± SD from triplicate samples). (B-C) The same 3 purified populations were cultured for 24 h with phosphate-buffered saline alone or each of the indicated TLR agonists. The samples were then examined for cytokine release measured by the cytometric bead array system (means ± SD from triplicate cultures) (B) and surface expression of CD40 (C). (D) The same 3 purified populations were pulsed for 60 min with the indicated form of OVA antigen and then co-cultured with OVA-specific CD4 T cells from OT-II TG mice (means ± SD from triplicate cultures). *P < .05, **P < .01, ***P < .001 compared with control group treated with PBS alone. Data are representative of at least 2 independent experiments.