MPO from mouse neutrophils decreases DC activation in vitro leading to reduced adaptive immune response in vivo. (A) BM-derived murine DCs were cultured (18 hours) with LPS (1 μg/mL), H₂O₂ (100 μM), and increasing doses of supernatant from CB/fMLP (± ABAH)-stimulated WT or Mpo^−/− neutrophils. DC activation was assessed by measuring production of IL-12 and IL-23 (ELISA) and expression of CD86 and CD40 (flow cytometry). Solid line represents DCs cultured with LPS/H₂O₂ only. Dotted line represents unstimulated DCs. (B) DCs were cultured with LPS/H₂O₂ and (Bi-ii) increasing concentrations of purified enzymatically active mouse MPO or (Biii-iv) with or without MPO (0.125 μg/mL) ± ABAH. (C) OVA_323-339-coated DCs, which had been cultured with LPS/H₂O₂ and (Ci-ii) with or without supernatant (5% v/v) from CB/fMLP-stimulated WT or Mpo^−/− neutrophils or (Ciii-iv) with or without purified MPO (0.125 μg/mL) ± ABAH, were injected subcutaneously into WT recipients (n = 6-8/group). Two recipients received uncoated DCs cultured only with LPS/H₂O₂. OVA-specific skin DTH and serum IgG (1/50 dilution; ELISA) were measured 9 days later. Data are representative of 2-3 independent experiments. Each group in (A-B) contained 4 replicates. *P < .05, **P < .01, ***P < .001. SN, supernatant.