Figure 6.

Efferocytosis is impaired in NRP2‐deficient AM. (A, B) AM from Nrp2^fl/fl or LysMcre/Nrp2^fl/fl mice were cultured with PKH26‐labeled apoptotic Jurkat cells at a ratio of 5:1 apoptotic cells:AM. Two hours later, AM was washed and efferocytosis of apoptotic cells (AC) was determined by flow cytometry. (A) Representative cytograms showing efferocytosis of PKH26‐labeled apoptotic cells by AM from Nrp2 ^fl/fl mice (blue histogram) or LysMcre/Nrp2 ^fl/fl mice (red histogram). AM cultured without apoptotic cells (gray histogram) were included as a negative control for gating. (B) Phagocytosis index for Nrp2^fl/fl or LysMcre/Nrp2^fl/fl AM. Bars represent mean ± SEM ( = 5 mice per group). (C) AM was isolated from Nrp2^fl/fl or LysMcre/Nrp2^fl/fl  mice that had been sensitized with OVA + LPS and challenged with OVA as in Figure 2A. Efferocytosis of apoptotic Jurkat cells was determined as in (A). Bars represent mean ± SEM (n = 5 mice per group). (D) Phagocytosis index for NRP2‐deficient (RAW‐NRP2KO) and control (RAW‐Cas9) RAW 264.7 macrophages. Bars represent mean ± SEM of triplicate wells. Representative data from one of two independent experiments is shown. (E) AM from Nrp2^fl/fl  or LysMcre/Nrp2^{fl /fl}  mice were cultured with apoptotic cells at a ratio of 5:1 apoptotic cells:AM for 24 h. Levels of IL‐10 and TGF‐β in cell culture supernatants were measured by ELISA. **p < .01, ***p < .001, Student's t‐test. AM, alveolar macrophages; ELISA, enzyme‐linked immunosorbent assay; IL‐10, interleukin‐10; LPS, lipopolysaccharide; NRP2, neuropilin‐2; ns, not significant; OVA, ovalbumin; TGF‐β, transforming growth factor‐β