Figure 5.

CD47 is involved in the regulation of hemoglobin expression in alveolar macrophages

Mice were i.n. infected with IAV. aMФ were isolated from WT and CD47^−/− mice 3 dpi.

(A) Transcriptome analysis of isolated WT and CD47^−/− aMФ were assessed by an Affymetrix MicroArray. Hemoglobin alpha (HBA) (B) and beta (HBB) (C) chains were detected by flow cytometry analysis in aMФ isolated from WT and CD47^−/− mice. Representative histograms for HBA and HBB are shown. Mean fluorescence intensity (MFI) for HBA and HBB from aMФ are shown (n = 4).

(D) Whole hemoglobin (HB) concentration was assessed in BALF at indicated time points by ELISA (n = 7–17).

(E) aMФ were isolated from infected WT and CD47^−/− mice 3 dpi and stimulated ex vivo with thrombospondin-1 (TSP-1). After 24 h, HB concentration in the supernatants was measured by ELISA (n = 4).

(F) Type II alveolar epithelial cells (AEC II) were isolated from the lung of naive WT mice and incubated with 1 μg/mL Alexa Fluor 647 (AF647)-labeled HB for 30 min. Representative flow cytometry plots are shown. F) Percentages of HB⁺ AEC II cells are shown (n = 4).

(G) Isolated cells from lungs of WT mice were incubated with 1 μg/mL AF647-labelled HB for 30 min.

Percentages of HB⁺ aMФ, iMФ, and DCs are shown. Results depicted were obtained from one (A) or two independent experiments (C, D, E, G, and H). Data shown are mean ± SEM. One-way ANOVA with Tukey’s multiple-comparisons post-test (D and E) or Student’s t test (F) were performed. ∗ = p < 0.05, ∗∗ = p < 0.01, ∗∗∗∗ = p < 0.0001.