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Type I Interferon Signaling Regulates Ly6C^hi Monocytes and Neutrophils during Acute Viral Pneumonia in Mice

Figure 5

Ifnar1^−/− bone marrow (BM) cells are unable to differentiate into MCP-1-producing Ly6C^hi monocytes.

(A) BM cells were isolated from WT and Ifnar1^−/− mice pre- and post-influenza infection and analyzed by flow cytometry. (B) Monocytes (Mo) and neutrophils (N) were isolated, cytospun, and stained by H&E (×40). (C) MFI of Ly6C expression of monocytes in the BM of WT and Ifnar1^−/− mice following influenza infection (1×10⁵ pfu). Mean values are in parenthesis. (D) BM cells from naïve WT and Ifnar1^−/− mice were prepared and stimulated in vitro with BALF from PR8 virus (1×10⁵ pfu)-infected WT mice or with PR8 virus (0.5 multiplicity of infection) alone. After 5 days of culture, subsets of Ly6C^hi and Ly6C^int were counted and the percentage of Ly6C^hi among Ly6C^pos was determined. (E) After BM cells were prepared from naïve WT mice, cells were infected with PR8 virus and cultured for 5 days with/without anti-IFNAR1 blocking Ab (100 ng/ml). (F) BM cells infected with PR8 and cultured in vitro for 5 days. Cultured cells were further sorted into Ly6C^neg, Ly6C^int, and Ly6C^hi. Each cell subset was cultured in vitro for 4 h and chemokine levels in the supernatant were analyzed. Values are mean + SD and data are representative of at least two independent experiments.

doi: https://doi.org/10.1371/journal.ppat.1001304.g005