Activation Of α7nAChR Promotes IL-4-induced Macrophages M2 Polarization Under Influenza A Virus Infection

Background : Macrophages can be polarized towards M1 phenotype upon stimulation with pathogens, LPS or IFN-γ, and this process promotes proinflammatory responses; on the contrary, macrophages can switch to M2 phenotype in response to Th2 cytokines IL-4 or IL-13 by which limits development of inflammation. It has been reported that stimulation of vagus nerve could release acetylcholine by which activates α7 n ACh R in the macrophages to dampen proinflammatory cytokines, but not affect M2 polarization under triggered by LPS or bacteria challenge. Whether activation of α7 n ACh R would regulate IL-4 induced M2 polarization in influenza-infected macrophages is less investigated.Objective:Whether and how activation of α7 n ACh R regulates IL-4-induced M2 polarization in influenza-infected macrophages?Methods:BMDM were pretreated with α7 n ACh R agonist GTS-21 for 30 min, and then challenged with influenza A virus. Six hours later, the cells were stimulated with IL-4. At 24 h, cells were collected to analyze M2 macrophage marker-Arg 1 at both protein and m RNA levels by western blot and Q-PCR respectively. Wild type, α7 n ACh R or Akt1 deficient BMDMs were used for confirming the results of the experiments. In the in vivo study, α7 n ACh R or Akt1 deficient mice were infected with PR8 and sacrificed at 3 or 5 dpi to examine Arg 1 protein levels in the BAL cell lysates and lung homogenates.Results: Activation of α7 n ACh R significantly enhanced IL-4-induced Arg 1 in PR8-infected BMDM without affecting quantity of its transcription factors, such as p-STAT6. Activation of α7 n ACh R could dose-dependently reduce phosphorylation of Akt in PR8-infected BMDM. Knockout of α7 n ACh R markedly reduced IL-4-induced Arg 1 in PR8-infected BMDM and lungs. Without functional Akt1 in PR8-infected BMDM, activation of α7 n ACh R could not efficiently enhance IL-4 induced Arg 1 expression at m RNA level. In an IAV-induced acute lung injury mouse model, lack of Akt1 enhanced Arg 1 at both protein and m RNA levels in both BAL cell lysates and the supernatants of lung homogenate.Conclusion: Activation of α7 n ACh R promotes Arg 1 transcription in IL-4-stimulated PR8-infected BMDM. Phosphorylation of Akt1 or itself may play different roles in mediating Arg 1 expression induced by IL-4 in PR8-infected BMDM.