title: Classically Activated Macrophages Protect against Lipopolysaccharide-induced Acute Lung Injury by Expressing Amphiregulin in Mice.
abstract: BACKGROUND
  Alveolar macrophages (AMs) activated into M1 phenotype are involved in the development of lipopolysaccharide-induced acute lung injury (ALI). However, whether AMs express amphiregulin and what roles amphiregulin plays in lipopolysaccharide-induced ALI remain poorly understood.
  
  
  METHODS
  Acute lung injury was induced by intratracheal instillation of lipopolysaccharide in male C57BL/6 mice. Lung injury scores, level of protein, and level of neutrophils in bronchial alveolar lavage fluid of lipopolysaccharide-induced ALI mice were compared with those in mice challenged with recombinant exogenous amphiregulin and antiamphiregulin antibody. Amphiregulin expression in macrophages and neutrophils in bronchial alveolar lavage fluid of lipopolysaccharide-induced ALI mice was determined by using immunofluorescence technique and further detected in M0, M1, and M2 phenotypes of both peritoneal macrophages and AMs. The effect of amphiregulin on apoptosis of MLE12 cells and activation of epithelial growth factor receptor-AKT pathway were, respectively, examined by using flow cytometry and western blotting.
  
  
  RESULTS
  Alveolar macrophages were found to highly express amphiregulin in ALI mice. Amphiregulin neutralization aggravated, whereas recombinant exogenous amphiregulin attenuated lipopolysaccharide-induced ALI in mice (n = 6). In cultured AMs and peritoneal macrophages, amphiregulin was mainly generated by M1, rather than M0 or M2 phenotype (n = 5). Apoptosis ratio of lipopolysaccharide-challenged MLE12 cells was significantly reduced by recombinant exogenous amphiregulin from 16.60 ± 1.82 to 9.47 ± 1.67% (n = 5) but significantly increased from 17.45 ± 1.13 to 21.67 ± 1.10% (n = 5) after stimulation with supernatant of M1-polarized AM media conditioned with amphiregulin-neutrolizing antibody. Western blotting revealed that amphiregulin activated epithelial growth factor receptor and AKT in the lung tissues and MLE12 cells (n = 5).
  
  
  CONCLUSIONS
  Different from the common notion that classically activated AMs have just a detrimental effect on the lung tissues, the results of this study showed that classically activated AMs also exerted a protective effect on the lung tissues by producing high-level amphiregulin in lipopolysaccharide-induced ALI.
authors.affiliation: From the Department of Anesthesiology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, China.
authors.forename: Younian
authors.initials: Y
authors.lastname: Xu
authors.forename: Chen
authors.initials: C
authors.lastname: Meng
authors.forename: Guilin
authors.initials: G
authors.lastname: Liu
authors.forename: Dong
authors.initials: D
authors.lastname: Yang
authors.forename: Lisha
authors.initials: L
authors.lastname: Fu
authors.forename: Min
authors.initials: M
authors.lastname: Zhang
authors.forename: Zhao
authors.initials: Z
authors.lastname: Zhang
authors.forename: Huimin
authors.initials: H
authors.lastname: Xia
authors.forename: Shanglong
authors.initials: S
authors.lastname: Yao
authors.forename: Shihai
authors.initials: S
authors.lastname: Zhang
mesh_terms: D055371:Acute Lung Injury / Q000139:chemically induced / Q000378:metabolism*; D066258:Amphiregulin; D000818:Animals; D019138:Antibodies, Blocking / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects; D001992:Bronchoalveolar Lavage Fluid / Q000166:cytology; D002478:Cells, Cultured; D004002:Clodronic Acid / Q000494:pharmacology; D066255:EGF Family of Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000494:pharmacology; D008070:Lipopolysaccharides; D008168:Lung / Q000473:pathology; D008262:Macrophage Activation; D016676:Macrophages, Alveolar / Q000187:drug effects / Q000378:metabolism*; D017737:Macrophages, Peritoneal / Q000187:drug effects; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism
chemical_list: D066258:Amphiregulin; D019138:Antibodies, Blocking; C587849:Areg protein, mouse; D066255:EGF Family of Proteins; D008070:Lipopolysaccharides; D004002:Clodronic Acid; D051057:Proto-Oncogene Proteins c-akt