Dysregulated Macrophages Are Present in Bleomycin-Induced Murine Laryngotracheal Stenosis

Otolaryngol Head Neck Surg. 2015 Aug;153(2):244-50. doi: 10.1177/0194599815589106. Epub 2015 Jun 17.

Abstract

Objective: To define the inflammatory cell infiltrate preceding fibrosis in a laryngotracheal stenosis (LTS) murine model.

Study design: Prospective controlled murine study.

Setting: Laboratory.

Subjects and methods: Chemomechanical injury mice (n = 44) sustained bleomycin-coated wire-brush injury to the laryngotracheal complex while mechanical injury controls (n = 42) underwent phosphate-buffered saline (PBS)-coated wire-brush injury. Mock surgery controls (n = 34) underwent anterior transcervical tracheal exposure only. Inflammatory and fibrosis protein and gene expression were assessed in each condition. Immunohistochemistry served as a secondary outcome.

Results: In chemomechanical injury mice, there was an upregulation of collagen I (P < .0001, P < .0001), Tgf-β (P = .0023, P = .0008), and elastin (P < .0001, P < .0001) on day 7; acute inflammatory gene Il1β (P = .0027, P = .0008) on day 1; and macrophage gene CD11b (P = .0026, P = .0033) on day 1 vs mechanical and mock controls, respectively. M1 marker inducible nitric oxide synthase (iNOS) expression decreased (P = .0014) while M2 marker Arg1 (P = .0002) increased on day 7 compared with mechanical controls. Flow cytometry demonstrated increased macrophages (P = .0058, day 4) and M1 macrophages (P = .0148, day 4; P = .0343, day 7; P = .0229, day 10) compared to mock controls. There were similarities between chemomechanical and mechanical injury mice with an increase in M2 macrophages at day 10 (P = .0196).

Conclusions: The bleomycin-induced LTS mouse model demonstrated increased macrophages involved with the development of fibrosis. Macrophage immunophenotype suggested that dysregulated M2 macrophages have a role in abnormal laryngotracheal wound healing. These data delineate inflammatory cells and signaling pathways in LTS that may potentially be modulated to lessen fibroblast proliferation and collagen deposition.

Keywords: airway epithelial injury; laryngotracheal stenosis; mouse model; subglottic stenosis; trachea.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Bleomycin
  • Collagen / analysis
  • Disease Models, Animal
  • Elastin / analysis
  • Flow Cytometry
  • Gene Expression
  • Immunohistochemistry
  • Laryngostenosis / chemically induced
  • Laryngostenosis / pathology*
  • Larynx / injuries
  • Macrophages / pathology*
  • Mice
  • Mice, Inbred C57BL
  • Prospective Studies
  • Trachea / injuries
  • Tracheal Stenosis / chemically induced
  • Tracheal Stenosis / pathology*
  • Transforming Growth Factor beta / analysis

Substances

  • Transforming Growth Factor beta
  • Bleomycin
  • Collagen
  • Elastin