Effect of Ambroxol and Beclomethasone on Lipopolysaccharide-Induced Nitrosative Stress in Bronchial Epithelial Cells

Respiration. 2015;89(6):572-82. doi: 10.1159/000381905. Epub 2015 May 12.

Abstract

Background: Nitrosative stress is involved in different airway diseases. Lipopolysaccharide (LPS) induces neutrophil-related cytokine release and nitrosative stress in human bronchial epithelial (BEAS-2B) cells alone or with human polymorphonuclear neutrophils (PMNs). Ambroxol protects against oxidative stress, and beclomethasone dipropionate is an anti-inflammatory drug.

Objectives: We evaluated the ability of ambroxol and/or beclomethasone dipropionate to inhibit LPS-induced expression/release of RANTES, IL-8, inducible NO synthase (iNOS), myeloperoxidase (MPO) and 3-nitrotyrosine (3-NT: nitrosative stress biomarker) in BEAS-2B ± PMNs stimulated with LPS (1 μg/ml).

Methods: The effect of ambroxol and/or beclomethasone dipropionate on IL-8, RANTES and iNOS levels was assessed by Western blot analysis; IL-8, MPO and 3-NT levels were measured by ELISA. Cell viability was assessed by the trypan blue exclusion test.

Results: In BEAS-2B alone, LPS (at 12 h) increased RANTES/iNOS expression and IL-8 levels (p < 0.001). Ambroxol suppressed LPS-induced RANTES expression and IL-8 release (p < 0.001), whilst inhibiting iNOS expression (p < 0.05). Beclomethasone dipropionate had no effect on RANTES but halved iNOS expression and IL-8 release. Coculture of BEAS-2B with PMNs stimulated IL-8, MPO and 3-NT production (p < 0.001), potentiated by LPS (p < 0.001). Ambroxol and beclomethasone dipropionate inhibited LPS-stimulated IL-8, MPO and 3-NT release (p < 0.05). Ambroxol/beclomethasone dipropionate combination potentiated the inhibition of IL-8 and 3-NT production in BEAS-2B with PMNs (p < 0.05 and p < 0.01, respectively). Ambroxol and/or beclomethasone dipropionate inhibited nitrosative stress and the release of neutrophilic inflammatory products in vitro.

Conclusion: The additive effect of ambroxol and beclomethasone dipropionate on IL-8 and 3-NT inhibition suggests new therapeutic options in the treatment of neutrophil-related respiratory diseases such as chronic obstructive pulmonary disease and respiratory infections.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Ambroxol / pharmacology*
  • Beclomethasone / pharmacology*
  • Bronchi / cytology
  • Cell Line
  • Chemokine CCL5 / drug effects
  • Chemokine CCL5 / metabolism
  • Epithelial Cells / drug effects*
  • Expectorants / pharmacology*
  • Glucocorticoids / pharmacology*
  • Humans
  • Interleukin-8 / drug effects
  • Interleukin-8 / metabolism
  • Lipopolysaccharides / toxicity
  • Neutrophils / drug effects
  • Neutrophils / metabolism
  • Nitric Oxide Synthase Type II / drug effects
  • Nitric Oxide Synthase Type II / metabolism
  • Nitrosation / drug effects
  • Peroxidase / drug effects
  • Peroxidase / metabolism
  • Respiratory Mucosa / cytology
  • Stress, Physiological / drug effects*
  • Tyrosine / analogs & derivatives
  • Tyrosine / drug effects
  • Tyrosine / metabolism

Substances

  • CCL5 protein, human
  • CXCL8 protein, human
  • Chemokine CCL5
  • Expectorants
  • Glucocorticoids
  • Interleukin-8
  • Lipopolysaccharides
  • Ambroxol
  • 3-nitrotyrosine
  • Tyrosine
  • Peroxidase
  • NOS2 protein, human
  • Nitric Oxide Synthase Type II
  • Beclomethasone