Abstract
BackgroundCeramide, a bioactive lipid, plays an essential role in the development of several pulmonary inflammatory diseases. Matrix metallopeptidase 9 (MMP-9) regulates the synthesis and degradation of extracellular matrix, and is associated with airway remodeling and tissue injury. This study was conducted to investigate the effects and underlying mechanisms of ceramide on MMP-9 expression in airway epithelium.MethodsBEAS-2B cells, normal human bronchial epithelium cell lines, were pretreated with AG490, a selective janus tyrosine kinase 2 (JAK2) inhibitor, or Stattic, a selective signal transducer and activator of transcription 3 (STAT3) inhibitor. The cells were then stimulated with C6-ceramide. The levels of MMP-9 were determined by ELISA and real-time quantitative PCR (RT-qPCR). JAK2, phosphorylated JAK2 (p-JAK2), STAT3, and phosphorylated STAT3 (p-STAT3) expression was examined by Western blotting. BALB/c mice were pretreated with AG490 or Stattic before intratracheally instillated with C6-ceramide. Pathological changes in lung tissues were examined by Hematoxylin and Eosin staining, Periodic-acid Schiff staining, and Masson’s trichrome staining. MMP-9, JAK2, p-JAK2, STAT3, and p-STAT3 expression in the lung tissues was examined by Western blotting.ResultsThe expression of MMP-9, p-JAK2 and p-STAT3 in BEAS-2B cells was significantly increased after the treatment of C6-ceramide. Furthermore, the increased expression of MMP-9 induced by C6-ceramide was inhibited by AG490 and Stattic. Similar results were obtained in the lung tissues of C6-ceramide-exposed mice which were treated with AG490 or Stattic.ConclusionsCeramide could up-regulate MMP-9 expression through the activation of the JAK2/STAT3 pathway in airway epithelium. Targeted modulation of the ceramide signaling pathway may offer a potential therapeutic approach for inhibiting MMP-9 expression. This study points to a potentially novel approach to alleviating airway remodeling in inflammatory airway diseases.
Highlights
Airway epithelium is a pivotal structure with the greatest surface exposed to a large number of external stimulus, including respiratory viruses, air pollutants, and allergens
C6-ceramide increased matrix metalloproteinases (MMPs)-9 expression in BEAS-2B In order to study the effects of ceramide on Matrix metallopeptidase 9 (MMP-9) expression, human bronchial epithelial BEAS-2B cells were treated with C6-ceramide, a synthetic cell-permeable ceramide analog
Compared to cells incubated with DMSO only, treatment with 10 μM, 5 μM, and 2.5 μM of C6-ceramide increased the relative MMP-9 mRNA levels from 1.00 ± 0.09 to 6.62 ± 0.65 (P < 0.01), 4.68 ± 0.32 (P < 0.01), and 3.82 ± 0.15 (P < 0.01), respectively (Fig. 1a), and increased the MMP-9 protein levels from 387.57 ± 40.79 pg/mL to 4481.76 ± 506.40 pg/mL (P < 0.01), 3361.10 ± 139.19 pg/mL (P < 0.01), and 1536.25 ± 53.05 pg/mL (P < 0.01), respectively (Fig. 1b)
Summary
Airway epithelium is a pivotal structure with the greatest surface exposed to a large number of external stimulus, including respiratory viruses, air pollutants, and allergens. Growing evidence has showed that pulmonary disorders, such as asthma, chronic obstructive pulmonary disease (COPD), and emphysema, are associated with epithelial dysfunction [2,3,4,5]. Ceramide is significantly increased in the lungs and serum of patients with asthma and COPD [14, 15]. Ceramide is associated with the phenotypes of asthma and COPD, and may contribute to exacerbation and poor disease control by worsening airway inflammation [16, 17]. Studies by Oyeniran et al showed that intratracheal delivery of ceramide in mice promoted pulmonary inflammation and tissue remodeling, and caused airway flow obstruction [18]. A bioactive lipid, plays an essential role in the development of several pulmonary inflammatory diseases. This study was conducted to investigate the effects and underlying mechanisms of ceramide on MMP-9 expression in airway epithelium
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