No abstract

RATIONALE: Flagellin which is abundant in gram-negative bacteria including pseudomonas is reported to be related with development of asthma. However, its effect on airway epithelial cells regarding asthma pathogenesis is not elucidated yet. We investigated the effect of flagellin on the production of cytokines, chemokines and metalloproteinase related with airway inflammation and remodeling in the primary human epithelial cells. METHODS: Primary human bronchial epithelial cells were grown and differentiated in air-liquid interface culture for 14-16 days. The cells were treated with flagellin in vitro at 10 and 100ng/ml for 3, 24, and 48hours. The conditioned media and cells were harvested to measure cytokines, chemokines and metalloproteinase related with asthma and their mRNA expressions by ELISA or western blot and quantitative PCR, respectively. RNA-sequencing was performed to screen the up-regulated and downregulated transcriptomes by flagellin. RESULTS: Flagellin upregulated mRNA expressions of GM-CSF at 3hrs of treatment and increased GM-CSF secretion significantly at 24hr in conditioned media. Flagellin enhanced mRNA expressions and secretions of CCL5, CXCL10, CXCL11 chemokines, which were detected in the RNA-sequencing. In addition, mRNA expressions and protein productions of MMP9 and MMP13 were also enhanced by flagellin. TGF-b1 and TGF-b2 augmented the mRNA expressions of GM-CSF, MMP9 and MMP13 induced by flagellin. CONCLUSIONS: Flagellin is an inducer of cytokines, chemokines and metalloproteinase contributing to airway inflammation and remodeling in bronchial epithelial cells. Further studies are needed to clarify their interaction with other inflammatory and structural cells and clinical implications in terms of asthma.Abstracts AB187 SUNDAY

Foreign molecules, including viruses and bacteria-derived toxins, can also induce airway inflammation. However, to the best of our knowledge, the roles of these molecules in the development of airway inflammation have not been fully elucidated. Herein, we investigated the precise role and synergistic effect of virus-mimicking double-stranded RNA (dsRNA) and staphylococcal enterotoxin B (SEB) in macrophages and epithelial cells. To identify cytokine expression profiles, both the THP-1-derived macrophages and BEAS-2B epithelial cells were stimulated with dsRNA or SEB. A total of 21 cytokines were evaluated in the culture supernatants. We observed that stimulation with dsRNA induced cytokine production in both cell types. However, cytokine production was not induced in SEB-stimulated epithelial cells, compared to the macrophages. The synergistic effect of dsRNA and SEB was evaluated observing cytokine level and intracellular phospho-signaling. Fifteen different types were detected in high-dose dsRNA-stimulated epithelial cells, and 12 distinct types were detected in macrophages; those found in macrophages lacked interferon production compared to the epithelial cells. Notably, a synergistic effect of cytokine induction by co-stimulation of dsRNA and SEB was observed mainly in epithelial cells, via activation of most intracellular phosphor-signaling. However, macrophages only showed an accumulative effect. This study showed that the type and severity of cytokine productions from the epithelium or macrophages could be affected by different intensities and a combination of dsRNA and SEB. Further studies with this approach may improve our understanding of the development and exacerbation of airway inflammation and asthma.