Abstract
BackgroundMucus overproduction is an important feature of asthma. Interleukin (IL)-4 is required for allergen-induced airway inflammation and mucus production. MUC5AC gene expression is regulated by transcript factors NF-κB. The intracellular Ca2+ ([Ca2+]i) signal is required for activation of NF-κB. The transient receptor potential canonical 1 (TRPC1) channel has been shown to contribute for agonist-stimulated Ca2+ influx in some types of cells. However, the relationships among IL-4, TRPC1 and mucus overproduction in bronchial epithelial cells (BECs) in asthma are poorly understood.MethodsBECs were isolated from large bronchial airway of rats and used as cell model. To present changes of lipid raft, caveolin-1 and TRPC1, immunofluorescence staining and sucrose gradient centrifugation were performed. [Ca2+]i was measured after loading with Fura-2. NF-κB activities were measured by an ELISA-based assay. MUC5AC mRNA and protein levels were detected by real-time quantitative RT-PCR, ELISA analysis and immunofluorescence staining respectively.ResultsIL-4 induced Ca2+ influx in BECs, and this was blocked by a Ca2+ influx inhibitor (2-APB). 2-APB also prevented MUC5AC protein synthesis induced by IL-4. Depletion of extracellular Ca2+ resulted in partial decrease in expression of MUC5AC in IL-4 treated cells. NF-κB rather than STAT6 activation mediated IL-4-induced MUC5AC protein synthesis. Then the mechanism of Ca2+ influx was investigated. Immunofluorescence staining and sucrose gradient centrifugation revealed that caveolin-1-containing lipid rafts aggregation was involved in TRPC1 activation and Ca2+ influx in BECs. Lastly, the data revealed that blocking lipid rafts aggregation exactly prevented Ca2+ influx, NF-κB activation and MUC5AC synthesis induced by IL-4.ConclusionsOur results indicate that IL-4-induced caveolin-1-containing lipid rafts aggregation at least partly contributes to MUC5AC synthesis in BECs.
Highlights
Mucus overproduction is an important feature of asthma
We found that IL-4 caused caveolin-1 containing lipid rafts aggregation and TPRC1 colocalization, and disruption of lipid rafts prevented IL-4 induced NF-κB activation and MUC5AC expression
Caveolin-1 containing lipid rafts aggregation was involved in transient receptor potential canonical 1 (TRPC1) activation by IL-4 in bronchial epithelial cells (BECs) To explore the potential mechanism underlying extracellular Ca2+ influx induced by IL-4, we focused on the lipid rafts clustering in BECs
Summary
Interleukin (IL)-4 is required for allergeninduced airway inflammation and mucus production. There is over-expression of the major mucin glycoprotein, MUC5AC [2, 3]. Th2 cells predominantly secrete cytokines interleukin (IL)-4, IL-5, IL9 and IL-13, which play a central role in the pathophysiology of asthma. These type 2 cytokines are targets for pharmaceutical intervention of asthma. Studies showed that IL-4 is required for allergen-induced airway inflammation and mucus production [5,6,7]. IL-4 binds to IL-4 receptor in airway cells and can trigger STAT6 and NF-κB signal pathway which is likely to be involved in the regulation of MUC5AC expression
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