Repeated and alternate stimulations with dsRNA and SEB alter responses in macrophages and epithelial cells

Introduction

The innate immune system is activated by foreign molecules via pattern recognition receptors and other surveillance systems, producing diverse cytokines that recruit and activate other immune cells. Recent studies have shown that once activated by foreign molecules, the innate immune system exhibits altered responses upon subsequent exposure to the same or different infectious agents, such as lipopolysaccharides (LPS) or bacteria. However, as these alterations in response to viral infection and staphylococcal enterotoxin B (SEB) in the airways have not been fully elucidated, we focused on the changes in immune responses induced by repeated stimulation of macrophages and epithelial cells with foreign molecules.

Methods

THP-1-derived macrophages and BEAS-2B epithelial cells were stimulated with dsRNA (double-stranded RNA) or SEB and cultured in fresh complete medium for 4 days. Subsequently, the cells were re-stimulated with different doses of dsRNA or SEB, and the cytokine and signal phosphorylation levels were evaluated.

Results

Repeated stimulation with high dose of dsRNA or SEB, induced an increase in IL-10, CCL2, CCL22, CCL24, CXCL10, and CXCL11 in macrophages, while only repeated stimulation with dsRNA stimulation resulted in an increase in IL-6, CCL2, CCL5, CCL24, CXCL11, and TGF-β in epithelial cells. Cross-stimulation with SEB-dsRNA induced an increase in CCL5, CCL20, CCL22, CCL24, CXCL10, and CXCL11 levels in macrophages. However, in epithelial cells, SEB-dsRNA stimulation increased the levels of CCL5, CXCL11, and TGF-β, while dsRNA-SEB stimulation elevated CCL1, CCL20, CXCL10, and CXCL11. These cytokine changes were driven by distinct phosphorylation patterns in macrophages and epithelial cells, depending on the type and intensity of stimuli.

Conclusion

Repeated stimulation with the same or cross-over stimuli induced alterations in the immune response of macrophages and epithelial cells. These observations indicated that persistent airway stimulation can lead to changes in airway inflammation, potentially leading to asthma.