Activation of Airway Epithelial Cells by Toll-Like Receptor Agonists

Abstract

Toll-like receptors (TLR) play an important role in pathogen recognition and innate immunity. We investigated the presence and function of TLRs in the BEAS-2B airway epithelial cell line and primary bronchial epithelial cells. Standard real-time reverse transcriptase-polymerase chain reaction (RT-PCR) analysis and Taqman RT-PCR revealed that BEAS-2B cells express mRNA for TLR1-10. Several TLR ligands were tested for their ability to activate gene expression in BEAS-2B cells using limited microarray analyses focusing on genes of the chemokine and chemokine receptor family, cytokines, and signaling pathways. While the TLR3 ligand double-stranded RNA was the most effective epithelial activator, clear responses to flagellin, lipopolysaccharide, CpG, peptidoglycan, and zymosan were also observed. RT-PCR and/or enzyme-linked immunosorbent assay were used to confirm results obtained with microarrays for five of the induced genes: interleukin-8, serum amyloid A, TLR3, macrophage inflammatory protein-3alpha, and granulocyte-macrophage colony-stimulating factor. Stimulation of epithelial cells with double-stranded RNA induced levels of interleukin-8 exceeding 20 ng/ml and levels of serum amyloid A exceeding 80 ng/ml. Double-stranded RNA, lipopolysaccharide, zymosan A, and flagellin also induced expression of macrophage inflammatory protein-3alpha and granulocyte-macrophage colony-stimulating factor, which may facilitate immature dendritic cell migration and maturation. These results suggest that airway epithelial cells express several TLRs and that they are functionally active. Epithelial expression of TLRs may be of importance in inflammation and immunity in the airways in response to inhaled pathogens.

Lipoteichoic Acid Increases TLR and Functional Chemokine Expression while Reducing Dentin Formation in In Vitro Differentiated Human Odontoblasts

“…including keratinocytes (24), intestinal epithelial cells (25), bronchial epithelial cells (26), and gingival epithelial cells (27). Interestingly, the pattern of TLRs expressed by odontoblasts was similar to the one reported for gingival fibroblasts in primary cultures (27).…”

Radioresistant cells expressing TLR5 control the respiratory epithelium's innate immune responses to flagellin

“…A variety of studies have demonstrated that lung and intestinal epithelial cells respond to flagellin by producing PMN-specific chemokines (CXCL1, CXCL5 or CXCL8), cytokines (TNF-a and IL-6) and antimicrobial molecules (NO and h-bD2) [18]. These findings agree with TLR5's predominant expression in lung and intestinal epithelial cells; hence, the epithelium is considered to be one of the main TLR5-responsive compartments [34,35]. Here, we used Salmonella typhimurium flagellin and its cognate detector TLR5 as models for investigating the contribution of radioresistant and radiosensitive cells to lung innate immunity.…”

“…These data are corroborated by experiments on animals lacking the TRIF adaptor molecule used by TLR4. Hence, the flagellin-mediated lung innate response depends solely on the TLR5 receptor and the MyD88 adaptor.Airway epithelial cells also produce chemokines (such as CCL20) that attract monocytes/dendritic cells and that may potentiate adaptive immunity in a mucosal adjuvant mechanism [34,[51][52][53][54][55]. In contrast, flagellin may directly activate lung dendritic cells [48,56].…”

Regulation of Airway Mucin Gene Expression

“…P. aeruginosa also has a protein called flagellin that can activate airway epithelial cells and MUC2 expression (34). Although flagellin normally binds to TLR5 and activates NF-κB in airway cells (35), the induction of MUC2 appears to work through the binding of flagella by ASGM1, a membrane ganglioside. After binding ASGM1, ATP is released and binds to an extracellular nucleotide receptor that signals through phospholipase C activation and the generation of inositol triphosphate and Ca 2+ .…”