Multiple TLRs activate EGFR via a signaling cascade to produce innate immune responses in airway epithelium

Abstract

Toll-like receptors (TLRs) are critical for the recognition of inhaled pathogens that deposit on the airway epithelial surface. The epithelial response to pathogens includes signaling cascades that activate the EGF receptor (EGFR). We hypothesized that TLRs communicate with EGFR via epithelial signaling to produce certain innate immune responses. Airway epithelium expresses the highest levels of TLR2, TLR3, TLR5, and TLR6, and here we found that ligands for these TLRs increased IL-8 and VEGF production in normal human bronchial epithelial cells. These effects were prevented by treatment with a selective inhibitor of EGFR phosphorylation (AG-1478), a metalloprotease (MP) inhibitor, a reactive oxygen species (ROS) scavenger, and an NADPH oxidase inhibitor. In an airway epithelial cell line (NCI-H292), TNF-alpha-converting enzyme (TACE) small interfering RNA (siRNA) was used to confirm that TACE is the MP involved in TLR ligand-induced IL-8 and VEGF production. We show that transforming growth factor (TGF)-alpha is the EGFR ligand in this signaling cascade by using TGF-alpha neutralizing antibody and by showing that epithelial production of TGF-alpha occurs in response to TLR ligands. Dual oxidase 1 (Duox1) siRNA was used to confirm that Duox1 is the NADPH oxidase involved in TLR ligand-induced IL-8 and VEGF production. We conclude that multiple TLR ligands induce airway epithelial cell production of IL-8 and VEGF via a Duox1--> ROS--> TACE--> TGF-alpha--> EGFR phosphorylation pathway. These results show for the first time that multiple TLRs in airway epithelial cells produce innate immune responses by activating EGFR via an epithelial cell signaling cascade.

DUOX1 mediates persistent epithelial EGFR activation, mucous cell metaplasia, and airway remodeling during allergic asthma

“…DUOX1-dependent EGFR activation can promote airway epithelial production of the neutrophil chemokine IL-8 (27,30). Analysis of BAL fluids from HDM-challenged wild-type mice indicated marked and DUOX1-dependent increases of the mouse functional IL-8 homolog CXCL1, consistent with the observed DUOX1-dependent neutrophilia in this model ( Figure 4A).…”

“…An additional consequence of IL-33-mediated ILC2 recruitment and activation is the production of AREG, which forms an essential component of EGFR-mediated epithelial repair responses in the context of parasite/helminth infection (47) but may also contribute to sustained EGFR activation in chronic asthma. Moreover, DUOX1 contributes to EGFR activation not only by promoting EGFR ligand activation, as previously reported (23,27), but also by promoting ligand-induced activation of EGFR phosphorylation by direct cysteine oxidation within EGFR. Although chronic epithelial alterations during allergic asthma are commonly attributed to epigenetic changes, potentially involving important asthma-relevant cytokines such as IL-13 (8, 48, 49), our results indicate that such alterations are also mediated by redox-dependent mechanisms due to enhanced expression and activation of DUOX1 and can be attenuated by inhibition of DUOX1.…”

“…Similarly, dose-dependent activation of EGFR and Src in cultured mouse tracheal epithelial cells (mTECs) in response to Areg was attenuated in mTECs from Duox1 -/-mice compared with WT mice, which was closely associated with corresponding changes in cysteine oxidation within these kinases ( Figure 1F). Collectively, these findings indicate that persistent EGFR activation within epithelia of asthmatic subjects involves oxidative mechanisms originating from DUOX1, which can promote EGFR ligand shedding (23,27) and enhance EGFR tyrosine kinase activity by promoting cysteine oxidation with the kinase domain (25).…”

MyD88 signaling in nonhematopoietic cells protects mice against induced colitis by regulating specific EGF receptor ligands

“…In addition, the cell type in which EGFR signaling occurs to prevent experimental colitis needs to be identified. Some evidence from in vitro studies using colonic epithelial cell lines suggested the possibility that Toll-like receptor (TLR) signaling might induce expression of EGFR ligands (18)(19)(20)(21). Also, studies using a model of colitis-associated cancer demonstrate that TLR4-dependent tumorigenesis is associated with activation of EGFR signaling (18).…”

TACE/TGF- /EGFR regulates CXCL8 in bronchial epithelial cells exposed to particulate matter components

“…Accordingly, EGFR signalling has been suggested as a convergent pathway for regulation of CXCL8 and other immune responses [23]. Recent studies with endogenous and microbial ligands have shown that CXCL8 regulation by EGFR involves transforming growth factor (TGF)-a ectodomain shedding by the metalloprotease tumour necrosis factor-a-converting enzyme (TACE)/ADAM-17 [20,21,24,25]. Whether organic or inorganic PM components regulate CXCL8 through similar mechanisms remains to be elucidated.…”