Neutrophil Elastase Induces MUC5AC Mucin Production in Human Airway Epithelial Cells via a Cascade Involving Protein Kinase C, Reactive Oxygen Species, and TNF-α-Converting Enzyme

Abstract

Mucus hypersecretion is a prominent manifestation in patients with chronic inflammatory airway diseases and contributes to their morbidity and mortality by plugging airways and causing recurrent infections. Human neutrophil elastase (HNE) exists in high concentrations (1–20 μM) in airway secretions of these patients and induces overproduction of MUC5AC mucin, a major component of airway mucus. Previous studies showed that HNE induces MUC5AC mucin production involving reactive oxygen species (ROS) generation and TGF-α-dependent epidermal growth factor receptor (EGFR) activation in human airway epithelial cells. However, the molecular mechanisms involved in these responses are not defined. TNF-α-converting enzyme (TACE) cleaves pro-TGF-α into soluble TGF-α and can be activated by ROS. We hypothesize that HNE activates TACE via ROS generation, resulting in cleavage of pro-TGF-α, EGFR activation, and MUC5AC mucin expression in airway epithelial cells. Here we show that in human airway epithelial cells HNE increases TGF-α release, EGFR phosphorylation, and MUC5AC mucin expression, effects that were attenuated by TACE inhibitor TAPI-1 and by specific knockdown of TACE expression with small interfering RNA, implicating TACE in HNE-induced responses. These responses to HNE were also reduced by pretreatment with ROS scavengers, implicating ROS. Furthermore, we show that HNE causes protein kinase C (PKC) activation and translocation from cytosol to plasma membrane; blockade of this effect by PKC inhibitors reduced HNE-induced ROS generation and other responses, implicating PKC. We conclude that HNE induces MUC5AC mucin expression via a cascade involving PKC-ROS-TACE in human airway epithelial cells.

EGF receptor activation during allergic sensitization affects IL‐6‐induced T‐cell influx to airways in a rat model of asthma

“…Amphiregulin and epiregulin have been reported to induce migration of urothelial cells, renal tubular cells and mammary epithelial cells [31][32][33], so they could have some potential effects on the migration of inflammatory cells in this study. It is perhaps surprising that TGF-a was not altered in our study, given its implication in epithelial differentiation following exposure to lipopolysaccharide and neutrophil elastase [3,34]. These results suggest that the contributions of epithelial-derived specific EGFR ligands and those derived from immune effector cells may differ, since the latter cells predominate in BAL.…”

Regulation of Airway Mucin Gene Expression

“…The differences in signaling and mechanistic steps are difficult to sort out because different cell systems and treatments were used. However, there are compelling data that reactive oxygen species (ROS) formation is required upstream for NE-mediated MUC5AC expression (74)(75)(76). The generation of ROS by NE appears to be mediated by different types of NADPH oxidases (77,78) and may also require PKC activation (74).…”

“…However, there are compelling data that reactive oxygen species (ROS) formation is required upstream for NE-mediated MUC5AC expression (74)(75)(76). The generation of ROS by NE appears to be mediated by different types of NADPH oxidases (77,78) and may also require PKC activation (74). This may mean that ROS act upstream of NF-κB in MUC5AC expression because ROS activation of NF-κB in airway epithelial and smooth muscle cells has been observed (79)(80)(81).…”

“…Signaling through EGFR (the ErbB1 receptor) has long been known to play an important role in modulating the expression of mucin genes (82,83). Many stimuli, including cigarette smoke, NE, phorbol esters, eosinophil products, bacterial products, and human airway trypsin-like protease, can upregulate MUC5AC through EGFR (18,73,74,(84)(85)(86). EGFR is expressed on airway epithelial cells, and in many cases its ligands, such as TGF-α and amphiregulin, are also present on the membrane in their inactive forms (55).…”

Proteases and cystic fibrosis

“…NE regulates mucin MUC5AC gene expression by both transcriptional (Song et al, 2005, Shao and Nadel, 2005b, Kohri, 2002, Shao and Nadel, 2005a) and post-transcription mechanisms (Voynow, 1999). NE stimulates translocation of PKCδ to the plasma membrane, resulting in activation of dual oxidase 1 (DUOX1 (Shao and Nadel, 2005a) and subsequent activation of TNFα-converting enzyme (TACE) to cleave and release the EGFR ligand, TGFα, permitting activation of EGFR and down-stream signaling via RAS-Raf-MEK1/2-ERK1/2 and either NF-κB (Song et al, 2005) or Sp1 (Perrais et al, 2002) activation.…”