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Published on: May 28, 2021
German cockroach proteases regulate matrix metalloproteinase-9 in human bronchial epithelial cells
K Page1, V S Hughes, G W Bennett
1Division of Critical Care Medicine, Cincinnati Children's Hospital Medical Center and Cincinnati Children's Research Foundation, Cincinnati, OH 45229, USA.
Background:
Matrix metalloproteinases (MMPs) digest extracellular matrix proteins and may play a role in the pathogenesis of bronchial asthma. MMP-9 levels are increased in the bronchoalveolar lavage fluid and sputum of asthmatics compared with that of controls. As exposure to cockroaches is an environmental risk factor for asthma, we sought to investigate the role of German cockroach fecal remnants (frass) on MMP-9 expression.
Methods:
Human bronchial epithelial cells (16HBE14o-) and primary normal human bronchial epithelial cells were treated with cockroach frass in the absence or presence of tumor necrosis factor (TNF)alpha. MMP-9 mRNA, protein levels and pro-MMP-9 activity were determined using real-time polymerase chain reaction (PCR), enzyme-linked immunosorbent assay (ELISA) and zymogram assays. Pretreatment of frass with aprotinin abolished protease activity. PD98059, a chemical inhibitor of extracellular signal regulated kinase (ERK), and SLIGKV, an activator of protease-activated receptor (PAR)-2 were also used. AP-1DNA binding was determined by electrophoretic mobility shift assay (EMSA) and ERK phosphorylation by Western blot analysis.
Results:
Cockroach frass augmented TNFalpha-mediated MMP-9 mRNA and protein expression by a mechanism dependent on active serine proteases within frass and not on endogenous endotoxin. Frass increased ERK phosphorylation, and chemical inhibition of ERK attenuated cockroaches' effects on MMP-9. Serine proteases are known to activate the PAR-2 receptor. We found that selective activation of PAR-2 using the peptide SLIGKV augmented TNFalpha-induced MMP-9 protein levels and increased ERK phosphorylation. Frass and SLIGKV each increased AP-1 translocation and DNA binding.
Conclusions:
These data suggest that German cockroach frass contains active serine proteases which augment TNFalpha-induced MMP-9 expression by a mechanism involving PAR-2, ERK and AP-1.
Insights
German cockroach frass contains serine proteases that increase matrix metalloproteinase-9 (MMP-9) expression in asthma. This involves protease-activated receptor-2 (PAR-2), extracellular signal-regulated kinase (ERK), and AP-1 activation.
Area of Science:
- Immunology
- Environmental Health
- Molecular Biology
Background:
- Matrix metalloproteinases (MMPs), particularly MMP-9, are implicated in bronchial asthma pathogenesis.
- Elevated MMP-9 levels are observed in asthmatic patients' respiratory samples.
- German cockroach exposure is a known asthma risk factor, prompting investigation into its effects on MMP-9.
Purpose of the Study:
- To investigate the role of German cockroach fecal remnants (frass) in modulating MMP-9 expression.
- To elucidate the molecular mechanisms underlying cockroach frass-induced MMP-9 production.
Main Methods:
- Human bronchial epithelial cells were treated with cockroach frass and tumor necrosis factor-alpha (TNFα).
- MMP-9 mRNA, protein, and activity were quantified using RT-PCR, ELISA, and zymography.
- Involvement of serine proteases, protease-activated receptor-2 (PAR-2), extracellular signal-regulated kinase (ERK), and AP-1 was assessed using specific inhibitors and activators.
Main Results:
- Cockroach frass potentiated TNFα-induced MMP-9 expression via active serine proteases, independent of endotoxin.
- Frass increased ERK phosphorylation, and ERK inhibition attenuated MMP-9 induction.
- Activation of PAR-2 by frass or a specific peptide (SLIGKV) mimicked these effects, increasing MMP-9 and ERK phosphorylation.
- Both frass and SLIGKV enhanced AP-1 translocation and DNA binding.
Conclusions:
- German cockroach frass contains active serine proteases that augment TNFα-induced MMP-9 expression.
- The mechanism involves the activation of PAR-2, leading to ERK phosphorylation and AP-1 activation.
