Abstract
Gingival junctional epithelial cell apoptosis caused by periodontopathic bacteria exacerbates periodontitis. This pathological apoptosis is involved in the activation of transforming growth factor β (TGF-β). However, the molecular mechanisms by which microbes induce the activation of TGF-β remain unclear. We previously reported that Aggregatibacter actinomycetemcomitans (Aa) activated TGF-β receptor (TGF-βR)/smad2 signalling to induce epithelial cell apoptosis, even though Aa cannot bind to TGF-βR. Additionally, outer membrane protein 29 kDa (Omp29), a member of the Aa Omps family, can induce actin rearrangements via focal adhesion kinase (FAK) signalling, which also plays a role in the activation of TGF-β by cooperating with integrin. Accordingly, we hypothesized that Omp29-induced actin rearrangements via FAK activity would enhance the activation of TGF-β, leading to gingival epithelial cell apoptosis in vitro. By using human gingival epithelial cell line OBA9, we found that Omp29 activated TGF-βR/smad2 signalling and decreased active TGF-β protein levels in the extracellular matrix (ECM) of cell culture, suggesting the transactivation of TGF-βR. Inhibition of actin rearrangements by cytochalasin D or blebbistatin and knockdown of FAK or integrinβ1 expression by siRNA transfection attenuated TGF-βR/smad2 signalling activity and reduction of TGF-β levels in the ECM caused by Omp29. Furthermore, Omp29 bound to fibronectin (Fn) to induce its aggregation on integrinβ1, which is associated with TGF-β signalling activity. All the chemical inhibitors and siRNAs tested blocked Omp29-induced OBA9 cells apoptosis. These results suggest that Omp29 binds to Fn in order to facilitate Fn/integrinβ1/FAK signalling-dependent TGF-β release from the ECM, thereby inducing gingival epithelial cell apoptosis via TGF-βR/smad2 pathway.
© 2016 John Wiley & Sons Ltd.
MeSH terms
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Aggregatibacter actinomycetemcomitans / genetics*
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Aggregatibacter actinomycetemcomitans / metabolism
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Apoptosis / genetics
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Bacterial Outer Membrane Proteins / genetics*
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Bacterial Outer Membrane Proteins / metabolism
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Bacterial Outer Membrane Proteins / pharmacology
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Cell Line, Transformed
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Cytochalasin D / pharmacology
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Epithelial Cells / drug effects
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Epithelial Cells / metabolism
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Epithelial Cells / microbiology*
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Fibronectins / genetics*
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Fibronectins / metabolism
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Focal Adhesion Kinase 1 / genetics*
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Focal Adhesion Kinase 1 / metabolism
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Gene Expression Regulation
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Gingiva / metabolism
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Gingiva / microbiology
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Heterocyclic Compounds, 4 or More Rings / pharmacology
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Host-Pathogen Interactions
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Humans
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Integrin beta1 / genetics*
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Integrin beta1 / metabolism
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Protein Serine-Threonine Kinases / genetics
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Protein Serine-Threonine Kinases / metabolism
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RNA, Small Interfering / genetics
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RNA, Small Interfering / metabolism
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Receptor, Transforming Growth Factor-beta Type I
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Receptors, Transforming Growth Factor beta / genetics
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Receptors, Transforming Growth Factor beta / metabolism
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Recombinant Proteins / genetics
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Recombinant Proteins / metabolism
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Recombinant Proteins / pharmacology
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Signal Transduction
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Smad2 Protein / antagonists & inhibitors
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Smad2 Protein / genetics
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Smad2 Protein / metabolism
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Transforming Growth Factor beta / antagonists & inhibitors
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Transforming Growth Factor beta / genetics*
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Transforming Growth Factor beta / metabolism
Substances
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Bacterial Outer Membrane Proteins
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Fibronectins
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Heterocyclic Compounds, 4 or More Rings
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Integrin beta1
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Omp29 protein, Actinobacillus actinomycetemcomitans
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RNA, Small Interfering
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Receptors, Transforming Growth Factor beta
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Recombinant Proteins
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SMAD2 protein, human
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Smad2 Protein
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Transforming Growth Factor beta
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blebbistatin
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Cytochalasin D
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Focal Adhesion Kinase 1
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PTK2 protein, human
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Protein Serine-Threonine Kinases
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Receptor, Transforming Growth Factor-beta Type I