Conclusion
P. gingivalis supernatant increased the migratory ability of monocytes, in part, by increasing activation and expression of MMP-9.
Material and methods
Human THP1 monocytes were treated with culture supernatant derived from P. gingivalis (ATCC 33277) for 24 h. Zymography, western blot analysis and quantitative PCRs were performed to analyse protein and mRNA levels of MMP-9. Protein and mRNA levels of TIMP-1 from monocytes treated with or without P. gingivalis were determined as well. Transwell migration assay was carried out to analyse the effect of P. gingivalis on the migration of human peripheral blood CD14-positive monocytes. An MMP inhibitor (GM6001) and a proteinase inhibitor (leupeptin) were used to determine the role of MMP-9 in P. gingivalis supernatant- and lipopolysaccharide-induced monocyte migration.
Methods
Human THP1 monocytes were treated with culture supernatant derived from P. gingivalis (ATCC 33277) for 24 h. Zymography, western blot analysis and quantitative PCRs were performed to analyse protein and mRNA levels of MMP-9. Protein and mRNA levels of TIMP-1 from monocytes treated with or without P. gingivalis were determined as well. Transwell migration assay was carried out to analyse the effect of P. gingivalis on the migration of human peripheral blood CD14-positive monocytes. An MMP inhibitor (GM6001) and a proteinase inhibitor (leupeptin) were used to determine the role of MMP-9 in P. gingivalis supernatant- and lipopolysaccharide-induced monocyte migration.
Objective
The migration of monocytes into the local environment is crucial for their maturation into macrophages or osteoclasts in the pathogenesis of periodontal disease. The objective of this study was to investigate the role and mechanisms mediated by Porphyromonas gingivalis in promoting the migration of monocytes by regulating MMP-9 and TIMP-1 expression. Material and
Results
In zymography and western blot, an 82 kDa band of active MMP-9 emerged in P. gingivalis-treated monocyte culture media in a dose-dependent manner, in addition to the MMP-9 proenzyme (92 kDa) band expressed in control cell culture media. P. gingivalis supernatant increased both the protein and the mRNA levels of MMP-9 and TIMP-1. P. gingivalis supernatant, but not its lipopolysaccharide, increased the migratory ability of CD14-positive monocytes. The increased migratory ability of P. gingivalis-treated monocytes was partly inhibited by leupeptin (200 μg/mL) and completely antagonized by the MMP inhibitor GM6001 (100 nm). Lipopolysaccharide of P. gingivalis increased protein and mRNA levels of MMP-9 in monocytes, but had no effect on the migratory ability or MMP-9 activation.