Porphyromonas gingivalis affects host collagen degradation by affecting expression, activation, and inhibition of matrix metalloproteinases.
ABSTRACT Studies have shown that Porphyromonas gingivalis and host matrix metalloproteinases (MMPs) play important roles in the tissue destruction associated with periodontal disease. It is still unclear which MMPs or their inhibitors are regulated by P. gingivalis at the transcriptional and/or at the protein levels. Therefore, this study was conducted to determine what effects P. gingivalis supernatant has on the collagen degrading ability of human gingival fibroblasts (HGFs) and how it regulates the activation, mRNA expression, and inhibition of MMPs.
Culture supernatant from P. gingivalis ATCC 33277 was added to HGFs cultured in six-well plates coated with Type I collagen. At certain time intervals, the cell conditioned media was collected for zymography and/or western blot analyses to determine the MMP and tissue inhibitor of MMPs (TIMP) protein levels. The cells were then removed and the collagen cleavage visualized by Coomassie blue staining. The mRNA expression of multiple MMPs and TIMPs by the treated and untreated HGFs was determined by reverse transcription-polymerase chain reaction.
The collagen in the six-well plates was degraded more rapidly by the HGFs treated with 10% v/v P. gingivalis supernatant. More active MMP-1, MMP-2, MMP-3, and MMP-14 were detected in the conditioned media from the HGFs treated with the P. gingivalis supernatant. TIMP-1, but not TIMP-2, was decreased in the presence of the P. gingivalis supernatant. MMP-1 mRNA expression by the treated HGFs increased more than two-fold over the untreated HGFs. MMP-3 mRNA was unchanged, MMP-2 mRNA had a slight increase, MMP-14 mRNA decreased, and MMP-15 increased. MMP-12 mRNA was induced in the P. gingivalis treated HGFs. TIMP-1 and TIMP-2 mRNA had a slight increase with P. gingivalis treatment.
Porphyromonas gingivalis increased the collagen degrading ability of HGFs, in part, by increasing MMP activation and by lowering the TIMP-1 protein level, as well as by affecting the mRNA expression of multiple MMPs and TIMPs.
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ABSTRACT: In vitro cytotoxicity of six contemporary commercial dental filling restoratives on human dental primary cells, pulp cells (HPCs) and human gingival fibroblasts (HGFs), were tested using WST-1 assay. Continuous 3T3 mouse fibroblast cell lines were used for comparison. The results show that conventional glass-ionomer cement (GIC) Fuji II is not cy-totoxic to all the cells. Resin-modified GIC (RMGIC) Fuji II LC is not cytotoxic to both HPCs and HGFs but cytotoxic to 3T3 cells. RMGIC Vitremer and resin composite Z100 are very cytotoxic to all the cells. Resin composite P60 is cy-totoxic but much less cytotoxic than Z100. Polycarboxylate cement Durelon is the most cytotoxic among the six tested materials. It was found that continuous 3T3 cell lines were more vulnerable to leachable cytotoxic components than primary HPCs and HGFs. It was also found that the cytotoxcity of the tested materials was dose-dependent.
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ABSTRACT: Generalized aggressive periodontitis (GAgP) is a complex periodontal disease affecting the entire dentition with a rapid destruction of the periodontium and resulting in loss of teeth. We hypothesized that better clinical healing of adjunctive use of amoxicillin plus metronidazole combination may be related to the effect of this combination therapy to restore imbalance between matrix metalloproteinases (MMP) and their tissue inhibitors (TIMP) which is associated with connective tissue and alveolar bone destruction in patients with GAgP. Twenty-eight subjects diagnosed with GAgP were recruited. Patients were randomly assigned to test or control groups. MMP-1/TIMP-1 ratio was compared between groups receiving scaling and root planning (SRP) alone (control) or in combination with amoxicillin plus metronidazole (test). Clinical periodontal variables were measured. Gingival crevicular fluid samples were obtained and analyzed for MMP-1 and TIMP-1. Measurements were taken at baseline and repeated at 3 and 6 months after therapy. Total MMP-1 levels were significantly decreased in both groups (P < 0.05) at 3 and 6 months. MMP-1 concentration levels showed a similar pattern to MMP-1 total levels decreasing significantly at 3 months (P < 0.05). TIMP-1 concentration levels increased in the test group throughout the study period, while the difference did not reach statistical significance (P > 0.05). TIMP-1/MMP-1 balance was restored in test group at 6 months significantly better than the control group (P < 0.05). The results of this study suggest that metronidazole and amoxicillin combination as an adjunct to SRP results in better clinical healing through restoring TIMP-1/MMP-1 balance.
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ABSTRACT: The objective of this study is to demonstrate the molecular action of Porphyromonas gingivalis cysteine proteases such as gingipains (R1, R2 and K) upon human molecules.Materials and methodsUsing the information on protein structure and function available in international databases (UniProtKB and Merops Database), the molecular interactions already described between gingipains and host molecules were clarified.ResultsPossible cleavage sites were identified in host-produced elastase inhibitors and in pro-Matrix MetalloProteinase (MMP)1. Analysis of the results leads to the suggestion that the elastase inhibitor alpha1-antitrypsin is also degraded by interpain A, a cystein protease of Prevotella intermedia sharing a high homology with the PrtT and periodontain of P. gingivalis.Conclusion The information obtained suggests a synergistic molecular mechanism by which cysteine proteases of different bacteria can be responsible for the clinical manifestations of periodontal disease, and illustrates the use of bioinformatics to establish and predict molecular mechanisms.10/2012; 53(4):240-245. DOI:10.1016/j.rpemd.2012.07.002