Actinobacillus actinomycetemcomitans lipopolysaccharide activates matrix metalloproteinase-2 and increases receptor activator of nuclear factor-kappaB ligand expression in human periodontal ligament cells.
BACKGROUND
The lipopolysaccharide (LPS) of A. actinomycetemcomitans is one of the major pathogenic factors in periodontal disease. It induces secretion of proinflammatory cytokines and is involved in alveolar bone destruction. We hypothesized that the LPS of A. actinomycetemcomitans could affect the activation of matrix metalloproteinase (MMP)-2 and the expression of receptor activator of nuclear factor kappa B ligand (RANKL) and osteoprotegerin in human periodontal ligament (HPDL) cells leading to the destruction of periodontium.


METHODS
HPDL cells were cultured in serum-free medium with or without the LPS of A. actinomycetemcomitans for 36 hours. The activation of MMP-2 was analyzed by zymography. Changes of the expression of RANKL and osteoprotegerin (OPG) were examined by reverse transcription-polymerase chain reaction and supported by Western blot analysis.


RESULTS
The activation of MMP-2 could be induced by the LPS of A. actinomycetemcomitans in HPDL cells and could be inhibited by a serine protease inhibitor. This result suggested that the LPS might activate MMP-2 through a serine protease-dependent pathway. This activation was also blocked by NF-kappaB inhibitor, which indicated the involvement of NF-kappaB. The upregulation of RANKL but not OPG by the LPS was found in both transcription and translation and could be reduced by indomethacin. In addition, serine protease inhibitor also inhibited the upregulation of RANKL, suggesting the activity of serine protease.


CONCLUSIONS
The effect of the LPS of A. actinomycetemcomitans on HPDL cells is serum-independent and the induction of the activation of MMP-2 and the expression of RANKL are serine protease-dependent pathways. The results suggest the role of HPDL cells in the pathogenesis of periodontitis.
Tiranathanagul
Siriluck
S
Department of Anatomy, Faculty of Dentistry, Chulalongkorn University, Bangkok, Thailand.
Yongchaitrakul
Tussanee
T
Pattamapun
Kassara
K
Pavasant
Prasit
P
D016976:Aggregatibacter actinomycetemcomitans / Q000737:chemistry / Q000472:pathogenicity*; D016301:Alveolar Bone Loss / Q000209:etiology*; D015153:Blotting, Western; D002352:Carrier Proteins / Q000096:biosynthesis*; D002478:Cells, Cultured; D002851:Chromatography, High Pressure Liquid; D004789:Enzyme Activation / Q000187:drug effects; D006023:Glycoproteins / Q000096:biosynthesis; D006801:Humans; D008024:Ligands; D008070:Lipopolysaccharides / Q000494:pharmacology; D020778:Matrix Metalloproteinase 2 / Q000378:metabolism*; D008562:Membrane Glycoproteins / Q000096:biosynthesis*; D016328:NF-kappa B / Q000378:metabolism; D053244:Osteoprotegerin; D010513:Periodontal Ligament / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear / Q000096:biosynthesis; D018124:Receptors, Tumor Necrosis Factor; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012697:Serine Endopeptidases / Q000378:metabolism
D002352:Carrier Proteins; D006023:Glycoproteins; D008024:Ligands; D008070:Lipopolysaccharides; D008562:Membrane Glycoproteins; D016328:NF-kappa B; D053244:Osteoprotegerin; D053245:RANK Ligand; D053246:Receptor Activator of Nuclear Factor-kappa B; D018160:Receptors, Cytoplasmic and Nuclear; D018124:Receptors, Tumor Necrosis Factor; C505686:TNFRSF11A protein, human; C506030:TNFRSF11B protein, human; C505688:TNFSF11 protein, human; D012697:Serine Endopeptidases; D020778:Matrix Metalloproteinase 2